Project description:The ability for bacteria to attach to and detach from various substrata is important for colonization, survival and transitioning to new environments. An opportunistic human pathogen, Vibrio vulnificus, can cause potentially fatal septicemia after ingestion of undercooked seafood. Based on genetic polymorphisms, strains of this species are subtyped into clinical (C) and environmental (E) genotypes. Vibrio vulnificus readily associates with chitin, thus we investigated chitin detachment dynamics in these disparate genotypes. We found that C-genotypes detach significantly more than E-genotypes after 24 hours in aerobic as well as anaerobic conditions. Furthermore, expression of genes involved in type IV pilin production was significantly downregulated in C-genotypes compared to E-genotypes, suggesting an importance in detachment. Interestingly, gbpA, a gene that has been shown to be important in host colonization in V. cholerae, was upregulated in the C-genotypes during detachment. Additionally, we found that C-genotypes detached to a greater extent, and produced more quorum-sensing (QS) autoinducer-2 molecules relative to E-genotypes, which suggests a role for QS in detachment. These findings suggest that for V. vulnificus, QS-mediated detachment may be a potential mechanism for transitioning into a human host for C-genotypes, while facilitating E-genotype maintenance in the estuarine environment.
Project description:Vibrio vulnificus multiply rapidly in host tissues under iron overloaded conditions. To understand the effects of iron in the physiology of this pathogen we performed a genome-wide transcriptional analysis of this bacterium growing under three different iron concentrations. V.vulnificus CMCP6 cells were grown under three different iron concentrations (TSBS + EDDA 50uM, TSBS and TSBS + FAC 250 ug/ml) and samples taken at log phase. Keywords: Response to the iron concentration of the media
Project description:Vibrio vulnificus causes rare but frequently fatal septicemia associated with raw oyster consumption by persons with underlying hepatic or immune system dysfunction. The virulence potential of environmental reservoirs appears widely distributed, because most strains are virulent in animal models; however, several investigations recently demonstrated genetic divergence among strains from clinical versus environmental origin at independent genetic loci. The present study used PCR to screen DNA polymorphisms in strains from environmental (n = 35) or clinical (n = 33) sources, and genomic relationships were determined by repetitive extragenic palindromic DNA PCR (rep-PCR) typing. Significant (P < 0.01) association was observed for typical "clinical" or "environmental" polymorphism profiles based on strain origin. Most oyster isolates (88%), including all of those with the "environmental" profile, also formed a single rep-PCR genogroup. Clinical isolates within this group did not have the typical "clinical" profile. On the other hand, clinical isolates with the typical polymorphism profile were distributed among multiple rep-PCR genogroups, demonstrating greater genetic diversity than was evident by profiling genetic polymorphisms. Wound isolates were genetically distinct from typical blood isolates by all assays. Strains from an outbreak of wound infections in Israel (biotype 3) were closely related to several U.S. strains by rep-PCR, indicating potential reservoirs of emerging disease. Strains genetically related to blood isolates appeared to be relatively rare in oysters, as only one had the "clinical" polymorphism profile or clustered by rep-PCR. However, this study was not an extensive survey, and more sampling using rep-PCR for sensitive genetic discrimination is needed to determine the virulence potential of environmental reservoirs.
Project description:Vibrio vulnificus multiply rapidly in host tissues under iron overloaded conditions. To understand the effects of iron in the physiology of this pathogen we performed a genome-wide transcriptional analysis of this bacterium growing under three different iron concentrations. V.vulnificus CMCP6 cells were grown under three different iron concentrations (TSBS + EDDA 50uM, TSBS and TSBS + FAC 250 ug/ml) and samples taken at log phase. Keywords: Response to the iron concentration of the media Strains were grown to an OD600nm of 0.6 to 0.8 in TSBS, TSBS with the addition of 250 μg/ml FAC or TSBS with the addition of 50 μM EDDA. Three independent cultures of the V. vulnificus cells grown in each media, were combined and treated as a single sample for the RNA extraction to minimize culture variation. Two samples per condition were used for the microarray analysis. Cells were centrifuged and the pellets resuspended in RNAWiz reagent (Ambion®, Austin, TX). Total RNA was extracted from each strain according to the manufacturerâ??s instructions
Project description:Vibrio vulnificus, a gram-negative pathogenic bacterium, transmitted via undercooked seafood or contaminated seawater, causes septicemia and wound infections. In this study, we analyzed 15 clinical and 11 environmental isolates. In total, 20 sequence types (STs), including eight novel STs, were identified. Antibiotic resistance gene analysis commonly detected the cyclic AMP receptor protein (CRP) in both the clinical and environmental isolates. Interestingly, clinical and environmental isolates were non-susceptible to third-generation cephalosporins, such as ceftazidime and cefotaxime, complicating the treatment of V. vulnificus infection. Multiple antibiotic resistance (MAR) index ranged from 0.1 to 0.5, with clinical isolates showing a higher mean MAR index than the environmental isolates, indicating their broader spectrum of resistance. Notable, no quantitative (124.3 vs. 126.5) and qualitative (adherence, antiphagocytosis, and chemotaxis/motility) differences in virulence factors were observed between the environmental and clinical strains. The molecular characteristics identified in this study provide insights into the virulence of V. vulnificus strains in South Korea, highlighting the need for continuous surveillance of antibiotic resistance in emerging V. vulnificus strains.
