RESUMO
Endocannabinoids are host-derived lipid hormones that fundamentally impact gastrointestinal (GI) biology. The use of cannabis and other exocannabinoids as anecdotal treatments for various GI disorders inspired the search for mechanisms by which these compounds mediate their effects, which led to the discovery of the mammalian endocannabinoid system. Dysregulated endocannabinoid signaling was linked to inflammation and the gut microbiota. However, the effects of endocannabinoids on host susceptibility to infection has not been explored. Here, we show that mice with elevated levels of the endocannabinoid 2-arachidonoyl glycerol (2-AG) are protected from enteric infection by Enterobacteriaceae pathogens. 2-AG directly modulates pathogen function by inhibiting virulence programs essential for successful infection. Furthermore, 2-AG antagonizes the bacterial receptor QseC, a histidine kinase encoded within the core Enterobacteriaceae genome that promotes the activation of pathogen-associated type three secretion systems. Taken together, our findings establish that endocannabinoids are directly sensed by bacteria and can modulate bacterial function.
Assuntos
Endocanabinoides/metabolismo , Enterobacteriaceae/patogenicidade , Animais , Ácidos Araquidônicos/química , Ácidos Araquidônicos/metabolismo , Aderência Bacteriana , Proteínas de Bactérias/metabolismo , Sistemas de Secreção Bacterianos/metabolismo , Citrobacter rodentium/patogenicidade , Colo/microbiologia , Colo/patologia , Endocanabinoides/química , Infecções por Enterobacteriaceae/microbiologia , Feminino , Microbioma Gastrointestinal , Glicerídeos/química , Glicerídeos/metabolismo , Células HeLa , Interações Hospedeiro-Patógeno , Humanos , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Monoacilglicerol Lipases/metabolismo , Salmonella/patogenicidade , VirulênciaRESUMO
Antibacterial autophagy (xenophagy) is an important host defense, but how it is initiated is unclear. Here, we performed a bacterial transposon screen and identified a T3SS effector SopF that potently blocked Salmonella autophagy. SopF was a general xenophagy inhibitor without affecting canonical autophagy. S. Typhimurium ΔsopF resembled S. flexneri ΔvirAΔicsB with the majority of intracellular bacteria targeted by autophagy, permitting a CRISPR screen that identified host V-ATPase as an essential factor. Upon bacteria-caused vacuolar damage, the V-ATPase recruited ATG16L1 onto bacteria-containing vacuole, which was blocked by SopF. Mammalian ATG16L1 bears a WD40 domain required for interacting with the V-ATPase. Inhibiting autophagy by SopF promoted S. Typhimurium proliferation in vivo. SopF targeted Gln124 of ATP6V0C in the V-ATPase for ADP-ribosylation. Mutation of Gln124 also blocked xenophagy, but not canonical autophagy. Thus, the discovery of SopF reveals the V-ATPase-ATG16L1 axis that critically mediates autophagic recognition of intracellular pathogen.
Assuntos
Proteínas Relacionadas à Autofagia/metabolismo , Proteínas de Bactérias/genética , Macroautofagia , Salmonella/metabolismo , ATPases Vacuolares Próton-Translocadoras/metabolismo , Fatores de Virulência/genética , ADP-Ribosilação , Proteínas Relacionadas à Autofagia/deficiência , Proteínas Relacionadas à Autofagia/genética , Proteínas de Bactérias/metabolismo , Sistemas CRISPR-Cas/genética , Edição de Genes , Células HeLa , Humanos , Proteínas Associadas aos Microtúbulos/metabolismo , Ligação Proteica , Salmonella/patogenicidade , Sistemas de Secreção Tipo III/metabolismo , ATPases Vacuolares Próton-Translocadoras/genética , Fatores de Virulência/metabolismoRESUMO
Understanding the dynamic evolution of Salmonella is vital for effective bacterial infection management. This study explores the role of the flexible genome, organised in regions of genomic plasticity (RGP), in shaping the pathogenicity of Salmonella lineages. Through comprehensive genomic analysis of 12,244 Salmonella spp. genomes covering 2 species, 6 subspecies, and 46 serovars, we uncover distinct integration patterns of pathogenicity-related gene clusters into RGP, challenging traditional views of gene distribution. These RGP exhibit distinct preferences for specific genomic spots, and the presence or absence of such spots across Salmonella lineages profoundly shapes strain pathogenicity. RGP preferences are guided by conserved flanking genes surrounding integration spots, implicating their involvement in regulatory networks and functional synergies with integrated gene clusters. Additionally, we emphasise the multifaceted contributions of plasmids and prophages to the pathogenicity of diverse Salmonella lineages. Overall, this study provides a comprehensive blueprint of the pathogenicity potential of Salmonella. This unique insight identifies genomic spots in nonpathogenic lineages that hold the potential for harbouring pathogenicity genes, providing a foundation for predicting future adaptations and developing targeted strategies against emerging human pathogenic strains.
