scholarly journals Mapping and Regulation of Genes within Salmonella Pathogenicity Island 12 That Contribute toIn VivoFitness of Salmonella enterica Serovar Typhimurium

2013 ◽  
Vol 81 (7) ◽  
pp. 2394-2404 ◽  
Author(s):  
Ana M. Tomljenovic-Berube ◽  
Brandyn Henriksbo ◽  
Steffen Porwollik ◽  
Colin A. Cooper ◽  
Brian R. Tuinema ◽  
...  
Keyword(s):  
Transcriptional Activation ◽  
Salmonella Enterica ◽  
Pathogenic Bacteria ◽  
Pathogenicity Island ◽  
Bacterial Survival ◽  
Content Type ◽  
Reverse Transcription Pcr ◽  
Serovar Typhimurium ◽  
Bacterial Fitness ◽  
Virulence Regulator

ABSTRACTSalmonellapathogenicity island 12 (SPI-12) ofSalmonella entericaserovar Typhimurium is a 15-kb region that encompasses genesSTM2230toSTM2245and encodes a remnant phage known to contribute to bacterial virulence. In mouse infection experiments and replication assays in macrophages, we demonstrated a role for four genes in SPI-12 for bacterial survival in the host. STM2239, a potential Q antiterminator, showed a prominent contribution to bacterial fitness. Transcriptional reporter experiments, quantitative reverse transcription-PCR (RT-PCR), and immunoblotting demonstrated that the virulence regulator SsrB and STM2239 contribute to transcriptional activation of genes in SPI-12. SsrB was found to indirectly regulate this locus by transcriptional read-through from thesspH2(STM2241) promoter. Chromatin immunoprecipitation showed that STM2239 copurified with the promoter regulatingSTM2237, suggesting that STM2239 may function as an antiterminator to activate adjacent genes. These results demonstrate that bacteriophage genes may be adapted by pathogenic bacteria to improve fitness in the host.

scholarly journals (p)ppGpp-dependent regulation of the nucleotide hydrolase PpnN confers complement resistance in Salmonella enterica serovar Typhimurium

2020 ◽  
Author(s):  
N. Y Elizabeth Chau ◽  
Deyanira Pérez-Morales ◽  
Wael Elhenawy ◽  
Víctor H. Bustamante ◽  
Yong E. Zhang ◽  
...  
Keyword(s):  
Salmonella Enterica ◽  
Pathogenic Bacteria ◽  
Stringent Response ◽  
Metabolic Reprogramming ◽  
Bacterial Survival ◽  
Complement Resistance ◽  
Serovar Typhimurium ◽  
Bacterial Fitness ◽  
Host Infection

The stringent response is an essential mechanism of metabolic reprogramming during environmental stress that is mediated by the nucleotide alarmones, guanosine tetraphosphate and pentaphosphate ((p)ppGpp). In addition to physiological adaptations, (p)ppGpp also regulates virulence programs in pathogenic bacteria including Salmonella enterica serovar Typhimurium. S. Typhimurium is a common cause of acute gastroenteritis, but it may also spread to systemic tissues resulting in severe clinical outcomes. During infection, S. Typhimurium encounters a broad repertoire of immune defenses that it must evade for successful host infection. Here, we examined the role of the stringent response in S. Typhimurium resistance to complement-mediated killing and found that the (p)ppGpp synthetase-hydrolase, SpoT, is required for bacterial survival in human serum. We identified the nucleotide hydrolase, PpnN, as a target of the stringent response that is required to promote bacterial fitness in serum. Using chromatography and mass spectrometry, we show that PpnN hydrolyzes purine and pyrimidine monophosphates to generate free nucleobases and ribose 5′-phosphate, and that this metabolic activity is required for conferring resistance to complement killing. In addition to PpnN, we show that (p)ppGpp is required for the biosynthesis of the very long and long O-antigen in the outer membrane known to be important for complement resistance. Our results provide new insights into the role of the stringent response in mediating evasion of the innate immune system by pathogenic bacteria.

