Identification of Vibrio alginolyticus virulent strain‐specific DNA regions by suppression subtractive hybridization and PCR

• Published in: Journal of applied microbiology 2020-12, Vol.129 (6), p.1472-1485
• Main Authors: Xie, Z.‐Y., Gong, X.‐X., Xu, X.‐D., Mei, B., Xuan, X.‐Z., Long, H., Zhang, X., Cai, X.‐N.
• Format: Article
• Language: English
• Subjects: Animals; Bacteria; Chemotaxis; Cloning; Deoxyribonucleic acid; DNA; DNA, Bacterial - genetics; Fish Diseases - microbiology; Fish farms; Fragments; Genome, Bacterial - genetics; Genomes; Harbors; Homology; Hybridization; Pathogens; PCR; Permease; Polymerase Chain Reaction; Proteins; Shells; Southern blot hybridization; Species Specificity; SSH; Subtractive Hybridization Techniques; Transcription; Transportation systems; V. alginolyticus; Vibrio alginolyticus; Vibrio alginolyticus - genetics; Vibrio alginolyticus - isolation & purification; Vibrio alginolyticus - pathogenicity; Vibrio Infections - microbiology; Vibrio Infections - veterinary; Virulence; Virulence - genetics; virulent strain‐specific DNA region; Waterborne diseases
• ISSN: 1364-5072; 1365-2672
• DOI: 10.1111/jam.14739

Aims Vibrio alginolyticus was frequently isolated from diseased farmed fish in the coaster waters of Hainan Island over the past two decades. In this study, we attempted to identify candidates of virulent strain‐specific DNA regions for this pathogen. Methods and Results Suppression subtractive hybridization (SSH) and PCR were successively performed between the typical virulent strain and avirulent strain of V. alginolyticus, in which they shared 99·54% homology of 16S rDNAs. Out of 2873 subtracted clones, nine clones were finally indicated to harbour virulent strain‐specific DNA fragments. The receivable functions of the major fragments in the nine clones were believed to encode methyl‐accepting chemotaxis protein (n = 1), type VI secretion system‐associated FHA domain protein TagH (n = 1), diguanylate cyclase (n = 1), AraC family transcriptional regulator (n = 1), ABC‐type uncharacterized transport system permease component (n = 1) and hypothetical proteins (n = 4). Two hypothetical proteins contain several disordered regions. Conclusions Some specific DNA regions existed in the virulent strain of V. alginolyticus, and the SSH assay could be a highly sensitive method for identifying virulent regions in pathogens. Significance and Impact of the Study This report is the first to describe the identification of virulent strain‐specific DNA regions in the V. alginolyticus genome, which is helpful in developing virulent strain‐specific rapid detection methods and is a pivotal precondition for clarifying the molecular virulence mechanism of V. alginolyticus.