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Letters in Applied Microbiology Jun 2014Vibrio vulnificus and Vibrio parahaemolyticus are Gram-negative marine bacteria naturally found in estuaries such as the Gulf of Mexico and can be pathogenic to humans....
UNLABELLED
Vibrio vulnificus and Vibrio parahaemolyticus are Gram-negative marine bacteria naturally found in estuaries such as the Gulf of Mexico and can be pathogenic to humans. We quantified both of these organisms in fish, oyster, sediment, and water using culture-independent (quantitative PCR; qPCR) and culture-dependent (direct plating-colony hybridization; DP-CH) techniques during the transition period between winter and spring. We correlated these levels to environmental conditions and to abundance of total bacteria and total vibrio. By qPCR, fish intestine samples had the highest V. vulnificus densities and were 2·7, 3·5, and 4·2 logs greater than oyster, sediment and water samples, respectively. Densities of V. parahaemolyticus in fish samples by qPCR were 1·0, 2·1, and 3·1 logs greater than oyster, sediment and water samples, respectively. Similar differences between sample types were also observed by DP-CH. The difference between the more favourable and less favourable environmental conditions identified in this study was small (mean salinity 4·3 vs. 13 ppt). However, V. vulnificus and V. parahaemolyticus were consistently detected in fish intestines, but infrequently detected and at lower levels in oysters and during the less favourable period. This trend was observed by qPCR and DP-CH, indicating fish intestines are a significant source of pathogenic vibrios in the environment.
SIGNIFICANCE AND IMPACT OF THE STUDY
This is the first study to report the densities of Bacteria 16S rRNA, Vibrio 16S rRNA, Vibrio vulnificus, and V. parahaemolyticus in fish intestine, oyster, sediment and water samples, as well as compare these values through culture-dependent and culture-independent methodology. Vibrio vulnificus and V. parahaemolyticus were detected in samples of fish intestines by qPCR and colony hybridization when conditions were less favourable for their occurrence in the environment. In contrast, V. vulnificus and V. parahaemolyticus were infrequently detected and at lower levels in other niches examined. These results indicate that fish intestinal tracts are a significant source of these pathogens.
Topics: Animals; Fishes; Gastrointestinal Tract; Geologic Sediments; Gulf of Mexico; Humans; Molecular Typing; Ostreidae; RNA, Bacterial; RNA, Ribosomal, 16S; Rivers; Salinity; Seasons; Shellfish; Vibrio parahaemolyticus; Vibrio vulnificus; Virulence; Water Microbiology
PubMed: 24571291
DOI: 10.1111/lam.12226 -
FEMS Microbiology Letters Apr 2015Vibrio vulnificus causes rapid disseminating septicemia by oral infection in infected individuals who have an underlying disease, especially chronic liver diseases....
Vibrio vulnificus causes rapid disseminating septicemia by oral infection in infected individuals who have an underlying disease, especially chronic liver diseases. Although the elucidation of specific risk factors for V. vulnificus infection in patients with liver diseases is of urgent importance, no appropriate experimental animal model that mimics the liver diseases in this bacterial infection has been available so far. To discover these risk factors, we generated a liver disordered mouse by performing bile duct ligation (BDL). Hepatitis developed in the BDL mice; however, this did not affect mortality in mice after orogastric administration of V. vulnificus, suggesting that the liver disorders caused by the BDL were not risk factors for V. vulnificus septicemia. When the dead and surviving mice were compared, V. vulnificus could be detected from the spleen only in the dead group. Furthermore, significantly higher numbers of V. vulnificus were detected from the intestines in the dead group than in the surviving group ( P < 0.001). These findings suggested that proliferation of the challenge inoculum in the intestine was needed for the oral infection with V. vulnificus, and that the elimination of V. vulnificus in the liver and/or spleen plays a critical role in survival of the host.
Topics: Animals; Bile Ducts; Disease Models, Animal; Hepatitis; Intestines; Liver; Mice; Mouth; Risk Factors; Spleen; Vibrio Infections; Vibrio vulnificus
PubMed: 25790509
DOI: 10.1093/femsle/fnv005 -
The Journal of Veterinary Medical... Jul 2015Vibrio vulnificus is the causative agent of primary septicemia, wound infection and gastroenteritis in immunocompromised people. In this study, signature-tagged...
