scholarly journals Identification of the Vibrio cholerae Enterobactin Receptors VctA and IrgA: IrgA Is Not Required for Virulence

2002 ◽  
Vol 70 (7) ◽  
pp. 3419-3426 ◽  
Author(s):  
Alexandra R. Mey ◽  
Elizabeth E. Wyckoff ◽  
Amanda G. Oglesby ◽  
Eva Rab ◽  
Ronald K. Taylor ◽  
...  
Keyword(s):  
Vibrio Cholerae ◽  
Mutant Strain ◽  
Genomic Sequence ◽  
Iron Acquisition ◽  
Lethal Dose ◽  
Single Mutation ◽  
Periplasmic Binding Protein ◽  
Infant Mouse ◽  
Abc Transport ◽  
Important Virulence Factor

ABSTRACT The gram-negative enteric pathogen Vibrio cholerae requires iron for growth. V. cholerae has multiple iron acquisition systems, including utilization of heme and hemoglobin, synthesis and transport of the catechol siderophore vibriobactin, and transport of several siderophores that it does not itself make. One siderophore that V. cholerae transports, but does not make, is enterobactin. Enterobactin transport requires TonB and is independent of the vibriobactin receptor ViuA. In this study, two candidate enterobactin receptor genes, irgA (VC0475) and vctA (VCA0232), were identified by analysis of the V. cholerae genomic sequence. A single mutation in either of these genes did not significantly impair enterobactin utilization, but a strain defective in both genes did not use enterobactin. When either irgA or vctA was supplied on a plasmid, the ability of the irgA vctA double mutant to use enterobactin was restored. This indicates that both VctA and IrgA transport enterobactin. We also identify the genes vctPDGC, which are linked to vctA and encode a periplasmic binding protein-dependent ABC transport system that functions in the utilization of both enterobactin and vibriobactin (VCA0227-0230). An irgA::TnphoA mutant strain, MBG40, was shown in a previous study to be highly attenuated and to have a strong colonization defect in an infant mouse model of V. cholerae infection (M. B. Goldberg, V. J. DiRita, and S. B. Calderwood, Infect. Immun. 58:55-60, 1990). In this work, a new irgA mutation was constructed, and this mutant strain was not significantly impaired in its ability to compete with the parental strain in infant mice and was not attenuated for virulence in an assay of 50% lethal dose. These data indicate that the virulence defect in MBG40 is not due to the loss of irgA function and that irgA is unlikely to be an important virulence factor.

scholarly journals Characterization of Ferric and Ferrous Iron Transport Systems in Vibrio cholerae

2006 ◽  
Vol 188 (18) ◽  
pp. 6515-6523 ◽  
Author(s):  
Elizabeth E. Wyckoff ◽  
Alexandra R. Mey ◽  
Andreas Leimbach ◽  
Carolyn F. Fisher ◽  
Shelley M. Payne
Keyword(s):  
Vibrio Cholerae ◽  
Transport System ◽  
Iron Transport ◽  
Iron Acquisition ◽  
Shigella Flexneri ◽  
Transport Systems ◽  
Infant Mouse ◽  
Mouse Intestine ◽  
Dependent Transport

ABSTRACT Vibrio cholerae has multiple iron acquisition systems, including TonB-dependent transport of heme and of the catechol siderophore vibriobactin. Strains defective in both of these systems grow well in laboratory media and in the infant mouse intestine, indicating the presence of additional iron acquisition systems. Previously uncharacterized potential iron transport systems, including a homologue of the ferrous transporter Feo and a periplasmic binding protein-dependent ATP binding cassette (ABC) transport system, termed Fbp, were identified in the V. cholerae genome sequence. Clones encoding either the Feo or the Fbp system exhibited characteristics of iron transporters: both repressed the expression of lacZ cloned under the control of a Fur-regulated promoter in Escherichia coli and also conferred growth on a Shigella flexneri mutant that has a severe defect in iron transport. Two other ABC transporters were also evaluated but were negative by these assays. Transport of radioactive iron by the Feo system into the S. flexneri iron transport mutant was stimulated by the reducing agent ascorbate, consistent with Feo functioning as a ferrous transporter. Conversely, ascorbate inhibited transport by the Fbp system, suggesting that it transports ferric iron. The growth of V. cholerae strains carrying mutations in one or more of the potential iron transport genes indicated that both Feo and Fbp contribute to iron acquisition. However, a mutant defective in the vibriobactin, Fbp, and Feo systems was not attenuated in a suckling mouse model, suggesting that at least one other iron transport system can be used in vivo.