scholarly journals SlyA and HilD Counteract H-NS-Mediated Repression on the ssrAB Virulence Operon of Salmonella enterica Serovar Typhimurium and Thus Promote Its Activation by OmpR

2019 ◽  
Vol 201 (8) ◽  
Author(s):  
María M. Banda ◽  
Crispín Zavala-Alvarado ◽  
Deyanira Pérez-Morales ◽  
Víctor H. Bustamante
Keyword(s):  
Salmonella Enterica ◽  
Salmonella Enterica Serovar Typhimurium ◽  
Growth Conditions ◽  
The Other ◽  
Regulatory Mechanisms ◽  
Two Component System ◽  
Content Type ◽  
Additive Action ◽  
Serovar Typhimurium ◽  
Bacterial Fitness

ABSTRACT H-NS-mediated repression of acquired genes and the subsequent adaptation of regulatory mechanisms that counteract this repression have played a central role in the Salmonella pathogenicity evolution. The Salmonella pathogenicity island 2 (SPI-2) is an acquired chromosomal region containing genes necessary for Salmonella enterica to colonize and replicate in different niches of hosts. The ssrAB operon, located in SPI-2, encodes the two-component system SsrA-SsrB, which positively controls the expression of the SPI-2 genes but also other many genes located outside SPI-2. Several regulators have been involved in the expression of ssrAB, such as the ancestral regulators SlyA and OmpR, and the acquired regulator HilD. In this study, we show how SlyA, HilD, and OmpR coordinate to induce the expression of ssrAB under different growth conditions. We found that when Salmonella enterica serovar Typhimurium is grown in nutrient-rich lysogeny broth (LB), SlyA and HilD additively counteract H-NS-mediated repression on ssrAB, whereas in N-minimal medium (N-MM), SlyA antagonizes H-NS-mediated repression on ssrAB independently of HilD. Interestingly, our results indicate that OmpR is required for the expression of ssrAB independently of the growth conditions, even in the absence of repression by H-NS. Therefore, our data support two mechanisms adapted for the expression of ssrAB under different growth conditions. One involves the additive action of SlyA and HilD, whereas the other involves SlyA, but not HilD, to counteract H-NS-mediated repression on ssrAB, thus favoring in both cases the activation of ssrAB by OmpR. IMPORTANCE The global regulator H-NS represses the expression of acquired genes and thus avoids possible detrimental effects on bacterial fitness. Regulatory mechanisms are adapted to induce expression of the acquired genes in particular niches to obtain a benefit from the information encoded in the foreign DNA, as for pathogenesis. Here, we show two mechanisms that were integrated for the expression of virulence genes in Salmonella Typhimurium. One involves the additive action of the regulators SlyA and HilD, whereas the other involves SlyA, but not HilD, to counteract H-NS-mediated repression on the ssrAB operon, thus favoring its activation by the OmpR regulator. To our knowledge, this is the first report involving the coordinated action of two regulators to counteract H-NS-mediated repression.

scholarly journals Quinazoline-Based Antivirulence Compounds Selectively Target Salmonella PhoP/PhoQ Signal Transduction System

2019 ◽  
Vol 64 (1) ◽  
Author(s):  
María Ayelén Carabajal ◽  
Christopher R. M. Asquith ◽  
Tuomo Laitinen ◽  
Graham J. Tizzard ◽  
Lucía Yim ◽  
...  
Keyword(s):  
Signal Transduction ◽  
Salmonella Enterica ◽  
Pathogenic Bacteria ◽  
Catalytic Domain ◽  
Kinase Inhibitor ◽  
Adaptive Responses ◽  
Antibacterial Compounds ◽  
Content Type ◽  
Two Component ◽  
Serovar Typhimurium

