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

Abstract

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 bla_IMP-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 bla_IMP-26, mcr-9, and two copies of bla_TEM-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 bla_TEM-1B gene. Remarkably, this study first identified a multidrug-resistant S. marcescens strain co-harboring bla_IMP-26 and mcr-9 on a megaplasmid pYL4.1 and also included a proposed evolutionary pathway of epidemic megaplasmids carrying bla_IMP-26.

Keywords: Serratia marcescens; blaIMP-26; evolutionary pathway; mcr-9; megaplasmids IMP-26; multidrug resistance.

Figure 1

Genetic features of the pYL4.1 plasmid. Inner and outer circles correspond to average G+C content (black circle) and GC skew information (green and purple circles, respectively). The colored circles represent different plasmids (details are in the legend), and the Genbank numbers are as follows: pIMP26 (MH399264), pEHZJ1 (CP033103), pEC-IMPQ (EU855788), pGMI14-002 (CP028197), p505108-MDR (KY978628). An outer cyan-blue circle shows where resistance genes and IntI are located. The location of the resistance genes (magenta), insert elements (blue), and replication origins (orange) are also shown outside the round and tagged by different colors.

Figure 2

Genetic content of bla_IMP-26. Comparison of the genetic environment of bla_IMP-26 between different plasmids. Magenta arrows indicate resistance genes, and blue arrows indicate insertion sequences. Shading regions denote homologous regions (>95% nucleotide identity).

Figure 3

Comparison of the genetic environment of mcr-9 in different plasmids. Magenta arrows indicate resistance genes, blue arrows indicate insertion sequences, green arrows indicate identified ORFs, and gray arrows represent hypothetical genes. Shaded regions denote homologous regions (>95% nucleotide identity).

Figure 4

Evolutionary path between pEHZJ1 (CP033103), pYL4 (CP083755), and pIMP-26 (MH399264). The arrows indicate ORFs that have been confirmed or putatively confirmed and their orientations. Length of the arrow corresponds to the predicted ORF. The color code represents: replication initiation protein genes (orange), integrase genes (red), transposase genes and insertion elements (blue), antibiotic resistance genes (magenta), toxin-antitoxin system genes (yellow), heavy metal resistance genes (green), conjugal transfer genes (cray), putative, hypothetical, and other genes are represented by gray arrows. Shaded gray areas denote homologous regions with up to 95% nucleotide identity. When the shadow areas overlap, the color of the shaded region is darkened.

Figure 5

Structure of phage-like plasmid pYL4.2 carrying bla_TEM-1B from S. marcescens isolate YL4. The arrows with different colors represent open reading frames (ORFs), with purple, orange, and blue representing mobile elements, resistance genes, and ORFs, respectively. Inner and outer circles correspond to average G+C content (black circle) and GC skew information (green and purple circles), respectively.