IncHI2 Plasmid Research Articles

Genotypic Diversity among Salmonella Typhi Isolated from Children Living in Informal Settlements in Nairobi, Kenya.

The persistence of multidrug-resistant (MDR) Salmonella Typhi (S. Typhi) is a challenge especially in regions where typhoid is endemic. Surveillance of circulating genotypes of MDR S. Typhi is crucial in typhoid acute cases and carriers. This study aimed to investigate genotypic diversity of S. Typhi from symptomatic and asymptomatic children in endemic settings in Nairobi, Kenya. Symptomatic and asymptomatic individuals' ≤ 16 years were recruited at four health facilities and tested for typhoid through stool cultures. The S. Typhi isolates were subjected to antibiotic susceptibility testing to investigate multidrug resistance. The MDR S. Typhi isolates' DNA was extracted and illumina sequenced. Raw reads were de novo assembled and analyzed by pathogen-watch. From the 90 sequenced isolates, 60 (67%) were confirmed to be S. Typhi (sequence Type 1 and genotype 4.3.1). Out of the 60 S. Typhi strains; 39 (65%) had plasmids, from these 38 (97%) had IncHI1 plasmids alone. Out of the 60, 59 (98%) S. Typhi isolates had bla TEM-1D . Point mutations conferring reduced susceptibility to quinolones were detected in 42 (70%) of S. Typhi isolates, from these; 14 (33%) had gyrA S83Y, and 28 (67%) gyrB S464F genes, respectively. This study reports 4.3.1 (H58) as the most dominant S. Typhi genotype responsible for spread of MDR phenotypes carried on IncHI1 plasmids. Presence of MDR S. Typhi with resistance genes such as bla TEM-1D and reduced susceptibility to ciprofloxacin especially among asymptomatic individuals, reiterates the need for use of typhoid conjugate vaccine among vulnerable children as a control and prevention measure against typhoid.

Genomic characterisation of Escherichia coli isolated from poultry at retail through Sink Surveillance in Dhaka, Bangladesh reveals high levels of multi-drug resistance.

The surveillance of antimicrobial resistance (AMR) in commensal Escherichia coli from livestock at slaughter is widely employed to assess the potential for risk to humans. There is currently a limited understanding of AMR in Bangladesh poultry at retail in live bird markets, with studies focussing solely on phenotypic characterisation of resistance. To address this evidence gap we performed antimicrobial susceptibility testing and whole genome sequencing on E. coli obtained from chickens from live bird markets in Dhaka in 2018 (n = 38) and 2020 (n = 45). E. coli were isolated from caeca samples following ISO guidelines and sequenced using short and long read methods. Multidrug resistance was extremely common (n = 77) and there was excellent concordance between AMR phenotype and the presence of corresponding AMR genes or mutations. There was considerable genomic diversity, with 43 different sequence types detected. Public health considerations included the high occurrence of resistance to ciprofloxacin (n = 75) associated with plasmid-residing qnrS or mutations in the gyrA and parC chromosomal genes; and the detection of a tigecycline resistant isolate harbouring tet(X4) on an IncHI1A/B-IncFIA mosaic plasmid. Thirty-nine isolates were resistant to azithromycin and harboured mphA, with a significant increase in the incidence of resistance between 2018 and 2020. Although azithromycin is banned for veterinary use in Bangladesh it remains an important treatment option for humans. Interestingly, mphA confers high-level resistance to azithromycin and erythromycin, and the latter is commonly used on poultry farms in Bangladesh. Seven isolates were colistin resistant and carried mcr1. For two isolates hybrid assemblies revealed that mcr1 resided on a highly conserved IncHI2 plasmid that had 93% nucleotide identity to a plasmid from the published genome of an E. coli isolate of Bangladeshi human origin. Six isolates had resistance to third generation cephalosporins, associated with plasmid-residing bla CTX-M-55, bla CTX-M-65, or bla DHA-1. By employing phenotypic and genomic approaches for AMR surveillance we have provided new insights into the potential for One Health AMR linkages in Bangladesh. Employing similar approaches in human and environmental sectors will help inform the One Health approach to addressing AMR, and generate evidence to support mitigation measures such as improved antimicrobial stewardship.

