The virulence regulator AbsR in avian pathogenic Escherichia coli has pleiotropic effects on bacterial physiology

Avian pathogenic Escherichia coli(APEC)belonging to extraintestinal pathogenic E.coli(ExPEC)can cause severe infections in extraintestinal tissues in birds and humans,such as the lungs and blood.MprA(microcin production regulation,locus A,herein renamed AbsR,a blood survival regulator),a member of the MarR(multiple antibiotic resistance regulator)transcriptional regulator family,governs the expression of capsule biosynthetic genes in human ExPEC and represents a promising druggable target for antimicrobials.However,a deep understanding of the AbsR regulatory mechanism as well as its regulon is lacking.In this study,we present a systems-level analysis of the APEC AbsR regulon using ChIP-Seq(chromatin immunoprecipitation sequencing)and RNA-Seq(RNA sequencing)methods.We found that AbsR directly regulates 99 genes and indirectly regulates 667 genes.Furthermore,we showed that:1)AbsR contributes to antiphagocytotic effects by macrophages and virulence in a mouse model for systemic infection by directly activating the capsular gene cluster;2)AbsR positively impacts biofilm formation via direct regulation of the T2SS(type II secretion system)but plays a marginal role in virulence;and 3)AbsR directly upregulates the acid tolerance signaling system EvgAS to withstand acid stress but is dispensable in ExPEC virulence.Finally,our data indicate that the role of AbsR in virulence gene regulation is relatively conserved in ExPEC strains.Altogether,this study provides a comprehensive analysis of the AbsR regulon and regulatory mechanism,and our data suggest that AbsR likely influences virulence primarily through the control of capsule production.Interestingly,we found that AbsR severely represses the expression of the type I-F CRISPR(clustered regularly interspaced short palindromic repeats)-Cas(CRISPR associated)systems,which could have implications in CRISPR biology and application.

Distribution of Virulence-Associated Genes of Avian Pathogenic Escherichia coli Isolates in China

216 avian pathogenic Escherichia coli （APEC） isolates were obtained from poultry with colibacillosis in different areas of China. Among them, 195 were serotyped as 078, 088, and 093. Thirteen virulence-associated genes, including fimC, iucD, iss, tsh, fyuA, irp2, eaeA, hlyE, colV, papC, stx2f, vat, and astA, were submitted to PCR amplification. The fimC gene was the most prevalent with a detection rate of 93.6%, followed by iucD （70.8%）, iss （58.8%）, and tsh （51.4%） in APEC isolates. The detection rate of high pathogenicity islands （HPI）-associatedfyuA and irp2 genes were both 44.9%, with no LEE （the locus of enterocyte effacement） island-associated gene eaeA detected. In terms of distribution patterns of the 13 virulence-associated genes, 5 isolates harborbed 10 genes, 19 isolates contained onlyfimC gene, and only 4 isolates had no virulence-associated gene detected. Different correlations of the virulence-associated genes with O serotypes were also investigated and 50% 078 isolates had a gene distribution patterns of fimC^＋iucD^＋irp2^＋fyuA^＋iss^＋colV^＋tsh^＋.

Fosfomycin Resistance in Avian Pathogenic Escherichia coli Isolates

Fosfomycin, a broad-spectrum antibiotic against both Gram-positive and Gram-negative bacteria, is very important in the clinic but many fosfomycin-resistant bacteria have been isolated from patients. In this study, the resistance mechanism of three fosfomycin-resistant avian pathogenic Escherichia coli （APEC） strains （JE1, IF7 and CD11） isolated from septicemic chickens were analyzed. The results showed that their fosfomycin-resistance mechanisms were different. An alteration in the glpT transport system was the main reason of the fosfomycin-resistance mechanisms of strain IF7. Compared with the control stain BL21, the capacity of fosfomycin-uptake was low in all these three stains （JE 1 〉IFT〉CD 11）. Sequence results of murA showed that there were more than 10 sites of nucleotide mutation, but only one amino acid mutation T116A showed in CD11. Real-time detection test showed that the expression level of the murA gene of the three stains was significantly increased （four times increase in strain CD11 and two times increase in strains JE1 and IFT）. The transformation and recombinant test showed that the recombinant bacteria with the murA of JE1 and CD11 showed high minimal inhibitory concentration （MIC） against fosfomycin. From the results of this research, it showed that most of the fosfomycin- resistance mechanisms once showed in patient bacteria have appeared in the APEC strains and the fosfomycin-resistance mechanism of the three APEC isolates was different.

Dietary coated essential oil and organic acid mixture supplementation improves health of broilers infected with avian pathogenic Escherichia coli

Colibacillosis caused by avian pathogenic Escherichia coli(APEC)is a very prevalent disease in poultry farms in China.The exploration of effective non-antibiotic substances is of great significance for the control of APEC infections.This experiment evaluated the efficacy of coated essential oil and organic acid(EOA)supplementation to prevent E.coli O78 infection in broiler chickens.A total of 288 one-day-old male broiler chicks were randomly distributed into 4 groups with 6 replicates per group.Chickens were fed a diet either supplemented with EOA(500 mg/kg feed)or not,and either uninfected or infected with E.coli O78 intratracheally.Results showed that E.coli O78 infection reduced body weight gain,increased mortality and the ratio of feed to gain along with cecal and liver E.coli load,damaged gut mucosa,induced local and systemic inflammation,and altered cecal microbial composition,diversity and function(P<0.05).Supplemental EOA improved feed conversion efficiency,lowered gross lesion scores and cecal E.coli population,enhanced intestinal goblet cells and serum IgG concentration,and tended to decrease serum IL-12 production(P<0.05).Essential oil and organic acid addition downregulated IFN-γmRNA,tended to decrease mucin-2 mRNA levels while upregulating IL-10 mRNA,and tended to increase ZO-1 gene expression in the jejuna of infected birds at 7 d after E.coli O78 challenge(P<0.05).The 16S rRNA gene sequencing indicated that both EOA addition and E.coli O78 challenge altered the diversity and composition of the cecal microbiota community.Furthermore,infected birds fed EOA showed decreased Bacteroidetes and genus Lactobacillus abundance compared with the infected control.LEfSe analysis showed that Firmicutes,Ruminococcaceae,Clostridiales,Clostridia,Lactobacillus,Lactobacilaceae,and cc-115 were enriched in the non-infected but EOA-treated group(P<0.05).Collectively,dietary EOA supplementation could mildly alleviate E.coli-induced gut injury and inflammation.

生物膜、毒力基因和耐药基因对APEC的致病性作用

禽致病性大肠杆菌病(Avian Pathogenic Escherichia coli,APEC)是危害养禽业发展的主要细菌性病原之一,给养禽业的发展带来了严重危害。为研究APEC的致病性与耐药性之间的关系,从毒力基因的分布、生物膜的形成能力、耐药基因的分布等三个方面探讨了生物膜的形成能力与毒力基因、耐药基因在致病性中的作用,为后期研究禽大肠杆菌的致病机理及耐药机制等提供理论基础,并为开展禽大肠杆菌病的流行病学调查及监测、防控禽大肠杆菌病的流行提供理论数据。