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Nitric oxide antagonizes the acid tolerance response that protects Salmonella against innate gastric defenses.

Bourret TJ, Porwollik S, McClelland M, Zhao R, Greco T, Ischiropoulos H, Vázquez-Torres A - PLoS ONE (2008)

Bottom Line: NO prevented Salmonella from mounting a de novo ATR, but was unable to suppress an already functional protective response, suggesting that RNS target regulatory cascades but not their effectors.Transcriptional and translational analyses revealed that the PhoPQ signaling cascade is a critical ATR target of NO in rapidly growing Salmonella.Since PhoPQ-regulated acid resistance is widespread in enteric pathogens, the RNS-mediated inhibition of the Salmonella ATR described herein may represent a common component of innate host defenses.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, University of Colorado Health Sciences Center, Aurora, Colorado, United States of America.

ABSTRACT

Background: Reactive nitrogen species (RNS) derived from dietary and salivary inorganic nitrogen oxides foment innate host defenses associated with the acidity of the stomach. The mechanisms by which these reactive species exert antimicrobial activity in the gastric lumen are, however, poorly understood.

Methodology/principal findings: The genetically tractable acid tolerance response (ATR) that enables enteropathogens to survive harsh acidity was screened for signaling pathways responsive to RNS. The nitric oxide (NO) donor spermine NONOate derepressed the Fur regulon that controls secondary lines of resistance against organic acids. Despite inducing a Fur-mediated adaptive response, acidified RNS largely repressed oral virulence as demonstrated by the fact that Salmonella bacteria exposed to NO donors during mildly acidic conditions were shed in low amounts in feces and exhibited ameliorated oral virulence. NO prevented Salmonella from mounting a de novo ATR, but was unable to suppress an already functional protective response, suggesting that RNS target regulatory cascades but not their effectors. Transcriptional and translational analyses revealed that the PhoPQ signaling cascade is a critical ATR target of NO in rapidly growing Salmonella. Inhibition of PhoPQ signaling appears to contribute to most of the NO-mediated abrogation of the ATR in log phase bacteria, because the augmented acid sensitivity of phoQ-deficient Salmonella was not further enhanced after RNS treatment.

Conclusions/significance: Since PhoPQ-regulated acid resistance is widespread in enteric pathogens, the RNS-mediated inhibition of the Salmonella ATR described herein may represent a common component of innate host defenses.

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Related in: MedlinePlus

RNS suppress the increased oral virulence of ATR-adapted Salmonella.C57BL/6 mice were inoculated orally with ∼5×105 CFU of NonAd, Ad and Ad+NO Salmonella grown as described in figure 1. (A) Fecal shedding of nalidixic acid resistant Salmonella was monitored in individual mice for 3 days after oral inoculation. Panel B shows the % of mice that survived after oral challenge with Salmonella. Data represent 10 mice per group from 2 separate experiments.
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pone-0001833-g003: RNS suppress the increased oral virulence of ATR-adapted Salmonella.C57BL/6 mice were inoculated orally with ∼5×105 CFU of NonAd, Ad and Ad+NO Salmonella grown as described in figure 1. (A) Fecal shedding of nalidixic acid resistant Salmonella was monitored in individual mice for 3 days after oral inoculation. Panel B shows the % of mice that survived after oral challenge with Salmonella. Data represent 10 mice per group from 2 separate experiments.

Mentions: To determine whether the acid sensitivity seen in Salmonella treated with spermine NONOate affects passage through the gastrointestinal tract, C57BL/6 mice were infected per orally with ∼5×105 CFU of the nalidixic acid resistant Salmonella strain IR175 grown under the acid tolerance assay conditions described above. This inoculum, which has been used by multiple investigators, is close to the oral LD50 for wild-type S. Typhimurium [31]–[33]. Shedding of Salmonella in the feces shortly after oral challenge was used as a marker for successful passage through the gastrointestinal tract. A higher percentage of mice challenged with Salmonella adapted in EG medium, pH 4.4 shed bacteria in feces than controls challenged with nonadapted bacteria (one-way ANOVA, p<0.01) (fig. 3A). To the best of our knowledge, these data demonstrate for the first time that bacteria that have mounted an ATR exhibit higher oral virulence. Treatment of Salmonella with 250 µM spermine NONOate during culture in EG medium, pH 4.4 significantly (one-way ANOVA, p<0.01) abrogated the increased shedding seen in adapted controls (fig. 3A). Fecal shedding of Salmonella reflected oral virulence. About 10% of mice infected with acid adapted Salmonella survived after 9 days of infection, whereas survival rates reached 60% to 80% in mice infected with either nonadapted or spermine NONOate-treated, adapted controls (fig. 3B).


