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| 2. |
- Collin, Betty, 1976, et al.
(author)
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The origin of vibrio cholerae influences uptake and persistence in the blue mussel Mytilus edulis
- 2012
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In: Journal of Shellfish Research. - : National Shellfisheries Association. - 0730-8000 .- 1943-6319. ; 31:1, s. 87-92
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Journal article (peer-reviewed)abstract
- Vibrio cholerae may cause diarrheal diseases and wound infections, both of which have the potential to be fatal. Transmission to humans is often linked to consumption of contaminated shellfish/drinking water or dermal exposure to water (e.g. when swimming). In this study, we investigated whether different isolates of Vibrio cholerae differ in terms of accumulation, persistence, and viability when encountering blue mussels (Mytilus edulis). Mussel uptake and elimination of three different V. cholerae strains were compared: one fatal clinical non-O1/O139 isolate, one highly potent El Tor biotype, and one marine strain isolated from blue mussels. The results showed that the uptake of the marine strain was significantly higher than the clinical strain, but the elimination process of the marine strain was also more efficient. The El Tor strain was not at all ingested by the mussels. In addition, the survival of bacteria when incubated together with M. edulis hemocytes was tested in vitro. The viability of clinical strains was unaffected by the presence of hemocytes, and the marine strains were even more resistant and able to multiply. We conclude that the highly virulent El Tor biotype was not taken up by the mussels and could thereby escape the mussels' elimination process. The potentially fatal non-O1/O139 V. cholerae strain may accumulate in low numbers, but could be very persistent in mussels.
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| 3. |
- Ishikawa, Takahiko, et al.
(author)
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Quorum sensing regulation of the two hcp alleles in Vibrio cholerae O1 strains
- 2009
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In: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 4:8
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Journal article (peer-reviewed)abstract
- BACKGROUND: The type VI secretion system (T6SS) has emerged as a protein secretion system important to several gram-negative bacterial species. One of the common components of the system is Hcp, initially described as a hemolysin co-regulated protein in a serotype O17 strain of Vibrio cholerae. Homologs to V. cholerae hcp genes have been found in all characterized type VI secretion systems and they are present also in the serotype O1 strains of V. cholerae that are the cause of cholera diseases but seemed to have non-functional T6SS.METHODOLOGY/PRINCIPAL FINDINGS: The serotype O1 V. cholerae strain A1552 was shown to express detectable levels of Hcp as determined by immunoblot analyses using polyclonal anti-Hcp antiserum. We found that the expression of Hcp was growth phase dependent. The levels of Hcp in quorum sensing deficient mutants of V. cholerae were compared with the levels in wild type V. cholerae O1 strain A1552. The expression of Hcp was positively and negatively regulated by the quorum sensing regulators HapR and LuxO, respectively. In addition, we observed that expression of Hcp was dependent on the cAMP-CRP global transcriptional regulatory complex and required the RpoN sigma factor.CONCLUSION/SIGNIFICANCE: Our results show that serotype O1 strains of V. cholerae do express Hcp which is regarded as one of the important T6SS components and is one of the secreted substrates in non-O1 non-O139 V. cholerae isolates. We found that expression of Hcp was strictly regulated by the quorum sensing system in the V. cholerae O1 strain. In addition, the expression of Hcp required the alternative sigma factor RpoN and the cAMP-CRP global regulatory complex. Interestingly, the environmental isolates of V. cholerae O1 strains that showed higher levels of the HapR quorum sensing regulator in comparison with our laboratory standard serotype O1 strain A1552 where also expressing higher levels of Hcp.
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| 4. |
- Lindmark, Barbro, 1963-, et al.
(author)
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Evaluation of the presence of virulence-associated factors in Vibrio cholerae non-O1 non-O139 isolates from clinical and environmental sources
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Other publication (other academic/artistic)abstract
- The broad group non-O1 non-O139 Vibrio cholerae presumably causes clinical diseases due to properties distinct from V. cholerae O1 and O139 serogroups in which cholera toxin is the hallmark of the infection. In this study, we screened for the presence of virulence-associated factors in V. cholerae non-O1 non-O139 strains isolated in Sweden. Six isolates from patients with different clinical manifestations and two environmental isolates were studied. None of the V. cholerae non-O1 non-O139 isolates carried the ctx or tcpA genes, but gene sequences for other putative virulence factors such as OmpU, cytolysin (VCC) and RTX were present. Significant differences in serum resistance were observed among the isolates independent of their encapsulation. The isolates were tested on Caenorhabditis elegans as an alternative host for analysis of factors associated with protection from natural predator grazing and environmental survival of V. cholerae. Three isolates caused lethality to C. elegans and also showed the strongest ability to resist killing by serum. We also observed that actin cross-linking activity and the level of protease secretion were different among the strains.
