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1.
BMC Infect Dis ; 21(1): 310, 2021 Mar 31.
Artigo em Inglês | MEDLINE | ID: mdl-33789598

RESUMO

BACKGROUND: Recent seroepidemiological studies have suggested that tularemia could be an endemic bacterial zoonosis in Iran. METHODS: From January 2016 to June 2018, disease cases characterized by fever, cervical lymphadenopathy and ocular involvement were reported in Youzband Village of Kaleybar County, in the East Azerbaijan Province, northwestern Iran. Diagnostic tests included Francisella tularensis serology (including tube agglutination test and ELISA), PCR, and culture. RESULTS: Among 11 examined case-patients, the tularemia tube agglutination test was positive in ten and borderline in one. PCR detected the F. tularensis ISFtu2 elements and fopA gene in one rodent and a spring water sample from the same geographic area. CONCLUSIONS: Based on the clinical manifestations of the disease suggesting an oropharyngeal form of tularemia, serology results in case patients, and F. tularensis detection in the local fauna and aquatic environment, the water supply of the village was the likely source of the tularemia outbreak. Intervention such as dredging and chlorination of the main water storage tank of the village and training of villagers and health care workers in preventive measures and treatment of the illness helped control the infection.


Assuntos
Francisella tularensis/isolamento & purificação , Tularemia/diagnóstico , Adolescente , Adulto , Idoso , Testes de Aglutinação , Animais , Proteínas da Membrana Bacteriana Externa/genética , Criança , Pré-Escolar , DNA Bacteriano/metabolismo , Feminino , Francisella tularensis/genética , Água Doce/microbiologia , Humanos , Irã (Geográfico) , Masculino , Camundongos , Reação em Cadeia da Polimerase , Tularemia/microbiologia
2.
J Med Microbiol ; 70(3)2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-33616518

RESUMO

Introduction. Streptococcus dysgalactiae subspecies equisimilis (SDSE) is becoming increasingly recognized as an important human pathogen. Recurrent bacteremia with SDSE has been described previously.Aim. The aims of the study were to establish the genetic relatedness of SDSE isolates with emm-type stG643 that had caused recurrent bacteraemia in three patients and to search for signs of horizontal gene transfer of the emm gene in a collection of SDSE stG643 genomes.Hypothesis. Recurring SDSE bacteremia is caused by the same clone in one patient.Methodology. Whole genome sequencing of 22 clinical SDSE stG643 isolates was performed, including three paired blood culture isolates and sixteen isolates from various sites. All assemblies were aligned to a reference assembly and SNPs were extracted. A total of 53 SDSE genomes were downloaded from GenBank. Two phylogenetic trees, including all 75 SDSE isolates, were created. One tree was based on the emm gene only and one tree was based on all variable positions in the genomes.Results. The genomes from the three pairs of SDSE isolates showed high sequence similarity (1-17 SNPs difference between the pairs), whereas the median SNP difference between the 22 isolates in our collection was 1694 (range 1-11257). The paired isolates were retrieved with 7-53 months between episodes. The 22 SDSE isolates from our collection formed a cluster in the phylogenetic tree based on the emm gene, while they were more scattered in the tree based on all variable positions.Conclusions. Our results show that the paired isolates were of the same clonal origin, which in turn supports carriage between bacteraemia episodes. The phylogenetic analysis indicates that horizontal gene transfer of the emm-gene between some of the SDSE isolates has occurred.


Assuntos
Bacteriemia/microbiologia , Infecções Estreptocócicas/microbiologia , Streptococcus/genética , Antígenos de Bactérias/genética , Proteínas da Membrana Bacteriana Externa/genética , Proteínas de Transporte/genética , Genoma Bacteriano/genética , Humanos , Filogenia , Streptococcus/classificação , Streptococcus/isolamento & purificação
3.
Int J Mol Sci ; 22(3)2021 Feb 02.
Artigo em Inglês | MEDLINE | ID: mdl-33540627

RESUMO

In this study, we found that the loss of OmpR, the response regulator of the two-component EnvZ/OmpR system, increases the cellular level of Fur, the master regulator of iron homeostasis in Y. enterocolitica. Furthermore, we demonstrated that transcription of the fur gene from the YePfur promoter is subject to negative OmpR-dependent regulation. Four putative OmpR-binding sites (OBSs) were indicated by in silico analysis of the fur promoter region, and their removal affected OmpR-dependent fur expression. Moreover, OmpR binds specifically to the predicted OBSs which exhibit a distinct hierarchy of binding affinity. Finally, the data demonstrate that OmpR, by direct binding to the promoters of the fecA, fepA and feoA genes, involved in the iron transport and being under Fur repressor activity, modulates their expression. It seems that the negative effect of OmpR on fecA and fepA transcription is sufficient to counteract the indirect, positive effect of OmpR resulting from decreasing the Fur repressor level. The expression of feoA was positively regulated by OmpR and this mode of action seems to be direct and indirect. Together, the expression of fecA, fepA and feoA in Y. enterocolitica has been proposed to be under a complex mode of regulation involving OmpR and Fur regulators.


