Effects of ibeA Deletion on Virulence and Biofilm Formation of Avian Pathogenic Escherichia coli

Abstract

The ibeA gene is located on a genomic island, GimA, which is involved in the pathogenesis of neonatal meningitis Escherichia coli (NMEC) and avian pathogenic E. coli (APEC). The prevalence of ibeA in the APEC collection in China was investigated, and 20 of 467 strains (4.3%) were positive. In addition, analysis of the association of the E. coli reference (ECOR) groups with positive strains revealed that ibeA was linked to group B2. The ibeA gene in DE205B was analyzed and compared to those of APEC and NMEC, which indicated that the specificity of ibeA was not consistent along pathotypes. The invasion of chicken embryo fibroblast DF-1 cells by APEC DE205B and RS218 was observed, which suggested that DF-1 cells could be a model to study the mechanism of APEC invasion. The inactivation of ibeA in APEC DE205B led to the reduced capacity to invade DF-1 cells, defective virulence in vivo, and decreased biofilm formation compared to the wild-type strain. In addition, strain AAEC189 expressing ibeA exhibited enhanced invasion capacity and biofilm formation. The results of the quantitative real-time reverse transcription-PCR (qRT-PCR) analysis and animal system infection experiments indicated that the loss of ibeA decreased the colonization and proliferation capacities of APEC in the brain during system infection.Escherichia coli typically colonizes the mammalian and avian gastrointestinal tract and other mucosal surfaces. While many of these strains are commensal, certain pathogenic strains can cause severe diseases (33). Extraintestinal pathogenic E. coli (ExPEC) is a group of strains that have been implicated in a large range of infections in humans and animals, such as neonatal meningitis, urinary tract infections (UTIs), pneumonia, osteomyelitis, and septicemia (14,16,30,35). Among these, typical infections caused by ExPEC in humans are UTIs and neonatal meningitis (5). Similarly, systemic infections caused by avian pathogenic E. coli (APEC) are economically devastating to poultry industries (14, 16). APEC enters and colonizes the avian respiratory tract by inhalation of fecal dust, leading to localized infections, such as airsacculitis and pneumonia. In certain cases, they spread into various internal organs, typically causing pericarditis, perihepatitis, peritonitis, salpingitis, and other extraintestinal diseases. Systemic infection of poultry is characterized in its acute form by septicemia, commonly resulting in sudden death (14,16,52).Previous studies showed that certain subsets of ExPEC strains isolated from different host organisms are highly similar (17,31,44,51), thus increasing the need to study their zoonotic potential. Virulence determinants common to uropathogenic E. coli (UPEC), APEC, neonatal meningitis Escherichia coli (NMEC), or septicemia-associated E. coli (SEPEC), such as the aerobactin iron transport system, the K1 capsule, and type 1 and P fimbriae, have been identified (2,14,19,21,43,47,59). Furthermore, the function of the K1 capsule was similar in virulence to those of APEC and NMEC. The K1...

Characterization and Functional Analysis of AatB, a Novel Autotransporter Adhesin and Virulence Factor of Avian Pathogenic Escherichia coli

“…The APEC strain DE205B was isolated from the brain of a duck with septicemia and neurological symptoms and belonged to the phylogenetic E. coli reference (ECOR) group B2, as determined by phylogenetic analysis using multiplex PCR (37,38). This strain harbored the virulence-associated genes tsh, mat, fyuA, irp2, iucD, iutA, iss, aatA, aatB, vat, malX, fimC, ompA, ibeA, ibeB, yijp, gimB, and aslA but was negative for papC and hlyA by PCR analysis (38)(39)(40). Strain DE205B was used for infection studies, mutant construction, and functional assays.…”

“…For immunoblot analysis, protein samples were loaded into 12% SDS gels and transferred onto a polyvinylidene fluoride membrane (Amersham Pharmacia Biotech, Uppsala, Sweden) using a semidry blotting apparatus (TE77; Amersham Pharmacia Biotech), as described previously (23,38,39). Anti-AatB and anti-DE205B immune sera were used as primary antibodies.…”

“…The resulting plasmid, pET28a-aatB, was transformed into competent E. coli BL21(DE3) cells, and the AatB protein was expressed by isopropyl-beta-D-thiogalactopyranoside (IPTG) induction at a final concentration of 1 mM. Protein purification was performed Polyclonal anti-AatB antiserum was raised in New Zealand White rabbits by subcutaneous immunization, as described previously (23,38,39). The immunization dose was 300 g of purified AatB protein mixed with an equal volume of Montanide ISA 206 adjuvant (SEPPIC, Lyon, France).…”

Prevalence of Avian-Pathogenic Escherichia coli Strain O1 Genomic Islands among Extraintestinal and Commensal E. coli Isolates

“…1). IbeA is a recognized NMEC virulence factor (6) and also is involved in biofilm formation and avian fibroblast invasion (44). The ibeA gene has previously been shown to be a component of the GimA locus, thought to be an ancestral component of strains belonging to phylogenetic group B2 that has evolved via reductive evolution (10).…”

Genes of Virulence and Phylogenetic Group in Isolates of Avian Pathogenic Escherichia coli

“…Unlike E. coli of the intestinal biota APEC expresses enterobactins, aerobactins and yersiniabactins for the uptake and transport of iron [5,6]. Additionally, APEC synthesizes outer membrane proteins as Iss, Trat and OmpA1 that allow it to evade host defenses, as well as another proteins that contribute to the formation of biofilms [7].…”