Phage-like Plasmid Research Articles

Isolation and characterization of a multidrug-resistant carbapenem-resistant Klebsiella pneumoniae strain co-carrying IncFII/IncR-KPC-2 and phage-like plasmids

ObjectivesThe aim of this study was to characterize the co-occurrence of the blaSHV-182, blaKPC-2, blaCTX-M-65, and blaTEM-1B genes in a multi-resistant carbapenem carbapenem-resistant K. pneumoniae (CRKP) strain. MethodsThe multi-resistant CRKP strain was isolated from the bronchoalveolar lavage fluid (BAL) of a patient with acute exacerbation of chronic obstructive pulmonary disease (AECOPD), after cultured in lysogeny broth, the colonies with apparent phenotypic differences (size, shape, color, and surface states) were picked out for species identification by sequencing their 16S rDNA and BLASTN in the NCBI database, antimicrobial susceptibility was determined, Genomic DNA of overnight cultured K. pneumoniae D1a was extracted by using the Bacterial DNA Isolation Kit, the library was constructed by using NEBNext®UltraTM DNA Library Prep Kit for Illumina, and the whole-genome sequencing was performed on the Illumina HiSeq PE150 platform. The pair-end reads were assembled by SOAP denovo v2.04, SPAdes, and ABySS. Genome characteristics were analyzed by using bioinformatics methods. ResultsThe WGS of the multi-resistant CRKP strain D1a (CRKP D1a) yielded 84 scaffolds of genomic DNA (5, 492, 035 bp) and two circular plasmids, pD1a1 (173, 413 bp) and pD1a2 (57, 686 bp). CRKP D1a was classified as the sequence type 11, and harbored 16 antibiotic resistance genes and 30 multidrug efflux pump-encoding genes associated with the resistance to almost all kinds of commonly used antibiotics. The plasmid pD1a1 belonged to the IncFII/IncR hybrid plasmid family with blaSHV-182, blaKPC-2, blaCTX-M-65, and blaTEM-1B genes flanked by the IS6-like element. The plasmid pD1a2 was comprised of 95 phage-related genes but had no lysis activity on the tested K. pneumoniae strains. ConclusionThis study reports the isolation of a multidrug-resistant CRKP strain co-carrying IncFII/IncR-KPC2 plasmid and phage-like plasmid, the findings hold significant implications for further investigating the resistance mechanism and molecular diversity of CRKP.

IS26-Mediated Formation of a Hybrid Plasmid Carrying mcr-1.1.

The objective of this study was to elucidate the characteristics and mechanism of formation of the fusion plasmid pHNSHP24 carrying mcr-1.1. mcr-1.1-bearing Escherichia coli SHP24 and the corresponding transconjugant were subjected to whole-genome sequencing (WGS) combining the Illumina and MinION platforms to obtain the complete sequences of the fusion plasmid and its original plasmids. Complete sequence analysis and S1 nuclease-pulsed field gel electrophoresis (S1-PFGE) results indicated that E. coli SHP24 carried four plasmids: mcr-1.1-harboring phage-like plasmid pHNSHP24-3, F53:A-:B- plasmid pHNSHP24-4, pHNSHP24-1, and pHNSHP24-2. However, the plasmid pHNSHP24 carrying mcr-1.1 presents in the transconjugant differed from the four plasmids in the donor strain SHP24. Further analysis showed that pHNSHP24 may be the fusion product of pHNSHP24-3 and pHNSHP24-4 and is formed through a replicative transposition mechanism mediated by IS26 in E. coli SHP24. This study is the first to report the fusion of an mcr-1.1-harboring phage-like pO111 plasmid and an F53:A-:B- plasmid mediated by IS26. Our findings revealed the role of phage-like and fusion plasmids in the dissemination of mcr-1.1.

Molecular Characterization of pBOq-IncQ and pBOq-95LK Plasmids of Escherichia coli BOq 01, a New Isolated Strain from Poultry Farming, Involved in Antibiotic Resistance.

