Surveys for entomopathogenic bacteria and their associated nematode hostswere conducted locally (University of Bath campus) and across southernEngland. Sampling involved trialing a novel Android app. (Epicollect) tomanage sample collection data. Galleria larvae were used to bait UK soilsamples. Insects which became infected were placed on White traps to collectany emerging nematodes, from which bacteria were isolated. Bacteria werealso isolated from the haemolymph of any infected larvae. Bacterial isolateswere classified on the basis of 16s rDNA and recA gene sequences. Serratiaproteamaculans-like strains dominated the samples, and Multilocus sequenceanalysis (MLSA) was developed for the characterization of these Serratiaisolates. We determined the sequences of (350-450-bp) fragments from fivehousekeeping genes of 84 isolates of Serratia proteamaculans. MLSA wasshown to be effective for distinguishing closely related strains found in theinsects’ haemolymph and from different nematodes. goeBURST was used tovisualize the relationships between the STs, and the data showed a high levelof discrimination, resolving 69 STs from the 84 isolates. In addition, the dataderived from this study were represented in a phylogenetic network using theSplits Tree-network methods, to show the rate of recombination within andbetween the genes.From a total of 256 infected Galleria 23.04% were nematode positive. Thenematodes were identified based on 18S rDNA 19 isolates were closerelatives of the species Pristionchus entomophaga and Diplogasteriodesmagnus (Diplogastridae). A further 16 isolates were more closely related toSteinernema glaseri (Steinernematidae). All three nematode types wereisolated from diverse habitats and soil types, but were isolated morefrequently in cold seasonal conditions. The bacterial sequence data suggestthat the nematode- associated strains of bacteria belong to specific clades,distinct from the free living infective strains, which hints at ecological diversitywithin the S. proteamaculans population.Two of the Serratia proteamaculans-like strains had been chromosomallylabeled with GFP to confirm the specifics of their association with thenematode hosts. The associated S. proteamaculans-like isolates isolatedfrom Bath and Chepstow soils were examined further for their pathogenicity toGalleria mellonella and Manduca sexta larvae. Serratia Bath isolates, isolatedfrom Pristionchus were more virulent toward both insect hosts than theSerratia from the Chepstow isolates associated with Steinernema nematodes.This suggests that host specificity may play important role in the virulence ofthe strain.
|Date of Award||6 Sep 2013|
|Supervisor||Edward Feil (Supervisor) & Nicholas Waterfield (Supervisor)|
- multilocus sequence typing