BIOLOGY, ECOLOGY AND MANAGEMENT OF KEY PESTS OF 
                                                SATSUMA CITRUS IN ALABAMA 
 
by 
 
Yingfang Xiao 
 
 
 
 
A dissertation submitted to the Graduate Faculty of 
Auburn University 
in partial fulfillment of the 
requirements for the Degree of 
Doctor of Philosophy 
 
Auburn, Alabama 
December 18, 2009 
 
 
 
 
Keywords: Leptoglossus zonatus, Phyllocnistis citrella, and Panonychus citri, 
host preference, fruit damage, seasonal abundance, and biological control 
 
 
Copyright 2009 by Yingfang Xiao 
 
 
Approved by 
 
Henry Y. Fadamiro, Chair, Associate Professor of Entomology 
Arthur G. Appel, Alumni Professor of Entomology 
Charles Ray, Research Fellow in Entomology 
Elina Coneva, Assistant Professor of Horticulture  
 ii 
 
 
 
 
 
 
Abstract  
 
 
Satsuma mandarin (Citrus unshiu Marcovitch) production is an emerging industry in 
Alabama. Recent surveys identified Leptoglossus zonatus (leaffooted bug), Phyllocnistis citrella 
(citrus leafminer), and Panonychus citri (citrus red mite), as important pests of satsuma in 
Alabama. My dissertation comprised of three sections with the focus on the ecology and 
management of the above three key pests, and included laboratory and field studies. The first 
section (chapters II and III) focused on L. zonatus. Host preference and development of L. 
zonatus was studied in chapter II. The results showed that tomato was the most preferred fruit, 
but satsuma is also a suitable host. Damage to satsuma by L. zonatus was evaluated in chapter III. 
Feeding by L. zonatus on satsuma produced external damage, and resulted in significant fruit 
weight loss, fruit abortion, and reduced soluble solids content (SSC). Section 2 (chapters IV and 
V) focused on P. citrella. The seasonal phenology and natural enemy fauna of P. citrella in 
Alabama was investigated in chapter IV. The results showed multiple overlapping generations of 
P. citrella, and effective pheromone-baited traps were identified. At least 21 species of beneficial 
arthropods were recorded. In chapter V, I investigated the relative contributions of key beneficial 
arthropods to natural mortality of P. citrella in Alabama using exclusion techniques. Predation 
by spiders was the most important natural mortality factor, whereas parasitism was minimal. 
Section 3 (chapters VI and VII) focused on P. citri. Chapter VI presented a laboratory evaluation 
of three commercially available predacious mite species in the family Phytoseiidae (Phytoseiulus 
persimilis, Galendromus occidentalis, and Neoseiulus californicus), as potential biological 
 
 iii 
control agents of P. citri. All three species were effective in regulating P. citri density, but P. 
persimilis showed the highest predation potential followed by G. occidentalis. The results of 
field evaluations of small-scale releases of P. persimilis or G. occidentalis against P. citri 
(chapter VII) showed that two timed releases of either species at an appropriate release rate (100 
or more per tree) provided effective season-long suppression of P. citri. These results support the 
development of an IPM program for satsuma production in the Gulf Coast region.  
 
 iv 
 
 
 
 
 
 
Acknowledgements 
 
 
I am very grateful to my major advisor, Dr. Henry Fadamiro for his support, constant 
supervision, and patience during these years at Auburn. His scientific attitude and philosophy 
will influence me forever. His enthusiasm and generosity with time has contributed immensely to 
the success of this work. I am also indebted to my other committee members: Drs. Arthur G. 
Appel, Charles Ray, Elina Covena, and the outside reader, Dr. James Spiers for their genuine 
interest, time and valuable advice. I am also thankful to my course professors, Drs. Arthur Appel, 
Wayne Clark, Henry Fadamiro, Nannan Liu, Michael Williams and Joseph Kloepper, for their 
excellent teaching and encouragement. My sincere thanks also go to Mr. Monte Nesbitt and 
Terry Hargroder for assisting with field research. I am also grateful to all my former and present 
colleagues in the laboratory of Dr. Fadamiro in particular, Li Chen, Vincent Umeh, Ebenezer 
Onagbola, Hongyi Wei, Xiaofang He, Elly Maxwell, Esther Ngumbi, Shelia Boyt, Clement 
Akotsen-Mensah, Prithwiraj Das, Ramohan Rao Balusu, Timothy Nafziger, Kavita Sharma, and 
Kate Nangle for their encouragement, generosity and help. I would also like to acknowledge the 
Chair, faculty, staff and students of the Department of Entomology and Plant Pathology for their 
help. Special thanks to my wife for her selfless devotion, love and support. I am particularly 
grateful to God for the spirit of encouragement.  
 
 
 
 
 v 
 
 
 
 
 
 
Table of Contents  
 
 
Abstract???????????????????????????.???? ?? .......ii 
Acknowledgements??????????????????????????? ??... ..iv 
List of Tables?? ...........................................................................................................................x 
List of Figures??. ......................................................................................................................xiii 
Chapter 1 Introduction and Literature Review ...............................................................................1 
1.1 Satsuma Mandarin Production ............................................................................................1 
1.2 Major Arthropod Pests of Satsuma Mandarin in Alabama????????????..2  
1.2.1 Leaffooted Bugs, Leptoglossus zonatus......................................................................4 
1.2.1 Citrus Leafminer, Phyllocnistis citrella?????????????????. 6  
1.2.1 Citrus Red Mite, Panonychus citri???.???????? .???????...9     
1.3 Justification???......................................................................................................... ...12 
1.4 Dissertation Outline, Goals and Objectives.......................................................................12 
1.5 References..........................................................................................................................16 
Chapter 2 Host Preference and Development of the Leaffooted bug, Leptoglossus zonatus, on 
Satsuma Mandarin, Citrus unshiu?..............................................................................26                                                                                                               
2.1 Introduction........................................................................................................................26 
2.2 Materials and Methods ......................................................................................................27 
2.2.1 Rearing of L. zonatus ...............................................................................................27 
2.2.2 Host Preference of L. zonatus...................................................................................28 
 
 vi 
2.2.3 Development of L. zonatus on Satsuma Fruit??????????????...29  
2.3 Results ...............................................................................................................................30 
2.3.1 Host Preference of L. zonatus...................................................................................30 
2.3.2 Development of L. zonatus on Satsuma Fruits? ??????.? ??.. ..............32 
2.3.2.1 Basic Description of the Stages????????????...?????32  
2.3.2.2 Immature Development and Survival??????????????......32   
2.3.2.3 Adult Survival and Reproduction....................................................................32 
2.4 Discussion..........................................................................................................................33 
2.5 References..........................................................................................................................37 
Chapter 3 Evaluation of Damage to Satsuma Mandarin, Citrus unshiu, by the Leaffooted bug, 
Leptoglossus zonatus???????????????? ?????????. 45 
3.1 Introduction........................................................................................................................45 
3.2 Materials and Methods.......................................................................................................46 
3.2.1 Rearing L. zonatus....................................................................................................46 
3.2.2 Laboratory Tests?????????????????????????...47  
3.2.3 Field Tests.................................................................................................................48 
3.2.4 Statistical Analysis....................................................................................................49 
3.3 Results................................................................................................................................49 
3.3.1 Description of Fruit Damage....................................................................................49 
3.3.2 Laboratory Tests.......................................................................................................50 
3.3.3 Field Tests...........................................................................................................? ..51 
3.4 Discussion..........................................................................................................................52 
3.5 Acknowledgements............................................................................................................56 
3.6 References..........................................................................................................................57 
 
 vii 
Chapter 4 Seasonal Phenology and Natural Enemy Fauna of Citrus Leafminer, Phyllocnistis 
citrella, and Evaluation of Pheromone-Baited for Monitoring the Pest in Alabama 
Satsuma Orchards ??????....................................... ..........................................68                                   
4.1 Introduction........................................................................................................................68 
4.2 Materials and Methods ......................................................................................................70 
4.2.1 Seasonal Abundance of P. citrella Immatures??????????????.. 70 
4.2.2 Evaluation of Pheromone-Baited Traps and Adult Seasonal Phenology??.?..... 72 
4.3 Results and Discussion......................................................................................................73  
4.3.1 Seasonal Abundance of P. citrella Immatures?????????? .???.?73  
4.3.2 Natural Enemy Fauna???????????????????????...75  
4.3.3 Evaluation of Pheromone-Baited Traps???????????????........77  
4.3.4 Adult Seasonal Phenology ????????????????.......................78  
4.4 Acknowledgements............................................................................................................81 
4.5 References..........................................................................................................................82 
Chapter 5 Exclusion Experiments Reveal Relative Contribution of Natural Enemies to Mortality 
of Citrus Leafminer, Phyllocnistis citrella, in Alabama Satsuma Orchards ??? ?.. 96                                   
5.1 Introduction.......................................................................................................................96 
5.2 Materials and Methods .....................................................................................................99 
5.2.1 Study Sites ...............................................................................................................99 
5.2.2 Exclusion Experiments.............................................................................................99 
5.2.3 Evaluation of Mortality factors...............................................................................101 
5.2.4 Statistical Analyses ................................................................................................102 
5.3 Results..............................................................................................................................102 
5.4 Discussion........................................................................................................................104 
5.5 Acknowledgements..........................................................................................................109 
 
 viii 
5.6 References........................................................................................................................110 
Chapter 6 Functional Responses and Prey Stage Preferences of Three Species of Predaceous 
Mites of Citrus Red mites, Panonychus citri??????? ?. ???????. 123 
6.1 Introduction......................................................................................................................123 
6.2 Materials and Methods ....................................................................................................125 
6.2.1 Rearing of P. citri (the prey)...................................................................................125 
6.2.2 Rearing of Predacious Mites...................................................................................126 
6.2.3 Experiment 1. Functional Response.......................................................................126  
6.2.4 Experiment 2. Prey Stage Preference ....................................................................127  
6.2.5 Experiment 3. Effect of Starvation??????????????????.127  
6.3.6 Preference Quantification Methods........................................................................127 
6.2.7 Statistical Analyses????????????????????????.128  
6.3 Results..............................................................................................................................128 
6.3.1 Experiment 1. Functional Response?????????????????... 128 
6.3.1.1 Phytoseiulus persimilis????????????????????..129  
6.3.1.2 Galendromus occidentalis?????????????????....?.129   
6.3.1.3 Neoseiulus californicus????????????????????..129  
6.3.2 Experiment 2. Prey Stage Preference......................................................................130 
6.3.3 Experiment 3. Effect of Starvation??.................................................................131  
6.4 Discussion........................................................................................................................131 
6.5 References........................................................................................................................137 
Chapter 7 Evaluation of Small Scale Release of the Predaceous Mites, Phytoseiulus persimilis 
and Galendromus occidentalis for Biological Control of Citrus Red Mites, Panonychus 
citri in Alabama Satsuma Orchards ???????......?? ???????. ?..150  
7.1 Introduction......................................................................................................................150 
 
 ix 
7.2 Materials and Methods ...................................................................................................153 
7.2.1 Study Sites..............................................................................................................153 
7.2.2 Predacious Mites.....................................................................................................153 
7.2.3 General Procedure for Predacious Mites Release and Samplings????..??154   
7.2.4 Experiment 1. G. occidentalis Releases in 2007.....................................................155 
7.2.5 Experiment 2. G. occidentalis Releases in 2008???????...?.????.156  
7.2.6 Experiment 3. P. persimilis Releases in 2008????????.?...????.156  
7.2.7 Experiment 4. Releases of G. occidentalis or P. persimilis in 2008???. ?......157  
7.2.8 Statistical Analysis..................................................................................................157 
7.3 Results .............................................................................................................................158 
7.3.1 Experiment 1. G. occidentalis Releases in 2007.....................................................158 
7.3.2 Experiment 2. G. occidentalis Releases in 2008??????? ...?.????.158  
7.3.3 Experiment 3. P. persimilis Releases in 2008????????.?...????.159  
7.3.4 Experiment 4. Releases of G. occidentalis or P. persimilis in 2008.......................159 
7.3.5 Population Dynamics of P. citri on Experimental Trees??????????16 0 
7.4 Discussion........................................................................................................................160 
7.5 Acknowledgements..........................................................................................................165 
7.6 References........................................................................................................................166
 
 x 
 
 
 
 
 
 
List of Tables   
 
 
Chapter 2 
Table 1.  Mean (? SE) number of L. zonatus of different stages recorded on various fruit 
species 24h after releasing in multiple choice tests???????????......40   
Table 2.  Life history parameters of L. zonatus, (eggs and nymphs) on satsuma mandarin 
fruits under laboratory condition???????????????????..41  
Table 3.  Life history parameters of L. zonatus on satsuma under laboratory 
conditions????????????????????????????..42  
Chapter 3 
Table 1.  Two-way ANOVA testing for effects of insect density, feeding duration, and 
interactions of both factors on fruit weight loss (FWL), percent damaged sections 
(PDS), and soluble solid contents (SSC) of satsuma fruits infested by Leptoglossus 
zonatus in laboratory tests.??????...??????????????.?60  
Table 2.  Effect of insect density on fruit weight loss (FWL), percent damaged section   
(PDS), and soluble solid contents (SSC) of satsuma fruits infested by Leptoglossus 
zonatus for 14 days in laboratory tests????????????????......61  
Table 3. Evaluation of damage to satsuma fruit infested by Leptoglossus zonatus females in 
two south Alabama satsuma orchards in 2008?????????????......62  
Table 4. Evaluation of damage to satsuma fruit infested by Leptoglossus zonatus males in 
two south Alabama satsuma orchards in 2008??????????????..63  
 
 xi 
Chapter 4  
Table1. Relative abundance of P. citrella immature stages in Alabama satsuma  
             orchards surveyed during 2006 -2007???????????????....?.....88  
Table 2. Captures of Phyllocnistis citrella males in delta and wing traps baited with    
synthetic sex pheromone lures during 2006 and 2007 in two Alabama satsuma 
orchards.???????????????????...???.??????89  
Table 3. Beneficial arthropods observed in association with P. citrella in six Alabama 
satsuma citrus orchards in 2006???????????????????....90  
Table 4.  Abundance of key beneficial arthropods observed in association with P. citrella in 
six Alabama satsuma citrus orchards in 2006?????????????.?..92  
Chapter 5 
Table 1.  Mortality of Phyllocnistis citrella cohorts from first stage larvae to adult emergence 
on unprotected (control) branches of satsuma trees and branches protected with 
different exclusion techniques (sticky barrier or cage) during different stages of 
development at Coker orchard in 2007????????????????...116    
Table 2. Mortality of Phyllocnistis citrella cohorts from first stage larvae to adult emergence 
on unprotected (control) branches of satsuma trees and branches protected with 
different exclusion techniques (sticky barrier or cage) during different stages of 
development at GCREC orchard in 2007???????????????...118    
Table 3.  Mortality of Phyllocnistis citrella cohorts from first stage larvae to adult emergence 
on unprotected (control) branches of satsuma trees and branches protected with 
different exclusion techniques (sticky barrier or cage) during different stages of 
development at Coker orchard in 2008????????????????... 119 
 
 xii 
Table 4.  Mortality of Phyllocnistis citrella cohorts from first instar larvae to adult emergence 
on unprotected (control) branches of satsuma trees and branches protected with 
different exclusion techniques (sticky barrier or cage) during different stages of 
development at GCREC orchard in 2008???????????????... 120   
Chapter 6 
Table1. Predation capacity of three species of predacious mites at different densities of the 
prey, P. citri under laboratory conditions?..????? ?????????.143  
Table2.  Prey-stage preference of three species of predacious mites when offered eggs and 
nymphs of P. citri at different ratios for 24 h under laboratory conditions???144  
Table 3. Effect of starvation on the predation capability of three species of predacious mites 
when offered a 1:1 ratio of P. citri eggs and nymphs???????????.145  
Chapter 7 
Table 1. Effect of G. occidentalis releases on density of P. citri on satsuma trees (experiment 
1, spring 2007)??????????????????????????173  
Table 2. Effect of G. occidentalis releases on density of P. citri on satsuma trees (experiment 
2, spring 2008)?????????????????????????....174  
Table 3. Effect of P. persimilis releases on density of P. citri on satsuma trees (experiment    
3, spring 2008)???????????? ??????????????175  
Table 4. Effect of G. occidentalis or P. persimilis releases on density of P. citri on satsuma 
trees (experiment 4, spring 2008)??????????????????...176  
 xiii 
 
 
 
 
 
 
List of Figures   
 
 
Chapter 2 
Figure 1. Mean (? SE) of L. zonatus nymphs (A) and adult females (B) recorded on different 
fruit species (all ripened) at different periods after they were released in a multiple 
choice bioassay. Ten individuals were released per test and replicated eight times. 
Means for the same period having different letter in common are significantly 
different (P < 0.05, Tukey - Kramer HSD test)?????  ????????...43  
Figure 2. Mean (? SE) of L. zonatus nymphs (A) and adult females (B) recorded on 
unripened tomato (green) versus ripened satsuma, peach and lemon at different 
periods after they were released in a multiple choice bioassay. Ten individuals were 
released per test and replicated eight times. Means for the same period having 
different letter in common are significantly different (P < 0.05, Tukey-Kramer HSD 
test). ??????????????????????????????..44  
Chapter 3 
Figure 1.  Damage symptoms on satsuma fruits infested by L. zonatus. A (external damage): 
control fruit showing no damage (A1), ?green islands? on outer rind of infested fruit 
(A2); B (necrosis): control fruit showing no necrosis (B1), necrosis on inner rind of 
infested fruit (B2); C (internal damage): control fruit showing no internal damage 
(C1), infested fruit showing dried-out juice sacs (C2). ???????????64   
 xiv 
Figure 2.  Effects of insect density and feeding duration on fruit weight loss (% FWL) of 
satsuma fruits infested by Leptoglossus zonatus in laboratory tests in 2008. A: 
Females, B: Males; C: Nymphs. Figure shows mean (? SE) number of FWL (%) 
over different feeding periods???????????..?????????.65  
Figure 3.  Effects of insect density and feeding duration on (A) percent damaged sections 
(PDS) and (B) soluble solids content (SSC) of satsuma fruits infested by L. zonatus 
females in laboratory tests in 2008?????????......????????.66  
Figure 4.  Negative correlation between percent damaged sections (PDS) and soluble solids 
content (SSC) of satsuma fruits infested by L. zonatus females at different densities. 
A: Data from the laboratory test in 2008, B: Data from the field trial in Coker 
orchard in 2008.........................................................................................................67  
Chapter 4 
Figure 1. Seasonal abundance of Phyllocnistis citrella immatures in the Alabama satsuma 
orchards surveyed during 2006 (A) and 2007 (B). Figures shows mean (  SE) 
number of immatures per flush (~ 10 leaves) per sampling date (n = 25 flushes). 
Immatures = larvae + prepupae + pupae. GCREC = Gulf Coast Research and 
Extension Center????????????????????????.....93  
Figure 2.  Associating seasonal abundance of Phyllocnistis citrella immatures with 
abundance of leaf flushes in two Alabama satsuma orchards (A: 2006 data for 
Ladnier, B: 2007 data for Brantley). No. leaves = number of leaves per flush,      
No. P. citrella = total number of P. citrella immatures (larvae + prepupae + pupae) 
counted; % infested leaves = % leaves with at least one P. citrella (calculated by 
dividing number of infested leaves per flush by the total number of leaves per 
 
 xv 
flush)??????????????????????????????94  
Figure 3.  Seasonal phenology of Phyllocnistis citrella moths in two Alabama satsuma 
orchards during 2006 and 2007. Figure shows mean (? SE) weekly (A: 2006) or 
biweekly (B: 2007) captures of males per Delta trap baited with the synthetic sex 
pheromone lure. GCREC = Gulf Coast Research and Extension Center????.95  
Chapter 5 
Figure 1.  Survivorship curve of P. citrella immature stage over whole development periods 
in natural and protected condition in the two orchards in 2007. Figure showed mean 
(? SE) numbers of P. citrella. A: Coker and B: GCREC (Gulf Coast Research and 
Extension Center, Fairhope, AL)?????????.?????????...12 1 
Figure 2.  Survivorship curve of P. citrella immature stage over whole development periods 
in natural and protected condition in the two orchards in 2008. Figure showed mean 
(? SE) numbers of P. citrella. A: Coker and B: GCREC (Gulf Coast Research and 
Extension Center, Fairhope, AL)????????? ????????..?.12 2  
Chapter 6 
Figure 1.  Relationship between numbers of P. citri preyed on by a female of P. persimilis 
(A), G. occidentalis (B), or N. californicus (C) and the density of P. citri (nymphs) 
provided per day. For all three species, the data followed the type II convex 
functional response model in which the number of prey consumed increased with 
prey availability but began to decrease when a maximum point was reached.?.  146 
Figure 2.  Predation potential of three phytoseiid species on P. citri. Figure shows mean (? 
SE) number of eggs, nymphs, and total number of prey killed by P. persimilis, G. 
occidentalis, and N. californicus in 24 h.????????.???????..147  
 
 xvi 
Figure 3.  Prey-stage preferences of female P. persimilis (A), G. occidentalis (B), and N. 
californicus (C) when provided varying ratios (1:1, 1:2 or 2:1) of eggs and nymphs 
of P. citri. Figure shows mean (? SE) preference index (?)..???????.? 148 
Figure 4.  Effect of starvation on the prey-stage preferences of three species of female P. 
persimilis (A), G. occidentalis (B), and N. californicus (C) when provided varying 
starvation level of 0, 24 h, and 48 h at a ratio (1:1) of eggs and nymphs of P. citri. 
Figure shows mean (? SE) preference index (?)??????????.??...149  
Chapter 7 
Figure 1. Population dynamics of P. citri on control trees and trees on which predacious 
mites were released in spring 2008. (A): Data from experiment 2 for G. occidentalis 
releases; (B): Data from experiment 3 for P. persimilis releases. G50-2: two 
releases of P. persimilis at the rate of 50/tree; G100-1: one release at the rate of 
100/tree; G200-1: one release at the rate of 200/tree; G200-2: two releases at the 
rate of 200/tree; Control: no releases. Total number of P. citri = eggs + motiles 
(nymphs and adults). Arrows indicate predacious mite release dates?????177  
 
 
 
 1 
 
 
 
 
 
 
CHAPTER 1  
INTRODUCTION AND LITERATURE REVIEW 
 
INTRODUCTION 
Satsuma Mandarin Production 
 Citrus spp. (Sapindales: Rutaceae) is one of the most widely grown nutrient-rich fruit 
crops around the world, and also a major fruit crop with an average annual market value of about 
$10 billion in the United States.  In the United States (U.S.), citrus is produced mainly in Florida, 
California, and Texas. However, the last two decades have seen an increase in citrus acreage in 
southern Alabama and other parts of the Gulf Coast region of the U.S. (Campbell et al. 2004, 
Fadamiro et al. 2007, 2008)  
Satsuma mandarin (Citrus unshiu Marcovitch), a seedless and easy-peeling fruit, is one of 
the most appealing citrus species. Native to Asia, satsuma has been grown for over a century in 
the Gulf Coast of the U.S. (English and Turnipseed 1940). However, growth and expansion of 
the industry have been hampered by periodic freezes (Winberg 1948, Campbell et al. 2004). 
Since the early 1990s, satsuma production has significantly increased, particularly in the two 
coastal counties (Mobile and Baldwin) in southern Alabama. Renewed interest in satsuma 
production by the region growers is fueled by improved production and tree protection methods, 
as well as strong industry and state support (Campbell et al. 2004).  
 
 2 
Currently, satsuma mandarin is a minor citrus crop in the U.S. produced commercially in 
the states of Alabama, California, Florida, Louisiana, Mississippi and Texas. California is the top 
satsuma producing state with approximately 3,000 acres, and Louisiana is second with about 300 
acres (Boudreaux and Vaughn, 2006). Alabama is the third leading state for satsuma production 
in the U.S. with over 100 acres. Based on acreage of mature trees and production studies, 
Alabama?s current annual level of satsuma production is 1.4 to 1.8 million pounds with a market 
value of approximately $1 million per year (Fadamiro et al. 2007). The predominant variety of 
satsuma grown in Alabama is ?Owari?. Other varieties are also planted including ?Armstrong 
Early?, ?Brown?s Select?, and ?Early St. Ann? (Ebel et al. 2004, Fadamiro et al. 2007). In 
Alabama, satsuma is marketed entirely for fresh consumption. The public school system in 
Alabama is also a major market for satsumas; about one-third of the local Satsuma mandarin 
crop has been sold annually to the Alabama public school system since 2003. Growers also 
market fruit to produce brokers and to fresh produce market vendors.  
.  
Major Arthropod Pests of Satsuma Mandarin in Alabama 
As in other citrus growing regions, one of the key factors blocking the expansion of the 
budding Alabama citrus industry is insect and mite pest damages and management. The first 
published studies on life history and control of pests of Alabama satsuma mandarin were 
conducted in the early part of the last century (Dozier 1924, English and Turnipseed 1933, 1940). 
These studies identified several arthropods as key pests of the crop in Alabama including citrus 
whitefly, Dialeurodes citri (Ashmead) (Hemiptera: Aleyrodidae), purple scale, Lepidosaphes 
beckii (Newman) (Hemiptera: Diaspididae), Glover scale, L. gloveri (Packard) (Hemiptera: 
Diaspididae), citrus red mite, Panonychus citri (McGregor) (Acari: Tetranychidae); and citrus 
 
 3 
rust mite, Phyllocoptruta oleivora (Ashmead) (Acari: Eriophyidae). After these early 
publications, commercial production of satsuma mandarins in Alabama was largely abandoned 
due to severe freezes. The recent expansion of commercial satsuma production in the state has 
called for a renewed attention to pest management.  
A recent comprehensive survey of Alabama satsuma orchards identified at least twenty 
eight arthropod pest species (Fadamiro et al. 2007, 2008). These included 24 insect species from 
five orders: Hemiptera (18 species), Hymenoptera (1 species), Lepidoptera (2 species), 
Orthoptera (2 species), and Thysanoptera (1 species). In addition, four species of pest mites 
(Acari) were identified (Fadamiro et al. 2007, 2008). Seven species were identified as major 
pests of satsuma in Alabama including citrus whitefly, Dialeurodes citri (Ashmead) (Hemiptera: 
Aleyrodidae), purple scale, Lepidosaphes beckii (Newman) (Hemiptera: Diaspididae), Glover 
scale, L. gloveri (Packard) (Hemiptera: Diaspididae), citrus red mite, Panonychus citri 
(McGregor) (Acari: Tetranychidae); and citrus rust mite, Phyllocoptruta oleivora (Ashmead) 
(Acari: Eriophyidae); citrus leafminer, Phyllocnistis citrella, Stainton (Lepidoptera: 
Gracillariidae), and leaffooted bug, Leptoglossus zonatus. (Hemiptera: Coreidae). The minor 
(secondary) pests include citrus mealybug, Planoccocus citri (Risso), black citrus aphid, 
Toxoptera auranti (Fonscolombe), green citrus aphid, Aphis spiraecola Patch, cotton aphid, 
Aphis gossypii Glover, flower thrips, Frankliniella bispinosa Morgan, Brown stink bug, 
Euschistus servus Say, green stink bug, Acrosternum hilare Say, and red imported fire ant, 
Solenopsis invicta Buren, Orangedog, Papilio cresphontes (Cramer),  Citrus snow scale, Unaspis 
citri (Comstock), Cottony cushion scale, Icerya purchasi Maskell (Fadamiro et al. 2007, 2008). 
Most of the arthropod pests identified by Fadamiro et al. (2007, 2008) had first been identified in 
Alabama in the early 1900s (English and Turnipseed 1933, 1940). However, a few arthropod 
 
 4 
species which have recently become pests of satsuma in Alabama were also reported by 
Fadamiro et al. (2007, 2008) including leaffooted bugs, Leptoglossus spp., and citrus leafminer, 
P. citrella. In general, the arthropod fauna of satsuma mandarin in Alabama is similar to the pest 
fauna of citrus in Florida, Louisiana, and Texas, although some recently-introduced pests of 
citrus in Florida such as brown citrus aphid, Toxoptera citricida (Kirkaldy), and citrus root 
weevil, Diaprepes abbreviatus (L.) (Aerts and Mossler 2006), were not recorded in the Alabama 
orchards in the surveys by Fadamiro et al (2008). However, the Asian citrus psyllid, Diaphorina 
citri Kuwayama was first detected in Alabama in August 2008 on citrus at two locations in 
Baldwin County. D. citri was first found in the U.S. in Florida in 1998 and has also been recently 
discovered in Louisiana, Texas, Georgia, Mississippi, South Carolina, and California. This insect 
is the most efficient vector of citrus greening disease bacterium, Candidatus Liberobacter 
asiaticum. Citrus greening (also called Huanglongbing) is one of the most serious diseases of 
citrus causing reduced production and eventual tree death. Fortunately, so far none of the D. citri 
-infested citrus trees and insect samples tested positive for greening. Nevertheless, Baldwin 
County has since been placed under quarantine by the State Department of Agriculture. The 
Alabama Citrus Greening/Vector working group was recently established to formulate and 
coordinate a plan for managing the pest/disease complex in the state, including quarantine, 
education, research and management strategies. The following sections provide a detailed review 
of the three key pests of satsuma, which were the focus of this dissertation. 
 
Leaffooted bugs, Leptoglossus zonatus  
Leaffooted bugs, Leptoglossus spp. (Hemiptera: Coreidae), are polyphagous pests of 
various field, vegetable and fruit crops in the United States (Hussey 1953, Allen 1969, Hall and 
 
 5 
Teetes 1982, Johnson and Allain 1998, Schaefer and Panizzi 2000, Henne et al. 2003, Buss et al. 
2005, Fadamiro et al. 2008). Recently, two species, L. zonatus (Dallas) and L. phyllopus (L.), 
were identified as key direct pests of satsuma in the Gulf Coast region with the potential for 
major economic losses (Henne et al. 2003, Fadamiro et al. 2008). However, the predominant 
species in Alabama is L. zonatus. 
L. zonatus presents two large whitish-yellow spots on the anterior portion of the 
pronotum, and a zigzag white band across the hemelytra that easily distinguish it from related 
species (Hussey 1953, Allen 1969). L. zonatus is a pest of many economically important crops 
such as maize (Schaefer and Panizzi 2000), sorghum (Matrangolo et al. 1994), cotton (Jackson et 
al. 1995), citrus (Albrigo and Bullock 1977, Henne et al. 2003, Fadamiro et al. 2007, 2008), 
pecan (Ebeling 1950), and physic nut (Grimm and Guharay1998). Leptoglossus spp. feed 
primarily on fruits and developing seeds, occasionally feeding on vegetative tissues for moisture 
uptake (Schaefer and Mitchell 1983). Several cover-crop plants, e.g., watermelons, citrons, 
velvet beans mentioned by Griffiths and Thompson (1957), and thistles can also serve as 
important host plants for nymph development (Hubbard (1858).  In Alabama, L. zonatus 
typically move from crop fields (e.g., cotton, tomatoes, watermelons, etc) into adjacent satsuma 
orchards in the fall (September) when the fruit start to ripen. 
L. zonatus immatures pass through five nymphal stages, which are found in the same 
environment as the adults. Several aspects of the biology of L. zonatus documented in several 
crops showed various life history parameters on different hosts (Panizzi 1989, Jackson et al. 
1995, Grimm and Somarriba 1999). For example, development time on a meridic diet (Jackson 
et al. 1995) was different from that on physic nut (Grimm 1999). However, little is known about 
the development or life history of L. zonatus on satsuma mandarin. Fruit feeding by leaffooted 
 
 6 
bug immatures and adults can result in premature color break and fruit drop, and render the fruit 
unmarketable (Ebeling 1950, Griffiths and Thompson 1957, Albrigo and Bullock 1977, Grimm 
1999; Henne et al. 2003). In addition, Leptoglossus spp, can transmit a yeast disease pathogen 
(Nematocera coryli) to fruit, causing dryness in the affected wedges of the fruit and producing 
bad fruit flavor (Clarke and Wilde 1970, Henne et al. 2003). The economic damage due to 
Leptoglossus spp. has been estimated in sorghum (Hall and Teetes 1982) and physic nut (Grimm 
1999). Leaffooted bugs are primarily controlled using conventional insecticides Abudulai et al. 
2001, Grimm and Guharay 1998), but only a few products (mainly pyrethroids) are presently 
labeled for leaffooted bug control in the U.S. Despite their importance as pests of many crops, 
several aspects of the biology and management of leaffooted bugs have not been studied. In 
particular, little is known about their host preference, and their development on satsuma 
mandarin has not been evaluated. Knowledge of the biology and ecology of L. zonatus is 
necessary for the development of an effective IPM program for this pest.  
 