Extensively drug-resistant Enterobacter ludwigii co-harbouring MCR-9 and a multicopy of blaIMP-1 in South Korea

In this study, we describe an Enterobacter ludwigii clinical isolate that is resistant to both carbapenems and colistin in South Korea. Antimicrobial susceptibility testing revealed that E. ludwigii CRE2104-31 was non-susceptible to all tested antibiotics except fosfomycin. Whole genome sequencing identified a 323-kbp IncHI2 plasmid, pCRE2104-31a, that was co-harbouring mobile colistin resistance (mcr)-9.1 and blaIMP-1. In comparison with other full plasmids, pCRE2104-31a exhibited the closest similarity to a plasmid from the Klebsiella pneumoniae strain CNR48 from France, with 19.9% query coverage and 99% identity. Notably, we observed five tandem repeats of blaIMP-1 and aac(6′)-Il genes, accompanied by multiple attCs within a class I integron on the Tn402-like transposon. The unit of blaIMP-1-attC-aac(6′)-Il-attC might have accumulated due to multiple convergent events. In addition to mcr-9.1 and blaIMP-1, various other antibiotic resistance-associated genes were identified in the plasmid, as follows: blaTEM-1B, aph(3′)-I, aph(3′)-Ia, aac(6′)-Il, aac(6′)-IIc, aac(6′)-IIa, aph(6)-Id, aph(3′')-Ib, aadA2b, aac(6′)-Ib3, sul, dfrA19, qnrB2, aac(6′)-Ib-cr, ere(A), and qacE. A conjugation assay showed that the mcr-9.1/blaIMP-1-co-bearing plasmid was self-transmissible to E. coli J53. However, colistin and carbapenem resistance could not be transferred to E. coli due to high incompatibility. The convergence of mcr and carbapenemase genes is thought to be host-dependent among Enterobacteriaceae. The emergence of extensively drug-resistant E. ludwigii co-harbouring MCR-9.1 and a multicopy of blaIMP-1 would pose a significant threat within the compatible Enterobacteriaceae.

Salmonella Typhimurium with Eight Tandem Copies of blaNDM-1 on a HI2 Plasmid.

Carbapenem-resistant Salmonella has recently aroused increasing attention. In this study, a total of four sequence type 36 Salmonella enterica subsp. enterica serovar Typhimurium (S. Typhimurium) isolates were consecutively isolated from an 11-month-old female patient with a gastrointestinal infection, of which one was sensitive to carbapenems and three were resistant to carbapenems. Via antibiotic susceptibility testing, a carbapenemases screening test, plasmid conjugation experiments, Illumina short-reads, and PacBio HiFi sequencing, we found that all four S. Typhimurium isolates contained a blaCTX-M-14-positive IncI1 plasmid. One carbapenem-sensitive S. Typhimurium isolate then obtained an IncHI2 plasmid carrying blaNDM-1 and an IncP plasmid without any resistance genes during the disease progression. The blaNDM-1 gene was located on a new 30 kb multiple drug resistance region, which is flanked by IS26 and TnAs2, respectively. In addition, the ST_F0903R isolate contained eight tandem copies of the ISCR1 unit (ISCR1-dsbD-trpF-ble-blaNDM-1-ISAba125Δ1), but an increase in MICs to carbapenems was not observed. Our work further provided evidence of the rapid spread and amplification of blaNDM-1 through plasmid. Prompting the recognition of carbapenem-resistant Enterobacterales and the initiation of appropriate infection control measures are essential to avoid the spread of these organisms.

Molecular characteristic of mcr-1 gene in Escherichia coli from aquatic products in Guangdong, China

Aquatic ecosystems serve as a dissemination pathway and a reservoir of both antibiotic resistant bacteria (ARB) and antibiotic resistance genes (ARGs). This study aimed to determine the prevalence of colistin-resistant mcr-like genes in Enterobacteriales in aquatic products, which may be contribute to the transfer of ARGs in water environments. The mcr-1-positive Escherichia coli were recovered from 123 freshwater fish and 34 cultured crocodile cecum samples from 10 farmers' markets in Guangdong, China. Minimum inhibitory concentration (MIC) was determined using the agar dilution method. Genotyping was performed using pulsed-field gel electrophoresis (PFGE) and multi-locus sequence typing (MLST). Conjugation assay was carried out to investigate the transferability of mcr-1. Genomic information was obtained by whole genome sequencing (WGS) and bioinformatic analysis. Forty-four mcr-1 positive isolates showed co-resistance to tetracycline, trimethoprim/sulfamethoxazole, and gentamicin, while they were all sensitive to tigecycline, meropenem, and amikacin. They were typed into sixteen PFGE clusters. ST10 and ST117 were the most popular sequence types, followed by ST1114. S1-PFGE verified the presence of the mcr-1 gene on plasmids in sizes of ∼60 kb (n = 1) and ∼240 kb (n = 3). Whole genome sequencing-based analysis identified mcr-1 integrated in IncHI2 plasmid (n = 3), IncI2 plasmid (n = 2), and bacterial chromosome in two copies (n = 1). In addition to mcr-1, they carried several other antibiotic resistance genes, such as blaCTX-M-14, fosA3, and aac(6')-Ib-cr. These data suggest that aquatic products are an important antibiotic resistance reservoir and highlight possible risks regarding food safety.

