LuxS contributes to virulence in avian pathogenic Escherichia coli O78:K80:H9.

Avian pathogenic Escherichia coli (APEC) cause avian colibacillosis, a poultry disease characterized by multiple organ infections resulting in significant economic loss in the poultry industry. Several virulence factors are important for disease manifestation in APEC of which, role of quorum sensing has not been investigated. Quorum sensing is a population dependent cell-cell signaling system which modulates numerous physiological processes such as biofilm formation and virulence in multiple species. LuxS, a well-known controller in the QS, plays a role in regulating virulence in various bacterial species. Here we investigated the role of LuxS in regulating virulence in APEC O78:K80:H9. Mutation of luxS resulted in a significant reduction of virulence in APEC O78:K80:H9, evidenced by both in vivo and in vitro assays such as decreased invasion of internal organs in chicken embryo, reduced lethality in chicken embryo lethality assay, and altered lipopolysaccharide (LPS) profile. In addition, the abilities of the knockout strain to survive in chicken macrophage cell lines and to invade in chicken embryo fibroblast cells were significantly diminished. Further, structure and expression level of the LPS profile was significantly altered in the knockout strain, which may be one of the contributing factors for the persistence and virulence of APEC. Complementation of luxS gene in trans restored the virulence of the knockout strain to the level of wild-type bacteria. Taken together, these results show that LuxS contributes to the virulence in APEC O78:K80:H9 strain and partly explain the role played by LuxS in the pathogenesis of APEC strains.

Palaniyandi S ，Mitra A ，Herren CD ，Zhu X ，Mukhopadhyay S ... -  《-》

The luxS gene functions in the pathogenesis of avian pathogenic Escherichia coli.

Avian pathogenic Escherichia coli (APEC) causes avian colibacillosis, the most significant infectious bacterial disease of poultry worldwide. LuxS, the product of the luxS gene, mediates the quorum sensing (QS) mechanism. This involves the production of autoinducer-2 (AI-2), which regulates important physiological traits and a variety of adaptive processes in different bacteria. In this study, a luxS gene deleted APEC mutant strain, ΔDE17, was constructed using strain DE17. Analysis of bioluminescence indicated that deletion of the luxS gene abolished the production of the QS signal AI-2 in the bacteria. Further studies showed that deletion of the luxS gene in DE17 reduced the bacterial virulence by 31.5-fold in ducklings, based on the measurement of the 50% lethal dose. The mutant strain reduced significantly the abilities of adherence and invasion, by 50.0% and 40.7% respectively, compared with the wild strain DE17. The mutant strain also showed reduced survival in vivo: the bacterial loads of the mutant strain in infected liver, spleen and blood were 46.4-fold, 5.2-fold, and 3.7-fold reduced, respectively, compared with the wild-type strain DE17. Real-time polymerase chain reaction (PCR) demonstrated further that the mRNA levels of the virulence-related genes iucD, fyuA, vat, ompA, iss, fimC and tsh were significantly decreased in the mutant strain ΔDE17, when compared with DE17 (p < 0.05). In addition, the deletion of the luxS gene reduced the motility of the bacterium. This study suggests that the luxS gene functions in the pathogenesis of diseases caused by avian pathogenic E. coli.

Han X ，Bai H ，Liu L ，Dong H ，Liu R ，Song J ，Ding C ，Qi K ，Liu H ，Yu S ... -  《-》

Andrographolide interferes quorum sensing to reduce cell damage caused by avian pathogenic Escherichia coli.

Avian pathogenic Escherichia coli (APEC) induce septicemia in chickens by invading type II pneumocytes to breach the blood-air barrier. The virulence of APEC can be regulated by quorum sensing (QS). Andrographolide is a QS inhibitor of Pseudomonas aeruginosa (P. aeruginosa). Therefore, we investigate whether andrographolide inhibits the injury of chicken type II pneumocytes by avian pathogenic E. coli O78 (APEC-O78) by disrupting the bacterial QS system. The results showed that sub-MIC of andrographolide significantly reduced the release of lactate dehydrogenase (LDH), F-actin cytoskeleton polymerization, and the degree of the adherence to chicken type II pneumocytes induced by APEC-O78. Further, we found that andrographolide significantly decreased the autoinducer-2 (AI-2) activity and the expression of virulence factors of APEC-O78. These results suggest that andrographolide reduce the pathogenicity of APEC-O78 in chicken type II pneumocytes by interfering QS and decreasing virulence. These results provide new evidence for colibacillosis prevention methods in chickens.

