scholarly journals RNA Sequencing Reveals Differences between the Global Transcriptomes of Salmonella enterica Serovar Enteritidis Strains with High and Low Pathogenicities

2013 ◽  
Vol 80 (3) ◽  
pp. 896-906 ◽  
Author(s):  
Devendra H. Shah
Keyword(s):  
Rna Sequencing ◽  
Salmonella Enterica ◽  
Negative Binomial ◽  
Phenotypic Diversity ◽  
Functional Characterization ◽  
Salmonella Enterica Serovar Enteritidis ◽  
Content Type ◽  
Expression Of Genes

ABSTRACTSalmonella entericaserovar Enteritidis is one of the important causes of bacterial food-borne gastroenteritis worldwide. Field strains ofS. Enteritidis are relatively genetically homogeneous; however, they show extensive phenotypic diversity and differences in virulence potential. RNA sequencing (RNA-Seq) was used to characterize differences in the global transcriptome between several genetically similar but phenotypically diverse poultry-associated field strains ofS. Enteritidis grown in laboratory medium at avian body temperature (42°C). TheseS. Enteritidis strains were previously characterized as high-pathogenicity (HP;n= 3) and low-pathogenicity (LP;n= 3) strains based on bothin vitroandin vivovirulence assays. Using the negative binomial distribution-based statistical tools edgeR and DESeq, 252 genes were identified as differentially expressed in LP strains compared with their expression in the HP strains (P< 0.05). A majority of genes (235, or 93.2%) showed significantly reduced expression, whereas a few genes (17, or 6.8%) showed increased expression in all LP strains compared with HP strains. LP strains showed a unique transcriptional profile that is characterized by significantly reduced expression of several transcriptional regulators and reduced expression of genes involved in virulence (e.g.,Salmonellapathogenicity island 1 [SPI-1], SPI-5, and fimbrial and motility genes) and protection against osmotic, oxidative, and other stresses, such as iron-limiting conditions commonly encountered within the host. Several functionally uncharacterized genes also showed reduced expression. This study provides a first concise view of the global transcriptional differences between field strains ofS. Enteritidis with various levels of pathogenicity, providing the basis for future functional characterization of several genes with potential roles in virulence or stress regulation ofS. Enteritidis.

scholarly journals An Inducible and Secreted Eukaryote-Like Serine/Threonine Kinase of Salmonella enterica Serovar Typhi Promotes Intracellular Survival and Pathogenesis

2014 ◽  
Vol 83 (2) ◽  
pp. 522-533 ◽  
Author(s):  
Nagaraja Theeya ◽  
Atri Ta ◽  
Sayan Das ◽  
Rahul S. Mandal ◽  
Oishee Chakrabarti ◽  
...  
Keyword(s):  
Salmonella Enterica ◽  
Functional Characterization ◽  
Systemic Infection ◽  
Serine Threonine Kinase ◽  
Content Type ◽  
Threonine Kinase ◽  
Universal Substrate

Eukaryote-like serine/threonine kinases (eSTKs) constitute an important family of bacterial virulence factors. Genome analysis had predicted putative eSTKs inSalmonella entericaserovar Typhi, although their functional characterization and the elucidation of their role in pathogenesis are still awaited. We show here that the primary sequence and secondary structure of thet4519locus ofSalmonellaTyphi Ty2 have all the signatures of eukaryotic superfamily kinases.t4519encodes a ∼39-kDa protein (T4519), which shows serine/threonine kinase activitiesin vitro. Recombinant T4519 (rT4519) is autophosphorylated and phosphorylates the universal substrate myelin basic protein. Infection of macrophages results in decreased viability of the mutant (Ty2Δt4519) strain, which is reversed by gene complementation. Moreover, reactive oxygen species produced by the macrophages signal to the bacteria to induce T4519, which is translocated to the host cell cytoplasm. That T4519 may target a host substrate(s) is further supported by the activation of host cellular signaling pathways and the induction of cytokines/chemokines. Finally, the role of T4519 in the pathogenesis ofSalmonellaTyphi is underscored by the significantly decreased mortality of mice infected with the Ty2Δt4519strain and the fact that the competitive index of this strain for causing systemic infection is 0.25% that of the wild-type strain. This study characterizes the first eSTK ofSalmonellaTyphi and demonstrates its role in promoting phagosomal survival of the bacteria within macrophages, which is a key determinant of pathogenesis. This, to the best of our knowledge, is the first study to describe the essential role of eSTKs in thein vivopathogenesis ofSalmonellaspp.

