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p39R861-4, A Type 2 A/C2 Plasmid Carrying a Segment from the A/C1 Plasmid RA1
Sashindran Anantham1, Christopher J Harmer1, Ruth M Hall1
1School of Molecular Bioscience, The University of Sydney , Sydney, Australia .
Abstract:
The largest plasmid in the strain 39R861, which is used as a plasmid size standard, was recovered by conjugation and sequenced to determine its exact size. Plasmid p39R861-4 transferred at high frequency. Although reported to be the A/C1 plasmid RA1, p39R861-4 is a 155794-bp Type 2 A/C2 plasmid, in which a 39-kb segment, derived from RA1 that includes a relative of the RA1 resistance island, replaces 26.5 kb of the Type 2 backbone. p39R861-4 includes a single copy of IS10 and two resistance islands with a CR2-sul2 region in each of them. The 84 kb of backbone between the resistance islands is inverted relative to other known A/C plasmids and this inversion has arisen through recombination between the CR2-sul2 regions that are inversely oriented. The two resistance islands present before this inversion occurred were one related to but longer than that found in RA1, and one that is a form of the ARI-B island and identical to ARI-B in the A/C2 plasmid R55. They contain genes conferring resistance to tetracycline (tetA(D)), sulfonamides (sul2), and florfenicol and chloramphenicol (floR). The tet(D) determinant is flanked by two IS26 in a transposon-like structure named Tntet(D). Both resistance islands contain remnants of the two ends of the integrative element GIsul2, consistent with the sul2 gene being mobilized by GIsul2 rather than by CR2.
Insights
The A/C2 plasmid p39R861-4, a plasmid size standard, was sequenced revealing a novel structure with an inverted backbone and two resistance islands. This detailed characterization enhances understanding of plasmid evolution and antimicrobial resistance.
Area of Science:
- Microbiology
- Genomics
- Molecular Biology
Background:
- Plasmid p39R861-4 is utilized as a size standard.
- Previous reports identified it as the A/C1 plasmid RA1.
Purpose of the Study:
- To determine the exact size and sequence of plasmid p39R861-4.
- To elucidate the structural organization and genetic elements of p39R861-4.
Main Methods:
- Conjugation to recover the plasmid.
- Whole-genome sequencing to determine the complete nucleotide sequence.
Main Results:
- Plasmid p39R861-4 is a 155,794-bp Type 2 A/C2 plasmid, not A/C1 RA1.
- It contains a 39-kb segment from RA1 replacing part of the Type 2 backbone.
- An 84-kb backbone segment is inverted due to recombination between inversely oriented CR2-sul2 regions.
- Two resistance islands confer resistance to tetracycline (tetA(D)), sulfonamides (sul2), and florfenicol/chloramphenicol (floR).
- The tet(D) determinant is within a Tntet(D) structure flanked by IS26.
- Remnants of GIsul2 in resistance islands suggest sul2 mobilization by GIsul2.
Conclusions:
- Plasmid p39R861-4 exhibits a unique mosaic structure with significant backbone inversion.
- Its genetic makeup provides insights into the evolution of antimicrobial resistance genes and their mobilization.
- The findings refine the characterization of plasmid standards and contribute to understanding plasmid diversity.
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