Assuntos
Genoma Bacteriano , Salmonella , Salmonella/genética , Salmonella/patogenicidade , Genoma Bacteriano/genética , Virulência/genética , Humanos , Genômica/métodos , Família Multigênica , Filogenia , Plasmídeos/genética , Infecções por Salmonella/microbiologia , Prófagos/genética , Evolução MolecularRESUMO
In Escherichia coli and Salmonella, many genes silenced by the nucleoid structuring protein H-NS are activated upon inhibiting Rho-dependent transcription termination. This response is poorly understood and difficult to reconcile with the view that H-NS acts mainly by blocking transcription initiation. Here we have analyzed the basis for the up-regulation of H-NS-silenced Salmonella pathogenicity island 1 (SPI-1) in cells depleted of Rho-cofactor NusG. Evidence from genetic experiments, semiquantitative 5' rapid amplification of complementary DNA ends sequencing (5' RACE-Seq), and chromatin immunoprecipitation sequencing (ChIP-Seq) shows that transcription originating from spurious antisense promoters, when not stopped by Rho, elongates into a H-NS-bound regulatory region of SPI-1, displacing H-NS and rendering the DNA accessible to the master regulator HilD. In turn, HilD's ability to activate its own transcription triggers a positive feedback loop that results in transcriptional activation of the entire SPI-1. Significantly, single-cell analyses revealed that this mechanism is largely responsible for the coexistence of two subpopulations of cells that either express or do not express SPI-1 genes. We propose that cell-to-cell differences produced by stochastic spurious transcription, combined with feedback loops that perpetuate the activated state, can generate bimodal gene expression patterns in bacterial populations.
Assuntos
Proteínas de Bactérias , Proteínas de Ligação a DNA , Regulação Bacteriana da Expressão Gênica , Regiões Promotoras Genéticas , Salmonella , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Proteínas de Ligação a DNA/metabolismo , Inativação Gênica , Salmonella/genética , Salmonella/patogenicidade , Análise de Célula Única , Transcrição Gênica , Virulência/genéticaRESUMO
Type III and type VI secretion systems (T3/T6SS) are encoded in horizontally acquired genomic islands (GIs) that play crucial roles in evolution and virulence in bacterial pathogens. T3/T6SS expression is subjected to tight control by the host xenogeneic silencer H-NS, but how this mechanism is counteracted remains to be illuminated. Here, we report that xenogeneic nucleoid-associated protein EnrR encoded in a GI is essential for virulence in pathogenic bacteria Edwardsiella and Salmonella. We showed that EnrR plays critical roles in T3/T6SS expression in these bacteria. Various biochemical and genetic analyses demonstrated that EnrR binds and derepresses the promoter of esrB, the critical regulator of T3/T6SS, to promote their expression by competing with H-NS. Additionally, EnrR targets AT-rich regions, globally modulates the expression of â¼363 genes and is involved in various cellular processes. Crystal structures of EnrR in complex with a specific AT-rich palindromic DNA revealed a new DNA-binding mode that involves conserved HTH-mediated interactions with the major groove and contacts of its N-terminal extension to the minor groove in the symmetry-related duplex. Collectively, these data demonstrate that EnrR is a virulence activator that can antagonize H-NS, highlighting a unique mechanism by which bacterial xenogeneic regulators recognize and regulate foreign DNA.