Occurrence and characterization of NDM-5-producing Escherichia coli from retail eggs

The New Delhi Metallo-β-lactamase (NDM) producing Enterobacterales has been detected from diverse sources but has rarely been reported in retail eggs. In this study, 144 eggshell and 96 egg content samples were collected in 2022 from Guangdong province and were screened for NDM-producing strains. Four Escherichia coli strains (ST3014, ST10, ST1485, and ST14747) recovered from two (1.39%, 2 of 144) eggshells and two (2.08%, 2 of 96) egg content samples were identified as blaNDM−5-positive strains. Oxford Nanopore MinION sequencing and conjugation assays revealed that the blaNDM−5 gene was carried by IncX3 (n = 1), IncI1 (n = 1), and IncHI2 (n = 2). The IncI1-plasmid-carrying blaNDM−5 displayed high homology with one plasmid pEC6563-NDM5 from the human clinic, while the IncHI2 plasmid harboring blaNDM−5 shared highly similar structures with plasmids of animal origin. To the best of our knowledge, this is the first report on the identification of blaNDM−5-positive bacteria in retail eggs. NDM-producing E. coli could be transmitted to humans by the consumption of eggs or direct contact, which could pose a potential threat to human health.

Genetic characterization of a multidrug-resistant Salmonella enterica serovar Agona isolated from a dietary supplement in Germany.

Salmonella enterica subsp. enterica serovar Agona has a history of causing food-borne outbreaks and any emergence of multidrug-resistant (MDR) isolates in novel food products is of concern. Particularly, in food products frequently consumed without sufficient heating prior to consumption. Here, we report about the MDR isolate, 18-SA00377, which had been isolated from a dietary supplement in Germany in 2018 and submitted to the German National Reference Laboratory for Salmonella. WGS-based comparative genetic analyses were conducted to find a potential reservoir of the isolate itself or mobile genetic elements associated with MDR. As a phylogenetic analysis did not yield any closely related S. Agona isolates, either globally or from Germany, a detailed analysis of the largest plasmid (295,499 bp) was performed as it is the main carrier of resistances. A combined approach of long-read and short-read sequencing enabled the assembly of the isolate's chromosome and its four plasmids. Their characterization revealed the presence of 23 different antibiotic resistance genes (ARGs), conferring resistance to 12 different antibiotic drug classes, as well as genes conferring resistance to six different heavy metals. The largest plasmid, pSE18-SA00377-1, belongs to the IncHI2 plasmid family and carries 16 ARGs, that are organized as two distinct clusters, with each ARG associated with putative composite transposons. Through a two-pronged approach, highly similar plasmids to pSE18-SA00377-1 were identified in the NCBI database and a search for Salmonella isolates with a highly similar ARG resistance profile was conducted. Mapping and structural comparisons between pSE18-SA00377-1 and these plasmids and Salmonella isolates showed that both the plasmid backbone and identical or similar ARG clusters can be found not only in Salmonella isolates, originating mostly from a wide variety of livestock, but also in a diverse range of bacterial genera of varying geographical origins and isolation sources. Thus, it can be speculated that the host range of pSE18-SA00377-1 is not restricted to Salmonella and its spread already occurred in different bacterial populations. Overall, this hints at a complex history for pSE18-SA00377-1 and highlights the importance of surveilling multidrug-resistant S. enterica isolates, especially in novel food items that are not yet heavily regulated.

Genomics analysis of KPC-2 and NDM-5-producing Enterobacteriaceae in migratory birds from Qinghai Lake, China.