scholarly journals Microarray-Based Detection of Salmonella enterica Serovar Enteritidis Genes Involved in Chicken Reproductive Tract Colonization

2014 ◽  
Vol 80 (24) ◽  
pp. 7710-7716 ◽  
Author(s):  
R. Raspoet ◽  
C. Appia-Ayme ◽  
N. Shearer ◽  
A. Martel ◽  
F. Pasmans ◽  
...  
Keyword(s):  
Stress Responses ◽  
Salmonella Enterica ◽  
Reproductive Tract ◽  
Genomic Islands ◽  
Salmonella Enterica Serovar Enteritidis ◽  
Content Type ◽  
Library Screen ◽  
Chicken Oviduct

ABSTRACTSalmonella entericaserovar Enteritidis has developed the potential to contaminate table eggs internally, by colonization of the chicken reproductive tract and internalization in the forming egg. The serotype Enteritidis has developed mechanisms to colonize the chicken oviduct more successfully than other serotypes. Until now, the strategies exploited bySalmonellaEnteritidis to do so have remained largely unknown. For that reason, a microarray-based transposon library screen was used to identify genes that are essential for the persistence ofSalmonellaEnteritidis inside primary chicken oviduct gland cellsin vitroand inside the reproductive tractin vivo. A total of 81 genes with a potential role in persistence in both the oviduct cells and the oviduct tissue were identified. Major groups of importance include theSalmonellapathogenicity islands 1 and 2, genes involved in stress responses, cell wall, and lipopolysaccharide structure, and the region-of-difference genomic islands 9, 21, and 40.

scholarly journals Terminal Deoxynucleotidyl Transferase Is Not Required for Antibody Response to Polysaccharide Vaccines againstStreptococcus pneumoniaeandSalmonella entericaSerovar Typhi

2018 ◽  
Vol 86 (9) ◽  
Author(s):  
Vivek Belde ◽  
Matthew P. Cravens ◽  
Dania Gulandijany ◽  
Justin A. Walker ◽  
Isabel Palomo-Caturla ◽  
...  
Keyword(s):  
Antibody Response ◽  
B Cell ◽  
Salmonella Enterica ◽  
Passive Immunization ◽  
Terminal Deoxynucleotidyl Transferase ◽  
Human Infants ◽  
Content Type ◽  
Serovar Typhimurium

ABSTRACTB cell antigen receptor (BCR) diversity increases by several orders of magnitude due to the action of terminal deoxynucleotidyl transferase (TdT) during V(D)J recombination. Unlike adults, infants have limited BCR diversity, in part due to reduced expression of TdT. Since human infants and young mice respond poorly to polysaccharide vaccines, such as the pneumococcal polysaccharide vaccine Pneumovax23 and Vi polysaccharide (ViPS) ofSalmonella entericaserovar Typhi, we tested the contribution of TdT-mediated BCR diversity in response to these vaccines. We found that TdT+/−and TdT−/−mice generated comparable antibody responses to Pneumovax23 and survivedStreptococcus pneumoniaechallenge. Moreover, passive immunization of B cell-deficient mice with serum from Pneumovax23-immunized TdT+/−or TdT−/−mice conferred protection. TdT+/−and TdT−/−mice generated comparable levels of anti-ViPS antibodies and antibody-dependent, complement-mediated bactericidal activity againstS. Typhiin vitro. To test the protective immunity conferred by ViPS immunizationin vivo, TdT+/−and TdT−/−mice were challenged with a chimericSalmonella entericaserovar Typhimurium strain expressing ViPS, since mice are nonpermissive hosts forS. Typhi infection. Compared to their unimmunized counterparts, immunized TdT+/−and TdT−/−mice challenged with ViPS-expressingS. Typhimurium exhibited a significant reduction in the bacterial burden and liver pathology. These data suggest that the impaired antibody response to the Pneumovax23 and ViPS vaccines in the young is not due to limited TdT-mediated BCR diversification.

