Genetic characterization of trh positive Vibrio spp. isolated from Norway

Anette B Ellingsen,Narjol González-Escalona,Jaran S Olsen,Per E Granum,Liv M Rørvik

doi:10.3389/fcimb.2013.00107

Abstract

The thermostable direct hemolysin (TDH) and/or TDH-related hemolysin (TRH) genes are carried by most virulent Vibrio parahaemolyticus serovars. In Norway, trh+ V. parahaemolyticus constitute 4.4 and 4.5% of the total number of V. parahaemolyticus isolated from blue mussel (Mytilus edulis) and water, respectively. The trh gene is located in a region close to the gene cluster for urease production (ure). This region was characterized in V. parahaemolyticus strain TH3996 and it was found that a nickel transport operon (nik) was located between the first gene (ureR) and the rest of the ure cluster genes. The organization of the trh-ureR-nik-ure gene cluster in the Norwegian trh+ isolates was unknown. In this study, we explore the gene organization within the trh-ureR-nik-ure cluster for these isolates. PCR analyses revealed that the genes within the trh-ureR-nik-ure gene cluster of Norwegian trh+ isolates were organized in a similar fashion as reported previously for TH33996. Additionally, the phylogenetic relationship among these trh+ isolates was investigated using Multilocus Sequence Typing (MLST). Analysis by MLST or ureR-trh sequences generated two different phylogenetic trees for the same strains analyzed, suggesting that ureR-trh genes have been acquired at different times in Norwegian V. parahaemolyticus isolates. MLST results revealed that some pathogenic and non-pathogenic V. parahaemolyticus isolates in Norway appear to be highly genetically related.

Highlights

Vibrio parahaemolyticus is a halophilic, Gram-negative proteobacterium ubiquitous in the marine and estuarine environment worldwide (Iida and Honda, 2006)
The mechanism by which V. parahaemolyticus causes enteric disease is not fully understood, two virulence factors are usually associated with clinical isolates: the genes encoding for thermostable direct hemolysin (TDH), and TDH-related hemolysin (Gonzalez-Escalona et al, 2006)
Our study presents new information regarding the genetic relationship among trh+ isolates, and introduces a PCR screening method demonstrating that the organization of the trh-ureR-nik-ure gene cluster found in the Norwegian trh+ V. parahaemolyticus isolates is identical to the one previously described for strain TH3996 (Park et al, 2000)

Summary

Introduction

Vibrio parahaemolyticus is a halophilic, Gram-negative proteobacterium ubiquitous in the marine and estuarine environment worldwide (Iida and Honda, 2006). It is one of the main bacterial pathogens associated with raw and undercooked seafood in Asia and the U.S (Su and Liu, 2007). Several variants of tdh have been identified, all of which are about 98% identical (Nishibuchi and Kaper, 1995). In the case of trh, only two gene variants have been described, denoted trh and trh; these are about 84% identical in sequence (Kishishita et al, 1992). While V. parahaemolyticus can carry both genes and simultaneously express them, it appears that such isolates produces less TDH than trh negative isolates (Xu et al, 1994)

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