scholarly journals The Alternative Sigma Factor σE Plays an Important Role in Intestinal Survival and Virulence in Vibrio cholerae

2002 ◽  
Vol 70 (10) ◽  
pp. 5355-5362 ◽  
Author(s):  
Gabriela Kovacikova ◽  
Karen Skorupski
Keyword(s):  
Vibrio Cholerae ◽  
Sigma Factor ◽  
Parental Strain ◽  
Time Course ◽  
Lethal Dose ◽  
Alternative Sigma Factor ◽  
Growth And Survival ◽  
Infant Mouse

ABSTRACT The alternative sigma factor σΕ (RpoE) is involved in the response to extracytoplasmic stress and plays a role in the virulence of a variety of different bacteria. To assess the role of σΕ in Vibrio cholerae pathogenesis, a ΔrpoE mutant was constructed and analyzed using the infant mouse model. The results here show that σΕ contributes significantly to the virulence of V. cholerae. The ΔrpoE mutant was highly attenuated with a 50% lethal dose more than 3 logs higher than that for the parental strain, and its ability to colonize the intestine was reduced approximately 30-fold. A time course of infection revealed that the number of CFU of the ΔrpoE mutant was approximately 1 log lower than that of the parental strain by 12 h postinoculation and decreased further by 24 h. The defect in virulence in the ΔrpoE mutant thus appears to be a diminished ability to survive within the intestinal environment. The results here also show that σΕ is not required for growth and survival of V. cholerae in vitro at high temperatures but is required under other stressful conditions, such as in the presence of 3% ethanol. As in Escherichia coli, the expression of rpoE in V. cholerae is dependent upon two promoters located upstream of the gene, P1 and P2. P1 appears to be σ70 dependent, whereas the downstream promoter, P2, is positively autoregulated by σΕ.

scholarly journals Iron and Fur Regulation in Vibrio cholerae and the Role of Fur in Virulence

2005 ◽  
Vol 73 (12) ◽  
pp. 8167-8178 ◽  
Author(s):  
Alexandra R. Mey ◽  
Elizabeth E. Wyckoff ◽  
Vanamala Kanukurthy ◽  
Carolyn R. Fisher ◽  
Shelley M. Payne
Keyword(s):  
Gene Expression ◽  
Vibrio Cholerae ◽  
Iron Acquisition ◽  
Bacterial Species ◽  
Regulation Of Gene Expression ◽  
Transport Systems ◽  
Infant Mouse ◽  
Genes Encoding ◽  
Regulatory Functions

ABSTRACT Regulation of iron uptake and utilization is critical for bacterial growth and for prevention of iron toxicity. In many bacterial species, this regulation depends on the iron-responsive master regulator Fur. In this study we report the effects of iron and Fur on gene expression in Vibrio cholerae. We show that Fur has both positive and negative regulatory functions, and we demonstrate Fur-independent regulation of gene expression by iron. Nearly all of the known iron acquisition genes were repressed by Fur under iron-replete conditions. In addition, genes for two newly identified iron transport systems, Feo and Fbp, were found to be negatively regulated by iron and Fur. Other genes identified in this study as being induced in low iron and in the fur mutant include those encoding superoxide dismutase (sodA), fumarate dehydratase (fumC), bacterioferritin (bfr), bacterioferritin-associated ferredoxin (bfd), and multiple genes of unknown function. Several genes encoding iron-containing proteins were repressed in low iron and in the fur mutant, possibly reflecting the need to reserve available iron for the most critical functions. Also repressed in the fur mutant, but independently of iron, were genes located in the V. cholerae pathogenicity island, encoding the toxin-coregulated pilus (TCP), and genes within the V. cholerae mega-integron. The fur mutant exhibited very weak autoagglutination, indicating a possible defect in expression or assembly of the TCP, a major virulence factor of V. cholerae. Consistent with this observation, the fur mutant competed poorly with its wild-type parental strain for colonization of the infant mouse gut.

