Isolation And Culture Of Lactobacillus From Human By Characteristics Of Inhibition On Helicobacter Pylori In Vitro

[Objective]Hp(Helicobacter pylori, Hp) is regarded as a major cause of chronic gastritis, peptic ulcer and even gastric malignancy, and there is no ideal measure for its eradication until present. Microecological treatment(therapy) can deal with many problem in traditional therapeutics from new point of views, and include promoting the health by anti-infection, improving immune system and modulating the intestinal ecology. Basing on the international experience of vaccine development in Helicobacter pylori and some successful application of Lactobacillus to prevention and/or treatment of_Helicobacter pylori, Lactobacilli from gastrointestinal samples with characteristic of effective inhibition and/or killing ability on Helicobacter pylori and better adhesive ability to gastric mucosa is selected and identified: The identified Lactobacilli strains are able to be made into some Microecological modulator and correlative health food , which can more operatively inhibit Helicobacter pylori breeding. There are some better releasing system of carrier in through investigating and developing the Hp genetic enginerring live vaccine. At the same time, we are trying to find out the inhibitory mechanism that Helicobacter pylori inhibited by Lactobacilli. [Methods]1. Isolation and identification of lactobacilli: gastric biopsy or fluid from gastrosia patients and faces samples from health adult were collected and used to isolate lactobacilli with preliminary identification.2. Selection of Lactobacilli with characteristic of effective inhibition and/orkilling ability on Helicobacter pylori type strain SSI and good adhesive ability to gastric mucosa:2.1 The well test was used to select Lactobacillus with the best growth inhibitory activity.2.2 The selected Lactobacillus strains were then identified by 19 biochemical events.2.3 The experiment of adhesive ability to gastric epithelial cell SGC7901 were done.2.4 The ability that Lactobacillus strains were able to endure the differ concentration of hydrochloric acid and cholic acido3. Study of inhibitory mechanism of Lactobacilli on Helicobacter pylori: through the experiment that Helicobacter pylori adhesion cell model with type stain SSI and gastric epithelial cell line SGC7901, the experimentation that Lactobacilli restrained Helicobacter pylori was established using indirect immunofluorescence antibody test, the ability of Lactobacilli inhibiting SSI to SGC7901 was observed by microscope and scanning electron microscopy.4. Definitive identification of selected Lactobacilli: combining with the colony morphous and biochemical events results, API50CHL, a standard lactobacillus system identify card, was used to identify the selected lactobacilli to species.[Results]1. 470 samples from gastric mucosa of clinical patients and stools of healthy adults were collected to isolate Lactobacilli.2. Combining adhesive test results and adhesion inhibitory test results, four strains of Lactobacilli were selected, which displaying both good adhesive ability to gastric epithelial cell and/or strong restraining Helicobacter pylori growth ability in vitro, from one hundred seventy—eight Lactobacilli strains .3. Both viable and non-viable states of the four Lactobacilli strains can strongly inhibit Helicobacter pylori SSI binding to SGC7901.4. Using API50CHL, a standard Lactobacillus system identification card , three of the four Lactobacilli were identified as Lactobacillus salivarius, and the otherone as Lactobacillus plantarum. [Conclusion]1. The four strains were isolated and identified from four hundred seventy samples of gastric mucosa , and furthermore, the strains were able to endure the differ concentration of hydrochloric acid and cholic acido2. The viable and non-viable states of the four Lactobacilli strains could strongly inhibit Helicobacter pylori SSI binding to SGC7901, and it was remarkable that the inhibitory ability of Lactobacilli in the viable states is as the same as in the non-viable states.3. The new test means came into being that Lactobacilli restrained Helicobacter pylori from adhering to SGC7901 in vitro, whereas the homologous experiment hadn't been done at home.4. By systemic identification, three of the four Lactobacilli strains were identified as Lactobacillus salivarius, and the other one as Lactobacillus plantarum.The four Lactobacilli strains through the selected experiment may be excellent strains, which have the potential to be used as microbial preparation and carrier for live engineering vaccine with both micorecological modulating and immune stimulating function.