The study examined the epidemiological characteristics of carbapenem-resistant Enterobacteriaceae (CRE) isolated from migratory birds and surroundings in Qinghai Lake, China. We identified 69 (15.7%) CRE isolates from a total of 439 samples including 29 (6.6%) blaNDM-5 Escherichia coli and 40 (9.1%) blaKPC-2 Klebsiella pneumoniae. WGS analysis indicated that ST746, ST48, ST1011, and ST167 were the primary sequence types (ST) for blaNDM-5 E. coli, while all blaKPC-2K. pneumoniae were ST11 and harbored numerous antibiotic resistance gene types including blaCTX-M, qnrS, and rmtB. A phylogenetic tree based on core genomes revealed that blaNDM-5 E. coli was highly heterogeneous while the blaKPC-2K. pneumoniae was highly genetically similar within the group and to human Chinese isolates. IncX3, IncHI2, and IncFIB-HI2 plasmid replicon types were associated with blaNDM-5 spread, while IncFII-R and IncFII plasmids mediated blaKPC-2 spread. We also identified IncFII-R hybrid plasmids most likely formed by IS26-mediated integration of IncFII into IncR plasmid backbones. This also facilitated the persistence of IncFII-R plasmids and antibiotic resistance genes including blaKPC-2. In addition, all of the blaKPC-2K. pneumoniae isolates harbored a pLVKP-like virulence plasmid carrying a combination of two or more hypervirulence markers that included peg-344, iroB, iucA, rmpA, and rmpA2. This is the first description of ST11 K. pneumoniae that co-carried blaKPC-2- and pLVKP-like virulence plasmids from migratory birds. The blaKPC-2K. pneumoniae carried by migratory birds displayed high genetic relatedness to human isolates highlighting a high risk of transmission of these K. pneumoniae. KEY POINTS: • Multidrug resistance plasmids (blaKPC-2, bla436NDM-5, blaCTX-M, qnrS, and rmtB). • Co-occurrence of plasmid-mediated resistance and virulence genes. • High similarity between migratory bird genomes and humans.

Molecular characterization of highly prevalent Escherichia coli and Escherichia marmotae resistant to extended-spectrum cephalosporins in European starlings (Sturnus vulgaris) in Tunisia.

European starlings are widespread migratory birds that have already been described as carrying bacteria resistant to extended-spectrum cephalosporins (ESC-R). These birds are well known in Tunisia because they spend the wintertime in this country and are hunted for human consumption. The goal of our study was to estimate the proportion of ESC-R in these birds and to characterize the collected isolates using whole-genome sequencing. Results showed that 21.5% (42/200) of the birds carried either an extended-spectrum beta-lactamase (ESBL) or an acquired AmpC gene. Diverse bla CTX-M genes were responsible for the ESBL phenotype, bla CTX-M-14 being the most prevalent, while only bla CMY-2 and one bla CMY-62 were found in AmpC-positive isolates. Likewise, different genetic determinants carried these resistance genes, including IncHI2, and IncF plasmids for bla CTX-M genes and IncI1 plasmids for bla CMY-2 genes. Three chromosomally encoded bla CTX-M-15 genes were also identified. Surprisingly, species identification revealed a large proportion (32.7%) of Escherichia marmotae isolates. This species is phenotypically indistinguishable from Escherichia coli and has obviously the same capacity to acquire ESC-R genes. Our data also strongly suggest that at least the IncHI2/pST3 plasmid can spread equally between E. coli and E. marmotae. Given the potential transmission routes between humans and animals, either by direct contact with dejections or through meat preparation, it is important to closely monitor antimicrobial resistance in European starlings in Tunisia and to set up further studies to identify the sources of contamination of these birds. IMPORTANCE The One Health concept highlighted knowledge gaps in the understanding of the transmission routes of resistant bacteria. A major interest was shown in wild migratory birds since they might spread resistant bacteria over long distances. Our study brings further evidence that wild birds, even though they are not directly submitted to antibiotic treatments, can be heavily contaminated by resistant bacteria. Our results identified numerous combinations of resistance genes, genetic supports, and bacterial clones that can spread vertically or horizontally and maintain a high level of resistance in the bird population. Some of these determinants are widespread in humans or animals (IncHI2/pST3 plasmids and pandemic clones), while some others are less frequent (atypical IncI1 plasmid and minor clones). Consequently, it is essential to be aware of the risks of transmission and to take all necessary measures to prevent the proportions of resistant isolates from increasing uncontrollably.

Molecular epidemiological characteristics of E. coli strains carrying the mcr-1 gene isolatedfrom healthy human feces, domestic animal feces,food and water in Thanh Ha commune, Thanh Liem district, Ha Nam province