The increase in antimicrobial resistance has raised questions about how to use these drugs safely, especially in veterinary medicine, animal nutrition, and agriculture. Escherichia coli is an important human and animal pathogen that frequently contains plasmids carrying antibiotic resistance genes. Extra chromosomal elements are required for various functions or conditions in microorganisms. Several phage-like plasmids have been identified, which are important in antibiotic resistance. In this work, the molecular characterization of the pBOq-IncQ (4.5 kb) and pBOq-95LK (95 kb) plasmids found in the E. coli strain BOq 01, a multidrug resistant bacteria isolated from a poultry farm, are considered. Plasmid pBOq-IncQ belongs to the incQ incompatibility plasmid family and is involved in sulfonamide resistance. Plasmid pBOq-95LK is a lytic phage-like plasmid that is involved in the lysis of the E. coli BOq 01 strain and carries a bleomycin resistance gene and a strain cured of this plasmid shows bleomycin sensitivity. Induction of the lytic cycle indicates that this phage-like plasmid is an active phage. This type of plasmid has been reported to acquire genes such as mcr-1, which codes for colistin resistance and bacterial persistence and is a significant public health threat. A genome comparison, a pangenomic and phylogenomic analysis with other phage-like plasmids reported in the literature were performed to understand better the evolution of this kind of plasmid in bacteria and its potential importance in antibiotic resistance.

Occurrence of Serratia marcescens Carrying blaIMP-26 and mcr-9 in Southern China: New Insights in the Evolution of Megaplasmid IMP-26.

The spread of multidrug-resistant enterobacteria strains has posed a significant concern in public health, especially when the strain harbors metallo-beta-lactamase (MBL)-encoding and mobilized colistin resistance (mcr) genes as such genetic components potentially mediate multidrug resistance. Here we report an IncHI2/2A plasmid carrying blaIMP-26 and mcr-9 in multidrug-resistant Serratia marcescens human isolates YL4. Antimicrobial susceptibility testing was performed by the broth microdilution method. According to the results, S. marcescens YL4 was resistant to several antimicrobials, including β-lactams, fluorquinolones, sulfanilamide, glycylcycline, and aminoglycosides, except for amikacin. To investigate the plasmid further, we conducted whole-genome sequencing and sequence analysis. As shown, S. marcescens YL4 possessed a circular chromosome with 5,171,477 bp length and two plasmids, pYL4.1 (321,744 bp) and pYL4.2 (46,771 bp). Importantly, sharing high similarity with plasmids pZHZJ1 and pIMP-26, pYL4.1 has an IncHI2/2A backbone holding a variable region containing blaIMP-26, mcr-9, and two copies of blaTEM-1B. After comprehensively comparing relevant plasmids, we proposed an evolutionary pathway originating from ancestor pZHZJ1. Then, via an acquisition of the mcr-9 element and a few recombination events, this plasmid eventually evolved into pYL4.1 and pIMP-26 through two different pathways. In addition, the phage-like plasmid pYL4.2 also carried a blaTEM-1B gene. Remarkably, this study first identified a multidrug-resistant S. marcescens strain co-harboring blaIMP-26 and mcr-9 on a megaplasmid pYL4.1 and also included a proposed evolutionary pathway of epidemic megaplasmids carrying blaIMP-26.

Genomic Characterization of a Uropathogenic Escherichia coli ST405 Isolate Harboring bla CTX-M-15-Encoding IncFIA-FIB Plasmid, bla CTX-M-24-Encoding IncI1 Plasmid, and Phage-Like Plasmid.

