A comparison of cecal colonization of Salmonella enterica serotype Typhimurium in white leghorn chicks and Salmonella-resistant mice

<p>Abstract</p> <p>Background</p> <p>Salmonellosis is one of the most important bacterial food borne illnesses worldwide. A major source of infection for humans is consumption of chicken or egg products that have been contaminated with <it>Salmonella enterica </it>serotype Typhimurium, however our knowledge regarding colonization and persistence factors in the chicken is small.</p> <p>Results</p> <p>We compared intestinal and systemic colonization of 1-week-old White Leghorn chicks and <it>Salmonella</it>-resistant CBA/J mice during infection with <it>Salmonella enterica </it>serotype Typhimurium ATCC14028, one of the most commonly studied isolates. We also studied the distribution of wild type serotype Typhimurium ATCC14028 and an isogenic <it>invA </it>mutant during competitive infection in the cecum of 1-week-old White Leghorn chicks and 8-week-old CBA/J mice. We found that although the systemic levels of serotype Typhimurium in both infected animal models are low, infected mice have significant splenomegaly beginning at 15 days post infection. In the intestinal tract itself, the cecal contents are the major site for recovery of serotype Typhimurium in the cecum of 1-week-old chicks and <it>Salmonella</it>-resistant mice. Additionally we show that only a small minority of <it>Salmonellae </it>are intracellular in the cecal epithelium of both infected animal models, and while SPI-1 is important for successful infection in the murine model, it is important for association with the cecal epithelium of 1-week-old chicks. Finally, we show that in chicks infected with serotype Typhimurium at 1 week of age, the level of fecal shedding of this organism does not reflect the level of cecal colonization as it does in murine models.</p> <p>Conclusion</p> <p>In our study, we highlight important differences in systemic and intestinal colonization levels between chick and murine serotype Typhimurium infections, and provide evidence that suggests that the role of SPI-1 may not be the same during colonization of both animal models.</p

A comparison of cecal colonization of Salmonella enterica serotype Typhimurium in white leghorn chicks and Salmonella-resistant mice

<p>Abstract</p> <p>Background</p> <p>Salmonellosis is one of the most important bacterial food borne illnesses worldwide. A major source of infection for humans is consumption of chicken or egg products that have been contaminated with <it>Salmonella enterica </it>serotype Typhimurium, however our knowledge regarding colonization and persistence factors in the chicken is small.</p> <p>Results</p> <p>We compared intestinal and systemic colonization of 1-week-old White Leghorn chicks and <it>Salmonella</it>-resistant CBA/J mice during infection with <it>Salmonella enterica </it>serotype Typhimurium ATCC14028, one of the most commonly studied isolates. We also studied the distribution of wild type serotype Typhimurium ATCC14028 and an isogenic <it>invA </it>mutant during competitive infection in the cecum of 1-week-old White Leghorn chicks and 8-week-old CBA/J mice. We found that although the systemic levels of serotype Typhimurium in both infected animal models are low, infected mice have significant splenomegaly beginning at 15 days post infection. In the intestinal tract itself, the cecal contents are the major site for recovery of serotype Typhimurium in the cecum of 1-week-old chicks and <it>Salmonella</it>-resistant mice. Additionally we show that only a small minority of <it>Salmonellae </it>are intracellular in the cecal epithelium of both infected animal models, and while SPI-1 is important for successful infection in the murine model, it is important for association with the cecal epithelium of 1-week-old chicks. Finally, we show that in chicks infected with serotype Typhimurium at 1 week of age, the level of fecal shedding of this organism does not reflect the level of cecal colonization as it does in murine models.</p> <p>Conclusion</p> <p>In our study, we highlight important differences in systemic and intestinal colonization levels between chick and murine serotype Typhimurium infections, and provide evidence that suggests that the role of SPI-1 may not be the same during colonization of both animal models.</p

Linearized Siderophore Products Secreted via MacAB Efflux Pump Protect Salmonella enterica Serovar Typhimurium from Oxidative Stress

