scholarly journals Engineering and Preclinical Evaluation of Attenuated Nontyphoidal Salmonella Strains Serving as Live Oral Vaccines and as Reagent Strains

2011 ◽  
Vol 79 (10) ◽  
pp. 4175-4185 ◽  
Author(s):  
Sharon M. Tennant ◽  
Jin-Yuan Wang ◽  
James E. Galen ◽  
Raphael Simon ◽  
Marcela F. Pasetti ◽  
...  
Keyword(s):  
Salmonella Enterica ◽  
Lethal Dose ◽  
Invasive Disease ◽  
Oral Immunization ◽  
Oral Vaccines ◽  
Wild Type ◽  
Lethal Challenge ◽  
Antibiotic Resistant ◽  
Content Type ◽  
Serovar Typhimurium

ABSTRACTWhile nontyphoidalSalmonella(NTS) has long been recognized as a cause of self-limited gastroenteritis, it is becoming increasingly evident that multiple-antibiotic-resistant strains are also emerging as important causes of invasive bacteremia and focal infections, resulting in hospitalizations and deaths. We have constructed attenuatedSalmonella entericaserovar Typhimurium andSalmonella entericaserovar Enteritidis strains that can serve as live oral vaccines and as “reagent strains” for subunit vaccine production in a safe and economical manner. Prototype attenuated vaccine strains CVD 1921 and CVD 1941, derived from the invasive wild-type strainsS. TyphimuriumI77 andS. EnteritidisR11, respectively, were constructed by deletingguaBA, encoding guanine biosynthesis, andclpP, encoding a master protease regulator. TheclpPmutation resulted in a hyperflagellation phenotype. An additional deletion infliDyielded reagent strains CVD 1923 and CVD 1943, respectively, which export flagellin monomers. Oral 50% lethal dose (LD50) analyses showed that the NTS vaccine strains were all highly attenuated in mice. Oral immunization with CVD 1921 or CVD 1923 protected mice against lethal challenge with wild-typeS. TyphimuriumI77. Immunization with CVD 1941 but not CVD 1943 protected mice against lethal infection withS. EnteritidisR11. Immune responses induced by these strains included high levels of serum IgG anti-lipopolysaccharide (LPS) and anti-flagellum antibodies, with titers increasing progressively during the immunization schedule. SinceS. TyphimuriumandS. Enteritidisare the most common NTS serovars associated with invasive disease, these findings can pave the way for development of a highly effective, broad-spectrum vaccine against invasive NTS.

scholarly journals Role of sapA and yfgA in Susceptibility to Antibody-Mediated Complement-Dependent Killing and Virulence of Salmonella enterica Serovar Typhimurium

2017 ◽  
Vol 85 (9) ◽  
Author(s):  
Edna M. Ondari ◽  
Jennifer N. Heath ◽  
Elizabeth J. Klemm ◽  
Gemma Langridge ◽  
Lars Barquist ◽  
...  
Keyword(s):  
Salmonella Enterica ◽  
Protein Translocation ◽  
Invasive Disease ◽  
Immune Serum ◽  
Insertion Mutant ◽  
Content Type ◽  
Serovar Typhimurium ◽  
Serum Killing ◽  
Increased Sensitivity

ABSTRACT The ST313 pathovar of Salmonella enterica serovar Typhimurium contributes to a high burden of invasive disease among African infants and HIV-infected adults. It is characterized by genome degradation (loss of coding capacity) and has increased resistance to antibody-dependent complement-mediated killing compared with enterocolitis-causing strains of S. Typhimurium. Vaccination is an attractive disease-prevention strategy, and leading candidates focus on the induction of bactericidal antibodies. Antibody-resistant strains arising through further gene deletion could compromise such a strategy. Exposing a saturating transposon insertion mutant library of S. Typhimurium to immune serum identified a repertoire of S. Typhimurium genes that, when interrupted, result in increased resistance to serum killing. These genes included several involved in bacterial envelope biogenesis, protein translocation, and metabolism. We generated defined mutant derivatives using S. Typhimurium SL1344 as the host. Based on their initial levels of enhanced resistance to killing, yfgA and sapA mutants were selected for further characterization. The S. Typhimurium yfgA mutant lost the characteristic Salmonella rod-shaped appearance, exhibited increased sensitivity to osmotic and detergent stress, lacked very long lipopolysaccharide, was unable to invade enterocytes, and demonstrated decreased ability to infect mice. In contrast, the S. Typhimurium sapA mutants had similar sensitivity to osmotic and detergent stress and lipopolysaccharide profile and an increased ability to infect enterocytes compared with the wild type, but it had no increased ability to cause in vivo infection. These findings indicate that increased resistance to antibody-dependent complement-mediated killing secondary to genetic deletion is not necessarily accompanied by increased virulence and suggest the presence of different mechanisms of antibody resistance.