Vibrio vulnificus is the causative agent of primary septicemia, wound infection and gastroenteritis in immunocompromised people. In this study, signature-tagged mutagenesis (STM) was applied to identify the virulence genes of V. vulnificus. Using STM, 6,480 mutants in total were constructed and divided into 81 sets (INPUT pools); each mutant in a set was assigned a different tag. Each INPUT pool was intraperitoneally injected into iron-overloaded mice, and in vivo surviving mutants were collected from blood samples from the heart (OUTPUT pools). From the genomic DNA of mixed INPUT or OUTPUT pools, digoxigenin-labeled DNA probes against the tagged region were prepared and used for dot hybridization. Thirty tentatively attenuated mutants, which were hybridized clearly with INPUT probes but barely with OUTPUT probes, were negatively selected. Lethal doses of 11 of the 30 mutants were reduced to more than 1/100; of these, the lethal doses of 2 were reduced to as low as 1/100,000. Transposon-inserted genes in the 11 attenuated mutants were those for IMP dehydrogenase, UDP-N-acetylglucosamine-2-epimerase, aspartokinase, phosphoribosylformylglycinamidine cyclo-ligase, malate Na (+) symporter and hypothetical protein. When mice were immunized with an attenuated mutant strain into which IMP dehydrogenase had been inserted with a transposon, they were protected against V. vulnificus infection. In this study, we demonstrated that the STM method can be used to search for the virulence genes of V. vulnificus.
Topics: Animals; DNA Probes; Female; Genes, Bacterial; Mice; Mice, Inbred ICR; Mutagenesis, Insertional; Vibrio vulnificus; Virulence Factors
PubMed: 25755021
DOI: 10.1292/jvms.14-0655 -
BMC Microbiology Oct 2022A visual, rapid, simple method was developed based on a loop-mediated isothermal amplification (LAMP) assay to detect Vibrio vulnificus in aquatic products and...
BACKGROUND
A visual, rapid, simple method was developed based on a loop-mediated isothermal amplification (LAMP) assay to detect Vibrio vulnificus in aquatic products and aquaculture waters.
RESULTS
Genomic DNA was extracted from Vibrio vulnificus using the boiling method, and optimized primers were used to detect the gyrB gene using a visual LAMP method. The sensitivity of the assay was 10 fg/μL, and the obtained results were stable and reliable. Out of 655 aquatic product samples and 558 aquaculture water samples, the positive rates of Vibrio vulnificus detection were 9.01% and 8.60%, respectively, which are markedly higher than those of the traditional culture identification methods.
CONCLUSION
The relatively simple technical requirements, low equipment cost, and rapid detection make the visual LAMP method for the detection of Vibrio vulnificus a convenient choice for field detection in the aquaculture industry.
Topics: Vibrio vulnificus; Water; Sensitivity and Specificity; Nucleic Acid Amplification Techniques
PubMed: 36271365
DOI: 10.1186/s12866-022-02656-1 -
BMC Microbiology Mar 2020Vibrio vulnificus hemolysin (VVH) is a pore-forming toxin secreted by Vibrio vulnificus. Cellular cholesterol was believed to be the receptor for VVH, because...
BACKGROUND
Vibrio vulnificus hemolysin (VVH) is a pore-forming toxin secreted by Vibrio vulnificus. Cellular cholesterol was believed to be the receptor for VVH, because cholesterol could bind to VVH and preincubation with cholesterol inhibited cytotoxicity. It has been reported that specific glycans such as N-acetyl-D-galactosamine and N-acetyl-D-lactosamine bind to VVH, however, it has not been known whether these glycans could inhibit the cytotoxicity of VVH without oligomer formation. Thus, to date, binding mechanisms of VVH to cellular membrane, including specific receptors have not been elucidated.
RESULTS
We show here that VVH associates with ganglioside GM1a, Fucosyl-GM1, GD1a, GT1c, and GD1b by glycan array. Among them, GM1a could pulldown VVH. Moreover, the GD1a inhibited the cytotoxicity of VVH without the formation of oligomers.
CONCLUSION
This is the first report of a molecule able to inhibit the binding of VVH to target cells without oligomerization of VVH.
Topics: Animals; Bacterial Proteins; Binding Sites; CHO Cells; Cell Membrane; Cholesterol; Cricetulus; Gangliosides; Glycomics; Hemolysin Proteins; Microarray Analysis; Protein Binding; Protein Conformation; Protein Multimerization; Vibrio vulnificus
PubMed: 32228455
DOI: 10.1186/s12866-020-01755-1 -
MicrobiologyOpen Sep 2020Vibrio vulnificus is the leading cause of seafood-associated deaths worldwide. Despite the growing knowledge about the population structure of V. vulnificus, the...
