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Phenotypic resistance to fluoroquinolones due to mutational changes in the gyrA and parC genes is common among clinical Vibrio strains; the plasmid-mediated quinolone resistance (PMQR) qnrVC genes were also suggested to play a role in enhancing resistance development. This study investigated the prevalence of qnrVC genes in foodborne Vibrio strains collected in Shenzhen, China, during the period August 2015 and April 2017. A total of 1811 foodborne Vibrio strains were collected, mostly (73.8%) from shrimp samples and 20.2% of these strains were resistant to ciprofloxacin. Investigation of resistance mechanisms showed that mutations in the gyrA and parC genes were commonly associated with ciprofloxacin resistance. The presence of qnrVC genes was shown to enhance ciprofloxacin MIC in Vibrio strains and 69.7% of Vibrio strains that harbored target mutations also carried qnrVC genes, yet only 27.5% of the isolates not harboring such mutations carried the qnrVC genes. A total of 141 strains were found to carry the qnrVC alleles, with qnrVC5 and qnrVC1 being the most common types. Fourteen qnrVC variant genes that contained novel mutations were detectable, with 12 (85.7%) involving qnrVC5-like alleles. For the first time, we found a variant that was likely formed by the recombination of qnrVC1 and qnrVC5. The genetic context of the qnrVC genes found in this study was highly variable, with most being accompanied by mobile genetic elements and other resistance genes. The increasing prevalence of qnrVC genes in Vibrio and its contribution on mediating the development of ciprofloxacin resistance need to be further investigated.

Carbapenem-resistant and hypervirulent K. pneumoniae (CR-HvKP) strains that have emerged recently have caused infections of extremely high mortality in various countries. In this study, we discovered a conjugative plasmid that encodes carbapenem resistance and hypervirulence in a clinical ST86 K2 CR-HvKP, namely 17ZR-91. The conjugative plasmid (p17ZR-91-Vir-KPC) was formed by fusion of a non-conjugative pLVPK-like plasmid and a conjugative blaKPC-2-bearing plasmid and is present dynamically with two other non-fusion plasmids. Conjugation of p17ZR-91-Vir-KPC to other K. pneumoniae enabled them to rapidly express the carbapenem resistance and hypervirulence phenotypes. More importantly, genome analysis provided direct evidence that p17ZR-91-Vir-KPC could be directly transmitted from K2 CR-HvKP strain, 17ZR-91, to ST11 clinical K. pneumoniae strains to convert them into ST11 CR-HvKP strains, which explains the evolutionary mechanisms of recently emerged ST11 CR-HvKP strains., Carbapenem-resistant and hypervirulent Klebsiella pneumoniae strains are emerging. Here Xie et al. show that these phenotypes are carried on a plasmid formed from the fusion of a virulence plasmid with a conjugative plasmid.

Objectives To investigate the prevalence and genetic contexts of the blaCTX-M-14 gene harboured by foodborne isolates of Vibrio spp. in China. Methods A total of 1856 Vibrio spp. isolates collected from raw meat and shrimp samples in Guangdong Province of China were screened for blaCTX-M-14 by PCR. The blaCTX-M-14-positive isolates were characterized by MIC, PFGE, MLST, conjugation, S1-PFGE and Southern blotting and WGS using Illumina and Nanopore platforms. Results A total of 35 (1.9%) Vibrio isolates were positive for blaCTX-M-14, including 33 Vibrio parahaemolyticus strains and two Vibrio alginolyticus strains. MLST showed that most of the blaCTX-M-14-bearing isolates could be assigned into two major STs, with ST163 being more prevalent (n = 23), followed by ST180 (n = 6). Whole-genome analysis of these 35 isolates revealed that the blaCTX-M-14 gene was associated with ISEcp1 in the upstream region, of which 32 blaCTX-M-14 genes were located in the same loci of chromosome I, 1 blaCTX-M-14 gene was located in a novel chromosomal integrative conjugative element (ICE) belonging to the SXT/R391 family and 2 blaCTX-M-14 genes were located in the same type of plasmid, which belonged to the IncP-1 group. Conjugation experiments showed that only the plasmid-borne blaCTX-M-14 gene could be transferred to the recipient strain Escherichia coli J53. Conclusions The emergence of the novel ICE and IncP-1 plasmids has contributed to the variable genetic contexts of blaCTX-M-14 among strains of Vibrio spp. and facilitated the horizontal transfer of such genes between Vibrio spp. and other zoonotic pathogens, resulting in a rapid increase in the prevalence of blaCTX-M-14-bearing bacterial pathogens worldwide.

In this study, we identified a conjugative plasmid carrying a novel azithromycin resistance gene, erm (42), from a clinical K. pneumoniae strain. Conjugation of this plasmid into Salmonella conjugants conferred resistance to azithromycin, which is considered a choice for treating Salmonella infections.