Assuntos
Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Regulação Bacteriana da Expressão Gênica , Ferro/metabolismo , Proteínas Repressoras/genética , Transativadores/metabolismo , Yersinia enterocolitica/metabolismo , Proteínas da Membrana Bacteriana Externa/genética , Proteínas de Transporte/genética , Simulação por Computador , Homeostase , Proteínas de Ligação ao Ferro/genética , Regiões Promotoras Genéticas , Receptores de Superfície Celular/genética , Yersinia enterocolitica/genética
4.
Nat Commun ; 12(1): 1183, 2021 02 19.
Artigo em Inglês | MEDLINE | ID: mdl-33608518

RESUMO

Ice-nucleation active (INA) bacteria can promote the growth of ice more effectively than any other known material. Using specialized ice-nucleating proteins (INPs), they obtain nutrients from plants by inducing frost damage and, when airborne in the atmosphere, they drive ice nucleation within clouds, which may affect global precipitation patterns. Despite their evident environmental importance, the molecular mechanisms behind INP-induced freezing have remained largely elusive. We investigate the structural basis for the interactions between water and the ice-nucleating protein InaZ from the INA bacterium Pseudomonas syringae. Using vibrational sum-frequency generation (SFG) and two-dimensional infrared spectroscopy, we demonstrate that the ice-active repeats of InaZ adopt a ß-helical structure in solution and at water surfaces. In this configuration, interaction between INPs and water molecules imposes structural ordering on the adjacent water network. The observed order of water increases as the interface is cooled to temperatures close to the melting point of water. Experimental SFG data combined with molecular-dynamics simulations and spectral calculations show that InaZ reorients at lower temperatures. This reorientation can enhance water interactions, and thereby the effectiveness of ice nucleation.


Assuntos
Proteínas da Membrana Bacteriana Externa/química , Proteínas da Membrana Bacteriana Externa/metabolismo , Temperatura Baixa , Água/química , Atmosfera , Proteínas da Membrana Bacteriana Externa/genética , Óxido de Deutério , Congelamento , Gelo , Simulação de Dinâmica Molecular , Plantas/microbiologia , Pseudomonas syringae/metabolismo
5.
Proc Natl Acad Sci U S A ; 118(1)2021 01 05.
Artigo em Inglês | MEDLINE | ID: mdl-33443205

RESUMO

The type 6 secretion system (T6SS) is a dynamic organelle encoded by many gram-negative bacteria that can be used to kill competing bacterial prey species in densely occupied niches. Some predatory species, such as Vibrio cholerae, use their T6SS in an untargeted fashion while in contrast, Pseudomonas aeruginosa assembles and fires its T6SS apparatus only after detecting initial attacks by other bacterial prey cells; this targeted attack strategy has been termed the T6SS tit-for-tat response. Molecules that interact with the P. aeruginosa outer membrane such as polymyxin B can also trigger assembly of T6SS organelles via a signal transduction pathway that involves protein phosphorylation. Recent work suggests that a phospholipase T6SS effector (TseL) of V. cholerae can induce T6SS dynamic activity in P. aeruginosa when delivered to or expressed in the periplasmic space of this organism. Here, we report that inhibiting expression of essential genes involved in outer membrane biogenesis can also trigger T6SS activation in P. aeruginosa Specifically, we developed a CRISPR interference (CRISPRi) system to knock down expression of bamA, tolB, and lptD and found that these knockdowns activated T6SS activity. This increase in T6SS activity was dependent on the same signal transduction pathway that was previously shown to be required for the tit-for-tat response. We conclude that outer membrane perturbation can be sensed by P. aeruginosa to activate the T6SS even when the disruption is generated by aberrant cell envelope biogenesis.


Assuntos
Proteínas da Membrana Bacteriana Externa/metabolismo , Sistemas CRISPR-Cas , Membrana Celular/metabolismo , Genes Essenciais/fisiologia , Proteínas Periplásmicas/metabolismo , Pseudomonas aeruginosa/genética , Sistemas de Secreção Tipo VI/metabolismo , Proteínas da Membrana Bacteriana Externa/genética , Membrana Celular/genética , Membrana Celular/patologia , Sobrevivência Celular/genética , Técnicas de Silenciamento de Genes , Inativação Gênica , Genes Essenciais/genética , Genótipo , Proteínas Periplásmicas/genética , Fenótipo , Pseudomonas aeruginosa/citologia , Pseudomonas aeruginosa/crescimento & desenvolvimento , Pseudomonas aeruginosa/metabolismo , RNA-Seq , Transdução de Sinais/genética , Estresse Fisiológico , Vibrio cholerae/genética , Vibrio cholerae/crescimento & desenvolvimento
6.
Biomed Pharmacother ; 134: 111149, 2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-33385683

