Isolation of Clostridioides difficile from the atmospheric fine particulate matter

Clostridioides difficile is a spore−forming, obligate anaerobic bacillus commonly found in soil and the intestinal tracts of animals. It is a major cause of antibiotic−associated diarrhea (AAD) in humans. Our previous study suggested a possible environmental dissemination. To investigate whether atmospheric aerosols contribute to its spread, atmospheric aerosol samples were collected from ten geographic locations in East Asia, and the presence of C. difficile genomic sequences was assessed using 16S rRNA amplicon sequencing. Additionally, atmospheric PM2.5 samples were collected from two sites in Japan, and analyzed by 16S rRNA amplicon sequencing and selective culturing. C. difficile genomic sequences were detected in aerosols from 8 sampling sites, with relative abundances ranging from 0.03% to 9.88% of total prokaryotic reads. From PM2.5 samples, seven C. difficile strains were isolated, and classified into five genotypes. Three genotypes matched low−frequency clinical or environmental strains in Japan (0.1–0.6%), while two were previously unreported. Phylogenetic analysis showed that four genotypes were closely related to strains from North America, Europe, Africa, and Southeast Asia, respectively. These findings are consistent with the possibility that C. difficile may disperse globally via atmospheric aerosols. Successful colonization of C. difficile in the deposited environments likely depends on ecological adaptability. Seven C. difficile strains were isolated from atmospheric aerosols in Japan. Aerosol-derived strains showed close genetic ties to strains from other continents. Aerosols may serve as a pathway for the global spread of C. difficile.