Identification of Plant-Parasitic Nematodes Using FAME Analysis

Abstract

For several years, chromatographic analysis of fatty acid methyl esters (FAME) has been used to identify bacteria more quickly than traditional methods. The objective of this study was to determine the applicability of FAME analysis for the identification of plant-parasitic nematodes for use in diagnostic laboratories. Meloidogyne incognita, Rotylenchulus reniformis, and Heterodera glycines were statistically distinct (P < 0.0001) and could be identified in samples containing at least 100 total individuals for M. incognita and R. reniformis and 5 cysts for H. glycines. Two fatty acids (16:1 ω5c and 18:1 ω5c) indicate the presence of R. reniformis when comparing samples containing 100 or greater individuals of either R. reniformis or M. incognita. Significant variation (P < 0.0001) in the fatty acid profiles of M. incognita and R. reniformis was observed when either species was increased on tomato, cotton, or soybean plants, but variations of the FAME profile for each nematode allowed for identification of each species regardless of host. Mixed-species samples of M. incognita and R. reniformis could be differentiated (P < 0.0075) from one another and single-species samples of each species in 100% of comparisons when samples contained 5000 total individuals and in 95% of comparisons when samples contained 500 total individuals. In samples containing 500 total individuals, it was not possible to differentiate between a (50:50) ratio of M. incognita and R. reniformis or a sample with a (25:75) ratio of M. incognita and R. reniformis. However, all other comparisons, including these two ratios separately, were identifiable. Three Meloidogyne species (M. arenaria, M. hapla, and M. javanica) and three M. incognita races (races 1, 2, and 3) all produced distinct (P < 0.0001) fatty acid profiles and could be identified with 85.6% overall accuracy to the race level. Soil containing R. reniformis produced a fatty acid profile significantly different (P < 0.0001) from soil without R. reniformis and contained higher percentages of fatty acids found in nematodes. Certain fatty acids found in R. reniformis were found in higher quantities in soil infested with the nematode and may indicate its presence. The compiled library identified nematode samples with 90.4% total accuracy. Genus level identification was accurate at 100% for the samples studied and species level identification was 94.4% accurate. FAME analysis appears to be a promising alternative for identification of plant-parasitic nematodes.