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Microbiology (Reading, England) Dec 2020Rifampicin is a broad-spectrum antibiotic that binds to the bacterial RNA polymerase (RNAP), compromising DNA transcription. Rifampicin resistance is common in several...
Rifampicin is a broad-spectrum antibiotic that binds to the bacterial RNA polymerase (RNAP), compromising DNA transcription. Rifampicin resistance is common in several microorganisms and it is typically caused by point mutations in the gene encoding the β subunit of RNA polymerase, . Different mutations are responsible for various levels of rifampicin resistance and for a range of secondary effects. mutations conferring rifampicin resistance have been shown to be responsible for severe effects on transcription, cell fitness, bacterial stress response and virulence. Such effects have never been investigated in the marine pathogen , even though rifampicin-resistant strains of have been isolated previously. Moreover, spontaneous rifampicin-resistant strains of have an important role in conjugation and mutagenesis protocols, with poor consideration of the effects of mutations. In this work, effects on growth, stress response and virulence of were investigated using a set of nine spontaneous rifampicin-resistant derivatives of CMCP6. Three different mutations (Q513K, S522L and H526Y) were identified with varying incidence rates. These three mutant types each showed high resistance to rifampicin [minimal inhibitory concentration (MIC) >800 µg ml], but different secondary effects. The strains carrying the mutation H526Y had a growth advantage in rich medium but had severely reduced salt stress tolerance in the presence of high NaCl concentrations as well as a significant reduction in ethanol stress resistance. Strains possessing the S522L mutation had reduced growth rate and overall biomass accumulation in rich medium. Furthermore, investigation of virulence characteristics demonstrated that all the rifampicin-resistant strains showed compromised motility when compared with the wild-type, but no major effects on exoenzyme production were observed. These findings reveal a wide range of secondary effects of mutations and indicate that rifampicin resistance is not an appropriate selectable marker for studies that aim to investigate phenotypic behaviour in this organism.
Topics: Anti-Bacterial Agents; Bacterial Proteins; DNA-Directed RNA Polymerases; Drug Resistance, Bacterial; Genetic Fitness; Locomotion; Microbial Sensitivity Tests; Microbial Viability; Mutation; Rifampin; Stress, Physiological; Vibrio vulnificus
PubMed: 33186092
DOI: 10.1099/mic.0.000991 -
Biosensors May 2021and are two most reported foodborne pathogens related to seafood. Due to global ocean warming and an increase in seafood consumption worldwide, foodborne illnesses...
and are two most reported foodborne pathogens related to seafood. Due to global ocean warming and an increase in seafood consumption worldwide, foodborne illnesses related to infection of these two bacteria are growing, leading to food safety issues and economic consequences. Molecular detection methods targeting species-specific genes are effective tools in the fight against bacterial infections for food safety. In this study, a duplex detection biosensor based on isothermal recombinase polymerase amplification (RPA) and a three-segment lateral flow strip (LFS) has been established. The biosensor used gene of and gene of as the detection markers based on previous reports. A duplex RPA reaction for both targets were constructed, and two chemical labels, FITC and DIG, of the amplification products were carefully tested for effective and accurate visualization on the strip. The biosensor demonstrated good specificity and achieved a sensitivity of 10 copies per reaction or one colony forming unit (CFU)/10 g of spiked food for both bacteria. Validation with clinical samples showed results consistent with that of real-time polymerase chain reaction. The detection process was simple and fast with a 30-min reaction at 37 °C and visualization on the strip within 5 min. With little dependence on laboratory settings, this biosensor was suitable for on-site detection, and the duplex system enabled simultaneous detection of the two important foodborne bacteria. Moreover, the principle can be extended to healthcare and food safety applications for other pathogens.
Topics: Food Microbiology; Nucleic Acid Amplification Techniques; Real-Time Polymerase Chain Reaction; Recombinases; Sensitivity and Specificity; Vibrio cholerae; Vibrio vulnificus
PubMed: 34066017
DOI: 10.3390/bios11050151 -
Journal of Food Protection Dec 2021Desiccation is a routine farming practice used in off-bottom oyster aquaculture to reduce biofouling organisms and improve shell quality. This practice can increase...