ABSTRACT The rapid emergence of multidrug resistance among bacterial pathogens has become a significant challenge to human health in our century. Therefore, development of next-generation antibacterial compounds is an urgent need. Two-component signal transduction systems (TCS) are stimulus-response coupling devices that allow bacteria to sense and elaborate adaptive responses to changing environmental conditions, including the challenges that pathogenic bacteria face inside the host. The differential presence of TCS, present in bacteria but absent in the animal kingdom, makes them attractive targets in the search for new antibacterial compounds. In Salmonella enterica, the PhoP/PhoQ two-component system controls the expression of crucial phenotypes that define the ability of the pathogen to establish infection in the host. We now report the screening of 686 compounds from a GlaxoSmithKline published kinase inhibitor set in a high-throughput whole-cell assay that targets Salmonella enterica serovar Typhimurium PhoP/PhoQ. We identified a series of quinazoline compounds that showed selective and potent downregulation of PhoP/PhoQ-activated genes and define structural attributes required for their efficacy. We demonstrate that their bioactivity is due to repression of the PhoQ sensor autokinase activity mediated by interaction with its catalytic domain, acting as competitive inhibitors of ATP binding. While noncytotoxic, the hit molecules exhibit antivirulence effect by blockage of S. Typhimurium intramacrophage replication. Together, these features make these quinazoline compounds stand out as exciting leads to develop a therapeutic intervention to fight salmonellosis.

scholarly journals Dormant Intracellular Salmonella enterica Serovar Typhimurium Discriminates among Salmonella Pathogenicity Island 2 Effectors To Persist inside Fibroblasts

2013 ◽  
Vol 82 (1) ◽  
pp. 221-232 ◽  
Author(s):  
Cristina Núñez-Hernández ◽  
Ana Alonso ◽  
M. Graciela Pucciarelli ◽  
Josep Casadesús ◽  
Francisco García-del Portillo
Keyword(s):  
Salmonella Enterica ◽  
Pathogenicity Island ◽  
Effector Proteins ◽  
Intracellular Bacteria ◽  
Secretion Systems ◽  
Cultured Fibroblasts ◽  
Content Type ◽  
Type Iii Secretion Systems ◽  
Serovar Typhimurium

ABSTRACTSalmonella entericauses effector proteins delivered by type III secretion systems (TTSS) to colonize eukaryotic cells. Recentin vivostudies have shown that intracellular bacteria activate the TTSS encoded bySalmonellapathogenicity island-2 (SPI-2) to restrain growth inside phagocytes. Growth attenuation is also observedin vivoin bacteria colonizing nonphagocytic stromal cells of the intestinal lamina propria and in cultured fibroblasts. SPI-2 is required for survival of nongrowing bacteria persisting inside fibroblasts, but its induction mode and the effectors involved remain unknown. Here, we show that nongrowing dormant intracellular bacteria use the two-component system OmpR-EnvZ to induce SPI-2 expression and the PhoP-PhoQ system to regulate the time at which induction takes place, 2 h postentry. Dormant bacteria were shown to discriminate the usage of SPI-2 effectors. Among the effectors tested, SseF, SseG, and SseJ were required for survival, while others, such as SifA and SifB, were not. SifA and SifB dispensability correlated with the inability of intracellular bacteria to secrete these effectors even when overexpressed. Conversely, SseJ overproduction resulted in augmented secretion and exacerbated bacterial growth. Dormant bacteria produced other effectors, such as PipB and PipB2, that, unlike what was reported for epithelial cells, did not to traffic outside the phagosomal compartment. Therefore, permissiveness for secreting only a subset of SPI-2 effectors may be instrumental for dormancy. We propose that theS. entericaserovar Typhimurium nonproliferative intracellular lifestyle is sustained by selection of SPI-2 effectors that are produced in tightly defined amounts and delivered to phagosome-confined locations.

scholarly journals Flagellin Is Required for Host Cell Invasion and Normal Salmonella Pathogenicity Island 1 Expression by Salmonella enterica Serovar Paratyphi A

2015 ◽  
Vol 83 (9) ◽  
pp. 3355-3368 ◽  
Author(s):  
Dana Elhadad ◽  
Prerak Desai ◽  
Galia Rahav ◽  
Michael McClelland ◽  
Ohad Gal-Mor
Keyword(s):  
Host Cell ◽  
Cell Invasion ◽  
Salmonella Enterica ◽  
Pathogenicity Island ◽  
Dependent Manner ◽  
Content Type ◽  
Type Three Secretion System ◽  
Serovar Typhimurium ◽  
System 1 ◽  
Type Three Secretion

