Emergence of NDM-1- and CTX-M-3-Producing Raoultella ornithinolytica in Human Gut Microbiota.

Shuang Wang,Liuchen Xu,Zhenwang Bi,Hengjie Xie,Peibin Hou,Yan Li,Beiwen Zheng,Zengqiang Kou,Xiaohui Chi

doi:10.3389/fmicb.2019.02678

Abstract

Raoultella ornithinolytica is an opportunistic pathogen of the Enterobacteriaceae family and has been implicated in nosocomial infections in recent years. The aim of this study was to characterize a carbapenemase-producing R. ornithinolytica isolate and three extended-spectrum β-lactamase (ESBL)-producing R. ornithinolytica isolates from stool samples of adults in a rural area of Shandong Province, China. The species were identified using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and 16S rDNA sequence analysis. Antimicrobial susceptibility test showed that all four isolates were multidrug-resistant (MDR). The whole genome sequence (WGS) of these isolates was determined using an Illumina HiSeq platform, which revealed MDR-related genes. The S1 nuclease-pulsed-field gel electrophoresis (S1-PFGE) was used to characterize the plasmids carried by the R. ornithinolytica isolates. The blaNDM-1 and blaCTX-M-3 genes were probed using Southern blotting, which confirmed the location of both genes on the same plasmid with molecular weight of 336.5–398.4 kb. The transferability of blaNDM-1 and blaCTX-M was also confirmed by conjugation assays. Finally, BLAST analysis of both genes showed that mobile genetic elements were associated with the spread of drug resistance genes. Taken together, we report the presence of conjugative blaNDM-1 and blaCTX-M plasmids in R. ornithinolytica isolates from healthy humans, which indicate the possibility of inter-species transfer of drug resistance genes. To the best of our knowledge, this is the first study to isolate and characterize carbapenemase-producing R. ornithinolytica and ESBL-producing R. ornithinolytica isolates from healthy human hosts.

Highlights

Raoultella is a genus of encapsulated Gram-negative aerobic bacilli of the Enterobacteriaceae family (Luo et al, 2017) that was initially part of the genus Klebsiella, but later reclassified based on the 16S rDNA sequence and the rpoB, gyrA, and gyrB genes (Drancourt et al, 2001)
One carbapenemase-producing (ROF058) and three ESBLproducing (ROE007, ROE058, and ROI014) R. ornithinolytica strains were isolated from 1,380 fecal samples (Table 1) and confirmed by MALDI-TOF MS, 16S rDNA sequencing, and average nucleotide identity (ANI) analysis (Figure 1A)
The blaNDM-1, blaCTX-M-3, and blaCTX-M-14 genes were subsequently identified in all the strains, and multidrug resistance was confirmed by antimicrobial susceptibility tests

Summary

Introduction

Raoultella is a genus of encapsulated Gram-negative aerobic bacilli of the Enterobacteriaceae family (Luo et al, 2017) that was initially part of the genus Klebsiella, but later reclassified based on the 16S rDNA sequence and the rpoB, gyrA, and gyrB genes (Drancourt et al, 2001). R. ornithinolytica can cause pneumonia, biliary or urinary tract infections and bacteremia and such cases are being increasingly reported (Sekowska et al, 2015; Van Cleve et al, 2018). Both organ-specific and systemic R. ornithinolytica infections can be life-threatening, especially in cancer patients after abdominal surgery (Hajjar et al, 2018). It is vital to understand the pathogenic potential of R. ornithinolytica isolates in humans (De Petris and Ruffini, 2018). Cephalosporins, quinolones, and carbapenems are routinely used against this pathogen, and multi-drug resistant (MDR) R. ornithinolytica isolates have been reported in recent years (Wang et al, 2019)

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