Your browser doesn't support javascript.
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 10.012
Filtrar
1.
Nat Commun ; 11(1): 107, 2020 01 09.
Artigo em Inglês | MEDLINE | ID: mdl-31919364

RESUMO

Bacteria commonly form dense biofilms encased in extracellular polymeric substances (EPS). Biofilms are often extremely tolerant to antimicrobials but their reliance on shared EPS may also be a weakness as social evolution theory predicts that inhibiting shared traits can select against resistance. Here we show that EPS of Salmonella biofilms is a cooperative trait whose benefit is shared among cells, and that EPS inhibition reduces both cell attachment and antimicrobial tolerance. We then compare an EPS inhibitor to conventional antimicrobials in an evolutionary experiment. While resistance against conventional antimicrobials rapidly evolves, we see no evolution of resistance to EPS inhibition. We further show that a resistant strain is outcompeted by a susceptible strain under EPS inhibitor treatment, explaining why resistance does not evolve. Our work suggests that targeting cooperative traits is a viable solution to the problem of antimicrobial resistance.


Assuntos
Antibacterianos/farmacologia , Biofilmes/crescimento & desenvolvimento , Matriz Extracelular de Substâncias Poliméricas/metabolismo , Imidazóis/farmacologia , Salmonella typhimurium/crescimento & desenvolvimento , Farmacorresistência Bacteriana/genética , Farmacorresistência Bacteriana/fisiologia , Matriz Extracelular de Substâncias Poliméricas/efeitos dos fármacos , Salmonella typhimurium/genética
2.
Emerg Microbes Infect ; 9(1): 1-4, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31859589

RESUMO

CRISPR-based typing was performed to subtype isolates of S. Typhimurium and its monophasic variant Salmonella 4,[5],12:i:- from humans and animals between 2009 and 2017 in China. CRISPR typing classified all isolates into two lineages and four sub-lineages. All isolates from Lineage II and Lineage IB-1 were Salmonella Typhimurium. All of Salmonella 4,[5],12:i: - isolates were distributed in Lineage IA and Lineage IB-2, which all belonged to ST34 by MLST typing. Only Lineage IB-2 contained ST34 isolates from both Salmonella Typhimurium and Salmonella 4,[5],12:i:-. Among the isolates of ST34, TST4 was identified as the most common CRISPR type representing 86.5% of Salmonella 4,[5],12:i:- and 14.5 % of Salmonella Typhimurium mainly from pigs and humans. This study demonstrated that TST4-ST34 isolates were predominant in Salmonella 4,[5],12:i:-, and pig was the main reservoir for Salmonella 4,[5],12:i:- in China, which might have the potential to transmit to humans by pig production.


Assuntos
Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Reservatórios de Doenças , Técnicas de Genotipagem/métodos , Carne/microbiologia , Salmonella typhimurium/classificação , Salmonella typhimurium/isolamento & purificação , Sus scrofa , Animais , China , Diarreia/microbiologia , Eletroforese em Gel de Campo Pulsado , Humanos , Tipagem de Sequências Multilocus , Filogenia , Infecções por Salmonella/microbiologia , Salmonella typhimurium/genética
3.
PLoS Negl Trop Dis ; 13(12): e0007490, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31790394

RESUMO

Schistosoma mansoni threatens hundreds of millions of people in >50 countries. Schistosomulae migrate through the lung and adult worms reside in blood vessels adjacent to the intestinal mucosa. Current candidate vaccines aren't designed to elicit a mucosal response. We have repurposed an attenuated Salmonella enterica Typhimurium strain (YS1646) to produce such a vaccine targeting Cathepsin B (CatB), a digestive enzyme important for parasite survival. Promoter-Type 3 secretory signal pairs were screened for protein expression in vitro and transfected into YS1646 to generate candidate vaccine strains. Two strains were selected for in vivo evaluation (nirB_SspH1 and SspH1_SspH1). Female C57BL/6 mice were immunized twice, 3 weeks apart, using six strategies: i) saline gavage (control), ii) the 'empty' YS1646 vector orally (PO) followed by intramuscular (IM) recombinant CatB (20µg IM rCatB), iii) two doses of IM rCatB, iv) two PO doses of YS1646-CatB, v) IM rCatB then PO YS1646-CatB and vi) PO YS1646-CatB then IM rCatB. Serum IgG responses to CatB were monitored by ELISA. Three weeks after the second dose, mice were challenged with 150 cercariae and sacrificed 7 weeks later to assess adult worm and egg burden (liver and intestine), granuloma size and egg morphology. CatB-specific IgG antibodies were low/absent in the control and PO only groups but rose substantially in other groups (5898-6766ng/mL). The highest response was in animals that received nirB_SspH1 YS1646 PO then IM rCatB. In this group, reductions in worm and intestine/liver egg burden (vs. control) were 93.1% and 79.5%/90.3% respectively (all P < .0001). Granuloma size was reduced in all vaccinated groups (range 32.9-52.8 x103µm2) and most significantly in the nirB_SspH1 + CatB IM group (34.7±3.4 x103µm2vs. 62.2±6.1 x103µm2: vs. control P < .01). Many eggs in the vaccinated animals had abnormal morphology. Targeting CatB using a multi-modality approach can provide almost complete protection against S. mansoni challenge.