Project description:Vibrio vulnificus (V. vulnificus) is an opportunistic human pathogen known for causing various illnesses such as gastroenteritis, skin and muscle necrosis, septic shock, and sepsis. This halophilic estuarine bacterium's growth and infection process involves adaptation to both the natural briny environments and the host. OmpR, a response regulator in the EnvZ/OmpR two-component regulatory system (TCS), is crucial for environmental adaptation and pathogenicity. This study focused on investigating the impact of OmpR in V. vulnificus by creating an ompR knockout strain (ΔompR). The ΔompR strain exhibited reduced tolerance to alkaline stress, shorter flagella, and decreased virulence in epithelial cell and mouse models compared to the wild-type (WT) V. vulnificus. RNAseq analysis revealed the downregulation of genes involved in metabolism, flagellum-dependent motility, and transcription factors in the ΔompR strain. OmpR was found to repress the expression of aphB in alkaline conditions, impacting the acid resistance system CadBA, while also positively regulating the transcription of various flagellar genes. These findings suggest that OmpR acts as a global regulator, orchestrating the expression of multiple genes in response to different environments and during host invasion.
Project description:Vibrio vulnificus can be a highly invasive pathogen capable of spreading from an infection site to the bloodstream, causing sepsis and death. To survive and proliferate in blood, the pathogen requires mechanisms to overcome the innate immune defenses and metabolic limitations of this host niche. We created a high-density transposon mutant library in YJ016, a strain representative of the most virulent V. vulnificus lineage (or phylogroup) and used transposon insertion sequencing (TIS) screens to identify loci that enable the pathogen to survive and proliferate in human serum. Initially, genes underrepresented for insertions were used to estimate the V. vulnificus essential gene set; comparisons of these genes with similar TIS-based classification of underrepresented genes in other vibrios enabled the compilation of a common Vibrio essential gene set. Analysis of the relative abundance of insertion mutants in the library after exposure to serum suggested that genes involved in capsule biogenesis are critical for YJ016 complement resistance. Notably, homologues of two genes required for YJ016 serum-resistance and capsule biogenesis were not previously linked to capsule biogenesis and are largely absent from other V. vulnificus strains. The relative abundance of mutants after exposure to heat inactivated serum was compared with the findings from the serum screen. These comparisons suggest that in both conditions the pathogen relies on its Na+ transporting NADH-ubiquinone reductase (NQR) complex and type II secretion system to survive/proliferate within the metabolic constraints of serum. Collectively, our findings reveal the potency of comparative TIS screens to provide knowledge of how a pathogen overcomes the diverse limitations to growth imposed by serum.
Project description:IscR is a novel global regulator potentially contributing to the overall success in survival and pathogenesis of V. vulnificus by coordinating the regulation of various virulence factors. The profiles of transcripts from the V. vulnificus iscR mutant and the parental wild type were compared by using a V. vulnificus whole-genome microarray.
Project description:Vibrio vulnificus is a heterogeneous bacterial species that comprises virulent and avirulent strains from environmental and clinical sources that have been grouped into three biotypes. To validate the typing methods proposed to distinguish clinical from environmental isolates, we performed phenotypic (API 20E, API 20NE, and BIOLOG tests) and genetic (ribotyping and DNA polymorphism at several loci) studies with a large strain collection representing different biotypes, origins, and host ranges. No phenotypic method was useful for biotyping or grouping strains with regard to the origin of an isolate, and only the BIOLOG system was reliable for identifying the strains at the species level. DNA polymorphisms divided the population into three major profiles. Profile 1 strains were vcg type C, 16S rRNA type B, and vvh type 1 and included most of the biotype 1 human septicemic isolates; profile 2 strains were vcg type E, 16S rRNA type A, and vvh type 2 and included all biotype 2 isolates together with biotype 1 isolates from fish and water and some human isolates; and profile 3 strains were vcg type E, 16S rRNA type AB, and vvh type 2 and included biotype 3 strains. Ribotyping divided the species into two groups: one group that included profile 1 biotype 1 isolates and one group that included isolates of all three biotypes with the three profiles described above. In conclusion, no genotyping system was able to distinguish either clinical strains from environmental strains or biogroups within the species V. vulnificus, which suggests that new typing methodologies useful for public health have to be developed for this species.
Project description:In 1996 a common-source outbreak of severe soft tissue and bloodstream infections erupted among Israeli fish farmers and fish consumers due to changes in fish marketing policies. The causative pathogen was a new strain of Vibrio vulnificus, named biotype 3, which displayed a unique biochemical and genotypic profile. Initial observations suggested that the pathogen erupted as a result of genetic recombination between two distinct populations. We applied a whole genome shotgun sequencing approach using several V. vulnificus strains from Israel in order to study the pan genome of V. vulnificus and determine the phylogenetic relationship of biotype 3 with existing populations. The core genome of V. vulnificus based on 16 draft and complete genomes consisted of 3068 genes, representing between 59 and 78% of the whole genome of 16 strains. The accessory genome varied in size from 781 to 2044 kbp. Phylogenetic analysis based on whole, core, and accessory genomes displayed similar clustering patterns with two main clusters, clinical (C) and environmental (E), all biotype 3 strains formed a distinct group within the E cluster. Annotation of accessory genomic regions found in biotype 3 strains and absent from the core genome yielded 1732 genes, of which the vast majority encoded hypothetical proteins, phage-related proteins, and mobile element proteins. A total of 1916 proteins (including 713 hypothetical proteins) were present in all human pathogenic strains (both biotype 3 and non-biotype 3) and absent from the environmental strains. Clustering analysis of the non-hypothetical proteins revealed 148 protein clusters shared by all human pathogenic strains; these included transcriptional regulators, arylsulfatases, methyl-accepting chemotaxis proteins, acetyltransferases, GGDEF family proteins, transposases, type IV secretory system (T4SS) proteins, and integrases. Our study showed that V. vulnificus biotype 3 evolved from environmental populations and formed a genetically distinct group within the E-cluster. The unique epidemiological circumstances facilitated disease outbreak and brought this genotype to the attention of the scientific community.