Citrus Leafminer, Phyllocnistis citrella  
The citrus leafminer, Phyllocnistis citrella (Lepidoptera: Gracillariidae), is of southern 
Asian origin (Stainton 1856, Clausen 1931), and has now spread to all major citrus-growing 
areas around the world (Heppner 1993, Hoy and Nguyen 1997, Legaspi et al. 2001, Garcia-Mari 
2004, Diez et al. 2006). P. citrella is prevalent in most Alabama satsuma orchards (Fadamiro et 
al. 2007, 2008), and is generally regarded as an important pest in young citrus nurseries and top-
grafted trees (Knapp et al. 1995, Diez et al. 2006).   
The biology and ecology of P. citrella has been reported by a number of researchers in 
different countries (Clausen 1931, Ba-Angood 1978, Huang et al. 1989, Heppner 1993, Knapp et 
 
 7 
al. 1995). P. citrella goes through four developmental stages: eggs, larvae, pupae, and adults.  
Total developmental time from eggs to adult emergence was ~ 13-52 days, and had 6 -13 
generations per year depending on weather conditions (Clausen 1931, Lal 1950, Heppner 1993, 
Knapp et al. 1995). P. citrella attacks all citrus cultivars or related species within the family 
Rutaceae (Badawy 1967, Heppner 1993), including Citrus spp., grapefruit (Citrus paradisi 
Macfad.), and kumquat (Fortunella crassifolia Swingle) (Badawy 1967, Heppner 1993). Larvae 
of P. citrella feed on the leaf epidermis ingesting sap and causing chlorosis. They typically make 
serpentine tunnels called mines under the leaf cuticle, resulting in twisted and malformed leaves 
(Heppner 1993, Legaspi et al. 1999). The serpentine mines may reduce the photosynthetic 
capacity of leaves and increase their susceptibility to plant pathogens, such as citrus canker 
bacterium, Xanthomonas axopodis pv. citri (Sohi and Sandhu 1968, Cook 1988, Gottwald et al. 
1997, 2002). Virtually all citrus cultivars are susceptible to P. citrella (Heppner 1993, Legaspi 
and French 1999) and heavy infestations can result in significant yield loss (Pe?a et al. 2000).  
The seasonal dynamics of P. citrella and associated natural enemies have been 
documented in many regions (Chen et al. 1989, Pena et al. 1994, Pena 1998, Urbaneja et al. 2000, 
Legaspi et al. 2001, Diez et al. 2006, Lapointe and Leal 2007). The species is multivoltine with 
number of generations per year ranging from six in Japan (Clausen 1931) to 13 in Northcentral 
India (Lal 1950). In Florida, P. citrella is present throughout the season, but peak flight activity 
occurs from late March through early October, coinciding with flushes of vegetative growth 
(Lapointe and Leal 2007). The female-produced P. citrella sex pheromone was first partially 
identified by Ando et al. (1985). Recent studies have identified the full composition of this sex 
pheromone as consisting of three active components: (Z, Z, E)-7, 11, 13-hexadecatrienal, (Z, Z)-7, 
11-hexadecadienal, and (Z)-7-hexadecenal (Leal et al. 2006, Moreira et al. 2006). Follow-up 
 
 8 
field tests demonstrated strong attraction of P. citrella males to traps baited with a binary or 
tertiary lure of the active sex pheromone components (Mafi et al. 2005, Lapointe et al. 2006, 
Leal et al. 2006, Moreira et al. 2006, Lapointe and Leal 2007). A recent study also reported on 
the utility of traps baited with a commercially produced synthetic sex pheromone lure of P. 
citrella for monitoring the pest in Florida (Stelinski and Rogers 2008).  
P. citrella is usually controlled using insecticides but chemical control is rarely effective 
because the larvae are protected from insecticide by leaf cuticles (Legaspi et al. 2001). Thus, 
biological control is generally regarded as a sustainable management solution for P. citrella 
(Knapp et al. 1995, Hoy and Nguyen 1997, Legaspi et al. 2001, Garcia-Mari et al. 2004). Several 
predatory arthropods are known to feed on P. citrella, including lacewing larvae, ants, and 
hunting spiders (Argov et al. 1996, Pomerinke 1999, Amalin et al. 2001, Hoy et al. 2007, Xiao et 
al. 2007), and many studies have identified predation as the most important natural mortality 
factor acting on P. citrella in many parts of the world (Chen et al. 1989, Amalin et al. 1996, 2001, 
2002, Hoy et al. 2007, Xiao et al. 2007). In addition, many species of parasitoids have been 
reared from P. citrella worldwide (Hoy and Nguyen 1997, Schauff 1998, Legaspi et al. 1999), 
but indigenous parasitoids were found to provide only minimal levels of parasitism in Florida 
(Pena et al. 1996). Consequently, Ageniaspis citricola Logvinovskaya (Hymenoptera: 
Encyrtidae), an exotic specialist endo-parasitoid of P. citrella larvae, was recently introduced for 
classical biological control of the pest in Florida and Texas (Hoy and Nguyen 1997, Pomerinke 
and Stansly 1998). 
P. citrella is a key pest of satsuma mandarin in Alabama (Fadamiro et al. 2007, 2008) but 
little is known about its seasonal phenology, associated natural enemies, and key mortality 
factors in the Gulf Coast region.  Because pest phenology and response to pheromone traps may 
 
 9 
vary from region to region due to several factors including variations in climate and crop 
varieties (Fadamiro 2004a, b), data obtained from other citrus growing regions may not provide 
accurate information on the abundance and seasonal activity of P. citrella in southern Alabama. 
Several aspects of the biology and ecology of P. citrella in Alabama were studied in this 
dissertation. 
 
Citrus red mite, Panonychus citri   
Citrus red mite, Panonychus citri (McGregor) (Acari: Tetranychidae), was first identified 
on citrus in Florida in 1885 (English and Turnipseed 1940) and is now an important pest of citrus 
in many parts of the world (Gotoh and Kubota 1997, Jamieson et al. 2005, Childers et al. 2007). 
Like most spider mites, P. citri goes through five stages: egg, larvae, protonymph, deutonymph 
and adult (Krantz 1978). The oval eggs are laid on the leaf surface by females and the immatures 
resemble the adults (Krantz 1978). The female is larger than the male (Childers et al. 2007). The 
developmental time from egg to adult is ~12 days and adults live for ~10-23 days depending on 
environmental conditions (24 - 26 ?F and RH 50 -70 %) (Childers and Fasulo 2009).  Like other 
spider mites, P. citri adults deposit fine webbing to maintain beneficial environmental conditions 
on the leaf surface, as well as for protection (Krantz 1978). Adults and immatures feed primarily 
on leaves and fruits, but leaf damage is usually more common (Childers et al. 2007), producing 
stippling damage, and severe infestations can result in premature leaf drop, decreased plant vigor 
(Kranz 1977). 
P. citri is a major pest of satsuma mandarin in Alabama (English and Turnipseed 1940, 
Fadamiro et al. 2007, 2008). The results of recent field surveys identified P. citri as one of the 
most numerically abundant pests in Alabama satsuma orchards (Fadamiro et al. 2007, 2008). 
 
 10 
Two to three generations per year were recorded in southern Alabama and the pest was most 
abundant in the spring (Fadamiro et al. 2008), when population densities were typically greater 
than the economic threshold of five motiles per leaf, proposed by Childers et al. (2007). The 
results also showed that P. citri was more abundant in the exterior canopy than in the interior 
canopy, and infestations were more severe in conventionally sprayed orchards than in unsprayed 
orchards (Fadamiro et al. 2008).  
Traditionally, control of P. citri on citrus has been accomplished through the use of 
conventional acaricides (Childers 1994). However, there are some reports of documented or 
suspected cases of resistance of some phytophagous mites to acaricides (Omoto et al. 1995, 
Bergh et al. 1999). Furthermore, indiscriminate use of broad spectrum pesticides may induce or 
exacerbate populations of phytophagous mites by disrupting the activity of predatory mites and 
other natural enemies (Welty 1995, Antonelli et al. 1997, Jamieson et al. 2005).  
Many authors have reported on the biological control of phytophagous mites of citrus and 
other fruit crops with predacious mites, in particular those belonging to the families Phytoseiidae 
and Stigmaeidae (Childers et al. 1975, McMurtry 1983, Childers 1994, Wood et al. 1994, 
Jamieson et al. 2005). Like their spider mite prey, predacious mites also have five stages: egg, 
larva, protonymph, deutonymph and adult (Krantz 1978). Most phytoseiids are ovoid and often 
move much faster than spider mites. Phytoseiid mites have been reported to use kairomones from 
host plants to locate their prey (Sabelis 1985). Predacious mites often overexploit spider mites 
patches and are therefore forced to move to a new patch or die of starvation (Croft and Jung 
2001).  
Biological control with predacious mites may present an alternative to chemical control 
of P. citri in Alabama. A recent survey of the predacious mite fauna of satsuma mandarin in 
 
 11 
Alabama identified a total of 29 species from nine families, including 18 species in the family 
Phytoseiidae and one species in the family Stigmaeidae (Fadamiro et al. 2009). The dominant 
predacious mite species recorded were Typhlodromalus peregrinus Muma and Proprioseiopsis 
mexicanus (Garman) (Phytoseiidae), and Agistemus floridanus Gonzalez (Stigmaeidae) 
(Fadamiro et al. 2009). However, these predacious mites were recorded in the orchards at 
densities too low for effective suppression of P. citri (Fadamiro et al. 2009). Furthermore, recent 
attempts to mass rear T. peregrinus and other indigenous predacious mites for augmentative 
releases against P. citri have not been successful (unpublished data). This prompted our interest 
in the evaluation of some commercially available phytoseiids, such as, Phytoseiulus persimilis 
Athias Henriot, Galendromus occidentalis (Nesbitt), and Neoseiulus californicus (McGregor) as 
potential biological control agents of P. citri. Predacious mites in the family Phytoseiidae can be 
classified into four categories based on their feeding habits and related biological and 
morphological traits (McMurtry and Croft 1997). Type I phytoseiids (e.g., Phytoseiulus spp.) are 
specialized predators of spider mites, Tetranychus spp. Type II phytoseiids are selective 
predators of Tetranychus species. Examples include Galendromus spp. and some Neoseiulus spp. 
Type III phytoseiids (e.g., Amblyseius spp.) are generalist predators, while Type IV phytoseiids 
(e.g., Euseius spp) are specialized pollen feeders/generalist predators.  
In this dissertation, the above commercially available predacious mite species were 
evaluated in the laboratory and field to determine their potential as biological control agents of P. 
citri. 
 
 
 
 
 12 
JUSTIFICATION  
Historically, pest control in citrus and other fruit crops has largely been accomplished 
by multiple applications of conventional, broad-spectrum pesticides. In many cases, the 
pesticides have been applied on a calendar basis. This intensive chemical use has resulted in 
many drawbacks, including increasing concern over food safety, environmental pollution, pest 
resistances, and reducing activities of beneficial insects (Thistlewood 1991) The passage of the 
Federal Food Quality Protection Act (FQPA) of 1996 called for a significant reduction in 
pesticide use on food crops, and has resulted in the restriction or loss of many insecticides and 
miticides popularly used to control citrus pests.  For instance, use of Azinphos-methyl (Guthion) 
Carbaryl (Sevin), and AgriMek (Abamectin), is now restricted on many fruit crops 
Integrated pest management (IPM) programs based on pest sampling, biological control, 
and use of reduced-risk and selective pesticides have been developed in major citrus producing 
states or regions (Childers et al. 2007), but not in Alabama. In the absence of local scientific data 
on efficacy and suitability of alternative strategies, citrus growers in Alabama have been 
reluctant to adopt IPM. In southern Alabama, leaffooted bug, L. zonatus, citrus leafminer, P. 
citrella, and citrus red mite, P. citri are three pests with great concern to satsuma growers 
(Fadamiro et al. 2007, 2008).. However, little is known about several aspects of their biology and 
ecology. Knowledge of the biology and ecology of the pests is central to the development of an 
effective satsuma IPM program in Alabama.  
 
DISSERTATION OUTLINE, GOAL AND OBJECTIVES  
The goal of this dissertation is to support the development of effective and ecologically 
friendly pest management strategies for managing key pests of satsuma in Alabama through a 
 
 13 
knowledge of their biology, ecology and natural mortality factors. The dissertation is arranged 
under three sections with the basic aim of investigating the biology, ecology and management of 
three of the key pests of satsuma in Alabama, namely L. zonatus (leaffooted bug), P. citrella 
(citrus leafminer), and P. citri (citrus red mite).  Section 1 (Chapters II and III) focused on the 
host preference, development, and feeding damage of L. zonatus. The seasonal phenology and 
natural mortality factors of P. citrella were investigated in section 2 (Chapters IV and V). The 
focus of section 3 (Chapters VI and VII) was on biological control of P. citri.  
In Chapter II, I investigated host preference and suitability of satsuma fruit as host for L. 
zonatus under laboratory conditions. Tomato was the most preferred fruit by both the nymphs 
and adults, with satsuma a distant second. Development experiments showed that satsuma is a 
suitable host which can maintain modest to high populations of the pest.  
The main objective of Chapter III was to assess the injury and amount of damage to 
satsuma fruit caused by L. zonatus nymphs and adults at various densities (0-3 individuals per 
fruit) and feeding durations (0-14 days after infestation) in laboratory and field experiments. 
Feeding by L. zonatus on satsuma produced typical damage symptoms including the presence of 
green spots and dark spots on the outer rind and the collapsing and drying out of the juice 
vesicles in the inner rind. Insect density and feeding duration had significant effects on most 
damage parameters. Fruit weight loss, percentage of damaged fruit sections, fruit abortion and 
premature fruit ripening increased with insect density and feeding duration.  
The objective of Chapter IV was to evaluate the seasonal population dynamics of P. 
citrella and associated natural enemies in Alabama satsuma orchards. Multiple overlapping 
generations with at least three distinct peaks were recorded for P. citrella immatures in Alabama, 
and population densities progressively increased from spring to summer and declined during fall. 
 
 14 
At least 21 species of beneficial arthropods were also recorded, the most prevalent being spiders 
(13 species. e.g., Hibana sp.), ants (e.g., Solenopis invicta Buren), Chrysoperla sp., Harmonia 
axyridis Pallas, and two parasitoid species (Ageniaspis citricola Logvinovskaya and Cirrospilus 
sp.). Additional studies were conducted to evaluate two popular trap types (Pherocon VI ?Delta? 
trap versus Pherocon 1C ?Wing? trap) baited with a commercially available sex pheromone lure 
for monitoring P. citrella. No significant differences were recorded in captures of male moths in 
both types of traps. The highest trap captures were recorded from August to October in 2006 and 
from June to October in 2007. The phenology of P. citrella moths generally followed the same 
pattern as that of the immatures: highest densities were recorded in summer.  
In Chapter V, I investigated the relative contributions of key beneficial arthropods 
identified in Chapter IV to natural mortality of P. citrella in Alabama using cage or sticky barrier 
exclusion methods. Overall mortality of P. citrella on unprotected (control) satsuma tree 
branches ranged from 39% to 52 % depending upon location and year. Predation was the 
dominant natural mortality factor acting on P. citrella. In unprotected (control) satsuma tree 
branches, predation accounted for ~87-96 % of all deaths. In particular, predation by spiders was 
the single most important mortality element, which accounted for ~50-70 % of all deaths. 
Predation by ants was second, accounting for ~10-19 % of all deaths. Predation by predatory 
insect larvae accounted for ~ 3-27 % of all mortalities, while parasitism contributed the least (0-
10 %) to P. citrella mortality. Predation by spiders was excluded by a cage barrier, whereas a 
sticky barrier was more effective in excluding predation by ants. 
The objective of Chapter VI was a laboratory evaluation of the potential of three 
commercially available predacious mite species (Phytoseiidae), Phytoseiulus persimilis, 
Galendromus occidentalis, and Neoseiulus californicus, as biological control agents for P. citri. 
 
 15 
All three phytoseiids species were effective in regulating P. citri density, but very few eggs were 
laid by each. Regression analysis showed a functional type II (convex) response for all three 
species: the number of prey consumed increased with prey availability up to a maximum point 
after which it slowly began to decrease. Results from experiments on prey-stage preference 
showed that all three phytoseiids preferred nymphs to eggs of P. citri. Among the three species, 
P. persimilis was a slightly more effective predator, particularly at high prey densities.  
The objective of Chapter VII was to further investigate the potential of biological control 
of citrus red mite, P. citri, by conducting field trials to further evaluate the effectiveness of 
small-scale releases of G. occidentalis or P. persimilis, for suppression of P. citri in a 
commercial satsuma orchard in southern Alabama. Both predacious mite species were selected 
based on the results of Chapter VI in which they were identified as promising biological control 
agents for P. citri. In four separate experiments, releases of each single species at different 
release rates (0-200 per tree) and frequencies (1 versus 2 releases per season) were compared on 
satsuma trees with either high initial density (i.e. > 5 motiles per leaf) or moderate initial density 
(i.e. 3-4 motiles per leaf) of P. citri. The results showed that release rate and frequency were two 
key factors determining the ability of the predacious mites to maintain effective long-term 
suppression of the prey. In general, both predacious mite species were more effective at high 
release rates than at lower rates: the best treatment was two timed releases of either species at a 
rate of 200 per tree. Furthermore, predacious mite releases were more successful at moderate 
than at high prey densities. Both species showed similar efficacy in suppressing P. citri, although 
P. persimilis was slightly more effective in at least one trial. However, the two predacious mite 
species were detected at very low population densities throughout the trial, suggesting that they 
were unable to establish and proliferate in the orchard.  
 16 
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 26 
 
 
 
 
 
 
CHAPTER 2   
HOST PREFERENFCE AND DEVELOPMENT OF THE LEAFGOOTED BUG, 
LEPTOGLOSSUS ZONATUS (HEMIPTERA: COREIDAE) ON SATSUMA MANDARIN 
(CITRUS UNSHIU) 
 
INTRODUCTION 
Leaffooted bugs, Leptoglossus spp. (Hemiptera: Coreidae), are polyphagous pests of 
various field, vegetable and fruit crops in the United States (U.S.) (Hussey 1953, Allen 1969, 
Hall et al. 1982). L. zonatus (Dallas) is an important emerging pest of a wide range of crops in 
the southern U.S., including cotton, tomato, eggplant, peaches, citrus, watermelons, corn, pecans, 
and citrus (Albrigo et al. 1977, Johnson and Allain 1998, Schaefer and Panizzi 2000, Buss et al. 
2005). This species was also recently identified as a major pest of satsuma mandarin (Citrus 
unshiu Marcovitch) in the Gulf Coast region (Henne et al. 2003, Fadamiro et al. 2007, 2008). 
Feeding on satsuma fruit by nymphs and adults of L. zonatus or the closely related L. phyllopus 
(L.) can result in premature color break and fruit drop, and render the fruit unmarketable (Henne 
et al. 2003). Damage can also reduce the soluble solids content of satsuma fruit (Y. F. X., 
unpublished data). In addition, Leptoglossus spp. can transmit yeast disease pathogen 
(Nematocera coryli) to fruit, which causes dryness in the affected wedges of the fruit and 
produces bad fruit flavor (Henne et al. 2003).  
 
 27 
Certain aspects of the life history of L. zonatus have been documented in some host crops 
(Panizzi 1989, Matrangolo and Waquil 1994, Jackson et al. 1995, Grimm and Somarriba 1999), 
showing different developmental and survival rates for L. zonatus on different crops. For 
instance, total developmental time at ~ 30 ?C from second instars to final molt on maize was ~ 
42 days (Panizzi 1989) compared to ~ 21 days on physic nut (Jatropha curcas L.) (Grimm and 
Somarriba 1999). In southern Alabama, L. zonatus adults typically move from crop fields (e.g., 
cotton, tomatoes, watermelons, etc) into adjacent/nearby fruit orchards (e.g., satsuma, peach, etc) 
in the fall when the fruits start to ripen. However, little is known about its host preference. 
Furthermore, it is not clear whether or not L. zonatus can complete its life cycle on satsuma. The 
development of an effective program for managing L. zonatus on satsuma requires knowledge of 
several aspects of its ecology and biology, including host preference and developmental biology. 
This study was conducted to determine: i) host preferences in L. zonatus by testing relative 
attraction of adults and nymphs to various fruit species in multiple choice tests, and ii) life 
history parameters of L. zonatus on satsuma fruit including developmental times, survival rates, 
female fecundity, and adult longevity. 
 
MATERIALS AND METHODS  
Rearing of L. zonatus  
Adults of L. zonatus collected from satsuma orchards in southern Alabama in the fall of 
2007 were used to start laboratory colonies, which were supplemented by adults collected from 
the field in 2008. Adults were reared in wooden sleeve cages (60 ? 40 ? 30 cm; 5 pairs per cage) 
with screened walls, on lima bean seedlings (3 potted plants per cage) supplemented with fresh 
ripened satsuma fruit (3 per cage), and water. The cages were checked daily to collect freshly 
 
 28 
laid eggs and to replace diet, as necessary. Lima bean seedlings were replaced monthly, whereas 
satsuma fruit were replaced biweekly. The rearing conditions were 25 ? 2 ?C, 50 ?10 % RH, and 
a photoperiod of 14:10 (L: D) h. 
 
Host Preference in L. zonatus  
Three separate multiple choice experiments were conducted to evaluate attraction of L. 
zonatus nymphs and adults to the fruit of the following crops: tomato (Solanum lycopersicum L. 
syn. Lycopersicon lycopersicum), satsuma mandarin (Citrus unshiu Marcovitch), peach (Prunus 
persica (L.) Batsch), kumquat (Fortunella spp.), and lemon (Citrus limon (L.) Burm. F.). In 
experiment 1, attraction of L. zonatus to the following five fruit treatments was compared: 
tomato (ripened, deep red color), satsuma (unripened, green), satsuma (ripened, orange), peach 
(ripened, yellow), and kumquat (ripened, yellow). In experiment 2, attraction of L. zonatus to the 
following four fruit treatments was compared: tomato (ripened, deep red color), satsuma (ripened, 
orange), peach (ripened, yellow), and lemon (ripened, yellow). Experiment 3 was conducted to 
test whether the attraction of L. zonatus to tomato (as observed in experiments 1 and 2) is related 
to color (deep red). For this experiment, green, unripened tomato was compared with the other 
three treatments tested in the second experiment. The tests were conducted in a wooden cage (60 
x 40 x 30 cm) with screened (nylon screen net) walls at the above conditions used for insect 
rearing. In experiment 1, the cage was divided into five sections (four corners and one middle 
section). One single fruit of each species/type (treatments) was placed in the bottom of one of the 
five sections. The experiment was replicated three times and the position of each treatment in the 
cage was determined randomly and rotated during each replication. L. zonatus of the following 
stages were tested: early instars (2nd - 3rd), late instars (4th - 5th), adult females, and adult males. 
 
 29 
For each stage, 10 individuals, previously starved for 8 h, were introduced into the center of the 
cage. The number of individuals on each fruit treatment 24 h later was counted.  
In experiments 2 and 3, the cage was divided into four corner sections. One fruit of each 
species/type (treatments) was placed in the bottom of one of the four corners of the cage (one 
corner for each treatment). Ten individuals of late stage (4th - 5th) nymphs or adult females, 
previously starved for 8 h, were introduced into the center of the cage. Each experiment was 
replicated eight times. The replication and treatment rotation schemes ensured that each 
treatment was located in each of the four corners of the cage twice. The number of individuals on 
each treatment was counted at 1, 6, 12, and 24 h after the insects were released into the cage. 
Data obtained from the three experiments were first normalized by using the square-root 
transformation (? x + 0.5). Significant differences in the number of individuals recorded on each 
treatment were established using one-way analysis of variance (ANOVA) followed by Tukey-
Kramer honestly significant difference (HSD) comparison test (P < 0.05, JMP Version 7.01, 
SAS Institute 2007).  
 
Development of L. zonatus on Satsuma Fruit  
The life history parameters of L. zonatus on satsuma fruit were determined under 
laboratory conditions [25 ? 2 ?C, 50 ? 10 % RH and a photoperiod of 14:10 (L: D) h)], to 
evaluate the suitability of the fruit as host for the pest. Since it is difficult to obtain large number 
of eggs of L. zonatus on satsuma fruit in the laboratory, egg masses freshly laid on leaves of lima 
beans seedlings were collected daily and placed (with the leaves) into a glass jar (18 long  7.5 
cm diameter), where they were allowed to hatch and molt into second instars. Each observation 
consisted of 15 - 24 eggs with five replications for a total of 94 eggs. The durations of the egg 
 
 30 
stage and the first instars (both on lima bean) were recorded. Newly molted second instars were 
reared on satsuma fruit in a glass jar (dimensions above) until they emerged as adults. The 
number and stage of the nymphs were recorded daily until adult emergence. From these data the 
following life cycle parameters were computed: development time for each stage, total 
development time from egg to adult, stage-specific survivorship (survival of the different stages), 
and cumulative survivorship (calculated by dividing the total number of individuals observed at 
each stage (survivors) by the total number of eggs observed at the beginning of the experiment = 
94 eggs). Newly emerged adults were sexed and the data used to calculate sex ratio, and then 
placed in pairs (male and female) in a glass jar (dimensions above) and reared on satsuma fruit at 
the above stated conditions. Twelve pairs of adults were observed and the following data were 
recorded: pre-oviposition time, oviposition period, longevity, female fecundity (mean number of 
eggs oviposited), and egg hatch rate (%). Data were checked for normality and then subjected to 
one-way ANOVA followed by Tukey-Kramer HSD test (P < 0.05, JMP Version 7.01, SAS 
Institute 2007), to determine significant differences in developmental time among the different 
stages, and stage-specific survivorship. Significant sexual differences in adult longevity were 
established using Student?s t test (P < 0.05, JMP Version 7.01, SAS Institute 2007). 
 
RESULTS  
Host Preference of L. zonatus  
When simultaneously presented with the different fruit species (tomato, satsuma, peach, 
and lemon) in the three multiple choice experiments, nymphs and adults of L. zonatus were 
always found in greater numbers on tomato than on the other fruit species.  In experiment 1, 
significantly higher numbers of L. zonatus of all stages were recorded on tomato compared to the 
 
 31 
other four treatments, 24 h after insects were released in the cage (early nymphs: F = 18.14; df = 
4,10; P < 0.0001; late nymphs: F = 6.55; df = 4,10; P = 0.007; females: F = 11.88; df = 4,10; P = 
0.0008; males: F = 6.10; df = 4,10; P = 0.009; Table 1). In general, no significant differences 
were recorded in the numbers of L. zonatus of all stages found on unripened versus ripened 
satsuma (Table 1).  
In experiment 2, significantly more late stage nymphs were found on tomato (ripened, red) 
than on the remaining three fruit treatments (all ripened) at 6 h (F = 16.55; df = 3, 28; P = 
0.0001), 12 h (F = 46.4; df = 3, 28; P = 0.0001), and 24 h (F = 51.8; df = 3, 28; P = 0.0001) after 
the insects were released in the cage (Fig. 1A). Similarly, greater number of females were 
recorded on tomato (ripened, red) than on the remaining treatments at 6 h (F = 7.59; df = 3, 28; P 
= 0.0007), 12 h (F = 46.01; df = 3, 28; P = 0.0001), and 24 h (F = 84.15; df = 3, 28; P = 0.0001) 
after insect release (Fig. 1B). The next most attractive fruit species was satsuma, on which was 
recorded greater numbers of nymphs and adult females compared to peach and lemon, at 24 h 
after insect release (Figs.1A and B). In experiment 3, unripened (green) tomato was tested to 
determine whether the attraction of L. zonatus to tomato documented in experiments 1 and 2 is 
related to its deep red color. The results were generally similar for both the nymphs and adult 
females. The numbers of L. zonatus found on unripened tomato were greater than the numbers 
found on each of the remaining three fruit treatments (all ripened) at 6 h (F = 5.5; df = 3, 28; P = 
0.0042), 12 h (F = 89.72; df = 3, 28; P = 0.0001), and 24 h (F = 25.9; df = 3, 28; P = 0.0001) 
after insect release (late stage nymphs, Fig. 2A), and at 6 h (F = 7.7; df = 3, 28; P = 0.0001), 12 h 
(F = 12.03; df =3, 28; P = 0.0001), and 24 h (F = 34.4; df = 3,28; P = 0.0001) after insect release 
(adult females, Fig. 2B). No significant differences were recorded at 1 h after insect release. As 
recorded in the second experiment, the numbers of nymphs and adult females recorded on 
 
 32 
satsuma, 24 h after insect release, were significantly greater than the numbers recorded on peach 
or lemon.  
 
Development of L. zonatus on Satsuma Fruit 
Basic Description of the Stages. The egg masses of L. zonatus are golden brown color, 
cylindrical and flattened, and laid in a straight line on the hosts. An egg mass contained 19.4 
eggs on average (range: 15-32 eggs).  Five nymphal stages (instars) were recorded. First stage 
nymphs typically congregate under leaves (lima bean seedlings) or on the fruit (fruit crops), and 
feeding was initiated by the second stage nymphs. The nymphs were bright red and do not have 
wings.  
          Immature Development and Survival. Under our rearing conditions an egg took 14.7 days 
to hatch. The mean developmental times for the 1st, 2nd, 3rd, 4th, and 5th instars were 3.5, 5.1, 5.9, 
9.1, and 11.4 days, respectively (Table 2). Total developmental time from eggs through the fifth 
instars was ~ 50 days (Table 1). Development time of early instars was significantly shorter 
compared to the eggs and late instars (F = 490.11; df = 5, 465; P = 0.0001).  High survivorship 
was recorded for all the stages but the fourth instars had 100% survivorship. Cumulative 
survivorship from eggs through the fifth instars was 75.6 % (Table 2). 
          Adult Survival and Reproduction. Mean pre-oviposition period was 13 days, and eggs 
were laid over ~39 days (Table 3). Mean female longevity (72.6 days) was significantly greater 
than mean male longevity (57 days) (t = 2.45; df = 1, 11; P = 0.039). Percentage female 
emergence was 49.3%, which resulted in a female: male sex ratio of 1:1.03. Mean fecundity was 
~39 eggs per female, with a hatch rate of ~ 98 % (Table 3).  
 
 
 33 
DISCUSSION 
The results from the three multiple choice experiments indicated that tomato was the 
most preferred by L. zonatus of all tested fruit species, with satsuma a distant second. L. zonatus 
and related species are polyphagous insects with a wide host range. Schaefer and Mitchell (1983) 
listed host plants in 28 families for L. phyllopus, and three families for L. zonatus. Leaffooted 
bugs are economic pests of important crops including maize (Schaefer et al. 2000, Panizzi 2004), 
sorghum (Hall et al. 1982, Matrangolo et al. 1994), cotton (Essig 1926), and have recently 
emerged as key pests of satsuma mandarin (Henne et al. 2003, Fadamiro et al. 2007, 2008) and 
other fruit and vegetable crops in the U.S. Some species of plant bugs in the family Coreidae are 
known to feed preferentially on vegetative tissues like shoots, petioles and foliages, whereas 
others preferentially attack reproductive organs (Schaefer and Mitchell 1983). L. zonatus feed 
primarily on fruits and developing seeds, but also feed occasionally on vegetative tissues for 
moisture uptake (Schaefer and Mitchell 1983). In contrast, the principal host of L. phyllopus is 
thistles, Cirsium spp (Hubbard 1885). We are not aware of any previous studies on host 
preference in leaffooted bugs, and in particular studies which compared the fruit species tested in 
our study.  
Our results are in agreement with observations on the field ecology and migration pattern 
of leaffooted bugs in the Gulf Coast region. Adults typically migrate from adjacent/nearby crop 
fields (e.g., cotton, tomatoes, watermelons, etc) and miscellaneous non crop hosts into citrus 
groves and other tree fruit crops at the time of blooming to feed on opening buds or tender shoots, 
and also later in the season when the fruits start to ripen (Ziegler and Wolfe 1961, personal 
observation). However, the movement pattern and seasonal abundance of leaffooted bugs in 
different host crops remain largely unquantified in the region.  
 