Assuntos
Antígenos de Helmintos/imunologia , Catepsina B/imunologia , Portadores de Fármacos , Salmonella typhimurium/genética , Schistosoma mansoni/imunologia , Esquistossomose mansoni/prevenção & controle , Vacinas Sintéticas/imunologia , Animais , Anticorpos Anti-Helmínticos/sangue , Antígenos de Helmintos/genética , Catepsina B/genética , Modelos Animais de Doenças , Feminino , Imunoglobulina G/sangue , Injeções Intramusculares , Camundongos Endogâmicos C57BL , Schistosoma mansoni/genética , Esquistossomose mansoni/patologia , Resultado do Tratamento , Vacinas Atenuadas/administração & dosagem , Vacinas Atenuadas/imunologia , Vacinas Sintéticas/administração & dosagem
4.
Pol J Microbiol ; 68(4): 439-447, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31880888

RESUMO

Salmonella infection is most common in patients with infected aortic aneurysm, especially in Asia. When the aortic wall is heavily atherosclerotic, the intima is vulnerable to invasion by Salmonella, leading to the development of infected aortic aneurysm. By using THP-1 macrophage-derived foam cells to mimic atherosclerosis, we investigated the role of three Salmonella enterica serotypes - Typhimurium, Enteritidis, and Choleraesuis - in foam cell autophagy and inflammasome formation. Herein, we provide possible pathogenesis of Salmonella-associated infected aortic aneurysms. Three S. enterica serotypes with or without virulence plasmid were studied. Through Western blotting, we investigated cell autophagy induction and inflammasome formation in Salmonella-infected THP-1 macrophage-derived foam cells, detected CD36 expression after Salmonella infection through flow cytometry, and measured interleukin (IL)-1ß, IL-12, and interferon (IFN)-α levels through enzyme-linked immunosorbent assay. At 0.5 h after infection, plasmid-bearing S. Enteritidis OU7130 induced the highest foam cell autophagy - significantly higher than that induced by plasmid-less OU7067. However, plasmid-bearing S. Choleraesuis induced less foam cell autophagy than did its plasmid-less strain. In foam cells, plasmid-less Salmonella infection (particularly S. Choleraesuis OU7266 infection) led to higher CD36 expression than did plasmid-bearing strains infection. OU7130 and OU7266 infection induced the highest IL-1ß secretion. OU7067-infected foam cells secreted the highest IL-12p35 level. Plasmid-bearing S. Typhimurium OU5045 induced a higher IFN-α level than did other Salmonella serotypes. Salmonella serotypes are correlated with foam cell autophagy and IL-1ß secretion. Salmonella may affect the course of foam cells formation, or even aortic aneurysm, through autophagy.Salmonella infection is most common in patients with infected aortic aneurysm, especially in Asia. When the aortic wall is heavily atherosclerotic, the intima is vulnerable to invasion by Salmonella, leading to the development of infected aortic aneurysm. By using THP-1 macrophage-derived foam cells to mimic atherosclerosis, we investigated the role of three Salmonella enterica serotypes ­ Typhimurium, Enteritidis, and Choleraesuis ­ in foam cell autophagy and inflammasome formation. Herein, we provide possible pathogenesis of Salmonella-associated infected aortic aneurysms. Three S. enterica serotypes with or without virulence plasmid were studied. Through Western blotting, we investigated cell autophagy induction and inflammasome formation in Salmonella-infected THP-1 macrophage-derived foam cells, detected CD36 expression after Salmonella infection through flow cytometry, and measured interleukin (IL)-1ß, IL-12, and interferon (IFN)-α levels through enzyme-linked immunosorbent assay. At 0.5 h after infection, plasmid-bearing S. Enteritidis OU7130 induced the highest foam cell autophagy ­ significantly higher than that induced by plasmid-less OU7067. However, plasmid-bearing S. Choleraesuis induced less foam cell autophagy than did its plasmid-less strain. In foam cells, plasmid-less Salmonella infection (particularly S. Choleraesuis OU7266 infection) led to higher CD36 expression than did plasmid-bearing strains infection. OU7130 and OU7266 infection induced the highest IL-1ß secretion. OU7067-infected foam cells secreted the highest IL-12p35 level. Plasmid-bearing S. Typhimurium OU5045 induced a higher IFN-α level than did other Salmonella serotypes. Salmonella serotypes are correlated with foam cell autophagy and IL-1ß secretion. Salmonella may affect the course of foam cells formation, or even aortic aneurysm, through autophagy.


