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Vibrio parahaemolyticus strengthens their virulence through modulation of cellular reactive oxygen species in vitro.

El-Malah SS, Yang Z, Hu M, Li Q, Pan Z, Jiao X - Front Cell Infect Microbiol (2014)

Bottom Line: Our results show that Vp adheres to cell monolayers and can invade non-phagocytic cells.It also survives and persists in non-phagocytic cells by modulating reactive oxygen species (ROS), allowing its replication, and resulting in complete cellular destruction.We conclude that the pathogenicity of Vp is based on its capacities for adhesion and invasion.

View Article: PubMed Central - PubMed

Affiliation: Jiangsu Key Laboratory of Zoonosis/Jiangsu Co-Innovation Center for the Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou University Yangzhou, China.

ABSTRACT
Vibrio parahaemolyticus (Vp) is one of the emergent food-borne pathogens that are commensally associated with various shellfish species throughout the world. It is strictly environmental and many strains are pathogenic to humans. The virulent strains cause distinct diseases, including wound infections, septicemia, and most commonly, acute gastroenteritis, which is acquired through the consumption of raw or undercooked seafood, especially shellfish. Vp has two type three secretion systems (T3SSs), which triggering its cytotoxicity and enterotoxicity via their effectors. To better understand the pathogenesis of Vp, we established a cell infection model in vitro using a non-phagocytic cell line. Caco-2 cells were infected with different strains of Vp (pandemic and non-pandemic strains) and several parameters of cytotoxicity were measured together with adhesion and invasion indices, which reflect the pathogen's virulence. Our results show that Vp adheres to cell monolayers and can invade non-phagocytic cells. It also survives and persists in non-phagocytic cells by modulating reactive oxygen species (ROS), allowing its replication, and resulting in complete cellular destruction. We conclude that the pathogenicity of Vp is based on its capacities for adhesion and invasion. Surprisingly's; enhanced of ROS resistance period could promote the survival of Vp inside the intestinal tract, facilitating tissue infection by repressing the host's oxidative stress response.

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Cytotoxic effects of Vibrio parahaemolyticus on Caco-2 cells. Caco-2 cells were infected with Vibrio parahaemolyticus [MOI] = 100:1, the supernatants were collected at specific time points, and the amounts of LDH released were measured. The percentage of dead cells and the type of death (proportion of apoptotic cells) were determined using annexin V–FITC and flow cytometry (A,C); and (B) LDH release was measured with the CytoTox 96 Non-Radioactive Cytotoxicity Assay (Promega). Percentage cytotoxicity was calculated using the formula: (test LDH release—spontaneous release)/maximal release. Test LDH release is the LDH released after infection with different Vibrio parahaemolyticus strains; spontaneous release is the baseline cell LDH release, with no bacterial infection; and maximal LDH release is the release of LDH when cells are lysed with lysis solution. The data are the means of three independent experiments ± SEM, and statistically significant differences between the mean values were detected with Two-Way ANOVA (*P < 0.05; **P < 0.01; ***P < 0.001).
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Figure 6: Cytotoxic effects of Vibrio parahaemolyticus on Caco-2 cells. Caco-2 cells were infected with Vibrio parahaemolyticus [MOI] = 100:1, the supernatants were collected at specific time points, and the amounts of LDH released were measured. The percentage of dead cells and the type of death (proportion of apoptotic cells) were determined using annexin V–FITC and flow cytometry (A,C); and (B) LDH release was measured with the CytoTox 96 Non-Radioactive Cytotoxicity Assay (Promega). Percentage cytotoxicity was calculated using the formula: (test LDH release—spontaneous release)/maximal release. Test LDH release is the LDH released after infection with different Vibrio parahaemolyticus strains; spontaneous release is the baseline cell LDH release, with no bacterial infection; and maximal LDH release is the release of LDH when cells are lysed with lysis solution. The data are the means of three independent experiments ± SEM, and statistically significant differences between the mean values were detected with Two-Way ANOVA (*P < 0.05; **P < 0.01; ***P < 0.001).

Mentions: We analyzed the cellular contents released during infection by measuring the levels of cytoplasmic LDH released into the media during the infection of Caco-2 with different strains. This release indicates the degree to which the integrity of the host-cell membrane is compromised at different times. The levels of LDH release caused by strains Vp024 and Vp029, followed by strain Vp005, were higher than that caused by RIMD over the entire time course of the experiment. After 3 h, the levels of LDH release stimulated by Vp024 and Vp029 decreased slightly, in contrast to the continuous increase in the cytotoxicity of the other strains (Figure 6B).