Assuntos
Proteínas de Bactérias/metabolismo , Proteínas de Ligação a DNA/metabolismo , Edwardsiella/patogenicidade , Ilhas Genômicas , Salmonella/patogenicidade , Sistemas de Secreção Bacterianos , Edwardsiella/genética , Regulação Bacteriana da Expressão Gênica , Inativação Gênica , Salmonella/genética , VirulênciaRESUMO
Rats are particularly important from an epidemiological point of view, because they are regarded as reservoirs for diverse zoonotic pathogens including enteric bacteria. This study is the first to report the emergence of Salmonella serovar Ohio in brown rats (Rattus norvegicus) and food-producing animals in Hungary. We first reveal the genomic diversity of the strains and their phylogenomic relationships in the context of the international collection of S. Ohio genomes. This pathogen was detected in 4.3% (4/92) of rats, captured from multiple sites in Hungary. A whole-genome-based genotype comparison of S. Ohio, Infantis, Enteritidis, and Typhimurium strains showed that 76.4% (117/153) of the virulence and antimicrobial resistance genes were conserved among these serovars, and none of the genes were specific to S. Ohio. All S. Ohio strains lacked virulence and resistance plasmids. The cgMLST phylogenomic comparison highlighted a close genetic relationship between rat and poultry strains of S. Ohio from Hungary. These strains clustered together with the international S. Ohio genomes from aquatic environments. Overall, this study contributes to our understanding of the epidemiology of Salmonella spp. in brown rats and highlights the importance of monitoring to minimize the public health risk of rodent populations. However, further research is needed to understand the route of infection and evolution of this serovar.
Assuntos
Genoma Bacteriano , Filogenia , Aves Domésticas , Salmonelose Animal , Salmonella , Animais , Ratos , Hungria , Salmonella/genética , Salmonella/classificação , Salmonella/patogenicidade , Salmonelose Animal/microbiologia , Salmonelose Animal/epidemiologia , Suínos , Aves Domésticas/microbiologia , Virulência/genética , Sequenciamento Completo do GenomaRESUMO
Salmonella diarizonae (IIIb) is frequently isolated from reptiles and less frequently from birds and mammals. However, its isolation from invasive human infections has not been widely reported. Migratory mallard ducks are excellent bioindicators of pathogen presence and pathogen antibiotic resistance (AMR). We present the first isolation from a mallard duck in central Europe of the antibiotic-resistant Salmonella enterica subsp. diarizonae with the unique antigenic pattern 58:r:z53 and report its whole-genome sequencing, serosequencing, and genotyping, which enabled the prediction of its pathogenicity and comparison with phenotypic AMR. The isolated strain was highly similar to S. diarizonae isolated from humans and food. Twenty-four AMR genes were detected, including those encoding aminoglycoside, fluoroquinolone, macrolide, carbapenem, tetracycline, cephalosporin, nitroimidazole, peptide antibiotic, and disinfecting agent/antiseptic resistance. Six Salmonella pathogenicity islands were found (SPI-1, SPI-2, SPI-3, SPI-5, SPI-9, and SPI-13). An iron transport system was detected in SPI-1 centisome C63PI. Plasmid profile analyses showed three to be present. Sequence mutations in the invA and invF genes were noted, which truncated and elongated the proteins, respectively. The strain also harbored genes encoding type-III secretion-system effector proteins and many virulence factors found in S. diarizonae associated with human infections. This study aims to elucidate the AMR and virulence genes in S. enterica subsp. diarizonae that may most seriously threaten human health.
Assuntos
Patos , Animais , Patos/microbiologia , Humanos , Salmonella/genética , Salmonella/patogenicidade , Salmonella/isolamento & purificação , Salmonella/efeitos dos fármacos , Sequenciamento Completo do Genoma , Ilhas Genômicas/genética , Salmonelose Animal/microbiologia , Antibacterianos/farmacologia , Salmonella enterica/genética , Salmonella enterica/patogenicidade , Salmonella enterica/isolamento & purificação , Salmonella enterica/efeitos dos fármacos , Farmacorresistência Bacteriana Múltipla/genética , Genoma Bacteriano , Filogenia , Farmacorresistência Bacteriana/genética , Plasmídeos/genéticaRESUMO
In recent years, as the paradigm of communication between cells has been clarified, the ability of bacteria to change their gene expression patterns in response to various extracellular signals has attracted great interest. In particular, intracellular and intercellular communication between bacterial populations, called quorum sensing (QS), is essential for coordinating physiological and genetic activities. QS studies are critical, particularly in elucidating the regulatory mechanisms of infectious processes in food-borne pathogens. Elucidating the QS mechanisms in Salmonella is effective in silencing the virulence factors in the fight against this bacterium. The aims of this study were; to create luxS gene mutants that play a vital role in the QS activity of Salmonella and to determine the effect of this mutation on the expression of virulence genes in the bacteria and to determine the impact of synthetic N-hexanoyl-homoserine lactone (C6HSL) on biofilm formation and AI-2 signaling pathway of Salmonella wild strain and luxS gene mutants. luxS gene mutants were constructed by recombining the gene region with the chloramphenicol gene cassette based on homologous region recombination. In the luxS mutants obtained in this way, the expression of eight different virulence genes (hilA, invA, inv, glgC, fimF, fliF, lpfA, gyrA), which have essential roles in Salmonella pathogenicity, was determined by quantitative real-time reverse transcriptase polymerase chain reaction (rRT-qPCR) method and compared with natural strains. As a result of these studies, it was determined that the expression of each gene examined was significantly reduced in luxS mutant strains. The relative AI-2 activities of Salmonella strains were analyzed depending on time. It was determined that the highest activity occurred at the fourth hour and the AI-2 activities of luxS mutants were reduced compared to the wild strain. Finally, it was determined that C6HSL increased the biofilm activity of Salmonella Typhimurium DMC4, SL1344 wild strains, and mutants, mainly at the 72nd hour. In conclusion, our results proved that C6HSL stimulated QS communication in all strains and increased biofilm of Salmonella formation and autoinducer activity. This situation determines that Salmonella responds to external signals by using QS systems. In addition, this research contributed to provide additional information on interspecies communication mechanisms to develop strategies to prevent biofilm formation of this pathogen.