A plasmid that harbored the virulence factors highly like those of the virulence plasmid commonly found in clinical hypervirulent Klebsiella pneumoniae strains was detected in a foodborne Escherichia coli strain EC1108 and designated p1108-IncFIB. This virulent-like plasmid was found to be common in E. coli from various sources. To understand the contribution of this plasmid to the virulence of E. coli, plasmid p1108-IncFIB in strain EC1108 was first cured to generate strain EC1108-PC. The virulence plasmid p15WZ-82_Vir in Klebsiella pneumoniae strain 15WZ-82 was then transmitted to EC1108-PC to produce the transconjugant, EC1108-PC-TC to assess the contribution of this virulence plasmid to the virulence level of E. coli. During the process of conjugation, p15WZ-82_Vir was found to be evolved into p15WZ-82_int, which underwent homologous recombination with a plasmid encoding a carbapenemase gene

Resistance of nontyphoidal Salmonella to fluoroquinolones such as ciprofloxacin is known to be mediated by target mutations. This study surveyed the prevalence of Salmonella strains recovered from 2,989 food products in Shenzhen, China, during the period 2012 to 2017 and characterized the genetic features of several PMQR gene-bearing plasmids and ciprofloxacin resistance-encoding DNA fragments., The incidence of ciprofloxacin resistance in Salmonella has increased dramatically in the past decade. To track the evolutionary trend of ciprofloxacin resistance-encoding genetic elements during this period, we surveyed the prevalence of Salmonella in food products in Shenzhen, China, during the period of 2012 to 2017 and performed whole-genome sequencing and genetic analysis of 566 ciprofloxacin-resistant clinical Salmonella strains collected during this survey. We observed that target gene mutations have become much less common, with single gyrA mutation currently detectable in Salmonella enterica serovar Typhimurium only. Multiple plasmid-mediated quinolone resistance (PMQR) genes located in the chromosome and plasmids are now frequently detectable in ciprofloxacin-resistant Salmonella strains of various serotypes. Among them, the qnrS1 gene was often harbored by multiple plasmids, with p10k-like plasmids being the most dominant. Importantly, p10k-like plasmids initially were not conjugative but became transmissible with the help of a helper plasmid. Ciprofloxacin resistance due to combined effect of carriage of the qnrS1 gene and other resistance mechanisms is common. In S. Typhimurium, carriage of qnrS1 is often associated with a single gyrA mutation; in other serotypes, combination of qnrS1 and other PMQR genes located in the chromosomal fragment or plasmid is observed. Another major mechanism of ciprofloxacin resistance, mainly observable in S. Derby, involves a chromosomal fragment harboring the qnrS2–aac(6′)lb-cr–oqxAB elements. Intriguingly, this chromosomal fragment, flanked by IS26, could form a circular intermediate and became transferrable. To conclude, the increase in the incidence of various PMQR mobile genetic elements and their interactions with other resistance mechanism contribute to a sharp increase in the prevalence of ciprofloxacin-resistant clinical Salmonella strains in recent years. IMPORTANCE Resistance of nontyphoidal Salmonella to fluoroquinolones such as ciprofloxacin is known to be mediated by target mutations. This study surveyed the prevalence of Salmonella strains recovered from 2,989 food products in Shenzhen, China, during the period 2012 to 2017 and characterized the genetic features of several PMQR gene-bearing plasmids and ciprofloxacin resistance-encoding DNA fragments. The emergence of such genetic elements has caused a shift in the genetic location of ciprofloxacin resistance determinants from the chromosomal mutations to various mobile genetic elements. The distribution of these PMQR plasmids showed that they exhibited high serotype specificity, except for the p10k-like plasmids, which can be widely detected and efficiently transmitted among Salmonella strains of various serotypes by fusing to a new conjugative helper plasmid. The sharp increase in the prevalence of ciprofloxacin resistance in recent years may cause a predisposition to the emergence of multidrug-resistant Salmonella strains and pose huge challenges to public health and infection control efforts.

Mechanisms of azithromycin resistance have rarely been reported. In this study, an IncFIB/IncHI1B plasmid that confers resistance to azithromycin was recovered from a clinical Klebsiella pneumoniae strain. This plasmid could be efficiently disseminated to Escherichia coli, Salmonella, and other Gram-negative bacterial pathogens through conjugation. This plasmid was shown to carry three macrolide resistance genes

Increased usage of daptomycin to treat infections caused by Gram-positive bacterial pathogens has resulted in emergence of resistant mutants. In a search for more effective daptomycin analogues through medicinal chemistry studies, we found that methylation at the nonproteinogenic amino acid kynurenine in daptomycin could result in significant enhancement of antibacterial activity. Termed "kynomycin," this new antibiotic exhibits higher antibacterial activity than daptomycin and is able to eradicate methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant Enterococcus (VRE) strains, including daptomycin-resistant strains. The improved antimicrobial activity of kynomycin was demonstrated in in vitro time-killing assay, in vivo wax worm model, and different mouse infection models. The increased antibacterial activity, improved pharmacokinetics, and lower cytotoxicity of kynomycin, compared to daptomycin, showed the promise of the future design and development of next-generation daptomycin-based antibiotics.

