Visualization technology developed for the rapid observation of bacterial adaptive evolution

A research group led by Doctoral Student Kojiro Uemura at Sapporo Medical University Graduate School of Medicine, along with Instructor Soh Yamamoto, Associate Professor Noriko Ogasawara, Professor Satoshi Takahashi, Professor Hirofumi Chiba, and Professor Shin-ichi Yokota from Sapporo Medical University School of Medicine, Assistant Professor Kotaro Aoki from Toho University Faculty of Medicine, Professor Takayuki Wada from Osaka Metropolitan University Graduate School of Human Life and Ecology, and Associate Professor Toyotaka Sato from Hokkaido University Graduate School of Veterinary Medicine, has developed a method called RIBEA (Rapid and Integrated Bacterial Evolution Analysis). This method can rapidly adapt and evolve bacteria in a short period of time - i.e., within 20 days - and comprehensively extract and predict, from among the numerous mutations that emerge during evolution, genetic mutations related to pathogenicity and antimicrobial resistance. Their study was published in Nature Communications.

In bacterial infections, understanding the patterns of bacterial pathogenicity and antimicrobial resistance development and elucidating their mechanisms at the genetic level is extremely important, but observing bacterial evolution traditionally requires long-term experiments.

Analyzing bacteria currently causing problems in clinical settings is relatively easy. However, it has been impossible to comprehensively investigate which bacterial factors and genetic mutations are actually involved in the development of pathogenicity and antimicrobial resistance when the acquisition mechanisms are unknown.

The research group developed the RIBEA method using Klebsiella pneumoniae, a bacterium that causes clinical problems in respiratory and bloodstream infections. To develop this method, the research group integrated four techniques: evolution experiments using easily mutating bacterial strains, whole-genome sequencing (WGS), transposon-directed sequencing (TraDIS), and in vivo evaluation using mouse models, to observe bacterial evolution in a short period.

First, through evolution experiments using easily mutating bacterial strains, they succeeded in creating a mutant strain of K. pneumoniae lacking DNA repair function, thus accelerating the speed of bacterial evolution by more than 800 times. When this mutant strain was repeatedly subcultured in the presence of human serum or clinically important antimicrobial agents, resistant mutants appeared within 7 days. The group performed WGS analysis of these evolved bacteria and extracted all the genetic mutations that appeared and accumulated during the evolution process. Simultaneously, using the TraDIS method, they comprehensively extracted the genes involved in resistance to serum and antimicrobial agents possessed by K. pneumoniae. By integrating these analytical results, they successfully identified, from among the numerous genetic mutations that appeared during the evolution process, genetic mutations that were actually involved in serum and antimicrobial resistance. Through in vivo evaluation using mouse models, they revealed that the identified genetic mutations and evolved bacteria actually affect pathogenicity and antimicrobial treatment efficacy in living organisms.

The RIBEA established in this study is a powerful tool for quickly evaluating how bacteria evolve, cause severe disease, and acquire antimicrobial resistance. In the future, it is expected to help prevent the aggravation and spread of infections through the early detection of dangerous bacteria and the implementation of appropriate countermeasures in medical settings worldwide. Additionally, since RIBEA can be applied not only to clinical settings but also to elucidate the ecology and adaptation patterns of many bacteria, it is expected to contribute to the overall development of bacteriology.

Journal Information
Publication: Nature Communications
Title: Rapid and Integrated Bacterial Evolution Analysis unveils gene mutations and clinical risk of Klebsiella pneumoniae
DOI: 10.1038/s41467-025-58049-1