The Native Microbial Community of Gastropod-Associated Phasmarhabditis Species Across Central and Southern California.

Nematodes in the genus Phasmarhabditis can infect and kill slugs and snails, which are important agricultural pests. This useful trait has been commercialized by the corporation BASF after they mass produced a product labeled Nemaslug®. The product contains Phasmarhabditis hermaphrodita, which has been cultured with Moraxella osloensis, a bacterial strain that was originally thought to be responsible for causing mortality in slugs and snails. The exact mechanism leading to death in a Phasmarhabditis infected host is unknown but may involve contributions from nematode-associated bacteria. The naturally occurring microbial community of Phasmarhabditis is unexplored; the previous Phasmarhabditis microbial community studies have focused on laboratory grown or commercially reared nematodes, and in order to obtain a deeper understanding of the parasite and its host interactions, it is crucial to characterize the natural microbial communities associated with this organism in the wild. We sampled Phasmarhabditis californica, Phasmarhabditis hermaphrodita, and Phasmarhabditis papillosa directly from their habitats in Central and Southern California nurseries and garden centers and identified their native microbial community via 16S amplicon sequencing. We found that the Phasmarhabditis microbial community was influenced by species, location, and possibly gastropod host from which the nematode was collected. The predominant bacteria of the Phasmarhabditis isolates collected included Shewanella, Clostridium perfringens, Aeromonadaceae, Pseudomonadaceae, and Acinetobacter. Phasmarhabditis papillosa isolates exhibited an enrichment with species belonging to Acinetobacter or Pseudomonadaceae. However, further research must be performed to determine if this is due to the location of isolate collection or a species specific microbial community pattern. More work on the natural microbial community of Phasmarhabditis is needed to determine the role of bacteria in nematode virulence.

Dose dependence of Phasmarhabditis isolates (P. hermaphrodita, P. californica, P. papillosa) on the mortality of adult invasive white garden snails (Theba pisana).

Theba pisana is an invasive snail pest which has established itself in San Diego County and some areas of Los Angeles County, California. The snail has grown to large populations in some areas and mitigation is becoming necessary to stop the spread of the species. In a previous study, three US strains of Phasmarhabditis species (P. californica, P. papillosa, and P. hermaphrodita) effectively killed juvenile (0.25 gram each, 4-6 mm wide) T. pisana in laboratory conditions at 5 times (150 IJs/cm2) the recommended dose. Based on laboratory assays, we demonstrated that the same three US strains of Phasmarhabditis can effectively kill larger adult T. pisana (0.4-1.2 gram, 11.5-15mm wide) in two weeks at the same dose. The strains were more efficient at killing T. pisana than the compared molluscicide Sluggo Plus®. Results further showed that the most virulent P. californica did not effectively kill T. pisana at lower doses of 30 IJs/cm2 and 90 IJs/cm2. Additional research is needed to develop the most efficient means of application of Phasmarhabditis to mitigate T. pisana in the field.

Distribution of Phasmarhabditis (Nematode: Rhabditidae) and Their Gastropod Hosts in California Plant Nurseries and Garden Centers.

Three species of Phasmarhabditis were recovered from 75 nurseries and garden centers in 28 counties in California during fall and winter 2012-2021. A total of 18 mollusk species were recovered, most of them invasive. Nematodes were identified by sequencing the D2-D3 expansion segments of the large subunit (LSU or 28S) rRNA. Based on these surveys, P. californica was the most widespread species (37 isolates, 53.6% recovery); followed by P. hermaphrodita (26 isolates; 37.7% recovery); P. papillosa and a closely related P. papillosa isolate (6 isolates; 8.7% recovery). Nematode isolates were mainly collected from four invasive slugs (Deroceras reticulatum, D. laeve, Arion hortensis agg, Ambigolimax valentianus) and snails (Oxychilus spp. and Discus spp.). Results suggest that P. californica and P. hermaphrodita share an ecological niche in Northern, Central, Coastal, and Southern California, north of Los Angeles County.

TARANTOBELUS JEFFDANIELSI N. SP. (PANAGROLAIMOMORPHA; PANAGROLAIMIDAE), A NEMATODE PARASITE OF TARANTULAS.

Multiple tarantula deaths for a wholesale breeder were reported in 2018. The breeder noticed white discharge in the oral cavities of the tarantulas. Upon inspection, it was discovered that the white discharge was a large group of nematodes intertwined inside the tarantula's oral cavity. We examined the nematodes and propose a new species, Tarantobelus jeffdanielsi n. sp., in the currently monotypic genus Tarantobelus based on a combination of morphological and morphometrical data and unique nuclear rDNA 28S and 18S sequences. Based on phylogenetic analyses, the previously described Tarantobelus arachnicida was relocated, along with T. jeffdanielsi, into the family Panagrolaimidae. We also provide evidence of the ability of T. jeffdanielsi to parasitize Galleria mellonella larvae and the tarantula Grammostola pulchra. The life span and fecundity of the new species were also assessed, resulting in an 11.2-d average life span, and a total fertility rate of 158 nematodes/adult.

Entomopathogenic nematode-gastropod interactions.

Entomopathogenic nematodes (EPNs) infect and kill insects and have been successfully used in the biological control of some insect pests. Slugs and snails are known to be significant pests of agriculture and serve as vectors for disease-causing microbes that can affect crops and humans. The potential of EPNs to be used in the biological control of gastropods has not been well-studied. The few studies that have been performed on the efficacy of EPNs in controlling gastropod pests and vectors were reviewed. Suggested criteria for further assessments of EPN-gastropod interactions are: Dose of EPNs used, length of infection assays, host biology, nematode biology and development, and Koch's postulates. There are provocative data suggesting that EPNs may be useful biological control agents against gastropod pests of agriculture and vectors of disease, though additional studies using the suggested criteria are needed, including the publication of negative data or studies where EPNs were not efficacious or successful in controlling gastropods.

Mortality of the invasive white garden snail Theba pisana exposed to three US isolates of Phasmarhabditis spp (P. hermaphrodita, P. californica, and P. papillosa).

Theba pisana is a serious snail pest in many parts of the world and affects diverse crops including grain, vegetables, grapevines, and ornamental plants and shrubs. Due to its gregarious nature, ability to reproduce rapidly, and the difficulty of controlling it by conventional methods, it has the potential to become a significant pest where introduced. Mitigating this pest is an important challenge that must be addressed. Phasmarhabditis hermaphrodita, is a gastropod-killing nematode that is commercially available only in Europe (Nemaslug ®) and Sub-Saharan Africa (Slugtech ® SP). The use of effective gastropod-killing nematodes in the genus Phasmarhabditis (P. hermaphrodita, P. californica and P. papillosa) in California may provide one strategy for alleviating the potential damage and further spread of these snails, which are currently limited to San Diego and Los Angeles counties. Laboratory assays demonstrated for the first time that US isolates of P. hermaphrodita, P. californica and P. papillosa at 150 DJs/cm2 caused significant mortality and are equally lethal to T. pisana. Molluscicidal efficacy of these nematodes are comparable with those of iron phosphate, at the recommended high dose of 4.88 kg/m2. Additional trials are needed to determine their effects at lower dose and whether they are dependent on the size or age of the snails.