Molecular epidemiology of clinical isolates of Pseudomonas aeruginosa isolated from horses in Ireland

A Tazumi,T Buckley,M Matsuda,Bc Millar,Ce Goldsmith,Js Elborn,Y Maeda,Je Moore,Jsg Dooley

doi:10.1186/2046-0481-62-7-456

Abstract

Clinical isolates (n = 63) of Pseudomonas aeruginosa obtained from various sites in 63 horses were compared using ERIC2 RAPD PCR to determine their genetic relatedness. Resulting banding patterns (n = 24 genotypes) showed a high degree of genetic heterogeneity amongst all isolates examined, indicating a relative non-clonal relationship between isolates from these patients, employing this genotyping technique. This study characterised 63 clinical isolates into 24 distinct genotypes, with the largest cluster (genotype E) accounting for 10/63 (15.9%) of the isolates. ERIC2 RAPD PCR proved to be a highly discriminatory molecular typing tool of P. aeruginosa in isolates recovered from horses. With the adoption of several controls to aid reproducibility, this technique may be useful as an alternative to PFGE, particularly in epidemiological investigations of outbreaks where speed may be a significant parameter. This is the first report of clonal heterogeneity amongst P. aeruginosa from horses and demonstrated that ERIC RAPD PCR is a rapid method for the examination of this species in horses, which may be useful in outbreak analysis.

Highlights

Pseudomonas aeruginosa is an important Gram-negative bacterial pathogen in horses, associated with the genital tract (Samper and Tibary 2006), infection due to this causal agent may occur at virtually any anatomical site in the horse
We attempted to examine the genetic relatedness of a collection of 63 clinical isolates of P. aeruginosa, obtained from an equal number of horses throughout Ireland during the five year period, 20032007, employing an ERIC random amplification of polymorphic DNA (RAPD) PCR molecular technique
Several molecular typing schemes have been applied to help elucidate the molecular epidemiology of populations of P. aeruginosa in human clinical medicine and these methodologies have been reviewed comprehensively by Speert (2002), including pulsed-field gel electrophoresis (PFGE)

Summary

Introduction

Pseudomonas aeruginosa is an important Gram-negative bacterial pathogen in horses, associated with the genital tract (Samper and Tibary 2006), infection due to this causal agent may occur at virtually any anatomical site in the horse. Detailed genetic analysis at the subspecies (strain) level gives insights into the variability within a bacterial population and generates evidence on genome evolution, which in turn leads to bacterial adaptation to various environmental conditions This information can be used in a clinical setting to help identify clinical isolates that occur frequently in animals, so that further work may be undertaken to identify common virulence characteristics in equine strains and help, for example, in directing choice of strains to use in vaccine development. One of the PCR-based methods, namely random amplification of polymorphic DNA (RAPD)-PCR, known as arbitrarily primed-polymerase chain reaction (AP-PCR), has been described to be useful on account of its simplicity and utility for analysis of large throughput samples (Gürtlet and Mayall 2001) This technique utilises a variable short length arbitrary primer, and is advantageous, as it does not require any previous knowledge of the target DNA sequence data. The aim of this study was to examine the genetic relatedness of P. aeruginosa isolated from a variety of sites in Irish horses, over the five year period, 2003-2007, through employment of ERIC2-RAPD PCR

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Conclusion