Molecular detection of H. pylori antibiotic‐resistant genes and molecular docking analysis

Abstract

To explore the mutation characteristics of H. pylori resistance-related genes to antibiotics of clarithromycin (CAM), levofloxacin (LVX) and metronidazole, 23SrRNA, gyrA, gyrB, rdxA, and frxA genes were amplified and sequenced, respectively. Molecular docking study was performed to explore molecular interactions between chemotherapeutic agents and target proteins. In the CAM-resistant strains, the mutation rate in site A2143G was 74.2% (n = 23). The interactions in sites of G1949A, C1953T, and G2211T with CAM were weaker after mutation. In the LVX-resistant strains, the mutation rates in 87 (N to K/I) and 91 (D to N/Y/G) of gyrA were 28.6% (n = 16) and 12.5% (n = 7), respectively. We could infer by docking studies that N87 K/I, D91Y/G/N, D99N, and D143E mutations in GyrA protein all had weakened interaction with LVX. The mutation types of RdxA protein consisted of protein truncation caused by premature stop codons (n = 26, 33.3%), frameshift mutations (n = 8, 10.3%), FMN-binding sites (n = 16, 20.5%), and the others (n = 11, 14.1%).Docking analysis showed that some mutations in those four types of RdxA protein could reduce interaction with MNZ, and play a significant role in antibiotic resistance. What's more, we performed natural transformation with some of these mutated DNA fragments to see if they really impact susceptibility to antibiotics. Our study suggested that in addition to the reported mutation sites, H. pylori antibiotic resistance in this region may also be associated with changes in some new sites. Our study provides novel ideas and methods for the identification of H. pylori resistancerelated mutations, and also clues and basis for further investigation on specific molecular mechanism.

Mutations associated with Helicobacter pylori antimicrobial resistance in the Ecuadorian population

“…This limits the use of rdxA as a genetic marker for MTZ resistance (Chu et al, 2020). Therefore, phenotypic tests are needed to confirm that these mutations actually cause resistance to MTZ (Chu et al, 2020). In the case of the pbp1A gene, mutational changes are known to be the predominant cause of resistance to AMX in H. pylori.…”

“…However, in the case of the rdxA gene, mutations have been found in both sensitive and resistant strains or both. This limits the use of rdxA as a genetic marker for MTZ resistance (Chu et al, 2020). Therefore, phenotypic tests are needed to confirm that these mutations actually cause resistance to MTZ (Chu et al, 2020).…”

A Survey of Helicobacter pylori Antibiotic-Resistant Genotypes and Strain Lineages by Whole-Genome Sequencing in China

“… H. pylori genotypic resistance testing might be a good alternative to phenotypic resistance testing. Some genetic mutation loci have been reported to contribute to the H. pylori phenotype, and phenotypic resistance tests for clarithromycin and quinolones have been partially replaced by genotypic resistance tests ( 14 , 15 ). However, the loci of H. pylori conferring resistance to other antibiotics are controversial ( 16 , 17 ), and further studies are needed to determine which genetic mutations are responsible for resistance.…”

The outward shift of clarithromycin binding to the ribosome in mutantHelicobacter pyloristrains

“…Other supportive evidence comes from the molecular modeling research of telithromycin with A2058G mutant of 23S rRNA in Deinococcus radiodurans , which detected the antibiotic molecule distant from the position of G2058 and shifted toward the other side of the tunnel, to the vicinity of L22 71 . A similar weakening and repositioning of CLA in interaction with H pylori 23S rRNA were also observed in the molecular docking studies of A2143G mutant conformation 61 …”

“…We have also detected other mutations (T2182C, T2656A, and G2569C), which were outside of the drug‐interacting region and thus did not undergo our molecular docking studies. However, to our knowledge, there are very limited investigations, 61 exploring the molecular interactions underlying macrolide resistance in H pylori , caused by these key mutations.…”