Several slug species are highly pestiferous and threaten global sustainable agriculture. Current control methods rely heavily on metaldehyde pellets, which are often ineffective, harm nontarget organisms and have been banned in some countries. A viable alternative is the parasitic nematode Phasmarhabditis hermaphrodita (and recently P. californica), which has been formulated into a biological control agent (Nemaslug®) to control slugs across northern Europe. Nematodes are mixed with water and applied to soil where they seek out slugs, penetrate behind the mantle and kill them in 4–21 days. Phasmarhabditis hermaphrodita has been on the market since 1994 and since then there has been ample research on its use. Here we review the research carried out on P. hermaphrodita over the last 30 years since its development and release as a commercial product. We provide information on life cycle, worldwide distribution, history of commercialisation, gastropod immunity, host range, ecological and environmental factors that affect its success in the field, bacterial relationships, and summarise results of field trials. Finally, we suggest future directions for P. hermaphrodita research (and other Phasmarhabditis species) to enhance its use as a biological control agent to control slugs for the next 30 years. © 2023 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

The reproductive system regulates lifespan in insects, nematodes and vertebrates. In Caenorhabditis elegans removal of germline increases lifespan by 60% which is dependent upon insulin signaling, nuclear hormone signaling, autophagy and fat metabolism and their microRNA-regulators. Germline-deficient C. elegans are also more resistant to various bacterial pathogens but the underlying molecular mechanisms are largely unknown. Firstly, we demonstrate that previously identified genes that regulate the extended lifespan of germline-deficient C. elegans (daf-2, daf-16, daf-12, tcer-1, mir-7.1 and nhr-80) are also essential for resistance to the pathogenic bacterium Xenorhabdus nematophila. We then use a novel unbiased approach combining laser cell ablation, whole genome microarrays, RNAi screening and exposure to X. nematophila to generate a comprehensive genome-wide catalog of genes potentially required for increased lifespan and innate immunity in germline-deficient C. elegans. We find 3,440 genes to be upregulated in C. elegans germline-deficient animals in a gonad dependent manner, which are significantly enriched for genes involved in insulin signaling, fatty acid desaturation, translation elongation and proteasome complex function. Using RNAi against a subset of 150 candidate genes selected from the microarray results, we show that the upregulated genes such as transcription factor DAF-16/FOXO, the PTEN homolog lipid phosphatase DAF-18 and several components of the proteasome complex (rpn-6.1, rpn-7, rpn-9, rpn-10, rpt-6, pbs-3 and pbs-6) are essential for both lifespan and immunity of germline deficient animals. We also identify a novel role for genes including par-5 and T12G3.6 in both lifespan-extension and increased survival on X. nematophila. From an evolutionary perspective, most of the genes differentially expressed in germline deficient C. elegans also show a conserved expression pattern in germline deficient Pristionchus pacificus, a nematode species that diverged from C. elegans 250-400 MYA.

Exoskeletons have evolved 18 times independently over 550 MYA and are essential for the success of the Gastropoda. The gastropod shell shows a vast array of different sizes, shapes and structures, and is made of conchiolin and calcium carbonate, which provides protection from predators and extreme environmental conditions. Here, I report that the gastropod shell has another function and has been co-opted as a defense system to encase and kill parasitic nematodes. Upon infection, cells on the inner layer of the shell adhere to the nematode cuticle, swarm over its body and fuse it to the inside of the shell. Shells of wildCepaea nemoralis,C. hortensisandCornu aspersumfrom around the U.K. are heavily infected with several nematode species includingCaenorhabditis elegans. By examining conchology collections I show that nematodes are permanently fixed in shells for hundreds of years and that nematode encapsulation is a pleisomorphic trait, prevalent in both the achatinoid and non-achatinoid clades of the Stylommatophora (and slugs and shelled slugs), which diverged 90–130 MYA. Taken together, these results show that the shell also evolved to kill parasitic nematodes and this is the only example of an exoskeleton that has been co-opted as an immune system.

Essential oils are being increasingly utilised as a biorational element in integrated pest management regimes. Whereas there has been much research on the effects of these oils on mortality and behaviour of pestiferous molluscs, insects and nematodes, there has (to the present authors’ knowledge) been no research into their effects on the mortality and behaviour of beneficial nematodes. We address this lacuna by conducting laboratory assays on the behaviour (thrashing assays) and mortality of 13 essential oils plus controls of Tween and water on the malacophagous nematode Phasmarhabditis hermaphrodita and two species of entomopathogenic nematodes, namely: Steinernema feltiae and Heterorhabditis bacteriophora. Mortality results showed an “all or nothing” response with only three oils—pine oil, peppermint and lemongrass—displaying intermediate mortalities. Also, toxicity of essential oils was conserved across phylogenetically quite distinct beneficial nematode species with oil toxicity largely being repeated across the three beneficial nematodes. Thrashing assays confirmed the toxic effects of certain oils. We recommend that the effects of essential oils be tested on beneficial organisms in ecosystems before choices are made over which oils should be used. The present paper highlights some oils which are not toxic to beneficial nematodes but may affect their behaviour. Research should focus on these oils as part of a biorational control programme for pestiferous molluscs and insects.

The genetic mechanisms of how free-living nematodes evolved into parasites are unknown. Current genetic model nematodes (e.g.Caenorhabditis elegans) are not well suited to provide the answer, and mammalian parasites are expensive and logistically difficult to maintain. Here we propose the terrestrial gastropod parasitePhasmarhabditis hermaphroditaas a new alternative to study the evolution of parasitism, and outline the methodology of how to keepP. hermaphroditain the lab for genetic experiments. We show thatP. hermaphrodita(and several otherPhasmarhabditisspecies) are easy to isolate and identify from slugs and snails from around the UK. We outline how to make isogenic lines using ‘semi-natural’ conditions to reduce in-lab evolution, and how to optimize growth using nematode growth media (NGM) agar and naturally isolated bacteria. We show thatP. hermaphroditais amenable to forward genetics and thatuncandsmamutants can be generated using formaldehyde mutagenesis. We also detail the procedures needed to carry out genetic crosses. Furthermore, we show natural variation within ourPhasmarhabditiscollection, with isolates displaying differences in survival when exposed to high temperatures and pH, which facilitates micro and macro evolutionary studies. In summary, we believe that this genetically amenable parasite that shares many attributes withC. elegansas well as being in Clade 5, which contains many animal, plant and arthropod parasites, could be an excellent model to understand the genetic basis of parasitism in the Nematoda.