 34 
The results also showed that the relatively greater attraction of L. zonatus to tomato is not 
due mainly to color but may be mediated by chemical cues, possibly via olfaction or taste. Our 
observations also showed that tomato was almost always the first fruit species selected by both 
nymphs and adult females, further suggesting that olfaction is likely the main mechanism behind 
their preference for tomato. Host chemicals play a key role in mediating host preference in 
herbivorous insect species, most of which use host secondary chemicals (semiochemicals) as 
kairomones for host location (Duffey 1980, Brown 1984, Pasteels et al. 1988, Bowers 1990; 
Rank et al. 1998). Host biochemistry is dominant in the coevolution of plants and herbivorous 
insects, and thus in determination of diet breadth (Ehrlich and Murphy 1988). Our results suggest 
that L. zonatus may initially select a host based on chemical characteristics rather than 
morphological characteristics. The similarities in the data obtained for unripened versus ripened 
fruit of tomato and satsuma suggest that the chemical cues mediating attraction of L. zonatus to 
both fruit species also occur in unripened fruits, and that fruit maturity did not have a major 
effect on our results. Furthermore, results of an ongoing field study have demonstrated 
significant attraction and aggregation of L. zonatus on some noncrop plants in the family 
Solanaceae, a family which also includes tomato, suggesting that the chemical cues mediating 
this attraction are shared by members of this plant family. If confirmed, highly attractive noncrop 
Solanaceous plants could potentially be used as trap plants or sentinel monitoring plants for 
leaffooted bugs in fruit orchards and other economically important crops. Further studies 
including field tests are necessary to confirm our results and to identify the semiochemicals 
(kairomones) in tomato and satsuma that mediate the observed attraction of L. zonatus to both 
fruit crops. 
 
 35 
This study demonstrated that L. zonatus can survive, develop and reproduce on a diet 
consisting solely of satsuma fruit with no access to free water or supplementary food resources. 
Under laboratory conditions, Total developmental time from egg through the fifth stage was ~50 
days. High survivorship was recorded for all the immature stages ranging from 100% for fourth 
instars to 89.1% for first instars. A few studies have examined the development of L. zonatus on 
other crops or artificial diets (Panizzi 1989, Matrangolo and Waquil 1994, Jackson et al. 1995, 
Grimm and Somarriba 1999). Many of our data are similar to those reported in the above studies 
but with some key differences. For instance, the total developmental time for the nymphal stages 
recorded on satsuma in this study (36.4 days) is much longer than the 28.7 days on maize and 
31.6 days on sorghum reported by Matrangolo and Waquil (1994), or the 25.6 days recorded on 
physic nut (Grimm and Somarriba 1999). On the other hand, the 23.8 % mortality rate which we 
recorded for the nymphal stage on satsuma was much lower than the 53.8% and 55.1% recorded 
on maize and sorghum, respectively (Matrangolo and Waquil 1994), or the 59.7% mortality 
recorded on physic nut (Grimm and Somarriba 1999). The differences between our data and 
those previously reported may be related to differences in experimental conditions, and may not 
truly reflect the suitability of the different crops as hosts for L. zonatus.  
Female and male L. zonatus fed on satsuma fruit lived an average of  73 and  57 days, 
respectively, which are similar to the ~ 71 and ~ 54 days recorded on sorghum (Matrangolo and 
Waquil 1994), but shorter than the ~ 87 and 84 days recorded on physic nut, respectively 
(Grimm and Somarriba 1999). However, the ~39 eggs deposited per female on satsuma was less 
than the 96 eggs laid on sorghum (Matrangolo and Waquil 1994), the 229 eggs on physic nut 
(Grimm and Somarriba 1999), and the 348 eggs deposited on a meridic diet (Jackson et al. 1995).  
 
 36 
Altogether, these results suggest that satsuma fruit is an excellent host for development of L. 
zonatus, but not an optimal host for oviposition. The reduced oviposition on satsuma relative to 
those reported on other crops may be related to the waxy texture of the fruit, or to other yet 
unknown factors. Further studies are necessary to investigate the basis for the reduced 
oviposition by L. zonatus on satsuma and to compare oviposition and development on other fruit 
crops, such as tomato and peaches. Despite the lower number of eggs deposited on satsuma fruit 
in this study, our data, including the near 1:1 sex ratio of emerged adults recorded in this study, 
suggest that high populations of biologically fit L. zonatus can be maintained on this fruit crop.  
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 37 
REFERENCES  
Albrigo, L. G., and R. C. Bullock. 1977.   Injury to citrus fruit by leaf-footed and citrus plant 
bugs. Proc. Fla. State. Hort. Soc. 90: 63 - 67. 
Allen, R. C. 1969.  A revision of the genus Leptoglossus Guerin (Hemiptera: Coreidae).    
      Entomol. Am. 45: 35 - 140.  
Bowers, M. D. 1990.  Recycling plant natural products for insect defense. pp. 353-386. In D. L. 
Evans and J. O. Schmidt (eds.), Insect defenses: Adaptive mechanisms and strategies of prey 
and predators. State Univ. of New York, Albany. New York. 
Brown, K. S. 1984.  Adult-obtained pyrrolizidine alkaloids defend ithomiine butterflies against a 
spider predator. Nature (London). 309: 707 - 709. 
Buss, L., S. Halbert, and S. Johnson.  2005.  Leptoglossus zonatus - A new leaf-footed bug in 
Florida (Hemiptera: Coreidae). FDACS-DPI Pest Alerts. 
(http://www.doacs.state.fl.us/pi/enpp/ento/l.zonatus.html). 
Duffey, S. 1980.  Sequestration of natural plant products by insects. Ann. Rev. Entomol. 25: 447 
- 477. 
Ehrlich, P. R., and D. D. Murphy. 1988.   Plant chemistry and host range in insect herbivores. 
Ecology. 69: 908 - 909. 
Essig, E. O. 1926.   Insects of Western North America. The Macmillan Co., New York.   
Fadamiro, H. Y., M. Nesbitt, and C. Wall. 2007.  Crop profile for Satsuma mandarin in 
Alabama. (http://www.aces.edu/anr/ipm/old/crop_profiles/a_satsuma_citrus.pdf). 
Fadamiro, H. Y., Y. F Xiao, T. Hargroder, M. Nesbitt, V. Umeh, and C. Childers. 2008.  
Seasonal occurrence of key arthropod pests and associated natural enemies in Alabama 
Satsuma citrus. Environ. Entomol. 37: 555 - 567. 
 
 38 
Grimm, C., and A. Somarriba. 1999.  Suitability of physic nut, Jatropha curcas, as single host 
plant for the leaf-footed bug, Leptoglossus zonatus (Dallas), (Hemiptera: Coreidae). J. Appl. 
Entomol. 123: 347 - 350.  
Hall, D. G., and G. L Teetes. 1982.  Damage to grain sorghum green stink bug, conchuela and 
leaf-footed bug.  J. Econ. Entomol. 75: 620 - 625. 
Henne, D. C., S. J. Johnson, and W. J. Bourgeois. 2003.  Pest status of leaf-footed bugs 
(Hemiptera: Coreidae) on citrus in Louisiana. Proc. Ann. Meet. Fla. Sta. Hort. Soc. 116: 240 
- 241.  
Hubbard, H. G. 1885.   Insects affecting the orange. USDA, Division of Entomology, 
Washington, D. C. 
Hussey, R. F. 1953.   Concerning some North American Coreidae (Hemiptera). Bull. Brooklyn 
Entomol. Soc. 48: 29 - 34. 
Jackson, C. G., M. S. Tveten, and P. J. Figuli. 1995.   Development, longevity and fecundity 
of Leptoglossus zonatus on a meridic diet. Southwest. Entomol. 20: 43 - 48. 
Johnson, S. R., and L. K. Allain. 1998.  Observations on insect use of Chinese tallow (Sapium 
sebiferum (L.) Roxb.) in Louisiana and Texas. Castanea 63: 188 - 189.  
Matrangolo, W. J. R., and J. M. Waquil. 1994.  Biology of Leptoglossus zonatus (Dallas) 
 (Hemiptera: Coreidae) fed on maize and sorghum (English abstract). Ann. Soc. Entomol. Brasil. 
23: 419 - 423.  
Panizzi, A. R. 1989.   Desempenho de ninfas e adultos de Leptoglossus zonatus Dallas 
(Hemiptera: Coreidae) em diferentes alimentos (English abstract). Ann. Soc. Entomol. Brasil 
18: 375 - 389. 
 
 39 
Panizzi, A. R.  2004.  A possible territorial or recognition behavior of Leptoglossus zonatus 
(Dallas) (Hemioptera: Coreidae). Rev. Brasil Entomol. 48: 577 - 579. 
Pasteels, J. M., M. Rowell-Rahier, and M. J. Raupp. 1988.  Plant-derived defense in 
chrysomelid beetles. pp. 235 - 271. In P. Barbosa and D. K. Letourneau (eds.), Novel aspects 
of insect-plant interactions. Wiley, New York. 
Rank, N. E., A. Kopf, R. Julkunen-Tiitto, and J. Tahvanainen. 1998.   Host preference and 
larval performance of the salicylate-using leaf beetle phratora vitellinae. Ecology 79: 618-
631. 
SAS Institute, 2007.  JMP Statistics and Graphics Guide, Version 8.5. SAS Institute, Cary, NC. 
Schaefer, C.W., and P. L. Mitchell. 1983.   Food plants of the Coreoidea (Hemiptera: 
Heteroptera). Ann. Entomol. Soc. Am. 76: 591 - 615. 
Schaefer, C.W., and A. R. Panizzi. 2000.   Heteroptera of Economic Importance. CRC Press, 
Boca Raton, FL. 
Ziegler L. W., and H. S. Wolfe. 1961.   Citrus growing in Florida. University of Florida Press. 
Gainesville, Florida. 
 
 
 
 
 
 
 
 
 
 40 
Table 1.  Mean (? SE) number of L. zonatus of different stages recorded on various fruit 
species after they were released in multiple choice tests  
Stages  
 
Tomato 
(ripened) 
Satsuma 
(unripened) 
Satsuma 
(ripened) 
Kumquat 
(ripened) 
Peach  
(ripened) 
Early nymphs 2nd -3rd  6.3 ? 0.3a 0.0 ? 0.0b  1.0 ? 0.6b  0.3 ? 0.3b 0.0 ? 0.0b 
Late nymphs, 4th ? 5th   2.7 ? 0.6a 1.3 ? 0.0ab  0.6 ? 0.3ab 1.3 ? 0.0ab 0.0 ? 0.0b 
Females 2.7 ? 0.0a 0.0 ? 0.0b  0.7 ? 0.3b  0.3 ? 0.3b 0.0 ? 0.0b 
Males 1.7 ? 0.0a 0.0 ? 0.0b 1.0 ? 0.0ab 0.3 ? 0.3ab 0.3 ? 0.3b 
 Means (  SE) within the same row followed by different letters are significantly different (P < 
0.05, Tukey-Kramer HSD test). For each stage, ten individuals were released per test and 
replicated three times. 
 
 41 
Table 2.  Life history parameters of leaf footed bug, L. zonatus, (eggs and nymphs) on 
satsuma mandarin fruits under laboratory conditions 
Stage 
 
Duration (days) 
 
Total number 
observed (n) 
Stage-specific 
survivorship (%) 
Cumulative 
survivorship (%) 
Eggs 14.7 ? 0.2a 92 97.8 97.8 
1st 3.5 ? 0.1e 82 89.1 87.2 
2nd 5.1 ? 0.2d 78 95.1 82.9 
3rd 5.9 ? 0.2d 74 94.8 78.7 
4th 9.1 ? 0.2c 74 100.0 78.7 
5th 11.4 ? 0.7b 71 95.9 75.6 
Means (  SE) within the same column followed by different letters are significantly different (P 
< 0.05, Tukey-Kramer HSD test). Cumulative survivorship was calculated by dividing the total 
number of individuals observed at each stage (survivors) by the total number of eggs observed at 
the beginning of the experiment (94 eggs). 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 42 
Table 3.  Life history parameters of L. zonatus on satsuma mandarin fruit under laboratory 
conditions 
  Mean ? SE Range 
Female Pre-oviposition (days)         13.0 ? 1.1 11.0 -15.0 
 Oviposition (days)         38.5 ? 4.5 12.0 - 64.0 
 Fecundity (eggs/female)         38.8 ? 3.9  0.0 - 44.0 
 Egg hatch rate (%)         98.3 ? 1.7   91.7 - 100.0 
 Female emergence (%)         49.3 ? 2.0         41.0 - 53.0 
Female Longevity (days)         72.6 ? 8.0a   19.0 - 125.0 
Male Longevity (days)         57.0 ? 6.5b 21.0 - 93.0 
Significant sexual difference in longevity established using t - test (P < 0.05) 
 
 
 
 
 
 
 
 
 
 
 
 
 43 
Figure 1. Mean (? SE) of L. zonatus nymphs (A) and adult females (B) recorded on different 
fruit species (all ripened) at different periods after they were released in a multiple choice 
bioassay. Ten individuals were released per test and replicated eight times. Means for the same 
period having different letter in common are significantly different (P < 0.05, Tukey-Kramer 
HSD test). 
A
0
1
2
3
4
5
6
To m ato  (re d)
Satsu m a 
Pe ac h
Le m on
a
b
c
a
ab
b
b
a
b
c
c
c
B
M
ea
n (
? 
SE) n
umber o
f 
L. 
zo
na
tus
 
0
1
2
3
4
5
6
To m ato  (re d)
Satsu m a
Pe ac h
Le m on
a
b
c
c
a
b
c
c
a
b
b
b
Ho urs a ft er insect rel ea se
1
6
12
24
  
 
 
 44 
Figure 2. Mean (? SE) of L. zonatus nymphs (A) and adult females (B) recorded on unripened 
tomato versus ripened satsuma, peach and lemon at different periods after they were released in a 
multiple choice bioassay. Ten individuals were released per test and replicated eight times. 
Means for the same period having different letter in common are significantly different (P < 0.05, 
Tukey-Kramer HSD test). 
1 6 12
24
B 
Ho urs af ter inse ct rel ease
Mean 
(?
 SE) number of
 
L. z
o
natus
 
0
1
2
3
4
To m a to (g ree n )
S a ts u m a
Pea ch
Lem o n
a
b
c c
a
b
c     c
a
ab
bc
c
A
0
1
2
3
4
To m a to (g ree n )
S a ts u m a
Pea ch
Lem o n
a
b
c
c
a
b
bc
c
a
b
c
  
 
 
 45 
 
 
 
 
 
 
CHAPTER 3  
EVALUATION OF DAMAGE TO SATSUMA MANDARIN, CITRUS UNSHIU, BY THE 
LEAFFOOTED BUG, LEPTOGLOSSUS ZONATUS, (HEMIPTERA: COREIDAE)  
 
INTRODUCTION 
Satsuma mandarin, Citrus unshiu Marcovitch, has been grown for over a century in the 
Gulf Coast region of the United States (English and Turnipseed 1940). However, the last decade 
has recorded a major increase in the production of this crop in southern Alabama, fueled by 
improved production and tree protection methods, and strong industry and state support 
(Campbell et al. 2004).  Recent studies have identified several pest species capable of limiting 
the expansion of satsuma mandarin production in the region (Henne et al. 2003, Fadamiro et al. 
2007, 2008). In particular, two species of leaffooted bugs (Hemiptera: Coreidae), Leptoglossus 
zonatus (Dallas) and L. phyllopus (L.), were identified as key direct pests of satsuma with the 
potential for major economic losses (Henne et al. 2003, Fadamiro et al. 2008). In southern 
Alabama, the predominant Leptoglossus spp. on satsuma is L. zonatus (Dallas), whereas L. 
phyllopus (L.) is considered a minor pest (Fadamiro et al. 2008).  
Like many other Leptoglossus species, L. zonatus has recently risen in status as a pest of 
a wide range of crops in the southern U.S., including cotton, tomato, eggplant, sorghum, 
watermelons, corn, peaches, citrus, pecans (Albrigo and Bullock 1977, Matrangolo and Waquil 
 
 46 
1994, Johnson and Allain 1998, Schaefer and Panizzi 2000, Buss et al. 2005). In southern 
Alabama, L. zonatus adults typically move from crop fields (e.g., cotton, tomatoes, watermelons, 
etc) into adjacent or nearby satsuma orchards in the fall when the fruits start to ripen (personal 
observation). A recent study by our group suggests that satsuma fruit is a suitable host for L. 
zonatus, which can maintain modest to high populations of the pest, but tomato is preferred over 
satsuma (Xiao and Fadamiro in press). Feeding on citrus and other fruit by leaffooted bugs 
nymphs and adults can result in reduction in quality and quantity of fruits, including premature 
color break, fruit drop, and unmarketable fruit (Ebeling 1950, Griffiths and Thompson 1957, 
Albrigo and Bullock 1977, Grimm 1999, Henne et al. 2003). The insects can also transmit yeast 
disease pathogen, Nematocera coryli Peglion, to fruit, which causes dryness in the affected 
wedges of the fruit and produces bad fruit flavor (Henne et al. 2003). However, much of the 
background information necessary for effective management of L. zonatus on satsuma is missing, 
including the absence of a systematic assessment of injury and economic thresholds for L. 
zonatus on satsuma. The objectives of this study were to 1) characterize interior and exterior 
damage symptoms on satsuma fruit caused by L. zonatus; 2) determine relationship between 
insect density, feeding duration and fruit damage; and 3) propose an economic injury level for L. 
zonatus on satsuma based on the data from the above laboratory and field studies. It is hoped that 
the results will support the development of an effective integrated pest management program for 
managing L. zonatus on satsuma mandarin. 
 
MATERIALS AND METHODS 
Rearing of L. zonatus  
Adults of L. zonatus collected from satsuma orchards in south Alabama in the fall of 
 
 47 
2007 were used to start laboratory colonies. Adults were reared in wooden sleeve cages (60 ? 40 
? 30 cm; 5 pairs per cage) with screened walls, and provided lima bean seedlings (3 potted plants 
per cage), fresh mature satsuma fruits (3 per cage), fresh tomato (3 per cage) and water. The 
cages were checked daily to collect freshly laid eggs and to replace diet, as necessary. Lima bean 
seedlings were replaced monthly, whereas satsuma and tomato fruits were replaced biweekly and 
weekly, respectively. Satsuma fruits used were harvested from the field and stored in the 
refrigerator until use (fruits stored for up to 2 months). The rearing conditions were 25 ? 2 ?C, 50 
? 10% RH, and a photoperiods of 14:10 (L: D) h. 
 
Laboratory Tests  
Damage to satsuma fruit by L. zonatus was evaluated under laboratory conditions [25 ? 2 
?C, 50 ? 10% RH, and a photoperiod of 14:10 (L: D) h] during fall 2008.  Fresh, mature satsuma 
fruits were weighed and placed individually into glass jars (18 cm long ? 7.5 cm diameter, 1 fruit 
per jar). Newly-emerged L. zonatus females (2-5 days old) were introduced into each jar at one 
of four densities: 0 (control), 1, 2, or 3 per fruit. The jar was covered with screening nets. The set 
up consisted of a total of 48 jars distributed into four groups of 12 jars for each insect density. 
The fruit was weighed at 1 (12 replications), 3 (12 replications), 7 (9 replications), 10 (6 
replications), and 14 (3 replications) days after infestation. At 3, 7, 10, and 14 days after 
infestation, three fruits were selected from each group and dissected to determine interior 
damage. Fruit weight loss was calculated by comparing weight loss before and after infestation. 
The percentage of damaged sections (PDS) was calculated for each fruit by dividing the number 
of damaged sections by the total number of sections (usually 10 sections). The soluble solids 
content (SSC) of the fruit was then measured with a refractometer (Westover model RHB-
 
 48 
32ATC). Similar experiments were performed with L. zonatus males and late (4th - 5th) instars. 
Early instars (2nd - 3rd) were not tested based on a preliminary experiment which showed that 
they produce very little measurable damage to satsuma.  
 
Field Tests 
Field experiments were performed in two satsuma orchards located in Baldwin county, 
southern Alabama during 2007 (preliminary trial) and 2008: Gulf Coast Research and Extension 
Center (GCREC) orchard and Coker orchard. Both orchards were comprised mainly of satsuma 
mandarin (Owari variety) and were not conventionally sprayed during the trials. For the 2007 
trial, three premature satsuma fruits on a tree branch were randomly selected and covered with a 
cage (35 x 15 cm diameter) made from white color screening net. L. zonatus females were placed 
into the cages at an approximate density of 0 (control), 0.3, 0.7, 1.4, or 2 individuals per fruit for 
a period of 14 days. Each insect density treatment was replicated four times (4 cages) per orchard. 
On day 15, the fruits were harvested (by hand) and carefully transported (in plastic bag kept in 
ice) to the laboratory where they were weighed and inspected for external and interior damage. A 
similar methodology was used for the 2008 trial but with some modifications. Two premature 
fruits were caged (instead of 3 in 2007) and L. zonatus were placed in the cages at a density of 0, 
0.5, 1, 2 or 3 individuals per fruit. Also both females and males were tested separately in 2008. 
Each insect density treatment was also replicated four times (4 cages) per orchard. In both years, 
fruits were assessed for damage by collecting the following data: fruit color, premature fruit 
abortion (PFA), fresh fruit weight (FFW), percentage of damaged sections (PDS), and soluble 
solids contents (SSC). 
Statistical Analysis   
 
 49 
For the laboratory test, fruit weight loss was calculated as the difference in fruit weight 
before and after infestation. Fruit weight loss (FWL) data were converted to percentages and 
used in the statistical analyses. Data on damage fruit sections (PDS) and soluble solids contents 
(SSC) were also expressed as percentages. All data were first normalized by using the arcsine 
square-root transformation (? x + 0.5). Laboratory data (FWL, PDS, and SSC) were first 
analyzed with two-way analysis of variance (ANOVA) with insect density and feeding duration 
(days after infestation) as the main factors. Data collected on day 14 after infestation were further 
analyzed with one-way ANOVA to test for effect of insect density on FWL, PDS and SSC. Field 
data (premature fruit abortion or PFA, fresh fruit weight or FFW, PDS, and SSC) collected from 
each orchard on day 15 after infestation were analyzed with one-way ANOVA using insect 
density as the independent variable. In all tests, data for the different stages (females, males or 
nymphs) were analyzed separately. Significant differences between means were established by 
using Tukey-Kramer honestly significant difference (HSD) comparison test (P < 0.05, JMP 
Version 7.01, SAS Institute Inc. 2007).   
 
RESULTS 
Description of Fruit Damage  
L. zonatus pierced satsuma fruit skin with its mouthparts and sucked fluids directly from 
the underlying vesicles. A ?green island? on outer rind (Fig. 1A) and necrosis on inner rind (Fig. 
1B) often appeared on injured satsuma fruits. Dark spots were also observed on some injured 
fruits. Feeding may also result in interior fruit damage including collapsing and drying out of the 
juice vesicles (Fig. 1C). Premature ripening (yellowing) of infested fruit was also observed in the 
 
 50 
field tests. The symptoms expressed by fruit infested by L. zonatus females, males or nymphs 
were identical in appearance. 
 
Laboratory Tests  
For L. zonatus females, two-way ANOVA revealed significant effects of insect density 
and feeding duration (days after infestation) on all three parameters evaluated: fruit weight loss 
(FWL), percentage of damaged sections (PDS), and soluble solids content (SSC), as well as a 
significant density ? duration interaction on PDS and SSC (Table 1). Insect density and feeding 
duration each exerted a significant positive effect on FWL and PDS (increase fruit damage), but 
a negative effect on SSC (decline Soluble solids content). In other words, FWL and PDS 
increased with increasing insect density and feeding duration (Fig.2), while SSC decreased with 
both factors. Similar results were obtained for L. zonatus males and nymphs. The statistics for 
both sexes and the nymphs are shown in Table 1. For brevity, data obtained on day 14 of 
infestation (feeding duration of 14 days) were analyzed by one-way ANOVA and presented in 
Table 2. In general, FWL and PDS increased with increasing insect density and feeding duration, 
while SSC decreased with both factors. For instance, FWL increased from 19.6 to 24.3% and 
PDS from 3 to 71%, whereas SSC decreased from 8.4 to 6.3% at densities of 1 female/fruit and 3 
females/fruit, respectively (Table 2). The data also indicated that slightly higher fruit damage 
(FWL and PDS) were caused by L. zonatus females or males than by the nymphs (Fig. 3), 
although no statistical comparisons were made since the data for the different stages were not 
collected at the same time. 
 
 
 
 51 
Field Tests 
Feeding by L. zonatus females or males caused notable damage to satsuma fruits as 
determined by fruit color, premature fruit abortion (PFA), fresh fruit weight (FFW), percentage 
of damaged sections (PDS), and soluble solids contents (SSC). In the 2007 trial, only females 
were evaluated at both orchards. Analysis of the data collected at Coker orchard showed that 
female density had a significant positive effect on PFA (increased fruit abortion) (F = 23.3; df = 
4, 55; P < 0.0001) and PDS (increased percentage of damage) (F = 118.2; df = 4, 55; P < 
0.0001), but a negative effect on FFW ( (F = 19.6; df = 4, 55; P < 0.0001) and SSC (F = 21.8; df 
= 4, 55; P < 0.0001). Similar results were obtained at GCREC orchard: insect density had a 
significant positive effect on PFA (F = 33.4; df = 4, 55; P < 0.0001) and PDS (F = 96.4; df = 4, 
55; P = 0.0001) and a negative effect on FFW (F = 3.33; df = 4, 55; P = 0.01) and SSC (F = 
96.3; df = 4, 55; P = 0.001). Premature fruit yellowing was recorded at densities of 0.7 or more 
insects per fruit. In 2008, both females and males were evaluated at the two orchards and the 
results were generally similar to those obtained in 2007. At both orchards, female density had a 
significant positive effect on PFA and PDS, but a negative effect SSC (Table 3, Fig.4). Similar 
results were recorded also for males: increasing male density resulted in increased PFA and PDS 
and reduced SSC (Table 4). An effect of insect density on fruit color was also recorded at both 
locations and for both sexes. Premature fruit yellowing was recorded at densities of 1 or more 
insects per fruit, whereas fruits caged with ? 1 insects per fruit were still mainly green 14 days 
after infestation. However, no significant effect of insect density on fresh fruit weight (FFW) was 
recorded in 2008 (Tables 3 and 4). In general, no significant differences were recorded between 
the densities of 0 (control) and 0.5 insect per fruit for most damage parameters.  
 
 
 52 
DISCUSSION 
The symptoms of L. zonatus feeding damage on satsuma fruit recorded in this study have 
been previously reported on satsuma or other citrus species (Ebeling1950, Griffiths and 
Thompson 1957, Albrigo and Bullock 1977, Kubo and Filho 1992, Henne et al. 2003, Buss et al. 
2005). Henne et al. (2003) reported external damage to the rind of satsuma fruit infested by L. 
zonatus, in the form of green spots that remain for a time after the rind has turned orange. 
Leptoglossus spp.,have also been reported to cause similar damage to other crops (Wheeler and 
Miller 1990, Hall and Teetes 1982, Grimm 1999). The premature abortion of L. zonatus-infested 
fruit recorded in our field tests has also been documented in other citrus species (Griffiths and 
Thompson 1957; Albrigo and Bullock 1977), as well as in other crops (Ebeling 1950, Wiseman 
and McMillian 1971, DeBarr and Kormanik 1975, Bolkan et al. 1984, Grimm 1999). DeBarr and 
Kormanik (1975) reported that L. corculus (Say) nymphs fed on the cytoplasm of cells of the 
nucellar tissue of pine conelets resulting in total conelet abortion after only four days of feeding. 
Albrigo and Bullock (1977) suggested that citrus fruit drop from leaffooted bug feeding may be 
due to secondary infections by microorganisms. However, this hypothesis was not tested in the 
present study.  
In addition to direct feeding injury, L. zonatus and other related species have also been 
reported to transmit the yeast, Nematospora coryli, which causes further fruit damage including 
drying rot (Clarke and Wilde 1970, Henne et al. 2003). Henne et al. (2003) attributed the 
collapsing and drying out of the juice sacs inside the rind of satsuma fruit infested by L. zonatus 
to this yeast. Although, we did not attempt to isolate this pathogen and other microorganisms, it 
is possible that some of the symptoms of internal fruit damage observed in this study were due to 
yeast or similar pathogens.  
 
 53 
 As expected, our results showed major effects of L. zonatus density and feeding duration 
on most fruit damage parameters. In general, higher insect density and longer feeding duration 
resulted in increased fruit weight loss and percentage of damaged fruit sections (PDS), but 
reduced soluble solids content (SSC). The results were fairly consistent between laboratory and 
field experiments although minor differences were recorded. This is the first systematic 
evaluation of damage by L. zonatus or related species to satsuma or other citrus species. Similar 
results have been reported for some non-citrus crops damaged by Leptoglossus spp. (Hall and 
Teetes 1982, Grimm 1999). Hall and Teetes (1982) attributed the significant reduction in the 
yield of sorghum infested by L. phyllopus to direct seed feeding. Similarly, physic nut damaged 
by L. zonatus also showed a significant reduction in fruit weight and a slight reduction in oil 
contents (Grimm 1999).  
We were not surprised to find a negative correlation between the percentage of damaged 
sections (PDS) and soluble solids content (SSC) of satsuma fruit infested by L. zonatus. The SSC 
is commonly used as a measure of fruit juice quality (Widodo et al. 1996). Our data showed that 
a significant decline in SSC was not always recorded at densities lower than 2 individuals per 
fruit, suggesting that this parameter may not be a very reliable measure of L. zonatus feeding 
damage on satsuma. Also, SSC declined faster in the field trials than in the laboratory tests. For 
instance, the decrease in SSC was significant at the density of 1-2 individuals per fruit in the 
field trials, but a significant decrease in SSC was not recorded in the laboratory tests below the 
density of 3 individuals per fruit. These differences may be related to the hotter and unstable 
field conditions.  In general, L. zonatus females and males produced similar amounts of damage, 
which were slightly more than the damage produced by the nymphs. Grimm (1999) reported a 
 
 54 
similar finding on physic nut: L. zonatus adults of both sexes produced more damage than the 
nymphs. 
One of the objectives of this study was to extrapolate the fruit damage data to possibly 
propose an economic threshold for L. zonatus on satsuma. L. zonatus feeding affected fruit 
weight, yield, and quality, but the data showed that not all feeding resulted in significant 
measurable damage. For example, compared to uninfested (control) fruits, a significant fruit 
weight loss was not recorded below the density of 1 individual per fruit or at feeding durations of 
less than 10 days. Hall and Teetes (1982) reported a similar result on sorghum and concluded 
that two adults of L. phyllopus were generally not enough to cause significant reduction in the 
yield of green sorghum. Data from the laboratory tests showed that fruit weight loss (FWL) and 
PDS values of uninfested control fruits were not significantly different from values for fruits 
infested by L. zonatus at a density of one individual per fruit, but significantly different from 
values for fruits infested at a density of two or more individuals per fruit. In contrast, PDS and 
premature fruit abortion were significantly greater at the density of one individual per fruit 
compared to the control. We also observed premature ripening of infested fruit at densities of one 
or more individuals per fruit. Similar results on premature fruit abortion were reported for 
pistachio fruits infested by L. clypealis L., which resulted in an increase in premature fruit 
abortion from 3.8% in the control to 32.9 % in fruit clusters fed upon for 48 hours by a single L. 
clypealis (Bolkan et al. 1984). However, Grimm (1999) cautioned against reliance on premature 
fruit abortion to diagnose economic damage by bugs, since fruit abortion may be caused by other 
factors which are unrelated to insect feeding. For instance, Uma?a and Carballo (1995) reported 
that only 33-49 % of aborted macadamia nuts showed symptoms of bug feeding damage. Thus, 
our data on premature fruit abortion may not be used as a reliable indicator of economic damage 
 
 55 
by L. zonatus. In general, our field results corroborated our laboratory data. The minor 
differences between our laboratory and field results may be related to differences in 
environmental conditions, as well as the impact of other field factors. Although our data are a 
little conflicting, they suggest an economic injury level (lowest insect density that will result in 
economic damage) of 1-2 L. zonatus per satsuma fruit. At present, it is difficult to accurately 
propose an economic or action threshold (insect density that will trigger control measures to 
prevent the density from reaching economic injury level) for L. zonatus on satsuma, due to 
several factors including lack of effective monitoring and sampling techniques and the inability 
to accurately predict its movement from adjacent crop fields into satsuma orchards. Knowledge 
of these factors and the economics of managing the pest are requisite to the development of a 
reliable and practical economic threshold for L. zonatus on satsuma mandarin. 
 