ABSTRACT
Desiccation is a routine farming practice used in off-bottom oyster aquaculture to reduce biofouling organisms and improve shell quality. This practice can increase Vibrio parahaemolyticus and Vibrio vulnificus levels, leading to increased risk of illness for raw oyster consumers. Previous resubmersion studies were performed in geographic proximity to one another, so to better understand the broader applicability of resubmersion, the next step was to perform concurrent studies in multiple geographic locations within a region. This study evaluated the effect of variations in geographic location on the recovery time needed for elevated vibrio levels to return to ambient levels in desiccated oysters after resubmersion at Gulf Coast farms. Two trials were performed between May and August 2019 at sites spanning ∼100 km: three in Alabama and one in Florida. Oysters were deployed in OysterGro cages at each location, 2 weeks before each trial, and then either were desiccated for 24 h or remained submersed as controls. Triplicate samples were taken before and immediately following the desiccation period, as well as 7 and 14 days after resubmersion. Total and pathogenic V. parahaemolyticus and V. vulnificus levels were determined using most-probable-number (MPN) real-time PCR. Vibrio levels increased by 0.23 to 3.50 log MPN/g after desiccation. Recovery times varied among geographic locations by trial and Vibrio spp., with all vibrio counts recovering to levels not significantly higher than those in control oysters within 7 to 14 days of resubmersion (P ≥ 0.06). These results suggest a 14-day resubmersion period of cultured oysters allowed vibrio levels, elevated because of routine handling, to return to ambient levels at all farm sites studied.
Topics: Animals; Desiccation; Food Contamination; Gulf of Mexico; Ostreidae; Vibrio parahaemolyticus; Vibrio vulnificus
PubMed: 34383923
DOI: 10.4315/JFP-21-189 -
Frontiers in Cellular and Infection... 2021RtxA1 is a major cytotoxin of () causing fatal septicemia and necrotic wound infections. Our previous work has shown that RpoS regulates the expression and secretion of...
RtxA1 is a major cytotoxin of () causing fatal septicemia and necrotic wound infections. Our previous work has shown that RpoS regulates the expression and secretion of RtxA1 toxin. This study was conducted to further investigate the potential mechanisms of RpoS on RtxA1 secretion. First, TolCV1 and TolCV2 proteins, two TolC homologs, were measured at various time points by Western blotting. The expression of TolCV1 was increased time-dependently, whereas that of TolCV2 was decreased. Expression of both TolCV1 and TolCV2 was significantly downregulated in an deletion mutation. Subsequently, we explored the roles of TolCV1 and TolCV2 in pathogenesis. Western blot analysis showed that RtxA1 toxin was exported by TolCV1, not TolCV2, which was consistent with the cytotoxicity results. Furthermore, the expression of TolCV1 and TolCV2 was increased after treatment of the host signal bile salt and the growth of mutant was totally abolished in the presence of bile salt. A mutation resulted in significant reduction of induced-virulence in mice. Taken together, TolCV1 plays key roles in RtxA1 secretion, bile salt resistance, and mice lethality of , suggesting that TolCV1 could be an attractive target for the design of new medicines to treat infections.
Topics: Animals; Bacterial Proteins; Escherichia coli; Mice; Vibrio Infections; Vibrio vulnificus; Virulence
PubMed: 33996641
DOI: 10.3389/fcimb.2021.673222 -
Japanese Journal of Infectious Diseases Nov 2021Vibrio vulnificus (V. vulnificus) infection is rare but potentially fatal. This study explored new atypical manifestations and prognostic factors of V....
Vibrio vulnificus (V. vulnificus) infection is rare but potentially fatal. This study explored new atypical manifestations and prognostic factors of V. vulnificus-infected patients during hospitalization. We retrospectively reviewed the medical records of 33 patients diagnosed with V. vulnificus infection in Guangdong Province, China between 2010 and 2020. Multiple logistic regression and receiver operating characteristic (ROC) curve analyses were performed. The new atypical manifestations included cholangitis, urinary tract infection, and suppurative otitis media. Eleven of the 33 (33.3%) V. vulnificus-infected patients eventually died. Univariate analysis showed that patients with cardio-cerebrovascular diseases, lower platelet counts, and higher levels of C-reactive protein and procalcitonin (PCT) had statistically higher mortality. However, multivariate analysis showed that only the PCT level (P = 0.036) was statistically significant. In addition, the area under the ROC value estimate for PCT was 0.8816 (95% confidence interval (CI), 0.759-1.000; P = 0.0009). More than half of the patients with V. vulnificus infection died when PCT was > 20 ng/mL, while no patient died when PCT was ≤ 20 ng/mL. This study found new atypical manifestations of V. vulnificus infection. In addition, PCT was an effective and independent predictor of mortality in patients with V. vulnificus infection, allowing clinicians to conduct early risk stratification and determine the best therapeutic strategies.