Colistin is a critically important antimicrobial for the treatment of carbapenem-resistant gram-negative intestinal infections. E. coli strains carrying the colistin-resistant mediated colistin resistance (mcr-1)gene have been globally distributed, found in humans, animals and the environment with diverse resistance mechanisms, namely intrinsic, mutation, and horizontal gene transfer [1]. Evaluation of the epidemiological characteristics and transmission status of E. coli strains to carry the mcr-1 gene is urgent. Sequencing results of 87 community E. coli strains isolated from healthy human feces and environment showed that the mcr-1 gene of these strains in the study was found on both chromosomes and plasmids, the ratio of mcr-1 plasmid-mediated is much higher than on the chromosome (71.3% vs 26.4%, p<0.001). 100% of E. coli strains that carry the mcr-1 gene were subtype mcr-1.1. The plasmids diversified in size (from <50 kb to >300 kb). There were 7 types of single replicon carrying mcr-1 gene detected from 29 strains, including IncI2 (n=7, 10.9%), IncP (n=7, 10.9%), IncX4 (n=5, 7.8%), IncFIA (n=3, 4.7%), IncHI1B (n=3, 4.7%), IncN (n=1, 1.6%) and IncX1 (n=1, 1.6%), were found in healthy humans feces, animals and household water. There were also 21 different multi-replicons with combinations of IncFIA, IncFIB, IncHI1B, IncHI2, IncN, IncY, IncX1 and IncI2 plasmids, found in 31 strains of E.coli in humans, animals and water. Strains were classified into 56 different sequence types (STs). ST10 (n=10, 11%), ST48 (n=9, 9%) and ST206 (n=8, 8%) were common STs.

Characteristics of Two mcr-1-Harboring IncHI2 Plasmids from Clinical Salmonella Isolates in Jiaxing City.

Salmonella is a primary cause of foodborne diseases, and the increasing prevalence of mcr-1-carrying plasmids, which confer colistin resistance to Salmonella, poses significant global health concerns. As the frequency of occurrence of the mcr-1 gene is increasing globally, we studied the prevalence of mcr-1 in clinical Salmonella isolates by analyzing 195 clinical strains isolated in 2020. Of the 195 Salmonella isolates, 41 isolates were resistant to colistin. We found mcr-1 in two strains (Salmonella Typhimurium ZJJX20006 and Salmonella Kentucky ZJJX20014), which we analyzed in detail via whole-genome sequencing and antibiotic susceptibility testing. Two strains displayed resistance to ampicillin, ampicillin-sulbactam, tetracycline, chloramphenicol, and cotrimoxazole, while ZJJX20006 displayed resistance to colistin and ZJJX20014 was sensitive. Genomic analysis revealed that these strains had plasmid-encoded mcr-1 in IncHI2 plasmids, which were not similar to the mcr-1-IncX4 identified in 2016. These two strains also harbored other drug resistance genes, including blaOXA-1 and blaCTX-M-14. Our findings may help clarify the molecular mechanisms of mcr-1 dissemination among Salmonella strains in Jiaxing City and offer insights into the evolution of mcr-1 in Salmonella.

The characteristics of mcr-bearing plasmids in clinical Salmonella enterica in Sichuan, China, 2014 to 2017.

Salmonella is one of the most important zoonotic pathogens and a major cause of foodborne illnesses, posing a serious global public health hazard. The emergence of plasmid-mediated mcr genes in Salmonella has greatly reduced the clinical choice of salmonellosis treatment. The aim of this study was to investigate the plasmid characteristics of mcr-positive Salmonella identified from patients in Sichuan, China during 2014 to 2017 by whole genomes sequencing. In this study, a total of 12 mcr-positive isolates (1.15%, ; mcr-1, n=10; mcr-3, n=2) were identified from 1046 Salmonella isolates using PCR. Further characterization of these isolates was performed through antimicrobial susceptibility testing, conjugation assays, whole genome sequencing, and bioinformatics analysis. The mcr-1 gene in these isolates were carried by three types of typical mcr-1-bearing plasmids widely distributed in Enterobacteriaceae (IncX4, IncI2 and IncHI2). Of note, two mcr-1-harboring IncHI2 plasmids were integrated into chromosomes by insertion sequences. Two mcr-3-bearing plasmids were IncC and IncFIB broad-host-range plasmids respectively. Genetic context analysis found that mcr-1 was mainly located in Tn6330 or truncated Tn6300, and mcr-3 shared a common genetic structure tnpA-mcr-3-dgkA-ISKpn40. Overall, we found that mcr gene in clinical Salmonella were commonly carried by broad-host plasmids and have potential to transfer into other bacteria by these plasmids. Continuous surveillance of MDR Salmonella in humans and investigation the underlying transmission mechanisms of ARGs are vital to curb the current severe AMR concern.

Broad-Host Dissemination of Plasmids Coharboring the fos Operon for Fructooligosaccharide Metabolism with Antibiotic Resistance Genes.