Assuntos
Proteínas de Bactérias , Biofilmes , Liases de Carbono-Enxofre , Regulação Bacteriana da Expressão Gênica , Homosserina , Percepção de Quorum , Biofilmes/crescimento & desenvolvimento , Liases de Carbono-Enxofre/genética , Virulência , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Homosserina/análogos & derivados , Mutação , Fatores de Virulência/genética , 4-Butirolactona/análogos & derivados , 4-Butirolactona/metabolismo , Animais , Salmonella/patogenicidade , Salmonella/genéticaRESUMO
Tc toxins were originally identified in entomopathogenic bacteria, which are important as biological pest control agents. Tc toxins are heteromeric exotoxins composed of three subunit types, TcA, TcB, and TcC. The C-terminal portion of the TcC protein encodes the actual toxic domain, which is translocated into host cells by an injectosome nanomachine comprising the other subunits. Currently the pathogenic roles and distribution of Tc toxins among different bacterial genera remain unclear. Here we have performed a comprehensive genome-wide analysis, and established a database that includes 1,608 identified Tc loci containing 2,528 TcC proteins in 1,421 Gram-negative and positive bacterial genomes. Our findings indicate that TcCs conform to the architecture of typical polymorphic toxins, with C-terminal hypervariable regions (HVR) encoding more than 100 different classes of putative toxic domains, most of which have not been previously recognized. Based on further analysis of Tc loci in the genomes of all Salmonella and Yersinia strains in EnteroBase, a "two-level" evolutionary dynamics scenario is proposed for TcC homologues. This scenario implies that the conserved TcC RHS core domain plays a critical role in the taxonomical specific distribution of TcC HVRs. This study provides an extensive resource for the future development of Tc toxins as valuable agrochemical tools. It furthermore implies that Tc proteins, which are encoded by a wide range of pathogens, represent an important versatile toxin superfamily with diverse pathogenic mechanisms.
Assuntos
Proteínas de Bactérias/genética , Toxinas Bacterianas/genética , Evolução Biológica , Genoma Bacteriano , Salmonella/genética , Yersinia/genética , Proteínas de Bactérias/metabolismo , Toxinas Bacterianas/classificação , Toxinas Bacterianas/metabolismo , Células HEK293 , Células HeLa , Humanos , Salmonella/crescimento & desenvolvimento , Salmonella/patogenicidade , Yersinia/crescimento & desenvolvimento , Yersinia/patogenicidadeRESUMO
Large volumes of liquid and other materials from the extracellular environment are internalised by eukaryotic cells via an endocytic process called macropinocytosis. It is now recognised that this fundamental and evolutionarily conserved pathway is hijacked by numerous intracellular pathogens as an entry portal to the host cell interior. Yet, an increasing number of additional cellular functions of macropinosomes in pathologic processes have been reported beyond this role for fluid internalisation. It emerges that the identity of macropinosomes can vary hugely and change rapidly during their lifetime. A deeper understanding of this important multi-faceted compartment is based on novel methods for their investigation. These methods are either imaging-based for the tracking of macropinosome dynamics, or they provide the means to extract macropinosomes at high purity for comprehensive proteomic analyses. Here, we portray these new approaches for the investigation of macropinosomes. We document how these method developments have provided insights for a new understanding of the intracellular lifestyle of the bacterial pathogens Shigella and Salmonella. We suggest that a systematic complete characterisation of macropinosome subversion with these approaches during other infection processes and pathologies will be highly beneficial for our understanding of the underlying cellular and molecular processes.
