Cooperative Extension and Propagation for Ca?o Negro, Costa Rica 
by 
 
Rachel Allen Meriwether 
 
 
 
 
A thesis submitted to the Graduate Faculty of 
Auburn University 
in partial fulfillment of the 
requirements for the Degree of 
Master of Science 
 
Auburn, Alabama 
May 9, 2011 
 
 
 
 
Keywords: Costa Rica, propagation, scarification, palm, cacti, Acoelorraphe wrightii, 
Hylocereus 
 
 
Copyright 2010 by Rachel Allen Meriwether 
 
 
Approved by 
 
Amy Noelle Wright, Chair, Associate Professor of Horticulture 
Kenneth Tilt, Professor of Horticulture 
Robert Boyd, Professor of Biological Sciences 
 
 
ii 
 
Abstract 
 
 
A research relationship was established with colleagues in Costa Rica.  These included 
not only members of academia but also non-profit organization workers, government 
officials, community members, and private research partners.  After extensive 
consultation with an advisory committee formed by members from each of these entities, 
a research project was formed.  Ca?o Negro National Wildlife Refuge was identified as 
an optimal location to begin research by all parties, and two plant species native to this 
location were selected on which to begin propagation experiments.  Hylocereus 
stenopterus cuttings were subjected to several different treatments to determine the most 
effective propagation technique.  Half of the cuttings were stuck immediately, while the 
other half were allowed to remain uncovered on a bench in the greenhouse for 72 h 
(cicatrization).  Prior to sticking, the basal ends of half of the cuttings were treated with a 
commercial rooting hormone while the other half received no hormone.  Once stuck, half 
of the cuttings were covered with a plastic tent.  Thus, treatments were arranged in a 2 
hormone x 2 cicatrization x 2 tent factorial design or 8 total treatments with 20 
replications per treatment for a total of 160 cuttings.  Additional cuttings of Hylocereus 
undatus and Hylocereus costariensis received one of three hormone treatments: untreated 
(no hormone applied, control), 0.1% indole butyric acid (IBA) or 0.8% IBA.  H. 
stenopterus cuttings responded well to both rooting hormone and mist tent but showed 
very little response to cicatrization time.  However, cuttings from all three species 
iii 
 
showed excellent survival, vigorous root growth, as well as subsequent shoot growth 
regardless of treatment.  The same was true of H. undatus and H. costariensis.  H. 
costariensis had a 100% survival rate regardless of treatment.  Experiments were also 
conducted on the palm Acoelorraphe wrightii.  Seeds of this plant were subjected to 
several preplanting scarification techniques including manual scarification, hot water 
soaking, and sulfuric acid scarification.  In all experiments, seeds scarified by either acid 
or hot water had higher germination percentages and rates than untreated (control) or 
manually scarified seeds.  Highest germination percentages were observed in the 1:1 
peat:perlite mix.
iv 
 
Acknowledgments 
 
 
 The author would like to thank Dr. Amy Noelle Wright for seeing the potential of 
an unexpected graduate student and taking the chance on a unique project.  Without her 
constant guidance and encouragement there is no doubt that this endeavor would never 
have been completed.  Thanks also go out to Drs. Kenneth Tilt and Robert Boyd for all 
their patience, encouragement and contributions to this research.  Particular thanks go out 
to Warner Orozco-Obando for his tremendous contributions to the success of the author?s 
research.  Special thanks also go out to fellow graduate students and office mates that 
made each step of this process just that much more enjoyable and survivable.  The love 
and support offered by all friends and family of the author has been integral to not only 
the completion of this degree but also the confidence to pursue it at all.  So, to you all, 
eternal thanks and appreciation are offered. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
v 
 
Table of Contents 
 
 
Abstract ............................................................................................................................... ii 
Acknowledgments.............................................................................................................. iv  
List of Tables ..................................................................................................................... vi  
List of Figures ................................................................................................................... vii  
Literature Review ................................................................................................................1 
Development of a Cooperative Extension Program in Costa Rica  ...................................30 
Propagation of Hylocereus stenopterus and Acoelorraphe wrightii...................................70 
 
Appendix?????????????????????????????? ? 94 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
vi 
 
List of Tables 
 
 
Chapter III 
 
Table 1: Effect of scarification treatment and germination substrate on germination 
percentage of A. wrightii seeds grown in a greenhouse in Auburn, AL. ...............................89 
 
Table 2: Effect of scarification treatment and germination substrate on number of days 
until germination of A. wrightii seeds grown in a greenhouse in Auburn, AL. ???? 90 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
vii 
 
List of Figures 
 
 
Chapter II 
 
Figure 1: Map of research trips taken in September 2009 and May 2010 .........................66 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
1 
 
Chapter I 
Literature Review 
Costa Rica 
Ecotourism has become one of the most important contributors to the Costa Rican 
economy, surpassing in 1993 the earnings for export products such as coffee and bananas 
(ICT, 1993).  The rise of ecotourism to this point began in the 1950s when, around the 
world, international tourism began to increase (Latham, 1994).  In 1955 the Costa Rican 
Tourism Institute was established, and in 1960 tourism was officially designated an 
?industry? within the country (Campbell, 2002).  Over the next several decades, 
ecotourism expanded rapidly and Costa Rica established itself as the most popular Latin 
American ecotourism destination (Schluter, 1993).  Unfortunately, wherever it is found, 
ecotourism can be responsible for the loss of local employment when companies bring in 
outside employees as well as destruction of the environment upon which it relies (Wild, 
1994).  Some of the negative effects of ecotourism have been attributed to the lack of 
local ownership of not only hotels and hostels but also tour companies, restaurants, shops, 
and assorted associated businesses (Burton, 1998; Honey, 2008).  If owners were also 
local inhabitants, there might be a stronger local investment in the long term conservation 
goals of a sustainable ecotourism program.  This theoretical assumption goes hand-in-
hand with a general shift in conservation strategies away from exclusionary tactics 
toward consumptive use conservation, where local inhabitants are allowed access to and 
use of resources in a sustainable manner, and community based conservation 
2 
 
(CBC) projects.  CBC programs are those that focus efforts not only on conserving an 
area and its environment but doing so in a way that actively involves the local population 
in activities, planning sessions, and any eventual profits. 
Perhaps one of the most well developed consumptive use programs in Costa Rica 
to date is the Payments for Environmental Services (PES) program (Pagiola, 2006).  
Begun in 1997, this program offers a method for administrators and land owners to 
translate external, non-market services provided by the environment into actual financial 
incentives.  Currently, the PES program recognizes four services provided by Costa 
Rican forests: 1) mitigation of greenhouse gases; 2) hydrological services; 3) biodiversity 
conservation; and 4) provision of scenic beauty for recreation and tourism (Pagiola, 
2008).  Interested land owners can apply and, if accepted, will receive monetary 
compensation for allowing lands that provide at least one of the above services to remain 
in natural forest cover.  As of 2000, roughly 300,000 hectares of land were receiving 
funding through the PES program with a waiting list of similar magnitude (Pagiola, 
2008).   
One example of a CBC and consumptive use program is the legal harvest of green 
sea turtle eggs in the Ostional Wildlife Refuge (Campbell, 1999).  The village of Ostional 
lies within the boundaries of the Refuge, both of which are located along the Pacific coast 
of the country on the Nicoya Peninsula.  This area is home to large numbers of nesting 
sea turtles, particularly the olive ridley sea turtle (Lepidochelys olivacea Eschscholtz), 
and these turtles have become a successful tourist attraction (Campbell et al., 2007).  In 
conjunction with the tourism revenue, members of the community take part in a legal egg 
harvest.  Eggs are then packaged onsite and distributed nationwide as a food product.  
3 
 
Ostional community members and researchers from the University of Costa Rica worked 
together to generate a legal harvest limit based on natural egg losses due to unique olive 
ridley nesting behavior.  Olive ridley turtles at Ostional, unlike not only other species of 
sea turtle but also other groups of olive ridleys in different locations, congregate for mass 
nesting events (Hughes & Richard, 1974).  These last from three to ten days and usually 
occur once a month.  Eggs laid on the first days of these events would normally be lost as 
other turtles arrive and dig new nests on top of the ones already present.  Thus it was 
determined that harvesting the first wave of laid eggs did not affect the overall population 
of olive ridley sea turtles (Cornelius et al., 1991).  Researchers from the University of 
Costa Rica and Ostional community members lobbied the government for permission to 
harvest eggs, which they were granted upon meeting the condition of creating a 
governing body.  This body was called the Ostional Development Association and 
became active in 1985.  Since that time the project has yielded recognizable 
socioeconomic benefits, including, but not limited to, financial assistance for the 
community from the Costa Rican Tourism Institute, involvement of the community with 
the University of Costa Rica (which has provided a research lab and nature guide training 
to interested parties), and 70% of all households surveyed in the Ostional community in 
2007 reported egg harvesting as their primary source of income (Campbell et al., 2007). 
In northern Costa Rica the Ca?o Negro National Wildlife Refuge (NWR) is an 
area with the capacity for both conservation collaboration as well as the opportunity for 
economic development.  Though the majority of the country is relatively stable 
economically, this remote region is facing other challenges.  The Costa Rican economy 
currently relies heavily on agriculture, electronics exports, and tourism (CIA, 2010).  
4 
 
Neither of the first two options is feasible in the Ca?o Negro area: agriculturally because 
of the sensitive nature of the various ecosystems that dominate the region and electronics 
manufacture is unreasonable due to the remote nature of the village as well as the lack of 
necessary infrastructure.  Furthermore, the few available jobs in the region are subject to 
decreased wages due to extensive illegal immigration from neighboring Nicaragua.  It has 
been suggested, however, that there is potential for program development and 
capitalization of the ecotourism industry which, if handled properly, could be a boon to 
both the economy as well as the ecosystem (Orozco-Obando, 2009).  Internationally 
targeted cooperative extension programming can bring together not only the local and 
national resources of an area but also a global perspective that can aid the development of 
ecosystem assets and enhance both economic opportunities and conservation goals 
(Harder et al., 2010; Ludwig, 2002; Place et al., 2008). 
The Ca?o Negro NWR is also highly significant as an ecosystem.  The majority 
of the area is a tropical dry forest, the most endangered large ecosystem on Earth (Janzen, 
1988).  It also contains portions of a vast seasonal wetland that feeds Lake Nicaragua to 
the north and has extensive populations of birds, caimans, and unique plant life.  Ca?o 
Negro has been recognized by the Ramsar Convention as a wetland of international 
significance (RAMSAR, 2010) and is also part of the Mesoamerican biological hotspot 
established by the organization Conservation International (CI, 2010).  It is also one of 
the most remote parks in Costa Rica?s national park system and has, thus far, escaped the 
degradation of native plant and animal communities often associated with increased in 
visitor traffic.  Economic opportunities in the area are associated with the wetland and, as 
tourism continues to expand, improper planning might introduce these problems to Ca?o 
5 
 
Negro as well.   The area is also in danger from large scale agricultural interests.  As 
international pineapple growers are shifting out of Hawaii and into cheaper production 
areas, most notably Brazil and Costa Rica, the wetland areas of Ca?o Negro have come 
under particular attack (Niesse, 2006).  Portions of the wetland have already been drained 
to make way for pineapple fields, and Ca?o Negro community members are being 
actively approached with regard to selling their lands for more of the same (Ruiz, 2010).   
Program Development 
 The overall and ongoing goals of this program will be to facilitate rural 
community economic development and bring in supplemental income for local citizens 
by providing new crops with commercial value as well as to establish an ecotourism 
destination in the village of Ca?o Negro which is located within the Ca?o Negro NWR.  
The effect of ecotourism on both rural development and conservation goals has been 
studied in several countries with mixed results that are attributed to several variables 
(Bookbinder et al, 1998; Campbell, 1999; Hvenegaard & Dearden, 2002; Potvin et al., 
2003; Trejos & Chiang, 2009).  The proximity of a protected area to Ca?o Negro 
represents an opportunity for collaboration with larger government entities and promises 
a higher chance of success in the development of the local economy (Plummer & Fennell, 
2009).  Utilizing recommendations from previous research, our educational extension 
program will provide farmers and local residents an economic alternative to selling their 
lands to the large pineapple producers that are moving into the area (Ruiz, 2009).  This 
will help not only preserve a unique and beautiful community but further protect a 
globally significant area of species diversity. 
6 
 
To develop an extension program in Costa Rica, extensive research was done to 
identify appropriate collaborating entities.  These included the Universidad Nacional de 
Costa Rica (UNA), the Instituto Tecnologico de Costa Rica (ITCR), the Instituto 
Nacional de Biodiversidad (INBio), the Organization for Tropical Studies (OTS), 
ProNativas, the Ministerio de Ambiente y Energia (MINAE), and the Ministerio de 
Agricultura y Ganaderia (MAG).  Preliminary research to identify regional native plant 
species appropriate for this project was corroborated by consultation with Costa Rican 
academic colleagues, regional experiment stations, and community farmers.  Individuals 
from each of these groups have formed an informal working group and advisory 
committee that provided local and national educational and political contacts to facilitate 
both conservation and development activities.  
Administration and faculty from UNA were among the first contacts made when 
efforts began to evaluate the biological, political, and economic factors associated with 
the feasibility of this project.  A relationship had already been established in February 
2008 between Auburn University (AU) and UNA personnel.   Later that same year a 
memorandum of agreement between AU and UNA was drafted and signed in order to 
facilitate future cooperative efforts between these two institutions.  This project 
represents the next step in the development of that relationship. 
UNA is a public institution and therefore, within Costa Rica receives funding 
directly from the government (UNA, 2010).  It is a four year university with both 
undergraduate and graduate degrees in the social sciences and multiple fields within the 
biological sciences, as well as philosophy and liberal arts.  It includes six separate 
7 
 
campuses; the main campus is in San Jose with satellite campuses in Sarapaqui, Nicoya, 
Benjamin Nunez, Coto, Perez Zeledon, and Liberia. 
Another university contacted in Costa Rica that focuses on agriculture and the 
biological sciences in general is ITCR.  ITCR also concentrates on community and 
applied technologies.  Interdisciplinary studies are recognized as being of great benefit 
and every effort is made to foster collaboration with other universities.  There is a fully 
developed distance learning PhD program which is organized in conjunction with several 
universities in several countries.   
INBio is a nonprofit organization that has been expanding its influence across the 
globe.  Started in 1989, INBio has become an international authority on programs and 
projects that conserve and promote tropical biodiversity (INBio, 2010).  During the late 
1980?s Costa Rica officially recognized the need to protect its wealth of biodiversity by 
passing Executive Decree No. 19153.  Among other things, this decree established the 
Planning Commission for the National Institute of Biodiversity, a board composed of 
members from various governmental institutions and also from higher education centers 
and conservation organizations.  This board recommended to the Costa Rican 
government that a state biodiversity institute be created which functioned with a high 
degree of autonomy.  Due to issues within the government itself, this recommendation 
was ignored.  At this point members of the board decided to take the situation into their 
own hands and create a private non-profit organization.  This institution was granted 
official recognition by the standing government and has been supported by successive 
governments as well.  The members then went about the laborious process of securing 
funding, eventually finding it in the form of a loan for 80 million colones as well as 
8 
 
backing from the Swedish Cooperation Agency (SIDA) and the MacArthur Foundation.  
On 26 October 1989 a location was selected in Santo Domingo de Heredia, and this is 
where INBio is still housed today (INBio, 2010).  Several services are now offered 
through INBio including bioprospecting, consulting, education and training, as well as 
general information services.  In 2000, a theme park called INBioparque was added as 
well.  Visitors can tour an organic farm, view a green built house, enjoy a butterfly 
garden, as well as many other nature based attractions.  Atta, the database where all 
information generated and collected by INBio is stored, is an extensive resource and 
almost indispensible for someone working with tropical biodiversity. 
OTS is another major research association in Costa Rica.  AU has a long standing 
relationship with this organization and has been an OTS member institution since 1987.  
Classes are offered on a number of subjects pertaining to tropical ecology at both the 
graduate and undergraduate levels and in multiple countries.  There are three research 
stations in Costa Rica and each specializes in a particular area of tropical ecology.  La 
Selva research station near Sarapaqui was established in 1954 by Dr. Leslie Holdridge as 
an experimental farm and passed into the hands of OTS in 1968.  Since then it has 
become one of the most important sites in the world for research on tropical primary 
forests (OTS, 2010).  Many research projects are actively being pursued at this location, 
and a significant portion of them relate to conservation of native plants. 
ProNativas, a non-profit organization, is a consortium of individuals and 
organizations that are concerned with the conservation and use of native plants in Costa 
Rica.  Having only started within the last decade through the efforts of Willow 
Zuckowski, ProNativas is still in the process of establishing their presence in the 
9 
 