RESUMO

E. coli is associated with high rates of infection and resistance to drugs not only in China but also the rest of the world. In addition, the number of E. coli biofilm infections continue to increase with time. Notably, biofilms are attractive targets for the prevention of infections caused by multidrug-resistant bacteria. Moreover, the pgaABCD-encoded Poly-ß-1,6-N-acetyl-d-glucosamine (PNAG) plays an important role in biofilm formation. Therefore, this study aimed to explore the specific effect of the (R)-(+)-pulegone (PU) on growth and biofilm formation in multi-drug resistant E. coli. The molecular mechanisms involved were also examined. The results showed that PU had significant antibacterial and antibiofilm formation activity against E. coli K1, with MIC and MBC values of 23.68 and 47.35 mg/mL, respectively. On the other hand, the maximum inhibition rate for biofilm formation in the bacterium was 52.36 % at 94.70 mg/mL of PU. qRT-PCR data showed that PU significantly down-regulated expression of the pgaABCD genes (P < 0.05). PU was also broadly effective against biofilm formation in MG1655 and MG1655/ΔpgaABCD, exhibiting the maximum inhibition rates were 98.23 % and 93.35 %, respectively. In addition, PU destroyed pre-formed mature biofilm in both MG1655 and MG1655/ΔpgaABCD about 95.03 % and 92.4 %, respectively. The study therefore verified that pgaA was a potential and key target for PU in E. coli although it was not the only one. Overall, the findings indicated that PU is a potential and novel inhibitor of drug resistance, This therefore gives insights on new ways of preventing and treating biofilm-associated infections in the food industry as well as in clinical practice.


Assuntos
Antibacterianos/farmacologia , Proteínas da Membrana Bacteriana Externa/genética , Biofilmes/efeitos dos fármacos , Monoterpenos Cicloexânicos/farmacologia , Farmacorresistência Bacteriana Múltipla/genética , Escherichia coli K12/efeitos dos fármacos , Proteínas de Escherichia coli/genética , Amidoidrolases/genética , Biofilmes/crescimento & desenvolvimento , Escherichia coli K12/genética , Escherichia coli K12/crescimento & desenvolvimento , Regulação Bacteriana da Expressão Gênica , Testes de Sensibilidade Microbiana
7.
Lett Appl Microbiol ; 72(5): 496-508, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-33332656

RESUMO

The most common presentation of animal leptospirosis is the subclinical and silent chronic form, that can lead to important reproductive disorders. The diagnosis of this chronic form remains a challenge. The aim of the present study is to gather and critically analyse the current information about molecular tools applied to animal leptospirosis diagnosis, particularly the silent chronic presentation of the infection. Regarding clinical specimens, samples from urinary tract were the most used (69/102, 67·7%), while few studies (12/102, 11·8%) investigated samples from reproductive tract. Concerning the molecular methods applied, the most used is still the conventional polymerase chain reaction (PCR) (46/102, 45%), followed by real-time PCR (38/102, 37·2%). The lipL32 gene is currently the most common target used for Leptospira detection, with 48% of studies applying this genetic marker. From all the studies, only few (21/102, 20·5%) performed gene sequencing. According to the majority of authors, current evidence suggests that lipL32-PCR is useful for an initial screening for Leptospira DNA detection in animal clinical samples. Posteriorly, if DNA sequencing could be performed on positive lipL32-PCR samples, we encourage the use of secY gene as a genetic marker. The molecular methods appear as the most important tools for the diagnosis of the chronic silent leptospirosis on domestic animals, reinforcing its evident impact not only on animal reproduction but also on a One Health context.


Assuntos
Proteínas da Membrana Bacteriana Externa/genética , Leptospira/genética , Leptospirose/diagnóstico , Leptospirose/veterinária , Lipoproteínas/genética , Canais de Translocação SEC/genética , Animais , Animais Domésticos , Técnicas de Diagnóstico Molecular/métodos , Reação em Cadeia da Polimerase em Tempo Real/métodos , Sensibilidade e Especificidade , Análise de Sequência de DNA
8.
Int J Biol Macromol ; 168: 289-300, 2021 Jan 31.
Artigo em Inglês | MEDLINE | ID: mdl-33310091

RESUMO

Leptospirosis is a potentially fatal zoonosis that is caused by spirochete Leptospira. The signs and symptoms of leptospirosis are usually varied, allowing it to be mistaken for other causes of acute febrile syndromes. Thus, early diagnosis and identification of a specific agent in clinical samples is crucial for effective treatment. This study was aimed to develop specific monoclonal antibodies against LipL21 antigen for future use in leptospirosis rapid and accurate immunoassay. A recombinant LipL21 (rLipL21) antigen was optimized for expression and evaluated for immunogenicity. Then, a naïve phage antibody library was utilized to identify single chain fragment variable (scFv) clones against the rLipL21 antigen. A total of 47 clones were analysed through monoclonal phage ELISA. However, after taking into consideration the background OD405 values, only 4 clones were sent for sequencing to determine human germline sequences. The sequence analysis showed that all 4 clones are identical. The in silico analysis of scFv-lip-1 complex indicated that the charged residues of scFv CDRs are responsible for the recognition with rLipL21 epitopes. The generated monoclonal antibody against rLipL21 will be evaluated as a detection reagent for the diagnosis of human leptospirosis in a future study.


