RESUMO
The type 3 secretion system (T3SS) is essential for bacterial virulence through delivering effector proteins directly into the host cytosol. Here, we identified an alternative delivery mechanism of virulence factors mediated by the T3SS, which consists of the association of extracellularly secreted proteins from bacteria with the T3SS to gain access to the host cytosol. Both EspC, a protein secreted as an enteropathogenic Escherichia coli (EPEC) autotransporter, and YopH, a protein detected on the surface of Yersinia, require a functional T3SS for host cell internalization; here we provide biophysical and molecular evidence to support the concept of the EspC translocation mechanism, which requires (i) an interaction between EspA and an EspC middle segment, (ii) an EspC translocation motif (21 residues that are shared with the YopH translocation motif), (iii) increases in the association and dissociation rates of EspC mediated by EspA interacting with EspD, and (iv) an interaction of EspC with the EspD/EspB translocon pore. Interestingly, this novel mechanism does not exclude the injection model (i.e., EspF) operating through the T3SS conduit; therefore, T3SS can be functioning as an internal conduit or as an external railway, which can be used to reach the translocator pore, and this mechanism appears to be conserved among different T3SS-dependent pathogens.IMPORTANCE The type 3 secretion system is essential for injection of virulence factors, which are delivered directly into the cytosol of the host cells for usurping and subverting host processes. Recent studies have shown that these effectors proteins indeed travel inside an "injectisome" conduit through a single step of translocation by connecting the bacterium and host cell cytoplasms. However, all findings are not compatible with this model. For example, both YopH, a protein detected on the surface of Yersinia, and EspC, an autotransporter protein secreted by enteropathogenic E. coli, require a functional T3SS for host cell translocation. Both proteins have an intermediate extracellular step before their T3SS-dependent translocation. Here, we show an alternative delivery mechanism for these extracellularly secreted virulence factors that are then incorporated into the T3SS to enter the cells; this novel mechanism coexists with but diverges from the canonical injection model that involves the passage of the protein inside the injectisome.
Assuntos
Proteínas da Membrana Bacteriana Externa/metabolismo , Proteínas de Escherichia coli/metabolismo , Escherichia coli/metabolismo , Proteínas Tirosina Fosfatases/metabolismo , Sistemas de Secreção Tipo III/metabolismo , Yersinia/metabolismo , Transporte ProteicoRESUMO
Biodegradation of tributyltin (TBT) by four tin resistant Gram negative bacteria isolated from extremely contaminated river sediments in the Atacama Desert in Chile was studied. Moraxella osloensis showed the greatest resistance and degradation capability of TBT, producing less toxic by-products, such as dibutyltin (DBT) and inorganic tin. In 7 days, approximately 80 % of TBT degradation was achieved, generating close to 20 % of DBT as degradation product. The degradation rate constant (k) was 0.022 [day(-1)] and TBT half-life (t1/2) in culture was 4.3 days. Debutylation is stated a probable mechanism of TBT degradation.
Assuntos
Bactérias/metabolismo , Clima Desértico , Compostos Orgânicos de Estanho/análise , Estanho/análise , Compostos de Trialquitina/análise , Alcaligenes/metabolismo , Biodegradação Ambiental , Burkholderia cepacia/metabolismo , Chile , Farmacorresistência Bacteriana , Sedimentos Geológicos/química , Meia-Vida , Moraxella/metabolismo , Pseudomonas/metabolismo , Rios , Solo , Microbiologia do Solo , Poluentes do Solo/análise , Fatores de Tempo , Poluentes da Água/análise , Yersinia/metabolismoRESUMO
The ability of three psychrotrophic Gram-negative bacilli isolated from Chilean Patagonian cold freshwater rivers to produce bioactive metabolites was evaluated. The strains were isolated from cold waters rivers and identified by their biochemical properties and 16S rRNA gene analysis. The metabolites fractions showing antibacterial activity were obtained by solvent extraction and partially characterized by gas-mass chromatography (GC-MS). Antibacterial activity of the fractions was evaluated by an agar-well diffusion test upon 14 bacterial strains, both Gram positive and Gram negative. Thermal and proteolytic resistances of the antibacterial metabolites fractions were also evaluated. Molecular analysis allows the identification of the three Patagonian strains as Pseudomonas sp. RG-6 (Pseudomonas brenneri 99.6 % identity), Pseudomonas sp. RG-8 (Pseudomonas trivialis 99.6 % identity) and Yersinia sp. RP-3 (Yersinia aldovae 99.5 % identity). These extracts were able to inhibit both Gram-positive and Gram-negative bacteria but not Listeria monocytogenes. The antibacterial activity of the filtrated supernatants was lost at temperatures ≥60 °C, and was not affected by proteinase K treatment. The chemical structure of the active molecule remains to be elucidated, although the GC-MS analysis of the filtrates suggests that compounds like sesquiterpenes derivatives from ß-maaliene or δ-selinene could be responsible of this antibacterial activity. Pristine cold freshwater streams showed to be interesting sources of metabolites-producing microorganisms with antibacterial activity.
Assuntos
Antibacterianos/isolamento & purificação , Água Doce/microbiologia , Pseudomonas/isolamento & purificação , Pseudomonas/metabolismo , Yersinia/isolamento & purificação , Yersinia/metabolismo , Antibacterianos/farmacologia , Chile , Análise por Conglomerados , Temperatura Baixa , DNA Bacteriano/química , DNA Bacteriano/genética , DNA Ribossômico/química , DNA Ribossômico/genética , Cromatografia Gasosa-Espectrometria de Massas , Listeria monocytogenes/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Filogenia , Pseudomonas/classificação , Pseudomonas/genética , RNA Ribossômico 16S/genética , Análise de Sequência de DNA , Temperatura , Yersinia/classificação , Yersinia/genéticaRESUMO
The bacterial genus Yersinia belongs to the family Enterobacteriaceae and comprises 15 species. Species of the genus Yersinia are usually identified by their phenotypic characteristics. Thus, it is essential to establish a complete phenotypic classification for all species of the genus Yersinia. The species Yersinia massiliensis was proposed in 2008, based on 16S rRNA, gyrB, hsp60, rpoB and sodA gene sequences and some distinguishing phenotypic characteristics. In this study, four Yersinia strains classified as Y. massiliensis based on the sequencing of the loci mentioned above were subjected to a more detailed phenotypic characterization. This characterization revealed differences in the results of four tests previously reported as diagnostic for Y. massiliensis and the results of 18 additional tests provided new information about the biochemical diversity of this species. In the light of the results of the phenotypic characteristics of the four strains of Y. massiliensis, an emended description of Y. massiliensis is presented.