Topics: Aged; Anti-Bacterial Agents; Female; Humans; Male; Middle Aged; Prognosis; ROC Curve; Retrospective Studies; Tertiary Care Centers; Vibrio Infections; Vibrio vulnificus
PubMed: 33952769
DOI: 10.7883/yoken.JJID.2020.843 -
Journal of Applied Microbiology Nov 2014To construct statistical models to predict the presence, abundance and potential virulence of Vibrio vulnificus in surface waters of Chesapeake Bay for implementation in...
AIM
To construct statistical models to predict the presence, abundance and potential virulence of Vibrio vulnificus in surface waters of Chesapeake Bay for implementation in ecological forecasting systems.
METHODS AND RESULTS
We evaluated and applied previously published qPCR assays to water samples (n = 1636) collected from Chesapeake Bay from 2007-2010 in conjunction with State water quality monitoring programmes. A variety of statistical techniques were used in concert to identify water quality parameters associated with V. vulnificus presence, abundance and virulence markers in the interest of developing strong predictive models for use in regional oceanographic modeling systems. A suite of models are provided to represent the best model fit and alternatives using environmental variables that allow them to be put to immediate use in current ecological forecasting efforts.
CONCLUSIONS
Environmental parameters such as temperature, salinity and turbidity are capable of accurately predicting abundance and distribution of V. vulnificus in Chesapeake Bay. Forcing these empirical models with output from ocean modeling systems allows for spatially explicit forecasts for up to 48 h in the future.
SIGNIFICANCE AND IMPACT OF THE STUDY
This study uses one of the largest data sets compiled to model Vibrio in an estuary, enhances our understanding of environmental correlates with abundance, distribution and presence of potentially virulent strains and offers a method to forecast these pathogens that may be replicated in other regions.
Topics: Bays; Forecasting; Models, Statistical; Salinity; Temperature; Vibrio vulnificus; Virulence Factors; Water Microbiology
PubMed: 25139334
DOI: 10.1111/jam.12624 -
Microbiology and Immunology Sep 2020Vibrio vulnificus is a foodborne pathogen causing septicemia with high mortality rate. In this study, we explored how Escherichia coli, one of the commensal bacteria in...
Vibrio vulnificus is a foodborne pathogen causing septicemia with high mortality rate. In this study, we explored how Escherichia coli, one of the commensal bacteria in the human gastrointestinal tract, can interact with V. vulnificus. Our study results show that the amount of biofilm produced by V. vulnificus was reduced in the presence of E. coli ATCC 35218, although the growth of V. vulnificus L-180 remained unaffected. We also detected an antibiofilm effect of E. coli culture supernatant against V. vulnificus, which could not be reduced even after heat treatment. These findings indicate that E. coli and its culture supernatant may be suitable to prevent biofilm formation by V. vulnificus. By contrast, live cells of V. vulnificus could reduce the amount of preformed E. coli biofilm, but its culture supernatant could not. This suggests that the cell-associated factors contribute toward reduction in E. coli biofilm. Therefore, we speculate that ingestion of an infectious dose of V. vulnificus might induce dislodging of the commensal bacteria from the intestinal epithelia and thus can colonize to initiate the infection.
Topics: Biofilms; Culture Media, Conditioned; Escherichia coli; Microbial Interactions; Microbial Viability; Vibrio vulnificus
PubMed: 32603487
DOI: 10.1111/1348-0421.12829 -
Journal of AOAC International Sep 2023The Thermo Scientific™ SureTect™ Vibrio cholerae, Vibrio parahaemolyticus, and Vibrio vulnificus PCR Assay method is a real-time PCR method for the multiplex...
Validation of the Thermo Scientific™ SureTect™ Vibrio cholerae, Vibrio parahaemolyticus, and Vibrio vulnificus PCR Assay for the Detection of Vibrio cholerae, Vibrio parahaemolyticus, and Vibrio vulnificus in Seafood Matrixes: AOAC Performance Tested MethodsSM 022301.
BACKGROUND
The Thermo Scientific™ SureTect™ Vibrio cholerae, Vibrio parahaemolyticus, and Vibrio vulnificus PCR Assay method is a real-time PCR method for the multiplex detection of Vibrio cholerae, Vibrio parahaemolyticus, and Vibrio vulnificus in seafood.
OBJECTIVE
The Thermo Scientific SureTect Vibrio cholerae, Vibrio parahaemolyticus, and Vibrio vulnificus Assay was evaluated for AOAC Performance Tested MethodsSM certification.
METHOD
Inclusivity/exclusivity, matrix, product consistency/stability, and robustness studies were conducted to assess the method's performance. For the matrix study, the method was validated using the Applied Biosystems™ QuantStudio™ 5 Real-Time PCR Food Safety Instrument and the Applied Biosystems™ 7500 Fast Real-Time PCR Food Safety Instrument against the U.S. Food and Drug Administration Bacteriological Analytical Manual, Chapter 9 (2004), Vibrio and ISO 21872-1:2017 Microbiology of the food chain-Horizontal method for the determination of Vibrio spp.-Part 1: Detection of potentially enteropathogenic Vibrio parahaemolyticus, Vibrio cholerae, and Vibrio vulnificus reference methods.