scholarly journals Interaction of Antibiotics with Innate Host Defense Factors against Salmonella enterica Serotype Newport

mSphere ◽  
2017 ◽  
Vol 2 (6) ◽  
Author(s):  
George Sakoulas ◽  
Monika Kumaraswamy ◽  
Armin Kousha ◽  
Victor Nizet
Keyword(s):  
Innate Immunity ◽  
Immune System ◽  
Innate Immune System ◽  
Salmonella Enterica ◽  
Clinical Performance ◽  
Innate Immune ◽  
Content Type ◽  
Antimicrobial Assay

ABSTRACT It is becoming increasingly understood that the current paradigms of in vitro antimicrobial susceptibility testing may have significant shortcomings in predicting activity in vivo. This study evaluated the activity of several antibiotics alone and in combination against clinical isolates of Salmonella enterica serotype Newport (meningitis case) utilizing both conventional and physiological media. In addition, the interactions of these antibiotics with components of the innate immune system were evaluated. Azithromycin, which has performed quite well clinically despite high MICs in conventional media, was shown to be more active in physiological media and to enhance innate immune system killing. Alternatively, chloramphenicol did not show enhanced immune system killing, paralleling its inferior clinical performance to other antibiotics that have been used to treat Salmonella meningitis. These findings are important additions to the building understanding of current in vitro antimicrobial assay limitations that hopefully will amount to future improvements in these assays to better predict clinical efficacy and activity in vivo. This study examines the pharmacodynamics of antimicrobials that are used to treat Salmonella with each other and with key components of the innate immune system. Antimicrobial synergy was assessed using time-kill and checkerboard assays. Antimicrobial interactions with innate immunity were studied by employing cathelicidin LL-37, whole-blood, and neutrophil killing assays. Ceftriaxone and ciprofloxacin were found to be synergistic in vitro against Salmonella enterica serotype Newport. Ceftriaxone, ciprofloxacin, and azithromycin each demonstrated synergy with the human cathelicidin defense peptide LL-37 in killing Salmonella. Exposure of Salmonella to sub-MICs of ceftriaxone resulted in enhanced susceptibility to LL-37, whole blood, and neutrophil killing. The activity of antibiotics in vivo against Salmonella may be underestimated in bacteriologic media lacking components of innate immunity. The pharmacodynamic interactions of antibiotics used to treat Salmonella with each other and with components of innate immunity warrant further study in light of recent findings showing in vivo selection of antimicrobial resistance by single agents in this pathogen. IMPORTANCE It is becoming increasingly understood that the current paradigms of in vitro antimicrobial susceptibility testing may have significant shortcomings in predicting activity in vivo. This study evaluated the activity of several antibiotics alone and in combination against clinical isolates of Salmonella enterica serotype Newport (meningitis case) utilizing both conventional and physiological media. In addition, the interactions of these antibiotics with components of the innate immune system were evaluated. Azithromycin, which has performed quite well clinically despite high MICs in conventional media, was shown to be more active in physiological media and to enhance innate immune system killing. Alternatively, chloramphenicol did not show enhanced immune system killing, paralleling its inferior clinical performance to other antibiotics that have been used to treat Salmonella meningitis. These findings are important additions to the building understanding of current in vitro antimicrobial assay limitations that hopefully will amount to future improvements in these assays to better predict clinical efficacy and activity in vivo.

scholarly journals Poultry Body Temperature Contributes to Invasion Control through Reduced Expression of Salmonella Pathogenicity Island 1 Genes in Salmonella enterica Serovars Typhimurium and Enteritidis