scholarly journals Salmonella enterica Serovar Typhimurium Uses PbgA/YejM To Regulate Lipopolysaccharide Assembly during Bacteremia

2019 ◽  
Vol 88 (1) ◽  
Author(s):  
Melina B. Cian ◽  
Nicole P. Giordano ◽  
Revathi Masilamani ◽  
Keaton E. Minor ◽  
Zachary D. Dalebroux
Keyword(s):  
Salmonella Enterica ◽  
Lipid A ◽  
Salmonella Enterica Serovar Typhimurium ◽  
Transmembrane Domain ◽  
Wild Type ◽  
Content Type ◽  
Serovar Typhimurium ◽  
Periplasmic Domain ◽  
Inner Membrane Protein ◽  
Substitution Mutations

ABSTRACT Salmonella enterica serovar Typhimurium (S. Typhimurium) relies upon the inner membrane protein PbgA to enhance outer membrane (OM) integrity and promote virulence in mice. The PbgA transmembrane domain (residues 1 to 190) is essential for viability, while the periplasmic domain (residues 191 to 586) is dispensable. Residues within the basic region (residues 191 to 245) bind acidic phosphates on polar phospholipids, like for cardiolipins, and are necessary for salmonella OM integrity. S. Typhimurium bacteria increase their OM cardiolipin concentrations during activation of the PhoPQ regulators. The mechanism involves PbgA’s periplasmic globular region (residues 245 to 586), but the biological role of increasing cardiolipins on the surface is not understood. Nonsynonymous polymorphisms in three essential lipopolysaccharide (LPS) synthesis regulators, lapB (also known as yciM), ftsH, and lpxC, variably suppressed the defects in OM integrity, rifampin resistance, survival in macrophages, and systemic colonization of mice in the pbgAΔ191–586 mutant (in which the PbgA periplasmic domain from residues 191 to 586 is deleted). Compared to the OMs of the wild-type salmonellae, the OMs of the pbgA mutants had increased levels of lipid A-core molecules, cardiolipins, and phosphatidylethanolamines and decreased levels of specific phospholipids with cyclopropanated fatty acids. Complementation and substitution mutations in LapB and LpxC generally restored the phospholipid and LPS assembly defects for the pbgA mutants. During bacteremia, mice infected with the pbgA mutants survived and cleared the bacteria, while animals infected with wild-type salmonellae succumbed within 1 week. Remarkably, wild-type mice survived asymptomatically with pbgA-lpxC salmonellae in their livers and spleens for months, but Toll-like receptor 4-deficient animals succumbed to these infections within roughly 1 week. In summary, S. Typhimurium uses PbgA to influence LPS assembly during stress in order to survive, adapt, and proliferate within the host environment.

Oral immunization with a dam mutant of Yersinia pseudotuberculosis protects against plague

Microbiology ◽  
2005 ◽  
Vol 151 (6) ◽  
pp. 1919-1926 ◽  
Author(s):  
Victoria L. Taylor ◽  
Richard W. Titball ◽  
Petra C. F. Oyston
Keyword(s):  
Gene Expression ◽  
Bile Salts ◽  
Yersinia Pseudotuberculosis ◽  
Lethal Dose ◽  
Oral Immunization ◽  
Wild Type ◽  
Gene Encoding ◽  
Serovar Typhimurium ◽  
Dna Adenine Methylase ◽  
Methylase Gene

