N-Acyl-homoserine Lactones from Enterobacter sakazakii (Cronobacter spp.) and Their Degradation by Bacillus cereus Enzymes

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)A chemical study of acyl-homoserine lactones (acyl-HSLs) produced by Enterobacter sakazakii resulted in the identification of three molecules: (S)-N-heptanoyl-HSL, (S)-N-dodecanoyl-HSL and (S)-N-tetradecanoyl-HSL. Mixed cultures of E. sakazakii and Bacillus cereus depleted E. sakazakii acyl-HSLs, suggesting acyl-HSL degradation by B. cereus hydrolases (hydrolysis of the lactone or amide moiety). The expression of B. cereus acyl-HSL lactonase and acyl-homoserine acylase was confirmed by monitoring the biotransformation of (S)-N-dodecanoyl-HSL into (S)-N-dodecanoyl-homoserine, dodecanoic acid and homoserine in the presence of B. cereus whole cells, using electrospray-mass spectrometry (ESI-MS).602585592Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP

Possible involvement of an outer membrane protein in the pathogenicity of a chicken septicemic Escherichia coli isolate

A mutant variant of a septicemic Escherichia coli strain (L3) isolated from an outbreak in chickens was constructed by the insertion of TnphoA transposon. Seven mutant derivatives were analyzed regarding the pathogenicity. Two of them (XP2, XP4) were less pathogenic in the one-day-old chick pathogenicity assay. The expression of several outer membrane proteins of mutant XP2 strain was suppressed, and strain XP4 had a 47.8(kDa) protein that was not expressed. None of these proteins was correlated to the iron-acquisition system. Mutant XP2 could have suppression of a regulatory protein responsible for the expression of other proteins not related to pathogenicity but important for the rapid bacterial growth, while mutant XP4 did not express a 47.8(kDa) protein. We propose that the 47.8(kDa) protein could be associated to the pathogenicity process of Escherichia coli strains responsible for septicemia in poultry.29211311

Characterization of IcmF of the type VI secretion system in an avian pathogenic Escherichia coli (APEC) strain

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)The intracellular multiplication factor (IcmF) protein is a component of the recently described type VI secretion system (T6SS). IcmF has been shown to be required for intra-macrophage replication and inhibition of phagosome-lysosome fusion in Legionella pneumophila. In Vibrio cholerae it is involved in motility, adherence and conjugation. Given that we previously reported that two T6SS genes (hcp and clpV) contribute to the pathogenesis of a septicaemic strain (SEPT362) of avian pathogenic Escherichia coli (APEC), we investigated the function of IcmF in this strain. Further elucidation of the virulence mechanisms of APEC is important because this pathogen is responsible for financial losses in the poultry industry, and is closely related to human extraintestinal pathogenic E. coli(ExPEC) strains, representing a potential zoonotic risk, as well as serving as a reservoir of virulence genes. Here we show that an APEC icmF mutant has decreased adherence to and invasion of epithelial cells, as well as decreased intra-macrophage survival. The icmF mutant is also defective for biofilm formation on abiotic surfaces. Additionally, expression of the flagella operon is decreased in the icmF mutant, leading to decreased motility. The combination of these phenotypes culminates in this mutant being altered for infection in chicks. These results suggest that IcmF in APEC may play a role in disease, and potentially also in the epidemiological spread of this pathogen through enhancement of biofilm formation.1571029542962NIH [AI053067]Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)NIH [AI053067]FAPESP [2008/56739-6, 07/50432-3]CAPES [4062086]CAPES [041/2009

Rck of Salmonella enterica, subspecies enterica serovar Enteritidis, mediates Zipper-like internalization

International audienceSalmonella can invade non-phagocytic cells through its type III secretion system (T3SS-1), which induces a Trigger entry process. This study showed that Salmonella enterica, subspecies enterica serovar Enteritidis can also invade cells via the Rck outer membrane protein. Rck was necessary and sufficient to enable non-invasive E. coli and Rck-coated beads to adhere to and invade different cells. Internalization analysis of latex beads coated with different Rck peptides showed that the peptide containing amino acids 140-150 promoted adhesion, whereas amino acids between 150 and 159 modulated invasion. Expression of dominant-negative derivatives and use of specific inhibitors demonstrated the crucial role of small GTPases Rac1 and Cdc42 in activating the Arp2/3 complex to trigger formation of actin-rich accumulation, leading to Rck-dependent internalization. Finally, scanning and transmission electron microscopy with Rck-coated beads and E. coli expressing Rck revealed microvillus-like extensions that formed a Zipper-like structure, engulfing the adherent beads and bacteria. Overall, our results provide new insights into the Salmonella T3SS-independent invasion mechanisms and strongly suggest that Rck induces a Zipper-like entry mechanism. Consequently, Salmonella seems to be the first bacterium found to be able to induce both Zipper and Trigger mechanisms to invade host cells

Quorum sensing coordinates brute force and stealth modes of infection in the plant pathogen Pectobacterium atrosepticum

Quorum sensing (QS) in vitro controls production of plant cell wall degrading enzymes (PCWDEs) and other virulence factors in the soft rotting enterobacterial plant pathogen Pectobacterium atrosepticum (Pba). Here, we demonstrate the genome-wide regulatory role of QS in vivo during the Pba–potato interaction, using a Pba-specific microarray. We show that 26% of the Pba genome exhibited differential transcription in a QS (expI-) mutant, compared to the wild-type, suggesting that QS may make a greater contribution to pathogenesis than previously thought. We identify novel components of the QS regulon, including the Type I and II secretion systems, which are involved in the secretion of PCWDEs; a novel Type VI secretion system (T6SS) and its predicted substrates Hcp and VgrG; more than 70 known or putative regulators, some of which have been demonstrated to control pathogenesis and, remarkably, the Type III secretion system and associated effector proteins, and coronafacoyl-amide conjugates, both of which play roles in the manipulation of plant defences. We show that the T6SS and a novel potential regulator, VirS, are required for full virulence in Pba, and propose a model placing QS at the apex of a regulatory hierarchy controlling the later stages of disease progression in Pba. Our findings indicate that QS is a master regulator of phytopathogenesis, controlling multiple other regulators that, in turn, co-ordinately regulate genes associated with manipulation of host defences in concert with the destructive arsenal of PCWDEs that manifest the soft rot disease phenotype