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The emergence of the mobile tigecycline-resistance gene, tet(X4), poses a significant threat to public health. To investigate the prevalence and genetic characteristics of the tet(X4)-positive Escherichia coli in humans, 1101 human stool samples were collected from a tertiary class-A hospital in Beijing, China, in 2019. Eight E. coli isolates that were positive for tet(X4) were identified from clinical departments of oncology (n = 3), hepatology (n = 2), nephrology (n = 1), urology (n = 1), and general surgery (n = 1). They exhibited resistance to multiple antibiotics, including tigecycline, but remained susceptible to meropenem and polymyxin B. A phylogenetic analysis revealed that the clonal spread of four tet(X4)-positive E. coli from different periods of time or departments existed in this hospital, and three isolates were phylogenetically close to the tet(X4)-positive E. coli from animals and the environment. All tet(X4)-positive E. coli isolates contained the IncX1-plasmid replicon. Three isolates successfully transferred their tigecycline resistance to the recipient strain, C600, demonstrating that the plasmid-mediated horizontal gene transfer constitutes another critical mechanism for transmitting tet(X4). Notably, all tet(X4)-bearing plasmids identified in this study had a high similarity to several plasmids recovered from animal-derived strains. Our findings revealed the importance of both the clonal spread and horizontal gene transfer in the spread of tet(X4) within human clinics and between different sources. [ABSTRACT FROM AUTHOR]

Tsuyoshi Sekizuka, Yuba Inamine, Takaya Segawa, Makoto KurodaLaboratory of Bacterial Genomics, Pathogen Genomics Center, National Institute of Infectious Diseases, Shinjuku, Tokyo, JapanPurpose: New Delhi metallo-β-lactamase (NDM)-5-producing Enterobacteriaceae have been detected in rivers, sewage, and effluents from wastewater treatment plants (WWTPs). Environmental contamination due to discharged effluents is of particular concern as NDM variants may be released into waterways, thereby posing a risk to humans. In this study, we collected effluent samples from a WWTP discharged into a canal in Tokyo Bay, Japan.Methods: Testing included the complete genome sequencing of Escherichia coli GSH8M-2 isolated from the effluent as well as a gene network analysis.Results: The complete genome sequencing of GSH8M-2 revealed that it was an NDM-5-producing E. coli strain sequence type ST542, which carries multiple antimicrobial resistance genes for β-lactams, quinolone, tetracycline, trimethoprim-sulfamethoxazole, florfenicol/chloramphenicol, kanamycin, and fosfomycin. The blaNDM-5 gene was found in the IncX3 replicon plasmid pGSH8M-2-4. Gene network analysis using 142 IncX3 plasmid sequences suggested that pGSH8M-2-4 is related to both clinical isolates of E. coli and Klebsiella species in Eastern Asia. GSH8M-2 also carries the blaCTX-M-55 gene in IncX1 plasmid pGSH8M-2-3.Conclusion: This is the first report of environmental NDM-5-producing E. coli isolated from a WWTP in Japan. NDM-5 detection is markedly increasing in veterinary and clinical settings, suggesting that dual β-lactamases, such as NDM-5 and CTX-M-55, might be acquired through multiple steps in environment settings. Environmental contamination through WWTP effluents that contain producers of NDM variants could be an emerging potential health hazard. Thus, regular monitoring of WWTP effluents is important for the detection of antimicrobial-resistant bacteria that may be released into the waterways and nearby communities.Keywords: sewage, carbapenemase, NDM-5, IncX3, CTX-M-55, IncX1

The emergence of the mobile tigecycline-resistance gene, tet(X4), poses a significant threat to public health. To investigate the prevalence and genetic characteristics of the tet(X4)-positive Escherichia coli in humans, 1101 human stool samples were collected from a tertiary class-A hospital in Beijing, China, in 2019. Eight E. coli isolates that were positive for tet(X4) were identified from clinical departments of oncology (n = 3), hepatology (n = 2), nephrology (n = 1), urology (n = 1), and general surgery (n = 1). They exhibited resistance to multiple antibiotics, including tigecycline, but remained susceptible to meropenem and polymyxin B. A phylogenetic analysis revealed that the clonal spread of four tet(X4)-positive E. coli from different periods of time or departments existed in this hospital, and three isolates were phylogenetically close to the tet(X4)-positive E. coli from animals and the environment. All tet(X4)-positive E. coli isolates contained the IncX1-plasmid replicon. Three isolates successfully transferred their tigecycline resistance to the recipient strain, C600, demonstrating that the plasmid-mediated horizontal gene transfer constitutes another critical mechanism for transmitting tet(X4). Notably, all tet(X4)-bearing plasmids identified in this study had a high similarity to several plasmids recovered from animal-derived strains. Our findings revealed the importance of both the clonal spread and horizontal gene transfer in the spread of tet(X4) within human clinics and between different sources.