Assuntos
Aneurisma Aórtico/microbiologia , Infecções por Salmonella/microbiologia , Salmonella typhimurium/patogenicidade , Animais , Aneurisma Aórtico/genética , Aneurisma Aórtico/imunologia , Linhagem Celular , Humanos , Interferon-alfa/genética , Interferon-alfa/imunologia , Interleucina-12/genética , Interleucina-12/imunologia , Interleucina-1beta/genética , Interleucina-1beta/imunologia , Antígeno Ki-1/genética , Antígeno Ki-1/imunologia , Macrófagos/imunologia , Macrófagos/microbiologia , Monócitos/imunologia , Monócitos/microbiologia , Plasmídeos/genética , Plasmídeos/metabolismo , Infecções por Salmonella/genética , Infecções por Salmonella/imunologia , Salmonella typhimurium/genética , Salmonella typhimurium/fisiologia , Sorogrupo , Virulência
5.
Pol J Microbiol ; 68(4): 559-563, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31880899

RESUMO

We demonstrate here for the first time the use of an IncP-1ß plasmid, R751, as a gene capture vehicle for recombineering/conjugation strategies to clone large segments of bacterial genomes (20 - 100 + Kb). We designed R751 derivatives containing alternative markers for greater flexibility when using the R751 vehicle across different bacteria. These markers are removable if desired as part of the cloning procedure (with no extra steps needed). We demonstrated utility via cloning of 38 and 22 kb genomic segments from Salmonella enterica serovar Typhimurium and Escherichia coli, respectively. The plasmids expand the options available for use in recombineering/conjugation-based cloning applications.We demonstrate here for the first time the use of an IncP-1ß plasmid, R751, as a gene capture vehicle for recombineering/conjugation strategies to clone large segments of bacterial genomes (20 ­ 100 + Kb). We designed R751 derivatives containing alternative markers for greater flexibility when using the R751 vehicle across different bacteria. These markers are removable if desired as part of the cloning procedure (with no extra steps needed). We demonstrated utility via cloning of 38 and 22 kb genomic segments from Salmonella enterica serovar Typhimurium and Escherichia coli, respectively. The plasmids expand the options available for use in recombineering/conjugation-based cloning applications.


Assuntos
Clonagem Molecular , Conjugação Genética , Escherichia coli/genética , Plasmídeos/genética , Salmonella typhimurium/genética , DNA Bacteriano/genética , Recombinação Genética
6.
Ecotoxicol Environ Saf ; 186: 109822, 2019 Dec 30.
Artigo em Inglês | MEDLINE | ID: mdl-31634658

RESUMO

Nitroaromatic compounds (NACs) are an important type of environmental organic pollutants. However, it is lack of sufficient information relating to their potential adverse effects on human health and the environment due to the limited resources. Thus, using in silico technologies to assess their potential hazardous effects is urgent and promising. In this study, quantitative structure activity relationship (QSAR) and classification models were constructed using a set of NACs based on their mutagenicity against Salmonella typhimurium TA100 strain. For QSAR studies, DRAGON descriptors together with quantum chemistry descriptors were calculated for characterizing the detailed molecular information. Based on genetic algorithm (GA) and multiple linear regression (MLR) analyses, we screened descriptors and developed QSAR models. For classification studies, seven machine learning methods along with six molecular fingerprints were applied to develop qualitative classification models. The goodness of fitting, reliability, robustness and predictive performance of all developed models were measured by rigorous statistical validation criteria, then the best QSAR and classification models were chosen. Moreover, the QSAR models with quantum chemistry descriptors were compared to that without quantum chemistry descriptors and previously reported models. Notably, we also obtained some specific molecular properties or privileged substructures responsible for the high mutagenicity of NACs. Overall, the developed QSAR and classification models can be utilized as potential tools for rapidly predicting the mutagenicity of new or untested NACs for environmental hazard assessment and regulatory purposes, and may provide insights into the in vivo toxicity mechanisms of NACs and related compounds.


Assuntos
Poluentes Ambientais , Hidrocarbonetos Aromáticos , Mutagênicos , Nitrocompostos , Algoritmos , Simulação por Computador , Poluentes Ambientais/química , Poluentes Ambientais/toxicidade , Hidrocarbonetos Aromáticos/química , Hidrocarbonetos Aromáticos/toxicidade , Aprendizado de Máquina , Mutagênicos/química , Mutagênicos/toxicidade , Nitrocompostos/química , Nitrocompostos/toxicidade , Relação Quantitativa Estrutura-Atividade , Reprodutibilidade dos Testes , Salmonella typhimurium/efeitos dos fármacos , Salmonella typhimurium/genética
7.
PLoS Negl Trop Dis ; 13(10): e0007782, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31609964

RESUMO

BACKGROUND: Salmonella Typhimurium and Enteritidis are major causes of bloodstream infection in children in sub-Saharan Africa. This study assessed evidence for their zoonotic versus human reservoir. METHODS: Index patients were children with blood culture confirmed Salmonella infection recruited during a microbiological surveillance study in Nanoro, rural Burkina between May 2013 and August 2014. After consent, their households were visited. Stool from household members and livestock (pooled samples per species) as well as drinking water were cultured for Salmonella. Isolates with identical serotype obtained from index patient and any household sample were defined as "paired isolates" and assessed for genetic relatedness by multilocus variable number tandem-repeat analysis (MLVA) and whole-genome sequencing (WGS). RESULTS: Twenty-nine households were visited for 32/42 (76.2%) eligible index patients: two households comprised two index patients each, and in a third household the index patient had a recurrent infection. Among the 32 index patients, serotypes were Salmonella Typhimurium (n = 26), Salmonella Enteritidis (n = 5) and Salmonella Freetown (n = 1). All Typhimurium isolates were sequence type (ST)313. Median delay between blood culture sampling and household visits was 13 days (range 6-26). Salmonella was obtained from 16/186 (8.6%) livestock samples (13 serotypes) and 18/290 (6.2%) household members (9 serotypes). None of the water samples yielded Salmonella. Paired Salmonella Typhimurium isolates were obtained from three households representing four index patients. MLVA types were identical in two pairs and similar in the third (consisting of two index patients and one household member). WGS showed a strong genetic relatedness with 0 to 2 core genome SNPs difference between pairs on a household level. Livestock samples did not yield any Salmonella Typhimurium or Salmonella Enteritidis, and the latter was exclusively obtained from blood culture. Other serotypes shared by human and/or livestock carriers in the same household were Salmonella Derby, Drac, Tennessee and Muenster. CONCLUSIONS/SIGNIFICANCE: The current study provides further evidence of a human reservoir for invasive non-Typhoidal Salmonella (iNTS) in sub-Saharan Africa.