Assuntos
Antígenos de Bactérias/imunologia , Proteínas da Membrana Bacteriana Externa/imunologia , Técnicas de Visualização da Superfície Celular/métodos , Leptospira/imunologia , Lipoproteínas/imunologia , Animais , Anticorpos Monoclonais/imunologia , Antígenos de Bactérias/isolamento & purificação , Antígenos de Bactérias/metabolismo , Proteínas da Membrana Bacteriana Externa/genética , Proteínas da Membrana Bacteriana Externa/isolamento & purificação , Proteínas da Membrana Bacteriana Externa/metabolismo , Bacteriófagos/metabolismo , Ensaio de Imunoadsorção Enzimática/métodos , Epitopos/imunologia , Humanos , Imunoensaio/métodos , Leptospira/metabolismo , Leptospira interrogans/genética , Leptospirose/diagnóstico , Lipoproteínas/isolamento & purificação , Lipoproteínas/metabolismo , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes/genética , Anticorpos de Cadeia Única/imunologia
9.
Biochim Biophys Acta Biomembr ; 1863(1): 183488, 2021 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-33065135

RESUMO

Gram-negative bacteria export a large variety of antimicrobial compounds by forming two-membrane spanning tripartite multidrug efflux systems composed of an inner membrane transporter, an outer membrane channel and a periplasmic adaptor protein. Here we present the co-expression, purification and first electron microscopy insights of the Escherichia coli EmrAB-TolC tripartite Major Facilitator Superfamily (MSF) efflux system as a whole complex stabilized by Amphipol polymer. The structure reveals a 33 nm long complex delineated by the Amphipol belt at both extremities. Comparison of projection structures of EmrAB-TolC and AcrAB-TolC indicates that the outer membrane protein TolC linked to the periplasmic adaptor EmrA protein form an extended periplasmic canal. The overall length of EmrAB-TolC complex is similar to that of AcrAB-TolC with a probable tip-to-tip interaction between EmrA and TolC unveiling how the adaptor protein connects TolC and EmrB embedded in the inner membrane.


Assuntos
Proteínas da Membrana Bacteriana Externa/química , Proteínas de Escherichia coli/química , Escherichia coli/química , Proteínas de Membrana/química , Proteínas de Membrana Transportadoras/química , Complexos Multiproteicos/química , Proteínas da Membrana Bacteriana Externa/genética , Proteínas da Membrana Bacteriana Externa/metabolismo , Escherichia coli/genética , Escherichia coli/metabolismo , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Complexos Multiproteicos/genética , Complexos Multiproteicos/metabolismo , Estrutura Quaternária de Proteína
10.
PLoS One ; 15(10): e0239991, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33091006

RESUMO

Shedding of DNA of pathogenic Leptospira spp. has been documented in naturally infected cats in several countries, but urinary shedding of infectious Leptospira spp. has only recently been proven. The climate in Southern Chile is temperate rainy with high annual precipitations which represents ideal preconditions for survival of Leptospira spp., especially during spring and summer. The aims of this study were to investigate shedding of pathogenic Leptospira spp. in outdoor cats in Southern Chile, to perform molecular characterization of isolates growing in culture, and to assess potential risk factors associated with shedding. Urine samples of 231 outdoor cats from rural and urban areas in southern Chile were collected. Urine samples were investigated for pathogenic Leptospira spp. by 4 techniques: qPCR targeting the lipL32 gene, immunomagnetic separation (IMS)-coupled qPCR (IMS-qPCR), direct culture and IMS-coupled culture. Positive urine cultures were additionally confirmed by PCR. Multilocus sequence typing (MLST) was used to molecularly characterize isolates obtained from positive cultures. Overall, 36 urine samples (15.6%, 95% confidence interval (CI) 11.4-20.9) showed positive results. Eighteen (7.8%, 95% CI 4.9-12.1), 30 (13%, 95% CI 9.2-18), 3 (1.3%, 0.3-3.9) and 4 cats (1.7%; 95% CI 0.5-4.5) were positive in qPCR, IMS-qPCR, conventional culture, and IMS-coupled culture, respectively. MLST results of 7 culture-positive cats revealed sequences that could be assigned to sequence type 17 (6 cats) and sequence type 27 (1 cat) corresponding to L. interrogans (Pathogenic Leptospira Subgroup 1). Shedding of pathogenic Leptospira spp. by cats might be an underestimated source of infection for other species including humans. The present study is the first one reporting growth of leptospires from feline urine in culture in naturally infected cats in South-America and characterisation of culture-derived isolates. So far, very few cases of successful attempts to culture leptospires from naturally infected cats are described worldwide.