Inactivation of the gene encoding DNA adenine methylase (dam) has been shown to attenuate some pathogens such as Salmonella enterica serovar Typhimurium and is a lethal mutation in others such as Yersinia pseudotuberculosis strain YPIII. In this study the dam methylase gene in Yersinia pseudotuberculosis strain IP32953 was inactivated. Unlike the wild-type, DNA isolated from the mutant could be digested with MboI, which is consistent with an altered pattern of DNA methylation. The mutant was sensitive to bile salts but not to 2-aminopurine. The effect of dam inactivation on gene expression was examined using a DNA microarray. In BALB/c mice inoculated orally or intravenously with the dam mutant, the median lethal dose (MLD) was at least 106-fold higher than the MLD of the wild-type. BALB/c mice inoculated with the mutant were protected against a subcutaneous challenge with 100 MLDs of Yersinia pestis strain GB and an intravenous challenge with 300 MLDs of Y. pseudotuberculosis IP32953.

scholarly journals Alterations of Salmonella enterica Serovar Typhimurium Antibiotic Resistance under Environmental Pressure

2018 ◽  
Vol 84 (19) ◽  
Author(s):  
Mengfei Peng ◽  
Serajus Salaheen ◽  
Robert L. Buchanan ◽  
Debabrata Biswas
Keyword(s):  
Antibiotic Resistance ◽  
Resistance Genes ◽  
Salmonella Enterica ◽  
Tetracycline Resistance ◽  
Multidrug Resistant ◽  
Genetic Alterations ◽  
Antibiotic Resistant ◽  
Content Type ◽  
Farm Systems ◽  
Serovar Typhimurium

ABSTRACT Microbial horizontal gene transfer is a continuous process that shapes bacterial genomic adaptation to the environment and the composition of concurrent microbial ecology. This includes the potential impact of synthetic antibiotic utilization in farm animal production on overall antibiotic resistance issues; however, the mechanisms behind the evolution of microbial communities are not fully understood. We explored potential mechanisms by experimentally examining the relatedness of phylogenetic inference between multidrug-resistant Salmonella enterica serovar Typhimurium isolates and pathogenic Salmonella Typhimurium strains based on genome-wide single-nucleotide polymorphism (SNP) comparisons. Antibiotic-resistant S. Typhimurium isolates in a simulated farm environment barely lost their resistance, whereas sensitive S. Typhimurium isolates in soils gradually acquired higher tetracycline resistance under antibiotic pressure and manipulated differential expression of antibiotic-resistant genes. The expeditious development of antibiotic resistance and the ensuing genetic alterations in antimicrobial resistance genes in S. Typhimurium warrant effective actions to control the dissemination of Salmonella antibiotic resistance. IMPORTANCE Antibiotic resistance is attributed to the misuse or overuse of antibiotics in agriculture, and antibiotic resistance genes can also be transferred to bacteria under environmental stress. In this study, we report a unidirectional alteration in antibiotic resistance from susceptibility to increased resistance. Highly sensitive Salmonella enterica serovar Typhimurium isolates from organic farm systems quickly acquired tetracycline resistance under antibiotic pressure in simulated farm soil environments within 2 weeks, with expression of antibiotic resistance-related genes that was significantly upregulated. Conversely, originally resistant S. Typhimurium isolates from conventional farm systems lost little of their resistance when transferred to environments without antibiotic pressure. Additionally, multidrug-resistant S. Typhimurium isolates genetically shared relevancy with pathogenic S. Typhimurium isolates, whereas susceptible isolates clustered with nonpathogenic strains. These results provide detailed discussion and explanation about the genetic alterations and simultaneous acquisition of antibiotic resistance in S. Typhimurium in agricultural environments.

scholarly journals An Attenuated Salmonella enterica Serovar Typhimurium Strain and Galacto-Oligosaccharides Accelerate Clearance of Salmonella Infections in Poultry through Modifications to the Gut Microbiome