Background Carbapenemase-producing Vibrio spp., which exhibit an XDR phenotype, have become increasingly prevalent and pose a severe threat to public health. Objectives To investigate the genetic characteristics of NDM-1-producing Vibrio spp. isolates and the dissemination mechanisms of blaNDM-1 in Vibrio. Methods A total of 1363 non-duplicate Vibrio spp. isolates collected from shrimp samples in China were subjected to antimicrobial susceptibility tests and screened for blaNDM-1. The blaNDM-1-positive isolates were further characterized by PFGE, MLST, conjugation and WGS using Illumina and Nanopore platforms. Plasmid stability and fitness cost were assessed using Escherichia coli J53, Klebsiella pneumoniae Kpt80 and Salmonella spp. SA2051 as recipient strains. Results In total, 13 blaNDM-1-positive isolates were identified, all exhibiting MDR. WGS analysis revealed that the 13 blaNDM-1 genes were all associated with a derivative of Tn125. Plasmid analysis revealed that six blaNDM-1 genes were located in IncC plasmids and the other seven were carried by plasmids of two different novel types. Conjugation and plasmid stability assays showed that only the IncC plasmids could be transferred to all the recipient strains and could be stably maintained in the hosts. Conclusions The emergence of the novel plasmids has contributed to the variable genetic contexts of blaNDM-1 in Vibrio spp. and IncC plasmids harbouring the blaNDM-1 gene could facilitate the spread of such genes between Vibrio spp. and other zoonotic pathogens, leading to a rapid dissemination of blaNDM-1 in bacterial pathogens worldwide.

We previously reported the recovery of five ST11 carbapenem resistant hypervirulent

Carbapenem-resistant hypervirulent Klebsiella pneumoniae (CR-HvKP) strains have been increasingly reported, and it is important to understand the evolutionary mechanisms of these highly pathogenic and resistant bacterial pathogens. In this study, we characterized a ST11 carbapenem-resistant K. pneumoniae strain which harbored an IncFIB/IncHI1B type virulence plasmid and an IncFII/IncR type blaKPC–2-bearing plasmid. The virulence plasmid was found to be conjugative and harbored a 35-kbp fragment including aerobactin encoding cluster from virulence plasmid pLVPK and multiple resistance genes, resulting in a mosaic multi-drug resistance and virulence plasmid. This virulence plasmid could be transferred via conjugation to Escherichia coli and K. pneumoniae strains alone as well as together with the blaKPC–2-bearing plasmid. Co-transmission of virulence and blaKPC–2-bearing plasmids would directly convert a classic K. pneumoniae strain into CR-HvKP strain, leading to a sharp increase in the prevalence of CR-HvKP in clinical settings, which poses a great threat to human health.

Our objective was to characterize the genetic features of plasmids harbored by two genetically related, MCR-1 and NDM-5-producing Escherichia coli strains recovered from a chicken meat sample. The genetic profiles of all plasmids harbored by the two test strains, namely, 1106 and 1107, were determined by whole-genome sequencing, S1-pulsed-field gel electrophoresis (PFGE), Southern hybridization, and bioinformatics analysis. The transferability of plasmids harbored by the two strains was assessed by filter mating assay. Strains 1106 and 1107 were resistant to almost all the antibiotics, including colistin and fosfomycin, but remained susceptible to amikacin and tigecycline. The plasmids of p1107-NDM-5 and p1106-NDM-5 both contain a class I integron which lacks the ISAba125 element. The backbone of p1106-IncFII exhibited a high degree of similarity with that of p1106-NDM-5 and p1107-NDM-5, implying that events of plasmid fusion and resolution were involved in the formation of the two plasmids. The plasmids p1106-IncHI2MCR and p1107-IncHI2MCR belong to an IncHI2 replicon type, with three copies of ISApl1 being observed in p1106-IncHI2MCR, implying that the mcr-1 gene was transferable among bacteria that reside in the same food matrix. In this study, p1106-IncFIB, p1107-99K, p1107-111K, and p1107-118K were all found to be phage-like plasmids, with p1106-IncFIB and p1107-118K containing several virulence genes, including iroBCDEN, iucABCD, sitABCD, hlyF, and iss. Surprisingly, resistance genes such as aph(3′)-Ia, sul3, and aac(3′)-IId could also be found in p1107-118K, but resistance genes were not detected in other phage-like plasmids. In conclusion, enhanced surveillance is required to monitor and control the dissemination of various resistance determinants among foodborne pathogens. IMPORTANCE Carbapenem and colistin are last-resort antibiotics used to treat serious clinical infections caused by multidrug-resistant (MDR) bacterial pathogens. Plasmids encoding resistance to carbapenems and colistin have been reported in clinical pathogens in recent years, and yet few studies reported cocarriage of mcr and blaNDM genes in Escherichia coli strains of food origin. How plasmids encoding these two important resistance determinants are being evolved and transmitted in bacterial pathogens is not well understood. In this study, we investigated the genetic features of plasmids harbored by two nonclonal, mcr-1- and blaNDM-5-bearing E. coli strains (1106 and 1107) recovered from a fresh chicken meat sample to understand and provide evidence of the level and dynamics of MDR plasmid transmission. Our data confirmed that active plasmid fusion and resolution events were involved in the formation of plasmids that harbor multiple resistance genes, which provide insights into the further control of plasmid evolution in bacterial pathogens.