Assuntos
Derrame de Bactérias/fisiologia , Doenças do Gato/patologia , Leptospira/patogenicidade , Leptospirose/patologia , Animais , Proteínas da Membrana Bacteriana Externa/genética , Proteínas da Membrana Bacteriana Externa/isolamento & purificação , Doenças do Gato/microbiologia , Doenças do Gato/transmissão , Gatos , DNA Bacteriano/metabolismo , Feminino , Leptospira/genética , Leptospira/isolamento & purificação , Leptospirose/microbiologia , Leptospirose/transmissão , Leptospirose/veterinária , Lipoproteínas/genética , Lipoproteínas/isolamento & purificação , Masculino , Tipagem de Sequências Multilocus , Reação em Cadeia da Polimerase em Tempo Real , Fatores de Risco , Urina/microbiologia
11.
Proc Natl Acad Sci U S A ; 117(38): 23356-23364, 2020 09 22.
Artigo em Inglês | MEDLINE | ID: mdl-32879005

RESUMO

Much attention is being paid to conformational biases in the ensembles of intrinsically disordered proteins. However, it is currently unknown whether or how conformational biases within the disordered ensembles of foldable proteins affect function in vivo. Recently, we demonstrated that water can be a good solvent for unfolded polypeptide chains, even those with a hydrophobic and charged sequence composition typical of folded proteins. These results run counter to the generally accepted model that protein folding begins with hydrophobicity-driven chain collapse. Here we investigate what other features, beyond amino acid composition, govern chain collapse. We found that local clustering of hydrophobic and/or charged residues leads to significant collapse of the unfolded ensemble of pertactin, a secreted autotransporter virulence protein from Bordetella pertussis, as measured by small angle X-ray scattering (SAXS). Sequence patterns that lead to collapse also correlate with increased intermolecular polypeptide chain association and aggregation. Crucially, sequence patterns that support an expanded conformational ensemble enhance pertactin secretion to the bacterial cell surface. Similar sequence pattern features are enriched across the large and diverse family of autotransporter virulence proteins, suggesting sequence patterns that favor an expanded conformational ensemble are under selection for efficient autotransporter protein secretion, a necessary prerequisite for virulence. More broadly, we found that sequence patterns that lead to more expanded conformational ensembles are enriched across water-soluble proteins in general, suggesting protein sequences are under selection to regulate collapse and minimize protein aggregation, in addition to their roles in stabilizing folded protein structures.


Assuntos
Proteínas da Membrana Bacteriana Externa/química , Proteínas de Bactérias/química , Bordetella pertussis/metabolismo , Desdobramento de Proteína , Fatores de Virulência de Bordetella/química , Sequência de Aminoácidos , Proteínas da Membrana Bacteriana Externa/genética , Proteínas da Membrana Bacteriana Externa/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Bordetella pertussis/química , Bordetella pertussis/genética , Conformação Proteica , Dobramento de Proteína , Espalhamento a Baixo Ângulo , Fatores de Virulência de Bordetella/genética , Fatores de Virulência de Bordetella/metabolismo
12.
Am J Trop Med Hyg ; 103(4): 1427-1434, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32748767

RESUMO

Molecular data are required to improve our understanding of the epidemiology of leptospirosis in Africa and to identify sources of human infection. We applied molecular methods to identify the infecting Leptospira species and genotypes among patients hospitalized with fever in Tanzania and compared these with Leptospira genotypes detected among animals in Tanzania to infer potential sources of human infection. We performed lipL32 real-time PCR to detect the presence of pathogenic Leptospira in acute-phase plasma, serum, and urine samples obtained from study participants with serologically confirmed leptospirosis and participants who had died with febrile illness. Leptospira blood culture was also performed. In positive specimens, we performed species-specific PCR and compared participant Leptospira secY sequences with Leptospira reference sequences and sequences previously obtained from animals in Tanzania. We detected Leptospira DNA in four (3.6%) of 111 participant blood samples. We detected Leptospira borgpetersenii (one participant, 25.0%), Leptospira interrogans (one participant, 25.0%), and Leptospira kirschneri (one participant, 25.0%) (one [25%] undetermined). Phylogenetic comparison of secY sequence from the L. borgpetersenii and L. kirschneri genotypes detected from participants was closely related to but distinct from genotypes detected among local livestock species. Our results indicate that a diverse range of Leptospira species is causing human infection. Although our analysis suggests a close relationship between Leptospira genotypes found in people and livestock, continued efforts are needed to obtain more Leptospira genetic material from human leptospirosis cases to help prioritize Leptospira species and genotypes for control.


Assuntos
Leptospira/isolamento & purificação , Leptospirose/transmissão , Gado/microbiologia , Animais , Proteínas da Membrana Bacteriana Externa/genética , Reservatórios de Doenças , Genes Bacterianos , Técnicas de Genotipagem , Humanos , Leptospira/classificação , Leptospira/genética , Leptospirose/epidemiologia , Leptospirose/microbiologia , Lipoproteínas/genética , Patologia Molecular , Filogenia , Canais de Translocação SEC/genética , Tanzânia/epidemiologia , Zoonoses/epidemiologia
13.
Arch Microbiol ; 202(10): 2711-2726, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-32728830

