The micronutrient zinc inhibits EAEC strain 042 adherence, biofilm formation, virulence gene expression, and epithelial cytokine responses benefiting the infected host

Abstract

Enteroaggregative Escherichia coli (EAEC) is a major pathogen worldwide, associated with diarrheal disease in both children and adults, suggesting the need for new preventive and therapeutic treatments. We investigated the role of the micronutrient zinc in the pathogenesis of an E. coli strain associated with human disease. A variety of bacterial characteristics-growth in vitro, biofilm formation, adherence to IEC-6 epithelial cells, gene expression of putative EAEC virulence factors as well as EAEC-induced cytokine expression by HCT-8 cells-were quantified. At concentrations (≤ 0.05 mM) that did not alter EAEC growth (strain 042) but that are physiologic in serum, zinc markedly decreased the organism's ability to form biofilm (P<0.001), adhere to IEC-6 epithelial cells (P<0.01), and express putative EAEC virulence factors (aggR, aap, aatA, virK) (P<0.03). After exposure of the organism to zinc, the effect on virulence factor generation was prolonged (> 3 h). Further, EAEC-induced IL-8 mRNA and protein secretion by HCT-8 epithelial cells were significantly reduced by 0.05 mM zinc (P<0.03). Using an in vivo murine model of diet-induced zinc-deficiency, oral zinc supplementation (0.4 µg/mouse daily) administered after EAEC challenge (10 (10) CFU/mouse) significantly abrogated growth shortfalls (by>90%; P<0.01); furthermore, stool shedding was reduced (days 9-11) but tissue burden of organisms in the intestine was unchanged. These findings suggest several potential mechanisms whereby physiological levels of zinc alter pathogenetic events in the bacterium (reducing biofilm formation, adherence to epithelium, virulence factor expression) as well as the bacterium's effect on the epithelium (cytokine response to exposure to EAEC) to alter EAEC pathogenesis in vitro and in vivo. These effects may help explain and extend the benefits of zinc in childhood diarrhea and malnutrition.

The Human Antimicrobial Protein Calgranulin C Participates in Control of Helicobacter pylori Growth and Regulation of Virulence

“…Zinc availability is tightly regulated within the vertebrate host, and thus, zinc homeostasis is gaining greater appreciation for its important role in infectious disease processes (70). Bacteria have evolved strategies to sense and respond to zinc to gain a competitive advantage against resident microbiota (71,72). Additionally, zinc's regulatory effect on bacterial secretion has been demonstrated.…”

Zinc deficiency alters host response and pathogen virulence in a mouse model of enteroaggregativeescherichia coli-induced diarrhea

“…21 To determine if the lower levels of zinc in the intestine (see Fig. 2) altered the virulence of EAEC in vivo, we isolated colon contents from dC and dZD mice 3 d post-infection and isolated mRNA for qPCR analysis of EAEC virulence factor expression.…”

“…We have previously shown that sub-MIC zinc concentrations from 0.01-0.05mM decreases adherence to intestinal epithelial cells in vitro. 21 The EAEC gene aggR is a major regulator of EAEC virulence factor expression, including factors aap, aatK, and virK. 22,23 In vitro expression of these factors were significantly higher when grown in media without zinc.…”

“…Supplementation of the media with as little as 0.01 mM zinc greatly decreased virulence factor expression and biofilm formation without inhibiting growth. 21 Zinc concentrations 0.1 mM inhibited growth of the bacterium.…”

Early-life enteric infections: relation between chronic systemic inflammation and poor cognition in children

“…172 Although findings have been more contradictory, zinc supplementation has also been found to benefit the intestinal microbiota 173,174 and to modulate the intestinal inflammatory responses, 175,176 even reducing the virulence of pathogenic E. coli. 177,178 Zinc supplementation is still the only nutritional therapy clearly recognized to protect children with diarrheal diseases in low-income countries. 179 These findings highlight the benefits of certain nutrients in restoring or maintaining intestinal barrier function, but how these nutrients interact with the early-life intestinal microbiota when enteric pathogens are present is largely unknown.…”