Study On Culture Of Helicobacter Pylori And Its Survival In Artificially Inoculated Environments

Helicobacter pylori (H.pylori) is known as the pathogen of chronic gastritis, it is closelyrelated to peptic ulcer and gastric cancer. However, the routes of transmission are not veryclear. Contaminated water and food have been suggested as vehicles in the transmission ofH.pylori in recent years. For lack of strong evidence, it is still not sure for the survival ofH.pylori in the environment. In this study, we used the candle jar method to cultureHelicobacter pylori (NCTC11637) and explored the optimum culture method. The survivalof H.pylori that artificially inoculated into four kinds of environmental sample was observedin this study.Part Ⅰ Set up the curture method of Helicobacter pylori and compare the culturemediaObjective: To set up the candle jar method to culture Helicobacter pylori in the laboratory,and select the optimum medium.Methods: The candle jar method was used to culture Helicobacter pylori, then identifiedthe strain and did the preservation process. Same amount of H.pylori was inoculated ondifferent culture media, and then bacteria concentration on the plates was detected after5days’ culture in order to select the optimum medium. Different dilutions of H.pylori wereinoculated on the optimal culture medium to determine the culture effect of the medium.Results: The successful recovery of Helicobacter pylori in the experiment, and the candle jarmethod could be used as a good way of H. pylori cultivation. The growth of H.pylori on fourkinds of solid medium was different. The best medium in the experiment was Brucella brothagar adding7%newborn calf serum, and it presented a good culture effect. Conclusion: The best medium for culturing was Brucella broth agar adding7%newborncalf serum, which presented a good culture effect. It could be used for further isloation ofHelicobacter pylori from environmental samples.Part ⅡStudy on survival of Helicobacter pylori in artificially inoculated environmentsObjective: To study survival of Helicobacter pylori in different artificially inoculatedenvironmental samples.Methods: Water, milk, ground pork and carrots samples were designed in this study. H.pylori was artificially inoculated in25ml (g) samples to106cfu/ml (g). The sample flaskswere placed in room temperature and4℃respectively. Each Sample was inoculated ontoBrucella broth agar plates in duplicate for H. pylori at different sampling points, and thewater sample was stained in addition.Results: Helicobacter pylori could not grow in all the four environments; it could onlysurvive for a period of time. Survival time of H. pylori at4℃was longer than at roomtemperature. H. pylori could be seen gradually changed from the rod form to spherosomeunder the microscope. Comparing the four different environments, the longest survival timeof H. pylori was in milk, for more than10days at4°C and more than6days at roomtemperature. Survival time in pork was over6days at4°C, and more than3days at roomtemperature. H. pylori in4°C water survived more than5days, but2days later it could notbe separated from samples at room temperature. In the carrot sample, which the survivaltime was the shortest, it could only survive a few hours both at4°C and room temperature; H.pylori could not be isolated from carrot sample after one day.Conclusion: Helicobacter pylori could not grow in the environment; it could only survivefor a period of time. It was seen gradually changed to the coccoid form. H. pylori survivedthe longest time in milk at the same temperature and bacterial concentration. It wassuggested that milk may be an important vehicle in transmission of Helicobacter pylori, andthe role of pork, water and other environmental media play in the transmission of H. pyloricould not be ignored.