Identification and Characterization of a Novel Chromosomal Aminoglycoside 2'-N-Acetyltransferase, AAC(2')-If, From an Isolate of a Novel Providencia Species, Providencia wenzhouensis R33.

Kexin Zhou,Mengdi Gao,Junwan Lu,Weina Shi,Qiaoling Li,Mei Zhu,Xi Lin,Hailin Zhang,Jialei Liang,Xu Dong,Chunlin Feng,Qiyu Bao,Xueya Zhang,Kewei Li,Peiyao Zhang

doi:10.3389/fmicb.2021.711037

Abstract

Multidrug-resistant bacteria from different sources have been steadily emerging, and an increasing number of resistance mechanisms are being uncovered. In this work, we characterized a novel resistance gene named aac(2′)-If from an isolate of a novel Providencia species, Providencia wenzhouensis R33 (CCTCC AB 2021339). Susceptibility testing and enzyme kinetic parameter analysis were conducted to determine the function of the aminoglycoside 2′-N-acetyltransferase. Whole-genome sequencing and comparative genomic analysis were performed to elucidate the molecular characteristics of the genome and the genetic context of the resistance gene-related sequences. Among the functionally characterized resistance genes, AAC(2′)-If shares the highest amino acid sequence identity of 70.79% with AAC(2′)-Ia. AAC(2′)-If confers resistance to several aminoglycoside antibiotics, showing the highest resistance activity against ribostamycin and neomycin. The recombinant strain harboring aac(2′)-If (pUCP20-aac(2′)-If/DH5α) showed 256- and 128-fold increases in the minimum inhibitory concentration (MIC) levels to ribostamycin and neomycin, respectively, compared with those of the control strains (DH5α and pUCP20/DH5α). The results of the kinetic analysis of AAC(2′)-If were consistent with the MIC results of the cloned aac(2′)-If with the highest catalytic efficiency for ribostamycin (kcat/Km ratio = [3.72 ± 0.52] × 104 M–1⋅s–1). Whole-genome sequencing demonstrated that the aac(2′)-If gene was located on the chromosome with a relatively unique genetic environment. Identification of a novel aminoglycoside resistance gene in a strain of a novel Providencia species will help us find ways to elucidate the complexity of resistance mechanisms in the microbial population.

Highlights

Aminoglycoside antibiotics disturb translation and promote mistranslation of proteins, resulting in altering the integrity of bacterial cell membranes (Aguirre Rivera et al, 2021)
The major aminoglycoside resistance mechanism encountered in clinical isolates of gram-negative and gram-positive bacteria is most frequently associated with the expression of modifying enzymes (Wright, 1999; Vakulenko and Mobashery, 2003)
We report the identification and characterization of a novel aminoglycoside 2 -N-acetyltransferase designated AMEs include acetyltransferases (AACs)(2 )-If encoded in the chromosome of a Providencia wenzhouensis strain isolated from a rabbit

Summary

Introduction

Aminoglycoside antibiotics disturb translation and promote mistranslation of proteins, resulting in altering the integrity of bacterial cell membranes (Aguirre Rivera et al, 2021) They have a broad antimicrobial spectrum, and they often work in synergy with other antibiotics, making them valuable antiinfective agents (van Hoek et al, 2011). More than 100 aminoglycosidemodifying enzymes (AMEs) have been described and are broadly categorized into three groups based on the type of modification (Ramirez and Tolmasky, 2010) These three families of AMEs include acetyltransferases (AACs), nucleotidyltransferases or adenyltransferases (ANTs), and phosphotransferases (APHs). These classes are further divided into subtypes based on different region specificities of the enzymes for aminoglycoside modifications (Shaw et al, 1993; Ramirez and Tolmasky, 2010). The subclasses include four AACs, namely, AAC(1), AAC(2 ), AAC(3), and AAC(6 ); five ANTs, namely, ANT(2 ), ANT(3 ), ANT(4 ), ANT(6), and ANT(9); and seven APHs, namely, APH(2 ), APH(3 ), APH(3 ), APH(4), APH(6), APH(7 ), and APH(9) (Haas and Dowding, 1975; Shaw et al, 1993)

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Results

Conclusion