Characterization and Molecular Mechanism of Aminoglycoside-6-Adenyl Transferase Associated with Aminoglycoside Resistance from Elizabethkingia meningoseptica

Abstract

Shaoxing Zhang,1 Yuxin Zhang,1 Ruijie Liu,1 Shuying Yuan,2 Yanwen Chen,1 Wenjie Li,1 Xinrong Lu,3 Yongliang Tong,3 Linlin Hou,4 Li Chen,3 Guiqin Sun1 1School of Medical Technology and Information Engineering, Zhejiang Chinese Medical University, Hangzhou, Zhejiang, People’s Republic of China; 2Clinical Laboratory Department, Jiaxing Maternity and Child Health Care Hospital, Jiaxing, Zhejiang, People’s Republic of China; 3Key Laboratory of Medical Molecular Virology (MOE/NHC/CAMS), School of Basic Medical Sciences, Fudan University, Shanghai, People’s Republic of China; 4College of Veterinary Medicine, Qingdao Agricultural University, Qingdao, Shandong, People’s Republic of ChinaCorrespondence: Guiqin Sun, School of Medical Technology and Information Engineering, Zhejiang Chinese Medical University, 548 Binwen Road, Binjiang District, Hangzhou, Zhejiang, People’s Republic of China, Tel [phone], Fax [phone], Email [email protected] Li Chen, Key Laboratory of Medical Molecular Virology (MOE/NHC/CAMS), School of Basic Medical Sciences, Fudan University, 131 Dongan Road, Xuhui District, Shanghai, People’s Republic of China, Tel [phone], Fax [phone], Email [email protected]: Elizabethkingia meningoseptica (EM) is a multi-drug-resistant bacterium of global concern for its role in nosocomial infection and is generally resistant to aminoglycoside antibiotics. In the whole genome of an EM strain (FMS-007), an aminoglycoside-6-adenyl transferase gene (ant(6)FMS-007) was predicted. This study aimed to characterize the biochemical function of ANT(6)FMS-007 and analyze the relationship between genotype and phenotype of ant(6) in clinical EM isolates, so as to provide evidence for clinical precision drug use. This study could establish a method for the verification of known or unknown functionally resistant genes.Methods: A total of 42 EM clinical isolates were collected from clinical departments during 2015– 2023. The phenotype of aminoglycoside antibiotics was analyzed by broth microdilution (BMD) and Kirby-Bauer (K-B) methods. The whole-length ant(6) from EM clinical isolates was analyzed by polymerase chain reaction (PCR) and sequencing. The biochemical function of predictive ANT(6)FMS-007 from the FMS-007 whole genome was identified by 3D plate experiment and mass spectrometry analysis. Candidate active sites were predicted by multi-species sequence alignment and molecular docking, and other important sites were identified in the comparison of ant(6) genotypes and phenotypes of EM clinical isolates. Drug susceptibility test was used to verify the function of these sites.Results: The predictive ANT(6)FMS-007 protein could inactivate STR by modifying STR with ATP to form STR-AMP. Four active sites (Asp-38, Asp-42, Lys-95, and Lys-213) of ANT(6)FMS-007 were identified. Thirty-one EM clinical isolates (74%) carried the ant(6) gene. Eight EM clinical isolates containing the ant(6) gene had MIC values (

Keywords

• drug-resistance
• aminoglycoside-6-adenyl transferase
• characterization
• active sites
• elizabethkingia meningoseptica