The fos operon encoding short-chain fructooligosaccharide (scFOS) utilization enables bacteria of the family Enterobacteriaceae to grow and be sustained in environments where they would struggle to survive. Despite several cases of the detection of the fos operon in isolates of avian and equine origins, its global distribution in bacterial genomes remains unknown. The presence of the plasmid-harbored fos operon among resistant bacteria may promote the spread of antibiotic resistance. A collection of 11,538 antimicrobial-resistant Enterobacteriaceae isolates from various sources was screened for the fosT gene encoding the scFOS transporter. Out of 307 fosT-positive isolates, 80% of them originated from sources not previously linked to fosT (humans, wastewater, and animals). The chromosomally harbored fos operon was detected in 163/237 isolates subjected to whole-genome sequencing. In the remaining 74 isolates, the operon was carried by plasmids. Further analyses focusing on the isolates with a plasmid-harbored fos operon showed that the operon was linked to various incompatibility (Inc) groups, including the IncHI1, IncF-type, IncK2, IncI1, and IncY families. Long-read sequencing of representative plasmids showed the colocalization of fos genes with antibiotic resistance genes (ARGs) in IncHI1 (containing a multidrug resistance region), IncK2 (blaTEM-1A), IncI1 [sul2 and tet(A)], and IncY [aadA5, dfrA17, sul2, and tet(A)] plasmids, while IncF-type plasmids had no ARGs but coharbored virulence-associated genes. Despite the differences in the locations and structures of the fos operons, all isolates except one were proven to utilize scFOSs. In this study, we show that the fos operon and its spread are not strictly bound to one group of plasmids, and therefore, it should not be overlooked. IMPORTANCE It was believed that members of the family Enterobacteriaceae are unable to grow under conditions with short-chain fructooligosaccharides as the only source of carbon. Nevertheless, the first Escherichia coli isolate from chicken intestine was able to utilize these sugars owing to the chromosomally harbored fos operon. Studies on E. coli isolates from horses discovered the horizontal transfer of the fos operon on IncHI1 plasmids along with genes for antibiotic resistance. The first plasmid detected was pEQ1, originating from the feces of a hospitalized horse in the Czech Republic. Follow-up studies also revealed the dissemination of the IncHI1 plasmid-harbored fos operon in the Netherlands, Germany, Denmark, and France among healthy horses. Despite several cases of detection of the fos operon, its global distribution in bacterial genomes remains unknown. The fos operon possibly plays a role in the adaptation of plasmids among resistant bacteria and therefore may promote the spread of antibiotic resistance.

Persistence and dynamic structures of diverse cephalosporinase genes in nontyphoidal Salmonella in cross-sectional surveillance in Taiwan

Nontyphoidal Salmonella (NTS) is a major foodborne pathogen causing from acute gastroenteritis to bacteraemia, particularly in paediatric and elderly patients. Antimicrobial resistance of NTS, especially resistance to extended-spectrum cephalosporins, has emerged over the past decades. Thirteen NTS isolates resistant to ceftriaxone or cefotaxime were collected from a teaching hospital in Taipei, and another three from a tertiary hospital, in New Taipei City, Taiwan, from September 2018 to December 2019. Ten other archived isolates from 2000 to 2017 were also obtained. Complete genomes of the 26 isolates were obtained. Serovars, sequence types, resistomes, genetic relatedness, and sequence comparison of plasmids were analyzed. Serogroups B, C2 and E were significantly associated with ampicillin resistance. Over 90% of these 26 isolates are susceptible to carbapenems and colistin. Genomic epidemiology of these isolates shows that blaCMY-2-harbouring isolates in different serovars were prevalent over two decades, presumably resulting from highly mobile IncI1 plasmid harbouring blaCMY-2. One type of the IncI1 plasmids contained a mobile element, IS26, which might be involved in the acquisition of antimicrobial resistance genes. Two emerging serovars, S. Goldcoast ST358 harbouring blaCTX-M-55 on IncHI2 plasmids and S. Anatum ST64 harbouring blaDHA-1 on IncA/C2 plasmids persisted in Taiwan, possibly through the clonal spread. Integration of complete or partial plasmid sequences into host chromosomes or multiplications of the antimicrobial resistance genes also appears to be mediated by IS26, in the two emerging clones. The dynamic movement of cephalosporinase genes mediated by IS26 in NTS is of great concern.