scholarly journals Nonredundant Roles of Iron Acquisition Systems in Vibrio cholerae

2015 ◽  
Vol 84 (2) ◽  
pp. 511-523 ◽  
Author(s):  
Eric D. Peng ◽  
Elizabeth E. Wyckoff ◽  
Alexandra R. Mey ◽  
Carolyn R. Fisher ◽  
Shelley M. Payne
Keyword(s):  
Vibrio Cholerae ◽  
Diarrheal Disease ◽  
Iron Acquisition ◽  
Growth Conditions ◽  
Growth Defect ◽  
Null Mutant ◽  
Content Type ◽  
Infant Mouse ◽  
Iron Transporter

Vibrio cholerae, the causative agent of the severe diarrheal disease cholera, thrives in both marine environments and the human host. To do so, it must encode the tools necessary to acquire essential nutrients, including iron, under these vastly different conditions. A number ofV. choleraeiron acquisition systems have been identified; however, the precise role of each system is not fully understood. To test the roles of individual systems, we generated a series of mutants in which only one of the four systems that support iron acquisition on unsupplemented LB agar, Feo, Fbp, Vct, and Vib, remains functional. Analysis of these mutants under different growth conditions showed that these systems are not redundant. The strain carrying only the ferrous iron transporter Feo grew well at acidic, but not alkaline, pH, whereas the ferric iron transporter Fbp promoted better growth at alkaline than at acidic pH. A strain defective in all four systems (null mutant) had a severe growth defect under aerobic conditions but accumulated iron and grew as well as the wild type in the absence of oxygen, suggesting the presence of an additional, unidentified iron transporter inV. cholerae. In support of this, the null mutant was only moderately attenuated in an infant mouse model of infection. While the null mutant used heme as an iron sourcein vitro, we demonstrate that heme is not available toV. choleraein the infant mouse intestine.

scholarly journals FbpC Is Not Essential for Iron Acquisition inNeisseria gonorrhoeae

1999 ◽  
Vol 67 (6) ◽  
pp. 3141-3145 ◽  
Author(s):  
Shite Sebastian ◽  
Caroline Attardo Genco
Keyword(s):  
Iron Transport ◽  
Iron Acquisition ◽  
Transport Systems ◽  
Northern Analysis ◽  
Pcr Analysis ◽  
Ferric Nitrate ◽  
Periplasmic Binding Protein ◽  
Reverse Transcription Pcr ◽  
Abc Transport ◽  
Insertional Inactivation

ABSTRACT The fbpABC locus of Neisseria gonorrhoeaehas been proposed to encode a periplasmic protein-dependent iron transport system. Although the function of the gonococcal FbpA protein has been well characterized and its role as a periplasmic binding protein is well defined, little is known about the function of the FbpB and FbpC proteins. To define the function of the gonococcal FbpC protein, an N. gonorrhoeae F62 fbpC mutant was constructed by insertional inactivation with the kanamycin gene. TheN. gonorrhoeae F62 fbpC mutant was observed to grow with heme, transferrin, or ferric nitrate as the sole exogenous iron source, indicating that the gonococcal FbpC protein is not absolutely required for growth with these iron sources. In previous studies we were unable to detect fbpB- orfbpC-specific transcripts by Northern analysis. Reverse transcription-PCR analysis with RNA obtained from N. gonorrhoeae F62 grown under iron-replete and -depleted conditions detected fbpA and fbpAB transcripts but failed to detect fbpC or fbpBC transcripts. These results indicate that FbpC does not play a pivotal role in iron transport in N. gonorrhoeae and suggest that additional ABC transport systems are functional in the gonococcus for the acquisition of iron.

scholarly journals Transcutaneous Immunization with Toxin-Coregulated Pilin A Induces Protective Immunity against Vibrio cholerae O1 El Tor Challenge in Mice