Assuntos
Reservatórios de Doenças/microbiologia , Características da Família , Infecções por Salmonella/microbiologia , Salmonella/classificação , Salmonella/isolamento & purificação , Adolescente , Animais , Burkina Faso/epidemiologia , Criança , Pré-Escolar , Monitoramento Ambiental , Fezes/microbiologia , Feminino , Humanos , Lactente , Gado , Masculino , Tipagem de Sequências Multilocus , Filogenia , Salmonella/genética , Infecções por Salmonella/epidemiologia , Salmonella enterica/genética , Salmonella enterica/isolamento & purificação , Salmonella enteritidis/genética , Salmonella enteritidis/isolamento & purificação , Salmonella typhimurium/genética , Salmonella typhimurium/isolamento & purificação , Sorogrupo , Microbiologia da Água , Sequenciamento Completo do Genoma
8.
Infect Immun ; 88(1)2019 12 17.
Artigo em Inglês | MEDLINE | ID: mdl-31611277

RESUMO

To survive and replicate during infection, pathogens utilize different carbon and energy sources depending on the nutritional landscape of their host microenvironment. Salmonella enterica serovar Typhimurium is an intracellular bacterial pathogen that occupies diverse cellular niches. While it is clear that Salmonella Typhimurium requires access to glucose during systemic infection, data on the need for lipid metabolism are mixed. We report that Salmonella Typhimurium strains lacking lipid metabolism genes were defective for systemic infection of mice. Bacterial lipid import, ß-oxidation, and glyoxylate shunt genes were required for tissue colonization upon oral or intraperitoneal inoculation. In cultured macrophages, lipid import and ß-oxidation genes were required for bacterial replication and/or survival only when the cell culture medium was supplemented with nonessential amino acids. Removal of glucose from tissue culture medium further enhanced these phenotypes and, in addition, conferred a requirement for glyoxylate shunt genes. We also observed that Salmonella Typhimurium needs lipid metabolism genes in proinflammatory but not anti-inflammatory macrophages. These results suggest that during systemic infection, the Salmonella Typhimurium that relies upon host lipids to replicate is within proinflammatory macrophages that have access to amino acids but not glucose. An improved understanding of the host microenvironments in which pathogens have specific metabolic requirements may facilitate the development of targeted approaches to treatment.


Assuntos
Metabolismo dos Lipídeos , Macrófagos/microbiologia , Redes e Vias Metabólicas/genética , Salmonella typhimurium/crescimento & desenvolvimento , Salmonella typhimurium/metabolismo , Aminoácidos/metabolismo , Animais , Glucose/metabolismo , Camundongos , Viabilidade Microbiana , Salmonelose Animal/microbiologia , Salmonelose Animal/patologia , Salmonella typhimurium/genética
9.
Ecotoxicol Environ Saf ; 185: 109733, 2019 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-31580980

RESUMO

Presence of missing data points in datasets is among main challenges in handling the toxicological data for nanomaterials. As the processing of missing data is an important part of data analysis, we have introduced a read-across approach that uses a combination of supervised and unsupervised machine learning techniques to fill the missing values. A series of classification models (supervised learning) was developed to predict class label, and self-organizing map approach (unsupervised learning) was used to estimate relative distances between nanoparticles and refine results obtained during supervised learning. In this study, genotoxicity of 49 silicon and metal oxide nanoparticles in Ames and Comet tests. Collected literature data did not demonstrate significant variations related to the change of size including selected bulk materials. Genotoxicity-related features of nanomaterials were represented by ionic characteristics. General tendencies found in the current study were convincingly linked to known theories of genotoxic action at nano-level. Mechanisms of primary and secondary genotoxic effects were discussed in the context of developed models.


Assuntos
Dano ao DNA , Nanopartículas Metálicas/toxicidade , Modelos Teóricos , Mutagênicos/toxicidade , Aprendizado de Máquina não Supervisionado , Linhagem Celular , Ensaio Cometa , Humanos , Nanopartículas Metálicas/classificação , Mutagênicos/classificação , Óxidos/classificação , Óxidos/toxicidade , Relação Quantitativa Estrutura-Atividade , Salmonella typhimurium/genética
10.
J Basic Microbiol ; 59(11): 1143-1153, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31577373