landscaping network of Costa Rica.  Collaborative research by ProNativas on the use of 
certain native plants for both the private home owner as well as for ecosystem restoration 
is proceeding at the La Selva OTS station as well as several other locations including 
private residences, commercial nurseries, and universities. 
In Costa Rica, privately owned businesses and individuals are often deeply 
involved with environmental conservation.  Selva Verde Lodge in Chilimate is a Costa 
Rican ecolodge that is invested on several levels with the conservation of its surrounding 
environment.  Considered a pioneer of sorts, the lodge was built in the 1970?s and still 
remains committed to the tenents of sustainable tourism.  Currently it holds a rating of 
three leaves from the Certification for Sustainable Tourism (CST) Rating Program.  As a 
500 acre private reserve, Selva Verde provides protection to both primary and secondary 
rainforest ecosystems and the biodiversity contained within. 
The CST program was formed to help categorize tourism operations based on a 
single set of guidelines (Honey, 2008).  Certification is based on four criteria.  First, 
physiological-biological parameters are used to assess the interaction of the company 
with its surrounding habitat.  Second, the infrastructure and services are evaluated based 
on management policies and operational systems for lodging companies (this relates to 
energy conservation practices, waste disposal, and water pollution).  Third, the level to 
which the company interacts with its patrons to encourage participation in the 
sustainability practices of the establishment is assessed.  Finally, the socio-economic 
environment is evaluated.  In this lat category, the interaction of the company with the 
local community is evaluated based on the degree to which the tourist company assists 
with growth and development of the region and the generation of new employment.  Leaf 
10 
 
levels 1-5 are assigned yearly, and participation is entirely voluntary and open to any 
company that is interested.  Only seven lodges in Costa Rica currently hold a rating of 5 
leaves, and only four tour companies hold 5 leaf ratings.  The official entity in charge of 
this program is the Instituto Costarricense de Turismo (ICT).  The University of Costa 
Rica (UCR) represents the educational sector, while INBio and the Ministerio de 
Ambiente y Energia (MINAE) represent the public sector (CST, 2010).  All members are 
part of the evaluation process. 
 Several government agencies are involved in the process of environmental 
conservation as well.  The majority of responsibility and influence lies with two 
particular entities: MINAE and MAG.  MINAE is responsible for implementation, 
management and the administration of programs associated with biodiversity in Costa 
Rica (MINAE, 2010).  This government agency coordinates all issues regarding 
conservation and natural resources in the country.  Units within MINAE include (1) the 
Costa Rican Technical Unit, (2) Geology and Mining, (3) Meteorology, and (4) the 
Sistema Nacional de Areas de Conservacion (SINAC).  SINAC specializes in policies 
related to conservation, environmental management, and the network of conservation 
areas in Costa Rica.  During the past few years MINAE has been focused on ensuring 
that the information accumulated from various studies is actively utilized for the benefit 
of protected areas.  Ecotourism, biosphere projects, environmental service payments, and 
medicinal plantations are just some of the programs that have been implemented.  MAG, 
on the other hand, is specifically responsible for managing issues related with agricultural 
crops and livestock (MAG, 2010).  This includes issuing phytosanitary certificates for the 
export of any live plant or animal material out of the country.  Scientific research is also 
11 
 
conducted through this organization on topics relating to the fields of agriculture and 
livestock production; including pest and disease control, economic development, and 
marketing (to name a few). 
A number of other organizations in Costa Rica are actively participating in 
environmental conservation and/or ecotourism.  After careful research those described 
above presented the most promising opportunities for collaboration based on mutual 
interests and a common geographical focus.  This in no way permanently excludes any 
omitted organizations from future consideration nor guarantees a working relationship 
with those identified so far.  Two research trips to Costa Rica have been undertaken thus 
far to further determine the appropriateness of each organization and ascertain whether 
they would be amenable to working collaboratively with researchers from AU. 
Plant Propagation 
In the interests of facilitating both the expansion of conservation techniques as 
well as the overall conservation effectiveness of Costa Rica, propagation protocols for the 
following two species native to the Ca?o Negro area of northern Costa Rica were 
investigated: Hylocereus stenopterus (F.A.C. Weber) and Acoelorraphe wrightii Griseb. 
& H. Wendel. 
Hylocereus stenopterus 
H. stenopterus is a member of the Cactaceae, and was first described by Britton & 
Rose (1963).  It is a tropical hemiepiphyte (an epiphyte that can secondarily develop 
roots in the ground) that displays a liana-like growth habit, or growth where plants use 
trees and occasionally other objects for vertical support.  Development of adventitious 
roots allows this plant to acquire nutrients at microsites along the bodies of living plants 
12 
 
it climbs (Nobel and Barrera, 2002).  H. stenopterus is found only in Costa Rica.  Other 
members of the genus Hylocereus can be found throughout Latin America, where they 
are thought to have originated, as well as around the world in tropical and subtropical 
regions (Britton & Rose, 1963; LeBellec et al., 2006). There are currently 16 recognized 
species in the genus Hylocereus (Britton & Rose, 1963). These are all perennial plants 
that can grow quite rapidly and have high flower and fruit yield (Ben-Asher et al., 2006).  
As with many other members of the Cactaceae, these species utilize Crassulacean Acid 
Metabolism (CAM) (Britton & Rose, 1963).  The majority of species grow primarily in 
tropical dry forest habitats in the mid to lower canopy of forest trees (Britton & Rose, 
1963).  In Latin America several species in this genus, and their fruit, are collectively 
known as pithaya or pitahaya (other spellings can be found as well: pitahaja, pitajuia, or 
pitalla). In Vietnam it is called thang loy or dragon fruit (Nobel, 2002).  Though little 
known a decade ago, the pitahaya fruit is now an established item in the exotic fruit 
market (Mizrahi et al., 1997) as well as in domestic markets within production areas.  It 
continues to be recognized as a promising new crop (Janick, 1999), and the results of this 
research are intended to facilitate the advancement of its production not only as a cash 
crop for Ca?o Negro, Costa Rica but for other agriculturalists and horticulturists around 
the world. 
H. stenopterus consists of three-ribbed stems with a medium to large epigynous 
berry for a fruit (Nerd et al., 2002).  Britton and Rose (1963) describe it as a homoblastic 
species, meaning it displays few apparent changes in appearance between its juvenile and 
adult stages.  Stems are triangular, succulent, and feature spines 2-4 mm wide and 5-10 
mm long.  Flowers are hermaphroditic and 20-30 cm wide.  Most flowers of the various 
13 
 
species in the genus Hylocereus are creamy white, the two exceptions being H. 
stenopterus and H. extensus, which have red or pink blooms.  All bloom nocturnally.  
Though the cultivation of pitahaya is spreading, the majority of its consumption and use 
remains confined to the areas where it is grown.  As almost all commercial propagation 
within this genus has revolved around the one species Hylocereus undatus, the remainder 
of this review will focus on research done on this species. 
H. undatus is grown commercially in several countries in nurseries for retail sale 
as an ornamental plant as well as agriculturally for fruit production (Choi et al., 2007; 
Esquival et al., 2007; Merten, 2003; Yussof et al., 2008).  Large scale production in 
South and Central America occurs primarily in Columbia and Nicaragua with only a 
small amount occurring in Costa Rica (Barbeau, 1990), though several Asian countries 
now produce Hylocereus as well.  These include Thailand, Laos, Indonesia, Cambodia, 
and Japan with the majority of Asian production occurring in Vietnam (Nobel, 2002).  
Large operations have also begun to appear in Australia, Reunion Island and Israel 
(Jacobs, 1998; Le Bellec & Judith, 2002; Nerd & Mizrahi, 1997).  Other countries that 
produce H. undatus include the United States, Ecuador, Mexico, Guatemala, New 
Zealand, Peru, Spain, and the Philippines (Nobel & Barrera, 2002).  Overall, this species 
is considered to be fairly robust.  It can grow in temperatures from 12 to 40o C and at 
altitudes up to 2,750 m above sea level (De Dios, 2004).  All species can grow in various 
soils so long as they are well drained (Barcenas, 1994) and all species can survive and 
recover from drought with ease (Ben-Asher et al., 2006; Nobel & Barrera, 2002).  
Optimal light levels for specific species in this genus are unclear as studies show different 
levels for different species in the genus Hylocereus (Nerd et al., 2002; Raveh et al., 
14 
 
1998), but if grown in an extremely hot, dry climate some shade will be necessary (Raveh 
et al., 1998).    
In a commercial setting H. undatus is grown on some form of support, either 
vertical or inclined, and pruning is required (LeBellec et al., 2006).  Though these plants 
are capable of surviving drought, when regular flower and fruit set is desired regular 
watering is required (LeBellec, 2003; Nobel & Barrera, 2002; Nerd et al., 2002).  Le 
Bellec et al. (2006) recommend local micro-irrigation.  Suggested irrigation levels range 
from daily watering (Cavalcante & Martins, 2008) to watering once every two weeks 
(Thomson, 2002) and is affected by season and weather conditions in general.  Several 
papers explore methods for commercial propagation of this species though the practice 
only began around 15 years ago (Andrade et al., 2006; Cavalcante & Martins, 2008; De 
Andrade et al., 2007; De Andrade et al., 2008; Michel-Rosales & Farias-Larios, 2007; 
Mohamed-Yasseen, 2002).  Both asexual and sexual propagation have been attempted.  
Though propagation by seeds is possible, germination is often inconsistent (LeBellec, 
2003).  Pollination can be problematic in agricultural settings (Weiss et al., 1994) as the 
majority of pollination in the wild occurs via pollination by bats (Nerd & Mizrahi, 1997).  
Hand pollination is fairly easy to accomplish however and is only restricted by the labor 
associated with it for large scale production (Weiss et al., 1994).  Micropropagation has 
also met with success, but the cost of production is currently not met by the profit of 
distribution (Mohamed-Yasseen, 2002).  LeBellec (2003) concluded that asexual 
propagation by cuttings represents the most effective and promising form of propagation, 
at least for agricultural purposes.  He concluded that propagation by cuttings allows for 
reliable production of specific varieties, and the fruiting stage is reached three times as 
15 
 
quickly than with plants grown from seed.  This is due to ontogenetic aging, wherein the 
removed cuttings perpetuate the ontogenetic age of the parent plant and are already at a 
mature enough stage to begin flower and fruit production.  Cuttings can also be taken 
with ease directly in the field as opposed to other plants that might require special care, or 
the laborious process associated with producing and collecting seeds from this particular 
plant.   
Using cuttings, LeBellec (2003) achieved up to 90% rooting success.  Cavalcante 
& Martins (2008) suggested that cuttings be taken from juvenile growth with the basal 
end cut being made just above a joint in the fleshy green stem and the terminal end being 
an actual joint. Suggested cutting lengths vary between 25 and 50 cm (Cavalcante & 
Martins, 2008; LeBellec, 2003).  Bastos et al. (2006) concluded that application of a 
rooting hormone aids in rooting success, but Thomson (2002) reported no significant 
effect.  Thomson (2002) also suggest allowing cuttings to cure or cicatrize for up to 5 
days before planting while Cavalcante & Martins (2008) planted cuttings immediately.   
All these studies indicate that, for H. undatus, cicatrization time and application 
of rooting hormone may influence propagation success.  Further research is necessary to 
make more specific recommendations for asexual propagation of not only H. undatus but 
H. stenopterus and H. costariensis as well.  The current research will investigate stem 
cutting propagation for all three species. 
Accoelorraphe wrightii 
Aracaceae is an isolated and distinct group within the plant kingdom and is 
composed entirely of woody monocotyledonous plants collectively and commonly 
known as palms (Jones, 1995).  Palms are found around the world in tropical ecosystems.  
16 
 
There is no agreement on exactly how many species belong to this family: Meerow 
(2002) estimated 2600 species spread among about 200 genera.  All are perennial, and the 
family contains both monoecious and dioecious species.  Reproduction by palms in 
nature is almost exclusively by seed, and most palms have a fairly recognizable canopy 
with a prominent trunk and a characteristic crown of leaves. A few species are 
acaulescent (trunkless) and others are even climbers (Jones, 1995; Riffle & Craft, 2003).  
When trunks are present, they are found as a single stalk or as multiple clumping or 
caespitose trunks.   
Within this family, A. wrightii, also commonly known as Paurotis palm or 
Everglades palm, is a member of the Corypheae tribe and the Coryphoideae subfamily 
(Meerow, 2002).  It is found in Florida, the West Indies, and coastal or wetland areas of 
Central America (Farruggia, 2008; Jones, 1995; Meerow, 2002; Riffle & Craft, 2003).  
The genus Acoelorraphe is monotypic with A. wrightii the only species (Jones, 1995). 
This is a swamp-growing species with a clumping style growth habit.  It will often form 
dense thickets in brackish waters.  It has palmate fans that are divided more than half the 
way up the leaf into several slender sections (Meerow, 2002).  The trunk is covered with 
a think fibrous sheath that can be quite prickly (Jones, 1995; Meerow, 2002).  Moisture is 
extremely important to this species, and individuals in dry locations grow very slowly 
(Meerow, 2002).   
A. wrightii produces bisexual white flowers, and there are no published reports on 
its pollination method in the wild.  It is a relatively tolerant species when it comes to 
habitat, being one of the few palms that can grow in saturated soils (Meerow, 2002).  It 
17 
 
can have problems with manganese deficiencies but responds well to fertilization and has 
no known pest problems (Meerow, 2002). 
Though reproductive maturity is reached in four to five years by some species of 
palms, others can take up to forty or fifty years (Meerow, 2004).  Once flowering begins 
most palms are pleonanthic (flower regularly throughout their lives) though a few are 
monocarpic.  Pollination was historically considered to be by wind, but it is now thought 
that most pollination is done by beetles, flies, bees, and bats (Jones, 1995).  Seeds come 
in a range of sizes, with one species in this family (Lodoicea maldivica J. Gmelin) 
producing the largest seed of any plant in the world that can grow up to 40 pounds, yet 
seeds of some species are only a few millimeters long (Meerow, 2004).  This family also 
contains the species (Raphia regalis Becc) with the longest leaf of any plant at 24 m and 
the plant (Corypha Becc) with largest inflorescence, which can reach up to 7.5 m tall and 
contain thousands of tiny flowers (Jones, 1995). 
Propagation of palms is almost entirely done by seed, though vegetative 
propagation is possible by air layering, division, or even more recently, tissue culture 
(Jones, 1995).  The information on seed germination for palms tends to be inconsistent as 
much of it comes from hobbyists not researchers, and no clear conclusion can be drawn 
for more than only a few commercially important species.  Vegetative propagation is 
almost exclusively used by enthusiasts and collectors while tissue culture is used by 
researchers, collectors, and large scale commercial operations.  Neither tissue culture nor 
vegetative propagation is feasible for the average nursery or farm.   
Unlike A. wrightii, seed germination in most palms is notoriously difficult with 
some seeds taking years to germinate (Odetola, 1987).  It has been estimated that up to 
18 
 
25% of all palms take over 100 days to germinate and often achieve less than a 20% 
germination rate (Tomlinson, 1990).  Germination of A. wrightii usually occurs within 2 
to 4 months (Carpenter, 1988; Jones, 1995; Meerow, 2002).  Efforts to produce palms 
commercially persist, however, as certain species are significant in the ornamental and 
edible markets (Jones, 1995).  Some species are also important in indigenous lifestyles as 
fiber, food, and building material (Potvin et al., 2003). 
There is no clear evidence why this difficulty in germination occurs, as very few 
seed germination studies have been conducted for palms (Meerow, 2004).  Most palms 
do have small embryos compared to the size of the seed and the endosperm (Corner, 
1966).  Often, the embryo of palms is not fully developed when the seed is disseminated.  
Several species have an embryo that initially consists of a small disk of cells located near 
the operculum (Ginieis, 1957; Corner, 1966).  Only after a series of complicated chemical 
changes does the embryo break dormancy, develop fully and fill the entire seed 
(DeMason, 1985). 
Germination itself begins when certain enzymes reach the endosperm of the seed 
and mobilize the food resources there.  The endosperm itself can be easily recognized in 
the species Cocos nucifera L. as the white ?meat? and milk in a coconut.  In A. wrightii 
the endosperm is extremely hard and has the appearance and consistency of hard plastic 
(personal observation).  In general, the endosperm of palm seeds is made up primarily of 
cellulose, a non-nitrogenous storage material, (Orozco-Segovia et al., 2003) and lipids 
which are also an important source of energy in embryo development (DeMason, 1986).  
Germination in several species begins with the differentiation of the cotyledon into three 
structures; the tubular base, the petiole, and the haustorium (Tomlinson, 1990).  The 
19 
 
haustorium is the structure that absorbs nutrients from the endosperm while the embryo 
and cotyledon swell and develop.  Seed germination among palms occurs in several 
forms but can generally be divided into two initial categories; remote or cryptocotylar 
germination and adjacent or hypogeal germination (Meerow, 2004; Orozco-Segovia et 
al., 2003).  Seed germination in A. wrightii occurs by the remote germination pathway 
(Meerow, 2002).  With remote germination the first structure to emerge from the seed is 
called the cotyledonary petiole; this is commonly mistaken for the first seedling root.  
However, the first seedling root will actually emerge from this structure after it has 
grown down into the soil.  At the other end, the seedling shoot or plumule will emerge.  
The actual cotyledon remains inside as the absorptive structure and is known as the 
haustorium.  Adjacent germination involves only a small portion of the cotyledon 
emerging from the seed; this is known as the button (Meerow, 2004).  Then, the radicle 
and plumule emerge from the button.  The radicle in this type of germination is very short 
lived and is quickly replaced by adventitious roots that form around the stem base.  Like 
remote germination, the part of the cotyledon known as the haustorium remains inside the 
seed and absorbs nutrients from the endosperm.  These different germination strategies 
have been associated with establishment in different ecosystems (Orozco-Sergovia et al., 
2003). 
Seed dormancy has been identified as an important aspect of palm seed 
germination and may be a significant part of understanding the long and sporadic 
germination of many palm species (Orozco-Segovia et al., 2003).  Other possibilities that 
have been proposed to explain sporadic palm seed germination include embryo 
immaturity at time of dissemination, hormonal controls, or inadequate environmental 
20 
 