Genomic characterisation of a blaKPC-2- and mcr-10-co-harbouring Enterobacter kobei isolate with high-level resistance to colistin and carbapenems

The emergence and spread of colistin resistance in carbapenem-resistant Enterobacteriaceae pose a serious threat to human and animal health. This work aimed to characterize the genetic features of antimicrobial resistance of a carbapenem and colistin resistant Enterobacter kobei SCLZS19 from hospital sewage by using whole genome sequencing. Antimicrobial susceptibility tests were performed by using the disk diffusion method. Whole genome sequencing of SCLZS19 was carried out on the HiSeq 2000 combined with PacBio RSII platforms. Sequence type, plasmid incompatibility types, resistance genes, and insertion elements were identified using the MLST, PlasmidFinder, ResFinder, and ISfinder, respectively. Conjugation assays were performed using both broth- and filter-based methods with the azide-resistant Escherichia coli J53 as the recipient. The function of the mcr-9-like variant was determined by gene cloning. E. kobei SCLZS19 has a 4,862,177-bp circular chromosome, and nine circular plasmids ranging in size from 4,120 bp to 282,472 bp. It carries 11 antibiotic resistance genes (ARGs), and 10 of them are located on plasmids. The colistin resistance gene mcr-10 is located on a 118,766-bp non-transferable IncF (Y3:A-:B-) plasmid. The carbapenemase gene blaKPC-2 is carried by a self-transmissible IncP6 plasmid which is epidemic in China. In addition, SCLZS19 also carries a mcr-9-like variant on a IncHI2 (ST1) plasmid. Cloning assay shows that the mcr-9-like variant does not mediate colistin resistance in E. coli DH5α. The findings highlight that carbapenem- and colistin-resistant Enterobacterales from water environment may serve as a reservoir for clinically significant ARGs, and continuous surveillance is required.

Prevalence and molecular characterization of mcr-1-positive foodborne ST34-Salmonella isolates in China

Salmonella enterica serovar Typhimurium (S. Typhimurium) and S. 4,[5],12:i:- have become the most common serovars associated with human salmonellosis worldwide. Moreover, the emergence of mcr-carrying S. Typhimurium and S. 4,[5],12:i:- with multidrug resistance (MDR) patterns has posed a threat to public health. In this study, we retrospectively screened 2009–2022 laboratory-preserved strains for the presence of mcr genes. We obtained 16 mcr-1-positive S. Typhimurium and S. 4,[5],12:i:- strains with MDR that belonged to sequence type 34 (ST34). Whole-genome sequencing analysis revealed that the mcr-1 was located on the IncI2 or IncHI2 plasmids. The ISApl1 element downstream of mcr-1 was present in all pig-derived strains. Conjugation experiments confirmed that nine mcr-1-carrying IncHI2 plasmids could not be transferred to Escherichia coli due to loss of the conjugation region. Finally, core genome single nucleotide polymorphism (cgSNP) analyses of the 16 mcr-1-carrying strains and 77 mcr-carrying ST34-Salmonella genome sequences from the NCBI and ENA databases showed that five out of eight clusters contained strains from pig and pig products, revealing pigs and pig products as key reservoirs of mcr-1-positive ST34-Salmonella strains. The transmission of mcr-carrying ST34 Salmonella strains to humans via the pig food chain is a potential cause for public health concern in controlling human salmonellosis.

Emergence of chromosomally located blaCTX-M-14b and qnrS1 in Salmonella enterica serotype Kentucky ST198 in China

Highly fluoroquinolone-resistant Salmonella enterica serotype Kentucky has become widespread in recent years, largely associated with the spread of sequence type 198 (ST198), which often leads to multidrug resistance. Research on the genomic epidemiology of Salmonella Kentucky in China is currently uncommon. In this study, we analysed the genomic epidemiology and antimicrobial resistance characteristics of Salmonella Kentucky ST198 collected from foodborne disease surveillance in Shenzhen, China, during 2010-2021, using whole-genome sequencing and antibiotic susceptibility testing. In addition, 158 global Salmonella Kentucky ST198 genomes were included for comparison. Among 8559 Salmonella isolates, 43 Salmonella Kentucky ST198 isolates were detected during 2010-2021. The global Salmonella Kentucky ST198 evolutionary tree was divided into five clades, with Shenzhen isolates distributed in clades 198.1, 198.2-1 and 198.2-2, mainly clustered with Chinese strains. Strains in clade 198.2 dominated in Shenzhen and all of them showed multidrug resistance. Nine strains showed high resistance to ceftriaxone, which was associated with blaCTX-M-14b in clade 198.2-1, which was demonstrated to be located on the chromosome. Fifteen strains showed high resistance to ciprofloxacin, which was associated with carriage of qnrS1 in clade 198.2-2. qnrS1 was first located on an IncHI2 plasmid and then transferred into the chromosome. Here we report the genomic and antimicrobial resistance characterisation of Salmonella Kentucky ST198 in Shenzhen. Of particular concern, we identified for the first time a clade 198.2-1 isolate carrying blaCTX-M-14b as well as chromosomally located qnrS1 in clade 198.2-2 of Salmonella Kentucky ST198 in China, highlighting the necessity of surveillance of clade 198.2.

