Abstract
BACKGROUND: The gastric microbiome is closely associated with gastric cancer, and single-region 16S rRNA sequencing has limitations in analyzing its characteristics, necessitating the search for a better sequencing method. AIM: To evaluate the effectiveness of multi-region 16S rRNA gene sequencing in studying the microbiome of gastric cancer tissues. METHODS: Patients with gastric cancer (n = 118) who underwent surgery at Liyang People's Hospital from January 2022 to December 2024 were enrolled. Fifty-nine paraffin-embedded and 59 fresh tissue samples were obtained. The ZymoBIOMICS(TM) microbial community standard and Escherichia coli ATCC 25922 were used as positive controls. Multi-region and single-region 16S rRNA gene sequencing were performed. Species identification, detection rates at varying microbial abundances, operational taxonomic unit (OTU) counts, and alpha diversity indices in gastric cancer tissues were compared between the two methods. RESULTS: Multi-region 16S rRNA sequencing identified more species (eight species and eight genera) in the positive controls compared with single-region sequencing (one species and six genera). Detection rates at concentrations of 10(3), 10(2), and 10 CFU/mg were significantly higher using multi-region sequencing (P < 0.05). Multi-region sequencing also revealed significantly higher OTU counts and alpha diversity indices (Shannon, Simpson, and Chao1) in gastric cancer tissues (P < 0.05). CONCLUSION: Compared with single-region sequencing, multi-region 16S rRNA gene sequencing demonstrates superior species resolution and detection sensitivity, providing a more comprehensive profile of microbial diversity in gastric cancer tissues.