2017 ◽  
Vol 84 (5) ◽  
Author(s):  
M. Andrea Azcarate-Peril ◽  
Natasha Butz ◽  
Maria Belen Cadenas ◽  
Matthew Koci ◽  
Anne Ballou ◽  
...  
Keyword(s):  
United States ◽  
Gut Microbiota ◽  
Gut Microbiome ◽  
Salmonella Enterica ◽  
The United States ◽  
Wild Type ◽  
Content Type ◽  
Salmonella Infections ◽  
Serovar Typhimurium ◽  
The Impact

ABSTRACT Salmonella is estimated to cause one million foodborne illnesses in the United States every year. Salmonella -contaminated poultry products are one of the major sources of salmonellosis. Given the critical role of the gut microbiota in Salmonella transmission, a manipulation of the chicken intestinal microenvironment could prevent animal colonization by the pathogen. In Salmonella , the global regulator gene fnr ( f umarate n itrate r eduction) regulates anaerobic metabolism and is essential for adapting to the gut environment. This study tested the hypothesis that an attenuated Fnr mutant of Salmonella enterica serovar Typhimurium (attST) or prebiotic galacto-oligosaccharides (GOS) could improve resistance to wild-type Salmonella via modifications to the structure of the chicken gut microbiome. Intestinal samples from a total of 273 animals were collected weekly for 9 weeks to evaluate the impact of attST or prebiotic supplementation on microbial species of the cecum, duodenum, jejunum, and ileum. We next analyzed changes to the gut microbiome induced by challenging the animals with a wild-type Salmonella serovar 4,[5],12:r:− (Nal r ) strain and determined the clearance rate of the virulent strain in the treated and control groups. Both GOS and the attenuated Salmonella strain modified the gut microbiome but elicited alterations of different taxonomic groups. The attST produced significant increases of Alistipes and undefined Lactobacillus , while GOS increased Christensenellaceae and Lactobacillus reuteri . The microbiome structural changes induced by both treatments resulted in a faster clearance after a Salmonella challenge. IMPORTANCE With an average annual incidence of 13.1 cases/100,000 individuals, salmonellosis has been deemed a nationally notifiable condition in the United States by the Centers for Disease Control and Prevention (CDC). Earlier studies demonstrated that Salmonella is transmitted by a subset of animals (supershedders). The supershedder phenotype can be induced by antibiotics, ascertaining an essential role for the gut microbiota in Salmonella transmission. Consequently, modulation of the gut microbiota and modification of the intestinal microenvironment could assist in preventing animal colonization by the pathogen. Our study demonstrated that a manipulation of the chicken gut microbiota by the administration of an attenuated Salmonella strain or prebiotic galacto-oligosaccharides (GOS) can promote resistance to Salmonella colonization via increases of beneficial microorganisms that translate into a less hospitable gut microenvironment.

scholarly journals Oral Immunization with an rfaH Mutant Elicits Protection against Salmonellosis in Mice

2004 ◽  
Vol 72 (7) ◽  
pp. 4297-4301 ◽  
Author(s):  
Gábor Nagy ◽  
Ulrich Dobrindt ◽  
Jörg Hacker ◽  
Levente Emödy
Keyword(s):  
Type Strain ◽  
Wild Type Strain ◽  
Lethal Dose ◽  
Fold Increase ◽  
Oral Immunization ◽  
Oral Infection ◽  
Wild Type ◽  
Virulence Attenuation ◽  
Serovar Typhimurium

ABSTRACT Loss of the transcriptional antiterminator RfaH results in virulence attenuation (>104-fold increase in 50% lethal dose) of the archetypal Salmonella enterica serovar Typhimurium strain SL1344 by both orogastric and intraperitoneal routes of infection in BALB/c mice. Oral immunization with the mutant efficiently protects mice against a subsequent oral infection with the wild-type strain. Interestingly, in vitro immunoreactivity is not confined to strain SL1344; rather, it is directed also towards other serovars of S. enterica and even Salmonella bongori strains.

scholarly journals Use of Attenuated but Metabolically Competent Salmonella as a Probiotic To Prevent or Treat Salmonella Infection