 56 
ACKNOWLEDGEMENTS 
We thank Mr. Monte Nesbitt for helping with field collection of leaffooted bugs used in 
starting our laboratory colonies. We also thank our commercial satsuma grower cooperators in 
southern Alabama. Funding for this study was provided by the Alabama Agricultural Experiment 
Station and Auburn University Horticulture Line Item grants program. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 57 
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Albrigo, L. G., and R. C. Bullock. 1977.   Injury to citrus fruit by leaf-footed and citrus plant 
bugs. Proc. Fla. Sta. Hort. Soc. 90: 63 - 67. 
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bug (Hemiptera: Coreidae) and epicarp lesion of pistachio fruits. J. Econ. Entomol. 77: 1163 
- 1165. 
Buss, L., S. Halbert, and S. Johnson. 2005.   Leptoglossus zonatus - A new leaf-footed bug in 
Florida (Hemiptera: Coreidae). FDACS-DPI Pest Alerts. 
(http://www.doacs.state.fl.us/pi/enpp/ento/l.zonatus.html). 
Campbell, B. L., R. G. Nelson, R. C. Ebel, W. A. Dozier, J. L. Adrian, and B. R. Hockema. 
2004.  Fruit quality characteristics that affect consumer preferences for satsuma mandarins.  
HortScience 39: 1664 -1669. 
Clarke, R. G., and G. E. Wilde. 1970.  Association of the green stink bug and the yeast spot 
disease organism of soybeans. II. Frequency of transmission to soybeans, transmission from 
insect to insect, isolation from field population.  J. Econ. Entomol. 63: 355 - 357. 
DeBarr, G. L., and P. P. Kormanik. 1975.   Anatomical basis for conelet abortion on Pinus 
echinata following feeding by Leptoglossus corculus (Hemiptera: Coreidae). Can. Entomol. 
107: 81 - 86. 
Ebeling, W. 1950.   Subtropical Entomology (fruit pests). Lithotype Process Co., San Francisco, 
CA.  
Fadamiro, H. Y., M. Nesbitt, and C. Wall. 2007.  Crop profile for satsuma mandarin in 
Alabama. (http://www.aces.edu/anr/ipm/old/crop_profiles/a_satsuma_citrus.pdf). 
 
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Fadamiro, H. Y., Y. F. Xiao, T. Hargroder, M.  Nesbitt, V. Umeh, and C. Childers. 2008.  
Seasonal occurrence of key arthropod pests and associated natural enemies in Alabama 
satsuma citrus. Environ. Entomol. 37: 555 - 567. 
Griffiths, J. T., and W. L. Thompson. 1957.  Insects and mites found on Florida citrus. Fla. 
Agr. Exp. Sta. Bull., Gainesville, Fla. 591: 1 - 86. 
Grimm, C. 1999.  Evaluation of damage to physic nut, Jatropha curcas, by true bugs. Entomol. 
Exp. Appl. 92: 127 - 136. 
Hall, D. G., and G. L. Teetes. 1982.   Damage to grain sorghum by southern green stink bug, 
conchuela and leaf-footed bug.  J. Econ. Entomol. 75: 620 - 625. 
Henne, D. C., S. J. Johnson, and W. J. Bourgeois. 2003.  Pest status of leaf-footed bugs 
(Hemiptera: Coreidae) on citrus in Louisiana. Proc. Ann. Meet. Fla. Sta. Hort. Soc. 116: 240 
- 241.  
Johnson, S. R., and L. K. Allain. 1998.   Observations on insect use of Chinese tallow (Sapium 
sebiferum (L.) Roxb.) in Louisiana and Texas. Castanea 63: 188 - 189.  
Kubo, R. K., and A. Batista Filho. 1992.   Ocorrencia e danos provocados por Leptoglossus 
zonatus (Dallas) (Hemiptera: Coreidae) em citros. Ann. Soc. Entomol. Brasil 21: 467 - 470. 
Matrangolo, W. J. R., and J. M. Waquil. 1994.  Biology of Leptoglossus zonatus (Dallas) 
(Hemiptera: Coreidae) fed on maize and sorghum (English abstract). Ann. Soc. Entomol. 
Brasil. 23:  419 - 423.  
SAS Institute. 2007.  JMP Statistics and Graphics Guide, Version 7.0.1.  SAS Institute, Cary, 
NC. 
Schaefer, C. W., and A. R. Panizzi. 2000.   Heteroptera of Economic Importance. CRC Press, 
Boca Raton, FL. 
 
 59 
Steinbauer, M. J., G. S. Taylor, and J. L. Madden. 1997.   Comparison of damage to 
Eucalyptus caused by Amorbus obscuricornis and Gelonus tasmanicus.  Entomol. Exp. Appl. 
82: 175 - 180. 
Uma?a, M. E., and V. Carballo. 1995.   Biolog?a de Antiteuchus tripterus L. (Hemiptera: 
Pentatomidae) y su parasitoide Trissolcus radix (Johnson) (Hymenoptera: Scelionidae) en 
macadamia. Manejo Integrado Plagas (Costa Rica) in English abstract 38: 16 - 19. 
Wheeler, A. G. and G. L. Miller. 1990.  Leptoglossus fulvicornis (Heteroptera: Coreidae), a 
specialist on magnolia fruits: seasonal history, habits, and descriptions of immature stages. 
Ann. Entomol. Soc. Am. 83: 753 - 765. 
Widodo, S. E., M. Shiraishi, and S. Shiraishi. 1996.  On the interpretation of Brix value for the 
juice of acid citrus. J. Sci. Food Agri. 71: 537 - 540. 
Wiseman, B. R., and W.W. McMillian. 1971.  Damage to sorghum in south Georgia by 
Hemiptera.  J.  Ga. Entomol. Soc. 6: 229 - 236. 
 
 
 
 
 
 
 
 
 
 
 
 60 
Table 1.  Two-way ANOVA testing for effects of insect density, feeding duration, and 
interactions of both factors on fruit weight loss (FWL), percent damaged sections (PDS), 
and soluble solid contents (SSC) of satsuma fruits infested by Leptoglossus zonatus in 
laboratory tests. 
 Effect Df Females Males Nymphs 
   F P F P F P 
FWL Density 3 74.8 0.0001 37.1 0.0001 16.4 0.0001 
 Duration 4 1424.8 0.0001 738 0.0001 599 0.0001 
 Density ? Duration  12 2.8 0.0016 1.55 0.112 0.59 0.84 
PDS Density 3 166.7 0.0001 62.9 0.0001 59.7 0.0001 
 Duration 3 70.9 0.0001 39.0 0.0001 42.6 0.0001 
 Density ? Duration  9 35.1 0.0001 12.9 0.0001 14.2 0.0001 
SSC Density 3 7.4 0.0007 18.2 0.0001 25.7 0.0001 
 Duration 3 4.9 0.006 9.1 0.0002 10.8 0.0001 
 Density ? Duration  9 1.4 0.217 1.2 0.34 4.1 0.001 
Four levels of insect density (0, 1, 2 or 3 per fruit) were evaluated at feeding durations of 1, 3, 7, 
10, and 14 days after infestation for FWL and at 3, 7, 10 and 14 days after infestation for PDS 
and SSC. 
 
 
 61 
Table 2.  Effect of insect density on fruit weight loss (FWL), percent damaged section 
(PDS), and soluble solid contents (SSC) of satsuma fruits infested by Leptoglossus zonatus 
for 14 days in laboratory tests. 
Stage Density  
(insect no. per fruit) 
Mean (? SE) per fruit 
FWL (%) PDS (%) SSC (%) 
Females 0.0 15.9 ? 0.3c 0c 8.7 ? 0.2a 
 1.0   19.6 ? 1.2bc  3.0 ? 3.0c 8.4 ? 0.2a 
 2.0   21.6 ? 0.5ab 17.2 ? 0.5b 8.1 ? 0.2a 
 3.0 24.3 ? 1.3a 71.1 ? 4.4a 6.3 ? 0.7b 
   F = 13.84 
       P = 
0.0015 
F = 98.32 
P = 0.0001 
F = 2.86 
P = 0.104 
Males 0.0 15.9 ? 1.5c 0c 8.5 ? 0.1a 
 1.0   18.4 ? 0.4bc       5.8 ? 2.9bc   8.1 ? 0.1ab 
 2.0   20.9 ? 0.9ab      23.3 ? 6.7b   7.8 ? 0.1ab 
 3.0    23.3 ? 1.1a      63.1 ? 6.8a         6.7 ? 0.6b 
  F = 9.4 
    P = 0.005 
F = 31.04 
  P = 0.0001 
          F = 4.8 
          P = 0.032 
Nymphs 0.0 14.4 ? 1.1b 0c 8.8 ? 0.2a 
 1.0   17.3 ? 1.4ab    3.0 ? 1.5bc 8.7 ? 0.2a 
 2.0   19.6 ? 0.6ab 14.4 ? 3.1b 8.3 ? 0.2a 
 3.0  20.7 ? 1.3a 42.1 ? 4.1a 6.8 ? 0.4b 
   F = 5.71 
  P = 0.021 
F = 39.11 
P = 0.001 
 F = 12.36 
         P = 0.002 
Means (  SE) within the same column for each stage followed by different letters are 
significantly different (P < 0.05, Tukey-Kramer HSD test; df = 3, 8). 
 
 
 
 62 
Table 3. Evaluation of damage to satsuma fruit infested by Leptoglossus zonatus females   
in two south Alabama satsuma orchards in 2008 
Location Density  
(insect no. 
per fruit) 
Mean (? SE) per fruit 
Fruit 
Color 
PFA 
(%) 
FFW 
(g) 
PDS 
(%) 
SSC    
(%) 
Coker 0 G 0b 103.9 ? 3.5       0d 9.1 ? 0.2a 
 0.5 G 0b   85.7 ? 3.1   3.6 ? 1.8d 8.7 ? 0.2a 
 1 Y 87.5 ? 12.5a   97.8 ? 8.3 15.2 ? 2.3c 8.2 ? 0.2a 
 2 Y 75.0 ? 16.2a   88.3 ? 7.7 31.9 ? 3.9b 6.0 ? 0.4b 
 3 Y  100 ? 0a   80.8 ? 7.5 56.8 ? 6.8a 5.0 ? 0.2bc 
     F = 28.0 
  P < 0.0001 
F = 2.14 
     P = 0.09 
 F = 41.34 
P < 0.0001 
 F = 45.99 
P < 0.0001 
GCREC 0 G 0c 113.2 ? 6.7 0d 9.4 ? 0.3a 
 0.5 G 12.5 ? 6.5bc 105.7 ? 3.9   3.0 ? 1.7cd 8.6 ? 0.3a 
 1 Y 75.0 ? 16ab 102.3 ? 1.3 11.0 ? 2.7c 7.1 ? 0.2b 
 2 Y 62.5 ? 18.2a 106.7 ? 6.8 37.0 ? 2.4b 5.8 ? 0.3c 
 3 Y  100 ? 0.0a 108.3 ? 6.0 71.0 ? 3.2a 5.1 ? 0.3c 
    F = 11.8 
 P < 0.0001 
F = 0.512 
P = 0.691 
F = 122.38 
P < 0.0001 
F = 39.99 
P < 0.0001 
Means (  SE) within the same column for each location followed by different letters are 
significantly different (P < 0.05, Tukey-Kramer HSD test; df = 4, 35). GGREC = Gulf Coast 
Research and Extension Center, Fairhope, AL; G = green color, Y = yellow color; PFA = 
premature fruit abortion (%); FFW = fresh fruit weight (in grams); SSC = soluble solids content 
(%); PDS = damaged fruit sections (%). 
 
 
 
 63 
Table 4. Evaluation of damage to satsuma fruit infested by Leptoglossus zonatus males in 
two south Alabama satsuma orchards in 2008 
Location Density  
(insect no. 
per fruit) 
Mean (? SE) per fruit 
Fruit 
Color 
PFA 
(%) 
FFW 
(g) 
PDS 
(%) 
SSC    
(%) 
Coker 0 G 0c 100.7 ? 4.3        0d 9.5 ? 0.3a 
 0.5 G 0c   90.8 ? 2.3   2.1 ? 1.4d 8.6 ? 0.1a 
 1 Y 37.5 ? 18.2bc   90.8 ? 4.5 17.2 ? 4.5c 6.8 ? 0.5b 
 2 Y 50.0 ? 18.9b   94.9 ? 4.8 34.3 ? 2.3b 6.3 ? 0.4b 
 3 Y  100 ? 0.0a 104.5 ? 3.9 64.8 ? 2.7a 5.3 ? 0.5b 
       F = 12.4 
   P < 0.0001 
F = 2.36 
   P = 0.07 
 F = 99.37 
 P < 0.0001 
F = 18.37 
P <0.0001 
GCREC 0 G 0c 101.0 ? 2.1 0c 8.9 ? 0.3a 
 0.5 G 0c   99.8 ? 1.9   1.4 ? 0.7c 8.4 ? 0.2a 
 1 Y  50.0 ? 18.8b 100.7 ? 2.2   8.1 ? 2.6c 6.9 ? 0.3b 
 2 Y  87.5 ? 12.5a 100.9 ? 3.6 34.5 ? 4.1b 6.9 ? 0.5b 
 3 Y   100 ? 0.0a   91.6 ? 3.1 65.9 ? 4.1a 5.0 ? 0.3c 
   F = 21.6 
   P < 0.0001 
   F = 2.34 
P = 0.09 
 F = 89.13 
P < 0.0001 
F = 27.12 
P < 0.0001 
 Means (  SE) within the same column for each location followed by different letters are 
significantly different (P < 0.05, Tukey-Kramer HSD test; df = 4, 35). GGREC = Gulf Coast 
Research and Extension Center, Fairhope, AL; G = green color, Y = yellow color; PFA = 
premature fruit abortion (%); FFW = fresh fruit weight (in grams); SSC = soluble solids content 
(%); PDS = % damaged fruit sections. 
 
 
 
 
 64 
 Figure 1.  Damage symptoms on satsuma fruits infested by L. zonatus. A (external damage): 
control fruit showing no damage (A1), ?green islands? on outer rind of infested fruit (A2); B 
(necrosis): control fruit showing no necrosis (B1), necrosis on inner rind of infested fruit (B2); C 
(internal damage): control fruit showing no internal damage (C1), infested fruit showing dried-
out juice sacs (C2).  
A 
                                
B 
           
C 
           
A1  A2  
C1  
B2  B1  
C2  
 
 65 
Figure 2.  Effects of insect density and feeding duration on percent fruit weight loss (% FWL) of 
satsuma fruits infested by Leptoglossus zonatus in laboratory tests in 2008. A: Females, B: Males; 
C: Nymphs. Figure shows mean (? SE) number of FWL (%) over different feeding periods. 
1
3
7
107
14
A:  Fe m a le s
0 2 4 6 8 10 12 14 16
M
ea
n 
(?
 S
E) 
FWL 
(%
)
0
5
10
15
20
25
30
0  (co nt ro l)
1   per  f ruit
2   per  f ruit
3   per  f ruit
B : Ma le s
X D a ta
0 2 4 6 8 10 12 14 16
0
5
10
15
20
25
30
0  (co nt ro l)
1   per  f ruit
2   per  f ruit
3   per  f ruit
C:  Ny m ph s
D a y s a ft er insect  in fest a tio n
0
5
10
15
20
25
0  (co nt ro l)
1   per  f ruit
2   per  f ruit
3   per  f ruit
  a b
bc
ab
 c
 a
ab
bc
 c
   a
ab
 b
c
 a
ab
ab
 b
a
ab
b
a
bc
ab
c
a
 
 
 
 66 
Figure 3.  Effects of insect density and feeding duration on (A) percent damaged sections (PDS) 
and (B) soluble solids content (SSC) of satsuma fruits infested by L. zonatus females in 
laboratory tests in 2008. 
0
10
20
30
40
50
60
70
80
M
e
an
 (
?
 S
E
)
 P
D
S
 (
%)
0 1 2 3 0 1 2 3 0 1 2 3
F e m a l e s M a l e s N y m p h s
3  d
1 4  d
A :  PD S 3  d 7  d 1 0  d 1 4  d
F
e
e
d
i
n
g
 d
u
r
a
t
i
o
n
 (
d
a
y
s
)
0
2
4
6
8
10
M
e
an
 (
?
 S
E
)
 S
S
C
 (
%)
0 1 2 3 0 1 2 3 0 1 2 3
F e m a l e s M a l e s N y m p h s
3  d
1 4  d
D e n s i t y /  f r u i t
B :  S S C 3  d 7  d 1 0  d 1 4  d
F
e
e
d
i
n
g
 d
u
r
a
t
i
o
n
 (
d
a
y
s
)
 
 
 
 
 
 
 
 
 
 67 
Figure 4.  Negative correlation between percent damaged sections (PDS) and soluble solids 
content (SSC) of satsuma fruits infested by L. zonatus females at different densities. A: Data 
from the laboratory test in 2008, B: Data from the field trial in Coker orchard in 2008. 
A
0
2
4
6
8
10
0 1 2 3
M
e
an
 (
?
 S
E
)
 S
S
C
 (
%)
0
10
20
30
40
50
60
70
80
M
e
an
 (
?
 S
E
)
 P
D
S
 (
%)
SSC PD S
 
B
0
2
4
6
8
10
0 1 2 3
D e n s i t y /  f r u i t
M
e
an
 (
?
 S
E
)
 S
S
C
 (
%)
0
10
20
30
40
50
60
70
 
M
e
an
 (
?
 S
E
)
 P
D
S
 (
%)
SSC PD S
  
 
 
 
 68 
 
 
 
 
 
 
CHAPTER 4  
SEASONAL PHENOLOGY AND NATURAL ENEMY FAUNA OF CITRUS 
LEAFMINER, PHYLLOCNISTIS CITRELLA (LEPIDOPTERA: GRACILLARIIDAE), 
AND EVALUATION OF PHEROMONE-BAITED FOR MONITORING THE PEST IN 
ALABAMA SATSUMA ORCHARD 
 
INTRODUCTION 
The citrus leafminer, Phyllocnistis citrella Stainton (Lepidoptera: Gracillariidae) is a 
major pest of citrus in many parts of the world including Africa, Australia, the Middle East, the 
Caribbean, Central, South, and North America, (Heppner 1993, Knapp et al. 1995, Heppner and 
Dixon 1995, Pe?a et al. 1996, Perales-Gutierrez et al. 1996, Hoy and Nguyen 1997, Legaspi et al. 
2001, Diez et al. 2006). Native to southeast Asia, P. citrella was first recorded in the United 
States (U.S.) in 1993 in citrus nurseries in Dade County, Florida and quickly spread rapidly 
throughout the state (Heppner 1995). The pest is now found in several citrus producing states in 
the U.S. including Alabama, Louisiana, Texas and California (Gill 1999, Legaspi et al. 1999). 
Satsuma mandarin production is an emerging fruit industry in the Gulf Coast region of the U.S. 
(Campbell et al. 2004). Recent surveys have identified P. citrella as a major pest of satsuma 
mandarin (Citrus unshiu Marcovitch) in Alabama (Fadamiro et al. 2007, 2008).  
Larvae of P. citrella feed on the leaf epidermis ingesting sap and causing chlorosis. They 
typically make serpentine tunnels called mines under the leaf cuticle, resulting in twisted and 
 
 69 
malformed leaves (Heppner 1993, Legaspi et al. 1999). The serpentine mines may reduce the 
photosynthetic capacity of leaves and increase their susceptibility to plant pathogens, such as the 
citrus canker bacterium, Xanthomonas axopodis pv. citri (Sohi and Sandhu 1968, Cook 1988, 
Schubert and Sun 1996, Gottwald et al. 1997, 2002). Virtually all citrus cultivars are susceptible 
to P. citrella (Heppner 1993, Legaspi et al. 1999) and heavy infestations can result in significant 
yield loss (Pe?a et al. 2000). The pest is potentially damaging to young nurseries and top-grafted 
trees since damage can prevent young leaves from expanding and sometimes cause leaves to fall 
(Diez et al. 2006).  
Several aspects of the biology and ecology of P. citrella have been studied in many citrus 
growing regions of the world (Ba-Angood 1978, Huang et al. 1989, Heppner 1993, Pe?a et al. 
1996, Pe?a 1998, Legaspi et al. 2001, Diez et al. 2006, Xiao et al. 2007, Stelinski and Rogers 
2008). Many of the above studies reported on the seasonal population dynamics of the pest and 
associated natural enemies in different regions. The species is multivoltine with number of 
generations per year ranging from six in Japan (Clausen 1931) to 13 in Northcentral India (Lal 
1950). In Florida, P. citrella is present season long but peak flight activity occurs from late 
March through early October, during the availability of new leaf flushes (Lapointe and Leal 
2007). The female produced sex pheromone of P. citrella was first partially identified by Ando 
et al. (1985). Recent studies have identified the full composition of this sex pheromone as 
consisting of three active components: (Z, Z, E)-7, 11, 13-hexadecatrienal, (Z, Z)-7, 11-
hexadecadienal, and (Z)-7-hexadecenal (Leal et al. 2006, Moreira et al. 2006). Follow-up field 
tests demonstrated strong attraction of P. citrella males to traps baited with a binary or tertiary 
lure of the active sex pheromone components (Mafi et al. 2005, Lapointe et al. 2006, Leal et al. 
2006, Moreira et al. 2006, Lapointe and Leal 2007). A recent study also reported on the utility of 
 
 70 
traps baited with a commercially produced synthetic sex pheromone lure of P. citrella for 
monitoring the pest in Florida (Stelinski and Rogers 2008).  
Phyllocnistis citrella is a perennial pest of satsuma mandarin in southern Alabama 
(Fadamiro et al. 2007, 2008) but little is known about its seasonal abundance and natural 
mortality factors in the region. Furthermore, the utility of pheromone-baited traps for monitoring 
P. citrella in southern Alabama and other parts of the Gulf Coast has not been demonstrated. 
Because pest phenology and response to pheromone traps may vary from region to region due to 
several factors including variations in climate and crop varieties (Fadamiro 2004a, b), data 
obtained from other citrus growing regions may not provide accurate information on the 
abundance and seasonal activity of P. citrella in southern Alabama. The objectives of this study 
were to 1) determine seasonal abundance of P. citrella immatures and associated natural enemies 
in Alabama satsuma orchards, and 2) evaluate the efficacy of two popular types of commercially 
available traps baited with synthetic sex pheromone lures for monitoring adult populations of the 
pest in Alabama. The results are expected to support the development of monitoring guidelines 
and management tools for P. citrella in southern Alabama and other parts of the Gulf Coast 
region. 
 
MATERIALS AND METHODS 
Seasonal Abundance of P. citrella Immatures  
The seasonal abundance of immature stages of P. citrella was studied during 2006 and 
2007 at several locations in southern Alabama. Six orchards (Brantley, Buck, Coker, Ladnier, 
Gulf Coast Research and Extension Center or GGREC, and McDaniel) were sampled in 2006 
and four (Brantley, Coker, GCREC, and McDaniel) in 2007. The survey orchards were located in 
 
 71 
Baldwin and Mobile counties, the two main citrus-growing regions in southern Alabama, and 
were comprised mainly of satsuma mandarin, with a few sweet orange (Citrus sinensis), 
grapefruit (C. paradise), and kumquat (Fortunella spp.). The predominant cultivar of satsuma 
mandarin was ?Owari?, with few trees each of ?Armstrong Early? and ?Brown?s Select?. Five of 
the orchards (Brantley, Buck, Coker, GCREC and Ladnier) were commercial farms typically 
managed using conventional practices including routine applications of pesticides, while 
McDaniel orchard was unsprayed prior to and during the surveys.  
Each orchard was repeatedly sampled biweekly (at 2-week intervals) by collecting 5 
flushes (each with ~ 10 leaves) from five randomly selected trees for a total of 25 flushes (or ~ 
250 leaves) per sampling date per orchard. The flushes were stored in properly-labeled paper 
bags, held in a cooler and transported to the laboratory, where they were examined under the 
microscope (10 ? magnification) to determine the presence of all stages of P. citrella. The 
number of P. citrella immatures (sum of larvae, prepupae and pupae) per flush was recorded. 
Mean numbers per flush per sampling date were then calculated using the 25 flushes as replicates, 
and then mean number were used to generate seasonal abundance charts. Observations on natural 
enemies of P. citrella were also periodically made in the six satsuma orchards surveyed in 2006. 
On three of the sampling dates (May, July, October), predators observed on the surveyed trees 
were captured by hand or sweep net, collected in glass vials containing ethyl alcohol (75 %) and 
transported to the laboratory for identification. In addition, the P. citrella infested leaves 
collected on the above three sampling dates were kept in properly labeled Petri dishes (9 cm 
diameter) in the laboratory [25 ? 2 ?C, 50 ? 10% RH, and a photoperiod of 14:10 (L: D) h] to 
determine parasitism and rear out the parasitoids. Final species determinations of predators and 
 
 72 
parasitoids were made by Dr. Charles Ray (Auburn University Plant Diagnostic Laboratory), and 
voucher specimens were deposited in the Auburn University Entomology museum.  
 
Evaluation of Pheromone-Baited Traps and Adult Seasonal Phenology  
A second study was conducted at two locations (GCREC and Coker orchards) in southern 
Alabama during 2006 and 2007 to evaluate two types of sticky traps, Pherocon VI ?Delta? trap 
and Pherocon 1C ?Wing? trap (Tr?c? Inc., Salinas, CA) for monitoring adult P. citrella. Both 
traps types are popular among growers and are commonly used for monitoring adult Lepidoptera 
in North America (Riedl et al. 1976, Delisle, J. 1992, Fadamiro 2004a, b, Stelinski and Rogers 
2008). The two traps have similar liner catch surface area: 379 cm2 and 394 cm2 for Pherocon VI 
Delta trap (evaluated with its opening flap closed) and Pherocon 1C Wing trap, respectively. 
Each orchard was divided into four blocks where the trap treatments were replicated using a 
randomized complete block design. The following four treatments were evaluated in each block: 
i) Pheromone-baited Delta trap; ii) Pheromone-baited Wing trap; iii) Unbaited Delta trap, and iv) 
Unbaited Wing trap. Pheromone-baited traps were baited with a rubber septum (gray) lure loaded 
with 0.1 mg of Z7Z11E13-16: Ald and 0.033 mg of Z7Z11-16:Ald (Advanced Pheromone 
Technologies or APTIV, Portland, OR), which are the active sex pheromone components of P. 
citrella (Leal et al. 2006, Moreira et al. 2006). The unbaited traps of each type (i.e. treatments iii 
and iv) served as non-lure controls. In each block, traps were randomly placed along a single row 
of satsuma trees at ~ 1.5 m above the ground and spaced apart by at least 20 m within a block 
and 40 m between blocks. Satsuma tree canopy heights at the two test orchards ranged from 2.5 
m to 3.5 m. Traps were deployed in the last week of April in 2006 and early March in 2007 and 
removed in mid December of each year, and were checked weekly (2006) or biweekly (2007), 
 
 73 
counting and recording the number of P. citrella males per trap. The dates of first and peak moth 
captures were also noted for each trap. Captures of nontarget insects were very minimal in both 
trap types and thus were not analyzed or presented. The position of each trap was re-randomized 
every four weeks to minimize potential effect of trap position on moth captures. Pheromone lures 
were replaced every four weeks in accordance with manufacturer?s recommendation, and trap 
liners were replaced after each weekly or biweekly data collection. Data for each orchard and 
year were analyzed and presented separately. Captures in unbaited traps were too low for 
analysis (near zero) and thus were not included in the comparison. Actual captures in 
pheromone-baited trap were calculated by subtracting captures in unbaited control traps of each 
type from captures in pheromone-baited traps. Weekly (2006) or biweekly (2007) data were used 
to general adult seasonal phenology charts. Seasonal mean capture per sampling date was 
calculated for each block (replicate) and then used to calculate the seasonal mean capture for 
each pheromone-baited trap treatment (means of four replicates per trap type). Seasonal mean 
trap captures were first transformed by using the square-root method (? x + 0.5), and then 
analyzed using the student?s t test (P < 0.05, JMP Version 7.01, SAS Institute 2007) to determine 
significant differences between the two trap types. 
 
RESULTS AND DISSCUSION 
Seasonal Abundance of P. citrella Immatures  
The seasonal mean number of P. citrella immatures per flush per sampling period (2 
weeks) in the surveyed orchards ranged from  3.1 to 5.1 in 2006 and from 4.3 to 6.4 in 2007 
(Table 1), indicating moderate to high infestation in all the orchards. Among the orchards, the 
highest population densities were recorded at Buck, Ladnier and McDaniel in 2006 and at 
 
 74 
Brantley, Coker and GCREC in 2007. Data for these top three locations for each year were used 
to generate seasonal phenology charts (Fig. 1). P. citrella immatures were recorded throughout 
the sampling period from April to November each year, indicative of continuous overlapping 
generations. However, the highest densities were recorded during summer: densities 
progressively increased from May to July and remained at high levels throughout the summer 
(Fig. 1). Population densities of P. citrella began to decline in October and eventually reached 
very low levels in November. At least three distinct peaks of P. citrella were recorded per year in 
each orchard (Fig. 1). The percentage of infested leaves per flush per sampling date was 
calculated for the top location for each year (i.e. Ladnier in 2006 and Brantley in 2007) to 
determine possible association between the number of leaves per flush (used as an estimate of 
flush abundance) and the severity of P. citrella infestation (Fig. 2). The number of leaves per 
flush increased slightly from ~ 8 - 10 in spring (April-May) to 10 - 12 in summer (June-
September) and then decreased in fall (October-November). The lowest numbers of leaves per 
flush (~ 5 ? 7) were recorded in November. The percentage of infested leaves per flush was very 
low in spring and sharply increased to very high levels in summer and decreasing again in fall 
(Fig. 2).  
In general, our results on the seasonal phenology of P. citrella in Alabama are similar to 
those reported in Florida (Pe?a et al. 1996), Mexico (Bautista-Mart?nez et al. 1998), and southern 
Texas (Legaspi et al. 1999): population density of P. citrella typically increased from spring to 
summer and declined during winter. A similar seasonal phenology was also reported in 
Argentina where the highest densities were recorded in spring and summer (Diez et al. 2006). 
The relatively greater abundance of P. citrella during spring and summer has been attributed to 
greater availability of leaf flushes and new shoots, as well as higher temperatures (Pe?a et al. 
 