Acinetobacter sp. is among the ESKAPE organisms which represent the major nosocomial pathogens that exhibited a high resistance rate. A. pittii, frequently associated with antimicrobial resistance particularly to carbapenems, is one of the most common Acinetobacter species causing invasive infection. Pandrug resistant A. pittii has rarely been reported. Here, we report the case of a patient with acute exacerbations of chronic obstructive pulmonary disease three years after double lung transplantation and developed severe pneumonia associated with pandrug resistant A. pittii infection. Phenotypic and genomic characteristics of this pandrug resistant isolate (17-84) was identified, and the mechanisms underlying its resistance phenotypes were analyzed. Isolate 17-84 belonged to ST63, carried a non-typable and non-transferable plasmid encoding multiple acquired resistance genes including carbapenemase gene blaOXA-58. Point mutations and acquired resistance genes were identified which were associated with different drug resistance phenotypes. To our knowledge, this is the first detailed phenotypic and genomic characterization of PDR A. pittii causing severe infections in clinical settings. Findings from us and others indicate that A. pittii could serve as a reservoir for carbapenem determinants. The emergence of such a superbug could pose a serious threat to public health. Further surveillance of PDR A. pittii strains and implementation of stricter control measures are needed to prevent this emerging pathogen from further disseminating in hospital settings and the community.

Colistin resistance mediated by mcr-1-bearing plasmids poses a new challenge to treatment of Salmonella infections. To probe the scale of the problem that colistin resistance mediated by mcr-1 plasmids among Salmonella, the prevalence of mcr-1 in foodborne Salmonella recovered from 2014 to 2017 in Shenzhen, China and genetic profile of mcr-1 positive isolates were investigated. All mcr-1 positives Salmonella strains were collected from food products, characterized by PCR and MALDI-TOF, and subjected to antimicrobial susceptibility testing, whole-genome sequencing, bioinformatics analysis, and conjugation. Twenty-eight mcr-1-positive Salmonella strains were recovered from pork. The rate of recovery displayed an increasing trend and was often accompanied by multidrug resistance. Salmonella Typhimurium was the most prevalent serotypes. Comparative genomic analysis indicated that the mcr-1 gene was located on the transferable IncX4 plasmids, as well as the IncHI2 plasmids, in which the gene was associated with ISApl1. All two types of plasmids were often detected in zoonotic pathogen. Transferable 251K mcr-1-bearing IncHI2 type plasmids were frequently reported in human and food-producing animals, but this is first time to detect a certain number in food. These findings show that dissemination of these two types of plasmids is responsible for the increase in the prevalence of colistin resistance in Salmonella strains in recent years, leading to rapid emergence of MDR Salmonella upon acquisition of these two mcr-1-bearing plasmids. Transmission of IncX4 and IncHI2 plasmids in Salmonella would cause huge public health concerns in controlling foodborne infections caused by Salmonella.

Our objective was to characterize the genetic features of plasmids harbored by two genetically related, MCR-1 and NDM-5-producing Escherichia coli strains recovered from a chicken meat sample. The genetic profiles of all plasmids harbored by the two test strains, namely, 1106 and 1107, were determined by whole-genome sequencing, S1-pulsed-field gel electrophoresis (PFGE), Southern hybridization, and bioinformatics analysis. The transferability of plasmids harbored by the two strains was assessed by filter mating assay. Strains 1106 and 1107 were resistant to almost all the antibiotics, including colistin and fosfomycin, but remained susceptible to amikacin and tigecycline. The plasmids of p1107-NDM-5 and p1106-NDM-5 both contain a class I integron which lacks the ISAba125 element. The backbone of p1106-IncFII exhibited a high degree of similarity with that of p1106-NDM-5 and p1107-NDM-5, implying that events of plasmid fusion and resolution were involved in the formation of the two plasmids. The plasmids p1106-IncHI2MCR and p1107-IncHI2MCR belong to an IncHI2 replicon type, with three copies of ISApl1 being observed in p1106-IncHI2MCR, implying that the mcr-1 gene was transferable among bacteria that reside in the same food matrix. In this study, p1106-IncFIB, p1107-99K, p1107-111K, and p1107-118K were all found to be phage-like plasmids, with p1106-IncFIB and p1107-118K containing several virulence genes, including iroBCDEN, iucABCD, sitABCD, hlyF, and iss. Surprisingly, resistance genes such as aph(3′)-Ia, sul3, and aac(3′)-IId could also be found in p1107-118K, but resistance genes were not detected in other phage-like plasmids. In conclusion, enhanced surveillance is required to monitor and control the dissemination of various resistance determinants among foodborne pathogens. IMPORTANCE Carbapenem and colistin are last-resort antibiotics used to treat serious clinical infections caused by multidrug-resistant (MDR) bacterial pathogens. Plasmids encoding resistance to carbapenems and colistin have been reported in clinical pathogens in recent years, and yet few studies reported cocarriage of mcr and blaNDM genes in Escherichia coli strains of food origin. How plasmids encoding these two important resistance determinants are being evolved and transmitted in bacterial pathogens is not well understood. In this study, we investigated the genetic features of plasmids harbored by two nonclonal, mcr-1- and blaNDM-5-bearing E. coli strains (1106 and 1107) recovered from a fresh chicken meat sample to understand and provide evidence of the level and dynamics of MDR plasmid transmission. Our data confirmed that active plasmid fusion and resolution events were involved in the formation of plasmids that harbor multiple resistance genes, which provide insights into the further control of plasmid evolution in bacterial pathogens.