factors to induce germination (Baskin & Baskin, 1998; Corner, 1966; Hussey, 1958; 
Nagao et al., 1980).  Moisture is one of the primary factors to consider when overcoming 
dormancy in palm seeds.  Indeed the most common form of scarification for palms is 
extended soaking in water for anywhere from 12 hours to up to 7 days (Broschat & 
Donselman, 1986; Kheong, 1992; Moussa et al., 1998).  This practice is thought to help 
soften the often hard seed coats of many palm seeds, although well documented studies 
on the effectiveness of this treatment are in short supply (Orozco-Segovia et al., 2003, 
Ehara et al., 2001).  In cases of extremely hard or thick seed coats, soaking in 
concentrated sulfuric acid can be effective for shortening germination time (Merlo et al., 
1993).  Physical scarification accomplished by nicking the seed coat has also been used 
to reduce germination time and increase germination rate (Holmquist & Popenoe, 1967; 
Nagao et al., 1980; Odetola, 1987).   
Temperature can also play an important part in palm seed germination.  Most 
palms are thermophilous with optimum germination temperatures between 30-40o C 
(Carpenter, 1987; Ehara et al., 1998; Odetola 1987; Rees, 1962).  Cold stratification has 
been used in a few instances, but heating is far more common (Hussey, 1958; Nagao et 
al., 1980; Rees, 1962).  In general, the interaction of several factors affecting seed 
dormancy and germination is very poorly understood for palm seeds. 
In nurseries, appropriate germination temperatures are often achieved by the use 
of heated greenhouses and/or bottom heat cables.  Other methods reported as effective 
include covering containers with plastic or placing containers on heat-retaining surfaces 
(Meerow, 2004).  Appropriate light levels are important as well.  The majority of palm 
seeds can germinate and will grow in full sun but a few do require shade as they grow 
21 
 
naturally in the understory of thick rainforests (Meerow, 2004).  Planting depth can be 
adjusted to help compensate for overly sunny conditions.  Generally palm seeds can be 
planted at a depth equal to the diameter of the seed, but this is not always the case.  For 
many of the very large palm seeds, it is best to simply push the seed down into the 
substrate while part of the seed remains exposed (Jones, 1995; Meerow, 2004).  If a palm 
species usually grows in a shady location it can be planted at a deeper depth in a sunny 
location and still germinate effectively (Meerow, 2004).  Substrate for palms needs to be 
well-drained yet have good moisture holding capacity.  Germinating palms do not 
respond well to alternating periods of dry and wet.  A common mix in the nursery 
industry is a 1:1 ratio of peat moss:perlite (Meerow, 2004).  Palms have also been 
germinated in perlite or peat moss, but a 1:1 mix of the two appears to be most effective 
(Carpenter, 1988; Jones, 1995; Koebernik, 1971).   
Containers for palm propagation come in many shapes and sizes with the primary 
consideration being that they allow adequate growing space for the seedling to develop 
and that they be well drained (Meerow, 2004).  Flats are not generally recommended 
because of shallow soil depth, therefore specially made individual cells or containers are 
used.  In the tropics, many palms are sown directly in large raised beds, and some palms 
can only be grown in the ground because as they grow a sinker root that can be up to a 
meter long before a leaf shows above ground.  Once this long root has developed, the 
plant will often not tolerate transplanting (Jones, 1995).   
Most palms require uniform moisture during germination and subsequent seedling 
development (Meerow, 2004).  It is important that the substrate stay damp but not be 
saturated since palm seeds can be susceptible to rot (Koebernik, 1971).  Fertilization of 
22 
 
palms is not important until late in seedling development.  For the first few months after 
germination the endosperm acts as the only source of nutrition for the developing 
seedling and added nutrients can damage the new seedling (Meerow, 2004).   
Only one reported research project directly studied propagation of A. wrightii.  It 
was concluded that A. wrightii will germinate best at temperatures above 35o C and 
within a range of 33o to 39o C (Carpenter, 1988).  The current research will investigate 
the effect of scarification method and sowing substrate on the germination rate and final 
germination percentage of Acoelorraphe wrightii seeds. 
Research Objective 
 Plant propagation and subsequent outplanting can be used as an effective 
conservation technique (Potvin et al., 2003), and ecotourism can benefit both local 
economies and the local environment (Campbell, 2002).  Based on these two 
observations, the objective of this research will be to determine effective propagation 
techniques for two plant species native to the Ca?o Negro NWR so that they may be 
produced, harvested, and transplanted by community members to stimulate the economy 
of Ca?o Negro village and help conserve the unique ecological community of the Ca?o 
Negro NWR. 
 
 
 
 
 
 
23 
 
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Agriculture & Environment 6:305-311 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
30 
 
Chapter II 
Development of a Cooperative Extension Program in Costa Rica 
Key Words: Costa Rica, International extension, Ca?o Negro, conservation 
Abstract: A joint extension program between Auburn University (AU) and the 
Universidad Nacional de Costa Rica (UNA) was developed in an effort to expand 
educational opportunities at participating universities, conserve global biodiversity, and 
support community level economic development.  AU researchers made two research 
trips to Costa Rica in order to develop this relationship in Sept. 2009 and May 2010.  In 
Sept. contact was made with several professional entities as well as community members 
of Ca?o Negro village to assess the feasibility and validity of the proposed research 
program.  Professional relationships were established with not only UNA Sarapaqui but 
also with the Instituto Nacional de Biodiversidad (INBio) and the Ministerio de Ambiente 
y Energia (MINAE).  Upon returning in May, a research bed was built at the UNA 
Sarapaqui campus, and relationships with all appropriate parties were reinforced. 
Introduction 
In northern Costa Rica the Ca?o Negro National Wildlife Refuge (NWR) is an 
area with the capacity for both conservation collaboration as well as the opportunity for 
economic development.  Though the majority of the country is relatively stable 
economically, this remote region is facing other challenges.  The Costa Rican economy 
currently relies heavily on agriculture, exports by the electronics industry, and tourism 
(CIA, 2010).   
31 
 
Neither of the first two options is feasible in the Ca?o Negro area: agriculturally 
because of the sensitive nature of the various ecosystems that dominate the region and 
industrially because electronics manufacture is unreasonable due to the remote nature of 
the village as well as the lack of necessary infrastructure.  Furthermore, the few available 
jobs in the region are subject to decreased wages due to extensive illegal immigration 
from Nicaragua.  It has been suggested, however, that there is potential for program 
development and capitalization of the ecotourism industry which, if handled properly, 
could be a boon to both the economy as well as the ecosystem (Orozco-Obando, 2009).  
Internationally targeted cooperative extension programming can bring together not only 
the local and national resources of an area but also add a global perspective that can aid 
the development of ecosystem assets and enhance both economic opportunities and 
conservation goals (Harder et al., 2010; Ludwig, 2002; Place et al., 2008). 
The Ca?o Negro NWR is also highly significant as an ecosystem.  The majority 
of the area is a tropical dry forest, the most endangered ecosystem on Earth (Janzen, 
1988).  It also contains portions of a vast seasonal wetland that feeds Lake Nicaragua to 
the north and has extensive populations of birds, caimans, and unique plant life.  Ca?o 
Negro has been recognized by the Ramsar Convention (RAMSAR, 2010) as a wetland of 
international significance and is also part of the Mesoamerican biological hotspot 
established by the organization Conservation International (CI, 2010).  Additionally it is 
one of the most remote parks in Costa Rica?s national park system and has, thus far, 
escaped the degradation of native plant and animal communities often associated with an 
increase in visitor traffic.  Economic opportunities in the area are associated with the 
wetland.  As tourism continues to expand, improper planning might introduce these 
32 
 
problems to Ca?o Negro as well.   The area is also in danger from large scale agricultural 
interests.  As international pineapple growers are moving out of Hawaii and into cheaper 
production areas, most notably Brazil and Costa Rica, the wetland areas of Ca?o Negro 
have come under particular attack (Niesse, 2006).  Portions of the wetland have already 
been drained to make way for pineapple fields and Ca?o Negro community members are 
being asked to sell their lands for more of the same (Ruiz, 2010).   
Los Chiles is the only municipal area in close proximity to the refuge that could 
be considered to be larger than a village.  This is the northernmost town in the Huetar 
Norte Region of Costa Rica, lying only 10 km south of the Nicaraguan border.  From this 
location, tourists can take boat tours along the San Juan River, the official border between 
Nicaragua and Costa Rica.  However, this is a border that is still at times violently 
contested, with many Nicaraguans claiming that the entire river belongs to their country, 
and Costa Ricans claiming that the border is along the middle of the river.  One of the 
guides encountered during lunch in Los Chiles said that there is even occasional gunfire 
exchanged along the river.  This issue has been ostensibly resolved by the governments 
of these neighboring countries, but reality is not so cut and dry. 
The overall and ongoing goals of this project will be to facilitate rural community 
economic development and bring in supplemental income for local citizens by providing 
a new crop with commercial value as well as establish an ecotourism destination in the 
village of Ca?o Negro which is located within the Ca?o Negro NWR.  The effect of 
ecotourism on both rural development and conservation goals has been studied in several 
countries with mixed results that are attributed to several variables (Bookbinder et al, 
1998; Campbell, 1999; Hvenegaard & Dearden, 2002; Potvin et al., 2003; Trejos & 
33 
 
Chiang, 2009).  Propagation of native plants and subsequent out planting has been an 
effective conservation tool (Potvin et al., 2003).  In Nepal, it was observed that 
ecotourism did not contribute significantly to the local economy (Bookbinder et al., 1998) 
but other studies are more optimistic about the benefits of ecotourism (Campbell, 1999; 
Hvenegaard & Dearden, 2002).  Challenges to development include finding ways to keep 
monetary benefits within a community and targeting the focus of that community on how 
their natural environmental resources can be utilized to enhance their economic 
opportunities.  The proximity of a protected area to Ca?o Negro represents an opportunity 
for collaboration with larger government entities and promises a higher chance of success 
in the development of the local economy (Plummer & Fennell, 2009).  Utilizing 
recommendations from research based models, our educational extension program will 
provide farmers and local residents an economic alternative to selling their lands to the 
large pineapple producers that are moving into the area (Ruiz, 2009).  This will help not 
only preserve a unique and beautiful community but further protect a globally significant 
area of species diversity. 
To develop a program in Costa Rica, extensive research was done to identify 
appropriate collaborative entities.  These included the Universidad Nacional de Costa 
Rica (UNA), the Instituto Tecnologico de Costa Rica (ITCR), the Instituto Nacional de 
Biodiversidad (INBio), the Organization for Tropical Studies (OTS), ProNativas, the 
Ministerio de Ambiente y Energia (MINAE) and the Ministerio de Agricultura y 
Ganaderia (MAG) among others.  Preliminary research to identify regional native plant 
species appropriate for this project was verified by consultation with Costa Rican 
academic colleagues, regional experiment stations and community farmers.  Individuals 
34 
 
from each of these groups have formed an informal working group and advisory 
committee that provided local and national educational and political contacts to facilitate 
both conservation and development activities.  
Administration and faculty from UNA were among the first contacts made when 
efforts began to evaluate the biological, political, and economic factors associated with 
the feasibility of this project.  A relationship had already been established in February 
2008 between Auburn University (AU) and UNA personnel.   Later that year, a 
memorandum of agreement between AU and UNA was drafted and signed to facilitate 
future cooperative efforts between these institutions.  This project represents the next step 
in the development of that relationship. 
UNA is a public institution and receives funding directly from the Costa Rican 
government (UNA, 2010).  It is a four year university with both undergraduate and 
graduate degrees including the social sciences and multiple fields in the biological 
sciences, as well as philosophy and liberal arts.  It includes six separate campuses; the 
main campus is in San Jose with satellite campuses in Sarapaqui, Nicoya, Benjamin 
Nunez, Coto, Perez Zeledon, and Liberia. 
Another university contacted in Costa Rica that focuses on agriculture and the 
biological sciences in general is ITCR.  ITCR also concentrates on community and 
applied technologies.  Interdisciplinary studies are recognized as being of great benefit 
and every effort is made to foster collaboration with other universities.  There is a fully 
developed distance learning PhD program which is organized in conjunction with several 
universities in several countries.   
35 
 
INBio is a nonprofit organization that has been expanding its influence across the 
globe.  Started in 1989, INBio has become an international authority on programs and 
projects that conserve and promote tropical biodiversity (INBio, 2010).  During the late 
1980?s Costa Rica officially recognized the need to protect its wealth of biodiversity by 
passing Executive Decree No. 19153.  Among other things, this decree established the 
Planning Commission for the National Institute of Biodiversity, a board composed of 
members from various governmental institutions and also from higher education centers 
and conservation organizations.  This board recommended to the Costa Rican 
government that a state biodiversity institute be created which functioned with a high 
degree of autonomy.  Due to issues within the government itself, this recommendation 
was ignored.  At this point members of the board decided to take the situation into their 
own hands and create a private non-profit organization.  This institution was granted 
official recognition by the standing government, and has been supported by successive 
governments as well.  The members then went about the laborious process of securing 
funding, eventually finding it in the form of a loan for 80 million colones as well as 
backing from the Swedish Cooperation Agency (SIDA) and the MacArthur Foundation.  
On October 26th of 1989 a location was selected in Santo Domingo de Heredia and this is 
still where INBio is housed today (INBio, 2010).  Several services are now offered 
through INBio including bioprospecting, consulting, education and training, as well as 
general information services.  In 2000, a theme park called INBioparque was added as 
well.  Visitors can tour an organic farm, view a green built house, enjoy a butterfly 
garden, as well as many other nature based attractions.  ATTA, the database where all 
36 
 
information generated and collected by INBio is stored, is an extensive resource and 
almost indispensible for someone working with tropical biodiversity. 
OTS is another major research association in Costa Rica.  AU has a long standing 
relationship with this organization and has been an OTS member institution since 1987.  
Classes are offered on a number of subjects pertaining to tropical ecology at both the 
graduate and undergraduate levels and in multiple countries.  There are three research 
stations in Costa Rica and each specializes in a particular area of tropical ecology.  La 
Selva research station near Sarapaqui was established in 1954 by Dr. Leslie Holdridge as 
an experimental farm and passed into the hands of OTS in 1968.  Since then it has 
become one of the most important sites in the world for research on tropical primary 
forests (OTS, 2010).  Many research projects are actively being pursued at this location 
and a good portion of them relate to conservation of native plants. 
ProNativas, a non-profit organization, is a consortium of individuals and 
organizations that are all concerned with the state of native plants in Costa Rica.  Having 
only started within the last decade through the efforts of Willow Zuckowski, ProNativas 
is still in the process of establishing their presence in the landscaping network of Costa 
Rica.  Joint research on the use of certain native plants for both the private home owner 
as well as for ecosystem restoration is proceeding at the La Selva OTS station as well as 
several other locations including private residences, commercial nurseries, and 
universities. 
In Costa Rica, privately owned businesses and individuals are often deeply 
involved with environmental conservation.  Selva Verde Lodge in Chilimate is a Costa 
Rican ecolodge that is invested on several levels with the conservation of its surrounding 
37 
 
environment.  Considered a pioneer of sorts, the lodge was built in the 1970?s and still 
remains committed to the tenents of sustainable tourism.  Currently it holds a rating of 
three leaves from the Certification for Sustainable Tourism (CST) Rating Program.  As a 
500 acre private reserve, Selva Verde provides protection to both primary and secondary 
rainforest ecosystems and the biodiversity contained within. 
The CST program was formed to help categorize tourism operations based on a 
single set of guidelines (Honey, 2008).  Certification is based on four criteria.  First, 
physiological-biological parameters are used to assess the interaction of the company 
with its surrounding habitat.  Second, the infrastructure and services are evaluated based 
on management policies and operational systems for lodging companies (this relates to 
energy conservation practices, waste disposal, and water pollution).  Third, the level to 
which the company interacts with its patrons to encourage participation in the 
sustainability practices of the establishment is assessed.  Finally, the socio-economic 
environment is evaluated.  In this lat category, the interaction of the company with the 
local community is evaluated based on the degree to which the tourist company assists 
with growth and development of the region and the generation of new employment.  Leaf 
levels 1-5 are assigned yearly and participation is entirely voluntary and open to any 
company that is interested.  Only seven lodges in Costa Rica currently hold a rating of 5 
leaves and only four tour companies hold 5 leaf ratings.  The official entity in charge of 
this program is the Costa Rican Board of Tourism (ICT).  The University of Costa Rica 
(UCR) represents the educational sector while INBio and the Ministerio de Ambiente y 
Energia (MINAE) represent the public sector (CST, 2010).  All members are part of the 
evaluation process. 
38 
 