RESUMO

Salmonella enterica serovar Typhimurium (S. Tm) is a major intracellular pathogen that infects humans and animals, and its survival and growth in macrophages is essential for its pathogenicity. More than 50 putative regulatory proteins are encoded by the S. Tm genome, but the functions of these regulatory proteins in mediating S. Tm pathogenicity are largely unknown. In this study, we investigated the biological function of the STM0030 gene, which encodes a putative LysR-type transcriptional regulator. We found that STM0030 is upregulated 2.8-5.7-fold during S. Tm growth in macrophages. Further, mutating this gene decreased bacterial growth in macrophages and attenuated virulence in mice. RNA-sequencing to investigate the regulatory function of STM0030 in S. Tm revealed that 447 genes were differentially expressed between the mutant and the wild-type strains; 429 of these genes were downregulated, suggesting that STM0030 mainly acts as a transcriptional activator. Moreover, the expression of gluconate, maltose, and hexose-p transport genes, as well as allantoin utilization genes were downregulated in the STM0030 mutant; this might be associated with the observed decrease in intracellular replication and pathogenicity of the mutant. Our findings suggest that STM0030 is a new pathogenicity-associated regulatory protein that broadens our understanding of the virulence regulatory network of S. Tm.


Assuntos
Proteínas de Bactérias/metabolismo , Salmonella typhimurium/patogenicidade , Fatores de Transcrição/metabolismo , Fatores de Virulência/metabolismo , Animais , Proteínas de Bactérias/genética , Feminino , Regulação Bacteriana da Expressão Gênica , Macrófagos/microbiologia , Camundongos , Camundongos Endogâmicos BALB C , Mutação , Células RAW 264.7 , Salmonelose Animal/microbiologia , Salmonella typhimurium/genética , Salmonella typhimurium/crescimento & desenvolvimento , Fatores de Transcrição/genética , Virulência , Fatores de Virulência/genética
11.
BMC Genomics ; 20(1): 731, 2019 Oct 12.
Artigo em Inglês | MEDLINE | ID: mdl-31606035

RESUMO

BACKGROUND: TetR-family transcriptional regulators (TFTRs) are DNA binding factors that regulate gene expression in bacteria. Well-studied TFTRs, such as AcrR, which regulates efflux pump expression, are usually encoded alongside target operons. Recently, it has emerged that there are many TFTRs which act as global multi-target regulators. Our classical view of TFTRs as simple, single-target regulators therefore needs to be reconsidered. As some TFTRs regulate essential processes (e.g. metabolism) or processes which are important determinants of resistance and virulence (e.g. biofilm formation and efflux gene expression) and as TFTRs are present throughout pathogenic bacteria, they may be good drug discovery targets for tackling antimicrobial resistant infections. However, the prevalence and conservation of individual TFTR genes in Gram-negative species, has to our knowledge, not yet been studied. RESULTS: Here, a wide-scale search for TFTRs in available proteomes of clinically relevant pathogens Salmonella and Escherichia species was performed and these regulators further characterised. The majority of identified TFTRs are involved in efflux regulation in both Escherichia and Salmonella. The percentage variance in TFTR genes of these genera was found to be higher in those regulating genes involved in efflux, bleach survival or biofilm formation than those regulating more constrained processes. Some TFTRs were found to be present in all strains and species of these two genera, whereas others (i.e. TetR) are only present in some strains and some (i.e. RamR) are genera-specific. Two further pathogens on the WHO priority pathogen list (K. pneumoniae and P. aeruginosa) were then searched for the presence of the TFTRs conserved in Escherichia and Salmonella. CONCLUSIONS: Through bioinformatics and literature analyses, we present that TFTRs are a varied and heterogeneous family of proteins required for the regulation of numerous important processes, with consequences to antimicrobial resistance and virulence, and that the roles and responses of these proteins are frequently underestimated.


Assuntos
Biologia Computacional/métodos , Farmacorresistência Bacteriana , Bactérias Gram-Negativas/classificação , Fatores de Transcrição/genética , Proteínas de Bactérias/genética , Sequência Conservada , Escherichia coli/genética , Regulação Bacteriana da Expressão Gênica , Bactérias Gram-Negativas/genética , Klebsiella pneumoniae/genética , Funções Verossimilhança , Família Multigênica , Filogenia , Salmonella typhimurium/genética
12.
PLoS Genet ; 15(10): e1008425, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31589608

RESUMO

Evolutionarily conserved NusG protein enhances bacterial RNA polymerase processivity but can also promote transcription termination by binding to, and stimulating the activity of, Rho factor. Rho terminates transcription upon anchoring to cytidine-rich motifs, the so-called Rho utilization sites (Rut) in nascent RNA. Both NusG and Rho have been implicated in the silencing of horizontally-acquired A/T-rich DNA by nucleoid structuring protein H-NS. However, the relative roles of the two proteins in H-NS-mediated gene silencing remain incompletely defined. In the present study, a Salmonella strain carrying the nusG gene under the control of an arabinose-inducible repressor was used to assess the genome-wide response to NusG depletion. Results from two complementary approaches, i) screening lacZ protein fusions generated by random transposition and ii) transcriptomic analysis, converged to show that loss of NusG causes massive upregulation of Salmonella pathogenicity islands (SPIs) and other H-NS-silenced loci. A similar, although not identical, SPI-upregulated profile was observed in a strain with a mutation in the rho gene, Rho K130Q. Surprisingly, Rho mutation Y80C, which affects Rho's primary RNA binding domain, had either no effect or made H-NS-mediated silencing of SPIs even tighter. Thus, while corroborating the notion that bound H-NS can trigger Rho-dependent transcription termination in vivo, these data suggest that H-NS-elicited termination occurs entirely through a NusG-dependent pathway and is less dependent on Rut site binding by Rho. We provide evidence that through Rho recruitment, and possibly through other still unidentified mechanisms, NusG prevents pervasive transcripts from elongating into H-NS-silenced regions. Failure to perform this function causes the feedforward activation of the entire Salmonella virulence program. These findings provide further insight into NusG/Rho contribution in H-NS-mediated gene silencing and underscore the importance of this contribution for the proper functioning of a global regulatory response in growing bacteria. The complete set of transcriptomic data is freely available for viewing through a user-friendly genome browser interface.