2015 ◽  
Vol 81 (23) ◽  
pp. 8192-8201 ◽  
Author(s):  
Bryan Troxell ◽  
Nicholas Petri ◽  
Caitlyn Daron ◽  
Rafaela Pereira ◽  
Mary Mendoza ◽  
...  
Keyword(s):  
Body Temperature ◽  
Foodborne Pathogens ◽  
Salmonella Enterica ◽  
Pathogenicity Island ◽  
Avian Host ◽  
Content Type ◽  
Poultry Products ◽  
Epithelial Cell Layer

ABSTRACTSalmonella entericaserovars Typhimurium (S. Typhimurium) and Enteritidis (S. Enteritidis) are foodborne pathogens, and outbreaks are often associated with poultry products. Chickens are typically asymptomatic when colonized by these serovars; however, the factors contributing to this observation are uncharacterized. Whereas symptomatic mammals have a body temperature between 37°C and 39°C, chickens have a body temperature of 41°C to 42°C. Here,in vivoexperiments using chicks demonstrated that numbers of viableS. Typhimurium orS. Enteritidis bacteria within the liver and spleen organ sites were ≥4 orders of magnitude lower than those within the ceca. When similar doses ofS. Typhimurium orS. Enteritidis were given to C3H/HeN mice, the ratio of the intestinal concentration to the liver/spleen concentration was 1:1. In the avian host, this suggested poor survival within these tissues or a reduced capacity to traverse the host epithelial layer and reach liver/spleen sites or both.Salmonellapathogenicity island 1 (SPI-1) promotes localization to liver/spleen tissues through invasion of the epithelial cell layer. Followingin vitrogrowth at 42°C, SPI-1 genessipC,invF, andhilAand the SPI-1rtsAactivator were downregulated compared to expression at 37°C. Overexpression of thehilAactivatorsfur,fliZ, andhilDwas capable of inducinghilA-lacZat 37°C but not at 42°C despite the presence of similar levels of protein at the two temperatures. In contrast, overexpression of eitherhilCorrtsAwas capable of inducinghilAandsipCat 42°C. These data indicate that physiological parameters of the poultry host, such as body temperature, have a role in modulating expression of virulence.

scholarly journals Transcriptional Regulator LsrB of Sinorhizobium meliloti Positively Regulates the Expression of Genes Involved in Lipopolysaccharide Biosynthesis

2014 ◽  
Vol 80 (17) ◽  
pp. 5265-5273 ◽  
Author(s):  
Guirong Tang ◽  
Ying Wang ◽  
Li Luo
Keyword(s):  
Sinorhizobium Meliloti ◽  
Sodium Dodecyl ◽  
Host Cells ◽  
Content Type ◽  
Expression Of Genes ◽  
New Findings ◽  
Promoter Dna ◽  
Transcriptional Start Sites

ABSTRACTRhizobia induce nitrogen-fixing nodules on host legumes, which is important in agriculture and ecology. Lipopolysaccharide (LPS) produced by rhizobia is required for infection or bacteroid survival in host cells. Genes required for LPS biosynthesis have been identified in severalRhizobiumspecies. However, the regulation of their expression is not well understood. Here,Sinorhizobium melilotiLsrB, a member of the LysR family of transcriptional regulators, was found to be involved in LPS biosynthesis by positively regulating the expression of thelrp3-lpsCDEoperon. AnlsrBin-frame deletion mutant displayed growth deficiency, sensitivity to the detergent sodium dodecyl sulfate, and acidic pH compared to the parent strain. This mutant produced slightly less LPS due to lower expression of thelrp3operon. Analysis of the transcriptional start sites of thelrp3andlpsCDEgene suggested that they constitute one operon. The expression oflsrBwas positively autoregulated. The promoter region oflrp3was specifically precipitated by anti-LsrB antibodiesin vivo. The promoter DNA fragment containing TN11A motifs was bound by the purified LsrB proteinin vitro. These new findings suggest thatS. melilotiLsrB is associated with LPS biosynthesis, which is required for symbiotic nitrogen fixation on some ecotypes of alfalfa plants.

