Diversity Of VacA Gene Of Helicobacter Pylori And Evaluation Of Infected Mouse Model By ～(14)C-urea Breath Test

AIMS (1) To investigate the diversity of vacA and cagA of Helicobacter pylon (Hp) from patients from Guangzhou city or around area, and analyze characteristic of genotype distribution of H. Pylon strains and the relationship between the genotype and pathogenesis. (2) To establish mouse model infected with Hp and Helicobacter heilmanii (Hh). (3) To develop a method of 14C-urea breath test (UBT) for the detection of Hp and Hh without hurting the animal. METHODS (1) The vacA signal sequences and middle-region alleles and cagA of 92 Hp isolates were amplified by the method of polymerase chain reaction (PeR). (2) Neutral red absorb method was used to measure the cytotoxicity of Hp culture concentrated supernatants on gastric cancer cells strain SGC-7901 cells. (3) The method of PCR was applied to amplify ureB gene fragment and another unidentified fragment from Hh infected human and from Hh-Iike Helicobacter infected dog, and the unidentified fragments were sequenced. (4) 26 fresh isolates of Np and Sydney strain (SSI) were inoculated respectively to Balb/c mice to establish mouse model. (5) The gastric mucus tissue from a patient infected with Nh or from a dog infected with Nh-like Helicobacter were ground and inoculated directly to Balb/c mice to establish mouse model respectively. (6) UBT was used to detect Hp and Nh in mouse model. RESULTS (1) 92 clinical isolates of H. pylon were characterized by vacA typing. 14 of 92 isolates (15.2%) were sla/ml, 6 (6.5%) were slalmla, 55 (59.8%) were slaIm2, 2 (2.2%) were sib/mi, 5 (5.4%) were slb/m2, 8 (8.7%) were s2/m2, only 2 could not be typed. slb/mla or s2/mlor s2/mla isolates had not been found. (2) With regard to the Hp isolates from patients with peptic ulceration and chronic gastnitis, the signal sequences between two groups was statistically significant (PO.05). sla genotype Hp associated with peptic ulceration. (3) 83 of the 92 Hp isolates (90.2%) were cagA positive. The frequency of cagA positive in Hp isolates from patients with gastric cancer and peptic ulceration was more higher than those chronic gastnitis (P<0.05), which suggested that cagA-positive isolates correlated with gastric cancer and peptic ulceration. (4) The frequency of cagA-positive isolates associated with vacA signal sequence type, especially sla type, but no relation to midregion type. (5) There were relations between the level of in vitro cytotoxin activity of isolates and the signal sequence, midregion type, cagA gene status. (6) Both Rh from a patient with chronic gastritis and Rh-like Helicobacter from a dog could been amplified uniformly with 2 pairs of different primers in turn by the method of PCR, and there was 97.4% nucleotide identity between the Rh and Hh-like Helicobacter, which indicated that the two Helicobacter were the same bacteria, Rh. (7) Both Rh and Hh-like Helicobacter were prone to colonize in gastric mucosa of all experimental Balb/c mice. The fresh Np isolates and 551 strain could colonize in some of the experimental Balb/c mice also, but the colonization of Hp was poor. (8) The histological features among the fresh isolates, SS 1, Rh and Rh-like HeI.icobacter infection in mice were similar. But the changes of histological features between the Rh and Rh-like Helicobacter infection in mice were more ahead and severer than Np infection. After 4 weeks of infection, there were a few lymphocytes, neutrophilic leukocytes, acidophilic...