PMC6067848_01
Clinical description of a patient undergoing antibiotic treatment.. A. The history of antibiotic exposure for a patient admitted into the hospitalThe colored text and bars represent the sources from which the bacterial species were isolated. S-E. coli, sensitive-E. coli; CR-E. coli, Carbapenem-resistant E. coli B. Dynamic alterations of body temperature and leukocyte counts of the patient recorded during the hospitalization period
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On 3 June 2017, a 58-year-old man was admitted to the Lishui People's Hospital of Zhejiang Province, China (Figure 1(A)), and was diagnosed to be suffering from Small Intestinal Perforation with Peritonitis (SIPWP). While in hospital, the inflammatory markers (temperature and Leukocyte count) of this patient were normal (Figure 1(B)). To probe the suspected bacterial agent, the specimen of peritoneal fluid from the patient (Figure 1(A)) was inoculated on a petri plate with MacConkey agar. Following 2-weeks of antibiotic therapy, which proved to be a clinically-successful anti-infection treatment (Figure 1(A)), the abdominal drainage and the blood of the patient were sampled for bacterial cultivation. Thus, it was appropriately designated as CR-E. coli LSB54 (Figure 1(A)).
Genomic map of the mcr-1-carrying plasmid pLSB54-mcr-1.. The plasmid map was generated using GenomeVx with automatic coloring.
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As we recently described with mcr-1-bearing plasmids carried by commensal E. coli, whole genome sequencing was performed, giving evidence that two unique plasmids coexist in the MDR-E. coli strain LSB54 (Figure 2-4). Among the two plasmids, pLSB54-mcr-1 is the mcr-1-harboring plasmid (Figures 2 and 4(A)), and pLSB54-NDM-5 is the NDM-5-carrying plasmid (Figures 3 and 4(A)). Subsequent plasmid replicon typing (https://cge.cbs.dtu.dk/services/PlasmidFinder/) revealed that pLSB54-mcr-1 (Acc. no.: MG773376) is a IncHI2-type plasmid with a genome size of 251.657 kb (Figure 2), whereas pLSB54-NDM-5 (Acc. no.: MG773377) is a IncX3-like plasmid whose genome is 46.19 kb long (Figure 3).
Genomic map of the NDM-5-producing plasmid pLSB54-NDM-5.. The plasmid map was generated using GenomeVx with automatic coloring.
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As we recently described with mcr-1-bearing plasmids carried by commensal E. coli, whole genome sequencing was performed, giving evidence that two unique plasmids coexist in the MDR-E. coli strain LSB54 (Figure 2-4). Among the two plasmids, pLSB54-mcr-1 is the mcr-1-harboring plasmid (Figures 2 and 4(A)), and pLSB54-NDM-5 is the NDM-5-carrying plasmid (Figures 3 and 4(A)). Subsequent plasmid replicon typing (https://cge.cbs.dtu.dk/services/PlasmidFinder/) revealed that pLSB54-mcr-1 (Acc. no.: MG773376) is a IncHI2-type plasmid with a genome size of 251.657 kb (Figure 2), whereas pLSB54-NDM-5 (Acc. no.: MG773377) is a IncX3-like plasmid whose genome is 46.19 kb long (Figure 3).
Comparative genomics of pLSB54-mcr-1 and pLSB54-NDM-5.. A. Cartoon description of the E. coli LSB54 strain carrying the two plasmids pLSB54-mcr-1 and pLSB54-NDM-5B. Colinear genome analyzes of three mcr-1-harboring plasmids pLSB54-mcr-1, pHNSHP45-2 (KU341381) and p2474-MCR1 (CP021209)C. Genomic comparison of the plasmid pLSB54-NDM-5 with two other NDM-5-expressing plasmids pCREC-591_4 (CP024825) and pNDM_MGR194 (KF220657)Boxed arrows represent the position and transcriptional direction of the ORFs. Regions of > 99% identity are given by green shading. Genes associated with the tra and pil loci are colored light blue, antibiotic resistance genes are colored red, insertion sequences are colored green, and other genes are colored black.
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As we recently described with mcr-1-bearing plasmids carried by commensal E. coli, whole genome sequencing was performed, giving evidence that two unique plasmids coexist in the MDR-E. coli strain LSB54 (Figure 2-4). Among the two plasmids, pLSB54-mcr-1 is the mcr-1-harboring plasmid (Figures 2 and 4(A)), and pLSB54-NDM-5 is the NDM-5-carrying plasmid (Figures 3 and 4(A)). Colinear alignment analyzes of the plasmid genomes indicated that pLSB54-mcr-1 is almost identical with the prototypical mcr-1-harboring plasmid pHNSHP45-2 (Acc. no.: KU341381) (Figure 4(B)), and possesses 90% coverage and 99% identity with p2474-MCR1 (Acc. no.: CP021209), another mcr-1-positive plasmid from a clinical E. coli isolate in China (Figure 4(B)). As for pLSB54-NDM-5, it presents 100% coverage with the NDM-5-producing IncX3 plasmid pCREC-591_4 (Acc. no.: CP024825) of E. coli isolated from South Korea (Figure 4(C)) and exhibits a 99% identity to the plasmid pNDM_MGR194 (KF220657) of Klebsiella pneumoniae seen in India (Figure 4(C)). Whereas, the insertion loci of NDM-5 are quite different in the above three plasmids (Figure 4(C)). IncX3-X4 plasmid in E. coli, mcr-1 and blaNDM-5 are separately encoded by two distinct plasmids in the clinical E. coli isolate LSB54 (Figure 4(A)). In this clinical case, the MDR-E. coli which possesses resistance to colistin and carbapenem (two lines of last-resort antibiotics), emerges in a patient undergoing antibiotic treatment with
Genetic contents of mcr-1 (and/or blaNDM-5).. A. The genetic environment of mcr-1 and its neighboring lociB. The genetic environment of blaNDM-5 and its neighboring lociThe arrows indicate open reading frames, resistance genes, insertion sequences and accessory genes are separately shown with red, green and gray arrows.
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Fine mapping of the genetic environment surrounding the mcr-1(blaNDM-5) gene further illustrated that i) In the IncHI2-type plasmid pLSB54-mcr-1, the insertion sequence ISApl1 is located upstream of the mcr-1 gene (Figure 5(A)) and the ISApl1-mcr-1-pap2 fragment is inserted downstream of the terF gene, just like the plasmids pHNSHP45-2 (Acc. no.: KU341381) and p2474-MCR (Acc. no.: CP021209) (Figure 5(A)); ii) The NDM-5-containing fragment is identical in all three IncX3-like plasmids, including pLSB54-NDM-5 (Figure 5(B)), and the blaNDM-5 is adjacent to the insertion sequence IS5, which is flanked by a truncated ISAba125 sequence (Figure 5(B)).