Haemagglutinin/protease expression and mucin gel penetration in El Tor biotype Vibrio cholerae

Microbiology ◽  
2003 ◽  
Vol 149 (7) ◽  
pp. 1883-1891 ◽  
Author(s):  
Anisia J. Silva ◽  
Kim Pham ◽  
Jorge A. Benitez
Keyword(s):  
Gastrointestinal Tract ◽  
Vibrio Cholerae ◽  
Transcriptional Activation ◽  
Bile Salts ◽  
Promoter Activity ◽  
Protease Production ◽  
Independent Mechanism ◽  
A Cell ◽  
El Tor ◽  
Protease Expression

Vibrio cholerae of both biotypes produce a soluble Zn2+-dependent metalloprotease: haemagglutinin/protease (Hap), encoded by hapA. Hap has been shown to have mucinolytic and cytotoxic activity. These activities are likely to play an important role in the pathogenesis of cholera and the reactogenicity of attenuated vaccine strains. Production of Hap requires transcriptional activation by the HapR regulator and is repressed by glucose. The present study shows that mucin purified from two sources, bile salts, and growth at 37 °C enhanced Hap protease production. Analysis of hapA and hapR promoter fusions with the lacZ gene showed both promoters to be activated in a cell-density-dependent pattern. Glucose repressed and mucin induced the hapA promoter by a HapR-independent mechanism. Bile had no effect on either hapR or hapA promoter activity. Expression of hapA was required for vibrios to translocate through a mucin-containing gel. These results suggest Hap to play an important role in cholera pathogenesis by promoting mucin gel penetration, detachment and spreading of infection along the gastrointestinal tract.

scholarly journals Characterization of Vibrio cholerae O1 El TorgalU and galE Mutants: Influence on Lipopolysaccharide Structure, Colonization, and Biofilm Formation

2001 ◽  
Vol 69 (1) ◽  
pp. 435-445 ◽  
Author(s):  
Jutta Nesper ◽  
Crystal M. Lauriano ◽  
Karl E. Klose ◽  
Dagmar Kapfhammer ◽  
Anita Kraiß ◽  
...  
Keyword(s):  
Organic Acids ◽  
Vibrio Cholerae ◽  
Bile Salts ◽  
Short Chain ◽  
Core Oligosaccharide ◽  
O Antigen ◽  
Mouse Small Intestine ◽  
Vibrio Cholerae O1 ◽  
El Tor

ABSTRACT Recently we described the isolation of spontaneous bacteriophage K139-resistant Vibrio cholerae O1 El Tor mutants. In this study, we identified phage-resistant isolates with intact O antigen but altered core oligosaccharide which were also affected in galactose catabolism; this strains have mutations in the galU gene. We inactivated another gal gene, galE, and the mutant was also found to be defective in the catabolism of exogenous galactose but synthesized an apparently normal lipopolysaccharide (LPS). Both gal mutants as well as a rough LPS (R-LPS) mutant were investigated for the ability to colonize the mouse small intestine. The galU and R-LPS mutants, but not thegalE mutant, were defective in colonization, a phenotype also associated with O-antigen-negative mutants. By investigating several parameters in vitro, we could show that galU and R-LPS mutants were more sensitive to short-chain organic acids, cationic antimicrobial peptides, the complement system, and bile salts as well as other hydrophobic agents, indicating that their outer membrane no longer provides an effective barrier function. O-antigen-negative strains were found to be sensitive to complement and cationic peptides, but they displayed significant resistance to bile salts and short-chain organic acids. Furthermore, we found thatgalU and galE are essential for the formation of a biofilm in a spontaneous phage-resistant rugose variant, suggesting that the synthesis of UDP-galactose via UDP-glucose is necessary for biosynthesis of the exopolysaccharide. In addition, we provide evidence that the production of exopolysaccharide limits the access of phage K139 to its receptor, the O antigen. In conclusion, our results indicate involvement of galU in V. cholerae virulence, correlated with the observed change in LPS structure, and a role for galU and galE in environmental survival of V. cholerae.

scholarly journals A Constitutively Active Variant of the Quorum-Sensing Regulator LuxO Affects Protease Production and Biofilm Formation in Vibrio cholerae