Highly drug resistant clone of Salmonella Kentucky ST198 in clinical infections and poultry in Zimbabwe

A highly multidrug-resistant strain of Salmonella enterica serotype Kentucky (S. Kentucky) of sequence type (ST)198 emerged in North Africa and has since spread widely. To investigate the genetic diversity and phylogenetic relationship of S. Kentucky in Zimbabwe and identify potential sources of infection, the whole-genome sequence of 37 S. Kentucky strains isolated from human clinical infections and from poultry farms between 2017 and 2020 was determined. Of 37 S. Kentucky isolates, 36 were ST198 and one was ST152. All ST198 isolates had between six and fifteen antimicrobial resistance (AMR) genes, and 92% carried at least ten AMRs. All ST198 isolates harbored the Salmonella genomic island K-Israel variant (SGI1-KIV) integrated into the chromosome with aac(3)-ld, aac(6)-laa, aadA7, blaTEM-1, sul1, and tetA genes, with occasional sporadic loss of one or more genes noted from five isolates. All ST198 isolates also had mutations in the quinolone resistance-determining region of the gyrA and parC genes. The blaCTX-M-14.1 and fosA3 genes were present in 92% of the ST198 isolates, conferring resistance to extended-spectrum cephalosporins and fosfomycin, respectively, were present on an IncHI2 plasmid with the aadA2b, aadA1, aph(3’)-Ib, aph(6’)-Id, cmlA1 and sul3 AMR genes. S. Kentucky ST198 isolates from Zimbabwe formed a closely related phylogenetic clade that emerged from a previously reported global epidemic population. The close genetic relationship and population structure of the human clinical and poultry isolates of ST198 in Zimbabwe are consistent with poultry being an important source of highly resistant strains of S. Kentucky in Zimbabwe.

Prevalence and genetic diversity of multidrug-resistant Salmonella Typhimurium monophasic variant in a swine farm from China.

Salmonella 4,[5],12:i:-, a monophasic variant of S. Typhimurium, has become a global serovar causing animal and human infections since its first emergence in the late 1980's. Several previous studies showed the increasing prevalence of S. 4,[5],12:i:- in China, most of which were from swine with multidrug resistance (MDR) profiles. However, the molecular characteristic and evolution of S. 4,[5],12:i:- in the same swine farm are still unknown. In this study, a total of 54 S. enterica strains were isolated from different fattening pigs aged 1, 3, and 6 months, most of which belonged to S. 4,[5],12:i:-. Whole-genome sequencing revealed that all 45 S. 4,[5],12:i:- strains belonged to ST34 and were further divided into two different ribosomal STs and nine different core-genome STs. Phylogenetic analysis of 286 S. 4,[5],12:i:- strains in China, including 241 from the EnteroBase Salmonella database, revealed the genetic diversity of S. 4,[5],12:i:- and indicated that S. 4,[5],12:i:- in this swine farm might have multiple origins. Three different IncHI2 plasmids carrying various resistance genes were characterized by nanopore sequencing and could be conjugated to Escherichia coli. The colistin resistance gene mcr-1 and ESBLs gene blaCTX - M-14 were co-located on the chromosome of one strain. The dynamic changes in antimicrobial resistance regions and transferability of IncHI2 plasmids, as well as the chromosomal location of resistance genes, facilitated the diversity of the antimicrobial resistance characteristics in S. 4,[5],12:i:-. Since the swine farm is regarded as the important reservoir of MDR S. 4,[5],12:i:-, the prevalence and evolution of S. 4,[5],12:i:- from swine farms to pig products and humans should be continually monitored.

First Detection of blaNDM-1-Haboring IncHI2 Plasmid in Escherichia coli Strain Isolated from Goose in China.

Carbapenem-resistant Enterobacteriaceae infections are among the most serious threats to human and animal health worldwide. Of the 1013 strains of Escherichia coli isolated and identified in 14 regions of China from 2007 to 2018, seven strains were resistant to meropenem and all were positive for blaNDM. The seven New Delhi metallo-β-lactamase (NDM)-positive strains belonged to five different sequence types, indicating that most of the NDM-positive strains were nonclonal. An IncHI2 plasmid carrying the blaNDM-1 element was identified in the C1147 strain from a goose source and reported for the first time, showing a specific structure. Conjugation experiments revealed that the IncHI2 plasmid was conjugatable, and the horizontal propagation of the plasmid led to the rapid propagation of NDM in the same and different strains. This study revealed that waterfowl, as a potential transmission factor for carbapenem-resistant blaNDM-1, poses a threat to human health.