Studies on Aeromonas taxonomy.

Abstract

A diverse collection of 400 Aeromonas strains from clinical, veterinary and environmental sources worldwide has been assembled. A number of molecular-based typing methods: Amplified-Fragment Length Polymorphism (AFLP), Repetitive Extragenic Palindromic sequence Polymerase Chain Reaction (REP-PCR) and Randomly amplified polymorphic DNA (RAPD), have been adapted for this genus and used to assess the relatedness of isolates from different sources. AFLP proved to be the most discriminatory, RAPD was found to be of limited value for discriminating Aeromonas spp. Though less discriminatory, a novel rapid and inexpensive one-tube method of performing REP-PCR is described as well as adaptation of the technique for use with a DNA sequencer for the first time. A novel use of degenerate PCR primer antagonists for improving the specificity of REP-PCR that may be of more general applicability in PCR is also described. The AFLP data contributes much to the debate surrounding Aeromonas taxonomy. The data supports the hypothesis that A. popoffli and A.bestarium are stable and genetically distinct taxa. Evidence is presented for the first time that A.caviae HG4 may comprise two genetically distinct sub-groups. Further evidence is presented for the existence within HG7, of a group phenotypically similar to HG10. Though genetically distinct groups corresponding to A.encheleia, HG11 and A.eucrenophila were identified, they appeared to separate below the species level. A.encheleia, however, was found to be far more diverse than previously suspected. Strains implicated in diseases in different hosts were not strongly correlated with particular species. However the most apparently virulent strains appeared to be rather specialised and host-specific. In particular a genetically distinct group of HG3 isolates, geographically widespread and stable over several years, was recovered only from porpoises. The presence of Class 1 integron-specific sequences is reported for the first time Aeromonas spp. Though these appeared to have spread widely between strains isolated from different hosts, they were found to be comparatively rare in environmental isolates. Evidence of such widespread horizontal transfer has important implications for the population structure of the genus.