Investigating Possible Interspecies Communication of Plasmids Associated with Transfer of Third-Generation Cephalosporin, Quinolone, and Colistin Resistance Between Simultaneously Isolated Escherichia Coli and Klebsiella Pneumoniae

Abstract

The coinfection process producing multiple species of pathogens provides a specific ecological niche for the exchange of genetic materials between pathogens, in which plasmids play a vital role in horizontal gene transfer, especially for drug resistance, but the underlying transfer pathway remains unclear. Interspecies communication of the plasmids associated with the transfer of third-generation cephalosporins, quinolones, and colistin resistance has been observed in simultaneously isolated Escherichia coli and Klebsiella pneumoniae from abdominal drainage following surgery. The MICs of antimicrobial agents were determined by the broth microdilution method. The complete chromosome and plasmid sequences were obtained by combining Illumina paired-end short reads and MinION long reads. S1-PFGE, southern blot analysis and conjugation assay confirmed the transferability of the mcr-1-harboring plasmid. Both the E. coli isolate EC15255 and K. pneumoniae isolate KP15255 from the same specimen presented multidrug resistance. Each of them harbored one chromosome and three plasmids, and two plasmids and their mediated resistance could be transferred to the recipient by conjugation. Comparison of their genome sequences suggested that several genetic communication events occurred between species, especially among their plasmids, such as whole-plasmid transfer, insertion, deletion, amplification, or inversion. Exchange of plasmids or the genetic elements they harbor plays a critical role in antimicrobial resistance gene transmission and poses a substantial threat to nosocomial infection control, necessitating the continued surveillance of multidrug resistant pathogens, especially during coinfection. IMPORTANCE The genome sequence of bacterial pathogens commonly provides a detailed clue of genetic communication among clones or even distinct species. The intestinal microecological environment is a representative ecological niche for genetic communication. However, it is still difficult to describe the details of horizontal gene transfer or other genetic events within them because the evidence in the genome sequence is incomplete and limited. In this study, the simultaneously isolated Escherichia coli and Klebsiella pneumoniae from a coinfection process provided an excellent example for observation of interspecies communication between the two genomes and the plasmids they harbor. A complete genome sequence acquired by combining the Illumina and MinION sequencing platforms facilitated the understanding of genetic communication events, such as whole-plasmid transfer, insertion, deletion, amplification, or inversion, which contribute to antimicrobial resistance gene transmission and are a substantial threat to nosocomial infection control.

Keywords: IS26; colistin; interspecies; mcr-1; plasmid.

FIG 1

S1-digested plasmid DNA and Southern blot hybridization with the mcr-1 gene of E. coli isolate EC15255, K. pneumoniae isolate KP15255, and their transconjugants. The red arrows indicate positive signals via southern blot hybridization with the mcr-1 probes.

FIG 2

The schematic relationship among plasmids in E. coli and K. pneumoniae based on sequence similarities was visualized by Circos. For the circle, three arc segments from 7 to 11 o'clock clockwise (left part of the circle) represent the large, middle, and small plasmids in E. coli, respectively. Similarly, another three arc segments at the right part of the circle represent the three plasmids in K. pneumoniae. The ESBL gene bla_CTX-M-14, quinolone resistance gene qnrS1 and colistin resistance gene mcr-1.1 are marked with their relative position on the plasmid by small red arrows. The small triangles on the plasmid indicate the IS26 elements at multiple sites.

FIG 3

Comparison of the TnAs1-IS903B-bla_CTX-M-14-ISEc9 cassette between the plasmid pEC15255_2 and pKP15255_2 backgrounds. The yellow flag represents the direct repeats (DRs) (CTATATTG).