 Several government agencies are involved in the process of environmental 
conservation as well.  The majority of responsibility and influence lies with two 
particular entities: MINAE and MAG.  MINAE is responsible for implementation, 
management and the administration of programs associated with biodiversity in Costa 
Rica (MINAE, 2010).  This government agency coordinates all issues regarding 
conservation and natural resources in the country.  Units within MINAE include (1) the 
Costa Rican Technical Unit, (2) Geology and Mining, (3) Meteorology, and (4) the 
National System of Conservation Areas (SINAC).  SINAC specializes in policies related 
to conservation, environmental management, and the network of conservation areas in 
Costa Rica.  During the past few years MINAE has been focused on ensuring that the 
information accumulated from various studies is actively utilized for the benefit of 
protected areas.  Ecotourism, biosphere projects, environmental service payments, and 
medicinal plantations are just some of the programs that have been implemented.  MAG, 
on the other hand, is specifically responsible for managing issues related with agricultural 
crops and livestock (MAG, 2010).  This includes issuing phytosanitary certificates for the 
export of any live plant or animal material out of the country.  Scientific research is also 
conducted through this organization on topics relating to the fields of agriculture and 
livestock production; including pest and disease control, economic development, and 
marketing (to name a few). 
A number of other organizations in Costa Rica are actively participating in 
environmental conservation and/or ecotourism.  After careful research those described 
above presented the most promising opportunities for collaboration based on mutual 
interests and a common geographical focus.  This in no way permanently excludes any 
39 
 
omitted organizations from future consideration nor guarantees a working relationship 
with those identified so far. 
Materials and Methods  
The first exploratory mission to Costa Rica took place from 27 September 2009 
through 4 October 2009 (Figure 1).  A combination of teaching, research, and extension 
personnel were present including Dr. Amy Noelle Wright, horticulture associate 
professor and primary investigator, Rachel Allen Meriwether, horticulture graduate 
research assistant at Auburn University, Warner Orozco-Obando, horticulture graduate 
research assistant at Auburn University as well as liaison and translator for the trip, and 
Monica Jenkins DeTure, director of distance learning at Auburn University.   
 Monday, 28 Sept., was spent in meetings with various officials from both non-
profit and academic institutions.  On the first morning after arrival contact was made with 
the non-profit organization INBio.  A meeting was held with Nelson Zamora, Director of 
Botany, Alvaro Herrera, Director of Research, Jesus Ugalde-Gomez, Associate Director 
of Biodiversity, and Elena Galante Marcos, Director of the Rio Frio Project.  Discussion 
began with introductions by both groups (INBio and AU) highlighting their various 
programs and interests and proceeded into details on how a cooperative relationship 
might develop between INBio and Auburn.  Once the formal meeting was complete, 
everyone toured INBioparque. 
AU personnel then traveled to the campus of UNA in Heredia to discuss 
collaborative opportunities with this institution.  While visiting UNA, a presentation was 
given by the author outlining the project and the objectives for research in Costa Rica.  
Several faculty and staff members from UNA attended and were encouraged to make 
40 
 
suggestions and consider becoming involved with the project.  After the meeting, Victor 
Ruiz Obando, Regional Coordinator for the Huetar Norte Region, expressed interest in 
the project, even offering assistance during further travels through the country. 
Tuesday 29 Sept., the party left Heredia.  Driving north from the Central Valley, 
the travel ended in Chilimate at Selva Verde Lodge, one of the very first ecolodges built 
in Costa Rica.  While in Chilimate AU personnel were joined by Victor Ruiz and the 
opportunity became available to also visit the UNA Sarapaqui campus in Rio Frio for a 
tour of their facilities.  After the tour of UNA Sarapaqui on Tuesday, Ruiz and AU 
personnel traveled to the La Selva OTS field station for a basic nature tour with one of 
the OTS volunteers/guides. The following day was spent traveling to Batan where a tour 
was given of the new UNA aquaculture research center. 
 On 1 Oct., the group traveled north to Los Chiles and Ca?o Negro.  The first stop 
up north was made in the town of Los Chiles followed by a brief stop was made at the 
Nicaraguan border.  From the border, the next stop was the actual village of Ca?o Negro 
for two nights.  Upon arrival a meeting was scheduled with several community leaders 
from Ca?o Negro as well as the few resident biologists in the area.  It is extremely 
important when making decisions about development to be assured of community 
participation (Honey, 1999).  Those present included Don Lolo, one of the oldest 
inhabitants of Ca?o Negro village, Oscar Gutierez, Oscar?s young son, and Oscar?s wife 
Christina who is a biologist with the Ministerio de Medio Ambiente y Recursos Naturales 
(MARN).  MARN is an environmental organization based in El Salvador and is 
concerned with environmental assessments and international cooperation.  Also in 
attendance were Bernadita Aguilar Lopez, a woman who owns a small restaurant and 
41 
 
guest house called Tablitas located not far from Ca?o Negro and Victor Ruiz.  Ruiz is the 
equivalent of an extension agent for the area and is based out of the UNA Sarapaqui 
campus.  Mick Wigal, a member of the United States Peace Corp, was also present.  
Warner Orozco lead the meeting since it was almost exclusively in Spanish.  He began by 
describing the design and intention of the AU project to those present.  It was also made 
clear that the primary reason for calling this meeting was to receive suggestions from 
those present on how to make the project applicable and appropriate for Ca?o Negro.  
Once the introduction was complete, the discussion was opened up to questions and 
suggestions.  At this point Juan Diego Alfaro Hernandez, director of research for the 
Huertar Norte conservation area, and Carlos Alvarez arrived and joined the discussion. 
On the following day, 2 Oct., a visit was paid to the local MINAE office.  Juan 
Diego Alfaro Hernandez took the time to meet us in Ca?o Negro to discuss our plans and 
theirs for the area.  A presentation was given by Hernandez outlining plans for a large 
visitor and research center to be built in Ca?o Negro.  After the meeting at MINAE, a 
boat tour into the wildlife refuge was taken with a local tour company.  Kingfisher Lodge 
and Tours is owned by Antonio Sequeira and his wife, both of whom have lived in the 
Ca?o Negro area most of their lives.  The opportunity to see the lodge was not presented. 
Once the boat tour was completed a further tour was given by Don Hernandez of 
the building site where the new MINAE facility will be located.  After the tour of the 
proposed new facilities was complete, AU personnel left Ca?o Negro traveling west and 
stopped for lunch and further discussion and planning at Rancho las Tablitas, the 
restaurant and bar run by Bernadita Aguilar Lopez.  Following lunch, the group traveled 
to Cuidad Quesada (San Carlos), the home of the Instituto de Tecnologico de Costa Rica 
42 
 
(ITCR).  Overnight accommodations were provided by the school which has a hotel 
facility that is also a training area for Tourism students at ITCR. 
On 3 Oct., a meeting was held with Dr. Tomas de Jesus Guzman Hernandez, the 
director of the Doctoral program in Natural Sciences at ITCR.  A short tour was given of 
the campus by Ing. Oscar Lopez Villegas, a professor in computer sciences.  He works 
with small businesses primarily in the tourism industry and helps owners learn how to use 
the internet to advertise and develop their businesses.  This concluded activities at ITCR, 
and the remainder of the afternoon was spent traveling back to San Jose.  The final day 
spent in Costa Rica was 4 Oct. with all party members flying home that afternoon. 
A second research trip was conducted 5-16 May 2010 (Figure 1).  Along with 
following up on relationships established on the first trip, the intention of this trip was to 
establish research plots in Ca?o Negro.  Plans also included collecting cuttings of H. 
stenopterus to bring back to Auburn to continue propagation research there.  Participants 
were limited to Dr. Amy Wright and Rachel Meriwether.  Warner Orozco-Obando was 
once again of assistance in organizing the trip and acquiring supplies, though he was 
unable to participate in the actual trip.  Departure and arrival locations were the same as 
on the first trip. 
Immediately upon landing in San Jose a visit was made to INBio for a short visit 
with Nelson Zamora.  There was not time to conduct further business on this day, and the 
remainder of the evening was spent at Hotel Bougainvilla making sure all preparations 
had been made for the following two weeks.   
Early on the morning of 6 May a meeting was scheduled at the Ministerio de 
Agricultura y Ganaderia offices.  It had been discovered that this was the location where 
43 
 
the phytosanitary certificates necessary to bring plants from Costa Rica into the USA are 
issued.  The office is located on one of the UNA campuses in San Jose.  Ing. Magda 
Gonzalez Arroyo is a director for the office of Servicio Fitosanitario del Estado (State 
Phytosanitary Services) and was kind enough to give a presentation on a research project 
her office had conducted several years before.  At the conclusion of this meeting, Warner 
Orozco was delivered to the airport and returned to Auburn.  Wright and Meriwether 
continued on to Sarapaqui and Selva Verde Lodge with a driver and van provided by the 
travel company Costa Rica Ambiance. 
A visit was paid to UNA Sarapaqui on the morning of 7 May.  After touring the 
campus, a more formal meeting was held with the current director of the UNA Sarapaqui 
campus Alvaro Villalobos and the future director Esteban Araya.  During this visit 
permission was given to use the greenhouse for our cooperative research project.  Any 
supplies that had not already been purchased were acquired from a nearby hardware store 
and all materials were left at the UNA campus near the greenhouse area until work could 
commence on the propagation bed.  Due to the decision to use the pre-existing 
greenhouse, work was ahead of schedule and the rest of the day was spent at the hotel 
working on AU related tasks.   
On 8 May time was spent catching up on work and making sure everything was in 
readiness to travel north to Ca?o Negro the following day.  Unfortunately, due to 
complications with the van, it was not possible to take the trip to Ca?o Negro and an 
additional four nights were spent at Selva Verde Lodge.  On 9 May Wright and 
Meriwether participated in a guided hike that took travelers through the forests located on 
Selva Verde Lodge property.  On 10 May Wright and Meriwether met with Victor Ruiz 
44 
 
at Selva Verde to discuss plans for continued collaboration.  This was a short meeting, so 
the majority of discussion occurred late that night over dinner in Chilimate.  On 11 May 
an investigatory boat tour was taken down the Rio Frio to determine what plant life was 
visible in the area as well as assess the quality and style of services for tourists. 
Work began the following day, 12 May, putting in propagation experiments in the 
greenhouse at UNA Sarapaqui.  On the way to the campus from Selva Verde a visit was 
paid to the La Selva OTS station.  It had been learned during the course of the trip that a 
gentleman worked there who was also interested in propagation of native plants and was 
connected with Pro Nativas.  Orlando Vargas is the head of Scientific Operations at the 
La Selva Biological Station and has several projects with native plants in progress.  Once 
conversation was concluded, Mr. Vargas gave a brief tour of his onsite propagation 
facility.   
Once the visit to OTS was concluded, Wright and Meriwether proceeded to the 
UNA campus.  On this first day of work lumber for the raised bed was treated with Penta-
Cell, the mist system was constructed, and all necessary landscape fabric was cut.  Penta-
Cell is a liquid product that is mixed with either water or diesel fuel and seals lumber 
with active the active ingredients borate and chromate.  This also served to announce the 
beginning of construction in the greenhouse so that the following day, when work that 
required assistance would occur, the necessary people would be available.  Upon 
returning the following day, 13 May, it was communicated that UNA staff had learned of 
our presence and were available to assist with completing the experimental set up.  
Carmen Daly assisted with the project and conversation revealed that she is the manager 
for day to day operations in and around the greenhouse.  She also helps direct student 
45 
 
workers from the university that have duties in the greenhouse area.  Work on 
constructing the propagation beds in the greenhouse resumed almost immediately this 
day.  This included nailing together the frame, lining it with ground fabric, filling it with 
substrate, and putting the mist system in place.  Once this was complete, work began on 
preparing the seeds for planting.  A. wrightii seeds were planted with the assistance of 
Evelyn Jimenez, a student worker from UNA.  She was responsible for managing the 
propagation experiment once AU personnel departed.  A. wrightii seeds received one of 
two scarification treatments, or no scarification (control) before planting.  Scarification 
treatments were soaking in water and scratching with a rat tail file.  All were planted in 
100% perlite in traditional black square 4 inch pots.  Seeds of Inga sensiblis purchased a 
few days before from a roadside vendor were also planted.  These were planted in a 
mixed substrate consisting of equal parts sand and perlite and smaller but equal portions 
of organic compost, saw dust, and coconut fibers.  I. sensibilis seeds in the pod are 
covered by a thick white pulp that is the edible portion of the fruit.  One hundred and 
sixty seeds were planted, half with white pulp removed and half with white pulp left 
intact.  Seeds were planted in the black plastic bags that are typical for tree propagation in 
Costa Rica.  These bags appear to be made of number 4 low-density polyethylene plastic 
similar to typical trash can liners in the USA.  They were roughly 20 cm tall and 13 cm 
wide (when lying flat), with three rows of evenly spaced drainage holes running from top 
to bottom.  A portion of the H. stenopterus cutting that was gifted from Orlando Vargas 
was also planted.  This concluded work at UNA Sarapaqui.  All left over supplies were 
donated to the campus, and instructions for maintenance were provided.  Contact 
46 
 
information was exchanged with Ms. Daly, and arrangements were made for updates on 
the propagation experiments to be sent to Auburn. 
The morning of 14 May was spent traveling from Chilimate to San Jose.  Upon 
arrival in San Jose, a quick visit was made to INBio to visit with Nelson Zamora.  Seeds 
from A. wrightii were left with Zamora with the understanding that propagation 
experiments would begin on this plant in Costa Rica.   
On the last full day in Costa Rica, 15 May, the University of Costa Rica (UCR) 
botanic garden, Lankester Jardin Botanico, and Orosi, one of the more prominent 
agricultural regions of San Jose, were visited.  Travel back to the United States took place 
on 16 May. 
Results 
INBio 
During the first visit to INBio, it was discovered that this organization has an 
existing aquaculture research center in Ca?o Negro.  This project is jointly funded and 
administered by INBio and a Spanish company (the actual name of this company was not 
made available).  Plans for the aquaculture research center and the Ca?o Negro area in 
general focus on sustainable development, ecotourism, education and outreach, watershed 
management, and community development.  Unfortunately, there are limited funds 
available for this program so it is proceeding slowly.  Within the relationship, INBio has 
only an advisory role on this project and is primarily available to make scientific 
recommendations when necessary.  The Spanish company involved is particularly 
interested in production of organic food crops but is currently focused on 
commercialization and marketing of these crops rather than actual production.  Members 
47 
 
of INBio felt that this may become problematic in Ca?o Negro as many communities in 
the area only collect and sell crops but do not produce them.  There is apparently also a 
new interest in developing an urban landscapes program that seeks to educate the general 
public on the use of native plants in home gardens.  Propagation of native plants and 
expansion of nurseries that sell them would be needed in order for this project to develop.  
It was recognized that the AU project could facilitate this plan. 
 All the members of INBio present at the meeting felt that the AU project was 
extremely interesting and worth pursuing.  They felt that this project had the potential to 
begin filling in some of the knowledge gaps in their research on the Ca?o Negro area.  
The idea of being involved with in situ conservation also appealed to Nelson Zamora in 
particular.  They also suggested incorporating a global warming component into the 
research project somehow, perhaps as a way to elicit further funding. 
 During travel in May 2010 and the second visit to INBio current development of 
the project was discussed as well as progress on both sides.  Consideration was given to 
the possibility of establishing another propagation facility at INBio where research being 
conducted at AU and UNA can be replicated as well as work with native plants and their 
growth.  It was also agreed that on the next research trip of AU personnel to Costa Rica, 
Zamora would plan to travel north with us to Ca?o Negro to visit research sites and that 
further effort would be made to solidify the relationship between Auburn and INBio by 
beginning formal propagation experiments at the INBio offices.  Construction of a 
greenhouse at INBio was discussed. 
UNA 
48 
 
 The meeting and presentation at UNA in Heredia in May received mixed 
response.  Some individuals did seem interested in the project, but communication with 
them occurred primarily after the main meeting.  A few helpful suggestions were made, 
including the need for an economic analysis of the plants selected for research.  
Establishment of a relationship with Victor Ruiz Obando was the most significant benefit 
of this meeting.  Some discussion did occur about the possibility of having a student from 
UNA being directly involved in the project, however, to date no relationship has been 
established with the UNA campus in San Jose. 
Instead, an affiliation has been developed with UNA Sarapaqui.  The campus 
itself is only a few years old with the clear focus being one of extension and 
environmental programs of various kinds.  There are specific areas dedicated to various 
aspects of farming, agriculture, and ecotourism.  Construction and development of 
facilities is increasing rapidly.  Don Ruiz gave an exceptional tour that clearly 
demonstrated his own pride in the work there as well as his wish to develop collaborative 
research with AU.  Accommodations for future trips were offered on campus as well as 
research space in any and all facilities onsite.  This was reiterated the next day during a 
brief final visit in the morning with the director of the campus, Alvaro Villalobos.   
 On the second visit to UNA Sarapaqui it was clear that the campus had developed 
quite a bit since the previous visit in October.  This happened to include a recently 
completed greenhouse with full overhead irrigation.  It had been the intention of Wright 
and Meriwether to build a small hoop house in which to house propagation experiments 
but, upon finding the new structure, the aim became securing access to and use of this 
facility.  During the meeting with the campus directors it was established that everyone 
49 
 
was agreeable to research proceeding on the campus in general but that special 
permission would be needed to use the greenhouse.  Meryll Arias and a colleague were in 
charge of managing use of the greenhouse if not the actual maintenance of the structure 
and joined the discussion.  After fully explaining the project all UNA representatives 
were satisfied with allowing use of the greenhouse for the plant propagation project.  
Some interest was expressed in incorporating the forestry and agriculture programs into 
the project, possibly even using the tree propagation experiments for teaching purposes. 
 Over the evening meal with Don Ruiz on the trip in May, he expressed a 
particular interest in growing H. stenopterus on the campus of UNA Sarapaqui.  He 
explained that the majority of this fruit consumed in Costa Rica is actually imported at 
high cost from Columbia.  He felt that if dragon fruit were to be produced locally, it 
would be an excellent source of income for participating communities.  There is already 
an agriculture program as well as a focus on community development at UNA, and 
Victor Ruiz seemed to think that it would be possible to incorporate research on this plant 
into the pre-existing departments.  Though there was not much time to discuss future 
plans in detail, mutual agreement was reached that the project was proceeding as it 
should and both parties remained pleased with the relationship. 
During the trip in Oct. 2009, the OTS tour was not very successful.  Though the 
tour itself was somewhat informative and some wildlife was seen, the group left feeling 
slightly disappointed.  The opportunity to speak with actual researchers was never 
presented and so no contacts were made.  There was more success upon returning to OTS 
in May.  After a brief explanation of the AU project, Orlando Vargas expressed definite 
interest in being involved.  He was particularly interested in the palm and the cacti.  A. 
50 
 
wrightii was not a species with which he was familiar and he was quite excited to learn 
more.  H. stenopterus did turn out to be one of the plants he is working with and he was 
even kind enough to make a gift of a cutting from one of the cacti growing on a tree just 
outside his office.  This was the first time we were able to see this particular species of 
cactus growing in Costa Rica.  It was agreed that every attempt would be made for 
Orlando to travel north to Ca?o Negro on the next research trip to discuss collaborative 
work further.  Until then, communication would be maintained via the internet, and 
results of interest would be shared. 
On the trip in Oct. 2009 the tour of the aquaculture facility proved to be 
fascinating and informative.  Though aquaculture was neither directly related to the 
authors? research nor an area of expertise for anyone in the party, we were able to see its 
relevance to future projects and the potential for further collaboration with the AU 
Fisheries Department.  It also continued developing the relationships that will be crucial 
for success of the overall project.  UNA has thus far created an exceptional aquaculture 
facility that is maintaining a focus on community development and sustainable harvesting 
methods.  There are some long term goals that pertain to this area of research and the 
possibility of incorporating research on lotus (Nelumbo lutea), which is being conducted 
at Auburn University by Dr. Kenneth Tilt and Warner Orozco-Obando.  Ca?o Negro also 
is an area that could benefit from the research being done at the UNA aquaculture center.  
With an existing gar fish (or gaspar as it is locally known) production program in place, 
improved methods of gar farming are in high demand. 
Ca?o Negro 
51 
 