Salmonella entericaserovar Paratyphi A is a human-specific serovar that, together withSalmonella entericaserovar Typhi andSalmonella entericaserovar Sendai, causes enteric fever. Unlike the nontyphoidalSalmonella entericaserovar Typhimurium, the genomes ofS. Typhi andS. Paratyphi A are characterized by inactivation of multiple genes, including in the flagellum-chemotaxis pathway. Here, we explored the motility phenotype ofS. Paratyphi A and the role of flagellin in key virulence-associated phenotypes. Motility studies established that the human-adapted typhoidalS. Typhi,S. Paratyphi A, andS. Sendai are all noticeably less motile thanS. Typhimurium, and comparative transcriptome sequencing (RNA-Seq) showed that inS. Paratyphi A, the entire motility-chemotaxis regulon is expressed at significantly lowers levels than inS. Typhimurium. Nevertheless,S. Paratyphi A, likeS. Typhimurium, requires a functional flagellum for epithelial cell invasion and macrophage uptake, probably in a motility-independent mechanism. In contrast, flagella were found to be dispensable for host cell adhesion. Moreover, we demonstrate that inS. Paratyphi A, but not inS. Typhimurium, the lack of flagellin results in increased transcription of the flagellar and theSalmonellapathogenicity island 1 (SPI-1) regulons in a FliZ-dependent manner and in oversecretion of SPI-1 effectors via type three secretion system 1. Collectively, these results suggest a novel regulatory linkage between flagellin and SPI-1 inS. Paratyphi A that does not occur inS. Typhimurium and demonstrate curious distinctions in motility and the expression of the flagellum-chemotaxis regulon between these clinically relevant pathogens.

scholarly journals Protozoan Cysts Act as a Survival Niche and Protective Shelter for Foodborne Pathogenic Bacteria

2015 ◽  
Vol 81 (16) ◽  
pp. 5604-5612 ◽  
Author(s):  
Ellen Lambrecht ◽  
Julie Baré ◽  
Natascha Chavatte ◽  
Wim Bert ◽  
Koen Sabbe ◽  
...  
Keyword(s):  
Salmonella Enterica ◽  
Pathogenic Bacteria ◽  
Acanthamoeba Castellanii ◽  
Bacterial Survival ◽  
Free Living ◽  
Active Growth ◽  
Content Type ◽  
Foodborne Pathogenic Bacteria ◽  
Adverse Environmental Conditions ◽  
Species Specific

ABSTRACTThe production of cysts, an integral part of the life cycle of many free-living protozoa, allows these organisms to survive adverse environmental conditions. Given the prevalence of free-living protozoa in food-related environments, it is hypothesized that these organisms play an important yet currently underinvestigated role in the epidemiology of foodborne pathogenic bacteria. Intracystic bacterial survival is highly relevant, as this would allow bacteria to survive the stringent cleaning and disinfection measures applied in food-related environments. The present study shows that strains of widespread and important foodborne bacteria (Salmonella enterica,Escherichia coli,Yersinia enterocolitica, andListeria monocytogenes) survive inside cysts of the ubiquitous amoebaAcanthamoeba castellanii, even when exposed to either antibiotic treatment (100 μg/ml gentamicin) or highly acidic conditions (pH 0.2) and resume active growth in broth media following excystment. Strain- and species-specific differences in survival periods were observed, withSalmonella entericasurviving up to 3 weeks inside amoebal cysts. Up to 53% of the cysts were infected with pathogenic bacteria, which were located in the cyst cytosol. Our study suggests that the role of free-living protozoa and especially their cysts in the persistence and epidemiology of foodborne bacterial pathogens in food-related environments may be much more important than hitherto assumed.

scholarly journals Bistable Expression of CsgD in Salmonella enterica Serovar Typhimurium Connects Virulence to Persistence