Vibrio vulnificus is the leading cause of seafood-associated deaths worldwide. Despite the growing knowledge about the population structure of V. vulnificus, the evolutionary history and the ancestral relationships of strains isolated from various regions around the world have not been determined. Using the largest collection of sequence and isolate data of V. vulnificus to date, we applied ancestral character reconstruction to study the phylogeography of V. vulnificus. Multilocus sequence typing data from 10 housekeeping genes were used for the inference of ancestral states and reconstruction of the evolutionary history. The findings showed that the common ancestor of all V. vulnificus populations originated from East Asia, and later evolved into two main clusters that spread with time and eventually evolved into distinct populations in different parts of the world. While we found no meaningful insights concerning the evolution of V. vulnificus populations in the Middle East; however, we were able to reconstruct the ancestral scenarios of its evolution in East Asia, North America, and Western Europe.
Topics: Animals; Biological Evolution; Europe; Asia, Eastern; Fishes; Geologic Sediments; Humans; Multilocus Sequence Typing; Phylogeny; Phylogeography; Seawater; Shellfish; Spatio-Temporal Analysis; Vibrio Infections; Vibrio vulnificus
PubMed: 32779403
DOI: 10.1002/mbo3.1103 -
Diseases of Aquatic Organisms Apr 2014Vibrio vulnificus is a potentially zoonotic bacterial pathogen of fish, which can infect humans (causing necrotic fasciitis). We analysed 24 V. vulnificus isolates (from...
Vibrio vulnificus is a potentially zoonotic bacterial pathogen of fish, which can infect humans (causing necrotic fasciitis). We analysed 24 V. vulnificus isolates (from 23 severe eel disease outbreaks in 8 Dutch eel farms during 1996 to 2009, and 1 clinical strain from an eel farmer) for genetic correlation and zoonotic potential. Strains were typed using biotyping and molecular typing by high-throughput multilocus sequence typing (hiMLST) and REP-PCR (Diversilab®). We identified 19 strains of biotype 1 and 5 of biotype 2 (4 from eels, 1 from the eel farmer), that were subdivided into 8 MLST types (ST) according to the international standard method. This is the first report of V. vulnificus biotype 1 outbreaks in Dutch eel farms. Seven of the 8 STs, of unknown zoonotic potential, were newly identified and were deposited in the MLST database. The REP-PCR and the MLST were highly concordant, indicating that the REP-PCR is a useful alternative for MLST. The strains isolated from the farmer and his eels were ST 112, a known potential zoonotic strain. Antimicrobial resistance to cefoxitin was found in most of the V. vulnificus strains, and an increasing resistance to quinolones, trimethoprim + sulphonamide and tetracycline was found over time in strain ST 140. Virulence testing of isolates from diseased eels is recommended, and medical practitioners should be informed about the potential risk of zoonotic infections by V. vulnificus from eels for the prevention of infection especially among high-risk individuals. Additional use of molecular typing methods such as hiMLST and Diversilab® is recommended for epidemiological purposes during V. vulnificus outbreaks.
Topics: Anguilla; Animals; Anti-Bacterial Agents; Aquaculture; Disease Outbreaks; Drug Resistance, Bacterial; Fish Diseases; Genetic Variation; Netherlands; Vibrio Infections; Vibrio vulnificus
PubMed: 24695233
DOI: 10.3354/dao02703 -
Infection and Immunity Apr 2008Numerous secreted virulence factors have been proposed to account for the fulminating and destructive nature of Vibrio vulnificus infections. A mutant of V. vulnificus...
Numerous secreted virulence factors have been proposed to account for the fulminating and destructive nature of Vibrio vulnificus infections. A mutant of V. vulnificus that exhibited less cytotoxicity to INT-407 human intestinal epithelial cells was screened from a library of mutants constructed by random transposon mutagenesis. A transposon-tagging method was used to identify and clone an open reading frame encoding an RTX toxin secretion ATP binding protein, RtxE, from V. vulnificus. The deduced amino acid sequence of RtxE from V. vulnificus was 91% identical to that reported from Vibrio cholerae. Functions of the rtxE gene in virulence were assessed by constructing an isogenic mutant whose rtxE gene was inactivated by allelic exchanges and by evaluating the differences between its virulence phenotype and that of the wild type in vitro and in mice. The disruption of rtxE blocked secretion of RtxA to the cell exterior and resulted in a significant reduction in cytotoxic activity against epithelial cells in vitro. Also, the intraperitoneal 50% lethal dose of the rtxE mutant was 10(4) to 10(5) times higher than that of the parental wild type, indicating that RtxE is essential for the virulence of V. vulnificus. Furthermore, the present study demonstrated that the rtxBDE genes are transcribed as one transcriptional unit under the control of a single promoter, P(rtxBDE). The activity of V. vulnificus P(rtxBDE) is induced by exposure to INT-407 cells, and the induction requires direct contact of the bacteria with the host cells.