2006 ◽  
Vol 74 (10) ◽  
pp. 5834-5839 ◽  
Author(s):  
Julianne E. Rollenhagen ◽  
Anuj Kalsy ◽  
Francisca Cerda ◽  
Manohar John ◽  
Jason B. Harris ◽  
...  
Keyword(s):  
Cholera Toxin ◽  
Vibrio Cholerae ◽  
Lethal Dose ◽  
Infant Mouse ◽  
Type Iv ◽  
Transcutaneous Immunization ◽  
Structural Subunit ◽  
Vibrio Cholerae O1 ◽  
The Mean ◽  
El Tor

ABSTRACT Toxin-coregulated pilin A (TcpA) is the main structural subunit of a type IV bundle-forming pilus of Vibrio cholerae, the cause of cholera. Toxin-coregulated pilus is involved in formation of microcolonies of V. cholerae at the intestinal surface, and strains of V. cholerae deficient in TcpA are attenuated and unable to colonize intestinal surfaces. Anti-TcpA immunity is common in humans recovering from cholera in Bangladesh, and immunization against TcpA is protective in murine V. cholerae models. To evaluate whether transcutaneously applied TcpA is immunogenic, we transcutaneously immunized mice with 100 μg of TcpA or TcpA with an immunoadjuvant (cholera toxin [CT], 50 μg) on days 0, 19, and 40. Mice immunized with TcpA alone did not develop anti-TcpA responses. Mice that received transcutaneously applied TcpA and CT developed prominent anti-TcpA immunoglobulin G (IgG) serum responses but minimal anti-TcpA IgA. Transcutaneous immunization with CT induced prominent IgG and IgA anti-CT serum responses. In an infant mouse model, offspring born to dams transcutaneously immunized either with TcpA and CT or with CT alone were challenged with 106 CFU (one 50% lethal dose) wild-type V. cholerae O1 El Tor strain N16961. At 48 h, mice born to females transcutaneously immunized with CT alone had 36% ± 10% (mean ± standard error of the mean) survival, while mice born to females transcutaneously immunized with TcpA and CT had 69% ± 6% survival (P < 0.001). Our results suggest that transcutaneous immunization with TcpA and an immunoadjuvant induces protective anti-TcpA immune responses. Anti-TcpA responses may contribute to an optimal cholera vaccine.

scholarly journals The Global Regulator ArcA Modulates Expression of Virulence Factors in Vibrio cholerae

2003 ◽  
Vol 71 (10) ◽  
pp. 5583-5589 ◽  
Author(s):  
Nilanjan Sengupta ◽  
Kalidas Paul ◽  
Rukhsana Chowdhury
Keyword(s):  
Vibrio Cholerae ◽  
Mutant Strain ◽  
Virulence Factors ◽  
Response Regulator ◽  
Anaerobic Growth ◽  
Infant Mouse ◽  
Reverse Transcription Pcr ◽  
Concomitant Reduction ◽  
Aerobic And Anaerobic

ABSTRACT A Vibrio cholerae arcA mutant was constructed and used to examine the role of the global anaerobiosis response regulator ArcA in the expression of virulence factors in this important human pathogen. In V. cholerae, expression of the major virulence factors cholera toxin (CT) and toxin-coregulated pilus (TCP) is regulated by the transcriptional activator ToxT. toxT expression, in turn, is controlled by the transmembrane DNA binding proteins ToxR and TcpP. In the V. cholerae arcA mutant, although ToxR and TcpP were unaffected, Northern blot and reverse transcription-PCR analyses indicated that the expression of toxT was significantly decreased with concomitant reduction in the expression of CT and TCP. CT and TCP expression was completely restored in the V. cholerae arcA mutant strain by expressing a cloned toxT gene in the mutant. These results suggest that ArcA functions as a positive regulator of toxT expression under both aerobic and anaerobic conditions, although as expected, the effect was more pronounced during anaerobic growth. This was reflected in a reduction of virulence of the V. cholerae arcA mutant strain in the infant mouse cholera model.