2003 ◽  
Vol 71 (5) ◽  
pp. 2571-2576 ◽  
Author(s):  
Russell E. Vance ◽  
Jun Zhu ◽  
John J. Mekalanos
Keyword(s):  
Quorum Sensing ◽  
Vibrio Cholerae ◽  
Biofilm Formation ◽  
Protease Production ◽  
Aquatic Habitats ◽  
Wild Type ◽  
Protease Deficient ◽  
Active Variant ◽  
Protease Expression

ABSTRACT Vibrio cholerae normally inhabits aquatic habitats but can cause a severe diarrheal illness in humans. Its arsenal of virulence factors includes a secreted hemagglutinin (HA) protease. An HA protease-deficient mutant of V. cholerae was isolated and designated E7946 mpc. E7946 mpc was found to contain a point mutation in the luxO quorum-sensing regulator. In accordance with this finding, E7946 mpc exhibits a defect in quorum sensing. The mutant luxO allele [luxO(Con)] produces a protein with a leucine-to-glutamine substitution at amino acid 104. Transfer of the luxO(Con) allele to an otherwise wild-type background was sufficient to eliminate HA protease expression; conversely, deletion of luxO(Con) from E7946 mpc restored protease activity. We demonstrate that LuxO(Con) constitutively represses the transcription of hapR, an essential positive regulator of HA protease. Interestingly, strains harboring luxO(Con) form enhanced biofilms, and enhanced biofilm formation does not appear to be dependent on reduced HA protease expression. Taken together, the results confirm the role of LuxO as a central “switch” that coordinately regulates virulence-related phenotypes such as protease production and biofilm formation.

scholarly journals Enhancement by sodium chloride of the selectivity of thiosulfate citrate bile salts sucrose agar for isolating Vibrio cholerae biotype El Tor

1976 ◽  
Vol 4 (2) ◽  
pp. 133-136
Author(s):  
G K Morris ◽  
W E DeWitt ◽  
E J Gangarosa ◽  
W M McCormack
Keyword(s):  
Sodium Chloride ◽  
Vibrio Cholerae ◽  
Bile Salts ◽  
Selective Medium ◽  
Salt Tolerant ◽  
Normal Bowel ◽  
Inhibition Of Growth ◽  
El Tor

In this study we utilized the salt-tolerant characteristics of vibrios to develop a more selective medium by addition of NaCl to thiosulfate citrate bile salts sucrose (TCBS) agar. The effect of adding salt to TCBS agar varied greatly among brands of TCBS agar and between lots of the same brand. The addition of salt at concentrations as high as 1.5% (2.5% total NaCl) caused the inhibition of growth of three species of commonly encountered normal bowel flora and one strain of classical Vibrio cholerae but did not compromise the use of TCBS agar for isolation of V. cholerae biotype El Tor.

scholarly journals Cell Vacuolation, a Manifestation of the El Tor Hemolysin of Vibrio cholerae

2000 ◽  
Vol 68 (4) ◽  
pp. 1928-1933 ◽  
Author(s):  
Rupak Mitra ◽  
Paula Figueroa ◽  
Asish K. Mukhopadhyay ◽  
Toshio Shimada ◽  
Yoshifumi Takeda ◽  
...  
Keyword(s):  
Vibrio Cholerae ◽  
Pcr Amplification ◽  
Neutral Red ◽  
Partial Purification ◽  
Insertion Mutation ◽  
A Cell ◽  
Red Dye ◽  
Vacuolated Cells ◽  
El Tor ◽  
Culture Supernatants