Topics: Animals; Cell Line; Epithelial Cells; Gene Expression Regulation, Bacterial; Humans; Iron, Dietary; Mice; Mutation; Transcription, Genetic; Vibrio Infections; Vibrio vulnificus; Virulence; Virulence Factors
PubMed: 18250174
DOI: 10.1128/IAI.01503-07 -
Applied and Environmental Microbiology Nov 2020Oyster and seawater samples were collected from five sites in the Chesapeake Bay, MD, and three sites in the Delaware Bay, DE, from May to October 2016 and 2017....
Seasonal and Geographical Differences in Total and Pathogenic Vibrio parahaemolyticus and Vibrio vulnificus Levels in Seawater and Oysters from the Delaware and Chesapeake Bays Determined Using Several Methods.
Oyster and seawater samples were collected from five sites in the Chesapeake Bay, MD, and three sites in the Delaware Bay, DE, from May to October 2016 and 2017. Abundances and detection frequencies for total and pathogenic and were compared using the standard most-probable-number-PCR (MPN-PCR) assay and a direct-plating (DP) method on CHROMagar Vibrio for total ( ) and pathogenic ( and ) genes and total () and pathogenic () genes. The colony overlay procedure for peptidases (COPP) assay was evaluated for total DP had high false-negative rates (14 to 77%) for most PCR targets and was deemed unsatisfactory. Logistic regression models of the COPP assay showed high concordances with MPN-PCR for and and in oysters (85.7 to 90.9%) and seawater (81.1 to 92.7%) when seawater temperature and salinity were factored into the model, suggesting that the COPP assay could potentially serve as a more rapid method to detect vibrios in oysters and seawater. Differences in total and pathogenic abundances between state sampling sites over different collection years were contrasted for oysters and seawater by MPN-PCR. Abundances of and were ∼8-fold higher in Delaware oysters than in Maryland oysters, whereas abundances of were nearly identical. For Delaware oysters, 93.5% were both and , compared to only 19.2% in Maryland. These results indicate that pathogenic was more prevalent in the Delaware Bay than in the Chesapeake Bay. While and cause shellfish-associated morbidity and mortality among shellfish consumers, current regulatory assays for vibrios are complex, time-consuming, labor-intensive, and relatively expensive. In this study, the rapid, simple, and inexpensive COPP assay was identified as a possible alternative to MPN-PCR for shellfish monitoring. This paper shows differences in total and pathogenic vibrios found in seawater and oysters from the commercially important Delaware and Chesapeake Bays. isolates from the Delaware Bay were more likely to contain commonly recognized pathogenicity genes than those from the Chesapeake Bay.
Topics: Animals; Bays; Colony Count, Microbial; Delaware; Geography; Maryland; Ostreidae; Seasons; Seawater; Vibrio parahaemolyticus; Vibrio vulnificus
PubMed: 32978135
DOI: 10.1128/AEM.01581-20 -
Journal of Bacteriology Nov 2020Pathogenic species use many different approaches to subvert, attack, and undermine the host response. The toxins they produce are often responsible for the devastating... (Review)
Review
Pathogenic species use many different approaches to subvert, attack, and undermine the host response. The toxins they produce are often responsible for the devastating effects associated with their diseases. These toxins target a variety of host proteins, which leads to deleterious effects, including dissolution of cell organelle integrity and inhibition of protein secretion. Becoming increasingly prevalent as cofactors for toxins are proteins of the small GTPase families. ADP-ribosylation factor small GTPases (ARFs) in particular are emerging as a common host cofactor necessary for full activation of toxins. While ARFs are not the direct target of cholera toxin (CT), ARF binding is required for its optimal activity as an ADP-ribosyltransferase. The makes caterpillars floppy (MCF)-like and the domain X (DmX) effectors of the multifunctional autoprocessing repeats-in-toxin (MARTX) toxin also both require ARFs to initiate autoprocessing and activation as independent effectors. ARFs are ubiquitously expressed in eukaryotes and are key regulators of many cellular processes, and as such they are ideal cofactors for pathogens that infect many host species. In this review, we cover in detail the known toxins that use ARFs as cross-kingdom activators to both stimulate and optimize their activity. We further discuss how these contrast to toxins and effectors from other bacterial species that coactivate, stimulate, or directly modify host ARFs as their mechanisms of action.
Topics: ADP-Ribosylation Factors; Animals; Bacterial Toxins; Host-Pathogen Interactions; Humans; Multigene Family; Vibrio Infections; Vibrio vulnificus
PubMed: 32900828
DOI: 10.1128/JB.00278-20