Autoradiographic methods have been developed to detect metabolic activity of viable but nonculturable cells of Helicobacter pyloyi in water. Four strains of H. pylori were studied by using microcosms containing suspensions of 72-h cultures in water. The suspensions of aged, nonculturable cells of H. pylori were incubated with [3HJthymidine for 24 to 72 h, after which the cell suspensions were exposed to Kodak NTB2 emulsion for 3 to 28 days. Each sample was processed with three separate controls to rule out false-positive reactions. The organism remains viable and culturable under these conditions for up to 48 h and, in some cases, 20 to 30 days, depending on physical conditions of the environment. We found that temperature was a significant (P c 0.01) environmental factor associated with the viability ofH. pylori cells in water. Autoradiographs of tritium-labeled cells of H. pylori revealed aggregations of silver grains associated with uptake by H. pylori of radiolabelled substrate. Findings based on the autoradiographic approach give strong evidence supporting the hypothesis that there is a waterborne route of infection for H. pylori. The possibility that H. pylori may persist in water in a metabolically active stage but not actively growing and dividing is intriguing and relevant to public health concerns.
A human in vitro model to study the interaction between Helicobacter pylori and gastric epithelial cells was developed using primary cultures of gastric mucosal cells (isolated from gastric biopsies or operative specimen and maintained in culture for 2 weeks) as well as the well-differentiated human gastric carcinoma cell line HM02, the undifferentiated gastric tumour cell line HM51, and the laryngeal epithelial cell line HEp-2. Primary cultures and all cell lines were exposed to seven isolates of H. pylori isolated from gastritis and duodenal ulcer patients. Microbial adherence was assessed by microscopical evaluation of Giemsa-stained preparations and by culturing the viable bacteria attached to the epithelial cells. All H. pylori isolates adhered to the gastric cells in primary culture, to HM02 cells, and to HEp-2 cells with the greatest binding affinity found in primary gastric cells. No adherence was detected in HM51 cells. H. pylori adherence was dependent on bacterial load, incubation time, and temperature. There was no difference in microbial binding between H. pylori isolates derived from gastritis and duodenal ulcer patients. The effect of antiulcer drugs on H pylori adherence was investigated by pre-incubating isolates of H. pylori with omeprazole, cimetidine, and bismuth subcitrate. Omeprazole and cimetidine failed to significantly influence microbial adherence. In contrast, bismuth subcitrate already in concentrations below the MIC range decreased H. pylori adherence in gastric epithelial cells and in HEp-2 cells substantially. Our study shows that primary cultured human gastric mucosal cells and the human gastric carcinoma cell line HM02 provide suitable in vitro models for the study of the interactions between H. pylori and the gastric epithelium. This gastric cell model is characterized by a high affinity for H. pylori binding.
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