Molecular Detection of H.pylori Antibiotic-Resistant Genes and Bioinformatics Predictive Analysis

ABSTRACT To explore the mutation characteristics of H.pylori resistance-related genes to antibiotics of clarithromycin, levofloxacin and metronidazole. 23S rRNA, gyrA, gyrB, rdxA and frxA genes were amplified and sequenced, respectively. Their structural alteration after mutation was predicted using bioinformatics software. In the clarithromycin-resistant strains, the mutation rate in site A2143G was 74.2% (n=23). The mutations in sites C1883T, C2131T and T2179G might cause structural alteration. In the levofloxacin-resistant strains, the mutation rates in 87 (N to K/I) and 91 (D to N/Y/G) of gyrA were 28.6% (n=16) and 12.5% (n =7), respectively. Meanwhile, one of the mutation strains in site 91 was accompanied by D99N variation. Additionally, a D143E mutation was found in one drug-resistant strain. Some changes of tertiary structure occurred after these mutations. The mutation types of RdxA protein consisted of protein truncation caused by premature stop codons (n=26, 33.3%), frameshift mutations (n=8, 10.3%), FMN-binding sites (n=16, 20.5%) and the others (n=11, 14.1%). Predictive analysis showed that mutations in the first three groups and the A118S of the last group could lead to structural alteration. Our study suggested the clarithromycin-resistant sites of H.pylori were mainly located in A2143G of 23S rRNA. C1883T, C2131T and T2179G might also be related to resistance. Levofloxacin resistance was mainly based on the amino acid changes in 87 and 91 sites of gyrA. The new sites D99N and D143E might also be associated with resistance. Metronidazole resistance was related to RdxA protein truncation, frameshift, and FMN binding. The new site A118S might also be linked to drug resistance.