|Ph.D Student||Raz Nili|
|Subject||Development of Microarray Technology Based on Comparative|
Genomics for Typing of the Human Pathogen
|Department||Department of Biotechnology and Food Engineering||Supervisor||Professor Yechezkel Kashi|
|Full Thesis text|
Vibrio vulnificus is a halophile pathogenic bacterium, naturally found in marine and estuarine environments. Human infections are acquired through consumption of contaminated seafood or through skin wounds. Strains of V. vulnificus are classified into three different biotypes. The emergence of the recently identified biotype 3 in Israel is considered to be the result of genome hybridization of two different bacterial populations. As a new emerging pathogen group, it forms a genetically distinct and homogeneous clone. In the family Vibrionaceae, horizontal gene transfer is relatively frequent and responsible for the emergence of new pathogen groups. Our recent genomic study suggests the existence of an additional new virulent group of V. vulnificus, termed clade A.
In order to understand and follow the rapid evolution of the bacterium, we have developed a custom genome-wide SNP genotyping array implemented on the Illumina GoldenGate platform. We constructed a SNP-variation database by two approaches: (i) comparative in-silico data mining using an in-house genome-wide sequence comparison computer program to mine sequence variation of genes from the two available V. vulnificus genome sequences; (ii) in-vitro sequence analysis of selected genes among representative strains. A total of 1,071 SNPs were validated, of which the top-ranked 576 SNPs were selected for genotyping from 517 loci presenting both high and low mutation rates. 254 clinical and environmental V. vulnificus isolates with worldwide distribution recovered over a 30-year period from a wide variety of hosts and habitats were successfully genotyped by the array. This powerful high-throughput approach enabled us to simultaneously cover 576 SNPs randomly distributed throughout the genome of V. vulnificus among a diverse and large number of isolates.
Our data demonstrate that the selected SNPs are a ready source of high variation content, which could be utilized for accurate strain identification and for inferring phylogenetic relationships among strains, following in a detailed way the origin of genomic segments along the bacterial chromosome. Analyses of the array data together with our recently draft genome sequence of biotype 3 strain VVyb1(BT3), support the mosaic genome hypothesis, as well as the recent emergence of new pathogenic groups (clade A) within this species as a recurrent phenomenon. The results specifically contribute to the broad understanding of the evolution of this human pathogen. Moreover, the technology provides an important tool for haplotype analysis of bacterial genomes toward the development of the bacterial HapMaps database.