2015 ◽  
Vol 83 (6) ◽  
pp. 2312-2326 ◽  
Author(s):  
Keith D. MacKenzie ◽  
Yejun Wang ◽  
Dylan J. Shivak ◽  
Cynthia S. Wong ◽  
Leia J. L. Hoffman ◽  
...  
Keyword(s):  
Salmonella Enterica ◽  
Pathogenic Bacteria ◽  
Single Cells ◽  
Smooth Transition ◽  
Intestinal Cell ◽  
Planktonic Cells ◽  
Multicellular Aggregate ◽  
Content Type ◽  
Serovar Typhimurium ◽  
Cell Subpopulations

Pathogenic bacteria often need to survive in the host and the environment, and it is not well understood how cells transition between these equally challenging situations. For the human and animal pathogenSalmonella entericaserovar Typhimurium, biofilm formation is correlated with persistence outside a host, but the connection to virulence is unknown. In this study, we analyzed multicellular-aggregate and planktonic-cell subpopulations that coexist whenS. Typhimurium is grown under biofilm-inducing conditions. These cell types arise due to bistable expression of CsgD, the central biofilm regulator. Despite being exposed to the same stresses, the two cell subpopulations had 1,856 genes that were differentially expressed, as determined by transcriptome sequencing (RNA-seq). Aggregated cells displayed the characteristic gene expression of biofilms, whereas planktonic cells had enhanced expression of numerous virulence genes. Increased type three secretion synthesis in planktonic cells correlated with enhanced invasion of a human intestinal cell line and significantly increased virulence in mice compared to the aggregates. However, when the same groups of cells were exposed to desiccation, the aggregates survived better, and the competitive advantage of planktonic cells was lost. We hypothesize that CsgD-based differentiation is a form of bet hedging, with single cells primed for host cell invasion and aggregated cells adapted for persistence in the environment. This allowsS. Typhimurium to spread the risks of transmission and ensures a smooth transition between the host and the environment.

scholarly journals Sensitization of Intracellular Salmonella enterica Serovar Typhimurium to AminoglycosidesIn VitroandIn Vivoby a Host-Targeted Antimicrobial Agent

2014 ◽  
Vol 58 (12) ◽  
pp. 7375-7382 ◽  
Author(s):  
Jung-Hsin Lo ◽  
Samuel K. Kulp ◽  
Ching-Shih Chen ◽  
Hao-Chieh Chiu
Keyword(s):  
Salmonella Enterica ◽  
Antibacterial Activities ◽  
Eukaryotic Cell ◽  
Bacterial Survival ◽  
Content Type ◽  
Raw264.7 Macrophages ◽  
Serovar Typhimurium ◽  
Sensitizing Effect ◽  
Unique Ability

ABSTRACTAminoglycosides exhibit relatively poor activity against intracellularSalmonella entericaserovar Typhimurium due to their low permeativity across eukaryotic cell membranes. Previously, we identified the unique ability of AR-12, a celecoxib-derived small-molecule agent, to eradicate intracellularSalmonellaTyphimurium in macrophages by facilitating autophagosome formation and suppressing Akt kinase signaling. In light of this unique mode of antibacterial action, we investigated the ability of AR-12 to sensitize intracellularSalmonellato aminoglycosides in macrophages and in an animal model. The antibacterial activities of AR-12 combined with various aminoglycosides, including streptomycin, kanamycin, gentamicin, and amikacin, against intracellularS. Typhimurium in murine RAW264.7 macrophages were assessed. Cells were infected withS. Typhimurium followed by treatment with AR-12 or individual aminoglycosides or with combinations for 24 h. Thein vivoefficacies of AR-12, alone or in combination with gentamicin or amikacin, were also assessed by treatingS. Typhimurium-infected BALB/c mice daily for 14 consecutive days. Exposure ofS. Typhimurium-infected RAW264.7 cells to a combination of AR-12 with individual aminoglycosides led to a reduction in bacterial survival (P< 0.05), both intracellular and extracellular, that was greater than that seen with the aminoglycosides alone. This sensitizing effect, however, was not associated with increased aminoglycoside penetration into bacteria or macrophages. Moreover, daily intraperitoneal injection of AR-12 at 0.1 mg/kg of body weight significantly increased thein vivoefficacy of gentamicin and amikacin in prolonging the survival ofS. Typhimurium-infected mice. These findings indicate that the unique ability of AR-12 to enhance thein vivoefficacy of aminoglycosides might have translational potential for efforts to develop novel strategies for the treatment of salmonellosis.

