Heteroresistance of Helicobacter pylori from the same patient prior to antibiotic treatment

Antibiotic resistance among Helicobacter pylori strains has been increasing worldwide and has affected the efficacy of current treatments. The aim of this study was to evaluate whether treatment failure was due to the presence of antibiotic-susceptible and -resistant H. pylori simultaneously within the same host before eradication. In order to discover H. pylori with antibiotic heteroresistance in the same patient, we examined the antibiotic susceptibility of H. pylori isolated from 412 patients without H. pylori eradication. The E-test was used to determine the minimal inhibitory concentration of these strains. The results showed 19 (4.6%) of patients harbored antibiotic heteroresistant H. pylori, resistant to levofloxacin (5/19), clarithromycin (1/19) and metronidazole (16/19). Among them, three patients’ isolates showed heteroresistance to two antibiotics. The genetic diversity of each isolate was evaluated by random amplified polymorphic DNA PCR and the results showed that only 1 patient’ isolate (5.3%) had a different pattern while the others showed identical or similar fingerprinting patterns. Mutations in the genes responsible for antibiotic resistance were investigated by direct sequencing and compared between strains within each pair. All 5 levofloxacin-resistant isolates had mutations in GyrA at the QRDR region (N87 or D91). Strain 1571R with clarithromycin resistance had a A2042G substitution in its 23S rRNA. There were 15 metronidazole-resistant strains (100%) with isogenic variation of RdxA, and 6 strains (40%) contained FrxA variation (excluded pair 1159). These results suggest that the treatment failure of heteroresistant H. pylori mostly develops from high genomic variation of pre-existing strains through long term evolution rather than mixed infection with different strains.