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Stress response of Salmonella enterica serovar typhimurium to acidified nitrite.

ABSTRACT: The antimicrobial action of the curing agent sodium nitrite (NaNO2), which is added as a preservative to raw meat products, depends on its conversion to nitric oxide and other reactive nitrogen species under acidic conditions. In this study, we used RNA sequencing to analyze the acidified-NaNO2 shock and adaptive responses of Salmonella enterica serovar Typhimurium, a frequent contaminant in raw meat, considering parameters relevant for the production of raw-cured sausages. Upon a 10-min exposure to 150 mg/liter NaNO2 in LB (pH 5.5) acidified with lactic acid, genes involved in nitrosative-stress protection, together with several other stress-related genes, were induced. In contrast, genes involved in translation, transcription, replication, and motility were downregulated. The induction of stress tolerance and the reduction of cell proliferation obviously promote survival under harsh acidified-NaNO2 stress. The subsequent adaptive response was characterized by upregulation of NsrR-regulated genes and iron uptake systems and by downregulation of genes involved in anaerobic respiratory pathways. Strikingly, amino acid decarboxylase systems, which contribute to acid tolerance, displayed increased transcript levels in response to acidified NaNO2. The induction of systems known to be involved in acid resistance indicates a nitrite-mediated increase in the level of acid stress. Deletion of cadA, which encodes lysine decarboxylase, resulted in increased sensitivity to acidified NaNO2. Intracellular pH measurements using a pH-sensitive green fluorescent protein (GFP) variant showed that the cytoplasmic pH of S. Typhimurium in LB medium (pH 5.5) is decreased upon the addition of NaNO2. This study provides the first evidence that intracellular acidification is an additional antibacterial mode of action of acidified NaNO2.

SUBMITTER: Muhlig A

PROVIDER: S-EPMC4178661 | biostudies-literature | 2014 Oct

REPOSITORIES: biostudies-literature

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Stress response of Salmonella enterica serovar typhimurium to acidified nitrite.

Mühlig Anna A Behr Jürgen J Scherer Siegfried S Müller-Herbst Stefanie S

Applied and environmental microbiology 20140808 20

The antimicrobial action of the curing agent sodium nitrite (NaNO2), which is added as a preservative to raw meat products, depends on its conversion to nitric oxide and other reactive nitrogen species under acidic conditions. In this study, we used RNA sequencing to analyze the acidified-NaNO2 shock and adaptive responses of Salmonella enterica serovar Typhimurium, a frequent contaminant in raw meat, considering parameters relevant for the production of raw-cured sausages. Upon a 10-min exposur ...[more]

PMID: 25107963

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Project description:The intracellular pathogen Salmonella enterica serovar Typhimurium has emerged as a major cause of foodborne illness, representing a severe clinical and economic concern worldwide. The capacity of this pathogen to efficiently infect and survive inside the host depends on its ability to synchronize a complex network of virulence mechanisms. Therefore, the identification of new virulence determinants has become of paramount importance in the search of new targets for drug development. BolA-like proteins are widely conserved in all kingdoms of life. In Escherichia coli, this transcription factor has a critical regulatory role in several mechanisms that are tightly related to bacterial virulence. Therefore, in the present work we used the well-established infection model Galleria mellonella to evaluate the role of BolA protein in S Typhimurium virulence. We have shown that BolA is an important player in S Typhimurium pathogenesis. Specifically, the absence of BolA leads to a defective virulence capacity that is most likely related to the remarkable effect of this protein on S Typhimurium evasion of the cellular response. Furthermore, it was demonstrated that BolA has a critical role in bacterial survival under harsh conditions since BolA conferred protection against acidic and oxidative stress. Hence, we provide evidence that BolA is a determining factor in the ability of Salmonella to survive and overcome host defense mechanisms, and this is an important step in progress to an understanding of the pathways underlying bacterial virulence.IMPORTANCE BolA has been described as an important protein for survival in the late stages of bacterial growth and under harsh environmental conditions. High levels of BolA in stationary phase and under stresses have been connected with a plethora of phenotypes, strongly suggesting its important role as a master regulator. Here, we show that BolA is a determining factor in the ability of Salmonella to survive and overcome host defense mechanisms, and this is an important step in progress to an understanding of the pathways underlying bacterial virulence. This work constitutes a relevant step toward an understanding of the role of BolA protein and may have an important impact on future studies in other organisms. Therefore, this study is of utmost importance for understanding the genetic and molecular bases involved in the regulation of Salmonella virulence and may contribute to future industrial and public health care applications.

Project description:Flagellum-driven motility of Salmonella enterica serovar Typhimurium facilitates host colonization. However, the large extracellular flagellum is also a prime target for the immune system. As consequence, expression of flagella is bistable within a population of Salmonella, resulting in flagellated and nonflagellated subpopulations. This allows the bacteria to maximize fitness in hostile environments. The degenerate EAL domain protein RflP (formerly YdiV) is responsible for the bistable expression of flagella by directing the flagellar master regulatory complex FlhD4C2 with respect to proteolytic degradation. Information concerning the environmental cues controlling expression of rflP and thus about the bistable flagellar biosynthesis remains ambiguous. Here, we demonstrated that RflP responds to cell envelope stress and alterations of outer membrane integrity. Lipopolysaccharide (LPS) truncation mutants of Salmonella Typhimurium exhibited increasing motility defects due to downregulation of flagellar gene expression. Transposon mutagenesis and genetic profiling revealed that ?24 (RpoE) and Rcs phosphorelay-dependent cell envelope stress response systems sense modifications of the lipopolysaccaride, low pH, and activity of the complement system. This subsequently results in activation of RflP expression and degradation of FlhD4C2 via ClpXP. We speculate that the presence of diverse hostile environments inside the host might result in cell envelope damage and would thus trigger the repression of resource-costly and immunogenic flagellum biosynthesis via activation of the cell envelope stress response.IMPORTANCE Pathogenic bacteria such as Salmonella Typhimurium sense and adapt to a multitude of changing and stressful environments during host infection. At the initial stage of gastrointestinal colonization, Salmonella uses flagellum-mediated motility to reach preferred sites of infection. However, the flagellum also constitutes a prime target for the host's immune response. Accordingly, the pathogen needs to determine the spatiotemporal stage of infection and control flagellar biosynthesis in a robust manner. We found that Salmonella uses signals from cell envelope stress-sensing systems to turn off production of flagella. We speculate that downregulation of flagellum synthesis after cell envelope damage in hostile environments aids survival of Salmonella during late stages of infection and provides a means to escape recognition by the immune system.

Dataset Information

Stress response of Salmonella enterica serovar typhimurium to acidified nitrite.

Publications

Stress response of Salmonella enterica serovar typhimurium to acidified nitrite.

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