Whole-genome sequencing of clarithromycin resistant Helicobacter pylori characterizes unidentified variants of multidrug resistant efflux pump genes

Gut Pathogens

Takeshi Azuma¹  Masaru Yoshida¹  Shin Nishiumi¹  Koji Yamamoto¹  Lin Yang¹  Yoshiki Murakami⁴  Yoshihide Keida³  Kaoru Kikuchi³  Yukio Yoshida³  Rina Okada¹  Toshihito Tanahashi²  Akira Iwamoto¹ 

[1] Department of Internal Medicine, Division of Gastroenterology, Kobe University Graduate School of Medicine, 7-5-1 Kusunoki-cho, Chuo-ku, Kobe 650-0017, Japan;Department of Medical Pharmaceutics, Kobe Pharmaceutical University, 4-19-1 Motoyama-kita, Higashinada-ku, Kobe 658-8558, Japan;Division of Internal Medicine, Okinawa Prefectural Chubu Hospital, 281 Miyazato, Uruma-city, Okinawa 904-2293, Japan;Department of Hepatology, Osaka City University Graduate School of Medicine, 1-4-3 Asahimachi, Abeno-ku, Osaka 545-8585, Japan

关键词: TolC homolog;    Multidrug efflux;    Clarithromycin;    Helicobacter pylori;    Whole-genome sequencing;   

Others  :  817331
DOI  :  10.1186/1757-4749-6-27

 received in 2014-05-15, accepted in 2014-06-18,  发布年份 2014

【 摘 要 】

Background

Clarithromycin (CLR) is the key drug in 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 has been established in A2146G and A2147G mutations in the 23S rRNA gene, but evidence of the involvement of other genetic mechanisms is lacking. Using the MiSeq platform, whole-genome sequencing of the 19 clinical strains and the reference strain ATCC26695 was performed to identify single nucleotide variants (SNVs) of multi-drug resistant efflux pump genes in the CLR-resistant phenotype.

Results

Based on sequencing data of ATCC26695, 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 ranged from 1.82 to 10.8 million, and average coverage ranged from 90.9- to 686.3-fold, which were acceptable criteria for detecting SNVs. Utilizing the conventional approach of allele-specific PCR, point mutations in the 23S rRNA gene were detected in 12 clinical resistant isolates, but not in 7 clinical susceptible isolates. All sequencing reads of CLR-resistant strains had a G mutation in an identical position of the 23S rRNA gene. In addition, genetic variants of four gene clusters (hp0605-hp0607, hp0971-hp0969, hp1327-hp1329, and hp1489-hp1487) of TolC homologues, which have been implicated in multi-drug resistance, were examined. Specific SNVs were dominantly found in resistant strains.

Conclusions

Gene clusters of TolC homologues are involved in CLR susceptibility profiles in individual H. pylori strains. Whole-genome sequencing has yielded novel understanding of genotype-phenotype relationships.

【 授权许可】

   
2014 Iwamoto et al.; licensee BioMed Central Ltd.

【 图 表 】

Figure 2.

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