A novel mobile RND-type efflux pump gene cluster, tmexC3D2-toprJ3, confers tigecycline resistance in Pseudomonas alcaligenes

Tigecycline exhibits promising activity against multidrug-resistant gram-negative bacteria (MDR-GNB), whose infections are difficult to treat with antimicrobials. However, mobile tigecycline resistance genes, such as tmexCD-toprJ, have emerged in Enterobacterales isolated in China, Vietnam, and possibly other countries in the world. TMexCD-TOprJ complexes function as RND efflux pumps for multiple antimicrobials including tigecycline. To date, tmexCD1-toprJ1, tmexCD2-toprJ2, and tmexCD3-toprJ3 have been reported as variants of tmexCD-toprJ genes. Here, we identified a novel hybrid type of tmexCD-toprJ gene cluster, named tmexC3D2-toprJ3, in tigecycline- and carbapenem nonsusceptible isolates of Pseudomonas alcaligenes, a rare opportunistic human pathogen, obtained from medical sewage in Japan in 2020. Complete genomic analysis revealed that the P. alcaligenes isolate KAM426 co-harbors tmexC3D2-toprJ3 consisting of tmexC3, tmexD2, and toprJ3, along with two copies of the carbapenemase gene blaIMP-1 on the chromosome. tmexC3D2-toprJ3 in KAM426 was flanked by the IS5/IS1182 family transposase gene, suggesting that the gene cluster was acquired by horizontal gene transfer (HGT). Comparative genomic analysis using the public genome database showed that tmexC3D2-toprJ3 has spread to other Pseudomonas species such as Pseudomonas aeruginosa via HGT mediated by mobile gene elements such as a plasmid. This study suggests that diverse tmexCD-toprJ-like tigecycline resistance genes have spread among MDR-GNB worldwide and further epidemiological genomic studies are needed.