Assuntos
Disenteria Bacilar/microbiologia , Endossomos/microbiologia , Interações Hospedeiro-Patógeno , Infecções por Salmonella/microbiologia , Salmonella/patogenicidade , Shigella/patogenicidade , HumanosRESUMO
Many bacterial pathogens hijack macrophages to egress from the port of entry to the lymphatic drainage and/or bloodstream, causing dissemination of life-threatening infections. However, the underlying mechanisms are not well understood. Here, we report that Salmonella infection generates directional electric fields (EFs) in the follicle-associated epithelium of mouse cecum. In vitro application of an EF, mimicking the infection-generated electric field (IGEF), induces directional migration of primary mouse macrophages to the anode, which is reversed to the cathode upon Salmonella infection. This infection-dependent directional switch is independent of the Salmonella pathogenicity island 1 (SPI-1) type III secretion system. The switch is accompanied by a reduction of sialic acids on glycosylated surface components during phagocytosis of bacteria, which is absent in macrophages challenged by microspheres. Moreover, enzymatic cleavage of terminally exposed sialic acids reduces macrophage surface negativity and severely impairs directional migration of macrophages in response to an EF. Based on these findings, we propose that macrophages are attracted to the site of infection by a combination of chemotaxis and galvanotaxis; after phagocytosis of bacteria, surface electrical properties of the macrophage change, and galvanotaxis directs the cells away from the site of infection.
Assuntos
Trato Gastrointestinal/imunologia , Macrófagos/fisiologia , Resposta Táctica/fisiologia , Animais , Proteínas de Bactérias , Movimento Celular/fisiologia , Condutividade Elétrica , Eletricidade , Epitélio/imunologia , Epitélio/metabolismo , Feminino , Macrófagos/microbiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Fagocitose , Salmonella/patogenicidade , Infecções por Salmonella/metabolismo , Infecções por Salmonella/fisiopatologiaRESUMO
Salmonella enterica subsp. enterica is one of the leading causes of human foodborne infections and several outbreaks are now associated with the consumption of fresh fruit and vegetables. This study aims at evaluating whether Salmonella virulence can be linked to an enhanced ability to survive successive digestive environments. Thirteen S. enterica strains were selected according to high and low virulence phenotypes. Lettuce inoculated separately with each S. enterica strain was used as food matrix in the TNO gastrointestinal model (TIM-1) of the human upper gastrointestinal tract. During the passage in the stomach, counts determined using PMA-qPCR were 2-5 logs higher than the cultivable counts for all strains indicating the presence of viable but non-cultivable cells. Bacterial growth was observed in the duodenum compartment after 180 min for all but one strain and growth continued into the ileal compartment. After passage through the simulated gastrointestinal tract, both virulent and avirulent S. enterica strains survived but high virulence strains had a significantly (p = 0.004) better average survival rate (1003 %-3753 %) than low virulence strains (from 25 % to 3730%). The survival rates of S. enterica strains could be linked to the presence of genes associated with acid and bile resistance and their predicted products. The presence of single nucleotide polymorphisms may also impact the function of virulence associated genes and play a role in the resulting phenotype. These data provide an understanding of the relationship between measured virulence potential and survival of S. enterica during dynamic simulated gastrointestinal transit.
Assuntos
Trato Gastrointestinal/microbiologia , Salmonella/patogenicidade , Virulência , Humanos , Modelos BiológicosRESUMO
The adaptive in vivo mechanisms underlying the switch in Salmonella enterica lifestyles from the infectious form to a dormant form remain unknown. We employed Caenorhabditis elegans as a heterologous host to understand the temporal dynamics of Salmonella pathogenesis and to identify its lifestyle form in vivo. We discovered that Salmonella exists as sessile aggregates, or in vivo biofilms, in the persistently infected C. elegans gut. In the absence of in vivo biofilms, Salmonella killed the host more rapidly by actively inhibiting innate immune pathways. Regulatory cross-talk between two major Salmonella pathogenicity islands, SPI-1 and SPI-2, was responsible for biofilm-induced changes in host physiology during persistent infection. Thus, biofilm formation is a survival strategy in long-term infections, as prolonging host survival is beneficial for the parasitic lifestyle.