Nontyphoidal salmonellae (NTS) are exposed to reactive oxygen species (ROS) during their residency in the gut. To survive oxidative stress encountered during infection, salmonellae employ several mechanisms. One of these mechanisms involves the multidrug efflux pump MacAB, although the natural substrate of this pump has not been identified. MacAB homologs in pseudomonads secrete products of nonribosomal peptide synthesis (NRPS). In Salmonella enterica serovar Typhimurium, the siderophore enterobactin is produced by NRPS in response to iron starvation and this molecule can be processed into salmochelin and several linear metabolites. We found that Salmonella mutants lacking the key NRPS enzyme EntF are sensitive to peroxide mediated killing and cannot detoxify extracellular H2O2. Moreover, EntF and MacAB function in a common pathway to promote survival of Salmonella during oxidative stress. We further demonstrated that S. Typhimurium secretes siderophores in iron-rich media when peroxide is present and that these MacAB-secreted metabolites participate in protection of bacteria against H2O2. We showed that secretion of anti-H2O2 molecules is independent of the presence of the known siderophore efflux pumps EntS and IroC, well-described efflux systems involved in secretion of enterobactin and salmochelin. Both salmochelin and enterobactin are dispensable for S. Typhimurium protection against ROS; however, linear metabolites of enterobactin produced by esterases IroE and Fes are needed for bacterial survival in peroxide-containing media. We determined that linearized enterobactin trimer protects S. Typhimurium against peroxide-mediated killing in a MacAB-dependent fashion. Thus, we suggest that linearized enterobactin trimer is a natural substrate of MacAB and that its purpose is to detoxify extracellular reactive oxygen species

Linearized Siderophore Products Secreted via MacAB Efflux Pump Protect Salmonella enterica Serovar Typhimurium from Oxidative Stress

Nontyphoidal salmonellae (NTS) are exposed to reactive oxygen species (ROS) during their residency in the gut. To survive oxidative stress encountered during infection, salmonellae employ several mechanisms. One of these mechanisms involves the multidrug efflux pump MacAB, although the natural substrate of this pump has not been identified. MacAB homologs in pseudomonads secrete products of nonribosomal peptide synthesis (NRPS). In Salmonella enterica serovar Typhimurium, the siderophore enterobactin is produced by NRPS in response to iron starvation and this molecule can be processed into salmochelin and several linear metabolites. We found that Salmonella mutants lacking the key NRPS enzyme EntF are sensitive to peroxide mediated killing and cannot detoxify extracellular H2O2 Moreover, EntF and MacAB function in a common pathway to promote survival of Salmonella during oxidative stress. We further demonstrated that S. Typhimurium secretes siderophores in iron-rich media when peroxide is present and that these MacAB-secreted metabolites participate in protection of bacteria against H2O2 We showed that secretion of anti-H2O2 molecules is independent of the presence of the known siderophore efflux pumps EntS and IroC, well-described efflux systems involved in secretion of enterobactin and salmochelin. Both salmochelin and enterobactin are dispensable for S. Typhimurium protection against ROS; however, linear metabolites of enterobactin produced by esterases IroE and Fes are needed for bacterial survival in peroxide-containing media. We determined that linearized enterobactin trimer protects S. Typhimurium against peroxide-mediated killing in a MacAB-dependent fashion. Thus, we suggest that linearized enterobactin trimer is a natural substrate of MacAB and that its purpose is to detoxify extracellular reactive oxygen species. IMPORTANCE Nontyphoidal Salmonella bacteria induce a classic inflammatory diarrhea by eliciting a large influx of neutrophils, producing a robust oxidative burst. Despite substantial progress understanding the benefits to the host of the inflammatory response to Salmonella, little is known regarding how Salmonella can simultaneously resist the damaging effects of the oxidative burst. The multidrug efflux pump MacAB is important for survival of oxidative stress both in vitro and during infection. We describe a new pathway used by Salmonella Typhimurium to detoxify extracellular reactive oxygen species using a multidrug efflux pump (MacAB) to secrete a linear siderophore, a metabolite of enterobactin. The natural substrates of many multidrug efflux pumps are unknown, and functional roles of the linear metabolites of enterobactin are unknown. We bring two novel discoveries together to highlight an important mechanism used by Salmonella to survive under the oxidative stress conditions that this organism encounters during the classic inflammatory diarrhea that it also induces

De novo pyrimidine synthesis is necessary for intestinal colonization of Salmonella Typhimurium in chicks

© 2017 Yang et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. pyrE (STM3733) encodes orotate phosphoribosyltransferase (OPRTase; EC 2.4.2.10), the fifth enzyme of the de novo pyrimidine biosynthetic pathway. We identified a ΔpyrE mutant as under selection in screening of a Salmonella mutant library in 4-day old chicks. Here, we confirm that a ΔpyrE mutant colonizes 4-day old chicks poorly in competitive infection with isogenic wild type, and that the ability of this mutant to colonize chicks could be restored by providing a copy of pyrE in trans. We further show that our ΔpyrE mutant grows poorly in nutrient poor conditions in vitro, and that the ability of this mutant to grow is restored, both in vitro and in chicks, when precursors to the pyrimidine salvage pathway were provided. This finding suggests that the environment in the chick intestine during our infections lacks sufficient precursors of the pyrimidine salvage pathway to support Salmonella growth. Finally, we show that the colonization defect of a ΔpyrE mutant during infection occurs in to chicks, but not in CBA/J mice or ligated ileal loops in calves. Our data suggest that de novo pyrimidine synthesis is necessary for colonization of Salmonella Typhimurium in the chick, a nd that the salvage pathway is not used in this niche