Escherichia coli sequence type 405 is an emerging antibiotic-resistant clonal group associated with the global dissemination of extended-spectrum β-lactamase-producing E. coli. In this study, we report the genome assembly and characterization of a uropathogenic E. coli ST405 strain, SZESBLEC201, based on long and short reads obtained from the Nanopore and Illumina sequencing platforms, respectively. Whole-genome sequencing revealed that SZESBLEC201 harbors a 5,020,403 bp chromosome and three plasmids, namely, pSZESBLEC201-1, pSZESBLEC201-2, and pSZESBLEC201-3. pSZESBLEC201-1 (111,621 bp) belongs to the IncFIA-FIB type and harbors blaCTX-M-15. However, this plasmid does not harbor conjugative transfer-associated genes, rendering pSZESBLEC201-1 unable to be conjugatively transferred. pSZESBLEC201-2 (95,138 bp) is a phage-like plasmid that shows a strong genome synteny with Escherichia phage P1 but with the absence of mobile genetic elements and some regulatory genes. pSZESBLEC201-3 (92,865 bp) belongs to the IncI1 type and carries blaCTX-M-24. In contrast to pSZESBLEC201-1, pSZESBLEC201-3 retains its full active conjugation machinery and can be transferred via conjugation. The genetic features of the genome show that the SZESBLEC201 has a unique virulence pattern compared with genetically similar strains found in the same country (China). The plasmid backbones exhibit a high degree of similarity to those of geographically distant isolates, highlighting the global spread of blaCTX-M genes and the genome plasticity of this clonal group. The coexistence of two blaCTX-M variants in the same strain increases the risk of the emergence of new blaCTX-M variants. Further studies on phage-like plasmids are necessary to provide insights into their biological activities and clinical significance.

Complete Genetic Analysis of Plasmids Carried by Two Nonclonal blaNDM-5- and mcr-1-Bearing Escherichia coli Strains: Insight into Plasmid Transmission among Foodborne Bacteria.

ABSTRACTOur 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.

Characterisation of Colistin -Resistant Enterobacterales and Acinetobacter Strains Carrying mcr Genes from Asian Aquaculture Products.

Aquaculture systems are widely recognised as hotspots for horizontal gene transfer, and the need for screening for bacteria carrying antimicrobial resistance genes in aquaculture systems is becoming more important. In this study, we characterised seventeen bacterial strains (Escherichia coli, Klebsiella pneumoniae, Acinetobacter baumannii, and A. nosocomialis) resistant to colistin originating from retailed aquaculture products imported from Vietnam to the Czech Republic. The mcr-1.1 gene was found located on plasmid types IncHI2, IncI2, and IncX4, as well as on the rarely described plasmid types IncFIB-FIC and IncFIB(K), phage-like plasmid p0111, and on the chromosome of E. coli. One E. coli strain carried the mcr-3.5 gene on IncFII(pCoo) plasmid in addition to the mcr-1.1 gene located on IncHI2 plasmid. K. pneumoniae was found to carry the mcr-1.1 and mcr-8.2 genes on IncFIA(HI1) plasmid. The mcr-4.3 gene was found on similar untypeable plasmids of A. baumannii and A. nosocomialis strains, pointing to the possible interspecies transfer of plasmids carrying the mcr-4 gene. Our results highlight that some aquaculture products of Asian origin can represent an important source of variable plasmids carrying mcr genes. The results showed an involvement of phages in the incorporation of the mcr-1 gene into plasmids or the chromosome in E. coli strains from aquaculture. The detection of E. coli with the mcr-1 gene in the chromosome points to the risks associated with the stabilisation of the mcr genes in the bacterial chromosome.

What do we know about plasmids carried by members of the Acinetobacter genus?