Abstract We have identified an IncX1 plasmid named pQJDSal1 from Salmonella enterica subsp. enterica serovar Pullorum (S. Pullorum). The plasmid is 67,685 bp in size and has 72 putative genes. pQJDSal1 harbors a conserved IncX1-type backbone with predicted regions for conjugation, replication and partitioning, as well as a toxin/antitoxin plasmid addiction system. Two regions (A and B) that have not been previously reported in IncX1 plasmids are inserted into the backbone. Region A (10.7 kb), inserted between parA and taxD , consists of a new Tn 6168- like transposon containing an arsenic resistant operon arsB2CHR and sulfonamide resistance gene sul2. Region B contains another arsenic resistant operon arsADHR , resistance gene bla TEM-1B and three transposable elements. Conjugation experiments showed that pQJDSal1 could transfer from S. Pullorum to Escherichia coli (E. coli) J53. Statistical analysis of 70 sequenced IncX1 plasmids revealed that IncX1 plasmids harbored various antibiotic resistance genes. The results highlight the importance of IncX1 plasmids in disseminating antibiotic resistance genes. Highlights • The first time to identify an IncX1 plasmid in Salmonella Pullorum • A new Tn 6168 _like transposon is identified. • Tn 1772 is inserted into Tn 2. • Madding statistics on the antibiotic resistance genes in IncX1 plasmids [ABSTRACT FROM AUTHOR]

Objectives To determine the structure of two multidrug-resistant IncHI1 plasmids carrying blaCTX-M-1 in Escherichia coli isolates disseminated in an equine clinic in the Czech Republic. Methods A complete nucleotide sequencing of 239 kb IncHI1 (pEQ1) and 287 kb IncHI1/X1 (pEQ2) plasmids was performed using the 454-Genome Sequencer FLX system. The sequences were compared using bioinformatic tools with other sequenced IncHI1 plasmids. Results A comparative analysis of pEQ1 and pEQ2 identified high nucleotide identity with the IncHI1 type 2 plasmids. A novel 24 kb module containing an operon involved in short-chain fructooligosaccharide uptake and metabolism was found in the pEQ backbones. The role of the pEQ plasmids in the metabolism of short-chain fructooligosaccharides was demonstrated by studying the growth of E. coli cells in the presence of these sugars. The module containing the blaCTX-M-1 gene was formed by a truncated macrolide resistance cluster and flanked by IS26 as previously observed in IncI1 and IncN plasmids. The IncHI1 plasmid changed size and gained the quinolone resistance gene qnrS1 as a result of IS26-mediated fusion with an IncX1 plasmid. Conclusions Our data highlight the structure and evolution of IncHI1 from equine E. coli. A plasmid-mediated sugar metabolic element could play a key role in strain fitness, contributing to the successful dissemination and maintenance of these plasmids in the intestinal microflora of horses. [ABSTRACT FROM PUBLISHER]

Abstract: The large conjugative multidrug resistance (MDR) plasmid pOLA52 was sequenced and annotated. The plasmid encodes two phenotypes normally associated with the chromosomes of opportunistic pathogens, namely MDR via a resistance-nodulation-division (RND)-type efflux-pump (oqxAB), and the formation of type 3 fimbriae (mrkABCDF). The plasmid was found to be 51,602bp long with 68 putative genes. About half of the plasmid constituted a conserved IncX1-type backbone with predicted regions for conjugation, replication and partitioning, as well as a toxin/antitoxin (TA) plasmid addiction system. The plasmid was also classified as IncX1 with incompatibility testing. The conjugal transfer and plasmid maintenance regions of pOLA52 therefore seem to represent IncX1 orthologues of the well-characterized IncX2 plasmid R6K. Sequence homology searches in GenBank also suggested a considerably higher prevalence of IncX1 group plasmids than IncX2. The 21 kb ‘genetic load’ region of pOLA52 was shown to consist of a mosaic, among other things a fragmented Tn3 transposon encoding ampicillin resistance. Most notably the oqxAB and mrkABCDF cassettes were contained within two composite transposons (Tn6010 and Tn6011) that seemed to originate from Klebsiella pneumoniae, thus demonstrating the capability of IncX1 plasmids of facilitating lateral transfer of gene cassettes between different Enterobacteriaceae. [Copyright &y& Elsevier]

Escherichia coli, the main host of the CTX-M-15 extended-spectrum β-lactamase (ESBL) enzyme, is widely distributed and exchanged between the environment, animals and humans. Therefore, identification of bla CTX-M-15 -positive lineages in food has a significant impact on public health. In this regard, until the end of 1990s, ESBL-producing isolates were mainly associated with hospital-acquired infections, with a predominance of SHV- and TEM-type enzymes. In recent years, a new trend has been observed among ESBL-producers, where most isolates now harbour CTX-M-type, being further isolated from community-acquired infections. Nowadays, CTX-M-15 has been recognised as the most important ESBL variant, invading virtually all human and animal compartments, leading to a global pandemic. Thus, whilst the rapid emergence and dissemination of CTX-M-15 among E. coli isolates has generated a large genetic reservoir from which other members of the Enterobacteriaceae family can easily acquire this resistance gene, there are an increasing number of new reservoirs and transmission mechanisms that must be investigated. In this study, we present the draft genome sequence of a CTX-M-15-producing E. coli ST345 isolated from commercial chicken meat in Brazil. This draft genome can be used as a reference sequence for comparative analysis among CTX-M-15-producers., (Copyright © 2017. Published by Elsevier Ltd.)