In vitro liquid mass production of a South African isolate of Heterorhabditis zealandica

Abstract

Developing repeatable protocols for the in vitro liquid mass production of entomopathogenic nematodes (EPNs) is a difficult task and depends on the nematode species being cultured. Of critical importance is the establishment of a monoxenic population of nematodes as a stock culture for optimisation and experimental purposes. Establishing a new stock inoculum culture flask of pure infective juveniles (IJs) is challenging, particularly for the Heterorhabditis species, due to their affinity for developing into an amphimictic second generation that does not copulate in liquid culture flasks. Developing mass production protocols for multiple EPNs is advisable because different pest insects are susceptible to different species of EPN. This study attempted to mass-produce a South African isolate of Heterorhabditis zealandica and its symbiotic bacteria, Photorhabdus thracensis, using in vitro liquid culture technology methods previously developed for Steinernema species. The results indicate that the pre-culture protocols developed for Steinernema species are applicable to a H. zealandica isolate. Moreover, the results, in terms of the protein source optimisation experiments, confirm that different EPN species have different culture conditions and nutrient requirements, with H. zealandica seeming to prefer soy-based protein instead of egg yolk, having higher recovery and producing more hermaphrodites, using soy protein. This study illustrates the importance of developing dependable and infallible preculture methods, prior to the flask mass production process.