scholarly journals Efficiency of Conditionally Attenuated Salmonella enterica Serovar Typhimurium in Bacterium-Mediated Tumor Therapy

mBio ◽  
2015 ◽  
Vol 6 (2) ◽  
Author(s):  
Michael Frahm ◽  
Sebastian Felgner ◽  
Dino Kocijancic ◽  
Manfred Rohde ◽  
Michael Hensel ◽  
...  
Keyword(s):  
Salmonella Enterica ◽  
Tumor Targeting ◽  
Antitumor Effect ◽  
Treatment Options ◽  
Tumor Therapy ◽  
Therapeutic Benefit ◽  
Industrialized Countries ◽  
Content Type ◽  
Serovar Typhimurium ◽  
Therapeutic Molecules

ABSTRACTIncreasing numbers of cancer cases generate a great urge for new treatment options. Applying bacteria likeSalmonella entericaserovar Typhimurium for cancer therapy represents an intensively explored option. These bacteria have been shown not only to colonize solid tumors but also to exhibit an intrinsic antitumor effect. In addition, they could serve as tumor-targeting vectors for therapeutic molecules. However, the pathogenicS. Typhimurium strains used for tumor therapy need to be attenuated for safe application. Here, lipopolysaccharide (LPS) deletion mutants (ΔrfaL, ΔrfaG, ΔrfaH, ΔrfaD, ΔrfaP, and ΔmsbBmutants) ofSalmonellawere investigated for efficiency in tumor therapy. Of such variants, the ΔrfaDand ΔrfaGdeep rough mutants exhibited the best tumor specificity and lowest pathogenicity. However, the intrinsic antitumor effect was found to be weak. To overcome this limitation, conditional attenuation was tested by complementing the mutants with an inducible arabinose promoter. The chromosomal integration of the respective LPS biosynthesis genes into thearaBADlocus exhibited the best balance of attenuation and therapeutic benefit. Thus, the present study establishes a basis for the development of an applicably cancer therapeutic bacterium.IMPORTANCECancer has become the second most frequent cause of death in industrialized countries. This and the drawbacks of routine therapies generate an urgent need for novel treatment options. Applying appropriately modifiedS. Typhimurium for therapy represents the major challenge of bacterium-mediated tumor therapy. In the present study, we demonstrated thatSalmonellabacteria conditionally modified in their LPS phenotype exhibit a safe tumor-targeting phenotype. Moreover, they could represent a suitable vehicle to shuttle therapeutic compounds directly into cancerous tissue without harming the host.

scholarly journals Whole-Genome Sequence of Salmonella enterica subsp. enterica Serovar Typhimurium Strain UPM 260, Isolated from a Broiler Chicken in Perak, Malaysia

2017 ◽  
Vol 5 (46) ◽  
Author(s):  
Najwa Syahirah Roslan ◽  
Shagufta Jabeen ◽  
Nurulfiza Mat Isa ◽  
Abdul Rahman Omar ◽  
Mohd Hair Bejo ◽  
...  
Keyword(s):  
Genome Sequence ◽  
Salmonella Enterica ◽  
Broiler Chicken ◽  
Whole Genome Sequence ◽  
Whole Genome ◽  
Content Type ◽  
Typhimurium Strain ◽  
Serovar Typhimurium

ABSTRACT Salmonella enterica subsp. enterica serovar Typhimurium is one of several well-categorized Salmonella serotypes recognized globally. Here, we report the whole-genome sequence of S. Typhimurium strain UPM 260, isolated from a broiler chicken.

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