Several Acinetobacter spp. act as opportunistic pathogens causing healthcare-associated infections worldwide, and in this respect their ability to resist antimicrobial compounds has certainly boosted up their global propagation. Acinetobacter clinical strains have demonstrated a remarkable ability to evolve and become resistant to almost all available drugs in the antimicrobial arsenal, including the last-resort carbapenem β-lactams. The dissemination of antimicrobial resistant genes (ARG), heavy metals-detoxification systems and other traits such as virulence factors is facilitated by mobile genetic elements (MGE) through horizontal gene transfer. Among them, plasmids have been shown to play a critical role in this genus. Despite the continuous increase of Acinetobacter plasmid sequences present in databases, there are no reports describing the basic traits carried by these MGE. To fill this gap, a broad analysis of the Acinetobacter plasmidome was performed. A search for Acinetobacter complete plasmids indicated that 905 sequences have been deposited in the NCBI-GenBank public database, of which 492 are harbored by Acinetobacter baumannii strains. Plasmid-classification schemes based on Rep proteins homology have so far described 23 different groups for A. baumannii (GR1-23), and 16 Acinetobacter Rep3 Groups (AR3G1-16) for the complete genus. Acinetobacter plasmids size ranges from 1.3 to 400kb. Interestingly, widespread plasmids which are < 20kb make up 56% of the total present in members of this genus. This led to the proposal of Acinetobacter plasmid assignation to two groups according to their size (< 20kb and > 20kb). Usually, smaller plasmids are not self-transmissible, and thereby employ alternative mechanisms of dissemination. For instance, a subgroup of < 20kb-plasmids belonging to the pRAY-family, lack a rep gene, but encode a relaxase enabling their mobilization by conjugative plasmids. Other subgroup, including small GR2 Acinetobacter plasmids, does not encode a relaxase gene. However, they could still be mobilized by conjugative plasmids which recognize an oriT region carried by these small plasmids. Also, these < 20kb-plasmids usually carry accessory genes bordered by XerC/D-recombinases recognition sites which have been hypothesized to mediate plasmid plasticity. Conversely, many cases of larger plasmids are self-transmissible and might encode virulence factors and their regulators, thus controlling strain pathogenicity. The ARGs carried by the > 20kb-plasmids are usually encoded within other MGEs such as transposons, or as part of integrons. It has been recently noted that some of the > 20kb-plasmids are derived from excised phages, and thus dubbed as phage-like plasmids. All in all, the plethora of plasmids found in strains of this genus and the multiple strategies promoting their evolution and dissemination have certainly contributed to survival of the Acinetobacter members in different habitats, including the clinical environment.

Meta-analysis of Pandemic Escherichia coli ST131 Plasmidome Proves Restricted Plasmid-clade Associations

Extraintestinal multidrug resistant Escherichia coli sequence type (ST) 131 is a worldwide pandemic pathogen and a major cause of urinary tract and bloodstream infections. The role of this pandemic lineage in multidrug resistance plasmid dissemination is still scarce. We herein performed a meta-analysis on E. coli ST131 whole-genome sequence (WGS) databases to unravel ST131 plasmidome and specifically to decipher CTX-M encoding plasmids-clade associations. We mined 880 ST131 WGS data and proved that CTX-M-27-encoding IncF[F1:A2:B20] (Group1) plasmids are strictly found in clade C1, whereas CTX-M-15-encoding IncF[F2:A1:B-] (Group2) plasmids exist only in clade C2 suggesting strong plasmid-clade adaptations. Specific Col-like replicons (Col156, Col(MG828), and Col8282) were also found to be clade C1-associated. BLAST-based search revealed that Group1 and Group2 plasmids are narrow-host-range and restricted to E.coli. Among a collection of 20 newly sequenced Israeli ST131 CTX-M-encoding plasmids (2003–2016), Group1 and Group2 plasmids were dominant and associated with the expected clades. We found, for the first time in ST131, a CTX-M-15-encoding phage-like plasmid group (Group3) and followed its spread in the WGS data. This study offers a comprehensive way to decipher plasmid-bacterium associations and demonstrates that the CTX-M-encoding ST131 Group1 and Group2 plasmids are clade-restricted and presumably less transmissible, potentially contributing to ST131 clonal superiority.

Diversity of P1 phage-like elements in multidrug resistant Escherichia coli

The spread of multidrug resistance via mobile genetic elements is a major clinical and veterinary concern. Pathogenic Escherichia coli harbour antibiotic resistance and virulence genes mainly on plasmids, but also bacteriophages and hybrid phage-like plasmids. In this study, the genomes of three E. coli phage-like plasmids, pJIE250-3 from a human E. coli clinical isolate, pSvP1 from a porcine ETEC O157 isolate, and pTZ20_1P from a porcine commensal E. coli, were sequenced (PacBio RSII), annotated and compared. All three elements are coliphage P1 variants, each with unique adaptations. pJIE250-3 is a P1-derivative that has lost lytic functions and contains no accessory genes. In pTZ20_1P and pSvP1, a core P1-like genome is associated with insertion sequence-mediated acquisition of plasmid modules encoding multidrug resistance and virulence, respectively. The transfer ability of pTZ20_1P, carrying antibiotic resistance markers, was also tested and, although this element was not able to transfer by conjugation, it was able to lysogenize a commensal E. coli strain with consequent transfer of resistance. The incidence of P1-like plasmids (~7%) in our E. coli collections correlated well with that in public databases. This study highlights the need to investigate the contribution of phage-like plasmids to the successful spread of antibiotic resistant pathotypes.

