PRJDB2569
Helicobacter Whole-genome Sequencing
Clarithromycin (CLR) is the key drug of the eradication therapy of Helicobacter pylori (H. pylori) infection, and widespread use of CLR has led to an increase in primary CLR-resistant H. pylori. The known mechanism of CLR resistance was established in A2146G and A2147G mutations in the 23S rRNA gene, but there is no detailed information about the other genetic evidences in the H. pylori genome. Using MiSeq platform, we performed the whole-genome sequencing of the 19 clinical strains and their reference ATCC26695 to identify the single nucleotide variants (SNVs) of multi-drug resistant efflux pump genes involved in the CLR-resistant phenotype. Based on the six sequencing data of ATC26695, over one million sequencing reads with over 50-fold coverage were sufficient to detect SNVs, but not indels in the bacterial genome. Sequencing reads of the clinical isolates were ranged from 1.82 million to 10.8 million, and average coverage were ranged from 90.9- to 686.3-fold, which was accepted criteria of detecting SNVs. With the conventional approach of allele-specific PCR, point mutations of 23S rRNA gene were found in 12 isolates, but not in 7 wild-types. All sequencing reads of CLR-resistant strains were identical with the mutation nucleotide of G in the same position of 23S rRNA gene. Four gene clusters (HP0605-HP0607; hefABC, HP0971-HP0969; hefDEF, HP1327-HP1329; hefGHI, HP1489-HP1487) of TolC homologues associated with multi-drug resistance were examined to clarify the genetic variants in CLR-resistant strains. Specific SNVs were found in CLR-resistant strains contributed to the CLR susceptibility profiles for each strain. Whole-genome sequencing has the vast amount of the genomic information and yields a novel understanding of genotype-to phenotype relationships.