 75 
1996, Legaspi et al. 1999, Diez et al. 2006). However, our data which showed only a slight 
increase in the availability of leaf flushes in summer compared to the other periods suggest that 
the higher incidence of P. citrella recorded in summer cannot be fully explained by the higher 
abundance of new leaf flushes during this period. It is likely that lower temperatures starting 
from fall through early spring (October-May) contributed immensely to the lower incidence of P. 
citrella during this period. Pe?a et al. (1996) also attributed the decline in P. citrella populations 
during winter in Florida to low temperatures since they observed citrus flushes during this period. 
Similarly, the increase in the population of this pest during spring and summer in Argentina was 
ascribed more to increase in temperatures than to the presence of citrus flushes (Diez et al. 2006). 
Our data, which showed that the percentage of infested leaves per flush were much higher in 
summer (> 75%) than in spring and fall, further lend support to the proposition that changes in 
availability of leaf flushes played only a minor role in the observed seasonal population 
dynamics of P. citrella in Alabama. Although not statistically analyzed due to the low number of 
unsprayed orchards (n = 1), no consistent differences were recorded in the population densities 
of P. citrella in sprayed versus unsprayed orchards, as also reported by Diez et al. (2006). 
 
Natural Enemy Fauna  
At least 21 species of beneficial arthropods (19 predacious and 2 parasitoid species) were 
observed in the surveyed orchards (Table 3). However, abundance of the different species and 
types was not compared due to their occurrence at generally low densities (Table 4). Spiders 
were the most common predatory arthropods observed in all surveyed orchards (Table 4). Spider 
communities consisted of 13 species from nine families, including Araneidae (1 species), 
Anyphaenidae (1 species), Filistatidae (1 species), Lycosidae (1 species), Mitrugidae (1 species), 
 
 76 
Oxyopidae (1 species), Salticidae (4 species), Tetragnathidae (2 species), and Theridiidae (1 
species) (Table 3). The dominant predacious spider species were the yellow ghost spider, Hibana 
sp. (Anyphaenidae), the long-legged sac spider, Cheiracanthium sp. (Miturgidae), and Hentzia sp. 
(Salticidae), which were collected from at least three of the six orchards. Several studies have 
also identified spiders as important predators of P. citrella and other citrus pests (Muma 1965, 
Dondale et al. 1979, Mansour et al. 1982, Breene et al. 1993, Amalin et al. 2001, Ghavami and 
Amooz 2008). In south Florida citrus groves, the spider community consisted of 15 species from 
nine families (Amalin et al. 2001), while 34 species from 13 families were collected in southern 
Texas (Breene et al. 1993). Our results which showed high diversity and abundance of spiders 
relative to other beneficial arthropods (Table 4) suggest their importance as natural enemies of P. 
citrella and other pests in Alabama satsuma orchards. In addition, eight predacious insect species 
in the families Coccinellidae (1 species), Formicidae (2 species), Anthocoridae (1 species), 
Chrysopidae (1 species), and Phlaeothripidae (1 species) were observed. The key predacious 
insect species included ants (Hymenoptera: Formicidae), green lacewing, Chrysoperla spp. 
(Neuroptera: Chrysopidae), and multicolored Asian lady beetle, Harmonia axyridis Pallas 
(Coleoptera: Coccinellidae), which were observed feeding on P. citrella larvae in at least five of 
six surveyed orchards. These predators have also been recorded to feed on P. citrella (Chen et al. 
1989, Amalin et al. 2001, Xiao et al. 2007, Hoy et al. 2007).  
Two parasitoid species were reared out from P. citrella: Ageniaspis citricola 
Logvinovskaya (Hymenoptera: Encyrtidae) and Cirrospilus sp. (Hymenoptera: Eulophidae). 
Ageniaspis citricola, an exotic specialist endo-parasitoid parasitoid of P. citrella which has been 
introduced in Florida and Texas (Hoy and Nguyen 1997, Pomerinke and Stansly 1998), was 
reared out in very low numbers from infested leaves collected from one of the six orchards. We 
 
 77 
are not aware of any documented purposeful introduction of A. citricola in Alabama citrus. Thus, 
its occurrence is likely due to accidental introductions through movement of rootstock materials 
from Florida or Texas, or range expansion by natural dispersal. One species in the genus 
Cirrospilus was also reared out in low numbers from two of the six locations (Table 4). 
Cirrospilus spp., are indigenous generalist ecto-parasitoids of P. citrella in the U.S. (Pena 1996, 
Schauff et al. 1998).  
 
Evaluation of Pheromone-Baited Traps  
No significant differences were recorded in captures of P. citrella in both types of traps  
(Delta versus Wing traps) at the two locations in both years of the study (Table 2). However, the 
data showed a slightly higher male captures in Delta traps than in Wing traps. The dates of first 
moth capture were also similar for both trap types (Table 2), indicating that both traps are 
equally capable of detecting early flights of P. citrella males. In contrast to our results, Stelinski 
and Rogers (2008) reported that the Pherocon VI Delta trap (with opening flap closed) baited 
with a sex pheromone lure captured significantly more P. citrella males than identically baited 
Pherocon IC Wing trap. Other studies have also reported on the superiority of Pherocon VI Delta 
trap over Pherocon IC Wing trap for monitoring adult Lepidoptera of various species (e.g., 
Knodel and Agnello 1990, Fadamiro 2004a, b).  
Several factors may account for the different results obtained in the present study and 
those of Stelinski and Rogers (2008), in particular differences in population densities in the test 
orchards. The trap captures reported by Stelinski and Rogers (2008) were very low (averaged 19 
- 40 males per trap per week) compared to the present study in which much higher seasonal trap 
captures averaging about 70 - 200 or more males per trap per week were recorded (Table 2). This 
 
 78 
suggests that population densities of P. citrella were much higher in our test orchards than in the 
citrus grove used by Stelinski and Rogers (2008). Assuming that the Delta-style trap is indeed 
superior to the Wing-style trap, the absence of a significant trap effect in our study may be 
related to the higher population densities of the pest in our test orchards. This may have resulted 
in the quick saturation of the trap liners thereby impacting trap efficiency, given that our traps 
were checked and replaced on a biweekly basis. Future studies are needed to confirm the 
potential impact of insect population density on efficacy of pheromone-baited traps. Nevertheless, 
based on the report which showed that the Delta-style trap is more efficient than the Wing-style 
trap for monitoring P. citrella at low to moderate population densities (Stelinski and Rogers 
2008) and a consideration of other factors including trap durability and ease of deployment, the 
Delta-style trap baited with the sex pheromone lure is recommended for monitoring P. citrella in 
Alabama citrus. However, citrus growers who prefer the Wing-style trap may still achieve 
effective monitoring of the pest with it. 
 
Adult Seasonal Phenology  
The weekly or biweekly trap captures of P. citrella in pheromone-baited delta trap at both 
orchards are presented in Figure 3 to illustrate the seasonal phenology of the moths. In both years, 
P. citrella males were captured continuously in both orchards, indicating overlapping 
generations. However, the highest trap captures were recorded from August to October in 2006 
and from June to October in 2007. The data suggest at least two major flight peaks and possibly 
one or more minor peaks per year. In 2006 at the GCREC location, a small first peak was 
recorded in early June, followed by a second large peak in late August, and a third larger peak in 
late October. In 2007 at the same location, a large first peak was recorded in late May-early June, 
 
 79 
followed by a larger second peak in late September. Similar to our results, Lapointe and Leal 
(2007) reported about four flight peaks for P. citrella in Florida, including a small first peak in 
April, followed by a second larger peak in May-June, a large peak in July-August, and a small 
peak in October. The phenology of P. citrella moths generally followed the same pattern as that 
of the immatures, with peak densities of both recorded in summer. However, male moths were 
captured in the traps a little earlier than the occurrence of the immature stages, which was not 
unexpected.  
Variable numbers of generations per year have been reported for P. citrella in different 
regions, from six in Japan (Clausen 1931) to 13 in Northcentral India (Lal 1950). Although it is 
difficult to accurately determine the number of generations for this pest in southern Alabama due 
to the observed generations overlap, one could deduce the approximate number of generations 
from a combination of our data and published information on the developmental biology of P. 
citrella. Pandey and Pandey (1964) recorded developmental periods of 13 - 52 days for P. 
citrella depending on weather conditions, while Ba-Angood (1978) reported 47.5, 39.7, 28, and 
23.1 days at 20, 25, 30, and 35?C, respectively. Using a modest developmental period of about 
30 days in the hot, humid southern Alabama conditions, an estimate of six or more generations 
per year (from May to October) for P. citrella in Alabama is plausible. 
In general, densities of P. citrella males were lower in 2007 than in 2006 (Table 2) and 
the major flight peaks occurred earlier in 2007 than in the previous year (Fig. 3). It is unlikely 
that the reduced adult populations in 2007 is related to lower survival of overwintering 
populations in winter 2006 or reduced larval development or survival in spring 2007, since 
slightly higher numbers of immatures were recorded in 2007 compared to 2006 (Table 1). The 
data suggest the possibility of reduced flight activity of moths during 2007 probably due to lower 
 
 80 
temperatures or higher rainfall. To determine if temperature and rainfall could account for the 
lower adult populations recorded in 2007, historic weather data for Fairhope (nearest weather 
station to the orchards) were analyzed, comparing climate data for 2006 versus 2007. 
Temperature did not adequately explain the recorded difference between both years. However, 
key differences were recorded in the rainfall (precipitation) data with 2007 being a very wet year. 
The average total precipitation per month for the period when flight activity of P. citrella was 
highest (June - October) was ~ 3.98 and 6.40 for 2006 (June to Oct: 1.27, 3.23, 6.71, 5.27, and 
3.44) and 2007(June to October: 5.97, 6.47, 5.12, 6.09, and 8.33), respectively. In contrast, 
average rainfall per month in spring (April-May) was higher in 2006 (4.82) (3.48-6.18) than 
2007 (2.61) (1.86-3.35). Thus, higher rainfall in summer 2007 could account, at least in part, for 
the lower adult trap capture compared to summer 2006. Similarly, the rainfall data could also 
explain the differences in the spring activity of the pest in both years: the relatively lower spring 
activity of adults in spring 2006 compared to spring 2007 (Fig. 3) could be related to higher 
rainfall in spring 2006.  
In summary, our results confirm that P. citrella is an annual pest with multiple 
overlapping generations in Alabama. The low abundance of parasitoids in the surveyed orchards 
may suggest that the level of control provided by parasitoids is minimal, and this may partly 
explain the high abundance of P. citrella in Alabama orchards relative to Florida and other citrus 
growing regions. Studies are ongoing to determine the impact of key natural enemies of P. 
citrella in Alabama satsuma orchards. 
  
 
 81 
ACKNOWLEDGEMENTS   
The authors thank Mr. Monte Nesbitt (Gulf Coast Research and Extension Center, 
Fairhope, AL) for field assistance, and Drs. Art Appel, Charles Ray and Elina Coneva (Auburn 
University) for helping to review an earlier version of this paper. Dr. Charles Ray is also 
acknowledged for assisting with final species determinations of the specimens. We also thank 
our commercial satsuma grower cooperators in southern Alabama. Funding for this study was 
provided through grants by the Alabama Agricultural Experiment Station and Auburn University 
Horticulture Line Item grants program to HYF. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 82 
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Pe?a, J. E., R. Duncan, and H. Browning. 1996.  Seasonal abundance of Phyllocnistis citrella 
(Lepidoptera: Gracillariidae) and its parasitoids in south Florida citrus. Environ. Entomol. 25: 
698 - 702. 
Pe?a, J. E., Hunsberger, A., and Schaffer, B. 2000. Citrus leafminer (Lepidoptera: 
Gracillariidae) density: effect on yield of ?Tahiti? lime. J. Econ. Entomol. 93: 374 - 379.  
 
 87 
Perales-Gutierrez, M., H. C. Arredondo-Bernal,  E. Garza-Gonzalez, and A. Aguirre-Uribe. 
1996.  Native parasitoids of citrus leafminer, Phyllocnistis citrella Stainton in Colima, 
Mexico. Southwest. Entomol. 21: 349 - 350. 
Pomerinke, M. A., and P. A. Stansly. 1998.   Establishment of Ageniaspis citricola 
(Hymenoptera: Encyrtidae) for biological control of Phyllocnistis citrella (Lepidoptera: 
Gracillariidae) in Florida. Fla. Entomol. 81: 361 - 372.  
Riedl, H., B. A. Croft, and A. J.  Howitt. 1976.  Forecasting codling moth phenology based on 
pheromone trap catches and physiological time models. Can. Entomol. 108: 449 - 460. 
SAS Institute. 2007.  JMP Statistics and Graphics Guide, Version 7.0.1. SAS Institute, Cary, 
NC. 
Schauff, M. E., J. LaSalle, and G. A. Wijesekara. 1998.  The genera of chalcid parasites 
(Hymenoptera: Chalcidoidea) of citrus leafminer, Phyllocnistis citrella Stainton (Lepidoptera: 
Gracillariidae). J. Nat. Hist. 32: 1001 - 1056. 
Schubert, T. S. and X. Sun.  1996.  Bacterial Citrus Canker.  Plant Pathology Circular No. 377.            
     Fla. Dept. of Agriculture & Cons. Ser. Fla, Gainesville, Fla. 
Sohi, G. S., and M. S. Sandhu. 1968.  Relationship between citrus leafminer (Phyllocnistis 
citrella) injury and citrus canker (Xanthomonas citri (Hasse) Dowson) incidence on citrus 
leaves. J. Res. Punj Agri. Univ. 5: 66 - 69. 
Stelinski, L.L., and M. E. Rogers. 2008.  Factors affecting captures of male citrus leafminer, 
Phyllocnistis citrella, in pheromone-baited traps. J. Appl. Entomol. 132: 143 - 150. 
Xiao, Y. F., J. A. Qureshi, and P. A. Stansly. 2007.  Contribution of predation and  
     parasitism to mortality of citrus leafminer, Phyllocnistis citrella Stainton (Lepidoptera: 
Gracillariidae), populations in Florida. Biol. Control. 40: 396 - 404. 
 
 88 
Table 1. Relative abundance of Phyllocnistis citrella immature stages in the Alabama 
satsuma orchards surveyed during 2006 ? 2007 
Years 
 
Location 
 
Mean (? SE) number of P. citrella 
immatures per flush a  
Date of peak 
abundance  
Seasonal abundance          Peak abundance  
2006 
 
 
Brantley 3.1 ? 0.4 6.9 ? 0.6 Sep. 6 
Buck 3.8 ? 04 7.8 ? 0.5 Aug. 26 
Coker 3.7 ? 0.4 7.3 ? 0.4 Sep. 6 
Ladnier 5.1 ? 0.4       10.1 ? 0.7 Aug. 26 
GCREC 3.4 ? 0.3 8.1 ? 0.5 Aug. 12 
McDaniel 4.5 ? 04 8.9 ? 0.5 Sep. 6 
2007 
 
Brantley 6.4 ? 0.2       10.9 ? 0.7 Jul. 17 
Coker 5.1 ? 0.2       10.0 ? 0.6 Jul. 2 
GCREC 5.3 ? 0.2       10.4 ? 0.7 Sep. 6 
McDaniel 4.3 ? 0.2         8.1 ? 0.6 Aug. 24 
a Mean of 25 flushes per sampling date (biweekly). Immatures = larvae + prepupae + pupae. 
Note: immatures were detected as early as April 27 in 2006 and April 10 in 2007. In this and 
Table 2 and the figures, GCREC = Gulf Coast Research and Extension Center. 
 
 
 
 
 
 
 
 89 
Table 2.  Captures of Phyllocnistis citrella males in delta and wing traps baited with 
synthetic sex pheromone lures during 2006 and 2007 in two Alabama satsuma orchards  
Location  Year  Trap  Date of first 
moth 
capture b 
Mean (? SE) trap capture per 
sampling period a 
Date of 
peak moth 
capture Seasonal capture Peak capture 
 
 
 
GCREC 
 
2006 
Delta Apr. 27   223 ? 97 972 ? 125 Oct. 19  
Wing Apr. 27  213 ? 63 836 ? 273 Oct. 19 
  t = 0.44, df = 206  
P = 0.66 
t = 0.628, df = 6  
P = 0.553 
 
 
2007 
Delta Mar. 12 252 ? 32 849 ?  94 Sep. 6 
Wing Mar. 12 219 ? 33  769 ? 287 Sep. 6 
  t = 0.79, df = 150 
P = 0.4298 
t = 0.508, df = 6  
P = 0.629 
 
 
 
 
Coker 
 
2006 
Delta Apr. 27   164 ? 114 663 ? 215 Oct. 4 
Wing Apr. 27 194 ? 38 640 ? 175 Aug. 26 
   t = 0.69, df = 206 
P = 0.49 
t = 0.063, df = 6  
P = 0.95 
 
 
2007 
Delta Mar. 12   175 ? 21      425 ? 43   Sep. 6 
Wing Mar. 12    143 ? 20 527 ? 186 Sep. 6 
  t =1.113, df = 150 
P = 0. 2673 
t = 0.569, df = 6  
P = 0.59 
 
a Means of four replicates per trap type per sampling date (weekly in 2006 and biweekly in 
2007). In this and other tables and figures, GCREC = Gulf Coast Research and Extension Center. 
Captures in delta and wing traps were not significantly different (P > 0.05, Student?s t test).         
b Date of the first moth capture in 2006 may not signal the beginning of activity of the pest since 
activity may have commenced prior to trap deployment in the field. 
 
 
 
 90 
Table 3. Beneficial arthropods observed in association with P. citrella in six Alabama 
satsuma citrus orchards in 2006 
Order Family Species Feeding habit Distribution a 
Araneae Anyphaenidae Hibana sp. Predator +++++ 
 Araneidae Gasteracantha 
cancriformis (Linnaeus) 
Web builder ++ 
 Filistatidae Kukulcania hibernalis 
(Hentz) 
Web builder ++ 
 Lycosidae Schizocosa sp. Hunter ++ 
 Miturgidae Cheiracanthium sp.  Predator +++ 
 Oxyopidae Peucetia virescens 
(Hentz) 
 + 
 Salticidae Hentzia sp. Hunter +++ 
  Hentzia palmarum 
(Hentz) 
Hunter  
  Sitticus sp. Hunter + 
  Thiodina sylvana (Hentz) Hunter + 
   Tetragnathidae Leucauge venusta 
(Walckenaer) 
Web builder + 
  Nephila clavipes 
(Linnaeus) 
Web builder + 
 Theridiidae Latrodectes geometricus Web builder ++ 
 
 91 
(Koch) 
Coleoptera Coccinellidea Harmonia axyridis Pallas Predator ++++++ 
Hymenoptera Encyrtidae Ageniaspis citricola 
Heppner 
Parasitoid + 
 Eulophidae Cirrospilus sp. Parasitoid ++ 
 Formicidae Solenopsis invicta Buren Predator ++++++ 
  Brachymyrmex 
patagonicus Mayr  
Predator? ++ 
Hemiptera Anthocoridae Orius insidiosus (Say) Predator ++ 
Neuroptera Chrysopidae Chrysoperia sp. Predator +++++ 
Thysanoptera Phlaeothripidae Unknown species Predator ++ 
aDistribution in Alabama was computed based on the presence of beneficial species in the 
surveyed orchards (+ indicates presence in only one of the surveyed orchards while ++++++ 
indicates presence in six surveyed orchards). 
 
 
 
 
 
 
 
 
 
 
 92 
Table 4.  Abundance of key beneficial arthropods observed in association with P. citrella in 
six Alabama satsuma citrus orchards in 2006 
Location May July October 
 SP IP PA SP IP PA SP IP PA 
Brantley 6 1 0 8 2 0 2 1 1 
Buck 1 1 0 1 0 0 0 0 0 
Coker 4 3 0 6 3 2 0 4 1 
Ladnier 1 0 0 0 2 0 0 0 0 
GCREC 2 0 0 2 1 1 1 0 0 
McDaniel 4 0 0 9 1 0 4 2 0 
Table shows number of natural enemies of each type recorded on five trees per orchard per 
sampling date. SP: spiders; IP: insect predators (i.e. ants, lady beetles, lacewings, etc); PA: 
parasitoids 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 93 
Figure 1. Seasonal abundance of Phyllocnistis citrella immatures in the Alabama satsuma 
orchards surveyed during 2006 (A) and 2007 (B). Figures shows mean (  SE) number of 
immatures per flush (~ 10 leaves) per sampling date (n = 25 flushes). Immatures = larvae + 
prepupae + pupae. GCREC = Gulf Coast Research and Extension Center. 
A:  2 0 0 6
Ap r  May   J un   J ul  Au g   Sep  Oct  Nov   Dec  
0
2
4
6
8
10
12
Buck
La dn ie r
McDanie l
B: 2 0 0 7
Date
Ap r  May   J un   J ul  Au g   Sep  Oct  Nov   Dec  
Mean 
(? 
SE
) 
no.
 o
f 
im
mat
ure
s per flush
0
2
4
6
8
10
12
Br a nt le y
Cok er
GCR EC
 
 
 
 
 
 94 
Figure 2. Associating seasonal abundance of Phyllocnistis citrella immatures with abundance of 
leaf flushes in two Alabama satsuma orchards (A: 2006 data for Ladnier, B: 2007 data for 
Brantley). No. leaves = number of leaves per flush, No. P. citrella = total number of P. citrella 
immatures (larvae + prepupae + pupae) counted; % infested leaves = % leaves with at least one 
P. citrella (calculated by dividing number of infested leaves per flush by the total number of 
leaves per flush). 
A :  L a d n i e r  
0
2
4
6
8
10
12
14
27-
A p r
18-
M a y
2-
J u n
16-
J u n
29-
J u n
13-
J u l
27-
J u l
12-
A u g
26-
A u g
6-
S e p
21-
S e p
6-
O c t
19-
O c t
2-
N o v
15-
N o v
2006
M
e
an
 (
?
 S
E
)
 n
o. p
e
r
 f
lu
s
h
0
10
20
30
40
50
60
70
80
90
%  of
 l
e
ave
s
 i
n
f
e
s
t
e
d
N o .  l e a v e s N o .  P.  c i t r e l l a %  o f  l e a v e s  i n f e s t e d
 
B :  B r a n t l e y
0
2
4
6
8
10
12
14
10-
A p r
24-
A p r
10-
M a y
22-
M a y
5-
J u n
19-
J u n
2-
J u l
17-
J u l
1-
A u g
14-
A u g
24-
A u g
6-
S e p
20-
S e p
4-
O c t
18-
O c t
2-
N o v
2007
M
e
an
 (
?
 S
E
)
 n
o. p
e
r
 f
lu
s
h
0
20
40
60
80
100
%  of
 l
e
ave
s
 i
n
f
e
s
t
e
d
N o .  l e a v e s N o .  P.  c i t r e l l a %  o f  l e a v e s  i n f e s t e d
  
 
 
 
 
 
 
 95 
Figure 3.  Seasonal phenology of Phyllocnistis citrella moths in two Alabama satsuma orchards 
during 2006 and 2007. Figure shows mean (? SE) weekly (A: 2006) or biweekly (B: 2007) 
captures of males per Delta trap baited with the synthetic sex pheromone lure. GCREC = Gulf 
Coast Research and Extension Center. 
A:  2 00 6
Mar  Ap r  May  Jun  Jul  Au g  Sep  Oct  Nov  Dec  Ja n  
M
ea
n (
? 
SE) n
o
. 
o
f 
ma
les per t
ra
p
0
200
400
600
800
1000
1200
GC REC
Cok er  
B: 2 00 7
D elta  t ra p 
Mar  Ap r  May  Jun  Jul  Au g  Sep  Oct  Nov  Dec  Ja n  
0
200
400
600
800
1000
GC REC
Cok er
 
 
 
 
 
 
 96 
 
 
 
 
 
 
CHAPTER 5  
 EXCLUSION EXPERIMENTS REVEALED RELATIVE CONTRIBUTION OF 
NATURAL ENEMIES TO MORTALITY OF CITRUS LEAFMINER, PHYLLOCNISTIS 
CITRELLA, (LEPIDOPTERA: GRACILLARIIDAE) IN  
ALABAMA SATSUMA CITRUS ORCHARDS 
 
INTRODUCTION 
Satsuma mandarin (Citrus unshiu Marcovitch) production is a growing industry in 
southern Alabama and other parts of the Gulf Coast region of the United States (Campbell et al. 
2004). Recent surveys have identified the citrus leafminer, Phyllocnistis citrella Stainton 
(Lepidoptera: Gracillariidae) as a key pest of this specialty fruit crop in Alabama (Fadamiro et al. 
2007, 2008). Phyllocnistis citrella originated in southeast Asia and has become a global pest of 
citrus, having been found in Africa, Australia, the Middle East, the Caribbean, Central, South, 
and North America, (Heppner 1993, Heppner and Dixon 1995, Pe?a et al. 1996, Perales-
Gutierrez et al. 1996, Hoy and Nguyen 1997, Legaspi et al. 1999, Diez et al. 2006). In the United 
States, P. citrella was first recorded in 1993 in citrus nurseries in Dade County, Florida (Heppner 
1993). The pest is now found throughout the state of Florida, as well as in several other citrus 
producing states including Alabama, Louisiana, Texas and California (Legaspi et al. 1999, Gill, 
1999).  
 
 97 
Phyllocnistis citrella attacks all varieties of citrus, other Rutaceae, and several 
ornamental species (Heppner 1993, Legaspi et al. 1999). Females lay eggs on host leaves and 
eclosing larvae feed on leaf epidermis ingesting sap and causing chlorosis and curled leaves 
(Heppner 1993, Legaspi et al. 1999). Larvae of P. citrella make characteristic serpentine mines 
under the leaf cuticle, which may reduce photosynthesis (Cook 1988) The feeding tunnels 
produced by P. citrella larvae on citrus leaves may facilitate infection by the citrus canker 
bacterium, Xanthomonas axopodis pv. citri (Sohi and Sandhu, 1968, Cook 1988, Gottwald et al. 
1997, 2002). High population densities of P. citrella are usually recorded in spring and summer 
due to greater availability of leaf flushes and new shoots, as well as higher temperatures (Pe?a et 
al. 1996, Legaspi et al. 1999, Diez et al. 2006). Phyllocnistis citrella is an important pest in citrus 
nurseries and top-grafted trees (Diez et al. 2006), and heavy infestation can have significant 
impact on growth and yield (Pe?a et al. 2000, Browning et al. 2006).  
Control of P. citrella is typically accomplished through multiple applications of 
conventional insecticides, which are often ineffective because the larvae are usually concealed 
within the mines and thus are protected from insecticide sprays (Legaspi et al. 2001). Biological 
control is generally regarded as the most economically sound and environmentally sustainable 
management practice for P. citrella (Knapp et al. 1995, Hoy and Nguyen 1997). The population 
dynamics of P. citrella and associated natural enemies have been documented in several 
countries and regions (Chen et al. 1989, Pena et al. 1996, Pena 1998, Urbaneja et al. 2000, 
Legaspi et al. 2001, Diez et al. 2006, Lapointe and Leal. 2007). Several predatory arthropods are 
known to feed on P. citrella, including lacewing larvae, ants, and hunting spiders (Argov et al. 
1996, Pomerinke 1999, Amalin et al. 2001, Hoy et al. 2007, Xiao et al. 2007), and many studies 
have identified predation as the most important natural mortality factor acting on P. citrella in 
 
 98 
many parts of the world (Chen et al. 1989, Amalin et al. 1996, 2001, 2002, Hoy et al. 2007, Xiao 
et al. 2007). In addition, many species of parasitoids have been reared from P. citrella worldwide 
(Hoy and Nguyen 1997, Schauff et al. 1998, Legaspi et al. 1999), however indigenous 
parasitoids were found to provide only minimal levels of parasitism in Florida (Pena et al. 1996). 
Consequently, Ageniaspis citricola Logvinovskaya (Hymenoptera: Encyrtidae), an exotic 
specialist endo-parasitoid of P. citrella larvae, was introduced in the 1990s for classical 
biological control of the pest in Florida and Texas (Hoy and Nguyen, 1997 Pomerinke and 
Stansly 1998). 
In a recent study (Xiao and Fadamiro, in review), we documented the natural enemy 
fauna of P. citrella in southern Alabama as consisting of at least 21 species of beneficial 
arthropods, including various species of spiders (e.g., Hibana sp., Cheiracanthium sp. and 
Hentzia sp.) (Araneae), ants (Hymenoptera: Formicidae), Chrysoperla spp. (Neuroptera: 
Chrysopidae), and Harmonia axyridis (Coleoptera: Coccinellidae). Two parasitoid species were 
also detected in low numbers: A. citricola and Cirrospilus sp. (Hymenoptera: Eulophidae). 
Cirrospilus spp. are indigenous generalist ecto-parasitoids of P. citrella in the United States 
(Pena 1996, Schauff et al. 1998). However, little is known about the impact of these beneficial 
arthropods on P. citrella in Alabama. In this study, exclusion techniques (e.g., Smith and 
DeBach 1942, Xiao et al., 2007, Qureshi and Stansly 2009) were used to determine the relative 
contributions of key beneficial arthropods to natural mortality of P. citrella in Alabama satsuma 
orchards. The results should provide the baseline data necessary for development of an effective 
biological control program for managing P. citrella in the Gulf Coast region of the United States.  
 
 
 
 99 
MATERIALS AND METHODS 
Study Sites  
Experiments were conducted in July 2007 and August 2008 in two satsuma orchards 
located in Baldwin County in southern Alabama: Coker orchard and the Gulf Coast Research and 
Extension Center orchard (GGREC). Both orchards were comprised mainly of satsuma mandarin, 
with a few sweet orange (Citrus sinensis), grapefruit (C. paradise), and kumquat (Fortunella 
spp.). The predominant cultivar of satsuma mandarin was ?Owari?, with few trees each of 
?Armstrong Early? and ?Brown?s Select?. Both orchards were typically managed using 
conventional practices including routine applications of pesticides, but were not sprayed during 
this study.  
 
Exclusion Experiments 
Experimental trees in both orchards were first pruned to induce new flushes necessary for 
P. citrella infestation. Newly hatched first instar larvae were then located using a hand lens and 
identified by marking the adjacent leaf surface with dark ink. Two types of exclusion techniques 
(sticky barrier and cage barrier) were used to evaluate the relative contribution of natural 
enemies to P. citrella mortality, following the procedures described by Xiao et al. (2007) and 
Qureshi and Stansly (2009), but with some minor modifications. The use of both exclusion 
techniques was timed in order to determine the impact of various biotic factors attacking larvae 
of P. citrella at the different development stages. The aim of the sticky barrier exclusion 
treatments was to evaluate P. citrella mortality factors due to ants and other crawling predators 
(e.g., lacewing and lady beetle larvae) by comparing mortalities in P. citrella cohorts protected 
by a sticky barrier versus unprotected (control) cohorts. The cage barrier exclusion treatments 
 
 100 
were tested to separate the effect of other biotic factors (e.g., spiders and parasitoids) attacking 
larvae of P. citrella.  
In the first experiment (July 2007), 30 pruned branches located on mature satsuma trees 
were selected in each orchard. Each branch contained expanding leaves and 15 first stage larvae 
of P. citrella. The branches were randomly distributed among the following six treatments (i.e. 
each treatment was replicated five times): i) branch protected with sticky barrier from day 1 to 
day 12 (sticky barrier 1-12 or complete sticky barrier): ii) branch protected with sticky barrier 
from day 1 to day 5 with the sticky barrier removed on day 6 when the larvae had become 
second instars (sticky barrier 1-5 or early sticky barrier); iii) branch protected with sticky barrier 
from day 6 to day 12 when the larvae had become pupae (sticky barrier 6-12 or later sticky 
barrier); iv) branch caged from day 1 to day 5 with the cage removed on day 6 when the larvae 
had become second instars (cage barrier 1-5 or early cage barrier); v) branch caged from day 6 to 
day 12 when the larvae had become pupae (cage barrier 6 - 12 or later cage barrier); vi) control 
(branch not protected with sticky barrier or cage throughout the experiment). For the sticky 
barrier treatments, a 5-cm diameter sticky trap strip (Trece Inc, Salinas, CA) was applied to the 
branch to preclude ants and other crawling predators from reaching the larvae. For the cage 
barrier treatments, a sleeve cage made of fine mesh organdy (35 ? 15 cm diameter, white color 
made from screen netting) was used to exclude beneficial arthropods, such as spiders and 
parasitoids. The control (no sticky or cage barrier) ensured that no arthropods were excluded 
(Smith and DeBach, 1942).  
The experiment was repeated in August 2008 but with some minor modifications. At 
each location, 28 pruned branches, each containing expanding leaves and 15 first instar larvae of 
P. citrella, were randomly distributed among seven treatments (i.e. four replicates per treatment). 
 