Assuntos
Biofilmes , Caenorhabditis elegans/microbiologia , Imunidade Inata/fisiologia , Salmonella/fisiologia , Animais , Biomarcadores/metabolismo , Caenorhabditis elegans/crescimento & desenvolvimento , Intestinos/parasitologia , Larva/microbiologia , Salmonella/metabolismo , Salmonella/patogenicidade , VirulênciaRESUMO
Inflammatory responses have been shown to induce hyperglycemia, yet the underlying mechanism is still largely unclear. GLP-1 is an important intestinal hormone for regulating glucose homeostasis; however, few studies have investigated the influence of digestive tract Salmonella infection on enteroendocrine L cell secretions. In this study, we established a model of Salmonella-infected piglets by oral gavage in order to analyze the effects of Salmonella infection on enteroendocrine L cell function. Furthermore, in vitro lipopolysaccharide (LPS) was administered to STC-1 cells to clarify its direct effect on GLP-1 secretion. The results showed that significantly increased blood glucose in the group of Salmonella-infected piglets was observed, and Salmonella infection decreased blood GLP-1 content. Then, ileal epithelium damage was observed by histological detection, and this was further verified by TUNEL staining. We identified activation of TLR signaling demonstrating up-regulated expressions of TLR4 and nuclear factor-kappa B (NF-ΚB). Furthermore, it was shown that Salmonella induced pyroptosis of enteroendocrine L cells and enhanced the secretion of IL-1ß through augmenting gene and protein expressions of NOD-like receptor protein 3 (NLRP3), apoptosis-associated speck-like protein containing a carboxyl-terminal CARD (ASC), Caspase 1, and gasdermin D (GSDMD). Meanwhile, in vitro LPS treatment induced the pyroptosis of STC-1 cells and reduced the secretion of GLP-1. Altogether, the results demonstrated that Salmonella infection can reduce secretion of GLP-1 by inducing pyroptosis of intestinal L cells, which may eventually result in hyperglycemia. The results provided evidence for the cause of hyperglycemia induced by inflammation and shed new light on glucose homeostasis regulation.
Assuntos
Peptídeo 1 Semelhante ao Glucagon/metabolismo , Hiperglicemia/etiologia , Salmonelose Animal/metabolismo , Animais , Caspase 1/metabolismo , China , Células Enteroendócrinas/citologia , Células Enteroendócrinas/metabolismo , Hiperglicemia/patologia , Inflamassomos/metabolismo , Inflamação , Células L/metabolismo , Camundongos , NF-kappa B/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Piroptose/efeitos dos fármacos , Piroptose/fisiologia , Salmonella/patogenicidade , Transdução de Sinais , Suínos/microbiologiaRESUMO
Many bacterial pathogens and symbionts use type III secretion machines to interact with their hosts by injecting bacterial effector proteins into host target cells. A central component of this complex machine is the cytoplasmic sorting platform, which orchestrates the engagement and preparation of type III secreted proteins for their delivery to the needle complex, the substructure of the type III secretion system that mediates their passage through the bacterial envelope. The sorting platform is thought to be a dynamic structure whose components alternate between assembled and disassembled states. However, how this dynamic behavior is controlled is not understood. In S. Typhimurium a core component of the sorting platform is SpaO, which is synthesized in two tandemly translated products, a full length (SpaOL) and a short form (SpaOS) composed of the C-terminal 101 amino acids. Here we show that in the absence of SpaOS the assembly of the needle substructure of the needle complex, which requires a functional sorting platform, can still occur although with reduced efficiency. Consistent with this observation, in the absence of SpaOS secretion of effectors proteins, which requires a fully assembled injectisome, is only slightly compromised. In the absence of SpaOS we detect a significant number of fully assembled needle complexes that are not associated with fully assembled sorting platforms. We also find that although binding of SpaOL to SpaOS can be detected in the absence of other components of the sorting platform, this interaction is not detected in the context of a fully assembled sorting platform suggesting that SpaOS may not be a core structural component of the sorting platform. Consistent with this observation we find that SpaOS and OrgB, a component of the sorting platform, share the same binding surface on SpaOL. We conclude that SpaOS regulates the assembly of the sorting platform during type III secretion.
