211684) of the European Commission within its Seventh Framework Programme. The authors thank Dr Anna Rusznyak for critically reading the manuscript. Please note: Wiley-Blackwell is not responsible for the content or functionality of any supporting materials supplied by the authors. Any queries (other than missing material) should be directed to the corresponding
author for the learn more article. “
“Vibrio parahaemolyticus is an enteric pathogen, which can cause acute gastroenteritis in humans after consumption of raw or partially cooked seafood, and specific molecular markers are necessary for its accurate identification by PCR methods. In the present study, 23 protein-coding sequences were identified by the comparative genomics method Volasertib ic50 as V. parahaemolyticus-specific candidate markers. We targeted the irgB gene (vp2603), coding for iron-regulated virulence regulatory protein IrgB, in order to develop a PCR method for the detection of V. parahaemolyticus. PCR specificity was identified by amplification of 293 V. parahaemolyticus templates and by the loss of a PCR product with 11 strains from other Vibrio species and 35 non-Vibrio bacterial strains. The PCR assay had the 369-bp fragment and the sensitivity of 0.17 pg purified genomic DNA from V. parahaemolyticus. Furthermore, a multiplex PCR assay for the detection of total and virulent strains of V. parahaemolyticus
was developed by targeting irgB, tdh and trh genes. These data indicated that
the irgB gene is a new and effective marker for the detection of V. parahaemolyticus. In addition, this study demonstrates that genome sequence comparison has a powerful application in identifying specific markers for the detection and identification of bacterial pathogens. Vibrio parahaemolyticus is a Gram-negative bacterium commonly found in marine and estuarine environments around the world (Daniels et al., 2000). This organism may lead to acute gastroenteritis 4��8C characterized by diarrhea, headache, vomiting, nausea and low fever, after consumption of raw or partially cooked fish or shellfish (Tuyet et al., 2002; DePaola et al., 2003). Outbreaks of V. parahaemolyticus have been reported from many countries and regions such as China (Liu et al., 2004b), Japan (Alam et al., 2002), the United States (McLaughlin et al., 2005) and some European countries (Martinez-Urtaza et al., 2005). Therefore, early detection and identification of V. parahaemolyticus strains in clinical and food samples is essential for diagnosis and implementing timely risk management decisions. However, the detection of V. parahaemolyticus using conventional culture- and biochemical-based assays is time consuming and laborious, requiring more than 3 days. Those strains that produce thermostable direct hemolysin (TDH) and/or TDH-related hemolysin (TRH) are considered virulent for humans (Dileep et al., 2003; Zhang & Austin, 2005).