Generalized Bacteriophage Transduction in Serratia marcescens

© 2016, Springer Science+Business Media New York.Serratia marcescens is a Gram-negative bacterium from the Enterobacteriaceae family. As an opportunistic human pathogen and a causative agent of a range of nosocomial infections, S. marcescens represents a growing public health problem. Little is known about S. marcescens pathogenicity factors, in part due to the lack of well-developed functional genomic tools. Bacteriophage transduction is one of the main mechanisms of horizontal transfer of DNA in bacteria. S. marcescens bacteriophages previously used in transduction experiments are strain-specific, and no comprehensive studies have previously been done to evaluate the efficiency of transduction across different phages and host strains. In this study, we directly compared the ability of bacteriophages ΦOT8 and ΦIF3 to infect two S. marcescens strains, SM6 and SR41-8000. We successfully used bacteriophage ΦOT8 to transfer a mutation in tolC locus between two S. marcescens strains

The action of bacillus' metzinc metalloproteinase on cultured animal cell lines

Microbial enzymes with high activity are increasingly used in medical practice in development of drugs for treatment of burn injuries, diseases of gastrointestinal tract, as well as in systemic enzyme therapy. One of crucial requirements for such drugs is the absence of toxicity, both on cellular and organism levels. The aim of the present study was to evaluate cytotoxic effect of Bacillus pumilus metzinc metalloproteinase on animal cell lines. Cytological changes in LEK, Vero and PK-15 cell lines were observed after cell treatment with 155 μ/ml enzyme. However, lower enzyme concentrations - 75 and 15 μ/ml did not show any toxicity

Inactivation of Chromosomal Genes in Serratia marcescens

© 2016, Springer Science+Business Media New York.Gram-negative bacterium Serratia marcescens is a well-known environmental microorganism and the accepted clinical pathogen causing nosocomial infections. It attracts more attention in recent years due to the emergence of strains with multiple drug resistance. Standard recombinant techniques are difficult to apply to S. marcescens due to the presence of numerous hydrolytic enzymes, in particular, extracellular nuclease and restriction endonuclease, which degrade transforming DNAs. We overcame this obstacle by utilizing restrictionless nuclease-deficient mutant strain S. marcescens TT392. As a proof of principal, in this genetic background, we generated a knockout strain with deletion of macAB locus using lambda red technology. The resulting mutation could be easily moved to a new genetic background by generalized phage transduction. This strategy provides a good tool for evaluation of S. marcescens pathogenic potential

Effects of Bacillus Serine Proteases on the Bacterial Biofilms

© 2017 Olga Mitrofanova et al. Serratia marcescens is an emerging opportunistic pathogen responsible for many hospital-acquired infections including catheter-associated bacteremia and urinary tract and respiratory tract infections. Biofilm formation is one of the mechanisms employed by S. marcescens to increase its virulence and pathogenicity. Here, we have investigated the main steps of the biofilm formation by S. marcescens SR 41-8000. It was found that the biofilm growth is stimulated by the nutrient-rich environment. The time-course experiments showed that S. marcescens cells adhere to the surface of the catheter and start to produce extracellular polymeric substances (EPS) within the first 2 days of growth. After 7 days, S. marcescens biofilms maturate and consist of bacterial cells embedded in a self-produced matrix of hydrated EPS. In this study, the effect of Bacillus pumilus 3-19 proteolytic enzymes on the structure of 7-day-old S. marcescens biofilms was examined. Using quantitative methods and scanning electron microscopy for the detection of biofilm, we demonstrated a high efficacy of subtilisin-like protease and glutamyl endopeptidase in biofilm removal. Enzymatic treatment resulted in the degradation of the EPS components and significant eradication of the biofilms

Production of Siderophores by Serratia marcescens and the Role of MacAB Efflux Pump in Siderophores Secretion

© 2016, Springer Science+Business Media New York.Human opportunistic pathogen Serratia marcescens secrete siderophores to enable growth under iron-limiting conditions. Iron acquisition genes are among a few known virulence factors of S. marcescens. Siderophore export systems in Gram-negative bacteria are not fully understood. There is some evidence for involvement of efflux pumps in the export of synthesized enterobactin-like molecules. The goal of this study was to characterize siderophore production by two different strains of S. marcescens, SM6 and SR41-8000, and to evaluate the role of efflux pump MacAB in siderophore secretion by these strains. We showed that both strains produced siderophores in CAS agar assay. We further showed that both strains were able to secrete catecholate siderophores in response to iron starvation. MacAB efflux pump played a role in siderophore secretion of S. marcescens SR41-8000 strain but was dispensable for accumulation of these molecules in the culture supernatant of S. marcescens strain SM6