Detection of Helicobacter pylori Microevolution and Multiple Infection from Gastric Biopsies by Housekeeping Gene Amplicon Sequencing

Abstract

Despite the great efforts devoted to research on Helicobacter pylori, the prevalence of single-strain infection or H. pylori mixed infection and its implications in the mode of transmission of this bacterium are still controversial. In this study, we explored the usefulness of housekeeping gene amplicon sequencing in the detection of H. pylori microevolution and multiple infections. DNA was extracted from five gastric biopsies from four patients infected with distinct histopathological diagnoses. PCR amplification of six H. pylori-specific housekeeping genes was then assessed on each sample. Optimal results were obtained for the cgt and luxS genes, which were selected for amplicon sequencing. A total of 11,833 cgt and 403 luxS amplicon sequences were obtained, 2042 and 112 of which were unique sequences, respectively. All cgt and luxS sequences were clustered at 97% to 9 and 13 operational taxonomic units (OTUs), respectively. For each sample from a different patient, a single OTU comprised the majority of sequences in both genes, but more than one OTU was detected in all samples. These results suggest that multiple infections with a predominant strain together with other minority strains are the main way by which H. pylori colonizes the human stomach.

Whole-Genome Sequencing and Comparative Genomics of Three Helicobacter pylori Strains Isolated from the Stomach of a Patient with Adenocarcinoma

“…The ANI values of the B508 isolates were ≥99.58%, being consistent with the definition of species [ 26 ], and it is even more consistent considering they come from microevolution events. As shown by Palau et al [ 25 , 27 ], the strain B508A-T2A is the most distant among them. Therefore, the aim of the study was to search within these three strains for genetic markers that could be linked with the level of virulence, pathogenicity or the risk of developing gastric cancer.…”

“…Even though hundreds of H. pylori genomes have been published, few studies focused on different strains obtained from the same patient at a single point in time [ 24 ]. Recently, we suggested that the predominant pattern of H. pylori infections in humans are events of multiple infections, including a predominant strain and multiple minority H. pylori strains [ 25 ].…”

High prevalence of Helicobacter pylori mixed infections identified by multilocus sequence typing in Ningbo, China

“…Helicobacter pylori is the most common infectious pathogen causing gastrointestinal diseases in humans, and can cause chronic gastritis, peptic ulcer, and gastric cancer ( Blaser and Atherton, 2004 ; Burkitt et al, 2017 ; Camilo et al, 2017 ). Individuals can be infected by one or more types of H. pylori , which is known as multiple infections ( Didelot et al, 2013 ; Palau et al, 2020 ), co-infection ( Didelot et al, 2013 ; Seo et al, 2019 ; Mi et al, 2021 ), or mixed infections ( Kao et al, 2014 ; Lai et al, 2016 ). Ben Mansour et al (2016) distinguished the concepts of multiple infections and mixed infections, and defined multiple infections as an individual with two or more genetically different H. pylori isolates, whereas they defined mixed infections as an individual with two or more H. pylori isolates with different antibiotic sensitivity characteristics.…”

“…In this study, 12 different STs were obtained by MLST from H. pylori isolates isolated from 6 unrelated patient control groups and 6 ATCC controls, whereas only two different profiles from the clinical isolates were obtained by RAPD (data not shown), indicating that four isolates were not distinguished as different based on the RAPD type. However, almost all studies on mixed infections have not included controls and reference strains for H. pylori isolates from patients and ATCC strains ( Wong et al, 2001 ; Kim et al, 2003 ; Carroll et al, 2004 ; Kao et al, 2014 ; Ben Mansour et al, 2016 ; Lai et al, 2016 ; Seo et al, 2019 ; Palau et al, 2020 ; Mi et al, 2021 ).…”