Another unexpectedly interesting location was the international border between 
Costa Rica and Nicaragua; where the Costa Rican road literally ended in a field and the 
border itself was marked by a single strand of yellow caution tape stretched between four 
rusty barrels.  There was a guard station on site manned by four individuals.  A small 
camouflage hut was located just across the border on the Nicaraguan side.  It was unclear 
how many individuals were actually working at this guard house but at least one man was 
seen with a large automatic weapon.  There was no clear sign of an entrance road into the 
Nicaraguan side.  While there, several people walked across from Nicaragua into Costa 
Rica with no questions from the Costa Rican guards.  The people making this crossing 
were of normal Costa Rican appearance, being dressed in cotton clothing and primarily 
shod in flip-flops; one woman had on a flowered dress and seemed to be carrying a 
shopping bag of some sort.  There was no sign of any habitation or settlement on the 
Nicaraguan side.  The majority of the vegetation on the Nicaraguan side appeared to be 
orange plantations.  Upon speaking to the Costa Rican guards we were informed that 
these were indeed massive orange plantations on the other side of the border.  They 
further explained that land and labor in Nicaragua had recently become very cheap.  The 
fruit juice companies Dole and Ticofruit have for many years had large plantations in 
Costa Rica, but when prices began to fall in Nicaragua, they sold off their lands in Costa 
Rica and moved just across the border.  Apparently the original factories in Costa Rica 
are still in use, but the majority of the fruit and labor comes from Nicaragua now.  They 
said that many of the people in the area had lost jobs due to this migration, and some of 
them had little choice but to travel into Nicaragua to continue what they had been doing 
before, despite the fact that they were paid less and obviously had to travel further.  
52 
 
Though this was a short and unexpected stop it provided further insight into the 
agricultural economy of Costa Rica. 
Driving north through Agua Zarcas on the way to Los Chiles and the border it 
was clear that agriculture has become the primary economic activity of the area.  Aside 
from the occasional fields of ornamental plants, the land is dominated by sugar cane and 
pineapple production.  It also became clear upon arriving in Los Chiles that the town 
actively advertises itself as being the gateway to Ca?o Negro NWR, while in fact the 
town is not located in Ca?o Negro, and the boat tours never even enter the protected area.  
Many of the boats have the name Ca?o Negro stenciled on the side, and many other signs 
and stores indicate that this is Ca?o Negro. 
During meeting with community members and officials of Ca?o Negro, it was 
explained that sometime in the past the Ministerio de Recursos Naturales (MRN) had 
attempted a development project in the area, but when the administration changed within 
MRN no more support was received and the project was dropped.  There was quite a bit 
of concern over the same thing happening again with our project.  A clearer picture was 
also given of the gar project in the area as well as a river turtle project that is ongoing.  
The river turtle project in particular was used as an example of a successful project.  It 
was not clear what group started the project, but apparently they were very successful at 
involving and educating the local people about the project and its benefits.  It had been 
several years since the project had been handed over in full to the community members, 
and it was still operational and lucrative.  Each year, participating families collect turtle 
eggs from within the protected area.  These eggs are then hatched and the hatchlings 
raised in the research facility near the new MINAE offices.  Once the turtles have reached 
53 
 
the appropriate maturity level, one portion of the population is released back into the 
wildlife refuge and the other portion is sold for profit.  The entire operation is also used 
as a tourist attraction.  This is apparently not a community wide project, only about 3 or 4 
families are responsible for the majority of the work, but it appeared that everyone in the 
community is supportive and recognizes the benefits of this effort.   
The gar project is still in its infant stages, but those involved seemed excited 
about its prospects.  Apparently the plan will be to establish an aquaculture facility within 
the Ca?o Negro community that will fulfill multiple purposes.  First, it will act as a 
source of food and income for the residents.  Second, it will be used as an environmental 
restoration tool, as a portion of the fish that are raised will be released into the Ca?o 
Negro NWR and the San Juan River to build up the natural population.  This also has the 
potential to act as a source of revenue for the community if they are able to capitalize on 
the increased fish population through the attraction of sport fishermen, a sport which is 
quite popular in Costa Rica.  There are clearly similarities between these two projects and 
interestingly enough, both have the same similarities with the AU plant propagation and 
production project.  Ultimately the propagation of plants in Ca?o Negro would attempt to 
fulfill the same goals of generating food and revenue for the community while also 
providing material to begin environmental restoration efforts.   
Ca?o Negro community members are interested in developing the ecotourism 
industry in the area. Concern is mounting over the incursion of transnational pineapple 
corporations that are buying land on the edges of the wetland and draining it without 
concern or consideration being given to the environment or the community.  There is 
already some level of tourism though much business is lost to Los Chiles due to the 
54 
 
confusion about it being Ca?o Negro.  There is a large sign in the main square of Ca?o 
Negro village proclaiming it as the ?real Ca?o Negro?.  Most tourists do make the 
mistake of thinking that Ca?o Negro is just a wildlife refuge and not an actual village.  
The hope is that as more businesses related to tourism open up in Ca?o Negro, it will be 
easier for travelers to find the village. 
People present at the meeting were also adamant about the project remaining in 
the hands of local residents.  The one hotel in Ca?o Negro is owned by a foreigner, who 
does not contribute or participate in the community, so that tourism revenue does not 
remain in the area.  There are also other difficulties for Ca?o Negro as it is extremely 
remote and can be fairly hard to reach for the average traveler.  Thus it would be 
important to have businesses and attractions that will make it a sought after destination.  
Ca?o Negro is a young village having only been officially recognized in 1945 so many of 
the entities that tourists expect may not be present.  Perhaps the most common and 
lucrative tourism opportunity in the area is the boat tours that operate up and down the 
Rio Frio and San Juan River and take interested parties into the wildlife refuge.  Most of 
the tour boats are privately owned and operated by residents of Ca?o Negro village.  It 
would be hard to call any of these operations actual tour companies as no one person 
owns more than one or two boats, but the tours are run professionally and are quite 
enjoyable.  A few of the boat operators also offer a fishing tour option.  Boats that are 
used are quite typical for this sort of operation in Costa Rica.  They are 8 to 10 meter long 
flat bottom boats with rows of seats on each side, a roof, and are motor operated.  The 
majority of guides are at least partially bilingual.  There is also some hope that the native 
plant restoration efforts along the rivers will increase success of the boat tours.  As 
55 
 
previously mentioned, the river banks along the areas where the tour boats operate have 
become severely degraded and invasive plants continue to proliferate.  If native plants 
were to be outplanted along the banks, the natural beauty of the environment would be 
enhanced, particularly if the plants selected provided bold, ornamental features.  Plants 
that are good for attracting wildlife would also be beneficial as the majority of tourists are 
interested in viewing birds or monkeys rather than only the plants.  As with any 
ecosystem related project, there is no real end point or finite limit to the opportunities for 
positive impact.   
Suggestions were solicited and received during the meeting on which plant 
species would be most beneficial to the community.  There was agreement that the 
species already being considered as candidates for research were an acceptable 
beginning.  One additional species of tree in particular was mentioned, Lecythis ampla or 
the monkey pot tree.  A member of the Brazil nut tree family, this large tropical tree is 
quite rare.  Ultimately this dictated why this particular species was not selected for 
research due to the difficulty in finding an adequate number of stock plants for 
propagation.  A general suggestion was made to focus on tree species as these would be 
the most likely to have large environmental and ornamental impact and be lucrative for 
the community.  Additionally, it was noted that there are several rare tree species in the 
area.  These suggestions will be prioritized and considered not only during this project 
but in future projects as well. 
A small MINAE office/field station is currently in use in Ca?o Negro with a few 
permanent employees and volunteers on site at any one time.  Community members are 
also often involved with the activities of the office.  There are dorms where guest 
56 
 
researchers and volunteer firefighters can stay if necessary.  There was a decent lab 
building already present.  A large two story building with lab facilities on the bottom 
floor and office spaces on the second floor stands not far from the edge of the wetland 
itself and is much closer to the main square of the village than the current office space.  
This structure appears to be well built and will act as a separate research facility in the 
upcoming visitor complex.  This research center will be jointly administered by MINAE 
and UNA.  Behind this building were two large shade cloth tents that covered several 
raised benches and large aquatic tanks.  The benches are roughly a meter high and 4 
meters long.  They are filled with a peat based substrate in which the river turtle eggs are 
hatched before they are transferred to the tanks.  Everything was empty and not in daily 
use at this time.  It was explained that this is the area where much of the work for the 
river turtle project goes on, but that it only occurs during part of the year, and during the 
rest of the time it remains unused.  There was discussion on the possibility of using the 
same facility for plant propagation, as the benches used to hatch the turtle eggs could 
easily be converted into mist beds for propagation during the part of the year they are not 
being used for hatching. 
Plans for the new visitor center include areas for education, tourism, and research 
throughout the facility.  There was also a plan in place to landscape the facility using 
native plants and incorporate several green building features. 
Additional helpful suggestions were received while visiting the MINAE office, 
particularly with regard to Nymphaea ampla.  It was discussed that this plant would be 
excellent for mitigation of agricultural pollution in the wetland as well as helping to 
reduce total suspended solids (TSS), both of which are problems in the Ca?o Negro 
57 
 
wetland as well as the Rio Frio (Fernandez, 2009).  This represents additional 
opportunities for collaborative research and extension efforts.  Don Hernandez suggested 
that it would be of benefit for AU to work jointly with UNA to complete an initial water 
quality survey of the Ca?o Negro area before beginning research, both for project 
planning purposes as well as for comparisons with future data to see what effect 
outplanting may have on the ecosystem.   
Touring the Ca?o Negro NWR was informative and provided a critical visual 
impression of where and what research might be needed and appropriate.  Don Antonio, 
the boat tour guide, was obviously familiar not only with the larger fauna of the area but 
also the flora and all their many interactions.  When the project was described to him he 
agreed it was a good idea and even mentioned that he knew of only one location where a 
stand of the Acoelorraphe palm remained in the wetland.  Unfortunately it was not 
possible to view these as the water was too low to reach that area at that particular time of 
year.  Another tree was mentioned, a species of Inga, that Don Antonio said may be 
especially good for fisherman because this tree grows along river banks and drops its fruit 
in the water there.  This in turn attracts various fish that eat these fruit at the surface, 
making the fish easier to spot and catch.  During the boat tour it was clear that the banks 
along the Rio Frio had indeed become extremely eroded and were continuing to erode.  
Abundant wildlife was seen.  Caimans (Caiman crocodiles L.) were perhaps the most 
common large reptile seen.  There were a few basilisk or Jesus Christ lizards (Basiliscus 
basiliscus L.) but these were far outnumbered by the iguanas which could be seen at 
almost every turn, even swimming in the water.  The guide said he had seen caimans eat 
iguanas swimming across the open water.  Of course the bird population was extensive as 
58 
 
well.  Anhingas, or cormorants, (Anhinga anhinga L.) were by far the most frequently 
sighted bird on the tour: these are also known as snake birds because of the way they 
swim with their bodies under the water and their long necks protruding above the 
waterline.  Several green kingfishers (Chloroceryle americana Gmelin) were also seen.  
A few mantled howler monkeys (Alouatta palliata Gray) were seen, but they were very 
high up in the trees and napping during the heat of the day.  Some farm animals, cows 
and horses, were either in fields adjacent to the river or on drinking swales.  These swales 
or erosion ditches were almost completely devoid of plant life.  There are stream side 
management laws in Costa Rica to prevent this, but they are difficult to enforce and thus 
rarely followed (Ruiz, 2009).  Making the elimination of these swales economically 
advantageous would benefit the views and the environmental ecology of the area.  Plant 
life along the river banks was typical of the area with huge numbers of bromeliads 
growing in almost every tree.  Creeping vines occurred just as numerously, and large 
specimens of many tree species.  Several species of palm were seen along the banks as 
well, along with more than one species of Inga.  Once the wetland area was reached 
multiple species of water plants were seen.  No orchids or large flowering plants were 
seen along the bank, though this is not uncommon.  Perhaps most fascinating of all was 
the large number of epiphytic cacti seen in the trees.  One island in particular had several 
large Inga trees on which it was difficult to discern the trunks due to the masses of cacti 
hanging down from the branches.  It was not possible to stop and closely examine the 
cacti so there is no certainty as to what species were present but Hylocereus costariensis 
and Epiphyllum hookeri are the most likely candidates.  Identifying and mapping native 
plants along the river would be an excellent collaborative project for AU and UNA 
59 
 
graduate students.  Throughout the tour cacti were seen in multiple species of trees and at 
various heights in the canopy.  All species of these cacti are night blooming which could 
be used to promote night tours, a practice that is gaining popularity in Costa Rica.  If 
cacti were planted intentionally, stands could be monitored so as to know what nights 
blooms would be available for viewing during the flowering season. 
At Rancho las Tablitas, fresh tilapia from a neighbor?s aquaculture ponds is 
served in the restaurant.  The proprietor, Ms. Lopez, has a small botanic garden and even 
a labyrinth started behind the restaurant area.  Cabins are available for rent and spaces for 
camping are also available.  Most of the food served is raised on site and Bernadita Lopez 
is very involved with the community and its projects.  She currently works with a 
women?s group in the area that has been started in collaboration with the local Peace 
Corps volunteer.  Her reaction to the project and its goals was positive, and she did 
encourage contact with the women?s group.  This stop was a pleasant look into the 
everyday resources and activities of the area. 
Other Organizations 
During the visit to ITCR, specifics of the educational and degree process were 
presented.  The possibility of AU becoming a part of this venture though distance 
education was discussed and all agreed that the idea deserved further consideration and 
development.  It was interesting to learn Oscar Villegas? focus on small business 
development with a focus on technology application.  This would be an exceptionally 
good aspect of training for those developing the tourism industry in Ca?o Negro to 
receive.  
60 
 
