Next-Generation Sequencing-Based Study of Helicobacter pylori Isolates from Myanmar and Their Susceptibility to Antibiotics

Phawinee Subsomwong,Yoshio Yamaoka,Tomohisa Uchida,Kyaw Htet,Vo Phuoc Tuan,Kartika Afrida Fauzia,Dalla Doohan,Takashi Matsumoto,Junko Akada,Langgeng Agung Waskito,Than Than Yee,Takeshi Matsuhisa

doi:10.3390/microorganisms10010196

Abstract

Evaluation of Helicobacter pylori resistance to antibiotics is crucial for treatment strategy in Myanmar. Moreover, the genetic mechanisms involved remain unknown. We aimed to investigate the prevalence of H. pylori infection, antibiotic resistance, and genetic mechanisms in Myanmar. One hundred fifty patients from two cities, Mawlamyine (n = 99) and Yangon (n = 51), were recruited. The prevalence of H. pylori infection was 43.3% (65/150). The successfully cultured H. pylori isolates (n = 65) were tested for antibiotic susceptibility to metronidazole, levofloxacin, clarithromycin, amoxicillin, and tetracycline by Etest, and the resistance rates were 80%, 33.8%, 7.7%, 4.6%, and 0%, respectively. In the multidrug resistance pattern, the metronidazole–levofloxacin resistance was highest for double-drug resistance (16/19; 84.2%), and all triple-drug resistance (3/3) was clarithromycin–metronidazole–levofloxacin resistance. Twenty-three strains were subjected to next-generation sequencing to study their genetic mechanisms. Interestingly, none of the strains resistant to clarithromycin had well-known mutations in 23S rRNA (e.g., A2142G, A2142C, and A2143G). New type mutation genotypes such as pbp1-A (e.g., V45I, S/R414R), 23S rRNA (e.g., T248C), gyrA (e.g., D210N, K230Q), gyrB (e.g., A584V, N679H), rdxA (e.g., V175I, S91P), and frxA (e.g., L33M) were also detected. In conclusion, the prevalence of H. pylori infection and its antibiotic resistance to metronidazole was high in Myanmar. The H. pylori eradication regimen with classical triple therapy, including amoxicillin and clarithromycin, can be used as the first-line therapy in Myanmar. In addition, next-generation sequencing is a powerful high-throughput method for identifying mutations within antibiotic resistance genes and monitoring the spread of H. pylori antibiotic-resistant strains.

Highlights

The successful attempt to isolate and culture the gram-negative microaerophilic bacterium that infects the gastric epithelium has become an important event in the extensive study of Helicobacter pylori (H. pylori) [1]
We aimed to analyze the genetic mutations associated with the five common antibiotics used for H. pylori therapy in Myanmar using whole-genome sequencing techniques
We examined the association between antibiotic resistance and mutations in genes related to each antibiotic

Summary

Introduction

The successful attempt to isolate and culture the gram-negative microaerophilic bacterium that infects the gastric epithelium has become an important event in the extensive study of Helicobacter pylori (H. pylori) [1]. The prevalence of H. pylori infection varies according to geographical region [2]. H. pylori infection is estimated to affect half of the world’s population, and it has been suggested that low prevalence has been reported only in some developed countries [2,3]. As H. pylori infection has been attributed to poor sanitation, poor socioeconomic status, and overcrowding, no declining tendency has been noted in the majority of developing countries [4,5]. H. pylori eradication continues to become a challenge for clinicians because of the development of antibiotic resistance. The increase in H. pylori resistance to clarithromycin (CAM), metronidazole (MNZ), and levofloxacin (LVX) has been reported to be responsible for the decrease in cure rates worldwide [7]. The success of individual therapies mainly depends on information on the resistance patterns in a given region

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