A multidrug-resistant Vibrio alginolyticus isolate recovered from a shrimp sample with reduced carbapenem susceptibility produced a novel metallo-β-lactamase (MBL), VAM-1. That carbapenemase shared 67% to 70% amino acid identity with several VMB family subclass B1 MBLs, which were recently reported among some marine bacteria including Vibrio , Glaciecola , and Thalassomonas . The bla VAM-1 gene was located in a novel conjugative plasmid, namely, pC1579, and multiple copies of bla VAM-1 via an unusual mechanism of gene amplification were detected in pC1579.

Carbapenem-resistant hypervirulent Klebsiella pneumoniae (CR-HvKP) strains have been increasingly reported, and it is important to understand the evolutionary mechanisms of these highly pathogenic and resistant bacterial pathogens. In this study, we characterized a ST11 carbapenem-resistant K. pneumoniae strain which harbored an IncFIB/IncHI1B type virulence plasmid and an IncFII/IncR type blaKPC–2-bearing plasmid. The virulence plasmid was found to be conjugative and harbored a 35-kbp fragment including aerobactin encoding cluster from virulence plasmid pLVPK and multiple resistance genes, resulting in a mosaic multi-drug resistance and virulence plasmid. This virulence plasmid could be transferred via conjugation to Escherichia coli and K. pneumoniae strains alone as well as together with the blaKPC–2-bearing plasmid. Co-transmission of virulence and blaKPC–2-bearing plasmids would directly convert a classic K. pneumoniae strain into CR-HvKP strain, leading to a sharp increase in the prevalence of CR-HvKP in clinical settings, which poses a great threat to human health.

Objectives To test whether antibiotics of different functional categories exhibit differential potential in promoting transmission of MDR-encoding plasmids among members of the gut microbiome. Methods Rats inoculated with blaNDM-1-bearing Klebsiella pneumoniae were subjected to treatment with different types of antibiotics. The structural changes in the gastrointestinal (GI) tract microbiome were determined by 16S rRNA sequencing and analysis. In addition, the efficiency of transmission of blaNDM-1-bearing plasmids to different subtypes of GI tract Escherichia coli was also confirmed in vitro. Results We showed that drugs that are commonly used to treat Gram-negative bacterial infections, such as ampicillin and amoxicillin, could enrich both carbapenem-resistant Enterobacteriaceae (CRE) and antibiotic-susceptible E. coli in the GI tract, thereby promoting transmission of the blaNDM-1-bearing plasmid in the gut microbiome. In contrast, meropenem was found to minimize the population of CRE in the gut microbiome, hence treatment with this drug exhibited drastically lower potential to promote transmission of the blaNDM-1-bearing plasmid to the recipient strains. We further showed that an increased population size of Proteobacteria due to a suppressive effect on Firmicutes is a key factor in enhancing the efficiency of transmission of the blaNDM-1-bearing plasmid and hence dissemination of carbapenem-resistant strains. Conclusions This study depicted for the first time the effect of different antibiotics on the structure of the rat GI tract microbiome, which in turn determined the pattern and rate of transmission of the blaNDM-1-bearing plasmid. Such findings can help establish new guidelines for prudent antibiotic usage to minimize the chance of dissemination of mobile resistance elements among members of the GI tract microbiome.

This study reported for the first time and characterized in detail the genetic features of a mobile virulence-encoding fragment located in the chromosome of a clinical virulent K. pneumoniae strain and revealed the occurrence of a transposition event mediated by IS26. This genetic structure could mediate the transposition of the virulence-encoding fragment into various plasmid backbones and chromosomes through formation of a circular intermediate., Klebsiella pneumoniae is increasingly being regarded as a reservoir of diverse antibiotic resistance genes and a pathogen that causes severe infections in both the hospital and the community. In this study, we performed molecular characterization of a carbapenem-resistant and highly virulent K. pneumoniae strain recovered from a hospital patient. The virulence-encoding genes rmpA2 and iucABCDiutA were found to be located in the chromosome and are flanked by IS26 elements. These chromosomally located virulence genes, if not incurring fitness costs, are expected to be much more stable than plasmid-located genes. Detailed analysis of the fragment in which these virulence genes are located showed that the fragment can readily form a circular intermediate that may promote integration of this virulence-encoding fragment into various plasmid backbones and other chromosomal regions. Findings in this work provide new insights into mechanisms of transmission of virulence-encoding genes in K. pneumoniae. IMPORTANCE This study reported for the first time and characterized in detail the genetic features of a mobile virulence-encoding fragment located in the chromosome of a clinical virulent K. pneumoniae strain and revealed the occurrence of a transposition event mediated by IS26. This genetic structure could mediate the transposition of the virulence-encoding fragment into various plasmid backbones and chromosomes through formation of a circular intermediate. Therefore, findings in this work provide important insights into the transmission mechanisms of mobile virulence profiles in K. pneumoniae strains and lay the foundation for devising effective intervention approaches aimed at preventing the dissemination of these virulence-encoding elements.

