Jiamin Long,1,2,* Jiali Chen,3,* Yue Yuan,4,* Shaozhen Yang,5,* Xinya Xie,1,2 Xuelian Wu,1,2 Yuan Liu,1,2 Jinpeng Guo,2 Yong Chen,1,2 Changjun Wang,1,2 Xiong Liu2 

1School of Public Health, China Medical University, Shenyang, 110122, People’s Republic of China; 2Chinese PLA Center for Disease Control and Prevention, Beijing, 100071, People’s Republic of China; 3School of Medicine, NanKai University, Tianjin, 300071, People’s Republic of China; 4Fifth Medical Center of Chinese PLA General Hospital, Beijing, 100039, People’s Republic of China; 5School of Basic Medical Sciences, Army Medical University (Third Military Medical University), Chongqing, 400038, People’s Republic of China

*These authors contributed equally to this work

Correspondence: Xiong Liu; Changjun Wang, Chinese PLA Center for Disease Control and Prevention, Beijing, 100071, People’s Republic of China, Email liuxiong714@163.com; science2008@hotmail.com

Background: Klebsiella michiganensis is an emerging human pathogen that causes nosocomial infections. Its prevalence and spread in the environment should not be ignored. This study identified and characterized Klebsiella michiganensis co-harboring blaKPC-2 and TmexCD2-ToprJ2 in hospital wastewater samples.
Methods: Twelve K. michiganensis strains were isolated from wastewater samples collected at a tertiary hospital in Beijing, China. The genomic characteristics of K. michiganensis strains were analyzed using whole-genome sequences, providing information on the comparison between the genome of K. michiganensis strains and the reference genome, antibiotic resistance genes (ARGs), virulence genes, secretion systems, and mobile genetic elements (plasmids, insertion sequences [ISs], and prophages).
Results: Genome analysis showed that the twelve multi-drug resistant (MDR) strains carried a variety of ARGs and virulence genes, as well as four macromolecular secretion systems (T1SS, T2SS, T5aSS, T5bSS, and T4aP). The genetic environments of both the TmexCD2-ToprJ2 gene cluster and blaKPC-2 gene contained ISs. The plasmids carrying TmexCD2-ToprJ2 gene cluster of nine strains in clade 1 and two strains in clade 2 were annotated as IncR plasmid and rep_cluster_1254 type, respectively. The plasmids carrying blaKPC-2 in 10 strains in clade 1 were identified as IncU, and the plasmids carrying blaKPC-2 in the k11 and k12 strains in clade 2 were IncU and IncX6. The phylogenetic tree and heatmap revealed that the secretion system of type VI (T6SSi) existed in 10 strains in clade 1, and Type IV (T4SS) only existed in the k11 strain in clade 2. In addition, K. michiganensis strains carried 13 plasmids, 14 ISs, and 138 prophages.
Conclusion: In this study, the whole genome sequencing demonstrated the diversity of K. michiganensis genome despite 12 K. michiganensis strains from a hospital wastewater, which lays the foundation for further genetic research and drug resistance gene transmission.

Keywords: Klebsiella michiganensis, hospital wastewater, whole-genome sequencing, blaKPC-2, TmexCD2-ToprJ2