Two regulatory factors of Vibrio cholerae activating the mannose-sensitive haemagglutinin pilus expression is important for biofilm formation and colonization in mice

Microbiology ◽  
2021 ◽  
Vol 167 (10) ◽  
Author(s):  
Mengting Shi ◽  
Yue Zheng ◽  
Xianghong Wang ◽  
Zhengjia Wang ◽  
Menghua Yang
Keyword(s):  
Mouse Model ◽  
Vibrio Cholerae ◽  
Biofilm Formation ◽  
Type Species ◽  
Wild Type ◽  
Expression Library ◽  
Content Type ◽  
Genomic Expression ◽  
Infant Mouse ◽  
Link Type

Vibrio cholerae the causative agent of cholera, uses a large number of coordinated transcriptional regulatory events to transition from its environmental reservoir to the host intestine, which is its preferred colonization site. Transcription of the mannose-sensitive haemagglutinin pilus (MSHA), which aids the persistence of V. cholerae in aquatic environments, but causes its clearance by host immune defenses, was found to be regulated by a yet unknown mechanism during the infection cycle of V. cholerae . In this study, genomic expression library screening revealed that two regulators, VC1371 and VcRfaH, are able to positively activate the transcription of MSHA operon. VC1371 is localized and active in the cell membrane. Deletion of vc1371 or VcrfaH genes in V. cholerae resulted in less MshA protein production and less efficiency of biofilm formation compared to that in the wild-type strain. An adult mouse model showed that the mutants with vc1371 or VcrfaH deletion colonized less efficiently than the wild-type; the VcrfaH deletion mutant showed less colonization efficiency in the infant mouse model. The findings strongly suggested that the two regulators, namely VC1371 and VcRfaH, which are involved in the regulation of MSHA expression, play an important role in V. cholerae biofilm formation and colonization in mice.

scholarly journals The basis of α-hemolysis Negative Methicillin-resistant Staphylococcus Aureus Isolates from Beijing Children’s Hospital

2021 ◽  
Author(s):  
Lijuan Wang ◽  
Chen Sun ◽  
Suyun Qian ◽  
Yingchao Liu ◽  
Kaihu Yao ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Amino Acid ◽  
Methicillin Resistant Staphylococcus Aureus ◽  
Clonal Complex ◽  
Common Mutation ◽  
Rna Expression ◽  
Single Mutation ◽  
Methicillin Resistant ◽  
Significant Difference ◽  
Important Virulence Factor

Abstract Background. Methicillin-resistant Staphylococcus aureus (MRSA) Clonal Complex 59 (CC59) clone has spread among Chinese children, resulting in many Staphylococcus aureus infections. α-hemolysin (Hlα) is an important virulence factor of Staphylococcus aureus, but little research has been done on CC59 isolates with negative α-hemolysis. Results. During the 4 periods (2009-2011, 2012-2013, 2016, 2017), 291 MRSA isolates were collected. Isolates with β and δ hemolysis accounted for 60.47% among the MRSA isolates in 2009-2011; 56.41% in 2012-2013; 77.14% in 2016; and 56.25% in 2017. most ST59 isolates (94.38%), 9 ST338 isolates (100%) showed β and δ hemolysis, both ST59 and ST338 clone belong to CC59 clone. Twenty-two ST239 isolates (73.33%), 8 ST88 isolates (80%), 4 ST5 isolates (100%), 13 ST22 isolates (92.86%) and 6 ST398 isolates (85.71%) showed α and δ hemolysis. α hemolysin in most clinical isolates is highly conservative, each showed one amino acid locus variation, the most common mutation was threonine at position 275 instead of isoleucine, then glutamic acid replaced aspartic acid at 208. Seventeen ST59 and 2 ST338 isolates had no mutation, 3 ST59 isolates showed single mutation (C448G), and only one ST59 isolate showed multilocus mutation. Other ST typing, such as ST1, ST5, ST88, ST20, ST239 and ST398, all had multilocus mutations, sites were from 3 to 8, no conservative sequence was found among isolates with the same ST typing. The carrying rates of RNA III, Rot, agrA, SarR, SarU and SigB were all over 93%, the carrying rates of SarZ and SarA genes were 41.86% and 34.88% respectively. Trancriptional levels of hlα in isolates showed α and δ hemolysis and β and δ hemolysis were equal. USA300 and R23 produced Hlα, R23 didn’t showed α hemolysis phenotype.Conclusions. Most clinical CC59 isolates from children in China were α hemolysis negative. There was no statistically significant difference in hlα gene and RNA expression, they produced the protein. The reason for the phenotypic deletion probably related to β hemolysin (Hlβ).

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