2016 ◽  
Vol 84 (7) ◽  
pp. 2131-2140 ◽  
Author(s):  
Anice Sabag-Daigle ◽  
Henry M. Blunk ◽  
Juan F. Gonzalez ◽  
Brandi L. Steidley ◽  
Prosper N. Boyaka ◽  
...  
Keyword(s):  
Typhoid Fever ◽  
Salmonella Enterica ◽  
Probiotic Bacterium ◽  
Wild Type ◽  
Nutrient Source ◽  
Content Type ◽  
Nutrient Sources ◽  
Novel Approach ◽  
Serovar Typhimurium ◽  
Utilization Pathway

Salmonella entericais among the most burdensome of foodborne disease agents. There are over 2,600 serovars that cause a range of disease manifestations ranging from enterocolitis to typhoid fever. While there are two vaccines in use in humans to protect against typhoid fever, there are none that prevent enterocolitis. If vaccines preventing enterocolitis were to be developed, they would likely protect against only one or a few serovars. In this report, we tested the hypothesis that probiotic organisms could compete for the preferred nutrient sources ofSalmonellaand thus prevent or treat infection. To this end, we added thefralocus, which encodes a utilization pathway for theSalmonella-specific nutrient source fructose-asparagine (F-Asn), to the probiotic bacteriumEscherichia coliNissle 1917 (Nissle) to increase its ability to compete withSalmonellain mouse models. We also tested a metabolically competent, but avirulent,Salmonella entericaserovar Typhimurium mutant for its ability to compete with wild-typeSalmonella. The modified Nissle strain became more virulent and less able to protect againstSalmonellain some instances. On the other hand, the modifiedSalmonellastrain was safe and effective in preventing infection with wild-typeSalmonella. While we tested for efficacy only againstSalmonellaTyphimurium, the modifiedSalmonellastrain may be able to compete metabolically with most, if not all,Salmonellaserovars, representing a novel approach to control of this pathogen.

scholarly journals Palmitoylation State Impacts Induction of Innate and Acquired Immunity by the Salmonella enterica Serovar TyphimuriummsbBMutant

2011 ◽  
Vol 79 (12) ◽  
pp. 5027-5038 ◽  
Author(s):  
Qingke Kong ◽  
David A. Six ◽  
Qing Liu ◽  
Lillian Gu ◽  
Kenneth L. Roland ◽  
...  
Keyword(s):  
Salmonella Enterica ◽  
Lipid A ◽  
Inflammatory Responses ◽  
Outer Membrane Proteins ◽  
Acyl Chain ◽  
Wild Type ◽  
Proinflammatory Response ◽  
Content Type ◽  
Serovar Typhimurium

ABSTRACTLipopolysaccharide (LPS), composed of lipid A, core, and O-antigen, is a major virulence factor ofSalmonella entericaserovar Typhimurium, with lipid A being a major stimulator to induce the proinflammatory response via the Toll-like receptor 4 (TLR4)-MD2-CD14 pathway. WhileSalmonella msbBmutants lacking the myristate chain in lipid A were investigated widely as an anticancer vaccine, inclusion of themsbBmutation in aSalmonellavaccine to deliver heterologous antigens has not yet been investigated. We introduced themsbBmutation alone or in combination with mutations in other lipid A acyl chain modification genes encoding PagL, PagP, and LpxR into wild-typeS. entericaserovar Typhimurium. ThemsbBmutation reduced virulence, while thepagL,pagP, andlpxRmutations did not affect virulence in themsbBmutant background when administered orally to BALB/c mice. Also, all mutants exhibited sensitivity to polymyxin B but did not display sensitivity to deoxycholate. LPS derived frommsbBmutants induced less inflammatory responses in human Mono Mac 6 and murine macrophage RAW264.7 cellsin vitro. However, anmsbBmutant did not decrease the induction of inflammatory responses in mice compared to the levels induced by the wild-type strain, whereas anmsbB pagPmutant induced less inflammatory responsesin vivo. The mutations were moved to an attenuatedSalmonellavaccine strain to evaluate their effects on immunogenicity. Lipid A modification caused by themsbBmutation alone and in combination withpagL,pagP, andlpxRmutations led to higher IgA production in the vaginal tract but still retained the same IgG titer level in serum to PspA, a test antigen fromStreptococcus pneumoniae, and to outer membrane proteins (OMPs) fromSalmonella.