RESUMO

In recent years, bioremediation is considered as an efficient method to remove the pollutants from the industrial wastewater. In this study, quantitative gene expressions (Real-time RT-PCR) of mtr gene cluster (mtrA, mtrB, mtrC, mtrD, mtrE, mtrF and omcA) in five different uranium concentrations (0.1, 0.25, 0.5, 1 and 2 mM) were performed with ICP and microscopic live cell counting analysis under anaerobic condition, by Shewanella RCRI7 as a native bacterium. The results indicated that the amount of uranium removal and live-cell counting were decreased in the higher uranium concentrations (1 and 2 mM), due to the uranium toxicity, suggesting 0.5 mM as the optimum uranium concentration for Shewanella RCRI7 resistance. The expression of mtrCED and omcA genes presented increasing trend in the lower uranium concentrations (0.1, 0.25 and 0.5 mM) and a decreasing trend in 1 and 2 mM, while mtrABF, presented an inverse pattern, proving the alternative role of mtrF for mtrC and omcA, as the substantial multiheme cytochromes in Extracellular Electron Transfer (EET) pathway. These data are a proof of these gene vital roles in the EET pathway, proposing them for genetic engineering toward EET optimization, as the certain pathway in heavy metal bioremediation process.


Assuntos
Biodegradação Ambiental , Proteínas de Membrana Transportadoras/genética , Shewanella/genética , Shewanella/metabolismo , Urânio/análise , Poluentes Químicos da Água/análise , Proteínas da Membrana Bacteriana Externa/genética , Grupo dos Citocromos c/genética , Transporte de Elétrons/genética , Família Multigênica/genética , Oxirredução , Águas Residuárias/química , Poluição da Água/análise
14.
Nat Commun ; 11(1): 3571, 2020 07 16.
Artigo em Inglês | MEDLINE | ID: mdl-32678094

RESUMO

Pathogenic bacteria of the genus Bartonella can induce vasoproliferative lesions during infection. The underlying mechanisms are unclear, but involve secretion of an unidentified mitogenic factor. Here, we use functional transposon-mutant screening in Bartonella henselae to identify such factor as a pro-angiogenic autotransporter, called BafA. The passenger domain of BafA induces cell proliferation, tube formation and sprouting of microvessels, and drives angiogenesis in mice. BafA interacts with vascular endothelial growth factor (VEGF) receptor-2 and activates the downstream signaling pathway, suggesting that BafA functions as a VEGF analog. A BafA homolog from a related pathogen, Bartonella quintana, is also functional. Our work unveils the mechanistic basis of vasoproliferative lesions observed in bartonellosis, and we propose BafA as a key pathogenic factor contributing to bacterial spread and host adaptation.


Assuntos
Proteínas da Membrana Bacteriana Externa/metabolismo , Bartonella/patogenicidade , Neovascularização Patológica/metabolismo , Transdução de Sinais , Sistemas de Secreção Tipo V/metabolismo , Fatores de Crescimento do Endotélio Vascular/metabolismo , Fatores de Virulência/metabolismo , Animais , Proteínas da Membrana Bacteriana Externa/química , Proteínas da Membrana Bacteriana Externa/genética , Bartonella/classificação , Bartonella/genética , Proliferação de Células , Perfilação da Expressão Gênica , Células Endoteliais da Veia Umbilical Humana/citologia , Células Endoteliais da Veia Umbilical Humana/metabolismo , Células Endoteliais da Veia Umbilical Humana/microbiologia , Humanos , Camundongos , Neovascularização Patológica/genética , Neovascularização Patológica/microbiologia , Domínios Proteicos , Sistemas de Secreção Tipo V/química , Sistemas de Secreção Tipo V/genética , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Fatores de Virulência/química , Fatores de Virulência/genética
15.
Proc Natl Acad Sci U S A ; 117(31): 18737-18743, 2020 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-32675245

RESUMO

The outer membrane (OM) of gram-negative bacteria confers innate resistance to toxins and antibiotics. Integral ß-barrel outer membrane proteins (OMPs) function to establish and maintain the selective permeability of the OM. OMPs are assembled into the OM by the ß-barrel assembly machine (BAM), which is composed of one OMP-BamA-and four lipoproteins-BamB, C, D, and E. BamB, C, and E can be removed individually with only minor effects on barrier function; however, depletion of either BamA or BamD causes a global defect in OMP assembly and results in cell death. We have identified a gain-of-function mutation, bamA E470K , that bypasses the requirement for BamD. Although bamD::kan bamA E470K cells exhibit growth and OM barrier defects, they assemble OMPs with surprising robustness. Our results demonstrate that BamD does not play a catalytic role in OMP assembly, but rather functions to regulate the activity of BamA.