The incidence of ciprofloxacin resistance in

Klebsiella pneumoniae is increasingly being regarded as a reservoir of diverse antibiotic resistance genes and a pathogen that causes severe infections in both the hospital and the community. In this study, we performed molecular characterization of a carbapenem-resistant and highly virulent K. pneumoniae strain recovered from a hospital patient. The virulence-encoding genes rmpA2 and iucABCDiutA were found to be located in the chromosome and are flanked by IS26 elements. These chromosomally located virulence genes, if not incurring fitness costs, are expected to be much more stable than plasmid-located genes. Detailed analysis of the fragment in which these virulence genes are located showed that the fragment can readily form a circular intermediate that may promote integration of this virulence-encoding fragment into various plasmid backbones and other chromosomal regions. Findings in this work provide new insights into mechanisms of transmission of virulence-encoding genes in K. pneumoniae. IMPORTANCE This study reported for the first time and characterized in detail the genetic features of a mobile virulence-encoding fragment located in the chromosome of a clinical virulent K. pneumoniae strain and revealed the occurrence of a transposition event mediated by IS26. This genetic structure could mediate the transposition of the virulence-encoding fragment into various plasmid backbones and chromosomes through formation of a circular intermediate. Therefore, findings in this work provide important insights into the transmission mechanisms of mobile virulence profiles in K. pneumoniae strains and lay the foundation for devising effective intervention approaches aimed at preventing the dissemination of these virulence-encoding elements.

Objective The emergence of multidrug-resistant (MDR) and hypervirulent Klebsiella pneumoniae (hvKP) facilitates simultaneous dissemination of virulence and resistance in a single event, which poses serious threat to public health. Methods This study characterized the multidrug-resistant and moderately virulent ST11 K64 K. pneumoniae strain HB25-1 from a clinical case with microbiological and genomic approaches. Plasmids from strain HB25-1 were subjected to whole plasmid sequencing using both the Illumina NextSeq 500 sequencing platform and Nanopore MinION sequencer platforms. Klebsiella pneumoniae HB25-1 was subjected to a conjugation experiment and Galleria mellonella infection model to evaluate the transmission and virulence potential. Results We report the emergence of an ST11, serotype K64 K. pneumoniae isolate, which is resistant to third-generation cephalosporin and exhibited a moderate level of virulence. WGS revealed that this strain harboured a plasmid, pHB25-1, which carried multidrug resistance genes (blaDHA-1, qnrB4, dfrA12, aadA2, sul1, aac(3)-lld, blaTEM-1, mph(E)) and virulence-encoding genes (the regulator of mucoid phenotype A gene rmpA2 and the aerobactin gene cluster iutAiucABCD). Genomic analysis indicated that pHB25-1 was formed through co-integration of structural regions located in two different plasmids, enabling it to encode both resistance and virulent phenotypes. Conclusion Findings in this study provide evidence of active plasmid evolution in K. pneumoniae and suggest that surveillance of multidrug-resistant and hypervirulent K. pneumoniae is urgently needed.

Background: A pandemic outbreak caused by a novel coronavirus, 2019-nCoV, has originated from Wuhan, China and spread to many countries around the world. The outbreak has led to around 45 thousand cases and over one thousand death so far. Methods: Phylogenetic analysis and sequence alignment were used to align the whole genome sequence of 2019-nCoV with other over 200 sequences of coronaviruses to predict the origin of this novel virus. In addition, protein modeling and analysis were performed to access the potential binding of the spike protein of 2019-nCoV with human cell receptor, angiotensin-converting enzyme 2 (ACE2). Results: Detailed genomic and structure-based analysis of a new coronavirus, namely 2019-nCoV, showed that the new virus is a new type of bat coronavirus and is genetically fairly distant from the human SARS coronavirus. Structure analysis of the spike (S) protein of this new virus showed that its S protein only binds much weaker to the ACE2 receptor on human cells whereas the human SARS coronavirus exhibits strongly affinity to the ACE receptor. Conclusions: These findings suggest that the new virus should theoretically not be able to cause very serious human infection when compared to human SARS virus. However, the lower pathogenicity of this new virus may lead to longer incubation time and better adaption to human, which may favor its efficient transmission in human. These data are important to guide design of infection control policy and inform the public on the nature of threat imposed by 2019-nCov. Most importantly, using the analysis platform that we have developed, we should be able to predict whether the new mutations could lead to the increase of infectivity of the mutated virus in a very short time.

Detailed genomic and structure-based analysis of a new coronavirus, namely 2019-nCoV, showed that the new virus is a new type of bat coronavirus and is genetically fairly distant from the human SARS coronavirus. Structure analysis of the spike (S) protein of this new virus showed that its S protein only binds weakly to the ACE2 receptor on human cells whereas the human SARS coronavirus exhibits strongly affinity to the ACE receptor. These findings suggest that the new virus does not readily transmit between humans and should theoretically not able to cause very serious human infection. These data are important to guide design of infection control policy and inform the public on the nature of threat imposed by 2019-nCov when results of direct laboratory tests on this virus are not expected to be available in the near future.