 101 
The treatments consisted of the six treatments evaluated in 2007 plus a seventh treatment, a 
branch caged from day 1 to day 12 (cage barrier 1-12 or complete cage barrier). The average 
temperatures at both locations during the experiments in July 2007 and August 2008 were 
29.3?C (minimum: 24.5?C, maximum: 34?C) and 28.0?C (minimum: 23.4?C, maximum: 32.5?C), 
respectively. Average rainfall (precipitation) for July 2007 and August 2008 was 5.12 and 5.33, 
respectively. 
 
Evaluation of Mortality Factors 
Branches were inspected daily noting and recording the development and survival of the 
larvae, until all larvae had died or emerged. The number of dead larvae and the likely cause of 
the death were also noted. Pupae were checked for parasitism starting from day 10. Dead and 
missing larvae were classified according to the likely cause of death using the following criteria 
(Pomerinke 1999, Amalin et al. 2002, Xiao et al., 2007): (a) Spiders: spiders were observed to 
puncture immobile larvae over mines, sucking the body fluid or making a slit in the mine to 
remove larvae; (b) Ants: ants were observed to remove larvae from the mine through a small 
hole in the leaf cuticle, or through the back of the leaf over the pupa chamber; (c) Other crawling 
predatory insects (e.g., lacewing and lady beetle larvae) were observed to feed on  body parts of 
P. citrella larvae without removing the entire cadaver; (d) Parasitism by ecto-parasitoids (e.g., 
Cirrospilus spp.): presence of larval or pupal parasitoid inside the mine or pupal chamber of P. 
citrella; (e) Parasitism by  endo-parasitoids, (e.g., A. citricola): presence of multiple parasitoid 
pupae within the prepupae and inside pupal chamber of P. citrella; and (f) Physical mortality: 
absence of much of the cuticle over the mine.  
 
 
 102 
Statistical Analysis 
Daily survivorship was calculated for larvae in the different treatments. Percentage 
cumulative larval mortality (at the end of the experiment) was calculated for each treatment (by 
mortality factor) and used for statistical analysis. Data obtained were first normalized by using 
the arsine square-root transformation (? x + 0.5) and then analyzed with one-way analysis of 
variance (ANOVA) followed by the Tukey-Kramer honestly significant difference (HSD) test to 
determine significant treatments effects (P < 0.05, JMP Version 7.01, SAS Institute, 2007). Data 
for each orchard and year were analyzed and presented separately. 
 
RESULTS 
Daily survivorship of P. citrella larvae in the control and key exclusion treatments 
evaluated in 2007 and 2008 are shown in Figures 1 and 2. As expected, survivorship of larvae on 
unprotected (control) branches was lower than larval survival in the complete sticky barrier and 
cage exclusion treatments in which larvae were protected for the entire duration of the study. In 
general, significant differences in larval survival in the control versus exclusion treatments were 
recorded at GCREC orchard beginning as early as day 2 or 3 of the experiment. In 2007 at Coker 
orchard, however, similar larval survival rates were recorded from day 1 to day 6 of the 
experiment in the control versus the complete (1-12 days) sticky barrier treatment, after which 
larval survival in the control began to reduce significantly (Fig. 1A). The complete (1-12 days) 
cage barrier treatment was evaluated in 2008, which allowed for a comparison between the two 
complete exclusion (sticky barrier versus cage barrier) techniques. At both locations, daily larval 
survivorship was significantly greater in the complete cage barrier treatment than in the complete 
sticky barrier treatment beginning from day 2 until the end of the experiment (Fig. 2). For 
 
 103 
instance, percent survival of larvae on day 12 in the complete cage barrier treatment was ~ 92 % 
in both orchards compared with ~ 65-67 % in complete sticky barrier treatment and < 50 % in 
the control. These results suggest that the cage exclusion method was more effective in 
excluding key natural enemies, and consequently in reducing larval mortality. 
Percentage cumulative mortalities of P. citrella larvae recorded at the end of the 
experiment (12 days) in the different treatments are presented in Tables 1-4. In general, 
significant differences in mortalities were recorded among the treatments, with the highest 
mortalities recorded in the control. Overall mortality in the control was ~ 39-41 % in 2007 
(Tables 1 and 2) and ~ 48-52 % in 2008 (Tables 3 and 4) in both orchards. In both orchards in 
2007 (Tables 1 and 2), predation by spiders was the single most important mortality factor 
accounting for ~ 25-30 % mortality, whereas parasitism accounted for only ~ 1-3 % mortality in 
the control. All five exclusion (sticky barrier and cage barrier) treatments significantly reduced 
predation by spiders, which also resulted in significant reductions in overall larval mortalities 
compared with the control. However, the effects of the other mortality factors (i.e. ants, other 
crawling predatory insects and physical damage) were not significantly different between the 
control and any of the exclusion treatments. Furthermore, no significant differences in total 
larval mortalities were recorded among the five exclusion treatments, irrespective of whether the 
larvae were protected early (1-5 days), later (6-12 days) or throughout the experiment (1-12 
days). Comparison among the three sticky barrier treatments showed that only the complete (1-
12 days) sticky barrier treatment was very effective in excluding ants.  
In 2008 at Coker orchard, spiders also were the most important mortality factor which 
accounted for ~ 25% mortality in the control. In contrast, predation by spiders was completely 
excluded (0%) in the complete (1-12 days) cage barrier treatment, and significantly reduced (~ 
 
 104 
13%) in the later (6-12 days) cage barrier treatment, but not significantly reduced in the other 
treatments compared with the control (Table 3). Similarly, predation by ants and other predatory 
insects was completely (0%) excluded in the complete (1-12 days) cage barrier treatment. 
Predation by ants was also completely excluded (0%) in the complete (1-12 days) sticky barrier 
treatment. Parasitism was generally low ranging from 0% in the later cage barrier treatment to ~ 
5 % in the control. Compared with the control, overall mortality was significantly reduced in 
four of the five exclusion treatments, with the early (1-5 days) sticky barrier treatment being the 
exception. This suggests that the effect of predation by ants was greatest on early instar larvae. 
Among the three cage barrier treatments, the complete cage barrier treatment was the only 
treatment which effectively excluded spiders and ants (Table 3). No consistent differences were 
recorded between the early and the later cage barrier treatments, suggesting that P. citrella larvae 
are susceptible to predation by spiders and ants throughout their development. In general, similar 
results were recorded at the GCREC orchard: overall mortality was significantly reduced in five 
of the six exclusion treatments. The lowest mortality (~ 8 %) was recorded in the complete cage 
barrier treatment compared with 35 % mortality in the complete sticky barrier treatment and 
48 % mortality in the control (Table 4). 
 
DISCUSSION  
The results from both locations and years clearly showed that predation was the most 
significant biological effect, which accounted for ~ 87-96 % of all P. citrella larval mortalities in 
unprotected (control) trees in Alabama satsuma orchards. In particular, predation by spiders was 
the single most important natural mortality factor acting on early and late instars of P. citrella. 
The key predacious spider species recorded in our study included the yellow glost spider, Hibana 
 
 105 
sp. (Anyphaenidae), the long-leged sac spider, Cheiracanthium sp. (Miturgidae), and Hentzia sp. 
(Salticidae). Several authors have also reported on the role of spiders and insect predators as a 
key or dominant natural mortality factor acting on P. citrella in many parts of the world (Amalin 
et al. 1996, 2001, 2002; Browning and P?na 1995, Argov and Rosler 1996, Xiao et al. 2007, Hoy 
et al. 2007). Three species of sac spiders, Chiracanthium inclusum (Hentz) (Clubionidae), 
Hibana velox (Becker) (Anyphaenidae), and Trachelas volutes Gertsch (Corrinnidae) were 
reported to feed on P. citrella larvae and prepupae in Florida lime orchards (Amalin et al. 1996, 
2001). Further studies on their predatory habit showed that they are nocturnal species with the 
ability to detect their concealed prey by sensing movements (vibrations) of P. citrella larvae and 
prepupae with the leaf epidermis (Amalin et al. 2001). Prey preference studies also confirmed 
that certain predacious spider species prefer to feed on Lepidopteran and Homopteran pests in 
orchards (Jackson et al. 1977, Nyffeler et al. 1987, Amalin et al. 2001, Brown et al. 2003, 
Stephen and Berg 2008). Spiders are known to feed on P. citrella in two ways. They may 
directly puncture mines or remove the prey through open slit in the mines (Amalin et al. 2001, 
Xiao et al. 2007).  Thus, the specialized predatory habit and feeding behavior of spiders make 
them important predators of P. citrella and similar pests. In addition to directly feeding on the 
immature stages, spiders could disturb the larvae and cause them to drop from the plant, and the 
webs spun over the leaves may make them less suitable for oviposition and feeding by pests 
(Stephen and Berg 2008).  
Predation by ants was the second most important mortality factor, which accounted for ~ 
10-19 % of all P. citrella deaths on unprotected (control) branches. The key ant species recorded 
included Solenopsis invicta Buren and Brachymyrmex patagonicus Mayr, and both species were 
observed feeding on P. citrella larvae. Ants have also been reported as important predators of P. 
 
 106 
citrella in many parts of the world (Huang et al. 1989, Pomerinke 1999, Amalin et al. 2001, Xiao 
et al. 2007), but their impact appears to vary by region or climate. For instance, ants were the key 
predators of P. citrella in China, in particular during the dry and hot summer and fall seasons 
(Huang et al. 1989).  Also, predation by ants, such as Pseudomyrmex gracilis (Roger) and 
Crematogaster ashmeadi (Mayr), was the largest single cause of P. citrella mortality which 
accounted for > 30% of all mortality by natural enemies in a southwest Florida citrus grove 
(Xiao et al. 2007). In contrast, Urbaneja et al. (2004) reported no significant effect of ant 
exclusion on mortality of P. citrella in Spain. Ants have been observed to remove P. citrella 
larvae through a small hole made in the mine, resulting in missing cadavers (Amalin et al. 2001, 
Pomerinke 1999, Xiao et al. 2007). Predation by other predatory insects, such as larvae of 
lacewings (Chrysoperia sp.) and the multicolored Asian lady beetle (H. axyridis) accounted for ~ 
3-27 % of all P. citrella mortalities, as has been reported by other authors (Chen et al. 1989, 
Amalin et al. 2002).  
Two important parasitoids of P. citrella, Cirrospilus sp. and A. citricola, were recorded in 
this study. Cirrospilus spp., are indigenous generalist ecto-parasitoid of P. citrella in the U.S. 
(Pena 1996, Schauff et al. 1998), while A. citricola is an introduced parasitoid of P. citrella in 
Florida and Texas (Hoy and Nguyen 1997, Pomerinke and Stansly 1998). However, parasitism 
contributed only minimally (~ 0-10 %) to P. citrella mortality in the present study. Several 
authors have reported low to moderate rates of parasitism of P. citrella larvae in the field (Pe?a 
et al., 1996, Legaspi et al. 2001, Amalin et al. 2002, Diez et al. 2006, Xiao et al. 2007). Legaspi 
et al. (2001) recorded ~ 20 % parasitism in Mexico, with the dominant parasitoid being 
Zagrammosoma multilineatum (Ashmead) (Hymenoptera: Eulophidae). The introduced 
parasitoid, A. citricola was the dominant parasitoid recorded in southwest Florida, and accounted 
 
 107 
for ~ 8 - 29 % of P. citrella natural mortality (Xiao et al. 2007). We are not aware of any 
purposeful introduction of A. citricola in Alabama, and its occurrence is likely due to accidental 
introductions through movement of rootstock materials from Florida or Texas, or range 
expansion by natural dispersal. The relatively minor impact of parasitism recorded in the present 
study may simply be a reflection of the low endemic population densities of the identified 
parasitoids in Alabama citrus orchards. Since A. citricola has been reported to perform well in 
humid regions (Neale et al. 1995, Hoy and Nguyen 1997, Xiao et al. 2007), field augmentation 
of this parasitoid species is likely to be successful in the humid southern Alabama conditions. 
The sticky barrier and cage barrier exclusion methods evaluated in this study have been 
commonly used to assess the effectiveness and impact of natural enemies in the field 
(Grabenweger et al. 2005, Pomerinke 1999, Xiao et al. 2007, Qureshi and Stansly 2009). Our 
results which showed that predation by spiders was completely excluded in the complete cage 
barrier treatment, and reduced in the complete sticky barrier treatment, are not surprising given 
their behavior on trees (Stephen and Berg 2008). Spiders could move from branch to branch by 
ballooning and thus would not be that affected by a sticky barrier. As expected, predation by ants 
was excluded or significantly reduced in the complete sticky barrier treatment. Interestingly, 
predation by ants was also excluded or reduced in the cage barrier treatment. Similarly, key 
predatory larvae were also excluded or reduced in the complete cage barrier treatment but the 
numbers were sometimes too low to detect a significant effect. These results possibly indicate 
that the mesh size of the cage barrier was fine enough to exclude some ants (e.g., major ant 
workers) and predatory larvae. In general, the timed (early or later) barrier treatments were not as 
effective in excluding or reducing any of the key predatory arthropods. Also, parasitism rates 
were too low to detect a significant effect of the exclusion treatments. 
 
 108 
In summary, our results showed that predation by spiders and ants are very important 
natural mortality factors acting on P. citrella in Alabama citrus. Conservation of these key 
predators through the judicious use of pesticides and augmentation of field populations of key 
natural enemies are essential to the development of a sustainable pest management strategy for 
the pest in Alabama. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 109 
ACKNOWLEDGEMENTS 
We thank Mr. Monte Nesbitt (Gulf Coast Research and Extension Center, Fairhope, AL) 
for field assistance, and Drs. Art Appel, Charles Ray and Elina Coneva (Auburn University) for 
helping to review an earlier version of this paper. We also thank our commercial satsuma grower 
cooperators in southern Alabama. Funding for this study was provided through grants by the 
Alabama Agricultural Experiment Station and Auburn University Horticulture Line Item grants 
program to HYF. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 110 
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 116 
Table 1.  Mortality of Phyllocnistis citrella cohorts from first stage larvae to adult emergence on unprotected (control) 
branches of satsuma trees versus branches protected with different exclusion techniques (sticky barrier or cage) during 
different stages of development at Coker orchard in 2007.   
Exclusion treatments Percentage mortality (mean ? SE) due to each mortality factor  
Spiders Ants Other predators Parasitoids Physical Total mortality 
Control 29.3 ? 4.9a 6.7 ? 2.1 1.3 ? 1.3  2.7 ? 1.6 1.3 ? 1.3 41.3 ? 5.7a 
Complete sticky barrier  13.3 ? 1.6b    0 ? 0 6.7 ? 2.1  1.3 ? 1.3 2.7 ? 1.6 24.0 ? 3.4b 
Early sticky barrier 13.3 ? 0b 1.3 ? 1.3 5.3 ? 1.6  1.3 ? 1.6 1.3 ? 1.6 22.7 ? 1.6b 
Later sticky barrier   9.3 ? 1.6b 2.7 ? 1.6 4.0 ? 1.6  2.7 ? 1.6 2.6 ? 1.6 21.3 ? 2.5b 
Early cage barrier   9.3 ? 1.6b 1.3 ? 1.3 4.0 ? 1.6  4.0 ? 1.6 2.7 ? 1.6 21.3 ? 1.3b 
Later cage barrier 10.7 ? 1.6b 4.0 ? 2.6 1.3 ? 1.3  1.3 ? 1.6 2.7 ? 1.6 20.0 ? 3.0b 
ANOVA (F, P)  
df = 5, 24 
F = 10.7,    
P = 0.0001 
F = 1.96, 
P = 0.124 
F = 1.81, 
P = 0.14 
F = 0.24,  
P = 0.94 
F = 0.2, 
P = 0.95 
F = 4.99, 
P = 0.002 
 
 
 
 
 
 117 
Means in the same column followed by the same or no letter are not significantly different (P > 
0.05, Tukey-Kramer HSD test). In this and subsequent tables, complete sticky barrier = branch 
protected with sticky barrier from day 1 to day 12; early sticky barrier = branch protected with 
sticky barrier from day 1 to day 5; later sticky barrier = branch protected with sticky barrier from 
day 6 to day 12; complete cage barrier = branch caged from day 1 to day 12; early cage barrier = 
branch caged from day 1 to day 5; later cage barrier = branch caged from day 6 to day 12; and 
Control = branch not protected with sticky barrier or cage. Other predators = other crawling 
predacious insects (e.g., lacewing and lady beetle larvae). 
 
 118 
Table 2. Mortality of Phyllocnistis citrella cohorts from first stage larvae to adult emergence on unprotected (control) branches 
of satsuma trees versus branches protected with different exclusion techniques (sticky barrier or cage) during different stages 
of development at GCREC orchard in 2007.   
Exclusion treatments Percentage mortality (mean ? SE) due to each mortality factor  
Spiders Ants Other predators Parasitoids Physical Total mortality 
Control 25.3 ? 2.5a 4.0 ? 1.5 5.3 ? 1.3 1.3 ? 1.3 2.7 ? 1.3 38.6 ? 3.2a 
Complete sticky barrier  14.7 ? 2.5b 0 ? 0 5.3 ? 1.3 0 ? 0 2.7 ? 1.3 22.7 ? 2.6b 
Early sticky barrier 12.0 ? 1.5b 4.0 ? 1.6 8.0 ? 2.5 0 ? 0 2.7 ? 1.3 26.7 ? 3.6b 
Later sticky barrier   10.0 ? 1.6bc 2.7 ? 1.6 8.0 ? 1.3 5.3 ? 2.5 2.7 ? 1.1 28.7 ? 2.6b 
Early cage barrier   9.3 ? 2.6b 2.7 ? 1.6 5.3 ? 1.3 0 ? 0 2.7 ? 1.3 20.0 ? 2.9b 
Later cage barrier    8.0 ? 1.3bc 2.7 ? 1.6 5.3 ? 2.5 2.7 ? 2.6 4.0 ? 1.6 22.7 ? 3.4b 
ANOVA (F, P)  
df = 5, 24 
F = 10.4,    
P= 0.0001 
F =0.96, 
P = 0.46 
F = 0.58,    
P = 0.71 
F = 1.0,        
P = 0.44 
F = 0.11,  
P = 0.98 
F = 4.46,          
P = 0.005 
Means in the same column followed by the same or no letter are not significantly different (P > 0.05, Tukey-Kramer HSD test). 
GCREC = Gulf Coast Research and Extension Center, Fairhope, AL.
 
 119 
Table 3.  Mortality of Phyllocnistis citrella cohorts from first stage larvae to adult emergence on unprotected (control) 
branches of satsuma trees versus branches protected with different exclusion techniques (sticky barrier or cage) during 
different stages of development at Coker orchard in 2008.   
Exclusion treatments Percentage mortality (mean ? SE) due to each mortality factor  
Spiders Ants Other predators Parasitoids Physical Total mortality 
Control 25.3 ? 1.7a   9.7 ? 1.8ab 10.0 ? 1.3ab   5.0 ? 3.2   1.6 ? 0.8 51.6 ? 1.7a 
Complete sticky barrier  21.7 ? 1.7a      0 ? 0c   8.3 ? 1.7ab   3.3 ? 0.5      0 ? 0 33.3 ? 3.8b 
Early sticky barrier 18.3 ? 1.7ab   1.7 ? 0.8 13.3 ? 1.9a   3.3 ? 3.3   1.7 ? 0.9 38.3 ?3.2ab 
Later sticky barrier 18.3 ? 2.3ab 10.0 ? 1.6ab   3.3 ? 1.9bc   5.0 ? 1.7      0 ? 0 36.6 ? 3.3b 
Complete cage barrier      0 ? 0c      0 ? 0c      0 ? 0c   1.6 ? 0.5   6.7 ? 2.7   8.3 ? 3.2c 
Early cage barrier 15.0 ? 1.7a      0 ? 0c   3.3 ? 1.3bc   6.7 ? 1.9   3.3 ? 1.9 28.3 ? 7.4b 
Later cage barrier 13.3 ? 1.7b  6.7 ? 2.7bc   5.0 ? 0.5ab      0 ? 0   1.7 ? 0.8 26.7 ? 2.7b 
ANOVA (F, P)  
df = 6, 21 
F = 47.7,      
P = 0.0001 
F = 12.5,   
 P = 0.0001 
F = 6.03,   
 P = 0.009 
F = 0.69,    
P = 0.65 
F = 1.87, 
P = 0.13 
F = 11.05,  
P = 0.0001 
Means in the same column followed by the same or no letter are not significantly different (P > 0.05, Tukey-Kramer HSD test).   
 
 120 
Table 4.  Mortality of Phyllocnistis citrella cohorts from first stage larvae to adult emergence on unprotected (control) 
branches of satsuma trees versus branches protected with different exclusion techniques (sticky barrier or cage) during 
different stages of development at GCREC orchard in 2008   
Exclusion treatments Percentage mortality (mean ? SE) due to each mortality factor  
Spiders Ants Other predators Parasitoids Physical Total mortality 
Control 25.0 ? 3.2a  8.3 ? 3.2a 13.3 ? 2.3a     0 ? 0 1.7 ? 1.7 48.3 ? 1.7a 
Complete sticky barrier  20.0 ? 2.7a     0 ? 0b    10 ? 1.9ab  3.3 ? 1.9 1.7 ? 1.7 35.0 ? 1.7ab 
Early sticky barrier 18.3 ? 1.7a  3.3 ? 1.9b   5.0 ? 1.7bc     0 ? 0 3.3 ? 1.9 29.9 ? 3.3b 
Later sticky barrier 18.3 ? 1.7a  6.7 ? 0a   6.7 ? 2.7bc     0 ? 0    0 ? 0 31.7 ? 3.2b 
Complete cage barrier      0 ? 0b     0 ? 0b      0 ? 0c  3.3 ? 1.9 5.0 ? 1.7   8.3 ? 3.2c 
Early cage barrier 16.7 ? 1.9a  8.3 ? 1.7a   8.3 ? 1.7ab     0 ? 0    0 ? 0  33.3 ? 2.7b 
Later cage barrier 18.3 ? 1.7a  6.7 ? 2.7a   3.3 ? 1.9bc     0 ? 0 1.7 ? 1.7  30.0 ? 4.3b 
ANOVA (F, P)  
df = 6, 21 
F = 14.4,  
P = 0.0001 
 F= 4.18, P 
= 0.006 
F= 6.68, 
P=0.0005 
 F = 2.5,  
P = 0.054 
 F = 1.5,  
P = 0.225 
 F = 15.5,  
P = 0.0001 
Means in the same column followed by the same or no letter are not significantly different (P > 0.05, Tukey-Kramer HSD test). 
GCREC = Gulf Coast Research and Extension Center, Fairhope, AL.
 
 121 
Figure 1.  Survivorship curve of P. citrella immature stage over whole development periods in 
natural and protected condition in the two orchards in 2007.  Figure showed mean (? SE) 
numbers of P. citrella. A: Coker and B: GCREC (Gulf Coast Research and Extension Center, 
Fairhope, AL). 
0 2 4 6 8 10 12 14
Per
cen
t 
su
rv
ivo
rsh
ip 
(m
ea
n 
?
 S
E) 
50
60
70
80
90
100
110
Contro l 
Com plete  sti ck y  ba rr ie r
C o ho rt  a g e (d a y s)
0 2 4 6 8 10 12 14
50
60
70
80
90
100
110
Contro l
Com plete  sti ck y  ba rr ie r
A:  Co k er  
B : GCR E C
 
 
 
 122 
Figure 2.  Survivorship curve of P. citrella immature stage over whole development periods in 
natural and protected condition in the two orchards in 2008. Figure showed mean (? SE) 
numbers of P. citrella. A: Coker and B: GCREC (Gulf Coast Research and Extension Center, 
Fairhope, AL). 
 
A:  Cok er  
0 2 4 6 8 10 12 14
Percent s
urv
ivo
rship (
mea
n ?
 SE)
40
50
60
70
80
90
100
110
Cont ro l 
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0 2 4 6 8 10 12 14
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CHAPTER 6   
FUNCTIONAL RESPONSE AND PREY STAGE PREFERENCES OF THREE SPECIES 
OF PREDACEOUS MITES (ACARI: PHYTOSEIIDAE) WHEN OFFERRED CITRUS 
RED MITES, PANONYCHUS CITRI (ACARI: TETRANYCHIDAE) 
 
INTRODUCTION 
Citrus red mite, Panonychus citri (McGregor) (Acari: Tetranychidae), is a key pest of 
citrus in many parts of the world (Gotoh and Kubota 1997, Jamieson et al. 2005, Childers et al. 
2007). The adults and immatures feed primarily on leaves producing tiny grey or silvery spots 
known as stippling damage, which may inhibit photosynthesis (Kranz et al. 1977). High 
infestations can result in premature leaf drop, shoot dieback and decreased plant vigor, as well as 
fruit feeding and damage (Kranz et al. 1977, Jamieson et al. 2005). Panonychus citri is a major 
pest of satsuma mandarin (Citrus unshiu Marcovitch.) in Alabama (English and Turnipseed 
1940, Fadamiro et al. 2007, 2008). Satsuma mandarin production is a growing industry in 
southern Alabama and other parts of the Gulf Coast region of the United States (Campbell et al. 
2004). The results of recent field surveys identified P. citri as one of the most numerically 
abundant pests in Alabama satsuma orchards (Fadamiro et al. 2007, 2008). Two to three 
generations per year were recorded for P. citri in southern Alabama, and the pest was most 
abundant in the spring (Fadamiro et al. 2008), when population densities were typically greater 
than the economic threshold of five motiles per leaf, proposed by Childers et al. (2007). The 
 
 124 
results also showed that P. citri was more abundant in the exterior canopy than in the interior 
canopy, and infestations were more severe in conventionally sprayed orchards than in unsprayed 
orchards (Fadamiro et al. 2008).  
Predacious mites, in particular those belonging to the families Phytoseiidae and 
Stigmaeidae, have been widely used for biological control of pest mites in fruits and other crops 
worldwide (Childers et al. 1975, McMurtry 1983, Childers 1994, Wood et al. 1994, Opit et al. 
2004, Jamieson et al. 2005, Carmak et al. 2009, Authur et al. 2009), and may present an 
alternative to chemical control of P. citri in Alabama. A recent survey of the predacious mite 
fauna of satsuma mandarin in Alabama identified a total of 29 species from nine families, 
including 18 species in the family Phytoseiidae and one species in the family Stigmaeidae 
(Fadamiro et al. 2009). The dominant predacious mite species recorded were Typhlodromalus 
peregrinus Muma and Proprioseiopsis mexicanus (Garman) (Phytoseiidae), and Agistemus 
floridanus Gonzalez (Stigmaeidae) (Fadamiro et al. 2009). However, these predacious mites 
were recorded in the orchards at densities too low for effective suppression of P. citri (Fadamiro 
et al. 2009). Furthermore, recent attempts to mass rear T. peregrinus and other indigenous 
predacious mites for augmentative releases against P. citri have not been successful (Xiao and 
Fadamiro, unpublished data), prompting our interest in the evaluation of some commercially 
available phytoseiids (e.g., Phytoseiulus persimilis, Galendromus occidentalis, and Neoseiulus 
californicus) as potential biological control agents of P. citri. An assessment of the functional 
and numerical responses and prey-stage preferences of these phytoseiids when offered P. citri is 
a critical first step in determining their ability to regulate the prey.  
The functional response concept first described by Holling (1959) has been widely 
utilized to evaluate effectiveness of predacious mites (Laing and Osborn 1974, Everson 1980, 
 
 125 
Sabelis 1985, Badii et al. 2004, Reis et al. 2003, 2007).  In general, the functional response of a 
predator to a prey could follow one of three mathematical models: linear, convex or sigmoid. In 
the linear (type I) model, the number of prey consumed increases linearly with prey availability 
up to a maximum. In the convex (type II) model, the number of prey consumed increases with 
prey availability but begins to decrease when a maximum point is reached. In the sigmoid (type 
III) model, the relationship between numbers of prey consumed and prey availability follow a 
sigmoid form up to a maximum (Holling 1959, 1961).  This study was carried out to determine 
the potential of three commercially available phytoseiid species, P. persimilis, G. occidentalis 
and N. californicus, as biological control agents of P. citri by evaluating their functional 
responses and prey-stage preferences. Additional experiments were conducted to test the effect 
of starvation on the predation potential and prey-stage preferences of the three phytoseiid 
species.  
 
MATERIALS AND METHODS 
Rearing of P. citri (the prey) 
The starting colony of P. citri used as prey was collected from satsuma orchards in 
southern Alabama in spring 2008. Mixed stages of P. citri were reared on lima bean (Phaseolus 
lunatus L.) and/or satsuma leaves in transparent plastic containers (25 ? 10 ? 7 cm) with the top 
covered by screening net. Each container had about 10 fully expanded P.citri-infested lima bean 
or satsuma leaves placed ventral side up on a moistened paper towel, and then placed over a 
water-saturated sponge. This arrangement prevented mite escape and maintained leaf freshness 
for up to two weeks. The moistened paper towel and sponge were kept wet by adding water as 
 
 126 
necessary. All rearing and experiments were conducted under laboratory conditions at 25 ? 2?C, 
60 ? 10% RH and a photoperiod of 14:10 (L: D) h. 
 
Rearing of Predacious Mites  
The starting colonies of all three phytoseiid species (P. persimilis, G. occidentalis, and N. 
californicus) were purchased from Biocontrol Network, Inc., (Brentwood, TN) and reared on 
mixed stages of P. citri on satsuma leaves for two generations before the bioassays. The rearing 
method was as described above for P. citri. Adult females (~ 2 day old) of each species 
previously starved for 8 h were used for the bioassays. All experiments were conducted in 2008. 
 
Experiment 1. Functional Response  
The functional response of each of the three phytoseiid species on P. citri was 
investigated in separate bioassays using a protocol similar to that described by Reis et al. (2003).  
For each species, an adult female was confined for seven days in a 3-cm diameter satsuma leaf 
disc arena, placed on a moistened filter paper and over a thin wet sponge, inside a Petri dish (8.0 
cm diameter ? 1.5 cm depth). P. citri nymphs (motile immatures) were introduced as prey onto 
the leaf disc inside the Petri-dish at densities of 2, 5, 10, 20, 30, 40, 50, 75, 100, and 150 per 
arena. The Petri dish was then sealed with parafilm to prevent mite escape. Each density 
treatment was replicated four times.  The controls consisted of arenas with the same densities of 
P. citri nymphs but without predacious mite. The numbers of P. citri larvae killed and eggs laid 
by the predator were recorded daily for seven days. To maintain appropriate P. citri densities in 
the arenas, dead P. citri were replaced with live ones daily. 
 
 
 127 
Experiment 2. Prey-Stage Preference 
Experiments were conducted to determine prey stage (egg or nymph) is preferred by each 
phytoseiid species. The experimental protocol used was similar to that described by Blackwood 
et al. (2001).  For each species, an adult female was confined in a 3-cm diameter satsuma leaf 
disc arena (as described above) with P. citri eggs and nymphs at one of five ratios: 0:1, 1:0, 1:1, 
1:2 and 2:1 eggs: nymphs. The last two ratios (1:2 and 2:1) were tested to evaluate prey-stage 
switching. A total of 60 prey per arena was tested for the first three ratios and 90 prey per arena 
for the 1:2 and 2:1 ratios. The experiment was replicated 10 times per ratio per species. The 
numbers of each prey-stage consumed by the predator were recorded after 24 h.  
 
Experiment 3. Effect of Starvation 
Using the protocol described above, an adult female of each phytoseiid species was 
starved (held without food) for 0, 24, or 48 h before tests, and then introduced into a satsuma leaf 
arena containing 1:1 ratio of eggs and nymphs of P. citri (60 total prey per arena). The numbers 
of each prey stage consumed were recorded after 24 h. The experiment was replicated 10 times 
per species. 
 