Genomic Analysis of Multidrug-Resistant Escherichia coli from Surface Water in Northeast Georgia, United States: Presence of an ST131 Epidemic Strain Containing blaCTX-M-15 on a Phage-Like Plasmid.

Surface water is suspected of playing a role in the development and spread of antimicrobial-resistant (AR) bacteria, including human pathogens. In our previous study, 496 Escherichia coli isolates were recovered from water samples collected over a 2-year period from the Upper Oconee watershed, Athens, GA, United States, of which 34 (6.9%) were AR isolates. Of these, six isolates were selected based on their multidrug resistance (MDR) phenotypes, the presence of mobile genetic elements, and their pathogenic potential and were subjected to whole-genome sequence (WGS) analysis to enhance our understanding of environmental MDR E. coli isolates. This study is the first report on genomic characterization of MDR E. coli from environmental water in the United States through a WGS approach. The sequences of the six MDR E. coli isolates were analyzed and the locations of their AR genes were identified. One of the E. coli isolates was an ST131 epidemic strain, which also produced an extended-spectrum β-lactamase encoded by the blaCTX-M-15 gene, carried on a plasmid that is a member of a very rarely reported family of phage-like plasmids. This is the first time an in-depth sequence analysis has been done on a blaCTX-M-15- containing phage-like plasmid, the presence of which suggests a new emerging mechanism of AR gene transmission.

Emergence of a hybrid plasmid derived from IncN1-F33:A−:B− and mcr-1-bearing plasmids mediated by IS26

To characterize the complete sequences of four plasmids in MCR-1-producing clinical Escherichia coli strain D72, and to depict the formation mechanism and characteristics of the cointegrate plasmid derived from the pD72-mcr1 and pD72-F33 plasmids. The genetic profiles of plasmids in strain D72 and its transconjugant were determined by conjugation, S1-PFGE, Southern hybridization, WGS analysis and PCR. Plasmid sequences were analysed with bioinformatic tools. The traits of the fusion plasmid were characterized by cointegration, stability and conjugation assays. Strain D72, belonging to ST1114, contained four plasmids, including mcr-1-carrying pD72-mcr1, blaCTX-M-55-carrying pD72-F33, blaTEM-238-bearing pD72-IncP and pD72-IncX1 carrying aph(3')-Ia, qnrS2 and floR. A single plasmid, pD72C, in the transconjugant was found to be larger than any plasmid in the original strain D72. Sequence analysis showed that pD72C was the fusion product of pD72-mcr1 and pD72-F33, and the recombinant event involved an intermolecular replicative mechanism. Plasmid fusion occurred at a frequency of 1.75 × 10-4 cointegrates per transconjugant. The fusion plasmid presented a high stability and conjugation frequency of 8.00 × 10-3. To our knowledge, this is the first report of the IS26-mediated fusion of an IncN1-F33:A-:B- plasmid and an mcr-1-carrying phage-like plasmid, providing evidence for the important role of IS26 in the recombination of plasmids. The biological advantages of the fusion plasmid indicated that the fusion event presumably plays a potential role in the dissemination of mcr-1.

A Phage-Like Plasmid Carrying bla KPC-2 Gene in Carbapenem-Resistant Pseudomonas aeruginosa.