Assuntos
Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Sistemas de Secreção Tipo III/metabolismo , Proteínas de Bactérias/fisiologia , Proteínas de Membrana/fisiologia , Isoformas de Proteínas , Transporte Proteico/fisiologia , Salmonella/metabolismo , Salmonella/patogenicidade , Salmonella typhimurium/metabolismo , Sistemas de Secreção Tipo III/fisiologiaRESUMO
BACKGROUND: This study was undertaken to identify and functionally characterize virulence genes from Salmonella isolates in street food and stool cultures. From February 2017 to May 2018, clinical and food Salmonella strains were isolated in three regions in Burkina Faso. Salmonella was serotyped according to the White-Kauffmann-Le Minor method, and polymerase chain reaction (PCR) was used to detec invA, spvR, spvC, fimA and stn virulence genes commonly associated with salmonellosis in Sub-Saharan Africa. RESULTS: A total of 106 Salmonella isolates (77 human stools; 14 sandwiches) was analyzed using a serological identification with an O-group test reagent. The presence of Salmonella was confirmed in 86% (91/106) of the samples were reactive (OMA-positive/OMB-positive). Salmonella serogroup O:4,5 was the most common serogroup detected (40%; 36/91). Salmonella Enteritidis and Typhimurium represented 5.5% (5/91) and 3.3% (3/91), respectively and were identified only from clinical isolates. Furthermore, 14 serotypes of Salmonella (12/91 human strains and 2/15 sandwich strains) were evocative of Kentucky/Bargny serotype. For the genetic profile, 66% (70/106) of the Salmonella had invA and stn genes; 77.4% (82/106) had the fimA gene. The spvR gene was found in 36.8% (39/106) of the isolates while 48.1% (51/106) had the spvC gene. Among the identified Salmonella Enteritidis and Salmonella Typhimurium isolated from stools, the virulence genes detected were invA (3/5) versus (2/3), fimA (4/5) versus (3/3), stn (3/5) versus (2/3), spvR (4/5) versus (2/3) and spvC (3/5) versus (2/3), respectively. CONCLUSION: This study reports the prevalence of Salmonella serotypes and virulence genes in clinical isolates and in street foods. It shows that food could be a significant source of Salmonella transmission to humans. Our results could help decision-making by the Burkina Faso health authority in the fight against street food-related diseases, in particular by training restaurateurs in food hygiene.
Assuntos
Fast Foods/microbiologia , Fezes/microbiologia , Salmonella/isolamento & purificação , Fatores de Virulência/genética , Burkina Faso/epidemiologia , Diarreia/epidemiologia , Diarreia/microbiologia , Genes Bacterianos , Humanos , Prevalência , Salmonella/classificação , Salmonella/genética , Salmonella/patogenicidade , Sorogrupo , Sorotipagem , Virulência/genéticaRESUMO
An auxotrophic mutant of nontyphoidal Salmonella (NTS) strain (Salmonella Oslo) was phenotypically characterized in this study. The characterization was based on phenotype, morphology, motility, biofilm forming ability, growth kinetics, etc. The phenotypic results from the above experiments determined that the mutant showed variation in phenotypic characters from that of wild-type strain. Subsequently, mutant and wild-type NTS were subjected to epithelial cell invasion and intracellular replication assays. The real-time PCR analysis was also performed to analyse expression of tumor inhibiting cytokine genes and virulence genes post-bacterial infection in cell lines. The mutant showed highest invasion potential than wild-type NTS whereas the replication of mutant was slower in both the cell lines. Similar to the wild-type strain, the mutant also retained the cytotoxic potential when analysed in vitro. Furthermore, the expression of proinflammatory cytokine genes such as TNF-α and IL-1ß was upsurged with the downregulation of anti-inflammatory cytokine genes like TGF-ß, IL-6 and IL-10 post-infection of the mutant strain in cell lines. In addition, virulence genes of Salmonella pathogenicity island one and two of mutant were downregulated in vitro except invA in HeLa cell line. Therefore, the auxotrophic mutant showed positive attributes of a potential antitumor agent in terms of expressing tumor inhibiting cytokine genes when assessed in vitro. Though the study did not check the tumor inhibitory effect of NTS strain directly, findings of the study emphasizes on the development of a novel strain of NTS with less virulence and more immunogenic traits to inhibit tumor cells.