Assuntos
Proteínas de Bactérias/metabolismo , Proteínas de Ligação a DNA/metabolismo , Regulação Bacteriana da Expressão Gênica , Fatores de Alongamento de Peptídeos/metabolismo , Salmonella typhimurium/genética , Fatores de Transcrição/metabolismo , RNA Polimerases Dirigidas por DNA/metabolismo , Loci Gênicos , Fatores de Alongamento de Peptídeos/genética , RNA Bacteriano/metabolismo , Fator Rho/genética , Fator Rho/metabolismo , Salmonella typhimurium/patogenicidade , Fatores de Transcrição/genética , Terminação da Transcrição Genética , Regulação para Cima , Fatores de Virulência/genética
13.
Vet Res ; 50(1): 71, 2019 Sep 24.
Artigo em Inglês | MEDLINE | ID: mdl-31551081

RESUMO

Eggs and raw or undercooked egg-containing food items are frequently identified as the bacterial source during epidemiolocal investigation of Salmonella outbreaks. Multi-locus variable number of tandem repeats analysis (MLVA) is a widely used Salmonella typing method enabling the study of diversity within populations of the same serotype. In vivo passage, however, has been linked with changes in MLVA type and more broadly the Salmonella genome. We sought to investigate whether in vivo passage through layer hens had an effect on MLVA type as well as the bacterial genome and whether any mutations affected bacterial virulence. Layer hens were infected with either Salmonella Typhimurium DT9 (03-24-11-11-523) as part of a single infection or were co-infected with an equal amount of Salmonella Mbandaka. Salmonella shedding in both single and co-infected birds was variable over the course of the 16-week experiment. Salmonella Typhimurium and Salmonella Mbandaka were identified in feces of co-infected birds. Salmonella colonies isolated from fecal samples were subtyped using MLVA. A single change in SSTR-6 was observed in Salmonella Typhimurium strains isolated from co-infected birds. Isolates of Salmonella Typhimurium of both the parent (03-24-11-11-523) and modified (03-24-12-11-523) MLVA type were sequenced and compared with the genome of the parent strain. Sequence analysis revealed that in vivo passaging resulted in minor mutation events. Passaged isolates exhibited significantly higher invasiveness in cultured human intestinal epithelial cells than the parent strain. The microevolution observed in this study suggests that changes in MLVA may arise more commonly and may have clinical significance.


Assuntos
Galinhas , Genoma Bacteriano/genética , Mutação/fisiologia , Doenças das Aves Domésticas/fisiopatologia , Salmonelose Animal/fisiopatologia , Salmonella typhimurium/fisiologia , Animais , Células CACO-2 , Coinfecção/microbiologia , Coinfecção/fisiopatologia , Coinfecção/veterinária , Fezes/microbiologia , Feminino , Humanos , Repetições Minissatélites , Tipagem de Sequências Multilocus/veterinária , Doenças das Aves Domésticas/microbiologia , Salmonelose Animal/microbiologia , Salmonella enterica/fisiologia , Salmonella typhimurium/genética , Inoculações Seriadas , Virulência
14.
Biochimie ; 167: 152-161, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31563538

RESUMO

The intracellular pathogen Salmonella is an important cause of human foodborne diseases worldwide. Salmonella takes advantage of the phosphorelay regulatory systems to survive in the hostile environment of the host's gastrointestinal tract. It has been reported that the nitrate reductase Z (NR-Z), encoded by the narUZYV operon, is required during Salmonella transition to anaerobic environments and is constitutively produced at low levels, but little is known about the regulatory mechanism involved in the operon gene expression. In this work, we found that the RcsCDB system is activated by high concentrations of specific sugars as a carbon source. In this activation state, the RcsCDB system participates in the negative control of narUZYWV expression. This control strategy occurs during exponential growth when the carbon source is high, allowing for normal aerobic respiration. The RcsCDB system's participation in aerobic respiration is necessary to ensure efficient metabolism and optimal energy consumption when the bacteria are growing exponentially.