Background: A pandemic outbreak caused by a novel coronavirus, 2019-nCoV, has originated from Wuhan, China and spread to many countries around the world. The outbreak has led to around 45 thousand cases and over one thousand death so far. Methods: Phylogenetic analysis and sequence alignment were used to align the whole genome sequence of 2019-nCoV with other over 200 sequences of coronaviruses to predict the origin of this novel virus. In addition, protein modeling and analysis were performed to access the potential binding of the spike protein of 2019-nCoV with human cell receptor, angiotensin-converting enzyme 2 (ACE2). Results: Detailed genomic and structure-based analysis of a new coronavirus, namely 2019-nCoV, showed that the new virus is a new type of bat coronavirus and is genetically fairly distant from the human SARS coronavirus. Structure analysis of the spike (S) protein of this new virus showed that its S protein only binds much weaker to the ACE2 receptor on human cells whereas the human SARS coronavirus exhibits strongly affinity to the ACE receptor. Conclusions: These findings suggest that the new virus should theoretically not be able to cause very serious human infection when compared to human SARS virus. However, the lower pathogenicity of this new virus may lead to longer incubation time and better adaption to human, which may favor its efficient transmission in human. These data are important to guide design of infection control policy and inform the public on the nature of threat imposed by 2019-nCov. Most importantly, using the analysis platform that we have developed, we should be able to predict whether the new mutations could lead to the increase of infectivity of the mutated virus in a very short time.

We report the identification of a carbapenem-resistant, hypervirulent Klebsiella pneumoniae (hvKp) strain which produced the carbapenemase VIM-1. Genomic analysis showed that the strain belonged to sequence type ST23 and serotype K1, a major hvKp clone, and harbored three resistance-encoding plasmids. Among them, a bla VIM-1 -bearing plasmid was found to possess a mosaic structure presumably generated by multiple gene mobilization events.

This study aims to investigate and compare the complete nucleotide sequences of the multidrug resistance plasmids pVb0267 and pVb0499, which were recovered from foodborne Vibrio parahaemolyticus isolates, and analyze the genetic environment of blaCTX–M–55 to provide insight into the dissemination mechanisms of this resistance element. Analysis of the sequences of plasmids pVb0267 (166,467 bp) and pVb0499 (192,739 bp) revealed that the backbones of these two plasmids exhibited a high degree of similarity with pR148, a recognized type 1a IncC plasmid recovered from Aeromonas hydrophila (99% identity). The resistance genes, found in both plasmids, included qacH, aadB, arr2, blaOXA–10, aadA1, sul1, tet(A), and blaCTX–M–55, which were mostly arranged in a specific region designated ARI-A. Plasmid pVb0499 was found to possess a larger size of ARI-A than pVb0267, which lacked a mer determination region, a qnr A segment, an aacC3 gene and several mobility-encoding genes. Comparative analysis of resistance island (RI) of these plasmids and others revealed the potential evolution route of these RI sequences. In conclusion, plasmids harboring the blaCTX–M–55 gene has been recovered in Vibrio parahaemolyticus strains of food origin. It is alarming to find that IncC plasmids harboring resistance islands are disseminating in aquatic bacterial strains. The continuous evolution of resistance genes in conjugative plasmid in aquatic bacteria could be due to bacterial adaption to aquaculture environment, where antibiotics were increasingly used.

This report describes the first detection of a bla VEB-2 gene in a Vibrio parahaemolyticus strain isolated from a shrimp sample. The bla VEB-2 gene was carried on a novel Inc-type plasmid that was likely to have originated from aquatic organisms, as indicated by a comparison with other known genetic elements in the GenBank database. However, the plasmid contains resistance elements usually harbored by members of the family Enterobacteriaceae , suggesting that gene transfer events occurred and contributed to the formation of this multidrug resistance-encoding plasmid.

The human gut microbiome is the presumed site in which the emergence and evolution of antibiotic-resistant organisms constantly take place. To delineate the genetic basis of resistance formation in gut microbiome strains, we investigated the changes in the subpopulation structure of Escherichia coli in rat intestine before and after antimicrobial treatment. We observed that antibiotic treatment was selected for an originally minor subpopulation E. coli carrying the biofilm-forming genetic locus pgaABCD and the toxin-antitoxin system HipAB. Such strains possessed dramatically enhanced ability to withstand the detrimental effects of antibiotics, becoming a dominant subspecies upon antibiotic treatment and eventually evolving into resistant mutants. In contrast, E. coli strains that did not carry pgaABCD and HipAB were eradicated upon antibiotic treatment. Our findings, therefore, suggested that genes encoding biofilm-forming ability played an important role in conferring specific gut E. coli strains the ability to evolve into resistant strains upon a prolonged antibiotic treatment, and that such strains may therefore be considered bacterial antibiotic resistance progenitor cells in the gut microbiome.

The rapid dissemination of non-conjugative virulence plasmids among non-K1/K2 types of Klebsiella pneumoniae poses an unprecedented threat to human health, yet the underlying mechanisms governing dissemination of such plasmids is unclear. In this study, a novel 68 581 bp IncFIA plasmid is discovered that can be fused to a hypervirulence-encoding plasmid to form a hybrid conjugative virulence plasmid in conjugation experiments; such fusion events involve homologous recombination between a 241 bp homologous region located in each of the two plasmids. The fusion hypervirulence-encoding plasmid can be conjugated to both classic and blaKPC-2 -bearing carbapenem-resistant K. pneumoniae strains through conjugation, enabling such strains to acquire the ability to express the hypervirulence phenotype. Dissemination of this fusion virulence plasmid will impose an enormous burden on current efforts to control and treat infections caused by multidrug resistant and hypervirulent K. pneumoniae.