Background: Lateral gene transfer plays a central role in the dissemination of carbapenem resistance in bacterial pathogens associated with nosocomial infections, mainly Enterobacteriaceae and Pseudomonas aeruginosa. Despite their clinical significance, there is little information regarding the mobile genetic elements and mechanism of acquisition and propagation of lateral genes in P. aeruginosa, and they remain largely unknown.Objectives: The present study characterized the genetic context of blaKPC-2 in carbapenem-resistant P. aeruginosa strain BH9.Methods: Pseudomonas aeruginosa BH9 sequencing was performed using the long-read PacBio SMRT platform and the Ion Proton System. De novo assembly was carried out using the SMRT pipeline and Canu, and gene prediction and annotation were performed using Prokka and RAST.Results: Pseudomonas aeruginosa BH9 exhibited a 7.1 Mb circular chromosome. However, the blaKPC-2 gene is located in an additional contig composed by a small plasmid pBH6 from P. aeruginosa strain BH6 and several phage-related genes. Further analysis revealed that the beginning and end of the contig contain identical sequences, supporting a circular plasmid structure. This structure spans 41,087 bp, exhibiting all the Mu-like phage landmarks. In addition, 5-bp direct repeats (GGATG) flanking the pBH6 ends were found, strongly indicating integration of the Mu-like phage into the pBH6 plasmid. Mu phages are commonly found in P. aeruginosa. However, for the first time showing a potential impact in shaping the vehicles of the dissemination of antimicrobial (e.g., plasmid pBH6) resistance genes in the Pseudomonas genus.Conclusion: pBH6 captured the Mu-like Phage BH9, creating a co-integrate pBH6::Phage BH9, and this phage-plasmid complex may represent novel case of a phage-like plasmid.

A P7 Phage-Like Plasmid Carrying mcr-1 in an ST15 Klebsiella pneumoniae Clinical Isolate.

A Klebsiella pneumoniae clinical strain, named SCKP83, was isolated and found to be resistant to colistin thanks to the presence plasmid-borne colistin resistant gene mcr-1. The strain was subjected to whole genome sequencing and conjugation experiments. The subsequent analysis indicated that the strain belongs to ST15 and the capsular type K41. In SCKP83, mcr-1 was carried by a 97.4-kb non-self-transmissible plasmid, a 90.9-kb region of which was predicted as an intact phage. This phage was 47.79% GC content, encoded 105 proteins and contained three tRNAs. mcr-1 was located downstream of two copies of the insertion sequence ISApl1 (one complete and one truncated) and was inserted in the ant1 gene, which encodes a putative antirepressor for antagonizing C1 repression, in this phage. The phage is highly similar to phage P7 (77% coverage and 98% identity) from Escherichia coli. Several similar mcr-1-carrying plasmids have been found in E. coli at various locations in China, suggesting that these phage-like plasmids have circulated in China. The findings in this study suggest that the P7 phage-like plasmids are not restricted to E. coli and may represent new vehicles to mediate the inter-species spread of mcr-1.

Complete Sequence of pCY-CTX, a Plasmid Carrying a Phage-Like Region and an ISEcp1-Mediated Tn2Element fromEnterobacter cloacae

A plasmid pCY-CTX carrying a phage-like backbone from an extensively drug-resistant Enterobacter cloacae strain Guangzhou-ECL001 (previously known as CY01) was identified in this study. By Illumina MiSeq 2 × 250-bp paired-end sequencing, de novo assembly, and PCR, full sequence of pCY-CTX was obtained. Plasmid pCY-CTX was a circular plasmid with a length of 116,700 bp, harboring 136 putative open reading frames with the average G + C content of 50.8%. The backbone of pCY-CTX showed high identity to previously reported phage-like plasmid pHCM2 and phage SSU5. In addition, pCY-CTX contained a distinctive ISEcp1-mediated Tn2 region with two resistance genes blaTEM-1 and blaCTX-M-3. Transposition unit "ISEcp1- blaCTX-M-3- orf477" was inserted into the Tn2 structure, dividing Tn2 into two parts. This represents the first identification of a plasmid carrying a phage-like backbone and a distinctive ISEcp1-mediated Tn2 region within blaTEM-1 and blaCTX-M-3 in clinical E. cloacae. The finding of phage-like regions located in plasmids provides a new perspective in gene transfer associated with antimicrobial resistance.

AnCo3, a New Member of the Emerging Family of Phage-Like Plasmids.