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| 5. |
- Lindmark, Barbro, 1963-
(author)
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Modulators of Vibrio cholerae predator interaction and virulence
- 2009
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Doctoral thesis (other academic/artistic)abstract
- Vibrio cholerae, the causal agent of cholera typically encodes two critical virulence factors: cholera toxin (CT), which is primarily responsible for the diarrhoeal purge, and toxin-co-regulated pilus (TCP), an essential colonisation factor. Nontoxigenic strains expressing TCP can efficiently acquire the CT gene through lysogenic conversion with CTXΦ, a filamentous phage that encodes CT and uses TCP as a receptor. V. cholerae is a Gram-negative bacterium and a natural inhabitant of estuarine and coastal waters throughout both temperate and tropical regions of the world. In the aquatic environment, V. cholerae encounters several environmental stresses, such as change in salinity, UV stress, nutrient limitation, temperature fluctuations, viral infections and protozoan predation. To fully understand the pathogenic and virulence potential of V. cholerae, knowledge is required of its interactions with, not only human, but also environmental factors. By using the nematode Caenorhabditis elegans as host model, we were able to identify a previously uncharacterised protein, the extracellular protease PrtV. PrtV was shown to be required for the killing of. elegans and also necessary for survival from grazing by the ciliate Tetrahymena pyriformis and the flagellate Cafeteria roenbergensis. The PrtV protein, which belongs to a M6 family of metallopeptidases was cloned and purified for further characterisations. The purified PrtV was cytotoxic against the human intestinal cell line HCT8. By using human blood plasma, fibrinogen, fibronectin and plasminogen were identified as candidate substrates for the PrtV protease. Outer membrane vesicles (OMVs) are released to the surroundings by most Gram-negative bacteria through “bulging and pinching” of the outer membrane. OMVs have been shown to contain many virulence factors important in pathogenesis. Therefore, we investigated the association of PrtV with OMVs. PrtV was not associated with OMVs from the wild type O1 strain. In contrast, in an LPS mutant lacking two sugar chains in the core oligosaccharide PrtV was found to be associated with the OMVs. The OMV-associated PrtV was shown to be proteolytically and cytotoxically active. V. cholerae strains are grouped into >200 serogroups. Only the O1 and O139 serogroups have been associated with pandemic cholera, a severe diarrhoeal disease. All other serogroups are collectively referred to as non-O1 non-O139 V. cholerae. Non-O1 non-O139 V. cholerae can cause gastroenteritis and extraintestinal infections, but unlike O1 and O139 strains of V. cholerae, little is known about the virulence gene content and their potential to become human pathogens. We analysed clinical and environmental non-O1 non-O139 isolates for their putative virulence traits. None of them carry the genes encoding CT or the TCP, but other putative virulence factors were present in these isolates. The incidence of serum resistance was found to vary considerably and was independent of encapsulation. Three strains were strongly serum-resistant, and these same strains could also kill C. elegans.
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| 6. |
- Lindmark, Barbro, 1963-, et al.