Assuntos
Proteínas de Bactérias/genética , Nitrato Redutase/genética , Salmonella typhimurium/crescimento & desenvolvimento , Salmonella typhimurium/genética , Fatores de Transcrição/fisiologia , Regulação Bacteriana da Expressão Gênica , Óperon , Regiões Promotoras Genéticas , Transcrição Genética
15.
Biosci Trends ; 13(4): 342-350, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31527330

RESUMO

The retinoid-interferon-induced mortality-19 (GRIM-19) gene has been identified as a negative regulator associated with tumor development. The current study created a model of an orthotopically implanted hepatocarcinoma tumor to verify the inhibitory effect of GRIM-19 in vivo. After treatment with GRIM-19 carried by attenuated Salmonella, transplanted tumors were measured with an Imaging System. The expression of GRIM-19, Stat3/p-Stat3, cyclinD1, CDK4, PCNA, Bax/Bcl-2, cleaved caspase-9/3, VEGF, and MMP-2/9 was determined using immunohistochemistry and Western blot analysis. The cell cycle was assessed using flow cytometry (FCM). Apoptosis was determined using FCM and a TUNEL assay. Results indicated that GRIM-19 overexpression resulted in inhibition of peritoneal metastasis, induction of cell cycle arrest, and apoptosis in vivo. In addition, the expression of Stat3/p-Stat3 was down-regulated by GRIM-19. These results suggest that GRIM-19 overexpression could suppress the growth of orthotopically implanted hepatocarcinoma tumors by reversing the regulation of the Stat3 signaling pathway. This approach could potentially be a powerful treatment for hepatocarcinoma.


Assuntos
Vetores Genéticos/administração & dosagem , Neoplasias Hepáticas/terapia , NADH NADPH Oxirredutases/genética , Fator de Transcrição STAT3/metabolismo , Animais , Apoptose/genética , Carcinogênese/genética , Carcinoma Hepatocelular/prevenção & controle , Carcinoma Hepatocelular/secundário , Pontos de Checagem do Ciclo Celular/genética , Linhagem Celular Tumoral/transplante , Proliferação de Células/genética , Modelos Animais de Doenças , Regulação para Baixo , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Terapia Genética/métodos , Humanos , Neoplasias Hepáticas/patologia , Camundongos , Terapia de Alvo Molecular/métodos , NADH NADPH Oxirredutases/metabolismo , Neoplasias Peritoneais/prevenção & controle , Salmonella typhimurium/genética , Transdução de Sinais/genética , Resultado do Tratamento
16.
Food Chem Toxicol ; 134: 110815, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31520668

RESUMO

Genotoxicity of B. officinalis, G. officinalis, V. luteum and V. hirundinaria extracts, which demonstrated strong antioxidant capacity, was tested using chromosome aberration, sister chromatid exchange (SCE), cytokinesis-block micronucleus and alkaline single-cell gel electrophoresis (comet) assays in human lymphocytes in vitro and Ames Salmonella/microsome test. All tested extracts were not mutagenic in S. typhimurium strains TA98 and TA100 with and without metabolic activation and did not induce chromosome aberrations in human lymphocytes in vitro. Extract from G. officinalis was the only one, which induced significant increase in micronuclei, indicating possible aneugenic effect. All investigated plant extracts induced DNA damage evaluated by the comet assay, while B. officinalis and V. luteum extracts induced slight increase in SCE values. The determined variation in response might be due to the plant extract tested and donor susceptibility.


Assuntos
Lamiales/química , Mutagênicos/toxicidade , Extratos Vegetais/toxicidade , Stachys/química , Vincetoxicum/química , Ensaio Cometa , Humanos , Linfócitos/efeitos dos fármacos , Testes para Micronúcleos , Salmonella typhimurium/genética
17.
Nat Commun ; 10(1): 4280, 2019 09 19.
Artigo em Inglês | MEDLINE | ID: mdl-31537784

RESUMO

Bloodstream infections by Salmonella enterica serovar Typhimurium constitute a major health burden in sub-Saharan Africa (SSA). These invasive non-typhoidal (iNTS) infections are dominated by isolates of the antibiotic resistance-associated sequence type (ST) 313. Here, we report emergence of ST313 sublineage II.1 in the Democratic Republic of the Congo. Sublineage II.1 exhibits extensive drug resistance, involving a combination of multidrug resistance, extended spectrum ß-lactamase production and azithromycin resistance. ST313 lineage II.1 isolates harbour an IncHI2 plasmid we name pSTm-ST313-II.1, with one isolate also exhibiting decreased ciprofloxacin susceptibility. Whole genome sequencing reveals that ST313 II.1 isolates have accumulated genetic signatures potentially associated with altered pathogenicity and host adaptation, related to changes observed in biofilm formation and metabolic capacity. Sublineage II.1 emerged at the beginning of the 21st century and is involved in on-going outbreaks. Our data provide evidence of further evolution within the ST313 clade associated with iNTS in SSA.


Assuntos
Adaptação Fisiológica/efeitos dos fármacos , Antibacterianos/farmacologia , Farmacorresistência Bacteriana Múltipla/genética , Salmonella typhimurium/efeitos dos fármacos , Salmonella typhimurium/genética , Adaptação Fisiológica/genética , Animais , Azitromicina/farmacologia , Biofilmes/crescimento & desenvolvimento , Linhagem Celular , Ciprofloxacino/farmacologia , República Democrática do Congo , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Testes de Sensibilidade Microbiana , Plasmídeos/genética , Salmonella typhimurium/isolamento & purificação , Células THP-1 , Sequenciamento Completo do Genoma
18.
Microbiol Res ; 229: 126319, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31479952