Nitric oxide antagonizes the acid tolerance response that protects Salmonella against innate gastric defenses.

Bourret TJ, Porwollik S, McClelland M, Zhao R, Greco T, Ischiropoulos H, Vázquez-Torres A - PLoS ONE (2008)

RNS suppress the increased oral virulence of ATR-adapted Salmonella.C57BL/6 mice were inoculated orally with ∼5×105 CFU of NonAd, Ad and Ad+NO Salmonella grown as described in figure 1. (A) Fecal shedding of nalidixic acid resistant Salmonella was monitored in individual mice for 3 days after oral inoculation. Panel B shows the % of mice that survived after oral challenge with Salmonella. Data represent 10 mice per group from 2 separate experiments.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2266805&req=5

pone-0001833-g003: RNS suppress the increased oral virulence of ATR-adapted Salmonella.C57BL/6 mice were inoculated orally with ∼5×105 CFU of NonAd, Ad and Ad+NO Salmonella grown as described in figure 1. (A) Fecal shedding of nalidixic acid resistant Salmonella was monitored in individual mice for 3 days after oral inoculation. Panel B shows the % of mice that survived after oral challenge with Salmonella. Data represent 10 mice per group from 2 separate experiments.
Mentions: To determine whether the acid sensitivity seen in Salmonella treated with spermine NONOate affects passage through the gastrointestinal tract, C57BL/6 mice were infected per orally with ∼5×105 CFU of the nalidixic acid resistant Salmonella strain IR175 grown under the acid tolerance assay conditions described above. This inoculum, which has been used by multiple investigators, is close to the oral LD50 for wild-type S. Typhimurium [31]–[33]. Shedding of Salmonella in the feces shortly after oral challenge was used as a marker for successful passage through the gastrointestinal tract. A higher percentage of mice challenged with Salmonella adapted in EG medium, pH 4.4 shed bacteria in feces than controls challenged with nonadapted bacteria (one-way ANOVA, p<0.01) (fig. 3A). To the best of our knowledge, these data demonstrate for the first time that bacteria that have mounted an ATR exhibit higher oral virulence. Treatment of Salmonella with 250 µM spermine NONOate during culture in EG medium, pH 4.4 significantly (one-way ANOVA, p<0.01) abrogated the increased shedding seen in adapted controls (fig. 3A). Fecal shedding of Salmonella reflected oral virulence. About 10% of mice infected with acid adapted Salmonella survived after 9 days of infection, whereas survival rates reached 60% to 80% in mice infected with either nonadapted or spermine NONOate-treated, adapted controls (fig. 3B).

Bottom Line: NO prevented Salmonella from mounting a de novo ATR, but was unable to suppress an already functional protective response, suggesting that RNS target regulatory cascades but not their effectors.Transcriptional and translational analyses revealed that the PhoPQ signaling cascade is a critical ATR target of NO in rapidly growing Salmonella.Since PhoPQ-regulated acid resistance is widespread in enteric pathogens, the RNS-mediated inhibition of the Salmonella ATR described herein may represent a common component of innate host defenses.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, University of Colorado Health Sciences Center, Aurora, Colorado, United States of America.

ABSTRACT

Background: Reactive nitrogen species (RNS) derived from dietary and salivary inorganic nitrogen oxides foment innate host defenses associated with the acidity of the stomach. The mechanisms by which these reactive species exert antimicrobial activity in the gastric lumen are, however, poorly understood.

Methodology/principal findings: The genetically tractable acid tolerance response (ATR) that enables enteropathogens to survive harsh acidity was screened for signaling pathways responsive to RNS. The nitric oxide (NO) donor spermine NONOate derepressed the Fur regulon that controls secondary lines of resistance against organic acids. Despite inducing a Fur-mediated adaptive response, acidified RNS largely repressed oral virulence as demonstrated by the fact that Salmonella bacteria exposed to NO donors during mildly acidic conditions were shed in low amounts in feces and exhibited ameliorated oral virulence. NO prevented Salmonella from mounting a de novo ATR, but was unable to suppress an already functional protective response, suggesting that RNS target regulatory cascades but not their effectors. Transcriptional and translational analyses revealed that the PhoPQ signaling cascade is a critical ATR target of NO in rapidly growing Salmonella. Inhibition of PhoPQ signaling appears to contribute to most of the NO-mediated abrogation of the ATR in log phase bacteria, because the augmented acid sensitivity of phoQ-deficient Salmonella was not further enhanced after RNS treatment.

Conclusions/significance: Since PhoPQ-regulated acid resistance is widespread in enteric pathogens, the RNS-mediated inhibition of the Salmonella ATR described herein may represent a common component of innate host defenses.

Show MeSH
Related in: MedlinePlus