Assuntos
Proteínas da Membrana Bacteriana Externa , Membrana Externa Bacteriana , Proteínas de Escherichia coli , Mutação com Ganho de Função/genética , Membrana Externa Bacteriana/química , Membrana Externa Bacteriana/metabolismo , Proteínas da Membrana Bacteriana Externa/química , Proteínas da Membrana Bacteriana Externa/genética , Proteínas da Membrana Bacteriana Externa/metabolismo , Escherichia coli/química , Escherichia coli/genética , Escherichia coli/metabolismo , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo
16.
BMC Infect Dis ; 20(1): 507, 2020 Jul 13.
Artigo em Inglês | MEDLINE | ID: mdl-32660436

RESUMO

BACKGROUND: Group A streptococcus (GAS) is an important human pathogen responsible for a broad range of infections. Epidemiological surveillance has been crucial to detect changes in the geographical and temporal variation of the disease pattern. The objective of this study was to investigate the molecular epidemiological characteristics and antimicrobial resistance of GAS isolates from patients in Children's Hospital in Beijing. METHODS: From 2016 to 2017, pharyngeal swab samples were collected from the outpatients in Children's Hospital, Capital Institute of Pediatrics, who were diagnosed with scarlet fever. Antimicrobial susceptibility test was performed according to the distribution of conventional antibiotics and Clinical and Laboratory Standards Institute (CLSI) recommendations. The distribution of the macrolide-resistance genes (ermB, ermA, mefA), emm (M protein-coding gene) typing, and superantigens (SAg) gene profiling were examined by polymerase chain reaction (PCR). RESULTS: A total of 297 GAS isolates were collected. The susceptibility of the isolates to penicillin, ceftriaxone, and levofloxacin was 100%. The resistance rate to erythromycin and clindamycin was 98.3 and 96.6%, respectively. The dominant emm types were emm12 (65.32%), emm1 (27.61%), emm75 (2.69%), and emm89 (1.35%). Of the 297 isolates, 290 (97.64%) carried the ermB gene, and 5 (1.68%) carried the mefA gene, while none carried the ermA gene. The most common superantigen genes identified from GAS isolates were smeZ (96.97%), speC (92.59%), speG (91.58%), ssa (85.52%), speI (54.55%), speH (52.19%), and speA (34.34%). Isolates with the genotype emm1 possessed speA, speC, speG, speJ, speM, ssa, and smeZ, while emm12 possessed speC, speG, speH, speI, speM, ssa, and smeZ superantigens. CONCLUSIONS: The prevalent strain of GAS isolates in Beijing has a high resistance rate to macrolides; however, penicillin can still be the preferred antibiotic for treatment. Erythromycin resistance was predominantly mediated by ermB. The common emm types were emm12 and emm1. There was a correlation between emm and the superantigen gene. Thus, long-term monitoring and investigation of the emm types and superantigen genes of GAS prevalence are imperative.


Assuntos
Antibacterianos/uso terapêutico , Farmacorresistência Bacteriana/genética , Penicilinas/uso terapêutico , Escarlatina/tratamento farmacológico , Escarlatina/epidemiologia , Streptococcus pyogenes/imunologia , Adolescente , Antígenos de Bactérias/genética , Proteínas da Membrana Bacteriana Externa/genética , Proteínas de Bactérias/genética , Pequim/epidemiologia , Reanimação Cardiopulmonar , Proteínas de Transporte/genética , Criança , Pré-Escolar , Eritromicina/uso terapêutico , Feminino , Hospitais Pediátricos , Humanos , Lactente , Masculino , Testes de Sensibilidade Microbiana , Escarlatina/microbiologia , Streptococcus pyogenes/isolamento & purificação , Superantígenos/genética
17.
Nat Chem Biol ; 16(9): 1019-1025, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32572278

RESUMO

The ß-barrel assembly machinery (BAM) inserts outer membrane ß-barrel proteins (OMPs) in the outer membrane of Gram-negative bacteria. In Enterobacteriacea, BAM also mediates export of the stress sensor lipoprotein RcsF to the cell surface by assembling RcsF-OMP complexes. Here, we report the crystal structure of the key BAM component BamA in complex with RcsF. BamA adopts an inward-open conformation, with the lateral gate to the membrane closed. RcsF is lodged deep within the lumen of the BamA barrel, binding regions proposed to undergo outward and lateral opening during OMP insertion. On the basis of our structural and biochemical data, we propose a push-and-pull model for RcsF export following conformational cycling of BamA, and provide a mechanistic explanation for how RcsF uses its interaction with BamA to detect envelope stress. Our data also suggest that the flux of incoming OMP substrates is involved in the control of BAM activity.


Assuntos
Proteínas da Membrana Bacteriana Externa/química , Proteínas da Membrana Bacteriana Externa/metabolismo , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/metabolismo , Escherichia coli/química , Escherichia coli/metabolismo , Proteínas da Membrana Bacteriana Externa/genética , Cristalografia por Raios X , Escherichia coli/genética , Proteínas de Escherichia coli/genética , Bicamadas Lipídicas/química , Bicamadas Lipídicas/metabolismo , Modelos Moleculares , Conformação Proteica
18.
PLoS One ; 15(6): e0234306, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32555615

