Wide interguild relationships among entomopathogenic and free.living nematodes in soil as measured by real time qPCR

Abstract

Entomopathogenic nematodes (EPNs) are promising biological control agents of soil-dwelling insect pests of many crops. These nematodes are ubiquitous in both natural and agricultural areas. Their efficacy against arthropods is affected directly and indirectly by food webs and edaphic conditions. It has long been suggested that a greater understanding of EPN ecology is needed to achieve consistent biological control by these nematodes and the development of molecular tools is helping to overcome obstacles to the study of cryptic organisms and complex interactions. Here we extend the repertoire of molecular tools to characterize soil food webs by describing primers/probe set to quantify certain free-living, bactivorous nematodes (FLBNs) that interact with EPNs in soil. Three FLBN isolates were recovered from soil baited with insect larvae. Morphological and molecular characterization confirmed their identities as Acrobeloides maximum (RT-1-R15C and RT-2-R25A) and Rhabditis rainai (PT-R14B). Laboratory experiments demonstrated the ability of these FLBNs to interfere with the development of Steinernema diaprepesi, Steinernema riobrave and Heterorhabditis indica parasitizing the weevil Diaprepes abbreviatus (P < 0.001), perhaps due to resource competition. A molecular probe was developed for the strongest competitor, A. maximum. We selected the highly conserved SSU rDNA sequence to design the primers/ probe, because these sequences are more abundantly available for free-living nematodes than ITS sequences that can likely provide better taxonomic resolution. Our molecular probe can identify organisms that share P98% similarity at this locus. The use of this molecular probe to characterize soil communities from samples of nematode DNA collected within a citrus orchard revealed positive correlations (P < 0.01) between Acrobeloides-group nematodes and total numbers of EPNs (S. diaprepesi, H. indica and Heterorhabditis zealandica) as well as a complex of nematophagous fungi comprising Catenaria sp. and Monachrosporium gephyropagum that are natural enemies of EPNs. These relationships can be broadly interpreted as supporting Linford’s hypothesis, i.e., decomposition of organic matter (here, insect cadavers) greatly increases bactivorous nematodes and their natural enemies.