Assuntos
Citocinas/genética , Salmonella/genética , Animais , Células HEK293 , Células HeLa , Humanos , Fenótipo , Salmonella/patogenicidade , Virulência/genéticaRESUMO
According to the European Food Safety Authority (EFSA) and European Centre for Disease Prevention and Control (ECDC) annual report, human salmonellosis is mostly related to consumption of contaminated poultry products. Since 2003 in Europe, the Salmonella serovars considered relevant for human health and subject to control in breeding hens of Gallus gallus are: S. Enteritidis, S. Typhimurium (including the monophasic variant), S. Infantis, S. Hadar and S. Virchow. Herein, we investigated the Italian epidemiological situation from 2016 to 2018, comparing Salmonella serovar distributions in humans and poultry, in order to identify the target Salmonella serovars that, if controlled, would potentially have the largest public health impact in Italy. The results showed that control of S. Virchow and S. Hadar does no longer seem to be a priority in Italy and that S. Napoli and S. Derby, which are not included in the group of EU target serovars, are among the most frequent serovars isolated from humans in Italy. While S. Derby has its main reservoir in pigs, S. Napoli does not have a specific reservoir. However, because this serovar is frequently isolated from breeding poultry flocks and is characterised by causing severe human illness, it is a potential target Salmonella serovar in breeding hens of Gallus gallus in Italy.
Assuntos
Aves Domésticas/microbiologia , Infecções por Salmonella/microbiologia , Salmonella/genética , Salmonella/isolamento & purificação , Animais , Galinhas/microbiologia , Microbiologia de Alimentos , Humanos , Itália/epidemiologia , Prevalência , Salmonella/classificação , Salmonella/patogenicidade , Infecções por Salmonella/epidemiologia , SorogrupoRESUMO
As an environment-dependent pleiotropic gene regulator in Gram-negative bacteria, the H-NS protein is crucial for adaptation and toxicity control of human pathogens such as Salmonella, Vibrio cholerae or enterohaemorrhagic Escherichia coli. Changes in temperature affect the capacity of H-NS to form multimers that condense DNA and restrict gene expression. However, the molecular mechanism through which H-NS senses temperature and other physiochemical parameters remains unclear and controversial. Combining structural, biophysical and computational analyses, we show that human body temperature promotes unfolding of the central dimerization domain, breaking up H-NS multimers. This unfolding event enables an autoinhibitory compact H-NS conformation that blocks DNA binding. Our integrative approach provides the molecular basis for H-NS-mediated environment-sensing and may open new avenues for the control of pathogenic multi-drug resistant bacteria.
Assuntos
Proteínas de Bactérias/química , DNA Bacteriano/genética , Proteínas de Ligação a DNA/química , Desdobramento de Proteína , Proteínas de Bactérias/genética , DNA Bacteriano/química , Proteínas de Ligação a DNA/genética , Escherichia coli Êntero-Hemorrágica/genética , Escherichia coli Êntero-Hemorrágica/patogenicidade , Interação Gene-Ambiente , Humanos , Domínios Proteicos , Multimerização Proteica/genética , Salmonella/genética , Salmonella/patogenicidade , Temperatura , Vibrio cholerae/genética , Vibrio cholerae/patogenicidadeRESUMO
Cantaloupes contaminated with pathogens have led to many high-profile outbreaks and illnesses. Since bacterial virulence genes (VGs) can act in tandem with antibiotic-resistance and mobile genetic elements, there is a need to evaluate these gene reservoirs in fresh produce, such as cantaloupes. The goal of this study was to assess the distribution of antibiotic-resistance, virulence, and mobile genetic elements genes (MGEGs) in cantaloupe farm environments. A total of 200 samples from cantaloupe melons (n = 99), farm workers' hands (n = 66), and production water (n = 35) were collected in México. Each sample was assayed for the presence of 14 antibiotic-resistance genes, 15 VGs, and 5 MGEGs by polymerase chain reaction. Our results indicated that tetracycline (tetA and tetB) (18% of cantaloupe, 45% of hand samples) and sulfonamide (sul1) (30% of cantaloupe, 71% of hand samples) resistance genes were frequently detected. The colistin resistance gene (mcr1) was detected in 10% of cantaloupe and 23% of farm workers' hands. Among VGs, Salmonella genes invA and spiA were the most abundant. There was a significantly higher likelihood of detecting antibiotic-resistance, virulence, and MGEGs on hands compared with water samples. These results demonstrate a diverse pool of antibiotic-resistance and VGs in cantaloupe production.