RESULTS
Matrix studies showed equivalent or superior performance of the candidate method compared to the reference method and, overall, no difference between presumptive and confirmed results, except for one matrix due to high background flora. The inclusivity/exclusivity study correctly identified/excluded all strains analyzed. Robustness testing showed no statistically significant differences in assay performance under varied test conditions. Product consistency and stability studies demonstrated no statistically significant differences between assay lots with different expiration dates.
CONCLUSIONS
The data presented show that the assay constitutes a rapid and reliable workflow for the detection of V. cholerae, V. parahaemolyticus, and V. vulnificus in seafood matrixes.
HIGHLIGHTS
The SureTect PCR Assay method allows for fast, reliable detection of stipulated strains in seafood matrixes with results obtained in as little as 80 min post-enrichment.
Topics: Vibrio parahaemolyticus; Vibrio vulnificus; Vibrio cholerae; Real-Time Polymerase Chain Reaction; Seafood; Food Microbiology
PubMed: 37243669
DOI: 10.1093/jaoacint/qsad061 -
Microbiology Spectrum Jun 2015Vibrio vulnificus, carrying a 50% fatality rate, is the most deadly of the foodborne pathogens. It occurs in estuarine and coastal waters and it is found in especially...
Vibrio vulnificus, carrying a 50% fatality rate, is the most deadly of the foodborne pathogens. It occurs in estuarine and coastal waters and it is found in especially high numbers in oysters and other molluscan shellfish. The biology of V. vulnificus, including its ecology, pathogenesis, and molecular genetics, has been described in numerous reviews. This article provides a brief summary of some of the key aspects of this important human pathogen, including information on biotypes and genotypes, virulence factors, risk factor requirements and the role of iron in disease, association with oysters, geographic distribution, importance of salinity and water temperature, increasing incidence associated with global warming. This article includes some of our findings as presented at the "Vibrios in the Environment 2010" conference held in Biloxi, MS.
Topics: Animals; Bacterial Capsules; Foodborne Diseases; Global Warming; Humans; Iron; Ostreidae; Risk Factors; Salinity; Seawater; Sepsis; Shellfish; Temperature; Vibrio Infections; Vibrio vulnificus; Virulence Factors; Water Microbiology
PubMed: 26185084
DOI: 10.1128/microbiolspec.VE-0001-2014 -
PLoS Pathogens Jan 2023Many pathogenic bacteria form biofilms to survive under environmental stresses and host immune defenses. Differential expression (DE) analysis of the genes in biofilm...
Many pathogenic bacteria form biofilms to survive under environmental stresses and host immune defenses. Differential expression (DE) analysis of the genes in biofilm and planktonic cells under a single condition, however, has limitations to identify the genes essential for biofilm formation. Independent component analysis (ICA), a machine learning algorithm, was adopted to comprehensively identify the biofilm genes of Vibrio vulnificus, a fulminating human pathogen, in this study. ICA analyzed the large-scale transcriptome data of V. vulnificus cells under various biofilm and planktonic conditions and then identified a total of 72 sets of independently co-regulated genes, iModulons. Among the three iModulons specifically activated in biofilm cells, BrpT-iModulon mainly consisted of known genes of the regulon of BrpT, a transcriptional regulator controlling biofilm formation of V. vulnificus. Interestingly, the BrpT-iModulon additionally contained two novel genes, VV1_3061 and VV2_1694, designated as cabH and brpN, respectively. cabH and brpN were shared in other Vibrio species and not yet identified by DE analyses. Genetic and biochemical analyses revealed that cabH and brpN are directly up-regulated by BrpT. The deletion of cabH and brpN impaired the robust biofilm and rugose colony formation. CabH, structurally similar to the previously known calcium-binding matrix protein CabA, was essential for attachment to the surface. BrpN, carrying an acyltransferase-3 domain as observed in BrpL, played an important role in exopolysaccharide production. Altogether, ICA identified two novel genes, cabH and brpN, which are regulated by BrpT and essential for the development of robust biofilms and rugose colonies of V. vulnificus.
Topics: Humans; Vibrio vulnificus; Transcriptome; Biofilms; Vibrio; Genes, Bacterial; Gene Expression Regulation, Bacterial
PubMed: 36656902
DOI: 10.1371/journal.ppat.1011064