Vibrio vulnificus is a marine bacterial species that can cause fatal septicemia via ingestion of contaminated seafood and serious wound infection in humans, particularly those with severe chronic liver disorders like liver cirrhosis and hepatoma. To understand how can this bacterial pathogen cause such severe systemic infection in humans, we need to identify the bacterial factors associated with illness and even death of an infected animal, such as the mouse. A limiting factor for this research is that even the most up-to-date biochemical or genetic techniques can not cover all the components the bacterial cell produces. In 2003, the genome sequencing team lead by Professor Shih-Feng Tsai, in collaboration with my laboratory, completed the whole genome DNA sequence of a local V. vulnificus clinical strain, YJ016. This is the first whole genome sequence determined in Taiwan. Fig. 1 shows the features of V. vulnificus chromosomes deduced from the genome sequence by bioinformatic analyses, including the prediction of open-reading frames (ORFs, which are comparable to genes) and their possible functions by searching in the gene banks for those with similar DNA sequences. The results of this study revealed that, like V. cholerae and V. parahaemolyticus, the other two important Vibrio pathogens, V. vulnificus possesses two chromosomes; the big one contains about 3.4 megabase pairs and 3,300 ORFs and the small one contains about 1.85 megabase pairs and 1,700 ORFs. This strain also contains a plasmid, which was later shown by us to be dispensable for pathogenicity and drug resistance. By computational search, we further found the genes related to pathogenicity, such as capsule synthesis, iron acquisition and cytotoxins (Fig. 2). Later, a few local and overseas laboratories have identified potential virulence genes in this genome, and some of them have been experimentally confirmed as important virulence factors.
We have also compared the genome sequence of this bacterium with those of V. cholerae and V. parahaemolyticus to delineate the evolution of these bacterial pathogens. We found that these three Vibrio species evolved by active intra- and inter-chromosomal rearrangements and, compared to the large chromosomes, the small ones of these species show a higher degree of diversity. The genes associated with virulence are mostly located in the small chromosomes and possibly acquired from other organisms by the efficient gene transfer mechanisms of the bacteria. V. vulnificus appears to be more like V. parahaemolyticus than V. cholerae in terms of the genome size and gene organization.
On the other hand, the functions of about 34% of ORFs in this genome sequence are unknown; most of them are uniquely exist in V. vulnificus, meaning that many V. vulnificus-specific functions await further exploration, through which we could better understand the pathogenesis of diseases caused by this bacterial pathogen.
Fig. 1. The features of the chromosomes in V. vulnificus YJ016. Two circular chromosomes were derived from the genome DNA sequencing data. From outer to inner, circle 1 and circle 2: predicted ORFs with various functional categories shown by different colors; circle 3: local % (G + C) in relation to mean % (G + C) for the chromosome; circle 4: GC skew [(C-G)/(C+G)]; circle 5: super integron, type IV pilus genes and other pathogenicity-related genes; circle 6: tRNA genes; circle 7: rRNA genes.
Fig. 2. The rtx gene clusters in V. vulnificus YJ016. The gene or gene clusters similar to the rtx gene cluster of V. cholerae (VC1) were identified in the genome sequence of V. vulnificus YJ016 by bioinformatic analysis. Among them, gene cluster VVA1030-VVA1036 is most similar to the rtx gene cluster in V. cholerae. This gene cluster has been later confirmed experimentally to be closely associated with the virulence for mice.