Fengwei Jiang,Yan Yang,Zhao Mao,Wentong Cai,Ganwu Li

Microbiol Spectr.2023 Nov 2:e0152523.doi: 10.1128/spectrum.01525-23. Online ahead of print.

Abstract

Enterotoxigenic  Escherichia coli  (ETEC) cause severe diarrhea in humans and animals. These bacteria encode two major classes of virulence factors, adhesins that promote colonization of the small intestine and enterotoxins that induce fluid secretion and thus diarrhea. Oxygen and redox sensors are regulators of virulence expression in multiple bacterial pathogens. In this study, we show that microaerobic conditions enhance the expression of virulence factors in a porcine Shiga toxin-producing ETEC (STEC/ETEC) strain. We then investigated the role of the aerobic respiration control regulator ArcA in the pathogenicity and virulence regulation in STEC/ETEC. In a mouse model, deletion of  arcA  caused less severe pathology, and the  arcA  mutant displayed lower levels of colonization and obvious weight gain of the mice compared to the wild type (WT). In a cell culture model, a lack of  arcA  reduced the adherence of STEC/ETEC to porcine intestinal epithelial cells. Furthermore, ArcA positively regulated the expression of several key virulence factors, including F18 fimbriae ( fed ), heat-labile ( eltA ) and heat-stable ( estB ) toxins, Shiga toxin 2e ( stx2e ), and hemolysin ( hlyC ), under microaerobic conditions and  in vivo  conditions. We then found that ArcA positively regulated the expression of  eltA ,  estB , and  hlyC  by competing with a global repressor H-NS (histone-like nucleoid structuring). Mechanistically, we show that ArcA protein directly binds to the promoters of target genes, displaces H-NS silencing from the promoter and counteracts H-NS's repression. Collectively, our data established a key role for ArcA in the pathogenicity and virulence expression of porcine STEC/ETEC. IMPORTANCE Enterotoxigenic  Escherichia coli  (ETEC) cause severe diarrhea in humans and animals, leading to death and huge economic loss worldwide. Thus, elucidation of ETEC's pathogenic mechanisms will provide powerful data for the discovery of drugs serving as prevention or therapeutics against ETEC-caused diarrheal diseases. Here, we report that ArcA plays an essential role in the pathogenicity and virulence regulation in ETEC by positively regulating the expression of several key virulence factors including F18 fimbriae, heat-labile and heat-stable toxins, Shiga toxin 2e, and hemolysin, under microaerobic conditions and  in vivo . Moreover, we found that positive regulation of several virulence genes by ArcA requires a global repressor H-NS (histone-like nucleoid structuring), implying that ArcA may exert positive effects by antagonizing H-NS. Collectively, our data established a key role for ArcA in the pathogenicity of porcine ETEC and ETEC strains isolated from human infections. Moreover, our work reveals another layer of regulation in relation to oxygen control of virulence factors in ETEC.

Keywords: ArcA; Shiga toxin-producing enterotoxigenic Escherichia coli (STEC/ETEC); attenuation; virulence factor; virulence regulation.