Guo X ，Zhang LY ，Wu SC ，Xia F ，Fu YX ，Wu YL ，Leng CQ ，Yi PF ，Shen HQ ，Wei XB ，Fu BD ... -  《-》

Different activated methyl cycle pathways affect the pathogenicity of avian pathogenic Escherichia coli.

The activated methyl cycle (AMC) regulates the cellular levels of S-adenosyl-l-homocysteine (SAH) in bacteria, which plays a crucial role in bacterial pathogenicity. There are two AMC pathways in bacteria: one is a two-step reaction pathway (named the LuxS/Pfs pathway) in which LuxS and Pfs catalyze the conversion of SAH to l-homocysteine and autoinducer-2 (AI-2), and the other is a one-step reaction (named the SahH pathway) mediated by S-adenosyl-l-homocysteine hydrolase (SahH), which completes this cycle without producing AI-2. In this study, we evaluated the effects of different AMC pathways on the pathogenicity of avian pathogenic Escherichia coli (APEC). The plasmid pSTV-sahH (containing the sahH gene of Pseudomonas aeruginosa) was transformed into the wild-type APEC strain DE17 (containing the LuxS/Pfs pathway) and the pfs mutant strain DE17Δpfs, which lacks the LuxS/Pfs pathway, to create the strains SahH-DE17Δpfs (containing the SahH pathway) and SahH-DE17 (containing the LuxS/Pfs and SahH pathways). The results showed that the different AMC pathways had different effects on the growth rate, AI-2 activity, and motility in APEC. Furthermore, we showed that the 50% lethal doses of the DE17Δpfs and SahH-DE17Δpfs strains were reduced by 650-fold and 52-fold, respectively, in ducklings, compared with that of the DE17 strain. The DE17Δpfs strain exhibited significantly reduced adherence and invasion (p<0.01). In addition, the DE17Δpfs and SahH-DE17Δpfs strains also showed reduced survival in vivo, as evidenced by significant (p<0.01) reductions in their bacterial loads in infected liver, spleen, kidney, and blood. This study suggests that different AMC pathways affect the pathogenesis of APEC.

Xu D ，Zuo J ，Chen Z ，Lv X ，Hu J ，Wu X ，Qi K ，Mi R ，Huang Y ，Miao J ，Jiang W ，Wang S ，Wang C ，Han X ... -  《-》

SodA Contributes to the Virulence of Avian Pathogenic Escherichia coli O2 Strain E058 in Experimentally Infected Chickens.

Strains of avian pathogenic Escherichia coli (APEC), the common pathogen of avian colibacillosis, encounter reactive oxygen species (ROS) during the infection process. Superoxide dismutases (SODs), acting as antioxidant factors, can protect against ROS-mediated host defenses. Our previous reports showed that the sodA gene (encoding a Mn-cofactor-containing SOD [MnSOD]) is highly expressed during the septicemic infection process of APEC. sodA has been proven to be a virulence factor of certain pathogens, but its role in the pathogenicity of APEC has not been fully identified. In this study, we deleted the sodA gene from the virulent APEC O2 strain E058 and examined the in vitro and in vivo phenotypes of the mutant. The sodA mutant was more sensitive to hydrogen peroxide in terms of both its growth and viability than was the wild type. The ability to form a biofilm was weakened in the sodA mutant. The sodA mutant was significantly more easily phagocytosed by chicken macrophages than was the wild-type strain. Chicken infection assays revealed significantly attenuated virulence of the sodA mutant compared with the wild type at 24 h postinfection. The virulence phenotype was restored by complementation of the sodA gene. Quantitative real-time reverse transcription-PCR revealed that the inactivation of sodA reduced the expression of oxidative stress response genes katE, perR, and osmC but did not affect the expression of sodB and sodC Taken together, our studies indicate that SodA is important for oxidative resistance and virulence of APEC E058.IMPORTANCE Avian colibacillosis, caused by strains of avian pathogenic Escherichia coli, is a major bacterial disease of severe economic significance to the poultry industry worldwide. The virulence mechanisms of APEC are not completely understood. This study investigated the influence of an antioxidant protein, SodA, on the phenotype and pathogenicity of APEC O2 strain E058. This is the first report demonstrating that SodA plays an important role in protecting a specific APEC strain against hydrogen peroxide-induced oxidative stress and contributes to the virulence of this pathotype strain. Identification of this virulence factor will enhance our knowledge of APEC pathogenic mechanisms, which is crucial for designing successful strategies against associated infections and transmission.

Gao Q ，Xia L ，Wang X ，Ye Z ，Liu J ，Gao S ... -  《-》