ABSTRACT Culture supernatants of nontoxigenic nonepidemic clinical strains of Vibrio cholerae belonging to diverse serogroups were found to induce vacuolation of nonconfluent HeLa cells. The vacuoles became prominent 18 h after introduction of culture supernatant, and vacuolated cells survived for 48 h and then died. Only a fraction of the vacuolated cells took up neutral red dye, implying that there were differences in the vacuolar microenvironment. Further tests showed that the factor responsible for vacuolation was heat labile and proteinaceous. Vacuolating activity was completely neutralized by antibody to hemolysin of V. cholerae but not by antibody to vacuolating cytotoxin of Helicobacter pylori. Partial purification of the vacuolating factor led to elution of fractions, which showed both hemolytic and vacuolating activity. PCR amplification and cloning of the hemolysin structural gene (hlyA) intoEscherichia coli DH5α led to isolation of clones producing cell vacuolating factor in a cell-associated form. Further, a null insertion mutation in the hlyA gene of a high-vacuolating-factor-producing strain led to complete abolition of both cell vacuolating and hemolytic activities. These analyses establish vacuolation as a potentially important but previously unrecognized property of V. cholerae El Tor hemolysin.

scholarly journals The nucleoid occlusion protein SlmA is a direct transcriptional activator of chitobiose utilization in Vibrio cholerae

2017 ◽  
Author(s):  
Catherine A. Klancher ◽  
Chelsea A. Hayes ◽  
Ankur B. Dalia
Keyword(s):  
Cell Division ◽  
Vibrio Cholerae ◽  
Transcriptional Activation ◽  
Aquatic Environment ◽  
Point Mutations ◽  
Marine Zooplankton ◽  
Contaminated Food ◽  
A Cell

ABSTRACTChitin utilization by the cholera pathogen Vibrio cholerae is required for its persistence and evolution via horizontal gene transfer in the marine environment. Genes involved in the uptake and catabolism of the chitin disaccharide chitobiose are encoded by the chb operon. The orphan sensor kinase ChiS is critical for regulation of this locus, however, the mechanisms downstream of ChiS activation that result in expression of the chb operon are poorly understood. Using an unbiased transposon mutant screen, we uncover that the nucleoid occlusion protein SlmA is a regulator of the chb operon. SlmA has not previously been implicated in gene regulation. Also, SlmA is a member of the TetR family of proteins, which are generally transcriptional repressors. In vitro, we find that SlmA binds directly to the chb operon promoter, and in vivo, we show that this interaction is, surprisingly, required for transcriptional activation of this locus and for chitobiose utilization. Using point mutations that disrupt distinct functions of SlmA, we find that DNA-binding, but not nucleoid occlusion, is critical for transcriptional activation. This study identifies a novel role for SlmA as a transcriptional regulator in V. cholerae in addition to its established role as a cell division licensing factor.AUTHOR SUMMARYThe cholera pathogen Vibrio cholerae is a natural resident of the aquatic environment and causes disease when ingested in the form of contaminated food or drinking water. In the aquatic environment, the shells of marine zooplankton, which are primarily composed of chitin, serve as an important food source for this pathogen. The genes required for the utilization of chitin are tightly regulated in V. cholerae, however, the exact mechanism underlying this regulation is currently unclear. Here, we uncover that a protein involved in regulating cell division is also important for regulating the genes involved in chitin utilization. This is a newly identified property for this cell division protein and the significance of a common regulator for these two disparate activities remains to be understood.

scholarly journals VGJφ, a Novel Filamentous Phage of Vibrio cholerae, Integrates into the Same Chromosomal Site as CTXφ

2003 ◽  
Vol 185 (19) ◽  
pp. 5685-5696 ◽  
Author(s):  
Javier Campos ◽  
Eriel Martínez ◽  
Edith Suzarte ◽  
Boris L. Rodríguez ◽  
Karen Marrero ◽  
...  
Keyword(s):  
Vibrio Cholerae ◽  
Open Reading Frames ◽  
General Mechanism ◽  
Filamentous Phage ◽  
Type Iv ◽  
A Cell ◽  
Conserved Region ◽  
El Tor ◽  
Reading Frames ◽  
Filamentous Phages