scholarly journals Flavohemoglobin Hmp Protects Salmonella enterica Serovar Typhimurium from Nitric Oxide-Related Killing by Human Macrophages

2002 ◽  
Vol 70 (8) ◽  
pp. 4399-4405 ◽  
Author(s):  
Tânia M. Stevanin ◽  
Robert K. Poole ◽  
Eric A. G. Demoncheaux ◽  
Robert C. Read
Keyword(s):  
Nitric Oxide ◽  
Salmonella Enterica ◽  
Nitrosative Stress ◽  
Invasive Disease ◽  
Wild Type ◽  
Human Macrophages ◽  
Microbicidal Activity ◽  
Enhanced Sensitivity ◽  
Serovar Typhimurium

ABSTRACT Survival of macrophage microbicidal activity is a prerequisite for invasive disease caused by the enteric pathogen Salmonella enterica serovar Typhimurium. Flavohemoglobins, such as those of Escherichia coli, Salmonella, and yeast, play vital roles in protection of these microorganisms in vitro from nitric oxide (NO) and nitrosative stress. A Salmonella hmp mutant defective in flavohemoglobin (Hmp) synthesis exhibits growth that is hypersensitive to nitrosating agents. We found that respiration of this mutant exhibited increased inhibition by NO, whereas wild-type cells pregrown with sodium nitroprusside or S-nitrosoglutathione showed enhanced tolerance of NO. Most significantly, hmp mutants internalized by primary human peripheral monocyte-derived macrophages survived phagocytosis relatively poorly compared with similarly bound and internalized wild-type cells. That the enhanced sensitivity to macrophage microbicidal activity is due primarily to the failure of Salmonella to detoxify NO was suggested by the ability of l-N G-monomethyl arginine—an inhibitor of NO synthase—to eliminate the difference in killing between wild-type and hmp mutant Salmonella cells. These observations suggest that Salmonella Hmp contributes to protection from NO-mediated inhibition by human macrophages.

scholarly journals Binding of the RamR Repressor to Wild-Type and Mutated Promoters of theramAGene Involved in Efflux-Mediated Multidrug Resistance in Salmonella enterica Serovar Typhimurium

2011 ◽  
Vol 56 (2) ◽  
pp. 942-948 ◽  
Author(s):  
Sylvie Baucheron ◽  
Franck Coste ◽  
Sylvie Canepa ◽  
Marie-Christine Maurel ◽  
Etienne Giraud ◽  
...  
Keyword(s):  
Multidrug Resistance ◽  
Dna Binding ◽  
Binding Site ◽  
Salmonella Enterica ◽  
Transcriptional Start Site ◽  
Local Level ◽  
Wild Type ◽  
Binding Model ◽  
Content Type ◽  
Serovar Typhimurium

ABSTRACTThe transcriptional activator RamA is involved in multidrug resistance (MDR) by increasing expression of the AcrAB-TolC RND-type efflux system in several pathogenicEnterobacteriaceae. InSalmonella entericaserovar Typhimurium (S.Typhimurium),ramAexpression is negatively regulated at the local level by RamR, a transcriptional repressor of the TetR family. We here studied the DNA-binding activity of the RamR repressor with theramApromoter (PramA). As determined by high-resolution footprinting, the 28-bp-long RamR binding site covers essential features of PramA, including the −10 conserved region, the transcriptional start site oframA, and two 7-bp inverted repeats. Based on the RamR footprint and on electrophoretic mobility shift assays (EMSAs), we propose that RamR interacts with PramAas a dimer of dimers, in a fashion that is structurally similar to the QacR-DNA binding model. Surface plasmon resonance (SPR) measurements indicated that RamR has a 3-fold-lower affinity (KD[equilibrium dissociation constant] = 191 nM) for the 2-bp-deleted PramAof an MDRS.Typhimurium clinical isolate than for the wild-type PramA(KD= 66 nM). These results confirm the direct regulatory role of RamR in the repression oframAtranscription and precisely define how an alteration of its binding site can give rise to an MDR phenotype.

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