טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
Ph.D Thesis
Ph.D StudentEfimov Vera
SubjectVirulence Characterization of the Human Pathogen
Vibrio Vulnificus
DepartmentDepartment of Biotechnology and Food Engineering
Supervisor Professor Yechezkel Kashi
Full Thesis textFull thesis text - English Version


Abstract

Vibrio vulnificus is an aquatic bacterium and an important human pathogen, classified into three biotypes. Human infections are acquired through skin wounds or consumption of contaminated seafood, leading to necrosis, gastroenteritis and septicemia, with high fatality rates. The newly emerged biotype 3 appears to be rather clonal and geographically restricted to Israel, where it caused an outbreak of wound infections. The main aim of this work was to characterize virulence mechanisms of the Israeli group of V. vulnificus, biotype 3, compared to biotype 1, known to have a worldwide distribution. To understand the evolution of the bacterium's genome, we sequenced and analyzed the genome of biotype 3 strain VVyb1(BT3), and then conducted a microbial environmental survey of the hypothesized niche from which it probably evolved. The genome of this environmental isolate revealed higher similarity to the published biotype 1 genomes of clinical strains (90%) than to the environmental strains (87%), supporting the virulence of the biotype 3 group. Genotyping and genome sequence analysis indicated that biotype 3's genome was probably created based on the core genome of biotype 1 strain that horizontally gained a rather small number of genes from its natural environment. Biotype 3 was found to possess high virulence level in mice, similar to that of biotype 1 that originated from the same environmental niche in Israel, supporting their evolutionary relatedness. Moreover, the estimated lethal dose of the studied strains (~500 CFU) were three orders of magnitude lower than of the previously published biotype 1 clinical strains (p-value<0.5). Total transcriptome profiles of V. vulnificus biotype 3 and its possible ancestor biotype 1- clade B were studied during mice infection in comparison to logarithmic growth in culture. Results showed significant differences in the expression levels of over 1000 genes (~20% of the genome) between mice infection and culture growth, as well as between the two biotypes. Furthermore, results suggest biotype-specific virulence strategies. Interestingly, biotype 3 was found to express a large number of virulent genes during logarithmic growth, including a large cluster of motility and flagella associated genes. The higher motility of biotype 3 strain compared to biotype 1 strains (p-value<0.5) probably provides it an advantage during initial host invasion. This may explain the predominance of biotype 3 over biotype 1 in infectious cases in Israel. Finally, results of whole-transcriptome analysis showed high variation between the two biotypes, suggesting that changes in gene expression might also be the cause of the biochemical switch that led to the formation of new biotype - biotype 3. This multidisciplinary project that encompasses state-of-the-art methodologies in microbiology, bioinformatics, genomics and transcriptomics gives us a broader understanding of the unresolved evolution of V. vulnificus.