Preference Quantification Methods 
 Prey-stage preferences were quantified with the index ? (Manly et al., 1972), as 
described by Blackwood et al. (2001). 
1
1)/l n ( )'/'l n (
c
c
NN
NN  
 
 128 
N (eggs) and N? (larvae) are the numbers of each prey type provided, and Nc (eggs) and 
Nc? (larvae) are the numbers of each prey type consumed. This index assigns preference values 
from 0 to 1, where values 0.5 represent no preference. The ? value was calculated for each 
replicate and averaged to determine the mean ? value for each treatment.  
 
Statistical Aanalysis  
Data from the functional response experiment were first analyzed using regression 
analysis to determine the best fit model (Sigmplot 8, Systat Software, 2007). Data were then 
normalized by using the square-root transformation (? x + 0.5) and analyzed with analysis of 
variance (ANOVA) followed by Tukey-Kramer honest significant difference (HSD) test to 
determine significant differences among the three species (P < 0.05, JMP Version 7.0.1, SAS 
Institute Inc., 2007). Data from the experiments on prey-stage preference and effect of starvation 
were first normalized by using square-root transformation (? x + 0.5) and then analyzed with 
ANOVA. Differences among treatments in mean numbers of prey killed or mean preference 
index (?) were determined by using the Tukey-Kramer HSD test or Student?s t-test (P < 0.05, 
JMP Version 7.0.1, SAS Institute Inc., 2007).   
 
RESULTS 
Experiment 1. Functional Response 
The functional responses of the three phytoseiid species on P. citri are shown in Table 1 
and Fig.1. Natural mortality of P. citri observed in the control was minimal: 4.6%, 9.3 %, and 
6.7 % for P. persimilis, G.occidentalis, and N. californicus, respectively. This suggests that 
 
 129 
predation by the predacious mites was the dominant source of P. citri mortality recorded in this 
experiment. 
P. persimilis. The number of P. citri killed per day increased from ~ 0.2 at a prey density 
of 2 per arena to a maximum of ~ 14 (~ 19 % predation) at a prey density of 75 per arena (Table 
1, Fig.1A). The percentage of prey killed (or percentage of predation) ranged from ~ 11 % at the 
lowest prey density (2 per arena) to 24 % at the 5 prey per arena density. The generally low 
percentage of predation suggests that most of the prey provided were not consumed by the 
predator. The lowest percentage of predation recorded at the lowest prey density was probably 
related to the difficulty in finding the prey at very low densities. A plot of the relationship 
between number of prey provided and number killed by female P. persimilis revealed a positive 
and highly significant correlation (R2 = 0.9467; F = 151.06; P = 0.0001; Fig. 1A). The number of 
prey killed increased as a function of prey density up to a  maximum point (75 prey per arena) 
and then started to decrease, resulting in a convex (type II) functional response (Fig. 1A). 
 G. occidentalis. The number of P. citri killed per day also rose as a function of prey 
density from ~ 1 at a prey density of 2 to a maximum of 12 (~ 17 % predation) at a prey density 
of 75 (Table 1, Fig. 1B). The percentage of predation ranged from ~ 7 % at the highest prey 
density (150 per arena) to 37 % at a prey density of 10 per arena. The number of P. citri killed 
was positively correlated to the number provided (R2 = 0.921; F = 41.23; P = 0.0001), and was 
expressed by a convex (type II) functional response curve (Fig.1B).  
N. californicus. A similar trend was observed in the functional response of N. 
californicus on P. citri. The number of prey killed per day increased with density from 0.3 at a 
prey density of 2 per arena to a maximum of ~ 10 (~10 % predation) at a prey density of 100 
(Table 1, Fig.1C). The percentage of predation ranged from ~ 6 % at the highest prey density 
 
 130 
(150 per arena) to 22 % at a prey density of 10 per arena.  In general, predation by N. 
californicus was slightly lower than predation by P. persimilis and G. occidentalis. However, N. 
californicus demonstrated similar predation efficiency as the other two species at the lower prey 
densities of 30 or less per arena (Table 1 and Fig.1C). As with the other two species, a positive 
association (R2 = 0.99; F = 874.1; P = 0.0001) was recorded between the number of prey killed 
and the number provided up to a maximum point (100 prey per arena), which resulted in a 
convex (type II) functional response (Fig. 1C). 
 
Experiment 2. Prey-Stage Preference 
Prey-stage preferences of the three predacious mite species are summarized in Table 2 
and Fig. 2. When eggs and nymphs of P. citri were simultaneously presented, all three species 
fed on both stages but highly preferred nymphs, irrespective of the prey ratio (Table 2). This 
result was confirmed by the very high (> 0.65) preference index (?) recorded for all three species 
(Fig. 3): ? values greater than 0.5 represent preference for nymphs while those less than 0.5 
represent preference for eggs (Blackwood et al., 2001). When only eggs were provided, P. 
persimilis consumed more eggs (3.4 eggs) than G. occidentalis (2 eggs) and N. californicus (0.4 
eggs). Phytoseiulus persimilis also consumed more nymphs than the other two species, when 
only this prey stage was provided (Table 2). Similar results were recorded when the number of 
each prey stage consumed was pooled across all ratios and compared among the three species. 
Phytoseiulus persimilis consumed significantly greater number of nymphs (F = 45.7; df = 2, 87; 
P = 0.0001) than the other two species, and also significantly greater number of eggs than N. 
californicus (F = 5.7; df = 2, 87; P = 0.0047) (Fig. 2). Also, the total number of prey (eggs + 
nymphs) consumed by P. persimilis was significantly greater than that consumed by the other 
 
 131 
two species (F = 51.08; df = 2, 87; P = 0.0001). The preferences of each species when offered 
eggs and nymphs of P. citri at ratios of 1:1, 1:2 and 2:1 were compared to determine prey-
switching behavior, indicated by a significant change in ?. Significant prey-switching was 
recorded for G. occidentalis (F = 5.50; df = 2, 27; P = 0. 0099), and N. californicus (F = 7.31; df 
= 2, 27; P = 0.0029), but not for P. persimilis (F = 0.619; df = 2, 27; P = 0.546). Both G. 
occidentalis and N. californicus exhibited negative switching, indicated by an increased 
preference for nymphs in response to an increase in the relative abundance of eggs (Fig. 3). 
 
Experiment 3. Effect of Starvation 
  No significant effect of starvation was recorded on prey consumption by the phytoseiids 
(P. persimilis: eggs: F = 0.135, P = 0.874, nymphs: F = 1.77, P = 0.189, total prey: F = 1.25, P = 
0.30; G. occidentalis: eggs: F = 2.99, P = 0.067, nymphs: F = 0.51, P = 0.604, total prey: F = 
1.33, P = 0.28; N. californicus: eggs: F = 0.345, P = 0.71, nymphs: F = 2.71, P = 0.084, total 
prey: F = 3.04, P = 0.064; df = 2, 27) (Table 3). In other words, similar numbers of eggs and 
nymphs were consumed in the different starvation treatments. Furthermore, starvation had no 
significant effect on prey-stage preferences of phytoseiids: all three species showed similar 
preference for nymphs, irrespective of starvation duration. These results were also confirmed by 
the lack of significant change in ? with starvation (P. persimilis: F = 0.211, P = 0.81; G. 
occidentalis: F = 0. 965, P = 0.39; N. californicus: F = 0.764, P = 0.48; df = 2, 27) (Fig. 4).  
 
DISCUSSION 
The experimental results demonstrated the predation potential of the three 
 
 132 
phytoseiids, P. persimilis, G. occidentalis, and N. californicus, on P. citri. The functional 
responses of all three species followed the type II convex model in which the number of prey 
consumed increased with prey availability, but began to decrease when a maximum point was 
reached. These results are in agreement with previous studies in which the relationship between 
most predacious mite species and their prey was reported to follow the type II convex functional 
response model (Holling 1961, Mori and Chant 1966, Sandness and McMurtry 1970, Laing and 
Osborn 1974, Santos 1975, Ryoo 1986, Reis et al. 2003, Badii et al. 2004). For instance, Laing 
and Osborn (1974) reported a functional type II response curve for P. persimilis and G. 
occidentalis feeding on Tetranychus urticae Koch. Chant (1961) also reported a type II curve for 
P. persimilis on T. urticae, but observed a type I linear curve for G. occidentalis on the same 
prey. Interference or disturbance by prey has been proposed by various authors to explain the 
functional and numerical responses of predacious mites on pest mites (Mori and Chant 1966, 
Sandness and McMurtry 1970, Reis et al. 2003). For instance, Mori and Chant (1966) suggested 
that high prey densities may result in the disturbance of predacious mites and thus decrease their 
functional or numerical responses. On the other hand, absence of ?interference-stimulation? may 
explain the low predation efficiency recorded at low prey densities since contact with prey may 
have a stimulatory effect on predacious mites (Mori and Chant 1966, Sandness and McMurtry 
1970, Reis et al. 2003).  
Our results on prey-stage preference showed that all three phytoseiids (P. persimilis, G. 
occidentalis, and N. californicus) preferred nymphs to eggs of P. citri, and the nymphs were 
most often the first prey attacked by the predators. To our knowledge, this is the first 
documented evaluation of the predation potential of the three phytoseiids when offered P. citri as 
prey. However, previous studies in which the above or related phytoseiid species were evaluated 
 
 133 
on T. urticae have produced varied results (Burnett 1971, Takafuji and Chant, 1976, Fernando 
and Hassell 1980, Blackwood et al. 2001, 2004). Blackwood et al. (2001) reported that adult 
females of P. persimilis preferred T. urticae eggs over the nymphs, whereas G. occidentalis, and 
N. californicus showed no prey-stage preference. In contrast, Popov and Kondryakov (2008) 
reported that adult females of P. persimilis and G. occidentalis consumed more nymphs and 
males of Tetranychus spp. than the eggs or females. These contrasting results may be related to 
differences in experimental design and number of prey provided. A comparison of our results 
with those in which a Tetranychus spp. was used as prey (e.g., Blackwood et al. 2001, Popov and 
Kondryakov 2008) suggest that prey-stage preference of predacious mites may vary with 
different prey species. Our observed preference of the three phytoseiids for P. citri nymphs over 
eggs may be related to differences in the size and mobility of the two stages. The phytoseiids 
may have preferred the motile nymphs, possibly because they are larger and active, thus 
increasing the possibility of being encountered by a predacious mite (Sabelis 1985). Our results 
may also be explained by possible differences in the nutritional benefits of prey nymphs versus 
eggs. Previous work on the relative nutritional value of T. urticae eggs and nymphs suggest that 
eggs may be a more profitable prey stage for some predacious mites (Burnett 1971, Croft and 
McMurty 1972, McMurty and Rodriguez 1987). For instance, Croft and McMurty (1972) 
showed that G. occidentalis females fed on eggs or nymphs of Tetranychus pacifus McGregor 
consumed twice as many nymphs as eggs but with similar fecundities, suggesting that the 
predator obtained more nutritional benefits from eggs. Thus, only a minimal consumption of 
eggs in a diet of mixed-prey stages may be necessary to achieve maximum fecundity (McMurty 
and Rodriguez 1987), which may explain the minimal consumption of P. citri eggs in our 
experiments. In contrast, consumption of nymphs may actually be more profitable than egg 
 
 134 
consumption. Zaher and Shehata (1971) reported that Typhlodromus pyri Scheuten females had 
higher fecundity when feeding on mobile immatures of Tetranychus cinnabarinus Biosduval 
than on the eggs. Thus, our results may also suggest that P. citri nymphs are more profitable than 
the eggs to all three phytoseiids in terms of nutritional benefit and handling time. While no 
significant prey-switching was recorded for P. persimilis, both G. occidentalis and N. 
californicus exhibited negative prey switching, indicated by an increased preference for nymphs 
in response to an increase in the relative abundance of eggs. Blackwood et al. (2001) also 
reported significant prey-switching for some phytoseiid species on T. urticae, and suggested that 
this phenomenon may have been mediated by fixed preferences for particular ratios of prey 
stages. 
The data which showed no significant effect of starvation on the total number of prey 
consumed by the three phytoseiids or their prey-stage preferences, are in agreement with the 
report by Blackwood et al. (2001), in which starvation had no effect on the prey-stage preference 
of Neoseiulus fallacis (Garman). In contrast, starvation had a significant effect on the foraging 
behavior of P. persimilis (Zhang and Sanderson 1992). The lack of significant changes in the 
foraging behavior of the phytoseiids even after 48 h of food deprivation may suggest that the 
phytoseiids can survive a few days of starvation, or that the risk of starvation-induced death is 
not high after two days of food deprivation (Pratt et al. 1999, Blackwood et al. 2001). The ability 
to survive a few to several days of food deprivation in the field may be an important survival 
strategy for predacious mites, in particular those species with specialized diets. However, food 
deprivation for a day or more may have a negative impact on the fecundity of predacious mites 
(Ohnesorge, 1981), but this was not tested in this study. 
 
 135 
All three phytoseiids shared similar behavior and demonstrated strong potential in 
regulating P. citri. However, P. persimilis appeared to have a greater predation potential than the 
remaining two species. This conclusion is supported by a preponderance of evidence from the 
experiments which pointed to slightly greater predation efficiency for P. persimilis. For instance, 
P. persimilis consumed significantly greater number of prey than the other two species, in 
particular at high prey densities. However, very few eggs were laid by the phytoseiids, which 
made it difficult to compare their reproductive capabilities on P. citri.  
Our results on the predation potential of the three phytoseiids on P. citri may be 
explained by their feeding habits and degree of diet specialization. Predacious mites in the family 
Phytoseiidae can be classified into four categories based on their feeding habits and related 
biological and morphological traits (McMurtry and Croft 1997). Type I phytoseiids (e.g., 
Phytoseiulus spp.) are specialized predators of spider mites, Tetranychus spp. Type II 
phytoseiids are selective predators of Tetranychus species. Examples include Galendromus spp. 
and some Neoseiulus spp. Type III phytoseiids (e.g., Amblyseius spp.) are generalist predators, 
while Type IV phytoseiids (e.g., Euseius spp) are specialized pollen feeders/generalist predators. 
Thus, P. persimilis is a type I phytoseiid, while G. occidentalis, and N. californicus are type II 
phytoseiids (McMurtry and Croft 1997). It is therefore not surprising that P. persimilis, which is 
relatively more specialized on spider mites, showed a slightly greater predation potential than G. 
occidentalis, and N. californicus, which are less specialized type II predators. The relative size of 
the phytoseiids and their prey consumption rates may also play a role. Phytoseiulus species are 
relatively large and both nymphs and adults have a high prey consumption rate compared to most 
species in other genera (Gilstrap and Friese 1985, McMurtry and Croft 1997). In addition, we 
also observed that P. persimilis was more active than G. occidentalis and N. californicus, a 
 
 136 
behavior that may increase its probability of encountering a prey. 
In conclusion, our results demonstrated for the first time the potential of the 
commercially available phytoseiids, P. persimilis, G. occidentalis, and N. californicus, as 
biological control agents of P. citri. Studies are ongoing to evaluate small scale releases of the 
phytoseiids against P. citri in Alabama satsuma orchards.  
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 137 
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Soc.Am. 85, 775 - 783.
 
 143 
Table 1. Predation capacity of three species of predacious mites at different densities of the prey, P. citri under laboratory 
conditions. 
Densities 
/ arena 
Mean ? SE) number or percent per day 
P. persimilis G. occidentalis N. californicus 
No. prey 
killed 
(%) 
 
No. eggs 
laid 
No. prey 
killed 
(%) 
 
No. eggs 
laid 
No. prey 
killed 
(%) 
 
No. eggs 
laid 
2  0.2 ? 0.2 10.7 ? 11.2     0 ? 0 0.9 ? 0.6 46.4 ? 31.0    0 ? 0 0.3 ? 0.3 12.5 ? 14.4   0 ? 0 
5  1.2 ? 0.6 23.7 ? 13.2    0 ? 0 1.7 ? 0.8 34.6 ? 16.1    0 ? 0 0.9 ? 0.3 18.1 ? 6.7   0 ? 0 
10  2.1 ? 0.8  21.3 ? 8.6    0 ? 0 3.7 ? 1.3 37.0 ? 13.2    0 ? 0 2.2 ? 0.8 21.8 ? 7.7    0 ? 0 
20  4.3 ? 0.7  21.7 ? 3.5 0.1 ? 0.1 3.0 ? 1.1  14.8 ? 5.4    0 ? 0 3.3 ? 0.9 16.4 ? 4.6    0 ? 0 
30  4.4 ?  2.3  14.9 ? 7.6 0.6 ? 0.3 4.7 ? 1.1  15.7 ? 3.7    0 ? 0 3.9 ? 0.8 12.9 ? 2.8 0.1 ? 0.1 
40  7.5 ? 1.3  18.8 ? 3.2 0.1 ? 0.1 6.3 ? 3.2  15.8 ? 7.9 0.3 ? 0.2 5.1 ? 1.6 12.8 ? 3.9 0.1 ? 0.1 
50 10.8 ? 2.2  21.6 ? 4.3 0.4 ? 0.3 7.2 ? 1.2  14.4 ? 2.4     0 ? 0 7.0 ? 2.0 14.0 ? 4.0 0.2 ? 0.1 
75 14.3 ? 3.5  19.1 ? 4.7 0.7 ? 0.3 12.4 ? 2.1  16.5 ? 2.8 0.4 ? 0.3 7.6 ? 2.7 10.2 ? 3.6 0.1 ? 0.1 
100 13.8 ? 3.2  13.8 ? 3.2 0.7 ? 0.3 10.2 ? 2.9  10.2 ? 2.9 0.3 ? 0.2 9.8 ? 2.3   9.8 ? 2.3 0.3 ? 0.2 
150 10.7 ? 3.1    7.2 ? 2.1 0.4 ? 0.3 9.9 ? 2.4   6. 6 ? 1.6 0.6 ? 0.3 8.7 ? 1.7   5.8 ? 1.1 0.3 ? 0.3 
 
 144 
Table 2.  Prey-stage preference of three species of predacious mites when offered eggs and 
nymphs of P. citri at different ratios for 24 h under laboratory conditions.  
Species  Ratio  
(Eggs: Nymphs) 
Mean (? SE) number of P. citri killed t test 
t,  P Eggs Nymphs  
P. persimilis 1:0 3.4 ? 0.4 0 ? 0 - 
 0:1 0 ? 0 6.7 ? 0.8 - 
 1:1   0.9 ? 0.3b  7.3 ? 0.5a 10.2, 0.0001 
 1:2   1.2 ? 0.5b   8.1 ? 0.4a 9.07, 0.0001 
 2:1   1.0 ? 0.3b          7.6 ? 0a 9.31, 0.0001 
G. occidentalis 1:0       2.0 ? 0.5  0 ? 0 - 
 0:1 0 ? 0 4.7 ? 0.6 - 
 1:1   0.6 ? 0.3b   4.4 ? 0.5a 7.08, 0.0001 
 1:2   0.5 ? 0.2b   3.7 ? 0.3a 8.01, 0.0001 
 2:1   0.5 ? 0.2b   5.1 ? 0.6a 8.74, 0.0001 
N. californicus 1:0       0.4 ? 0.3 0 ? 0 - 
 0:1 0 ? 0 4.0 ? 0.6 - 
 1:1   0.0 ? 0.0b   1.6 ? 0.4a 9.01, 0.0001 
 1:2   0.3 ? 0.2b   3.0 ? 0.7a 3.68, 0.0001 
 2:1   0.3 ? 0.2b   4.6 ? 0.6a 8.78, 0.0001 
For each species, means in the same row followed by different letters are significantly different 
(P < 0.05, t-test). Note that numbers of eggs versus nymphs consumed were not compared at the 
first two ratios (0:1 and 1:0) 
 
 
 
 
 
 
 145 
Table 3. Effect of starvation on the predation capability of three species of predacious mites 
when offered a 1:1 ratio of P. citri eggs and nymphs. 
PM species Starvation duration 
(h)   
Mean (? SE) number of P. citri killed a 
Eggs Nymphs  Total  
P. persimilis 0 1.4 ? 0.3 5.2 ? 0.4 6.6 ? 0.5 
 24 1.5 ? 0.3 6.5 ? 0.7 8.0 ? 0.5 
 48 1.4 ? 0.5 5.8 ? 0.6 7.2 ? 0.8 
G. occidentalis 0 1.0 ? 0.3 6.0 ? 0.4 7.0 ? 0.6 
 24 0.3 ? 0.2 5.2 ? 0.6 5.5 ? 0.7 
 48 0.3 ? 0.2 5.8 ? 0.8 6.1 ? 0.8 
N. californicus 0 0.5 ? 0.3 4.5 ? 0.6 5.0 ? 0.7 
 24 0.7 ? 0.3 6.3 ? 0.5 7.0 ? 0.5 
 48 0.3 ? 0.2 5.7 ? 0.6 6.0 ? 0.5 
For each species, means in the same column followed by same or no letters are not significantly 
different (P < 0.05, Tukey HSD test). 
 
 
 
 
 
 
 
 
 146 
Figure 1.  Relationship between numbers of P. citri preyed on by a female of P. persimilis (A), 
G. occidentalis (B), or N. californicus (C) and the density of P. citri (nymphs) provided per day. 
For all three species, the data followed the type II convex functional response model in which the 
number of prey consumed increased with prey availability but began to decrease when a 
maximum point was reached.  
A:  P. p er s i m i l i s  
0
2
4
6
8
10
12
14
16
18
y  =  - 0 .0 0 1 1 x
2
 + 0 .2 4 9 x  -  0 .1 2 7 2
R
2
 =  0 .9 4 6 7 , F  =  1 5 1 .0 6 , P  =  0 .0 0 0 1
C:  N. ca l i f o r ni cus
Dens i t i es  o f  P. cit r i  ( nym ph s  /  a re na)
0 20 40 60 80 100 120 140 160
0
2
4
6
8
10
12
y  =  - 0 .0 0 0 7  x
2
 +  0 .1 6 4 4 x  -  0 .0 2 8 6
R
2 
=  0 .9 9 0 9 , F  =  8 7 4 .1 , P  =  0 .0 0 0 1  
B:  G. o cc i d ent a l i s
M
ea
n (
 ?
SE
)
 nu
m
ber 
o
f
 pre
y
 k
i
l
l
ed per
 da
y
 
0
2
4
6
8
10
12
14
y  =  - 0 .0 0 0 8  x
2
 +  0 .1 8 9 6 x  +  0 .5 5 4 9
R
2
 =  0 .9 2 1 , F  = 4 1 .2 3 , P  =  0 .0 0 0 1
 
 
 147 
Figure 2.  Predation potential of three phytoseiid species on P. citri. Figure shows mean (? SE) 
number of eggs, nymphs, and total number of prey killed by P. persimilis, G. occidentalis, and 
N. californicus in 24 h. 
a
2 D Gra ph  2
X D a ta
E g g s Nym ph s T o ta l
M
ea
n 
(?
 S
E) 
nu
mb
er 
o
f 
pr
ey
 k
il
led 
0
2
4
6
8
10
P. p ersimi li s
G. o cc id enta li s
N. ca li fo rn ic us a
b
c
a
ab
b
a
b
c
 
 
 
 
 
 
 
 
 
 
 
 
 148 
Figure 3.  Prey-stage preferences of female P. persimilis (A), G. occidentalis (B), and N. 
californicus (C) when provided varying ratios (1:1, 1:2 or 2:1) of eggs and nymphs of P. citri. 
Figure shows mean (? SE) preference index (?). 
A:  P. p ersimi li s
0 .4
0 .5
0 .6
0 .7
0 .8
0 .9
1 .0
B : G. o cc id enta li s
Pre
fere
nce (mean
 ?
 ?
 S
E) 
0 .4
0 .5
0 .6
0 .7
0 .8
0 .9
1 .0
C:  N. ca li fo rn ic us
Ra tio o ff er ed ( egg s: ny mphs)
1 :2 1 :1 2 :1
0 .4
0 .5
0 .6
0 .7
0 .8
0 .9
1 .0
 
 
 149 
Figure 4.  Effect of starvation on the prey-stage preferences of three species of female P. 
persimilis (A), G. occidentalis (B), and N. californicus (C) when provided varying starvation 
level of 0, 24 h, and 48 h at a ratio (1:1) of eggs and nymphs of P. citri. Figure shows mean (? 
SE) preference index (?). 
A:  P. pe rsimili s
0. 4
0. 5
0. 6
0. 7
0. 8
0. 9
1. 0
B: G. oc ci de ntal is
Pre
fere
nce
 (
me
a
n ? ?
 SE)
0. 4
0. 5
0. 6
0. 7
0. 8
0. 9
1. 0
C:  N. ca li fornicus
0. 4
0. 5
0. 6
0. 7
0. 8
0. 9
1. 0
0
24 48
            T i me  starve d (h our s)
  
 
 150 
 
 
 
 
 
 
CHAPTER 7   
EVALUATION OF SMALL SCALE RELEASE OF THE PREDACIOUS MITES, 
PHYTOSEIULUS PERSIMILIS AND GALENDROMUS OCCIDENTALIS (ACARI: 
PHYTOSEIIDAE) FOR BIOLOGICAL CONTROL OF CITRUS RED MITES, 
PANONYCHUS CITRI (ACARI: TETRANYCHIDAE) IN ALALBAMA SATSUMA 
CITRUS ORCHARDS 
 
INTRODUCTION 
Satsuma mandarin (Citrus unshiu Marcovitch) production is a growing industry in 
southern Alabama and other parts of the Gulf Coast region of the United States (Campbell et al. 
2004). This crop, which has been grown for over a century in Alabama (English and Turnipseed 
1940), has received renewed interests in the last two decades, partly because of strong industry 
and state support and development of new markets (Campbell et al. 2004). The citrus red mite, 
Panonychus citri (McGregor) (Acari: Tetranychidae) was first reported as an important pest of 
satsuma mandarin in Alabama by English and Turnipseed (1940). In recent surveys, P. citri was 
identified as one of the most abundant pests in Alabama satsuma orchards, in particular 
conventionally sprayed orchards (Fadamiro et al. 2007, 2008). In Alabama, P. citri is typically a 
spring pest (Fadamiro et al. 2008), and population densities of the pest during this period are 
usually above the economic threshold of 5 motiles per leaf, proposed by Childers (1994). 
Panonychus citri is also a key pest of citrus in many other parts of the world (Gotoh and Kubota 
 
 151 
1997, Jamieson et al. 2005, Childers et al. 2007). Both immatures and adults feed on citrus 
leaves. Severe leaf infestation may result in grey or silvery spots known as stippling damage, 
reduced photosynthesis, premature shoot dieback and decreased plant vigor (Kranz et al. 1977). 
High infestations may also result in fruit feeding and damage (Childers et al. 2007).  
Historically, P. citri has been controlled in the Gulf Coast region and other parts of the 
world through applications of conventional acaricides (Childers 1994, Jamieson et al. 2005, 
Childers et al 2007). However, use of broad spectrum acaricides may speed up phytophagous 
mite resistance (Shanks et al. 1992, Omoto et al. 1995, Bergh et al. 1999), disrupt predators 
(Thistlewood 1991, Welty 1995, Antonelli et al. 1997, Jamieson et al. 2005, Urbaneja et al. 
2008), and intensify food safety concerns. These concerns have resulted in renewed interests in 
the use of predacious mites for biological control of spider mites (McMurtry 1983, McMurtry 
and Croft 1997, Jamieson et al. 2005, Rhodes et al. 2006).  
Predacious mites, in particular those belonging to the families Phytoseiidae and 
Stigmaeidae, have been widely evaluated in the laboratory and field for biological control of 
phytophagous mites in many crop systems (Childers et al. 1975, McMurtry 1983, Childers 1994, 
Wood et al. 1994, Takano-Lee and Hoddle 2001, 2002; Colfer et al. 2004, Opit et al. 2004, Pratt 
et al. 2002, Jamieson et al. 2005, Rhodes et al. 2006, Fraduo and Liburd 2007, Arthur et al. 2009, 
Cakmak et al. 2009). The key species evaluated in many systems include three commercially 
available phytoseiids, Phytoseiulus persimilis Athias-Henriot, Galendromus occidentalis 
(Nesbitt), and Neoseiulus californicus (McGregor), but some indigenous species have also been 
evaluated (Muma and Denmark 1970, Childers et al. 1975, Childers 1994). Childers (1994) 
reported on the potential use of certain indigenous species of predacious mites for biological 
control of pest mites on citrus in Florida, including Typhlodromalus peregrinus (Phytoseiidae) 
 
 152 
and Agistemus floridanus (Stigmaeidae). A recent catalogue of the predacious mite fauna of 
satsuma mandarin in Alabama identified 29 species from nine families, with the dominant 
species being Typhlodromalus peregrinus Muma and Proprioseiopsis mexicanus (Garman) 
(Phytoseiidae), and Agistemus floridanus Gonzalez (Stigmaeidae) (Fadamiro et al. 2009). Many 
of these species (e.g., T. peregrinus and P. mexicanus) were found in association with P. citri 
(Fadamiro et al. 2008, 2009). However, abundance of predacious mites in local orchards was 
generally too low to effect significant suppression of phytophagous mites (Fadamiro et al. 2009). 
Attempts to mass rear the key indigenous predacious mites for augmentative releases against P. 
citri in Alabama orchards have not been successful (Xiao and Fadamiro, unpublished data), and 
thus our interest in evaluating commercially available phytoseiids as potential biological control 
agents for P. citri.  
In a recent laboratory study, we evaluated the predation potential of three commercially 
available phytoseiids, P. persimilis, G. occidentalis, and N. californicus, on P. citri (Xiao and 
Fadamiro in review). Phytoseiulus persimilis is a type I specialist predator of spider mites, while 
G. occidentalis, and N. californicus are type II selective predators of spider mites (McMurtry and 
Croft 1997, Blackwood et al. 2001, Croft and Jung 2001, Fraduo and Liburd 2007). Our study 
included a series of experiments to assess the numerical and functional responses and prey-stage 
preferences of the phytoseiids when offered P. citri as prey. The results showed that all three 
species were effective in regulating P. citri density, preferred P. citri nymphs over eggs, and 
showed a functional type II (convex) response in which the number of prey consumed increased 
with prey availability up to a maximum point after which it slowly began to decrease. However, 
the two most effective species were P. persimilis and G. occidentalis (Xiao and Fadamiro, in 
review). These initial findings coupled with their favorable life history traits (McMurtry and 
 
 153 
Croft 1997, Pratt and Croft 2000), aided our selection of both species for further evaluation in the 
field.  
In this study, field experiments were conducted to evaluate the effectiveness of small-
scale releases of P. persimilis or G. occidentalis at different rates and frequencies for suppression 
of P. citri in satsuma orchards in southern Alabama.  
 
MATERIALS AND METHODS 
Study Sites   
Field trials were conducted in Coker orchard (N 30? 33.598?, W 87? 48.026?) located in 
Baldwin county, south Alabama during 2007 and 2008. The orchard had ~ 250 citrus trees 
comprised primarily of satsuma mandarin, with a few sweet orange (Citrus sinensis (L.) Osbeck), 
grapefruit (C. paradisi Macfad), and kumquat (Fortunella spp.). The predominant cultivar of 
satsuma mandarin was ?Owari?. The orchard has a history of high infestation of P. citri and is 
typically managed using conventional practices including routine applications of pesticides. 
However, no pesticides were applied in the orchard during this study. Temperatures at this 
location during the experiments in spring (March-May) 2007 and 2008 were 17.1- 24 ?C 
(minimum: 10 ?C, maximum: 29.4 ?C), and 15.3 - 24.0?C (minimum: 9.5 ?C, maximum: 
28.6 ?C), respectively. Relative humidity in both years was around 65 -75 %.  
 
Predacious Mites  
The phytoseiids, P. persimilis and G. occidentalis were purchased from Biocontrol 
Network, Inc. (Brentwood, TN) and kept separately in a cooler (4 ?C) for 1-2 days, if the weather 
was unsuitable for immediate release.  
 