scholarly journals Salmonella Extracellular Matrix Components Influence Biofilm Formation and Gallbladder Colonization

2016 ◽  
Vol 84 (11) ◽  
pp. 3243-3251 ◽  
Author(s):  
Haley E. Adcox ◽  
Erin M. Vasicek ◽  
Varun Dwivedi ◽  
Ky V. Hoang ◽  
Joanne Turner ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Typhoid Fever ◽  
Salmonella Enterica ◽  
Biofilm Formation ◽  
Bacterial Survival ◽  
Content Type ◽  
Extracellular Matrix Components ◽  
Matrix Components

Salmonella enterica serovar Typhi, the causative agent of typhoid fever in humans, forms biofilms encapsulated by an extracellular matrix (ECM). Biofilms facilitate colonization and persistent infection in gallbladders of humans and mouse models of chronic carriage. Individual roles of matrix components have not been completely elucidated in vitro or in vivo . To examine individual functions, strains of Salmonella enterica serovar Typhimurium, the murine model of S . Typhi, in which various ECM genes were deleted or added, were created to examine biofilm formation, colonization, and persistence in the gallbladder. Studies show that curli contributes most significantly to biofilm formation. Expression of Vi antigen decreased biofilm formation in vitro and virulence and bacterial survival in vivo without altering the examined gallbladder pro- or anti-inflammatory cytokines. Oppositely, loss of all ECM components (Δ wcaM Δ csgA Δ yihO Δ bcsE ) increased virulence and bacterial survival in vivo and reduced gallbladder interleukin-10 (IL-10) levels. Colanic acid and curli mutants had the largest defects in biofilm-forming ability and contributed most significantly to the virulence increase of the Δ wcaM Δ csgA Δ yihO Δ bcsE mutant strain. While the Δ wcaM Δ csgA Δ yihO Δ bcsE mutant was not altered in resistance to complement or growth in macrophages, it attached and invaded macrophages better than the wild-type (WT) strain. These data suggest that ECM components have various levels of importance in biofilm formation and gallbladder colonization and that the ECM diminishes disseminated disease in our model, perhaps by reducing cell attachment/invasion and dampening inflammation by maintaining/inducing IL-10 production. Understanding how ECM components aid acute disease and persistence could lead to improvements in therapeutic treatment of typhoid fever patients.

scholarly journals Expression and Functional Characterization of the First Bacteriophage-Encoded Chaperonin

2012 ◽  
Vol 86 (18) ◽  
pp. 10103-10111 ◽  
Author(s):  
Lidia P. Kurochkina ◽  
Pavel I. Semenyuk ◽  
Victor N. Orlov ◽  
Johan Robben ◽  
Nina N. Sykilinda ◽  
...  
Keyword(s):  
Thermal Inactivation ◽  
Functional Characterization ◽  
Content Type ◽  
Chaperonin Groel ◽  
Phage Propagation ◽  
Physicochemical Methods ◽  
First Time

Chaperonins promote protein foldingin vivoand are ubiquitously found in bacteria, archaea, and eukaryotes. The first viral chaperonin GroEL ortholog, gene product 146 (gp146), whose gene was earlier identified in the genome of bacteriophage EL, has been shown to be synthesized during phage propagation inPseudomonas aeruginosacells. The recombinant gp146 has been expressed inEscherichia coliand characterized by different physicochemical methods for the first time. Using serum against the recombinant protein, gp146's native substrate, the phage endolysin gp188, has been immunoprecipitated from the lysate of EL-infected bacteria and identified by mass spectrometry.In vitroexperiments have shown that gp146 has a protective effect against endolysin thermal inactivation and aggregation, providing evidence of its chaperonin function. The phage chaperonin has been found to have the architecture and some properties similar to those of GroEL but not to require cochaperonin for its functional activity.

scholarly journals Infection of Mice by Salmonella enterica Serovar Enteritidis Involves Additional Genes That Are Absent in the Genome of Serovar Typhimurium