While visiting in May 2010, the meeting at MAG was highly informative and 
provided many ideas for subsequent cooperation with this agency.  The results of the 
study recently concluded by MAG and presented to AU personnel was an extensive 
investigation on what pests are most likely to be found on the most common export crops 
of Costa Rica, with a particular focus on ornamental plants.  Apparently there is a major 
problem because import standards imposed by other countries are too onerous for small 
farmers.  Consequently export profits have fallen.  The MAG study was intended to act as 
a recommendation reference for small farmers on how to effectively manage pests based 
on export standards.  At the conclusion of this presentation further discussion ensued on 
the possibilities for collaboration between Auburn and MAG.  These included 
internships, joint research, and study abroad programs, among other things.  Assistance 
from MAG on future research trips by AU was also offered. 
During the visit with Orlando Vargas, Wright and Meriwether learned that for 
several years he has been actively trying to remove invasive plant species from the OTS 
station property while replacing these with various native species.  In the future he hopes 
to turn this project into an educational program to help the public learn about problems 
and concerns pertaining to invasive plants.  He has also been instrumental in developing 
the La Selva online database of native plants, La Flora Digital de La Selva (La Selva, 
2010), which was begun with the help of an NSF grant.  Recently he has indeed become 
involved with Pro Nativas and has begun preliminary germination and propagation 
experiment on several native Costa Rican plants.  This work will be combined with the 
work on removal of invasive plants to promote the use of native plant species in the 
landscape.  His propagation area was composed of four raised beds and several plants 
61 
 
planted directly in the ground.  The area displayed a Pro Nativas sign.  A cutting of H. 
stenopterus was growing amongst these plants.  Vargas gave AU personnel a cutting 
from an H. stenopterus plant growing on the OTS campus 
Chilimate 
While on the hiking tour at Selva Verde, it was discovered that Selva Verde 
participates in the PES program, and the forest is well protected.  It was also well worth 
taking the boat tour in Chilimate in order to observe the contrast between it and the one 
taken in October in Ca?o Negro.  Transportation from the hotel to the site of the boat tour 
launch was organized through Selva Verde Lodge, as was the boat tour itself.  Boats left 
from a pier located in Chilimate and traveled up the river for about an hour.  The style of 
craft was the same as that seen in Ca?o Negro.  The guide for the trip was knowledgeable 
and spoke English.  Almost immediately it was obvious that this area had not experienced 
the same environmental degradation as that of the Ca?o Negro area.  Obvious erosion 
was not prevalent, and only one cow wallow was seen.  Not nearly as many caimans or 
iguanas were seen, but basilisk lizards were more plentiful.  A blooming bromeliad was 
seen, and the guide said that it was native only to the Rio Frio area.  Several epiphytic 
cacti were seen, probably Epiphyllum hookeri, but once again it was not possible to 
examine any of them closely enough to make a positive identification.  Kingfishers were 
once again present as were anhingas, though not in the same high numbers as in Ca?o 
Negro.  In general, though the animal life was not as obvious, the vegetation was thicker 
and more over grown in this southern location than further north in Ca?o Negro.  
Differences in the presence of various animal species can be attributed to differences in 
ecosystem type: the one in the north being a wetland and home to more crocodilians and 
62 
 
water fowl and the one in the south being a river.  Environmental status could be a 
reflection of the differences in history of the two areas.  Chilimate has been a center for 
tourism since the industry began to flourish in the early seventies.  Thus, for years the 
income of many of the local residents has been based on travelers coming to enjoy the 
beauty of the local area.  Therefore, it was in their best interests to protect that 
environment, whereas further north, in Ca?o Negro, agriculture was the primary industry 
and only very recently has tourism begun to expand there.  Protecting the environment 
was secondary to raising crops and cattle. 
The UCR botanic garden was well maintained, diverse, and well organized with 
several distinct styles and areas laid out.  Perhaps the most spectacular feature of the 
garden was the orchid house, where hundreds of varieties of orchids at every conceivable 
life stage could be seen.  No particular effort had been made to inform visitors about or 
encourage use of native plants, though plants were organized according to ecoregions.  
None of the plants currently being investigated were present. 
Orosi was also well worth seeing.  For many years this area has focused on 
agriculture and in particular shade grown coffee.  During the time of European 
colonization, immigrants settled primarily in this area as it is one of the first hospitable 
regions reached when traveling to the central valley from the Caribbean coast.  It is a 
good example of how agriculture is an integral part of Costa Rican society and not 
relegated to rural areas.   
Plans are being made for a return research trip.  It will be important to visit UNA 
Sarapaqui and check on the progress of the experiments in the greenhouse there.  There 
will also be a plan to begin similar propagation experiments at INBio in San Jose as well 
63 
 
as to pay a visit to Ca?o Negro, hopefully in the company of Orlando Vargas, Victor 
Ruiz, and Nelson Zamora.  An attempt will also be made to return with cactus cuttings 
from the Ca?o Negro area to continue experiments at AU. 
Discussion 
In retrospect it would have been impossible to make appropriate decisions about 
program development without having taken the two described research trips.  Though 
large amounts of useful information can be gleaned from reading books and articles on 
ecotourism and the dynamics of research in Costa Rica, it would be extremely difficult to 
fully understand the intricacies of sociological interactions without experiencing them 
firsthand. 
Working relationships have been established at this point with both UNA 
Sarapaqui and INBio.  Neither of these organizations were primary candidates for 
cooperation at the beginning of the first trip.  Though there was communication with 
UNA San Jose before the trips, it wasn?t even known that UNA Sarapaqui existed, and 
now it has evolved into the most promising candidate for a long term research partner.  
The only experiments implemented are located on the UNA Sarapaqui campus.  Victor 
Ruiz, its regional director, is now a personal friend and very involved with the project.  
Without the personal visit to the actual area, it is likely that this connection would never 
have been made.  The process progressed to jointly signing a Memorandum of 
Agreement with INBio to fully formalize and stimulate the relationship.  It is possible 
that this relationship as well would never have occurred if research trips had not been 
taken.  During preliminary research, this non-profit was simply considered a valuable 
information source as opposed to a research partner.  At this point, no formal experiments 
64 
 
have begun at INBio but Nelson Zamora is in possession of A. wrightii seeds.  The intent 
is to begin experiments at INBio later this year. 
Several positive meetings with potential partners were conducted during both 
research trips that have not yet translated into working relationships.  ITCR and OTS are 
two examples.  MAG has the potential of forming an excellent working relationship.  
There was talk of developing reciprocal internship programs between AU and MAG 
sometime in the future.  Work with OTS and Orlando Vargas offers a promising 
relationship for the future and is another example of the value of international travel to 
meet people and assess research goals and potential commitment to a collaborative 
program.  This connection may also establish a relationship with ProNativas.  Though no 
formal meeting has occurred to date, there has been communication via email with 
several members of Pro Nativas as well as its founder, Willow Zuckowski.  It is clear that 
the goals of the Auburn project and Pro Nativas are similar and there is a strong 
possibility for this relationship to develop into a long-term commitment. 
Though the overall goals of the project remain the same (to facilitate community 
development and conservation by encouraging and teaching propagation and use of 
native plants) the methods by which this will be accomplished have shifted to some 
degree.  There is less intention to focus solely on the community level.  Community 
participation and development is still of paramount importance, but more general 
program development may need to occur if there is to be a chance of success.  If anything 
the goals for the project have become more long-term and expansive, creating a program 
with the potential to grow beyond the Ca?o Negro area and become an integral part of 
two or more universities.  So the original goals have expanded to include the creation of a 
65 
 
multi-level program with an interdisciplinary influence that draws on resources from 
multinational sources; these sources being two educational institutions, multiple 
governmental agencies, and at least one non-profit organization.  Ultimately this program 
will include aspects of both education and research involving student and faculty study 
abroad and possible internships programs with concurrent and coordinated research in 
multiple locations.  Information and international models developed from this program 
will then be applied in rural communities such as Ca?o Negro and other international 
regions to further the goals of worldwide conservation and community development. 
 
 
 
66 
 
Figure 1: Map of research trips taken in September 2009 and May 2010. 
 
 
 
 
 