 154 
General Procedure for Predacious Mite Release and Sampling  
Four separate field experiments were conducted during 2007 and 2008 in the test orchard 
to determine the effectiveness of releases of P. persimilis or G. occidentalis at different rates and 
frequency in suppressing P. citri. First, satsuma trees in the orchard were sampled for P. citri 
using a protocol described by Fadamiro et al. (2008), to determine initial population densities. 
The trees were then classified into one of two groups based on the P. citri threshold proposed by 
Childers et al. (2007): i) trees with high initial P. citri density (i.e. > 120 motiles per 24 leaves, 
or > 5 motiles per leaf), and ii) trees with moderate initial P. citri density (i.e. ~ 72-96 motiles 
per 24 leaves or 3-4 motiles per leaf). The first three experiments were conducted on trees with 
high initial densities of P. citri, whereas the fourth experiment was conducted on trees with 
moderate initial prey densities. All selected trees were of similar size (canopy of 2 m height, 1.5 
m diameter) and had not previously been treated with pesticides during the season. Two test trees 
were separated by at least one ?buffer? tree to minimize wind-aided dispersal of mites between 
test trees. The experiment was a randomized complete block design and trees were randomly 
assigned to the different treatments. 
Each shipment of predacious mites from the supplier arrived in a plastic vial (6 cm high ? 
5 cm diameter) consisting of 500-1000 individuals on a carrier. The average number of 
predacious mites per carrier was estimated by rolling the vial to evenly disperse mites within the 
carrier, placing samples into a petri dish, and counting the number of predacious mites under a 
stereo dissecting microscope (20?). Prior to release, the predacious mites were tested for 
viability and activity by observing subset samples of ~ 20 individuals in a Petri-dish for about 10 
min. The releases rates evaluated were based on the supplier recommendation (i.e. ~ 2000/acre 
for field releases), published rates (e.g., ~5000/ha in cotton, Colfer et al. 2004), and cost 
 
 155 
considerations. For each species, individuals at the appropriate release rate were evenly 
distributed into four plastic containers (each with holes to allow predacious mites to exit and 
disperse), which were used as the release device. The four plastic containers were then hung on 
branches located on four sides (one container per side) of a test tree at ~ 1.5 m above the ground. 
Predacious mite releases were performed around 11 am to 2 pm when temperature was around 
25 ?C, at RH 65-70 %. 
To evaluate efficacy of predacious mite releases, 24 randomly selected leaves (6 per side) 
were taken from each test tree (trees on which predacious mites were released) per sampling date. 
The leaves were collected in properly-labeled paper bags, held in a cooler and transported to the 
laboratory where they were examined under a microscope at 20 ? magnification. The numbers of 
P. citri eggs and motiles (nymphs + adults) per leaf were counted and recorded. The total number 
of P. citri was calculated as the sum of eggs and motiles (i.e. all stages). Predacious mites were 
rarely observed on the leaves and thus were not recorded. 
 
Experiment 1: G. occidentalis releases in 2007 
The first trial in 2007 evaluated releases of G. occidentalis on satsuma trees infested with 
high densities (> 5 motiles per leaf) of P. citri, based on pre-release samples collected on 
February 28, one day before the first predacious mite release (March 1). The following five 
treatments testing releases of G. occidentalis at different rates and frequencies were evaluated: 1) 
one single release (in early March) at a rate of 25 G. occidentalis per tree (G25-1); 2) one single 
release (in early March) at a rate of 50 G. occidentalis per tree (G50-1); 3) two releases (in early 
March and late March, respectively) at a rate of 25 G. occidentalis per release per tree (G25-2);  
4) two releases (in early March and late March, respectively) at a rate of 50 G. occidentalis per 
 
 156 
release per tree (G50-2); and 5) no-release control. Each treatment was replicated three times and 
data were collected at 14, 28, 42, and 56 days after the first predacious mite release (DAR).  
 
Experiment 2: G. occidentalis releases in 2008 
A second experiment was conducted in 2008 to further evaluate releases of G. 
occidentalis on satsuma trees infested with high densities of P. citri. Higher release rates of the 
predacious mites were tested based on the results from the 2007 trial, which showed minimal 
efficacy at low release rates. The following treatments were evaluated: 1) two releases (in early 
March and late March, respectively) at a rate of 50 G. occidentalis per release per tree (G50-2); 2) 
two releases (in early March and late March, respectively) at a rate of 100 G. occidentalis per 
release per tree (G100-2); 3) two releases (in early March and late March, respectively) at a rate 
of 200 G. occidentalis per release per tree (G200-2);  4) one release (in early March) at a rate of 
200 G. occidentalis per release per tree (G200-1); and 5) no-release control. Each treatment was 
replicated three times and data were collected at 7, 14, 28, 35, 49, and 63 days after the first 
predacious mite release.  
 
Experiment 3: P. persimilis releases in 2008 
Similar to the G. occidentalis releases described in experiment 2, a third trial was 
conducted in 2008 to evaluate releases of P. persimilis on satsuma trees infested with high 
densities of P. citri. The following treatments were evaluated: 1) two releases (in early March 
and late March, respectively) at a rate of 50 P. persimilis per release per tree (P50-2); 2) two 
releases (in early March and late March, respectively) at a rate of 100 P. persimilis per release 
per tree (P100-2); 3) two releases (in early March and late March, respectively) at a rate of 200 P. 
 
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persimilis per release per tree (P200-2);  4) one release (in early March) at a rate of 200 P. 
persimilis per release per tree (P200-1); and 5) no-release control. Each treatment was replicated 
three times and data were collected at 7, 14, 28, 35, 49, and 63 days after the first predacious 
mite release.  
 
Experiment 4: Releases of G. occidentalis or P. persimilis in 2008  
A fourth experiment was conducted in 2008 to evaluate single releases of G. occidentalis 
or P. persimilis at two release rates (low and high) on satsuma trees infested with moderate 
densities (3-4 motiles per leaf) of P. citri. The following treatments were evaluated: 1) one single 
release of G. occidentalis (in late March) at a rate of 100 per tree (G100-1); 2) one single release 
of G. occidentalis (in late March) at a rate of 200 per tree (G200-1); 3) one single release of P. 
persimilis (in late March) at a rate of 100 per tree (P100-1);  4) one single release of P. persimilis 
(in late March) at a rate of 200 per tree (P200-1); and 5) no-release control. Each treatment was 
replicated three times and data were collected at 7, 21 and 35 days after predacious mite release.  
 
Statistical analysis   
Data for each experiment and predacious mite species were analyzed and presented 
separately. Mean number of P. citri (eggs, motiles and total) per sampling data were computed 
for each treatment and used for statistical analysis. Data were not normally distributed and thus 
were normalized by using the square-root transformation (? x + 0.5). The transformed data were 
then analyzed with one-way analysis of variance (ANOVA) followed by the Tukey-Kramer 
honestly significant difference (HSD) comparison test to test for significant treatment effect on 
each sampling data (P < 0.05, JMP Version 7.0.1, SAS Institute Inc. 2007).  
 
 158 
RESULTS 
Experiment 1: G. occidentalis Releases in 2007 
Pre-release sampling of P. citri on February 28 showed no significant differences among 
the treatments in the number of P. citri eggs, motiles, and the total number of all stages (Table 1). 
The first release of G. occidentalis was made on March 1. Data collected 14 days (March 14) 
after the first release (DAR) of G. occidentalis showed no significant differences among the 
treatments. However, significant differences were recorded among the treatments at 28 DAR. 
Significantly lower numbers of P. citri eggs and motiles were recorded in the treatment in which 
G. occidentalis was released twice at a rate of 50 per tree (G50-2) than in the control (Table 1), 
but no differences were recorded among the treatments at 42 (for motiles) and 56 DAR. Note 
that the second release of predacious mites was made in the appropriate treatments (G25-2 and 
G50-2) on March 28. Although densities of P. citri were lower in some treatments than in the 
control, high densities above the economic threshold (> 5 motiles per leaf) were recorded in all 
treatments in most sampling dates, with the exception of the last sampling date (56 DAR) when 
relatively lower densities (were recorded across the board (Table 1).  
 
Experiment 2: G. occidentalis Releases in 2008 
Data collected during the pre-release sampling of P. citri on February 29 showed no 
significant differences among the treatments in the density of P. citri (Table 2). The first release 
of G. occidentalis was made on March 1. Significant differences were recorded among the 
treatments as early as 7 DAR. Significant differences were also recorded at 14 DAR for eggs, 28 
DAR for eggs, 35 DAR, 49 DAR for motiles, and 63 DAR for eggs. In general, lower P. citri 
densities were recorded in all the treatments compared to control. For instance, the numbers of P. 
 
 159 
citri eggs were significantly lower in all the G. occidentalis treatments than in the control at 7, 14, 
28, and 35 DAR. At 63 DAR, significantly lower numbers of P. citri eggs were recorded in the 
G200-1 and G200-2 treatments than in the remaining treatments, suggesting that release rate of 
200 G. occidentalis per tree was more effective than the lower release rates (Table 2). The G200-
2 treatment also had the lowest number of P. citri motiles at 49 DAR. Note that the second 
release of G. occidentalis was made in the appropriate treatments (G50-2, G100-2 and G200-2) 
on March 28. 
 
Experiment 3: P. persimilis Releases in 2008 
The trends for the P. persimilis releases were generally similar to those recorded for G. 
occidentalis in Experiment 2. Pre-release sampling was conducted on February 29, followed by 
the first predacious mite release on March 1. The pre-release data showed no significant effect 
among treatments on P. citri density, but significant differences were recorded among the 
treatments at 7 (only for eggs), 14, 28, 35, 49, and 63 DAR (Table 3). In almost all the cases, 
densities of P. citri in the control were significantly greater than in the treatments. In general, the 
lowest densities were recorded in the P200-2 treatment, suggesting that this treatment may be 
more effective than the others. Surprisingly, the lowest number of P. citri motiles was recorded 
in the control at 63 DAR, but this may be due to a general crashing of P. citri populations at that 
time of the season. Note that the second release of P. persimilis was made in the appropriate 
treatments (G50-2, G100-2 and G200-2) on March 28. 
 
Experiment 4: Releases of G. occidentalis or P. persimilis in 2008  
 
 160 
In this experiment the two predacious mite species were released (in separate treatments) once on 
trees with initial moderate densities of P. citri. Pre-release data were collected on March 28, and 
the first predacious mite release was made on March 29. No significant differences were 
recorded among the treatments in the pre-release density of P. citri, and also at 7 DAR. However, 
significant differences in the numbers of P. citri eggs and motiles were recorded among the 
treatments at 21 and 35 DAR. In general, P. citri densities were higher in the control than in the 
treatments. No clear differences were recorded between G. occidentalis and P. persimilis 
releases, suggesting a similar efficacy for both predacious mite species.  
 
Population Dynamics of P. citri on Experimental Trees 
Data from Experiments 2 (G. occidentalis releases) and 3 (P. persimilis releases) were 
presented to illustrate the population dynamics of P. citri in the different treatments (Fig. 1). In 
general, P. citri density (total count of all stages) in the control increased from ~ 50 per leaf at 
the beginning of the experiment on March 6 to a high ~ 360 per leaf on April 4, and then 
declined to ~ 57 per leaf by the last sampling on May 2. In general, a similar trend in population 
dynamics of P. citri was recorded in the predacious mite treatments, only that densities of P. citri 
were much lower in some treatments (Fig. 1). 
 
DISCUSSION 
Several conclusions can be drawn based on the results from all four experiments. First, 
varying levels of success were achieved with releases of either G. occidentalis or P. persimilis on 
trees with moderate to high initial densities of P. citri, with release rate and frequency being the 
key factors determining the ability of the predacious mites to maintain effective long-term 
 
 161 
suppression of the prey. Second, predacious mites were more effective at high release rates than 
at lower rates. Third, releases were more successful at moderate than at high prey densities. 
Fourth, both predacious mite species showed similar efficacy in suppressing P. citri, but were 
detected at very low population densities throughout the trial, indicative of poor establishment 
and reproduction. Other authors have also reported that release rate/ratio, frequency and timing 
of release, and initial prey density are critical factors that may impact successful biological 
control of phytophagous mites by predacious mites (Hamlen and Lindquist 1981, Hoddle et al. 
2000, Pratt and Croft 2002, Shrewsbury and Hardin 2003, Opit et al. 2004, Fraulo and Liburd 
2007). In general, predacious mites were more effective at higher release rates and frequencies 
and at low initial prey densities. Furthermore, Fraulo and Liburd (2007) reported that release rate 
and frequency had a greater impact than release timing, on the ability of Neoseiulus californicus 
(McGregor) to provide effective and season-long suppression of spider mites on strawberry. 
Although results from the first three experiments suggest that the predacious mites were 
not consistently effective when released on trees with high population densities of P citri, data 
from the last experiment using trees with moderate P. citri density showed that one single 
releases of G. occidentalis or P. persimilis at a rate of 100 or more per tree provided effective, 
long-term suppression of P. citri below the economic threshold, whereas densities on  control 
trees dramatically increased during this period.  These results are consistent with the reports 
which showed that small to large scale releases of G. occidentalis, P. persimilis or other 
phytoseiids provided effective suppression of spider mite populations in many crop systems, 
including strawberries (McMurtry 1982, 1991, Van de Vrie and  Price 1994, Fraulo and Libburd 
2007, Cakmak et al. 2009), avocado (Hoddle et al. 2000, Takano-Lee and Hoddle 2001), hops 
(Strong and Croft 1995, 1996), ivy geranium (Opit et al. 2004), greenhouse vegetable crops 
 
 162 
(McMurtry 1991, Arthurs et al. 2009), and ornamental plants (Hamlen and Lindquist 1981, Pratt 
and Croft 1998). In contrast, a few authors reported little or no efficacy of predacious mite 
releases in suppression of spider mites in some crops (Shrewsburry and Hardin 2003, Colfer 
2004). Shrewsburry and Hardin (2003) reported that predator releases were more expensive and 
provided less effective suppression of spruce spider mites, Oligonychus ununguis (Jacobi) on 
juniper, compared with conventional pesticides. Similarly, Colfer et al. (2004) reported that 
large-scale releases of G. occidentalis did not provide economic control of spider mite in cotton. 
Our results which showed the field efficacy of both predacious mite species in 
suppressing P. citri, in particular at high release rates and moderate prey densities, are consistent 
with the results of a recent laboratory study which also demonstrated the ability of both species 
to regulate the prey (Xiao and Fadamiro, in review). The data which showed reduced efficacy of 
both species at high prey densities are also in agreement with previous reports by other authors 
(e.g., Shrewsbury and Hardin 2003, Colfer et al. 2004). Several factors may explain of the lack 
of efficacy of both predatory mite species recorded at high prey densities. In Alabama satsuma 
orchards, P. citri is typically a major pest in early spring (February to April) with populations 
starting to decline in late April and eventually crashing by early summer (mid-May) (Fadamiro et 
al. 2007, 2008). Thus, this temporal (8-9 weeks) seasonal occurrence may have provided limited 
time and opportunity for the establishment and reproduction of predacious mites (McMurtry and 
Croft 1997), and this may also explain our observation of very low population densities of both 
predacious mite species on the test trees throughout the trial.  Shrewsbury and Hardin (2003) also 
attributed the inability of multiple releases of N. fallacies and G. occidentalis to control O. 
ununguis on juniper, to the temporal seasonal abundance of the pest. Secondly, our data showing 
poor efficacy of both predatory mite species at high prey densities may be related to low predator: 
 
 163 
prey release ratios (Greco et al. 2005, Easterbrook et al. 2001, Opit et al. 2004, Fraudo and 
Liburd 2007). For example, Sato et al. (2007) reported that three releases of N. californicus 
without pesticides were not sufficient to significantly reduce populations of T. urticae, possibly 
because of low initial predator: prey release ratio. In addition, the observed poor efficacy of both 
predacious mites at high prey densities may possibly be due to intense interference between 
predator and prey, which may have negatively impacted establishment and performance of the 
predacious mites (Mori and Chant 1966, Pratt and Croft 2002, Shrewsbury and Hardin 2003). 
Although predacious mite releases at high rates resulted in lower densities of P. citri 
motiles, densities of prey eggs were consistently high in all treatments, suggesting that the 
predacious mites had minimal effects of P. citri eggs. These results are consistent with our 
findings in the laboratory which showed that both species preferred nymphs to eggs of P. citri 
(Xiao and Fadamiro in review). The fact that suppression of P. citri was achieved without any 
evidence of establishment of the predacious mites in the orchard is interesting and may suggest 
influence of other factors or interactions, other than predator release. Colfer et al. (2004) reported 
a similar result in which releases of G. occidentalis in cotton did enhance populations of the 
predator but resulted in lower spider mite densities, nonetheless. The authors proposed that the 
reduction in spider mite densities in plots in which the predator was released was probably 
caused by a chance event or a complicated set of effects which was triggered by predator mite 
releases (Colfer et al. 2004).  
In conclusion, the results of this study demonstrated efficacy of small-scale releases of P. 
persimilis or G. occidentalis at release rates of 100 or more predacious mites per tree, in 
suppression of P. citri at moderate initial densities. The two species showed similar efficacy 
against P. citri, although P. persimilis was slightly more effective in at least one trial. Previous 
 
 164 
studies suggest that both species can survive low to moderate starvation in the laboratory (Xiao 
and Fadamiro in review) and tolerate high field temperatures (McMurtry and Croft 1997).  
Moreover, both species are highly active with high prey searching efficiency (Pratt and Croft 
2000, Blackwood et al. 2001), and appeared adapted to disturbed habitats, such as intensively-
managed orchards (McMurtry and Croft 1997). These factors suggest that both species may fair 
well in the severe southern Alabama climate. Further field evaluations including cost analysis are 
necessary to determine the economic feasibility of large-scale biological control of P. citri with 
predacious mites. In this study, we evaluated releases of single predator species rather than 
multiple predator species. Previous studies that compared the effectiveness of single versus 
multiple predator species have produced contrasting results, ranging from negative effect 
(Rosenheim et al. 1995; Schausberger and Walzer 2001) to neutral (Denoth et al. 2002, Chow et 
al. 2008) or positive effect (Cakmak et al. 2009). Schausberger and Walzer (2001) found that 
interaction such as competition, interguild predation, and cannibalism affected the development 
and coexistence of mixed predator populations. In contrast, Cakmak et al. (2009) reported that 
combined releases of N. californicus and P. persimilis provided greater suppression of 
Tetranychus cinnabarinus (Boisd.) (Acari: Tetranychidae) on strawberry compared to releases of 
each predator species alone. Future studies will determine the efficacy of combined release of G. 
occidentalis and P. persimilis, as well as integration of predacious mites with petroleum oils (e.g., 
FC 435-66 oil) and other effective reduced-risk acaricides (Fadamiro et al. 2005), for managing 
P. citri in Alabama citrus orchards. 
 
 
 
 
 165 
ACKNOWLEDGEMENTS 
We thank Mr. Monte Nesbitt for helping with the field trials, and the satsuma grower 
cooperator (Coker orchard). Funding for this study was provided through grants by the Alabama 
Agricultural Experiment Station and Auburn University Horticulture Line Item grants program 
to HYF. 
 
 166 
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diverse pesticide programs. Can. Entomol. 123: 1163 - 1174. 
Urbaneja, A., S. Pascual-Ruiz, and T. Pina. 2008.  Efficacy of some acaricides against 
Tetranychus urticae (Acari: Tetranychidae) and their side-effects on selected natural enemies 
occurring in citrus orchards. Pest. Manag. Sci. 64: 834 - 842. 
Van de Vrie, M., and J. F. Price. 1994.  Manual for biological control of two spotted spider 
mites on strawberry in Florida. Univ. Fla. Agric. Exp. Sta. Rep. DOV. pp.10.  
Welty, C. 1995.   Survey of predators associated with European red mite (Panonychus ulmi 
(Acari:Tetranychidae) in Ohio apple orchards. Great Lake Entomol. 28: 171 - 184.  
Wood, L., D. A. Raworth, and M. Mackauer. 1994.   Biological control of the two spotted 
spider mite in raspberries with the predator mite, Phytoseiulus persimilis. J. Entomol. Soc. 
B.C. 91: 59 - 61. 
 
 
 173 
Table1. Effect of G. occidentalis releases on density of P. citri on satsuma trees (experiment 1, spring 2007) 
P. citri 
stage  
Treatment  
 
Mean (? SE) number of P. citri per 24 leaves  
 Pre-release 
(Feb 28) 
14 DAR 
(Mar 14) 
28 DAR 
(Mar 28) 
42 DAR 
(Apr 11) 
56 DAR 
(Apr 25) 
Eggs  G25-1 1438 ? 416 3308 ? 776 7383 ? 785a 3025 ? 744a 506 ? 167 
 G25-2  814 ? 216 1612 ? 626  5059 ? 848ab   732 ? 174b 316 ? 247 
 G50-1  816 ? 174 2393 ? 785  3807 ? 874ab 2523 ? 587a 620 ? 100 
 G50-2 984 ? 164 1244 ? 414    2976 ? 848b 2629 ? 390a      571 ? 90 
 Control 921 ? 191 777 ? 389   6225 ? 654ab  1729 ? 185ab 380 ? 253 
  
F = 0.871, 
P = 0.514 
F = 2.564, 
P = 0.103 
F = 4.840, 
P = 0.019 
F = 5.080, 
 P = 0.017 
F = 0.792, 
P = 0.556 
Motiles  G25-1 204 ? 77 516 ? 132 889 ? 235a 343 ? 52  218 ? 159 
 G25-2 132 ? 45 271 ? 107 350 ? 85ab 293 ? 33  48 ? 31 
 G50-1 209 ? 41    187 ? 98     430 ? 33ab  551? 85        69 ? 4 
 G50-2   97 ? 18    217 ? 44     289 ? 79b 263 ? 79 118 ? 36 
 Control 104 ? 21 202 ? 132     735 ? 58a  340 ? 81   99 ? 39 
  
F = 1.270, 
P = 0.344 
F = 1.350, 
P = 0.318 
F = 5.090, 
      P = 0.017 
F = 0.509, 
P = 0.731 
F = 0.661, 
P = 0.634 
Means within the same column (sampling date) for each stage with different letters are significantly different (P < 0.05, HSD, df = 4, 
10). G 25-1: one release of G. occidentalis at the rate of 25/tree; G25-2: two releases at the rate of 25/tree; G50-1: one release at the 
rate of 50/tree; G50-2: two releases at the rate of 50/tree; Control: no releases. Note: First and second releases of PM were made on 
March 1 and Mar 28, respectively. DAR: days after the first release of predacious mites.  
 
 174 
Table 2. Effect of G. occidentalis releases on density of P. citri on satsuma trees (experiment 2, spring 2008) 
P. citri 
stage  
 
Treatment 
Mean (? SE) number of P. citri per 24 leaves  
Pre-release 
(Feb 29) 
7 DAR 
(Mar 7) 
14 DAR 
(Mar 14) 
28 DAR 
(Mar 28) 
35 DAR 
(Apr 4) 
49 DAR 
(Apr 18) 
63 DAR 
(May 2) 
Eggs  G50-2  1253 ? 205    879 ? 121b  1603 ? 129b 3069 ? 160b 5984 ? 379b 4524 ? 758 1210 ? 156a 
 G100-2  1282 ? 377    956 ? 99b  1684 ? 140b 2536 ? 529b 4257 ? 420b 4110 ? 883 1022.? 159a 
 G200-1      925 ? 80    929 ? 102b  1388 ? 236b 2657 ? 426b 3757 ? 619b 3365 ? 303 524 ? 94b 
 G200-2 1225 ? 61    801 ? 44b   1642 ? 99b 1763 ? 136b 4088 ? 542b 3299 ? 345 453 ? 38b 
 Control  1092 ? 141  1586 ? 100a  3659 ? 456a 5473 ? 539a 7218 ? 729a 3503 ? 419 1309 ? 126a 
 
 F = 0.534, 
P = 0.714 
F = 11.08, 
P = 0.0011 
F = 14.32, 
P = 0.0004 
F = 10.74, 
  P = 0.0012 
F = 6.445, 
P = 0.007 
F = 0.735, 
P = 0.588 
F = 12.86, 
P = 0.0006 
Motiles  G50-2 201 ? 69   53 ? 39   379 ? 144  678 ? 132 672 ? 146b 1019 ?170ab 76 ? 4 
 G100-2 121 ? 73 38 ? 8     237 ? 54  569 ? 268 714  ? 26b 7 02 ? 131bc 78 ? 3 
 G200-1     155 ? 5 49 ? 8  220 ? 137  242 ? 58  592 ? 142b  591 ? 127bc  82 ? 24 
 G200-2 112 ? 24        77 ? 21     329 ? 52  460 ? 39  390 ? 154b   357 ? 123c 53 ? 4 
 Control 116 ? 37 156 ? 63  569 ? 126  953 ? 226 1399 ? 210a 1649 ? 312a 52 ? 9 
 
 F = 0.584, 
P = 0.681 
F = 3.042, 
P = 0.07 
F = 1.410, 
P = 0.297 
F = 2.830, 
   P = 0.083 
F = 9.050 
P = 0.002 
F = 9.581, 
P = 0.002 
F = 1.250, 
P = 0.349 
Means within the same column for each stage with different letters are significantly different (P < 0.05, HSD, df = 4, 10). G50-2: two 
releases of G. occidentalis at the rate of 50/tree; G100-1: one release at the rate of 100/tree; G200-1: one release at the rate of 200/tree; 
G200-2: two releases at the rate of 200/tree; Control: no releases. Note: First and second releases of PM were made on March 1 and 
March 28, respectively. DAR: days after the first release of predacious mites.  
 
 175 
Table 3. Effect of P. persimilis releases on density of P. citri on satsuma trees (experiment 3, spring 2008)  
P. citri 
stage 
Treatment 
 
Mean (? SE) number of P. citri per 24 leaves  
Pre-release 
(Feb 29) 
7 DAR 
(Mar 7) 
14 DAR 
(Mar 14) 
28 DAR 
(Mar 28) 
35 DAR 
(Apr 4) 
49 DAR 
(Apr 18) 
63 DAR 
(May 2) 
Eggs  P50-2 930 ? 64 915 ? 37b 1788 ? 76b 2947 ? 401b 4594 ? 591b 5393 ? 238a 1446 ? 75a 
 P100-2 1009 ? 223 868 ? 89b 1967 ? 139b 1899 ? 259b 4002 ? 336b  4525 ? 196ab 1430 ? 184a 
 P200-1 983 ? 90 843 ? 22b 1316 ? 58b 1986 ? 144b 4240 ? 303b  4108 ? 611ab 1418 ? 51a 
 P200-2 1276 ? 167 463 ? 52c 1437 ? 70b 2848 ? 620b 3417 ? 312b 3279 ? 459b   539 ? 123b 
 Control 1077 ? 136 1309 ? 100a 3636 ? 456a 6437 ? 539a 7198 ?729a 3501 ? 318b 1276 ? 96a 
  
F = 0.854, 
P = 0.523 
F = 17.34, 
P = 0.0001 
F = 24.14, 
P = 0.0001 
F = 10.35, 
P = 0.001 
F = 8.770, 
P = 0.003 
F = 4.860, 
P = 0.019 
F = 18.38, 
P = 0.002 
Motiles  P50-2 103 ? 8  49 ? 14 182 ? 23b 581 ? 105ab 715 ? 79ab   537 ? 55bc   121 ? 16ab 
 P100-2  155 ? 35 50 ? 3   327 ? 10ab  414 ? 62b 523 ? 133b 1156 ? 154ab     95 ? 11bc 
 P200-1    150 ? 15  43 ? 18   172 ? 3b  401 ? 22b 537 ? 194b    450 ? 40c   136 ? 210ab 
 P200-2 136 ? 96 43 ? 5  167 ? 13b  606 ? 44ab 410 ? 49b 514 ? 274c   175 ? 194a 
 Control    114 ? 9 157 ? 44  602 ?132a 924 ? 208a 1426 ? 186a 1492 ? 286a      55 ? 9c 
  
F = 0.526, 
P = 0.720 
F = 3.038, 
P = 0.070 
F = 5.250, 
P = 0.015 
F = 3.780, 
  P = 0.039 
F = 5.797, 
P = 0.011 
F = 11.84, 
P = 0.0001 
F = 10.60, 
P = 0.002 
Means within the same column for each stage with different letters are significantly different (P < 0.05, HSD, df = 4, 10). P50-2: two 
releases of P. persimilis at the rate of 50/tree; P100-1: one release at the rate of 100/tree; P200-1: one release at the rate of 200/tree; 
P200-2: two releases at the rate of 200/tree; Control: no releases. Note: First and second releases of PM were made on March 1 and 
March 28, respectively. DAR: days after the first release of predacious mites.  
 
 176 
Table 4. Effect of G. occidentalis or P. persimilis releases on density of P. citri on satsuma trees (experiment 4, spring 2008)  
P. citri  
stage 
Treatment 
 
Mean (? SE) number of P. citri per 24 leaves  
Pre-release  
(Mar 28) 
7 DAR 
(Apr 4) 
21 DAR    
(Apr 18) 
35 DAR 
(May 2) 
Eggs  G100-1 595 ? 70   522 ? 142  986 ? 29b   848 ? 99ab 
 G200-1 586 ? 92 438 ? 12    649 ? 73bc    375 ? 172b 
 P100-1   540 ? 106       442 ? 18    649 ? 85bc   518 ? 20b 
 P200-1   615 ? 110 421 ? 91 422 ? 9c   529 ? 69b 
 Control 562 ? 92 531 ? 55   2097 ? 210a   1610 ? 229a 
  
F = 0.103, 
P = 0.978 
F = 0.332, 
P = 0.851 
F = 45.22, 
P = 0.0001 
F = 7.84, 
 P = 00004 
Motiles  G100-1  87 ? 4  131 ? 38     101 ? 23b    54 ? 10ab 
 G200-1  87 ? 7    86 ? 20       74 ? 9b 33 ? 4b 
 P100-1    92 ? 11    76 ? 20  61 ? 19b   41 ? 14b 
 P200-1   85 ? 7    91 ? 32 79 ? 7b 39 ? 3b 
 Control   90 ? 2   162 ? 16 347 ? 26a 89 ? 9a 
  
F = 0.401, 
P = 0.804 
F = 2.180, 
P = 0.144 
F = 25.46, 
P = 0.0001 
F = 5.47, 
P = 0.013 
Means within the same column for each stage with different letters are significantly different (P < 0.05, HSD, df = 4, 10). G100-1: one 
release of G. occidentalis at the rate of 100/tree; G200-1: one release of G. occidentalis at the rate of 200/tree; P100-1: one release of 
P. persimilis at the rate of 100/tree; P200-1: one release of P. persimilis at the rate of 200/tree; Control: no releases. Note: Predacious 
mites were released on March 29. DAR: days after release of predacious mites.  
 
 177 
Figure 1. Population dynamics of P. citri on control trees and trees on which predacious mites 
were released in spring 2008. (A): Data from experiment 2 for G. occidentalis releases; (B): Data 
from experiment 3 for P. persimilis releases. G50-2: two releases of P. persimilis at the rate of 
50/tree; G100-1: one release at the rate of 100/tree; G200-1: one release at the rate of 200/tree; 
G200-2: two releases at the rate of 200/tree; Control: no releases. Total number of P. citri = eggs 
+ motiles (nymphs and adults). Arrows indicate predacious mite release dates. 
A:  G. o cc i d ent a l i s
Fe b  M a r  M a r  Ap r  Ap r  M a y   
Me
a
n 
(
? 
SE
)
 to
t
a
l
 n
um
be
r
 of
 
P. c
i
t
ri
 p
e
r
 24
 l
e
a
v
e
s
0
2000
4000
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10000
Cont ro l
G  5 0 - 2
G  1 0 0 - 2
G  2 0 0 - 2
G  2 0 0 - 1
Fe b 2 9 M a r 7 M a r 1 4 M a r 2 8
Ap r 4 Ap r 1 8 M a y  2
B :  P. p er s i m i l i s
0
2000
4000
6000
8000
10000
Cont ro l
P 5 0 - 2
P 1 0 0 - 2
P 2 0 0 - 2
P 2 0 0 - 1