ABSTRACT We describe a novel filamentous phage, designated VGJφ, isolated from strain SG25-1 of Vibrio cholerae O139, which infects all O1 (classical and El Tor) and O139 strains tested. The sequence of the 7,542 nucleotides of the phage genome reveals that VGJφ has a distinctive region of 775 nucleotides and a conserved region with an overall genomic organization similar to that of previously characterized filamentous phages, such as CTXφ of V. cholerae and Ff phages of Escherichia coli. The conserved region carries 10 open reading frames (ORFs) coding for products homologous to previously reported peptides of other filamentous phages, and the distinctive region carries one ORF whose product is not homologous to any known peptide. VGJφ, like other filamentous phages, uses a type IV pilus to infect V. cholerae; in this case, the pilus is the mannose-sensitive hemagglutinin. VGJφ-infected V. cholerae overexpresses the product of one ORF of the phage (ORF112), which is similar to single-stranded DNA binding proteins of other filamentous phages. Once inside a cell, VGJφ is able to integrate its genome into the same chromosomal attB site as CTXφ, entering into a lysogenic state. Additionally, we found an attP structure in VGJφ, which is also conserved in several lysogenic filamentous phages from different bacterial hosts. Finally, since different filamentous phages seem to integrate into the bacterial dif locus by a general mechanism, we propose a model in which repeated integration events with different phages might have contributed to the evolution of the CTX chromosomal region in V. cholerae El Tor.

scholarly journals Elucidating the correlation between the number of TTTTGAT heptamer repeats and cholera toxin promoter activity in Vibrio cholerae O1 pandemic strains

2021 ◽  
Author(s):  
Arindam Naha ◽  
Jeffrey H Withey ◽  
Piyali Mukherjee ◽  
Rudra Narayan Saha ◽  
Prosenjit Samanta ◽  
...  
Keyword(s):  
Cholera Toxin ◽  
Vibrio Cholerae ◽  
Transcriptional Activation ◽  
Promoter Activity ◽  
Watery Diarrhoea ◽  
Transcriptional Activators ◽  
Regulatory Cascade ◽  
Vibrio Cholerae O1 ◽  
Pandemic Strains ◽  
Heptad Repeats

A complex regulatory cascade controls expression of the cholera toxin genes (ctxAB) in Vibrio cholerae; which eventually leads to choleragen (CT) production and secretion, resulting in rice watery diarrhoea. The cholera toxin promoter (PctxAB) contains a series of heptad repeats (5′-TTTTGAT-3′); which have been previously shown to play crucial role in ctxAB transcriptional regulation by recruiting the transcriptional activators ToxT, ToxR, and the nucleoid-associated protein H-NS along the ctx promoter. The numbers of these repeats vary between the two biotypes of V. cholerae O1 strains, and even among strains of the same biotype. In this study, we examined PctxAB activation of V. cholerae O1 pandemic strains to understand the significance of the distal heptad repeats in regulating ctx expression. Interestingly, we found that ctx activation may depend on the number of TTTTGAT heptad repeats within PctxAB, and we posit that the occupation of the distal repeats by H-NS could further prevent transcriptional activation of ctx genes in V. cholerae. We hypothesize that ToxT-dependent transcriptional activation may not require entire displacement of H-NS and propose a revision in the currently accepted model of ToxT dependent PctxAB transcriptional activation.

scholarly journals Atypical and dual biotypes variant of virulent SA-NAG-Vibrio cholerae: an evidence of emerging/evolving patho-significant strain in municipal domestic water sources

2022 ◽  
Vol 72 (1) ◽  
Author(s):  
Bright E. Igere ◽  
Anthony I. Okoh ◽  
Uchechukwu U. Nwodo
Keyword(s):  
Vibrio Cholerae ◽  
Ground Surface ◽  
Polymyxin B ◽  
Protease Production ◽  
Eastern Cape Province ◽  
Eastern Cape ◽  
Domestic Water ◽  
Public Health Systems ◽  
Microbiological Methods ◽  
El Tor

