Abstract
BackgroundThe entire gastrointestinal tract is protected by a mucous layer, which contains complex glycoproteins called mucins. MUC2 is one such mucin that protects the colonic mucosa from invading microbes. The initial interaction between microbes and mucins is an important step for microbial pathogenesis. Hence, it was of interest to investigate the relationship between host (mucin) and pathogen interaction, including Shigella induced expression of MUC2 and IL-1β during shigellosis.MethodsThe mucin-Shigella interaction was revealed by an in vitro mucin-binding assay. Invasion of Shigella dysenteriae into HT-29 cells was analyzed by Transmission electron microscopy. Shigella induced mucin and IL-1β expression were analyzed by RT-PCR and Immunofluorescence.ResultsThe clinical isolates of Shigella were found to be virulent by a congo-red binding assay. The in vitro mucin-binding assay revealed both Shigella dysenteriae and Shigella flexneri have binding affinity in the increasing order of: guinea pig small intestinal mucin<guinea pig colonic mucin< Human colonic mucin. Invasion of Shigella dysenteriae into HT-29 cells occurs within 2 hours. Interestingly, in Shigella dysenteriae infected conditions, significant increases in mRNA expression of MUC2 and IL-1β were observed in a time dependent manner. Further, immunofluorescence analysis of MUC2 shows more positive cells in Shigella dysenteriae treated cells than untreated cells.ConclusionsOur study concludes that the Shigella species specifically binds to guinea pig colonic mucin, but not to guinea pig small intestinal mucin. The guinea pig colonic mucin showed a greater binding parameter (R), and more saturable binding, suggesting the presence of a finite number of receptor binding sites in the colonic mucin of the host. In addition, modification of mucins with TFMS and sodium metaperiodate significantly reduced mucin-bacterial binding; suggesting that the mucin-Shigella interaction occurs through carbohydrate epitopes on the mucin backbones. Overproduction of MUC2 may alter adherence and invasion of Shigella dysenteriae into human colonic epithelial cells.
Highlights
Mucosal surfaces employ a number of protective strategies to defend against noxious substances and pathogens found within the intestinal lumen
Congo-red binding assay, appearance of orange/pink color colonies of S. dysenteriae (Fig. 1a) and S. flexneri (Fig. 1b) suggest that these species are highly virulent when compared to pale white color colonies of plasmid DNA cured non-virulent strain of Shigella flexneri (Fig. 1c)
Bacterial binding to native mucin The binding of Shigella dysenteriae and Shigella flexneri to human colonic mucin and guinea pig colonic mucin was concentration dependent and saturable
Summary
Mucosal surfaces employ a number of protective strategies to defend against noxious substances and pathogens found within the intestinal lumen. The mucosal surface contains mucins, which are complex glycoproteins synthesized and secreted by epithelial cells of various organs to lubricate and protect luminal surfaces of the human body [1]. Adherence is recognized as an important initial step in other bacterial infections [5,6,7], relatively little information is available on the mechanisms of attachment of Shigellae to cells on the mucosal surface. A specific site on the host may be involved in the binding of pathogenic bacteria; for example, Hemophillus influenzae, a respiratory pathogen, binds to respiratory mucins but does not bind or cause disease in the intestinal tract. The initial interaction between microbes and mucins is an important step for microbial pathogenesis. It was of interest to investigate the relationship between host (mucin) and pathogen interaction, including Shigella induced expression of MUC2 and IL-1b during shigellosis
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