RESUMO

Moraxella catarrhalis is a human-adapted, opportunistic bacterial pathogen of the respiratory mucosa. Although asymptomatic colonization of the nasopharynx is common, M. catarrhalis can ascend into the middle ear, where it is a prevalent causative agent of otitis media in children, or enter the lower respiratory tract, where it is associated with acute exacerbations of chronic obstructive pulmonary disease in adults. Phase variation is the high frequency, random, reversible switching of gene expression that allows bacteria to adapt to different host microenvironments and evade host defences, and is most commonly mediated by simple DNA sequence repeats. Bioinformatic analysis of five closed M. catarrhalis genomes identified 17 unique simple DNA sequence repeat tracts that were variable between strains, indicating the potential to mediate phase variable expression of the associated genes. Assays designed to assess simple sequence repeat variation under conditions mimicking host infection demonstrated that phase variation of uspA1 (ubiquitous surface protein A1) from high to low expression occurs over 72 hours of biofilm passage, while phase variation of uspA2 (ubiquitous surface protein A2) to high expression variants occurs during repeated exposure to human serum, as measured by mRNA levels. We also identify and confirm the variable expression of two novel phase variable genes encoding a Type III DNA methyltransferase (modO), and a conserved hypothetical permease (MC25239_RS00020). These data reveal the repertoire of phase variable genes mediated by simple sequence repeats in M. catarrhalis and demonstrate that modulation of expression under conditions mimicking human infection is attributed to changes in simple sequence repeat length.


Assuntos
Regulação Bacteriana da Expressão Gênica/genética , Moraxella catarrhalis/genética , Aderência Bacteriana/genética , Proteínas da Membrana Bacteriana Externa/genética , Proteínas da Membrana Bacteriana Externa/metabolismo , Expressão Gênica/genética , Humanos , Repetições de Microssatélites/genética , Moraxella catarrhalis/patogenicidade , Infecções por Moraxellaceae , Otite Média/microbiologia , Sequências Repetitivas de Ácido Nucleico/genética
19.
Eur J Clin Microbiol Infect Dis ; 39(10): 1821-1830, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32557327

RESUMO

Helicobacter pylori (H. pylori) infection is associated with some gastric diseases, such as gastritis, peptic ulcer, and gastric cancer. CagA and VacA are known virulence factors of H. pylori, which play a vital role in severe clinical outcomes. Additionally, the expression of outer membrane proteins (OMPs) helps H. pylori attach to gastric epithelial cells at the primary stage and increases the virulence of H. pylori. In this review, we have summarized the paralogs of H. pylori OMPs, their genomic loci, and the different receptors of OMPs identified so far. We focused on five OMPs, BabA (HopS), SabA (HopP), OipA (HopH), HopQ, and HopZ, and one family of OMPs: Hom. We highlight the coexpression of OMPs with other virulence factors and their relationship with clinical outcomes. In conclusion, OMPs are closely related to the pathogenic processes of adhesion, colonization, persistent infection, and severe clinical consequences. They are potential targets for the prevention and treatment of H. pylori-related diseases.


Assuntos
Proteínas da Membrana Bacteriana Externa/genética , Proteínas da Membrana Bacteriana Externa/metabolismo , Infecções por Helicobacter/microbiologia , Helicobacter pylori/genética , Helicobacter pylori/patogenicidade , Fatores de Virulência/metabolismo , Células Epiteliais/microbiologia , Expressão Gênica , Humanos , Fatores de Virulência/genética
20.
Int J Infect Dis ; 98: 305-314, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32562850

RESUMO

OBJECTIVE: Scarlet fever epidemics caused by group A Streptococcus (GAS) have been ongoing in China since 2011. However, limited data are available on the dynamic molecular characterizations of the epidemic strains. METHOD: Epidemiological data of scarlet fever in Shanghai were obtained from the National Notifiable Infectious Disease Surveillance System. Throat swabs of patients with scarlet fever and asymptomatic school-age children were cultured. Illumina sequencing was performed on 39emm1 isolates. RESULTS: The annual incidence of scarlet fever was 7.5-19.4/100,000 persons in Shanghai during 2011-2015, with an average GAS carriage rate being 7.6% in school-age children. The proportion ofemm1 GAS strains increased from 3.8% in 2011 to 48.6% in 2014; they harbored a superantigen profile similar to emm12 isolates, except for the speA gene. Two predominant clones, SH001-emm12, and SH002-emm1, circulated in 66.9% of scarlet fever cases and 44.8% of carriers. Genomic analysis showed emm1 isolates throughout China constituted distinct clades, enriched by the presence of mobile genetic elements carrying the multidrug-resistant determinants ermB and tetM and virulence genes speA, speC, and spd1. CONCLUSION: A significant increase in the proportion ofemm1 strains occurred in the GAS population, causing scarlet fever in China. Ongoing surveillance is warranted to monitor the dynamic changes of GAS clones.


Assuntos
Antígenos de Bactérias/genética , Proteínas da Membrana Bacteriana Externa/genética , Proteínas de Transporte/genética , Escarlatina/microbiologia , Streptococcus pyogenes/isolamento & purificação , Adolescente , Antígenos de Bactérias/metabolismo , Proteínas da Membrana Bacteriana Externa/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Proteínas de Transporte/metabolismo , Criança , Pré-Escolar , China/epidemiologia , Exotoxinas/genética , Exotoxinas/metabolismo , Feminino , Humanos , Incidência , Masculino , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Escarlatina/diagnóstico , Escarlatina/epidemiologia , Streptococcus pyogenes/classificação , Streptococcus pyogenes/genética , Streptococcus pyogenes/metabolismo
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