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The monofunctional catalase KatE of Xanthomonas axonopodis pv. citri is required for full virulence in citrus plants.

Tondo ML, Petrocelli S, Ottado J, Orellano EG - PLoS ONE (2010)

Bottom Line: Moreover, Xac catalases expression pattern is modified in response to any stimuli associated with the plant or the microenvironment it provides.The catalase KatE has been shown to have an important function for the colonization and survival of the bacterium in the citrus plant during the pathogenic process.Our work provides the first genetic evidence to support a monofunctional catalase as a virulence factor in Xac.

View Article: PubMed Central - PubMed

Affiliation: Molecular Biology Division, Facultad de Ciencias Bioquímicas y Farmacéuticas, Instituto de Biología Molecular y Celular de Rosario, Consejo Nacional de Investigaciones Científicas y Técnicas, Universidad Nacional de Rosario, Rosario, Argentina.

ABSTRACT

Background: Xanthomonas axonopodis pv. citri (Xac) is an obligate aerobic phytopathogen constantly exposed to hydrogen peroxide produced by normal aerobic respiration and by the plant defense response during plant-pathogen interactions. Four putative catalase genes have been identified in silico in the Xac genome, designated as katE, catB, srpA (monofunctional catalases) and katG (bifunctional catalase).

Methodology/principal findings: Xac catalase activity was analyzed using native gel electrophoresis and semi-quantitative RT-PCR. We demonstrated that the catalase activity pattern was regulated in different growth stages displaying the highest levels during the stationary phase. KatE was the most active catalase in this phase of growth. At this stage cells were more resistant to hydrogen peroxide as was determined by the analysis of CFU after the exposition to different H(2)O(2) concentrations. In addition, Xac exhibited an adaptive response to hydrogen peroxide, displaying higher levels of catalase activity and H(2)O(2) resistance after treatment with sub-lethal concentrations of the oxidant. In the plant-like medium XVM2 the expression of KatE was strongly induced and in this medium Xac was more resistant to H(2)O(2). A XackatE mutant strain was constructed by insertional mutagenesis. We observed that catalase induction in stationary phase was lost meanwhile the adaptive response to peroxide was maintained in this mutant. Finally, the XackatE strain was assayed in planta during host plant interaction rendering a less aggressive phenotype with a minor canker formation.

Conclusions: Our results confirmed that in contrast to other Xanthomonas species, Xac catalase-specific activity is induced during the stationary phase of growth in parallel with the bacterial resistance to peroxide challenge. Moreover, Xac catalases expression pattern is modified in response to any stimuli associated with the plant or the microenvironment it provides. The catalase KatE has been shown to have an important function for the colonization and survival of the bacterium in the citrus plant during the pathogenic process. Our work provides the first genetic evidence to support a monofunctional catalase as a virulence factor in Xac.

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Effect of katE disruption on pathogenicity.Xac WT (WT), XackatE (katE−) and cXackatE (ckatE) cells were inoculated at 105 CFU ml−1 in 10 mM MgCl2 into the intercellular spaces of fully expanded orange leaves. (A) A representative leaf 20 days after inoculation is shown. Left panel, adaxial side; right panel, abaxial side. Dashed lines indicate the infiltrated area. (B) Bacterial growth of Xac cells in orange leaves. Values represent means of three independent samples; error bars represent standard deviations.
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pone-0010803-g008: Effect of katE disruption on pathogenicity.Xac WT (WT), XackatE (katE−) and cXackatE (ckatE) cells were inoculated at 105 CFU ml−1 in 10 mM MgCl2 into the intercellular spaces of fully expanded orange leaves. (A) A representative leaf 20 days after inoculation is shown. Left panel, adaxial side; right panel, abaxial side. Dashed lines indicate the infiltrated area. (B) Bacterial growth of Xac cells in orange leaves. Values represent means of three independent samples; error bars represent standard deviations.

Mentions: In order to assess the physiological role of KatE during the infection process, the mutant strain was tested for its ability to trigger disease in citrus leaves. Both wild-type bacteria and XackatE produced typical canker lesions upon infiltration at a concentration of 105 CFU ml−1, with no differences in the time of appearance of the first symptoms (water soaking). However, the magnitude of the lesions and the number of cankers were significantly diminished in the mutant strain compared to wild-type bacteria, even though the infiltration areas and the bacterial densities were equivalent for both strains. On the other hand, infiltration with the cXackatE strain caused the same symptoms and a similar percentage of necrotic area than wild-type cells (Figure 8A).