67 
 
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70 
 
Chapter III 
Propagation of Hylocereus stenopterus and Acoelorraphe wrightii 
Key Words: Hylocereus, Acoelorraphe wrightii, propagation, scarification, palm, cacti 
Abstract: Effects of rooting hormone, cicatrization time and use of a mist tent on 
rooting, root and stem growth, and fresh weight relative growth rate (RGR) of 
Hylocereus stenopterus cuttings was studied.  Effects of rooting hormone on rooting, root 
and stem growth, and fresh weight relative growth rate (RGR) of both Hylocereus 
undatus and H. costariensis cuttings was also studied.  Effects of scarification and 
substrate on germination rate of A. wrightii seeds was also studied.  All experiments took 
place in a glass house in Auburn, AL.  All three Hylocereus species showed increased 
root growth and RGR when rooting hormone was applied.  H. stenopterus cuttings also 
showed higher RGR when enclosed in a mist tent.  A. wrightii seeds showed highest 
germination percentages when propagated in a 1:1 peat:perlite mix.  Increased 
germination of A. wrightii seeds occurred in all experiments when seeds were scarified 
with water or sulfuric acid.  Manual scarification had lower germination percentages than 
other forms of scarification or unscarified (control) seeds. 
Introduction 
 Conservation of biodiversity has become an issue of international concern (CBD, 
2010).  Ecosystems do not adhere to political boundaries and the solution to slowing the 
loss of biodiversity must in some part involve international cooperation.  Research 
suggests that plant propagation and subsequent outplanting can be used as 
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an effective conservation technique (Potvin et al., 2003).  It has also been observed that 
ecotourism can benefit local economies as well as the local environment (Campbell, 
2002).  Opportunities exist for researchers at Auburn University (AU) to collaborate with 
colleagues in Costa Rica to develop propagation protocols for plant species native to the 
Ca?o Negro National Wildlife Refuge.  The research objective of this program is to 
develop reliable and clear propagation techniques for two multi-use (edible, economically 
viable, aesthetic, in need of conservation) plants native to the Ca?o Negro NWR so that 
they can be produced, harvested, and transplanted by community members to stimulate 
the economy of Ca?o Negro village and help conserve the unique ecological community 
of the Ca?o Negro NWR. 
As part of ongoing cooperation between AU and Universidad Nacional de Costa 
Rica (UNA), and in the interest of facilitating both the expansion of conservation 
techniques as well as the overall conservation of Costa Rican biodiversity, propagation 
protocols for the following two species native to the Ca?o Negro area of northern Costa 
Rica were investigated: pitaya or Hylocereus stenopterus (F.A.C. Weber) Britton & Rose 
and Paurotis palm or Acoelorraphe wrightii Griseb. & H. Wendel. 
Hylocereus sp. 
H. stenopterus is a member of the Cactaceae and was first described by Britton & 
Rose (1963).  It is a tropical hemiepiphyte (an epiphyte that can secondarily develop 
roots in the ground).  H. stenopterus consists of three-ribbed stems that are triangular, 
succulent, and feature spines 2-4 mm wide and 5-10 mm long (Nerd et al., 2002).  
Flowers are perfect and 20-30 cm wide.  Most flowers in the genus Hylocereus are 
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creamy white: two exceptions are H. stenopterus and H. extensus, which have red or pink 
flowers.  All bloom nocturnally.    
H. stenopterus is found only in Costa Rica.  Other members of the genus 
Hylocereus can be found throughout Latin America, where they are thought to have 
originated, as well as around the world in tropical and subtropical regions (Britton & 
Rose, 1963; LeBellec et al., 2006). There are currently 16 recognized species in the genus 
Hylocereus (Britton & Rose, 1963). In Latin America several plants in this genus, and 
their fruit, are collectively known as pithaya or pitahaya (other spellings can be found as 
well: pitahaja, pitajuia, or pitalla). In Vietnam it is called thang loy or dragon fruit 
(Nobel, 2002).  It continues to be recognized as a promising new crop (Janick, 1999).  
Results of this research are intended to facilitate the advancement of its production, not 
only as a cash crop for Ca?o Negro, Costa Rica, but for agriculturalists and horticulturists 
world-wide.  As almost all reports of commercial propagation within this genus have 
revolved around one species, Hylocereus undatus.  The remainder of this review will 
focus on research done on this species. 
H. undatus is grown commercially in several countries in nurseries for ornamental 
retail sale as well as horticulturally for fruit production (Choi et al., 2007; Esquival et al., 
2007; Merten, 2003; Yussof et al., 2008).  Several papers explore methods for 
commercial propagation of this species, though the practice only began around 15 years 
ago (Andrade et al., 2006; Cavalcante & Martins, 2008; De Andrade et al., 2007; De 
Andrade et al., 2008; Michel-Rosales & Farias-Larios, 2007; Mohamed-Yasseen, 2002).  
Both asexual and sexual propagation have been attempted.  Though propagation by seeds 
is possible, germination is often inconsistent (LeBellec, 2003).  Micropropagation has 
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also met with success, but the cost of production is currently not profitable (Garcia et al., 
2009; Mohamed-Yasseen, 2002).  LeBellec (2003) concluded that asexual propagation by 
cuttings represents the most effective and promising form of propagation, at least for 
horticulture purposes.   
Up to 90% rooting success can be achieved using cuttings (LeBellec, 2003).  
Cavalcante & Martins (2008) suggested that 25 to 50 cm cuttings be taken from juvenile 
growth with the basal end cut being made just above a joint in the fleshy green stem 
extending to the terminal end joint.  Bastos et al. (2006) concluded that application of a 
rooting hormone aided in rooting success, but Thomson (2002) reported no effect.  
Thomson (2002) also suggested allowing cuttings to cure or cicatrize for up to 5 days 
before planting while Cavalcante & Martins (2008) planted cuttings immediately.   
These studies indicate that, for H. undatus, cicatrization time and application of 
rooting hormone may influence propagation success.  Further research is necessary to 
make more specific recommendations for asexual propagation of not only H. undatus but 
H. stenopterus and H. costariensis as well.  Current research investigates stem cutting 
propagation for all three species. 
Accoelorraphe wrightii 
Aracaceae is an isolated and distinct group within the plant kingdom and is 
composed entirely of woody monocotyledonous plants collectively and commonly 
known as palms (Jones, 1995).  Palms are found around the world in tropical ecosystems 
and most have a fairly recognizable canopy with a prominent trunk and a characteristic 
crown of leaves.  All are perennial, and the family contains both monoecious and 
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dioecious species.  Reproduction by palms in nature is almost exclusively by seed 
(Meerow, 2002). 
Within this family, A. wrightii, also known as Paurotis palm or Everglades palm, 
is a member of the Corypheae tribe and the Coryphoideae subfamily (Meerow, 2002).  It 
is found in Florida, the West Indies, and coastal or wetland areas of Central America 
(Farruggia, 2008; Jones, 1995; Meerow, 2002; Riffle & Craft, 2003).  The genus 
Acoelorraphe is monotypic with A. wrightii the only species (Jones, 1995). It is a 
relatively tolerant species to habitat conditions, being one of the few palms that can grow 
in saturated soils (Meerow, 2002).  It can have problems with manganese deficiencies but 
responds well to fertilization and has no known pest problems (Meerow, 2002). 
Though reproductive maturity is reached in four to five years by some species of 
palms, others can take up to forty or fifty years (Meerow, 2004).  Unlike A. wrightii, seed 
germination in most palms is notoriously difficult, with some seeds taking years to 
germinate (Odetola, 1987).  It has been estimated that up to 25% of all palms take over 
100 days to germinate and often achieve less than a 20% germination rate (Tomlinson, 
1990).  Germination of A. wrightii usually occurs within 2 to 4 months (Carpenter, 1988; 
Jones, 1995; Meerow, 2002).  There is no clear evidence why this difficulty in 
germination occurs, as very few seed germination studies have been conducted for palms 
(Meerow, 2004).  Efforts to produce palms commercially persist, however, as certain 
species are significant in the ornamental and edible markets (Jones, 1995).   
Propagation of palms is almost entirely done by seed, though vegetative 
propagation is possible by air layering, division, or even more recently, tissue culture 
(Jones, 1995).  Vegetative propagation is almost exclusively used by enthusiasts and 
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collectors while tissue culture is used by researchers, collectors, and large scale 
commercial operations.  Neither tissue culture nor vegetative propagation is feasible for 
the average nursery or farm due to cost and equipment needs. 
Seed germination among palms occurs in several forms but can generally be 
divided into two initial categories; remote or cryptocotylar germination and adjacent or 
hypogeal germination (Meerow, 2004; Orozco-Segovia et al., 2003).  Seed germination 
in A. wrightii occurs by way of the remote germination pathway (Meerow, 2002).  With 
remote germination the first structure to emerge from the seed is called the cotyledonary 
petiole; this is commonly mistaken for the first seedling root (radicle).  However, the first 
seedling root will actually emerge from this structure after it has grown down into the 
soil.  At the other end, the seedling shoot or plumule will emerge.  The cotyledon remains 
inside as the absorptive structure and is known as the haustorium.  Adjacent germination 
involves only a small portion of the cotyledon emerging from the seed; this is known as 
the button (Meerow, 2004).  Then, the radicle and plumule emerge from the button.  The 
radicle in this type of germination is very short lived and is quickly replaced by 
adventitious roots that form around the stem base.  Like remote germination, the part of 
the cotyledon known as the haustorium remains inside the seed and absorbs nutrients 
from the endosperm.  These different germination strategies have been associated with 
establishment in different ecosystems (Orozco-Sergovia et al., 2003).  Information on 
seed germination for palms tends to be inconsistent as much of it comes from hobbyists 
observations and not controlled research.  No clear propagation protocols can be 
described for more than a few commercially important species. 
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Seed dormancy has been identified as an important aspect of palm seed 
germination and may be a significant part of understanding the long and sporadic 
germination of many palm species (Orozco-Segovia et al., 2003).  Several methods are 
used in plant propagation to break seed dormancies, the most common among palms 
being seed scarification.  Scarification for palms commonly utilizes extended soaks in 
water for 12 h up to several days (Broschat & Donselman, 1986; Kheong, 1992; Moussa 
et al., 1998; Odetola, 1987).  In cases of extremely hard or thick seed coats, soaking in 
concentrated sulfuric acid can be effective for shortening germination time (Merlo et al., 
1993).  Physical scarification accomplished by nicking the seed coat has also been used 
to reduce germination time and increase germination rate (Holmquist & Popenoe, 1967; 
Nagao et al., 1980; Odetola, 1987).   
Substrate for palms requires well-drained yet moist conditions.  Most palms 
require uniform moisture during germination and subsequent seedling development and 
do not respond well to alternating periods of dry and wet.  It is important that the 
substrate stay damp but not be saturated since palm seeds can be susceptible to rot 
(Koebernik, 1971).  A common substrate in the nursery industry is a 1:1 ratio of peat 
moss:perlite (Meerow, 2004).  Palms have also been germinated in only perlite or peat 
moss, but a 1:1 combination appears to be the most effective (Carpenter, 1988; Jones, 
1995; Koebernik, 1971).  Fertilization of palms is not important until late in seedling 
development.  For the first few months after germination the endosperm acts as the only 
source of nutrition for the developing seedling and further nutrients can damage the 
seedling (Meerow, 2004).   
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Only one study (Carpenter, 1988) directly investigated propagation of A. wrightii, 
and concluded that A. wrightii germinated best at temperatures above 35o C with an 
optimum range of 33o to 39o C.  Current research will investigate the effect of 
scarification method and sowing substrate on the germination rate and final germination 
percentage of Acoelorraphe wrightii seeds. 
Materials and Methods 
Hylocereus sp. 
All experiments with Hylocereus sp. were conducted in a glass house at the 
Paterson Greenhouse Complex on the Auburn University campus in Auburn, AL (32.6oN 
latitude).  Ground beds 1 m wide, 4.6 m long and 15.2 cm tall were used for propagation.  
Beds were lined with black fabric groundcloth and filled with 100% sand. 
Hylocereus stenopterus 
Cuttings of H. stenopterus were obtained from California in February 2010 
(Matt?s Landscape, San Juan Capistrano, CA).  Thirty-three cuttings, each measuring an 
average of 18 cm long, were harvested on 6 February, bundled in newspaper, mailed via 
USPS, and arrived in Auburn, AL on 16 February.  Upon receipt in Auburn, cuttings 
were sorted into groups based on stem diameter, recut into smaller pieces, each 
measuring approximately 3.8 cm in length, and fresh weight (FW) was recorded for each 
cutting.  Cuttings from each stem diameter were distributed evenly among treatments.  
Care was taken to maintain proper polarity of all cuttings.  Half of the cuttings were then 
stuck immediately, while the other half were allowed to remain uncovered on a bench in 
the greenhouse for 72 h (cicatrization).  Prior to sticking, the basal end of half of the 
cuttings were treated with 0.1% IBA (Hormodin?, OHP, Inc., Mainland, PA) while the 
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other half received no hormone.  Once stuck, half of the cuttings were covered with a 
plastic tent.  Mist tents were 91 cm tall, 127 cm wide, 178 cm long, and constructed of 1? 
Schedule 40 PVC? pipe and Husky? 3.5 mm clear plastic sheeting.  Mist tents were 
placed directly on top of the propagation bed.  Thus, treatments were in a 2 hormone x 2 
cicatrization x 2 tent factorial design for a total of 8 treatments with 20 replications per 
treatment for a total of 160 cuttings.  Cuttings were stuck such that the basal 1 cm was 
below the surface of the sand.  Experimental design was a split plot design with the 
presence or absence of plastic tent forming the main plot (total four main plots).  Within 
each main plot, treatments (hormone x cicatrization) were arranged randomly. 
Mist was applied initially for 20 s once an hour then adjusted as needed 
throughout the experiment.  For the first month cuttings were misted for 20 s every hour.  
This appeared to be too much moisture as rot began to occur on the cut surface of each 
cutting.  On 11 March mist was discontinued, and the substrate was allowed to dry for 4 
days.  Beginning on 15 March, cuttings then received mist once a day for 10 min a day 
between 8:00 a.m. and 10:00 a.m.  Beginning 14 June cuttings were fertilized at a rate of 
100 ppm N 20-20-20 Jack?s? Professional (J.R. Peters, Inc., Allentown, PA) water 
soluble fertilizer, applied to the bed once a week by hand.  This was increased to twice a 
week beginning 12 July.   
On 3 May, mortality, new shoot length, length of longest root, and total plant 
fresh weight were recorded.  To measure fresh weight, cuttings were carefully dug out of 
the sand by hand and the roots rinsed with water to remove sand before weighing.  To 
account for initial cutting size, relative growth rate (RGR) was calculated using the 
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formula (Final FW ? Initial FW)/Initial FW.  On 17 June shoot lengths were measured 
again.   
On 30 June, cuttings with new shoots longer than 5 cm were either removed from 
the bed and potted or the new shoot was removed from the original cutting, treated with 
hormone and stuck in the same sand bed adjacent to the original cutting.  Cuttings that 
were potted were planted either with the original cutting entirely below the substrate 
surface or with only the basal 1 cm of the cuttings below the surface (similar to position 
in the propagation bed).  Equal numbers of cuttings (37) were used for each ?replanting? 
technique.  On 22 July the original cuttings still in the bed were evaluated, and cuttings 
with new shoots over 5 cm were again either potted using the two potting methods 
described above or the new shoot was removed and stuck using the same procedures as 
on 30 June.  When potted, cuttings were planted in 3.8 L black plastic pots in 1:1:2 
perlite:peat moss:sand and placed on a raised bench in the same greenhouse as the sand 
ground bed.  Potted cuttings were fertigated by hand twice a week at the same rate and 
analysis of fertilizer previously given.  For all repotted and restuck cuttings, shoot lengths 
were measured on 24 Aug. and 29 Sept.  Relative growth rate of the shoots for cacti that 
were potted was calculated using the following formula ((Initial length-Final 
length)/Initial length)/Number of days between measurements.  Cuttings that were cut 
and stuck in the same bed were only observed for survival. 
H. undatus and H. costariensis 
Sixty cuttings of H. undatus and 15 cuttings of H. costariensis were harvested 22 
May 2010 in CA and shipped to Auburn, AL.  Experiment location, cutting source, and 
shipping method were the same as with H. stenopterus.  H. undatus cuttings were re-cut 
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so that each new cutting would have at least 3 nodes on at least 2 ribs.  Once recut, 
cuttings were sorted into three size classes based on fresh weight, and each size class was 
distributed evenly among each treatment.  H. costariensis cuttings were all of a similar 
size and there was no need to sort them based upon size.  Care was again taken to 
maintain proper polarity with all cuttings. 
For both species three hormone treatments were used: untreated (no hormone 
applied, control), 0.1% IBA or 0.8% IBA.  H. undatus had 30 replications in each 
treatment for a total of 90 cuttings.  H. costariensis had 9 cuttings per treatment for a total 
of 27 cuttings.  Experiments for both species were completely randomized.  All cuttings 
received overhead mist for 10 min a day between 8:00 a.m. and 10:00 a.m.  Beginning 14 
June all cuttings were fertigated once a week at the same rate and with the same fertilizer 
solution as that used previously.  Beginning 12 July cuttings were fertigated twice a week 
at the same rate. 
 Initial fresh weight and length of all cuttings were measured for both species.  
Mortality, length of new shoots, longest root length, and total plant fresh weight (as 
described above) were recorded for both species on 13 July.  RGR was calculated as 
described above.  Lengths of new shoots were measured again for H. undatus on 16 Sept.  
Lengths of new shoots, longest root length, and total fresh weight (as described above) of 
H. costariensis cuttings were measured again on 24 Aug. 
All growth and rooting data for all cacti were analyzed using PROC GLM in SAS 
9.1 (SAS Inc., Cary, NC).  Means were separated using PDIFF at ? = 0.05 (5%).  Results 
are not reported for any parameter showing no significant differences. 
Acoelorraphe wrightii 
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Experiments with this species began in February 2010.  Seeds were obtained from 
the Banana Tree (Easton, PA).  All experiments with A. wrightii were conducted in a 
glass house in Auburn, AL on raised metal benches at the Paterson Greenhouse Complex 
on the Auburn University campus (32.6oN latitude).  All containers were hand watered on 
a daily basis.  Seeds were stored in the cooler at the Paterson Greenhouse Complex at 
8oC.  For experiments 1 and 2 seeds were planted in 20 cm x 13 cm squat mum pots 
(Nursery Supplies, Inc. Kissimmee, FL).  In experiment 3 seeds were planted in 4? 
individual pots and placed in plastic mesh flats.  All seeds were planted at a depth of 1 
cm (1.5 times mean seed diameter).  For all experiments, experimental design was a 
completely randomized design. 
Experiment One 
To investigate the effect of scarification on germination of A. wrightii, seeds 
received one of three scarification treatments: non-scarified (control, unsoaked), abrasion 
scarified, and hot water scarified.  There were 10 pots per treatment with 5 seeds per pot.  
Abrasion scarification was accomplished by hand using a rat tail file to rasp out a 1-2 mm 
deep groove in a single location on each seed coat.  Seeds that were hot water scarified 
were placed in water initially heated to 65.5oC and left to soak for 24 h.  All seeds were 
planted on 23 February 2010 in a 1:1 peat:perlite mix.  Data collection included whether 
seeds germinated and when seedlings reached a height of 7 cm.   
Experiment Two 
To investigate the effect of scarification on germination of A. wrightii, seeds 
received one of eight scarification treatments: unscarified (control in which seeds were 
soaked in room temperature water for 30 min), immersion in 100% H2SO4 for 10, 20, or 
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30 min,  or immersion in 80%, 60%, 40%, or 20% H2SO4 for 30 min.  There were 10 pots 
per treatment with 3 seeds per pot.  All seeds were planted on 5 April 2010 in a 1:1 
peat:perlite mix.  Data collected included date of germination (defined as when new 
growth was visible above the soil surface) and date seedlings reached a height of 5 cm. 
Experiment Three 
To investigate the effect of scarification as well as substrate on germination of A. 
wrightii, seeds received one of twelve treatments.  There were four substrate treatments 
and three scarification treatments in a factorial design of 12 total treatments.  
Scarification treatments included unscarified (control), hot water scarification 
(methodology being the same as that for experiment one), and acid scarification of 30 
min immersion in 100% H2SO4.  Substrate treatments were a 1:1 peat:perlite mix, 100% 
perlite, 100% aged whole pine tree substrate (AP) (aged 9 months outdoors in 63 ft3 bulk 
bags), and 100% fresh whole pine tree substrate (FP) (harvested and processed the same 
week seeds planted).  There were 10 pots per treatment with 5 seeds per pot.  All seeds 
were planted on 11 June 2010.  Data collected were the same as that for experiment two. 
All data were analyzed using PROC GLM in SAS 9.1 (SAS Inc., Cary, NC).  
Means were separated using PDIFF at ? = 0.05 (5%).  Results are not reported for any 
parameter showing no significant differences. 
Results 
Hylocereus stenopterus 
For data collected 3 May, there was a significant interaction between rooting 
hormone and mist tent for root length.  Cacti that did not receive rooting hormone and 
were also not under a tent had less root length than other cuttings (2.5 cm (SE=0.69, 
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p=0.03) vs. 6 cm (SE=0.69, p<0.0001) average in all other treatments).  On 17 June a 
significant difference in RGR of shoots occurred between cacti cuttings that were covered 
by a tent (5.5 cm (SE=1.06, p<0.0001)) as compared to those that were not (2.8 cm 
(SE=1.06, p=0.007)).   
Cacti repotted on 30 June showed no significant differences among replanting 
techniques in average shoot length on 24 Aug. or 29 Sept.  Cacti repotted on 22 July 
showed no differences in RGR of shoots when data were taken on 24 Aug.  However, on 
29 Sept. RGR for shoots of cacti with the original cutting planted above the substrate 
surface were significantly higher than that for those planted below the substrate surface 
(45 cm and 31 cm, respectively (SE=3.1, p<0.0001)). 
H. undatus and H. costariensis 
On 13 July, for H. undatus, root length and date of new shoot development were 
different among treatments.  Root length was longer for untreated cuttings (16 cm 
(SE=1.36, p<0.0001)) than for cuttings receiving either treatment with 0.1% IBA or 0.8% 
IBA (12 cm (SE=1.08, p<0.0001) and 11.8 cm (SE=0.98, p<0.0001), respectively).  
Untreated cuttings also developed shoots in a significantly shorter period of time (25 days 
(SE=2.09, p<0.0001)) as compared to either treatment with 0.1% IBA or 0.8% IBA (31 
(SE=1.66, p<0.001) and 35 (SE=1.51, p<0.0001) days respectively). There were no 
significant differences among treatments for any of the measurements taken on 13 July 
for cuttings of H. costariensis. 
 On 16 Sept., RGR for new shoots of H. undatus cuttings treated with 0.8% IBA 
was higher (14.7 cm (SE=0.53, p<0.0001)) than for cuttings treated with 0.1% IBA (8.8 
cm (SE=0.70, p<0.0001)) and untreated cuttings (10.2 cm (SE=0.87, p<0.0001)).  There 
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was also a difference in the number of days until shoot development for cuttings treated 
with 0.1% IBA (46 days (SE=5.6, p<0.0001)) and 0.8% IBA (57 days (SE=4.2, 
p<0.0001)), but there was no difference between untreated cuttings (52 days (SE=6.9, 
p<0.0001)) and either of the two other treatments. 
 On 24 Aug., there was a difference in RGR of new shoots between untreated H. 
costariensis cuttings and those treated with 0.8% IBA (19 g (SE=2.3, p<0.0001) and 26 g 
(SE=2.3, p<0.0001) respectively). 
Acoelorraphe wrightii 
Experiment 1 
 Germination percentages for the control and water scarified seeds (40% 
(SE=0.059, p<0.0001) and 50% (SE=0.059, p<0.0001), respectively) were not 
significantly different from each other, while that for the abrasion scarified seeds (2% 
(SE=0.059, p=0.7)) was significantly lower than both treatments.  Of the seeds that 
germinated there were no significant differences among treatments as to the speed at 
which seeds developed to a height of 7 cm. 
Experiment 2   
 Seeds scarified in 100% sulfuric acid for 30 min had a higher germination 
percentage (80% (SE=0.05, p<0.0001)) than seeds in any other treatment.  Seeds 
scarified in 100% acid for 10 and 20 min had higher germination percentage (56% 
(SE=0.05, p<0.0001) and 50% (SE=0.05, p<0.0001), respectively) than seeds in all other 
treatments.  In all other treatments (seeds scarified for 30 m in 80%, 60%, 40%, and 20% 
acid and control seeds) germination percentages were not significantly different from 
each other (0%, 0%, 0%, 3%, and 0% respectively). 
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 Of the seeds that germinated, those scarified in 100% acid for 30 min germinated 
more quickly (65 days (SE=1.42, p<0.0001)) than seeds scarified in 100% acid for 20 
min (67 days (SE=1.95, p<0.0001)) and seeds scarified in 100% acid for 10 min (78 days 
(SE=1.75, p<0.0001)).  These treatment differences were the same for number of days 
after sowing until seedlings reached the height of 5 cm. 
Experiment 3 
 In experiment 3, there was an interaction between scarification and substrate 
treatments for germination percentage.  Germination percentage for seeds scarified in 
acid and planted in 1:1 peat:perlite was significantly higher (64% (SE=0.04, p<0.0001)) 
than all other substrate/scarification combinations (Table 1). 
 Of the seeds that germinated, those soaked in 100% acid and planted in 1:1 
peat:perlite had fewer number of days between sowing and germination (43 (SE=2.3, 
p<0.0001)) than all other treatments (Table 2).  There were no differences between 
treatments pertaining to number of days following germination until seedlings reached 5 
cm.   
Discussion 
Hylocereus sps. 
 Results for all three species of Hylocereus suggest that this genus has the potential 
to be an easily propagated and maintained crop for ornamental or edible field crop 
purposes.  High survival was observed for all three species regardless of treatment, 
cuttings rooted quickly and readily, growth was vigorous once cuttings were established, 
and plants were easily transplanted with only minimal mortality.  Like LeBellec (2003), 
high overall survival rates with cuttings were documented for all species regardless of 
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treatment (H. stenopterus: 85%, H. undatus: 77%, H. costariensis: 100%).  Within 
treatment, H. undatus had 97% survival rate for cuttings treated with 0.8% IBA.  One 
hundred percent survival was also seen for H. stenopterus cuttings that were covered by a 
mist tent but neither received rooting hormone nor were allowed to cicatrize.  In practice 
however, no treatment produced dramatic enough results to indicate a particular 
advantage when propagating this plant.  Both LeBellec (2003) and Cavalcante and 
Martins (2008) suggest using cuttings that are longer than 25 cm.  However, cuttings of 
H. stenopterus used for this study only average 3.8 cm long and were still successful.  
Rooting hormone did have an effect on the rooting and subsequent growth of 
cuttings from all the Hylocereus species investigated.  On 3 May, H. stenopterus cuttings 
that received rooting hormone had a significantly higher RGR and longer roots than other 
cuttings.  On 13 July H. undatus cuttings that were treated with 0.8% IBA had longer 
shoots than other treatments and cuttings treated with 0.1% IBA developed shoots more 
quickly than other treatments.  On 24 Aug. H. costariensis cuttings treated with 0.8% 
IBA had a higher RGR and longer shoots than all other treatments.  This is consistent 
with research by Bastos (2006), which suggested that the application of rooting hormone 
would help with survival of H. undatus cuttings. 
 Cicatrization did not affect survival subsequent growth of H. stenopterus cuttings.  
However, both Thomson (2002) and Cavalcante and Martins (2008) found it beneficial to 
allow cicatrization time.  During this investigation, on 3 May, cuttings that had cicatrized 
had a higher RGR than other treatments, but this was no longer the case when cuttings 
were measured again on 17 June. 
Acoelorraphe wrightii 
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 Research on this palm suggests that it would be a good candidate for commercial 
production within the ornamental market and for conservation oriented outplanting due to 
its relatively short germination time, relatively high germination percentages, robustness 
of seedlings, and ease of transplant.  Similar to results reported by Carpenter (1988), this 
research indicates that germination time for A. wrightii seeds, particularly when subjected 
to scarification, can be relatively short compared to other palm species (Meerow, 2004).  
Seeds scarified in 100% acid for 30 min in both Experiments 2 and 3 resulted in shorter 
germination times than other treatments.  Seedlings were robust after germination; no 
seedling mortality was observed in any experimental trials.  Speed of growth for 
seedlings was remarkably uniform as well.  Once seeds germinated, seedlings 
consistently took 9 to 12 days to reach a uniform height. 
Scarification of seeds affected not only germination time but also germination 
success.  In Experiment 1, seeds scarified in water had a higher germination percentage 
(50%) than either control seeds (40%) or seeds that were manually scarified (2%).  It is 
possible that the method used to abrasion scarify seeds damaged embryos.  The exact 
location of the embryo inside A. wrightii seeds is unclear so it is possible that 
scarification actually damaged the embryo.  Nagao et al. (1980) suggested a similar 
explanation for low germination percentages in palm seeds that were scarified manually 
with a file.  In all experiments some level of scarification, either by water or acid, 
increased percent germination.  This is consistent with findings from Odetola (1987) who 
reported that scarification improved germination in several other palm species.  In this, 
study the highest germination percentages were recorded when seeds were soaked in 
100% H2SO4 for 30 min. 
88 
 