(author)
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Outer membrane vesicle-mediated release of cytolethal distending toxin (CDT) from Campylobacter jejuni
- 2009
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In: BMC Microbiology. - : Springer Science and Business Media LLC. - 1471-2180. ; 16:9, s. 220-
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Journal article (peer-reviewed)abstract
- BACKGROUND: Cytolethal distending toxin (CDT) is one of the well-characterized virulence factors of Campylobacter jejuni, but it is unknown how CDT becomes surface-exposed or is released from the bacterium to the surrounding environment.RESULTS: Our data suggest that CDT is secreted to the bacterial culture supernatant via outer membrane vesicles (OMVs) released from the bacteria. All three subunits (the CdtA, CdtB, and CdtC proteins) were detected by immunogold labeling and electron microscopy of OMVs. Subcellular fractionation of the bacteria indicated that, apart from the majority of CDT detected in the cytoplasmic compartment, appreciable amounts (20-50%) of the cellular pool of CDT proteins were present in the periplasmic compartment. In the bacterial culture supernatant, we found that a majority of the extracellular CDT was tightly associated with the OMVs. Isolated OMVs could exert the cell distending effects typical of CDT on a human intestinal cell line, indicating that CDT is present there in a biologically active form.CONCLUSION: Our results strongly suggest that the release of outer membrane vesicles is functioning as a route of C. jejuni to deliver all the subunits of CDT toxin (CdtA, CdtB, and CdtC) to the surrounding environment, including infected host tissue.
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| 7. |
- Lindmark, Barbro, 1963-, et al.
(author)
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Role of LPS in vesicle mediated export of Vibrio cholerae PrtV protease
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Other publication (other academic/artistic)abstract
- Gram negative bacteria produce outer membrane vesicles (OMVs) during normal bacterial growth. Recent studies have shown that OMVs can transport biologically active toxins and enzymes to the environment and into the host. We have initiated analysis of OMV associated proteins in V. cholerae. In this study, V. cholerae wild type strain P27459 and the O-antigen ligase mutant waaL, which lacks the O-antigen of the LPS were analyzed for the OMV associated release of the secreted protease PrtV. OMVs from the waaL mutant showed a higher amount of associated secreted PrtV protein than the OMVs from wild type V. cholerae indicating that LPS might be involved in vesicle association of the PrtV protein. We showed that the OMV associated PrtV protein is biologically active.
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| 8. |
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| 9. |
- Ou, Gangwei, et al.
(author)
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Vibrio cholerae cytolysin causes an inflammatory response in human intestinal epithelial cells that is modulated by the PrtV protease
- 2009
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In: PLOS ONE. - San Francisco : Public Library of Science. - 1932-6203. ; 4:11
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Journal article (peer-reviewed)abstract
- We suggest that VCC is capable of causing an inflammatory response characterized by increased permeability and production of IL-8 and TNF-alpha in tight monolayers. Pure VCC at a concentration of 160 ng/ml caused an inflammatory response that reached the magnitude of that caused by Vibrio-secreted factors, while higher concentrations caused epithelial cell death. The inflammatory response was totally abolished by treatment with PrtV. The findings suggest that low doses of VCC initiate a local immune defense reaction while high doses lead to intestinal epithelial lesions. Furthermore, VCC is indeed a substrate for PrtV and PrtV seems to execute an environment-dependent modulation of the activity of VCC that may be the cause of V. cholerae reactogenicity.
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| 10. |
- Rompikuntal, Pramod Kumar, et al.
(author)
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Outer membrane vesicle-mediated export of PrtV protease from Vibrio cholerae
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Other publication (other academic/artistic)abstract
- Background: Gram-negative bacteria release large amounts of outer membrane vesicles (OMVs) during normal growth. OMVs from pathogenic bacteria are known to carry different biologically active toxins and enzymes into the surrounding environment. We hypothesized that OMVs may therefore be able to mediate the transport of bacterial products into host cells. We present here an analysis of the V. cholerae OMV-associated virulence factor PrtV. Methodology/Principal Findings: We observed that PrtV, a M6 family, zinc-binding metalloprotease, is secreted from V. cholerae wild type strain C6706 into the culture supernatant in association with OMVs. The association of PrtV with OMVs was determined by immunoblotting and electron microscopy using immunogold labeling. In addition, we observed that the PKD-domain(s) of PrtV has a role in the protein’s association with OMVs. We also demonstrated that OMV-associated PrtV was biologically active because HCT8 cells treated with OMVs from the wild type V. cholerae strain C6706 exhibited altered morphology, whereas cells treated with OMVs from the prtV isogenic mutant showed no morphological changes. Moreover, our data suggest that OMV-associated PrtV might be transported into target eukaryotic cells by a vesicle fusion mechanism in association with lipid raft microdomains in the plasma membrane.Conclusion/Significance: Our findings suggest that OMVs can deliver biologically active PrtV into target host cells.
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