2011 ◽  
Vol 80 (2) ◽  
pp. 839-849 ◽  
Author(s):  
Cecilia A. Silva ◽  
Carlos J. Blondel ◽  
Carolina P. Quezada ◽  
Steffen Porwollik ◽  
Helene L. Andrews-Polymenis ◽  
...  
Keyword(s):  
Salmonella Enterica ◽  
Phage Type ◽  
Pathogenicity Island ◽  
Genomic Islands ◽  
Enteric Infection ◽  
Type I ◽  
Modification System ◽  
Content Type

ABSTRACTSalmonella entericaserovar Enteritidis causes a systemic, typhoid-like infection in newly hatched poultry and mice. In the present study, a library of 54,000 transposon mutants ofS.Enteritidis phage type 4 (PT4) strain P125109 was screened for mutants deficient in thein vivocolonization of the BALB/c mouse model using a microarray-based negative-selection screening. Mutants in genes known to contribute to systemic infection (e.g.,Salmonellapathogenicity island 2 [SPI-2],aro,rfa,rfb,phoP, andphoQ) and enteric infection (e.g., SPI-1 and SPI-5) in this and otherSalmonellaserovars displayed colonization defects in our assay. In addition, a strong attenuation was observed for mutants in genes and genomic islands that are not present inS.Typhimurium or in most otherSalmonellaserovars. These genes include a type I restriction/modification system (SEN4290toSEN4292), thepegfimbrial operon (SEN2144AtoSEN2145B), a putative pathogenicity island (SEN1970 to SEN1999), and a type VI secretion system remnantSEN1001, encoding a hypothetical protein containing a lysin motif (LysM) domain associated with peptidoglycan binding. Proliferation defects for mutants in these individual genes and in exemplar genes for each of these clusters were confirmed in competitive infections with wild-typeS.Enteritidis. A ΔSEN1001mutant was defective for survival within RAW264.7 murine macrophagesin vitro. Complementation assays directly linked theSEN1001gene to phenotypes observedin vivoandin vitro. The genes identified here may perform novel virulence functions not characterized in previousSalmonellamodels.

scholarly journals In Salmonella enterica, the Gcn5-Related Acetyltransferase MddA (Formerly YncA) Acetylates Methionine Sulfoximine and Methionine Sulfone, Blocking Their Toxic Effects

2014 ◽  
Vol 197 (2) ◽  
pp. 314-325 ◽  
Author(s):  
Kristy L. Hentchel ◽  
Jorge C. Escalante-Semerena
Keyword(s):  
Salmonella Enterica ◽  
Physiological Role ◽  
Methionine Sulfoximine ◽  
Amino Acid Transporters ◽  
Content Type ◽  
Methionine Sulfone ◽  
Serovar Typhimurium ◽  
Acylation Reactions

Protein and small-molecule acylation reactions are widespread in nature. Many of the enzymes catalyzing acylation reactions belong to theGcn5-relatedN-acetyltransferase (GNAT; PF00583) family, named after the yeast Gcn5 protein. The genome ofSalmonella entericaserovar Typhimurium LT2 encodes 26 GNATs, 11 of which have no known physiological role. Here, we providein vivoandin vitroevidence for the role of the MddA (methioninederivativedetoxifier; formerly YncA) GNAT in the detoxification of oxidized forms of methionine, including methionine sulfoximine (MSX) and methionine sulfone (MSO). MSX and MSO inhibited the growth of anS. entericaΔmddAstrain unless glutamine or methionine was present in the medium. We used anin vitrospectrophotometric assay and mass spectrometry to show that MddA acetylated MSX and MSO. AnmddA+strain displayed biphasic growth kinetics in the presence of MSX and glutamine. Deletion of two amino acid transporters (GlnHPQ and MetNIQ) in a ΔmddAstrain restored growth in the presence of MSX. Notably, MSO was transported by GlnHPQ but not by MetNIQ. In summary, MddA is the mechanism used byS. entericato respond to oxidized forms of methionine, which MddA detoxifies by acetyl coenzyme A-dependent acetylation.

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