Abstract Introduction and purpose The recent cholera spread, new cases, and fatality continue to arouse concern in public health systems; however, interventions on control is at its peak yet statistics show continuous report. This study characterized atypical and patho-significant environmental Vibrio cholerae retrieved from ground/surface/domestic water in rural-urban-sub-urban locations of Amathole District municipality and Chris Hani District municipality, Eastern Cape Province, South Africa. Methods Domestic/surface water was sampled and 759 presumptive V. cholerae isolates were retrieved using standard microbiological methods. Virulence phenotypic test: toxin co-regulated pili (tcp), choleragen red, protease production, lecithinase production, and lipase test were conducted. Serotyping using polyvalent antisera (Bengal and Ogawa/Inaba/Hikojima) and molecular typing: 16SrRNA, OmpW, serogroup (Vc-O1/O139), biotype (tcpAClas/El Tor, HlyAClas/El Tor, rstRClas/El Tor, RS1, rtxA, rtxC), and virulence (ctxA, ctxB, zot, ace, cep, prt, toxR, hlyA) genes were targeted. Result Result of 16SrRNA typing confirmed 508 (66.9%) while OmpW detected/confirmed 61 (12.01%) V. cholerae strains. Phenotypic-biotyping scheme showed positive test to polymyxin B (68.9%), Voges proskauer (6.6%), and Bengal serology (11.5%). Whereas Vc-O1/O139 was negative, yet two of the isolates harbored the cholera toxin with a gene-type ctxB and hlyAClas: 2/61, revealing atypical/unusual/dual biotype phenotypic/genotypic features. Other potential atypical genotypes detected include rstR: 7/61, Cep: 15/61, ace: 20/61, hlyAElTor: 53/61, rtxA: 30/61, rtxC: 11/61, and prtV: 15/61 respectively. Conclusion Although additional patho-significant/virulent genotypes associated with epidemic/sporadic cholera cases were detected, an advanced, bioinformatics, and post-molecular evaluation is necessary. Such stride possesses potential to adequately minimize future cholera cases associated with dynamic/atypical environmental V. cholerae strains.

Genomic Variability of Pathogenicity Islands in Nontoxigenic Strains of Vibrio cholerae O1 Biotype El Tor

2020 ◽  
Vol 56 (9) ◽  
pp. 1055-1069
Author(s):  
N. I. Smirnova ◽  
A. A. Kritsky ◽  
J. V. Alkhova ◽  
E. Yu. Agafonova ◽  
E. Yu. Shchelkanova ◽  
...  
Keyword(s):  
Vibrio Cholerae ◽  
Pathogenicity Islands ◽  
Genomic Variability ◽  
Vibrio Cholerae O1 ◽  
El Tor

scholarly journals A Yeast taf17 Mutant Requires the Swi6 Transcriptional Activator for Viability and Shows Defects in Cell Cycle-Regulated Transcription

Genetics ◽  
2000 ◽  
Vol 154 (4) ◽  
pp. 1561-1576
Author(s):  
Neil Macpherson ◽  
Vivien Measday ◽  
Lynda Moore ◽  
Brenda Andrews
Keyword(s):  
Cell Cycle ◽  
Transcriptional Activation ◽  
Essential Gene ◽  
Temperature Sensitive ◽  
Permissive Temperature ◽  
Synthetic Lethal ◽  
Cycle Arrest ◽  
A Cell ◽  
Allelism Tests ◽  
Complementation Groups

Abstract In Saccharomyces cerevisiae, the Swi6 protein is a component of two transcription factors, SBF and MBF, that promote expression of a large group of genes in the late G1 phase of the cell cycle. Although SBF is required for cell viability, SWI6 is not an essential gene. We performed a synthetic lethal screen to identify genes required for viability in the absence of SWI6 and identified 10 complementation groups of swi6-dependent lethal mutants, designated SLM1 through SLM10. We were most interested in mutants showing a cell cycle arrest phenotype; both slm7-1 swi6Δ and slm8-1 swi6Δ double mutants accumulated as large, unbudded cells with increased 1N DNA content and showed a temperature-sensitive growth arrest in the presence of Swi6. Analysis of the transcript levels of cell cycle-regulated genes in slm7-1 SWI6 mutant strains at the permissive temperature revealed defects in regulation of a subset of cyclin-encoding genes. Complementation and allelism tests showed that SLM7 is allelic with the TAF17 gene, which encodes a histone-like component of the general transcription factor TFIID and the SAGA histone acetyltransferase complex. Sequencing showed that the slm7-1 allele of TAF17 is predicted to encode a version of Taf17 that is truncated within a highly conserved region. The cell cycle and transcriptional defects caused by taf17slm7-1 are consistent with the role of TAFIIs as modulators of transcriptional activation and may reflect a role for TAF17 in regulating activation by SBF and MBF.

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