Vibrio alginolyticus is a Gram-negative halophilic and mesophilic bacterium, particularly associated with severe epidemic vibriosis, which causes high mortality in marine animals, including fish, shellfish, and shrimp ([1][1]). The carbapenemase NDM-1 has been found mainly in Enterobacteriaceae

Objectives To investigate the genetic features of the blaVIM-1 gene first detected in a cephalosporin-resistant Vibrio alginolyticus isolate, Vb1978. Methods The MICs of V. alginolyticus strain Vb1978 were determined, and the β-lactamases produced were screened and analysed using conjugation, S1-PFGE and Southern blotting. The complete sequence of the blaVIM-1-encoding plasmid was also obtained using the Illumina and MinION sequencing platforms. Results V. alginolyticus strain Vb1978, isolated from a retail shrimp sample, was resistant to cephalosporins and exhibited reduced susceptibility to carbapenems. A novel blaVIM-1-carrying conjugative plasmid, designated pVb1978, was identified in this strain. Plasmid pVb1978 had 50 001 bp and comprised 59 predicted coding sequences (CDSs). The plasmid backbone of pVb1978 was homologous to those of IncP-type plasmids, while its replication region was structurally similar to non-IncP plasmids. The blaVIM-1 gene was found to be carried by the class 1 integron In70 and associated with a defective Tn402-like transposon. Conclusions A novel blaVIM-1-carrying conjugative plasmid, pVb1978, was reported for the first time in V. alginolyticus, which warrants further investigation in view of its potential pathogenicity towards humans and widespread occurrence in the environment.

Animal and human gastrointestinal tract is the key site from which resistant bacterial pathogens emerge. In this work, we tested whether antibiotics of different functional categories exhibited differential potential in promoting transmission of multidrug resistance-encoding plasmids among the gut microflora by measuring the population size of both donor and potential recipient strains before, during and after the treatment process. In this study, we showed that drugs that are commonly used to treat Gram negative bacterial infections such as ampicillin and amoxicillin could enrich both carbapenem resistant enterobacteriaceae (CRE) and antibiotic-susceptible E. coli in the gastrointestinal tract, thereby promoting transmission of the blaNDM-1-bearing plasmid in the gut microbiome. In contrast, meropenem was found to minimize the population of CRE in the gut microbiome, hence treatment with this drug exhibited drastically lower potential to promote transmission of the blaNDM-1-bearing plasmid to the recipient strains. We further showed that whether an antibiotic enriches or suppresses the population size of donor and recipient strains in the gastrointestinal tract is actually dependent on the relative inhibitory effects of the drug on the firmicutes or proteobacteria, and that an increased population size of proteobacteria due to a higher suppressive effect on firmicutes is a key factor in enhancing the efficiency of transmission of the blaNDM-1-bearing plasmid and hence dissemination of carbapenem resistant strains. In conclusion, this study identified and characterized the factors that affected the transmission of MDR plasmid in gut microbiome and its underlying mechanisms. Such findings can help establish new guidelines for prudent antibiotic usage to minimize the chance of dissemination of resistance mobile elements among the GI tract microflora. Funding Statement: The research was supported by the Collaborative Research Fund (C5026-16G) of the Hong Kong Research Grant Council and the Health and Medical Research Fund (HMRF) of the Food and Health Bureau, the Government of the Hong Kong SAR (14130402 to SC). Declaration of Interests: All authors declared no conflict of interests. Ethics Approval Statement: The experimental protocol was approved by the Research Animal Care and Use Committee of the Hong Kong Polytechnic University.

Objectives To investigate a set of MDR conjugative plasmids found in Vibrio species and characterize the underlying evolution process. Methods pAQU-type plasmids from Vibrio species were sequenced using both Illumina and PacBio platforms. Bioinformatics tools were utilized to analyse the typical MDR regions and core genes in the plasmids. Results The nine pAQU-type plasmids ranged from ∼160 to 206 kb in size and were found to harbour as many as 111 core genes encoding conjugative, replication and maintenance functions. Eight plasmids were found to carry a typical MDR region, which contained various accessory and resistance genes, including ISCR1-blaPER-1-bearing complex class 1 integrons, ISCR2-floR, ISCR2-tet(D)-tetR-ISCR2, qnrVC6, a Tn10-like structure and others associated with mobile elements. Comparison between a plasmid without resistance genes and different MDR plasmids showed that integration of different mobile elements, such as IS26, ISCR1, ISCR2, IS10 and IS6100, into the plasmid backbone was the key mechanism by which foreign resistance genes were acquired during the evolution process. Conclusions This study identified pAQU-type plasmids as emerging MDR conjugative plasmids among important pathogens from different origins in Asia. These findings suggest that aquatic bacteria constitute a major reservoir of resistance genes, which may be transmissible to other human pathogens during food production and processing.