 Choice of substrate affected rate of germination in Experiment 2.  Consistent with 
recommendations by Meerow (2004), a 1:1 peat:perlite substrate produced highest 
germination rates.  Koebernik (1971) suggests it is possible to grow palm seeds more 
successfully in 100% perlite.  This was not the final conclusion of this research, where 
seeds grown in 100% perlite had some of the lowest germination percentages over all. 
Conclusions 
All species of Hylocereus investigated in this research grow well in 100% sand.  
To speed up rooting, rooting hormone can be applied but is not necessary to assure final 
rooting success.  On latitudes even with or north of AU, it is necessary to grow cacti 
inside a structure that provides shelter from freezing temperatures.  However, the use of a 
mist tent is not crucial. 
A. wrightii seeds are easily stored at cool temperatures and respond with increased 
germination rates and percentages to scarification with water or acid.  The lack of 
seedling mortality and ease of transplant make this palm a good candidate for nursery 
production.  It is important to make sure these palms receive adequate water and are 
fertilized once seeds have finished germinating. 
Both these species appear to be good candidates for both commercial production 
as well as conservation oriented propagation and outplanting in Costa Rica.  All 
experiments indicate that both species are easy to work with, vigorous, attractive, and 
easy to propagate.  Collaborative entities in Costa Rica are also motivated to work with 
these species and remain cooperative toward ongoing research goals.
89 
 
Table 1: Effect of scarification treatment and germination substrate on germination 
percentage of A. wrightii seeds grown in a greenhouse in Auburn, AL. 
             Mean Germination Percentages (SE=0.04) 
Scarification 
Treatmentsz  Substrates
y   
 PP AP FP P 
 
Acid Scarification 
(100% H2SO4 for 
30 min) 
 
64Aa 16Ba 22Ba 2C 
Water Scarification 
(24 hr soak in water 
initially heated to 
65.5oC) 
22Ab 0Bb 13Bb 2B 
Control 
(Unscarified) 16Ab
 6Bb 2Cb 0C 
zTreatments included acid scarification (A) (soak in 100% sulfuric acid for 30 min), 
water scarification (W) (24 hr soak in water heated to 65.5oC), and unscarified (C) 
(control). 
ySubstrates used included 1:1 peat:perlite (PP), aged pine tree substrate (AP), fresh pine 
tree substrate (FP), and 100% perlite (P). 
Means separation using PDIFF at ? = 0.05 indicated by uppercase letters in rows and by 
lowercase letters in columns. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
90 
 
 
Table 2: Effect of scarification treatment and germination substrate on number of days 
until germination of A. wrightii seeds grown in a greenhouse in Auburn, AL. 
Mean Days Until Germination 
Scarification Treatmentsz  Substratesy  
 PP AP FP 
 
Acid Scarification (100% 
H2SO4 for 30 min) 
 
43Abx 
(SE=2.3) 
73B 
(SE=5.3) 
67B 
(SE=4.5) 
Water Scarification (24 hr 
soak in water initially heated 
to 65.5oC) 
 
94Aa 
(SE=4.5) --
w 66B 
(SE=5.4) 
Control (Unscarified) 85a (SE=5.2) -- -- 
zTreatments included acid scarification (A) (soak in 100% sulfuric acid for 30 min), 
water scarification (W) (24 hr soak in water heated to 65.5oC), and unscarified (C) 
(control). 
ySubstrates used included 1:1 peat:perlite (PP), aged pine tree substrate (AP), fresh pine 
tree substrate (FP), and 100% perlite (P). 
wData not available due to insufficient germination. 
xMeans separation using PDIFF at ? = 0.05 indicated by uppercase letters in rows and by 
lowercase letters in column
91 
 
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undatus) in an agrosystem of Yucatan, Mexico. Agrociencia 40:687-697 
Bastos, D.C., R. Pio, J.A. Scarpare Filho, M.N. Libardi, L.F.P. Almeida, T.P.D. Galuchi, 
and S.T. Bakker. 2006. Propagacao da pitaya ?vermelha? por estaquia.  Ciencia Agroteca 
30:1106-1109 
Borschat, T.K. and H. Donselman. 1986. Factors affecting storage and germination of 
Chrysalidocarpus lutescens seeds. Journal of the American Society for Horticulture 
111(6):872-877 
Britton, N.L. and J.N. Rose. 1963. Descriptions and illustrations of plants of the cactus 
family, Vol I and II. Dover Publications Inc. New York, NY 
Campbell, L.M. 2002. Conservation narratives in Costa Rica: conflict and co-existence. 
Development and Change 33:29-56 
Carpenter, W.J. 1988. Temperature affects seed germination of four Florida palm species. 
HortScience 23(2):336-337 
Cavalcante, I.H.L. and A.B.G. Martins. 2008. Effect of juvenility on cutting propagation 
of red pitaya. Fruits 63:277-283 
CBD, 2010. Convention on Biodiversity Diversity. Retrieved Sept. 2010, from CBD 
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Choi, H.W., I.M. Chung, M.H. Sin, Y.S. Kim, J.B. Sim, J.W. Kim, K.D. Kim, and S.C. 
Chun. 2007. The effect of spent mushroom sawdust compost mixes, calcium cyanamide 
and solarization on basal stem rot of the cactus Hylocereus trigonus caused by Fusarium 
oxysporum. Crop Protection 26:162-168 
De Andrade, R.A., A.B.G. Martins and M.T.H. Silva. 2007. Influence of the material 
source and the cicatrize time in vegetative propagation of red dragon fruit (Hylocereus 
undatus Haw). Revista Brasileira De Fruticultura 29:183-186 
De Andrade, R.A., A.B.G. Martins and M.T.H. Silva. 2008. Development of seedlings of 
red pitaya (Hylocereus undatus Haw) in different substrate volumes. Acta Scientiarum-
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Esquivel, P., F.C. Stintzing and R. Carle. 2007. Comparison of morphological and 
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Farruggia, F.T. 2008. A floristic description of a neotropical coastal savanna in Belize. 
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Holmquist, J.D. and J. Popenoe. 1967. The effect of scarification on the germination of 
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Janick, J. (ed). 1999. Perspectives on new crops and new uses. ASHS Press, Alexandria, 
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Jones, D.L. 1995. Palms throughout the world. Smithsonian Institution Press, Washington 
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94 
 
Appendix 
Nymphaea ampla 
Introduction 
 Preliminary research was begun on the propagation of the water lily Nymphaea 
ampla.  Trials were undertaken to investigate not only germination of N. ampla seeds but 
also subsequent growth and fertilization of seedlings.  Due to insufficient results no 
statistical analysis was performed and only a literature review and cursory results will be 
presented here. 
Literature Review 
N. ampla is a member of the Nymphaeaceae in the subgenus Brachyceras 
(Wunderlin, 1980).  This family contains seven genera and about 65 species, 12 of which 
are members of the subgenus Brachyceras (Muhlberg, 1981).  All are aquatic plants with 
rhizomes or tubers and leaves that are arranged in basal rosettes (Muhlberg, 1981).  Most 
grow in sheltered areas of standing or slowly flowing water and can even withstand short 
periods of drought (Speichert & Speichert, 2004).  Within the genus Nymphaea, an 
important distinction is made between tropical water lilies and hardy water lilies which 
can grow in cooler climates.  Hardy water lilies flower only during the daytime while 
tropical water lilies have both night and day blooming varieties (Slocum & Robinson, 
1996).   
N. ampla is a tropical water lily that flowers only during the day (Landon, 2009).  
Some tropical species also display viviparous germination (when seeds germinate while 
95 
 
still attached to the parent plant), but Slocum & Robinson (1996) assert that N. 
ampla is not one of them.  For this species all flowers are white and have a pronounced 
fragrance.  New leaves grow out as a bronzy red but quickly develop a deep green color.  
Leaves usually have dark splotches and prominent veins.  Leaf margins are dentate with a 
wavy perimeter.  This species is native to several countries in Central and South America 
as well as areas of the southern United States (Conard, 1905; Wunderlin, 1980). This 
plant has been used for centuries as a pharmaceutical and some of this use continues 
today (Bertol, 2004).  Aporphine is an active compound which can be found in the bulbs 
and roots of at least three species of Nymphaea including N. ampla (Bertol, 2004).  In the 
human body, aporphine is hydroxylated into the active compound apomorphine.  
Traditionally this has been used as a hallucinogen in rituals and religious ceremonies, not 
only in ancient times but regularly during the 1960s and 1970s. Today, research is 
ongoing as to its effectiveness to treat erectile dysfunction (Bertol, 2004). 
While it is possible to propagate N. ampla from seed or rhizomes, seed 
propagation is rarely used in industry (Purcell, 1995).  Reasons for this are not 
necessarily related to difficulties with seed propagation, but rather because named 
cultivars of water lilies are almost all hybrids and must be propagated from tubers or 
rhizomes in order to produce genetically identical individuals (Landon, 2009).  Some 
species have seeds which are extremely sensitive to desiccation and can only be 
germinated if planting occurs within a few weeks of harvesting (Conard, 1905).  Certain 
species in the subfamily Brachycerus are not among these and can withstand extended 
periods of drought (Conard, 1905).  Seed propagation on a commercial scale is restricted 
to the species Nymphaea pygmaea Salisb. and Nymphaea odorata Aiton (Case, 1991). 
96 
 
In general, fully mature seeds of tropical water lilies are hard and black and 
surrounded by a jelly-like substance which supposedly helps with flotation and dispersion 
in water (Speichert & Speichert, 2004).  Seeds of N. ampla measure only a few 
millimeters across, not including the jelly like substance.  Before planting, the jelly is 
removed by wiping it off to keep the seed from floating at the surface.  Then, growers 
generally use one of two methods.  First, seeds can be allowed to float in the water 
column and germinate before they are planted into a solid substrate.  Second, some 
growers will actually plant seeds by spreading them over soil or substrate, covering them 
with a fine layer of sand (no more than 2-3 mm deep), and finally covering again with 
several centimeters of water (Speichert & Speichert, 2004; Case, 1991).  For germination, 
most water lily seeds need to be submerged in 5 to 30 centimeters of water (Conard, 
1905).  Germination time is poorly documented, and reports vary widely for 
Nymphaeaceae in general.  In one experiment, one set of seeds from a single species 
germinated in two weeks while a second set of the same species germinated after more 
than a year (Conard, 1905).  Factors that appear to affect germination time are 
temperature, light level, crowding of seeds, storing time, sowing depth and water depth 
(Case, 1991; Conard, 1905; Slocum & Robinson, 1996; Speichert & Speichert, 2004). 
N. ampla has a small presence in the literature, but the closely related species 
Nymphaea odorata has been the subject of more research.  Else & Riemer (1984) 
investigated several aspects of germination for N. odorata: seed crowding, light level, 
stratification, freezing, desiccation, ethylene, and seed coat puncture.  Investigating seed 
crowding was meant to determine whether or not a germination inducer is released into 
the water by the seeds and, if present, what that substance might be.  Else and Reimer 
97 
 
(1984) determined that the level of seed crowding affects germination for this species, 
reporting that after replicating treatments over three growing seasons there was zero 
germination in experimental vials containing 20 seeds or less in 100 mL of water (1 
seed/5 mL).  The highest germination percentages in this research occurred in vials with 
100 seeds in 20 mL of water or 5 seeds/mL of water.  They also observed that when 
water from crowded vials was transferred to uncrowded vials, germination increased, 
indicating there was a substance released from the seed into the water that promoted 
germination.  After further experimentation it was hypothesized that the substance was a 
water soluble gas.  Other studies have shown that ethylene can affect germination in 
witchweed, cocklebur, clover, peanut, and a number of other species leading researchers 
to decide to investigate the effect of this gas on N. odorata (Egley & Dale, 1970; Egley, 
1980; Esashi & Leopold, 1964; Ketring & Morgan, 1969; Letham et al., 1978).  
Application of ethylene to tightly capped seed vials did produce higher germination 
percentages than in controls in all trials.  Restricted light level reduced germination 
percentages, and freezing severely reduced germination.  Periods of drying also 
significantly inhibited germination.  Seed coat puncturing did not assist germination.  
There did appear to be an interaction effect between cold stratification and seed 
crowding.  Stratified seeds showed increased germination in crowded situations after 5 
months of chilling but no effect in uncrowded situations.  Seeds stratified for 7 to 9 
months showed nearly 100% germination in crowded conditions and improved 
germination in uncrowded conditions.  Conard (1905) reported that seeds from the 
subgenus Castalia, of which N. odorata is a member, are particularly sensitive to drought 
and temperature fluctuations, but no such information is available for the subgenus 
98 
 
Brachyceras.  The current research will investigate the effect of crowding, light level, and 
sowing method on germination of Nymphaea ampla seeds. 
Results 
 Seeds were both ordered from an online supply company (The Banana Tree, 
Easton, PA) and provided by a private grower.  Seeds procured online were dry and had 
been stored for an unknown period of time.  Seeds provided by the grower were fresh 
from the plant and were never allowed to dry completely.  All seeds were germinated in 
petri dishes.  The only definitive result that can be reported is that dry seeds, barring any 
unknown dormancy breaking requirements, will not germinate.  Regardless of treatments 
applied, fresh seeds had almost 100% germination while not a single dry seed 
germinated.  Contrary to Slocum and Robinson (1996), viviparous germination was 
observed.  Fatal algal blooms were encountered when attempts were made to 
subsequently grow seedlings in pots.  Three different fertilization techniques were 
attempted with no success in any pots that actually received fertilizer.  Only unfertilized 
(control) pots did not produce algal blooms but seedlings also did not continue 
developing.   
 Rhizomes were not available at the time of this research but if some could be 
found this would perhaps be the best way to continue research on the growth of N. ampla.  
Future research on seedling growth may be more successful in containers that provide a 
larger body of water.  Communication with Ken Landon after initial attempts to grow 
seedlings failed indicated that due to evaporation, chemical levels in small amounts of 
water can reach toxic levels for water lily seedlings regardless of any other parameter. 
 
99 
 
Literature Cited 
Bertol, E., V. Fineschi, S.B. Karch, F. Mari and I. Riezzo. 2004. Nymphaea cults in 
ancient Egypt and the New World: a lesson in empirical pharmacology. Journal of Social 
Medicine 97(2):84-85 
Case, D. 1991. Water Garden Plants: The complete guide. Crowwood Press Ltd. 
Wiltshire, UK 
Conard, H.S. 1905. The waterlilies: A monograph of the genus Nymphaea. Lark 
Publications, Suffolk, UK 
Egley, G.H. and J.E. Dale. 1970. Ethylene 1-chloroethylphosphoric acid and witchweed 
germination. Weed Science 18:586-589 
Egley, G.H. 1980. Stimulation of common cocklebur (Xanthium pennsylvanicum) and 
redroot pigweed (Amaranthus retroflexus) seed germination by injections of ethylene into 
the soil. Weed Science 28:510-514 
Else, M.J. and D.N. Riemer. 1984. Factors affecting germination of seeds of fragrant 
waterlily (Nymphaea odorata). Journal of Aquatic Plant Management 22:22-25 
Esashi, Y and A.C. Leopold. 1964. Dormancy regulation in subterranean clover seed by 
ethylene. Plant Physiology 43:1470-1472 
Ketring, D.L. and P.W. Morgan. 1969. Ethylene as a component of the emanations from 
peanut seeds and its effect on Virginia-type peanuts. Plant Physiology 42:333-337 
Landon, K. 2009. Director of San Angelo International Water Lily Collection, San 
Angelo, TX. Personal communication, October 29, 2009 
Letham, D.S., P.B. Goodwin and T.J.V. Higgins. 1978. Phytohormones and related 
compounds: A comprehensive treatise. Elsevier/North Holland Biomedical Press, 
Amsterdam 
Muhlberg, H. 1981. The complete guide to water plants. EP Publishing Ltd. German 
Democratic Republic 
Purcell, J.K. 1995. Methods of propagation for aquatic plants. Combined Proceedings 
International Plant Propagators? Society 45:327-330 
100 
 
Slocum, P.D. and P. Robinson. 1996. Water gardening: Water lilies and lotuses. Timber 
Press. Portland, OR 
Speichert, G and S. Speichert. 2004. Encyclopedia of water garden plants. Timber Press. 
Portand, OR 
Wunderlin, R.P. 1980. Nymphaea ampla (Nymphaeaceae), a waterlily new to Florida. 
Phytologia 45(1):82-84