ABSTRACTA phage-like plasmid isolated from a clinical isolate of Salmonella enterica serovar Derby has strong nucleotide sequence identity to the phage-like plasmids pSTM_phi isolated from Salmonella enterica serovar Typhimurium L495, AnCo1 and AnCo2 from Escherichia coli 243 and Escherichia coli 244, and the virulent Salmonella-specific SSU5 bacteriophage.

Complete Genome Sequences of Two Phage-Like Plasmids Carrying the CTX-M-15 Extended-Spectrum β-Lactamase Gene.

ABSTRACTTwo similar phage-like plasmids carrying CTX-M-15 resistance cassettes were identified from two environmental Escherichia coli isolates. They demonstrate strong nucleotide sequence identity to the phage-like plasmid pECOH89 and Salmonella bacteriophage SSU5.

A Phage-Like IncY Plasmid Carrying the mcr-1 Gene in Escherichia coli from a Pig Farm in China.

We report here a new type of plasmid that carries the mcr-1 gene, the pMCR-1-P3 plasmid, harbored in an Escherichia coli strain isolated from a pig farm in China. pMCR-1-P3 belongs to the IncY incompatibility group and is a phage-like plasmid that contains a large portion of phage-related sequences. The backbone of this plasmid is different from that of other mcr-1-carrying plasmids reported previously.

Complete genetic analysis of plasmids carrying mcr-1 and other resistance genes in an Escherichia coli isolate of animal origin.

To investigate the genetic features of three plasmids recovered from an MCR-1 and ESBL-producing Escherichia coli strain, HYEC7, and characterize the transmission mechanism of mcr-1 . The genetic profiles of three plasmids were determined by PCR, S1-PFGE, Southern hybridization and WGS analysis. The ability of the mcr-1 -bearing plasmid to undergo conjugation was also assessed. The mcr-1 -bearing transposon Tn 6330 was characterized by PCR and DNA sequencing. Complete sequences of three plasmids were obtained. A non-conjugative phage P7-like plasmid, pHYEC7- mcr1 , was found to harbour the mcr-1 -bearing transposon Tn 6330 , which could be excised from the plasmid by generating a circular intermediate harbouring mcr-1 and the IS Apl1 element. The insertion of the circular intermediate into another plasmid, pHYEC7-IncHI2, could form pHNSHP45-2, the original IncHI2-type mcr-1 -carrying plasmid that was reported. The third plasmid, pHYEC7-110, harboured two replicons, IncX1 and IncFIB, and comprised multiple antimicrobial resistance mobile elements, some of which were shared by pHYEC7-IncHI2. The Tn 6330 element located in the phage-like plasmid pHYEC7- mcr1 could be excised from the plasmid and formed a circular intermediate that could be integrated into plasmids containing the IS Apl1 element. This phenomenon indicated that Tn 6330 is a key element responsible for widespread dissemination of mcr-1 among various types of plasmids and bacterial chromosomes. The dissemination rate of such an element may be further enhanced upon translocation into phage-like vectors, which may also be transmitted via transduction events.

Characterization of a large novel phage-like plasmid in Salmonella enterica serovar Typhimurium.

Salmonella enterica serovar Typhimurium is a food-borne pathogen and a leading cause of gastroenteritis in humans. Recently, we sequenced a phage-type DT108 strain (L945) and found reads with high similarity to both Salmonella typhi strain CT18 plasmid pHCM2 and bacteriophage SSU5. In this study, we completely sequenced the novel phage-like plasmid which was designated as pSTM_Φ. The presence of this phage-like plasmid was examined in a collection of 284 Salmonella Typhimurium isolates using PCR of the parB gene and only one other isolate (L946) was found to carry the phage-like plasmid suggesting that it is infrequently present amongst Salmonella Typhimurium isolates. pSTM_Φ is a circular phage-like plasmid of 107.7 kb encoding 132 coding regions (ORFs) with the majority of the ORFs encoding hypothetical proteins. Comparative analysis with other closely related phage-like plasmids and the SSU5 phage revealed that there were four divergent lineages of phage-like plasmids found in the family of Enterobacteriaceae. In conclusion, pSTM_Φ is a new member of an emerging family of phage-like plasmids.