Whole genome sequencing reveals novel genetic mutations of Helicobacter pylori associating with resistance to clarithromycin and levofloxacin

Abstract

<h3>Background</h3><p>Detection of mutations in one or a couple of genes may not provide enough data or cover all the genomic DNA variance related to antibiotic resistance of <em>Helicobacter pylori</em> to clarithromycin (CLA) and levofloxacin (LVX). We aimed to perform whole genome sequencing to explore novel antibiotic resistance-related genes to increase predictive accuracy for future targeted sequencing tests.</p><h3>Methods</h3><p>Gastric mucosal biopsies were taken during upper endoscopy in 27 <em>H</em>. <em>pylori</em>-infected patients. According to culture-based antibacterial susceptibility test, <em>H</em>. <em>pylori</em> strains were divided into three groups, with nine strains in each group: CLA single-drug resistance (group C), LVX single-drug resistance (group L), and strains sensitive to all antibacterial drugs (group S). Based on whole genome sequencing with group S being the control, group C and group L group-specific single nucleotide variants and amino acid mutations were screened, and potential candidate genes related to CLA and LVX resistance were identified.</p><h3>Results</h3><p>The median age of study subjects was 35 years (IQR: 31–40), and 17 (63.0%) were male. All nine CLA-resistant strains had A2143G mutations in 23<em>S rRNA</em>, while none of nine sensitive strains had the mutation. Six of nine strains in group L and six of nine strains in group S had 87th or 91st mutation in <em>gyrA</em>. After comparing sequencing data of strains among the three groups, we identified five mutated positions belonging to four genes related to CLA resistance, and 31 mutated positions belonging to 20 genes related to LVX resistance. Novel genetic mutations were detected for CLA resistance (including <em>fliJ</em> and <em>clpX</em>) and LVX resistance (including <em>fliJ</em>, <em>cheA</em>, <em>hemE</em>, Val360Ile, and HP0568). Missense mutations in <em>fliJ</em> and <em>cheA</em> gene were mainly involved in chemotaxis and flagellar motility to facilitate bacterial escape of antibiotics, while the functions of other novel gene mutations underpinning antibiotic resistance remain to be investigated.</p><h3>Conclusion</h3><p>Whole genome sequencing detected potential novel genetic mutations conferring resistance of <em>H. pylori</em> to CLA and LVX including <em>fliJ</em> and <em>cheA</em>. Further studies to correlate these findings with treatment outcome should be performed.</p>