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Rhodococcus equi's extreme resistance to hydrogen peroxide is mainly conferred by one of its four catalase genes.

Bidaud P, Hébert L, Barbey C, Appourchaux AC, Torelli R, Sanguinetti M, Laugier C, Petry S - PLoS ONE (2012)

Bottom Line: Moreover, katA deletion seems to significantly affect the ability of R. equi to survive within murine macrophages.In untreated bacteria, katB, katC and katD were overexpressed from 4.3 to 17.5 times in the stationary compared to the exponential phase.Taken together, our results show that KatA is the major catalase involved in the extreme H(2)O(2) resistance capability of R. equi.

View Article: PubMed Central - PubMed

Affiliation: Dozulé Laboratory for Equine Diseases, Unit Bacteriology and Parasitology, ANSES, Goustranville, France.

ABSTRACT
Rhodococcus equi is one of the most widespread causes of disease in foals aged from 1 to 6 months. R. equi possesses antioxidant defense mechanisms to protect it from reactive oxygen metabolites such as hydrogen peroxide (H(2)O(2)) generated during the respiratory burst of phagocytic cells. These defense mechanisms include enzymes such as catalase, which detoxify hydrogen peroxide. Recently, an analysis of the R. equi 103 genome sequence revealed the presence of four potential catalase genes. We first constructed ΔkatA-, ΔkatB-, ΔkatC-and ΔkatD-deficient mutants to study the ability of R. equi to survive exposure to H(2)O(2)in vitro and within mouse peritoneal macrophages. Results showed that ΔkatA and, to a lesser extent ΔkatC, were affected by 80 mM H(2)O(2). Moreover, katA deletion seems to significantly affect the ability of R. equi to survive within murine macrophages. We finally investigated the expression of the four catalases in response to H(2)O(2) assays with a real time PCR technique. Results showed that katA is overexpressed 367.9 times (± 122.6) in response to exposure to 50 mM of H(2)O(2) added in the stationary phase, and 3.11 times (± 0.59) when treatment was administered in the exponential phase. In untreated bacteria, katB, katC and katD were overexpressed from 4.3 to 17.5 times in the stationary compared to the exponential phase. Taken together, our results show that KatA is the major catalase involved in the extreme H(2)O(2) resistance capability of R. equi.

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ΔkatA is the most susceptible to elimination by macrophages.Mice were infected with 107 to 108 cells of R. equi WT, ΔkatA, ΔkatB, ΔkatC or ΔkatD by intraperitoneal injection. Intramacrophages bacteria viability was evaluated 24-, 48-and 72 h post infection. The data are the mean numbers of viable intracellular bacteria per 105 macrophages ± standard deviations (error bars) for three independent experiments with three wells in each experiment.
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pone-0042396-g003: ΔkatA is the most susceptible to elimination by macrophages.Mice were infected with 107 to 108 cells of R. equi WT, ΔkatA, ΔkatB, ΔkatC or ΔkatD by intraperitoneal injection. Intramacrophages bacteria viability was evaluated 24-, 48-and 72 h post infection. The data are the mean numbers of viable intracellular bacteria per 105 macrophages ± standard deviations (error bars) for three independent experiments with three wells in each experiment.

Mentions: During phagocytosis, phagocytic cells generate superoxide and other ROS involved in antibacterial activity [16]. This study compared the intracellular survival of the WT and ΔkatA, ΔkatB, ΔkatC and ΔkatD strains inside infected mouse peritoneal macrophages (Figure 3). No significant difference was observed in the levels of R. equi strains recovered 8 h post infection, suggesting that the different strains possessed similar abilities to infect macrophages (Figure 3). ΔkatA was shown to be the most susceptible mutant, completely eliminated by macrophages 72 h post infection (P<0.05). ΔkatB, ΔkatC and ΔkatD were shown to be more susceptible than the WT strain to macrophage killing at 48-and 72-h time points (P<0.05) but were not completely eliminated from macrophages 72 h post infection. The survival rate of each strain was similar by 24 h post infection but the WT strain’s ability to survive intracellularly was greater than that of ΔkatB, ΔkatC and ΔkatD at 48-and 72-h time points (P<0.05). The inability of ΔkatA to survive in macrophages beyond 72 h reveals KatA’s involvement in the intramacrophage resistance of R. equi to H2O2.