RESUMO

Methionine is critical for variety of metabolic processes in biological organisms, acting as a precursor or intermediate for many final products. The last step for the synthesis of methionine is the methylation of homocysteine, which is catalyzed by MetE. Here, we use Salmonella enterica serovar Typhimurium LT2 to study the regulation of the metE+ gene by an anaerobically induced small non-coding RNA-FnrS, the expression of which is strictly dependent on the anaerobic regulator-FNR. The MetE-HA protein was expressed at an increased level in the fnrS- and hfq- deficient strains under anaerobic conditions. The Hfq protein is predicted to stabilize the binding between small RNA(s) and their target mRNA(s). A transcriptional (op) and translational (pr) metE::lacZ fusion gene were separately constructed, with the metE+-promoter fused to a lacZ reporter gene. In an anaerobic environment, the metE::lacZ (pr) fusion gene and reverse transcription-PCR identified that FnrS and/or FNR negatively regulate metE+ mRNA levels in the rich media. Analysis of FnrS revealed a sequence complementary to the 5' mRNA translational initiation region (TIR) of the metE+ gene. Mutation(s) predicted to disrupt base pairing between FnrS and metE+ TIR were constructed in fnrS, and most of those resulted in the loss of repressive activity. When compensatory mutation(s) were made in metE+ 5' TIR to restore base pairing with FnrS, the repressive regulation was completely restored. Therefore, in this study, we identified that in anaerobic phase, there is a repression of metE+ gene expression by FnrS and that base-paring, between both expressive transcripts, plays an important role for this negative regulation.


Assuntos
Proteínas de Bactérias/genética , Regulação Bacteriana da Expressão Gênica , Metiltransferases/genética , RNA Bacteriano/genética , RNA Mensageiro/genética , Pequeno RNA não Traduzido/genética , Salmonella typhimurium/enzimologia , Proteínas de Bactérias/metabolismo , Pareamento de Bases , Sequência de Bases , Regulação Enzimológica da Expressão Gênica , Metiltransferases/química , Metiltransferases/metabolismo , Conformação de Ácido Nucleico , RNA Bacteriano/química , RNA Bacteriano/metabolismo , RNA Mensageiro/química , RNA Mensageiro/metabolismo , Pequeno RNA não Traduzido/química , Pequeno RNA não Traduzido/metabolismo , Salmonella typhimurium/genética , Salmonella typhimurium/metabolismo
19.
Microbiology ; 165(11): 1245-1250, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31486760

RESUMO

In Salmonella enterica serovar Typhimurium, the RcsCDB regulatory system controls the expression of genes involved in synthesis of colanic acid, formation of flagella and virulence. Here, we show that activation of the RcsCDB system downregulates expression of std, an operon that encodes fimbriae involved in Salmonella attachment to the mucus layer in the large intestine. Bioinformatic analysis predicts the existence of an RcsB-binding site located 180 bp upstream to the +1 transcription start site of the std promoter, and electrophoretic mobility shift assays confirm that RcsB binds the std promoter region in vitro. This study adds RcsB to the list of regulators of std transcription and provides an example of modulation of fimbriae synthesis by a signal transduction system.


Assuntos
Proteínas de Bactérias/metabolismo , Proteínas de Fímbrias/genética , Fímbrias Bacterianas/genética , Regulação Bacteriana da Expressão Gênica , Salmonella typhimurium/metabolismo , Transdução de Sinais , Aderência Bacteriana , Proteínas de Bactérias/genética , Sítios de Ligação , Mutação , Óperon , Regiões Promotoras Genéticas , Salmonella typhimurium/genética , Transcrição Genética
20.
Int J Mol Sci ; 20(18)2019 Sep 04.
Artigo em Inglês | MEDLINE | ID: mdl-31487790

RESUMO

Benzimidazole derivatives have a diverse range of biological activities, including antiulcer, antihypertensive, antiviral, antifungal, anti-inflammatory, and anticancer. Despite these activities, previous studies have revealed that some of the derivatives can induce mutations. This study aimed to screen for potential mutagenic activities of novel benzimidazole derivatives 1-4 using the Ames test and to study their structure-activity relationship (SAR). An Ames test was carried out on two strains of Salmonella typhimurium (TA98 and TA100) in the absence and presence of metabolic activation. Genetic analysis was performed prior to the Ames test to determine the genotypes of the bacterial tester strains. Both bacterial strains showed dependency on histidine with the presence of rfa mutation, uvrB deletion, and plasmid pKM101. Further, all derivatives tested showed no mutagenic activity in the absence of metabolic activation in both tester strains. However, in the presence of metabolic activation, compound 1 appeared to induce mutation at 2.5 µg/plate when tested against the TA98 strain. These results suggest that the absence of the -OH group at the ortho-position over the phenyl ring might be the cause of increased mutagenic activity in compound 1. Additionally, the presence of mutagenic activity in compound 1 when it was metabolically activated indicates that this compound is a promutagen.


Assuntos
Benzimidazóis/química , Resistência Microbiana a Medicamentos/genética , Mutagênicos/química , Ativação Metabólica , Benzimidazóis/metabolismo , Benzimidazóis/toxicidade , Genótipo , Mutagênicos/metabolismo , Mutagênicos/toxicidade , Salmonella typhimurium/efeitos dos fármacos , Salmonella typhimurium/genética , Relação Estrutura-Atividade
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA