The complete nucleotide sequence of the multi-drug resistance-encoding IncL/M plasmid pACM1
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Updated: Apr 22, 2026
Testing the Role of Multicopy Plasmids in the Evolution of Antibiotic Resistance
Published on: May 2, 2018
Karen E Preston1, Sandra A Hitchcock2, Abdullah Y Aziz2
1Center for Functional Genomics, University at Albany, SUNY, 1 Discovery Dr., Rensselaer, NY 12144-3452, USA.
Abstract:
The 89,977 bp nucleotide sequence of pACM1, isolated from a 1993 outbreak strain of cephalosporin-resistant Klebsiella oxytoca, has been completed and assigned GenBank accession number KJ541681. The plasmid has a single 31,842 bp mosaic multi-drug resistance-encoding (MDR) region comprising the mer resistance module of Tn1696, two integrons with a total of seven cassettes, one complete copy each of IS1R and IS26, and the bla(SHV-5)-carrying Tn2003 (with defective IS26 termini), all within a Tn1721-like element inserted into the mucB gene of the IncL/M plasmid backbone. The Tn1721-Tn1696 combination resembles sequence found in the chromosomal MDR islands of some Acinetobacter baumannii isolates. Among the completely sequenced IncL/M resistance plasmids, the Tn1721-based MDR region is unique, but data from older studies suggest that this type of plasmid was widespread in the 1990s. Since resistance gene dosage is affected by plasmid copy number (PCN), we used a relatively simple new "efficiency-corrected" qPCR assay to measure the PCN of pACM1. There are approximately three copies per chromosome in an Escherichia coli DH5α host, and two in the original Klebsiella oxytoca isolate. We could not find similar PCN data for other medically important plasmids for comparison. The study of this plasmid property and its effect on resistance levels should be facilitated in the future by the availability of qPCR instruments and complete genome sequences.
Insights
The pACM1 plasmid, carrying cephalosporin resistance genes, was sequenced. Its multi-drug resistance region is unique, and its plasmid copy number was measured using qPCR, providing insights into antibiotic resistance.
Area of Science:
- Microbiology
- Genomics
- Molecular Biology
Background:
- The pACM1 plasmid was isolated from a cephalosporin-resistant Klebsiella oxytoca strain in 1993.
- Plasmid sequence data is crucial for understanding antibiotic resistance mechanisms.
Purpose of the Study:
- To complete and analyze the nucleotide sequence of the pACM1 plasmid.
- To characterize the multi-drug resistance (MDR) region of pACM1.
- To measure the plasmid copy number (PCN) of pACM1 using a novel qPCR assay.
Main Methods:
- Whole-genome sequencing of the pACM1 plasmid.
- Bioinformatic analysis to identify resistance genes and genetic elements.
- Quantitative Polymerase Chain Reaction (qPCR) assay to determine PCN.
Main Results:
- The 89,977 bp sequence of pACM1 (GenBank accession KJ541681) was determined, featuring a unique 31,842 bp MDR region with multiple resistance cassettes and transposons.
- The MDR region contains mer, integrons, IS1R, IS26, and Tn2003 (blaSHV-5) within a Tn1721-like element inserted into the mucB gene of an IncL/M plasmid.
- The PCN of pACM1 was found to be approximately three copies per chromosome in E. coli DH5α and two copies in the original K. oxytoca isolate.
Conclusions:
- The pACM1 plasmid possesses a unique MDR region, distinct from other sequenced IncL/M plasmids.
- The developed qPCR assay provides a reliable method for determining PCN, which influences resistance gene dosage.
- Further studies on PCN and its impact on resistance levels are facilitated by complete genome sequences and qPCR technology.
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