Rhodococcus equi's extreme resistance to hydrogen peroxide is mainly conferred by one of its four catalase genes.

Bidaud P, Hébert L, Barbey C, Appourchaux AC, Torelli R, Sanguinetti M, Laugier C, Petry S - PLoS ONE (2012)

ΔkatA is the most susceptible to elimination by macrophages.Mice were infected with 107 to 108 cells of R. equi WT, ΔkatA, ΔkatB, ΔkatC or ΔkatD by intraperitoneal injection. Intramacrophages bacteria viability was evaluated 24-, 48-and 72 h post infection. The data are the mean numbers of viable intracellular bacteria per 105 macrophages ± standard deviations (error bars) for three independent experiments with three wells in each experiment.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3412833&req=5

pone-0042396-g003: ΔkatA is the most susceptible to elimination by macrophages.Mice were infected with 107 to 108 cells of R. equi WT, ΔkatA, ΔkatB, ΔkatC or ΔkatD by intraperitoneal injection. Intramacrophages bacteria viability was evaluated 24-, 48-and 72 h post infection. The data are the mean numbers of viable intracellular bacteria per 105 macrophages ± standard deviations (error bars) for three independent experiments with three wells in each experiment.
Mentions: During phagocytosis, phagocytic cells generate superoxide and other ROS involved in antibacterial activity [16]. This study compared the intracellular survival of the WT and ΔkatA, ΔkatB, ΔkatC and ΔkatD strains inside infected mouse peritoneal macrophages (Figure 3). No significant difference was observed in the levels of R. equi strains recovered 8 h post infection, suggesting that the different strains possessed similar abilities to infect macrophages (Figure 3). ΔkatA was shown to be the most susceptible mutant, completely eliminated by macrophages 72 h post infection (P<0.05). ΔkatB, ΔkatC and ΔkatD were shown to be more susceptible than the WT strain to macrophage killing at 48-and 72-h time points (P<0.05) but were not completely eliminated from macrophages 72 h post infection. The survival rate of each strain was similar by 24 h post infection but the WT strain’s ability to survive intracellularly was greater than that of ΔkatB, ΔkatC and ΔkatD at 48-and 72-h time points (P<0.05). The inability of ΔkatA to survive in macrophages beyond 72 h reveals KatA’s involvement in the intramacrophage resistance of R. equi to H2O2.

Bottom Line: Moreover, katA deletion seems to significantly affect the ability of R. equi to survive within murine macrophages.In untreated bacteria, katB, katC and katD were overexpressed from 4.3 to 17.5 times in the stationary compared to the exponential phase.Taken together, our results show that KatA is the major catalase involved in the extreme H(2)O(2) resistance capability of R. equi.

View Article: PubMed Central - PubMed

Affiliation: Dozulé Laboratory for Equine Diseases, Unit Bacteriology and Parasitology, ANSES, Goustranville, France.

ABSTRACT
Rhodococcus equi is one of the most widespread causes of disease in foals aged from 1 to 6 months. R. equi possesses antioxidant defense mechanisms to protect it from reactive oxygen metabolites such as hydrogen peroxide (H(2)O(2)) generated during the respiratory burst of phagocytic cells. These defense mechanisms include enzymes such as catalase, which detoxify hydrogen peroxide. Recently, an analysis of the R. equi 103 genome sequence revealed the presence of four potential catalase genes. We first constructed ΔkatA-, ΔkatB-, ΔkatC-and ΔkatD-deficient mutants to study the ability of R. equi to survive exposure to H(2)O(2)in vitro and within mouse peritoneal macrophages. Results showed that ΔkatA and, to a lesser extent ΔkatC, were affected by 80 mM H(2)O(2). Moreover, katA deletion seems to significantly affect the ability of R. equi to survive within murine macrophages. We finally investigated the expression of the four catalases in response to H(2)O(2) assays with a real time PCR technique. Results showed that katA is overexpressed 367.9 times (± 122.6) in response to exposure to 50 mM of H(2)O(2) added in the stationary phase, and 3.11 times (± 0.59) when treatment was administered in the exponential phase. In untreated bacteria, katB, katC and katD were overexpressed from 4.3 to 17.5 times in the stationary compared to the exponential phase. Taken together, our results show that KatA is the major catalase involved in the extreme H(2)O(2) resistance capability of R. equi.

Show MeSH
Related in: MedlinePlus