The monofunctional catalase KatE of Xanthomonas axonopodis pv. citri is required for full virulence in citrus plants.

Tondo ML, Petrocelli S, Ottado J, Orellano EG - PLoS ONE (2010)

Effect of katE disruption on pathogenicity.Xac WT (WT), XackatE (katE−) and cXackatE (ckatE) cells were inoculated at 105 CFU ml−1 in 10 mM MgCl2 into the intercellular spaces of fully expanded orange leaves. (A) A representative leaf 20 days after inoculation is shown. Left panel, adaxial side; right panel, abaxial side. Dashed lines indicate the infiltrated area. (B) Bacterial growth of Xac cells in orange leaves. Values represent means of three independent samples; error bars represent standard deviations.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2875408&req=5

pone-0010803-g008: Effect of katE disruption on pathogenicity.Xac WT (WT), XackatE (katE−) and cXackatE (ckatE) cells were inoculated at 105 CFU ml−1 in 10 mM MgCl2 into the intercellular spaces of fully expanded orange leaves. (A) A representative leaf 20 days after inoculation is shown. Left panel, adaxial side; right panel, abaxial side. Dashed lines indicate the infiltrated area. (B) Bacterial growth of Xac cells in orange leaves. Values represent means of three independent samples; error bars represent standard deviations.
Mentions: In order to assess the physiological role of KatE during the infection process, the mutant strain was tested for its ability to trigger disease in citrus leaves. Both wild-type bacteria and XackatE produced typical canker lesions upon infiltration at a concentration of 105 CFU ml−1, with no differences in the time of appearance of the first symptoms (water soaking). However, the magnitude of the lesions and the number of cankers were significantly diminished in the mutant strain compared to wild-type bacteria, even though the infiltration areas and the bacterial densities were equivalent for both strains. On the other hand, infiltration with the cXackatE strain caused the same symptoms and a similar percentage of necrotic area than wild-type cells (Figure 8A).

Bottom Line: Moreover, Xac catalases expression pattern is modified in response to any stimuli associated with the plant or the microenvironment it provides.The catalase KatE has been shown to have an important function for the colonization and survival of the bacterium in the citrus plant during the pathogenic process.Our work provides the first genetic evidence to support a monofunctional catalase as a virulence factor in Xac.

View Article: PubMed Central - PubMed

Affiliation: Molecular Biology Division, Facultad de Ciencias Bioquímicas y Farmacéuticas, Instituto de Biología Molecular y Celular de Rosario, Consejo Nacional de Investigaciones Científicas y Técnicas, Universidad Nacional de Rosario, Rosario, Argentina.

ABSTRACT

Background: Xanthomonas axonopodis pv. citri (Xac) is an obligate aerobic phytopathogen constantly exposed to hydrogen peroxide produced by normal aerobic respiration and by the plant defense response during plant-pathogen interactions. Four putative catalase genes have been identified in silico in the Xac genome, designated as katE, catB, srpA (monofunctional catalases) and katG (bifunctional catalase).

Methodology/principal findings: Xac catalase activity was analyzed using native gel electrophoresis and semi-quantitative RT-PCR. We demonstrated that the catalase activity pattern was regulated in different growth stages displaying the highest levels during the stationary phase. KatE was the most active catalase in this phase of growth. At this stage cells were more resistant to hydrogen peroxide as was determined by the analysis of CFU after the exposition to different H(2)O(2) concentrations. In addition, Xac exhibited an adaptive response to hydrogen peroxide, displaying higher levels of catalase activity and H(2)O(2) resistance after treatment with sub-lethal concentrations of the oxidant. In the plant-like medium XVM2 the expression of KatE was strongly induced and in this medium Xac was more resistant to H(2)O(2). A XackatE mutant strain was constructed by insertional mutagenesis. We observed that catalase induction in stationary phase was lost meanwhile the adaptive response to peroxide was maintained in this mutant. Finally, the XackatE strain was assayed in planta during host plant interaction rendering a less aggressive phenotype with a minor canker formation.

Conclusions: Our results confirmed that in contrast to other Xanthomonas species, Xac catalase-specific activity is induced during the stationary phase of growth in parallel with the bacterial resistance to peroxide challenge. Moreover, Xac catalases expression pattern is modified in response to any stimuli associated with the plant or the microenvironment it provides. The catalase KatE has been shown to have an important function for the colonization and survival of the bacterium in the citrus plant during the pathogenic process. Our work provides the first genetic evidence to support a monofunctional catalase as a virulence factor in Xac.

Show MeSH
Related in: MedlinePlus