Biological control (biocontrol) as a component of pest management strategies reduces reliance on synthetic chemicals, and seemingly offers a natural approach that minimizes environmental impact. However, introducing a new organism to new environments as a classical biocontrol agent can have broad and unanticipated biodiversity effects and conservation consequences. Nematodes are currently used in a variety of commercial biocontrol applications, including the use of Phasmarhabditis hermaphrodita as an agent targeting pest slug and snail species. This species was originally discovered in Germany, and is generally thought to have European origins. P. hermaphrodita is sold under the trade name Nema-slug®, and is available only in European markets. However, this nematode species was discovered in New Zealand and the western United States, though its specific origins remained unclear. In this study, we analyzed 45 nematode strains representing eight different Phasmarhabditis species, collected from nine countries around the world. A segment of nematode mitochondrial DNA (mtDNA) was sequenced and subjected to phylogenetic analyses. Our mtDNA phylogenies were overall consistent with previous analyses based on nuclear ribosomal RNA (rRNA) loci. The recently discovered P. hermaphrodita strains in New Zealand and the United States had mtDNA haplotypes nearly identical to that of Nemaslug®, and these were placed together in an intraspecific monophyletic clade with high support in maximum likelihood and Bayesian analyses. We also examined bacteria that co-cultured with the nematode strains isolated in Oregon, USA, by analyzing 16S rRNA sequences. Eight different bacterial genera were found to associate with these nematodes, though Moraxella osloensis, the bacteria species used in the Nemaslug® formulation, was not detected.

Phasmarhabditis bohemica n. sp. is described and illustrated from the body of Deroceras reticulatum from the village of Chelčice, the Czech Republic. Morphological and molecular data showed that the new isolate is close to other species of Phasmarhabditis, particularly P. californica and P. papillosa. Females are characterised by a body length of 2079 (1777-2222) μm and a long tapering tail with prominent papilliform phasmids located laterally in the mid-tail region. Males are 1683 (1515-1818) μm long. They have a peloderan bursa, with nine pairs of rays, 1/1/1/2/1/3, and a reflexed testis 495 (434-555) μm long. Dauer juveniles are thin, 553 (474-636) μm long, with prominent lateral fields consisting of two prominent ridges and three incisures. Small subunit (18S), ITS, and D2-D3 expansion segments of the large subunit of ribosomal DNA were used to analyse the phylogenetic relationships of sequenced species in Phasmarhabditis and other closely related species. Our preliminary observations suggest that the newly described species may be a facultative mollusc-parasitic nematode that is able to survive permanently in the saprobic phase on decaying organic matter. The ecology, morphology, and phylogenetic positions of P. bohemica n. sp. are discussed.

The effect of different growing substrates on the development of the slug parasite Phasmarhabditis hermaphrodita has been studied in a series of laboratory experiments. Wild, laboratory and Nemaslug strains of P. hermaphrodita and their monoxenic counterparts with Moraxella osloensis (CCM 5605T) were reared on homogenised pig kidney, Deroceras reticulatum, Arion lusitanicus, and Galleria mellonella, the faeces of D. reticulatum and A. lusitanicus, or leaf compost. Development time, yield, lipid reserves and the body length of nematodes were assessed. All P. hermaphrodita strains were able to grow and reproduce on all tested substrates; however, yields were markedly higher on invertebrate media. Lipid content and body size varied across the substrates and strains. A uniform response of all nematode strains and the most dramatic differences among substrates were recorded in yields, which could indicate that the quality of the substrate is expressed mainly in yield. Monoxenized strains had higher yield, length, lipid content and shorter development time. In general, the strain differences between the observed parameters were most likely due to different bacterial associates. The dramatic differences in yields on invertebrate substrates, in comparison to the others, illustrate the evolutionary advantage of the association of nematodes with invertebrates.