Raoultella planticola is an emerging pathogen causing several infections in humans, and its roles in the propagation of antibiotic resistance genes (ARGs) remain uncharacterized. In this study, a carbapenem and tigecycline-resistant R. planticola isolate was recovered from hospital sewage. It carried nine plasmids, bearing 30 ARGs, including one blaKPC-2 and two blaNDM-1. It also contained a plasmid-borne efflux pump gene cluster, tmexCD1-toprJ, conferring resistance to tigecycline. Analysis of plasmid sequences revealed that both blaNDM-1-carrying plasmids were highly similar to those recovered from humans, reinforcing the close relatedness of environmental and clinical isolates. We also identified that plasmid bearing blaNDM-1 or tmexCD1-toprJ1 was transferable, and can be stabilized in the host bacteria, indicating that the R. planticola isolate has a considerable potential in the dissemination of ARGs. Besides, we found that this isolate could produce biofilm and was virulent in a Galleria mellonella infection model. In conclusion, our study shows the convergence of virulence and multidrug resistance in a R. planticola isolate. This potentially virulent superbug may disseminate into its receiving rivers, and finally to humans through cross-contamination during recreation activities or daily use of water, which poses a risk to public health.

Multidrug-resistant (MDR) Proteus, especially those strains producing extended-spectrum β-lactamases (ESBL) and carbapenemases, represents a major public health concern. In the present work, we characterized 27 MDR Proteus clinical isolates, including 23 Proteus mirabilis, three Proteus terrae, and one Proteus faecis, by whole-genome analysis. Among the 27 isolates analyzed, SXT/R391 ICEs were detected in 14 strains, and the complete sequences of nine ICEs were obtained. These ICEs share a common backbone structure but also have different gene contents in hotspots and variable regions. Among them, ICEPmiChn2826, ICEPmiChn2833, ICEPmiChn3105, and ICEPmiChn3725 contain abundant antibiotic resistance genes, including the ESBL gene blaCTX-M-65. The core gene phylogenetic analysis of ICEs showed their genetic diversity, and revealed the cryptic dissemination of them in Proteus strains from food animals and humans on a China-wide scale. One of the isolates, FZP3105, acquired an NDM-1-producing MDR plasmid, designated pNDM_FZP3105, which is a self-transmissible type 1/2 hybrid IncC plasmid. Analysis of the genetic organization showed that pNDM_FZP3105 has two novel antibiotic resistance islands bearing abundant antibiotic resistance genes, among which blaNDM-1 is located in a 9.0 kb ΔTn125 bracketed by two copies of IS26 in the same direction. In isolates FZP2936 and FZP3115, blaKPC-2 was detected on an IncN plasmid, which is identical to the previously reported pT211 in Zhejiang province of China. Besides, a MDR genomic island PmGRI1, a variant of PmGRI1-YN9 from chicken in China, was identified on their chromosome. In conclusion, this study demonstrates abundant genetic diversity of mobile genetic elements carrying antibiotic resistance genes, especially ESBL and carbapenemase genes, in clinical Proteus isolates, and highlights that the continuous monitoring on their transmission and further evolution is needed.

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The emergence of carbapenem-resistant Proteus represents a serious threat to global public health due to limited antibiotic treatment options. Here, we characterize a Proteus isolate NMG38-2 of swine origin that exhibits extensive drug resistance, including carbapenems. Whole-genome sequencing based on Illumina and MinION platforms showed that NMG38-2 contains 24 acquired antibiotic resistance genes and three plasmids, among which, pNDM_NMG38-2, a pPvSC3-like plasmid, is transferable and co-carries blaNDM-1 and lnu(G). Sequence analysis of pPvSC3-like plasmids showed that they share a conserved backbone but have a diverse accessory module with complex chimera structures bearing abundant resistance genes, which are facilitated by transposons and/or homologous recombination. The acquisition of blaNDM-1 in pNDM_NMG38-2 was due to the ISCR1-mediated integration event. Comprehensive analysis of the lnu(G)-bearing cassettes carried by bacterial plasmids or chromosomes revealed a diversification of its genetic contexts, with Tn6260 and ISPst2 elements being the leading contributors to the dissemination of lnu(G) in Enterococcus and Enterobacteriaceae, respectively. In conclusion, this study provides a better understanding of the genetic features of pPvSC3-like plasmids, which represent a novel plasmid group as a vehicle mediating the dissemination of blaNDM-1 among bacteria species. Moreover, our results highlight the central roles of Tn6260 and ISPst2 in the spread of lnu(G).

Three NDM-5-producing Enterobacteriaceae (Escherichia coli, Klebsiella pneumoniae, and Citrobacter braakii, one each) were isolated during a screening study for the presence of carbapenemase-producing Enterobacteriaceae (CPE) strains in urban rivers in China. The aim of the present study was to characterize these NDM-5-producing isolates by using whole-genome analysis. Methods: In vitro susceptibility testing was performed using the broth microdilution method. Conjugation assay was carried out to investigate the transferability of bla NDM-5harboring plasmids. Whole-genome sequencing was performed using an Illumina HiSeq combined with the PacBio RSII system. The genetic characteristics of the bla NDM-5harboring plasmids were analyzed. Antimicrobial resistance genes and virulence genes were identified from the genome sequences. Phylogenetic analysis was performed based on core genome. Results: Antimicrobial susceptibility testing showed that all three isolates were resistant to carbapenems, cephalosporins, quinolones, and aminoglycosides, and susceptible to colistin. Whole-genome sequencing showed that each isolate carried multiple antibiotic resistance genes mediating multidrug resistance, and harbored numerous virulence genes, some of which were located on plasmids. In these isolates, bla NDM-5 was carried by an IncX3 plasmid in K. pneumoniae and C. braakii, and on an IncR/IncX1 plasmid in E. coli. Conjugation experiments showed that these bla NDM-5 -haboring plasmids were successfully transferred to E. coli J53. Phylogenetic analysis revealed that E. coli SCLZR49 was present in a cluster with isolates of different origin, K. pneumoniae SCLZR50 was mainly clustered with clinical isolates, and C. braakii SCLZR53 had closely genetic relationship with environmental isolates. Conclusion:This study revealed contamination of the urban river ecosystems by clinically significant carbapenemase gene bla NDM-5 , raising the possibility of plasmid transmission into the environmental from humans and highlighting the need for a constant surveillance of CPE in the environment under the "One-Health" perspective.