Vibrio parahaemolyticus strengthens their virulence through modulation of cellular reactive oxygen species in vitro.

El-Malah SS, Yang Z, Hu M, Li Q, Pan Z, Jiao X - Front Cell Infect Microbiol (2014)

Cytotoxic effects of Vibrio parahaemolyticus on Caco-2 cells. Caco-2 cells were infected with Vibrio parahaemolyticus [MOI] = 100:1, the supernatants were collected at specific time points, and the amounts of LDH released were measured. The percentage of dead cells and the type of death (proportion of apoptotic cells) were determined using annexin V–FITC and flow cytometry (A,C); and (B) LDH release was measured with the CytoTox 96 Non-Radioactive Cytotoxicity Assay (Promega). Percentage cytotoxicity was calculated using the formula: (test LDH release—spontaneous release)/maximal release. Test LDH release is the LDH released after infection with different Vibrio parahaemolyticus strains; spontaneous release is the baseline cell LDH release, with no bacterial infection; and maximal LDH release is the release of LDH when cells are lysed with lysis solution. The data are the means of three independent experiments ± SEM, and statistically significant differences between the mean values were detected with Two-Way ANOVA (*P < 0.05; **P < 0.01; ***P < 0.001).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4269196&req=5

Figure 6: Cytotoxic effects of Vibrio parahaemolyticus on Caco-2 cells. Caco-2 cells were infected with Vibrio parahaemolyticus [MOI] = 100:1, the supernatants were collected at specific time points, and the amounts of LDH released were measured. The percentage of dead cells and the type of death (proportion of apoptotic cells) were determined using annexin V–FITC and flow cytometry (A,C); and (B) LDH release was measured with the CytoTox 96 Non-Radioactive Cytotoxicity Assay (Promega). Percentage cytotoxicity was calculated using the formula: (test LDH release—spontaneous release)/maximal release. Test LDH release is the LDH released after infection with different Vibrio parahaemolyticus strains; spontaneous release is the baseline cell LDH release, with no bacterial infection; and maximal LDH release is the release of LDH when cells are lysed with lysis solution. The data are the means of three independent experiments ± SEM, and statistically significant differences between the mean values were detected with Two-Way ANOVA (*P < 0.05; **P < 0.01; ***P < 0.001).
Mentions: We analyzed the cellular contents released during infection by measuring the levels of cytoplasmic LDH released into the media during the infection of Caco-2 with different strains. This release indicates the degree to which the integrity of the host-cell membrane is compromised at different times. The levels of LDH release caused by strains Vp024 and Vp029, followed by strain Vp005, were higher than that caused by RIMD over the entire time course of the experiment. After 3 h, the levels of LDH release stimulated by Vp024 and Vp029 decreased slightly, in contrast to the continuous increase in the cytotoxicity of the other strains (Figure 6B).

Bottom Line: Our results show that Vp adheres to cell monolayers and can invade non-phagocytic cells.It also survives and persists in non-phagocytic cells by modulating reactive oxygen species (ROS), allowing its replication, and resulting in complete cellular destruction.We conclude that the pathogenicity of Vp is based on its capacities for adhesion and invasion.

View Article: PubMed Central - PubMed

Affiliation: Jiangsu Key Laboratory of Zoonosis/Jiangsu Co-Innovation Center for the Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou University Yangzhou, China.

ABSTRACT
Vibrio parahaemolyticus (Vp) is one of the emergent food-borne pathogens that are commensally associated with various shellfish species throughout the world. It is strictly environmental and many strains are pathogenic to humans. The virulent strains cause distinct diseases, including wound infections, septicemia, and most commonly, acute gastroenteritis, which is acquired through the consumption of raw or undercooked seafood, especially shellfish. Vp has two type three secretion systems (T3SSs), which triggering its cytotoxicity and enterotoxicity via their effectors. To better understand the pathogenesis of Vp, we established a cell infection model in vitro using a non-phagocytic cell line. Caco-2 cells were infected with different strains of Vp (pandemic and non-pandemic strains) and several parameters of cytotoxicity were measured together with adhesion and invasion indices, which reflect the pathogen's virulence. Our results show that Vp adheres to cell monolayers and can invade non-phagocytic cells. It also survives and persists in non-phagocytic cells by modulating reactive oxygen species (ROS), allowing its replication, and resulting in complete cellular destruction. We conclude that the pathogenicity of Vp is based on its capacities for adhesion and invasion. Surprisingly's; enhanced of ROS resistance period could promote the survival of Vp inside the intestinal tract, facilitating tissue infection by repressing the host's oxidative stress response.

Show MeSH
Related in: MedlinePlus