Diverse new plasmid structures and antimicrobial resistance in strains isolated from perianal abscess patients
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Zhen Xu, Lulu Shi, Tao Meng, Mei Luo, Jiaming Zhu, Mingyu Wang, Wenlong Shen
Some bacteria carry small, separate DNA circles that can pick up useful traits and pass them on quickly. In this study, those circles included previously unseen routes to resistance against a last-resort antibiotic.
Introduction: Plasmids, the most important and versatile bacterial extrachromosomal DNA Molecules, has have been a center central topic for bacterial genetics and biology. However, the inability of short-read high-throughput sequencing methods to reliably assemble plasmids makes it difficult to investigate the diversity of plasmid structures and functions. Methods: In this work, we used the long-read Nanopore sequencing method to address this issue, by producing high quality whole genome sequences of 33 bacterial strains from 11 perianal abscess-suffering patients. Results and discussion: Successful high quality assemblies were generated with this method, including 20 perfect assemblies out of 33 genomes. A total of 47 plasmids were identified from the bacterial strains, including 12 unique, newly identified, high quality circular plasmids. These plasmids were further subject to structural analysis, leading to the finding of significant diversification from previously known plasmids, suggesting the diversity of plasmid structure and function. Particularly, two mcr10.1-harboring conjugative plasmids were found from Citrobacter portucalensis and Enterobacter kobei, which were not previously reported. This works shows the feasibility of using long-read sequencing to identify plasmids, and the high diversity of plasmid structure and function that awaits further surveillance.
Transcript
Some bacteria carry small, separate DNA circles that can pick up useful traits and pass them on quickly. In this study, those circles included previously unseen routes to resistance against a last-resort antibiotic. Besides the main chromosome, bacteria normally often carry circular DNA molecules called plasmids for additional functions that are not encoded on the chromosome.
Plasmids are the variable parts of the bacterial genome, bringing quick adaptation and beneficial traits while keeping the main genome small. Think of them like detachable instruction cards passed between workers: instead of waiting for a whole organization to change, one worker can copy a useful card and hand it to another.
Plasmids can be replicated and transferred between bacterial cells, leading to very quick acquisition of functions. The work aimed to understand how much remains unknown about plasmid diversity and function, and how improved long-read sequencing technologies could address that problem.
The approach was to identify plasmids in bacteria from clinical samples, check how many had not been observed before, and identify new plasmid structures and functions. Samples from people with perianal abscesses were used because the disease is caused by bacterial infection, and commonly used antibiotic treatment might create stress linked to enriched plasmid occurrence.
Long-read sequencing produced high-quality whole-genome sequences for all the isolated bacterial strains, with an average of 3.15 assembled pieces per bacterium. A total of 12 new plasmids were found, which was quite a surprise. The new plasmids had a size distribution close to all the plasmids found, with an average size of 58.80 kilobases.
All 12 new plasmids were assembled into circles, confirming the quality of the sequences. The three newly identified large plasmids were mobile, resistant to multiple drugs, and carried systems that can help move material between bacterial cells.
All of them carried multiple replication proteins, suggesting that repeated joining and rearrangement events helped create these complex plasmids. All three plasmids belonged to the same broad plasmid group. The presence of antibiotic-resistance genes was investigated in the isolated strains.
Only one bacterial strain did not carry any antibiotic-resistance genes. Most of the antibiotic-resistance genes were carried on plasmids, suggesting that resistance was widespread. One particularly interesting finding was a new mobile plasmid carrying a resistance gene in a bacterial species not previously reported to host it.
That gene gives resistance to polymyxin, an antibiotic of last resort that has received tremendous attention in recent years. The plasmid also carried the machinery needed for transfer between bacteria, expanding knowledge of the range of bacterial hosts that can carry this resistance.
The study found 12 plasmids that had not been observed before, possibly because long-read sequencing made accurate discovery and surveillance possible. Compared with their closest known counterparts, these new plasmids showed significant differences, confirming that plasmids change strongly and contribute to the flexibility of bacterial genomes.
Two mobile plasmids carrying resistance to polymyxin were found in two bacterial species, including the first reported finding of this resistance gene in one of them. The study found that bacterial DNA circles are far more varied than routine reading methods reveal, including mobile resistance carried by an unexpected bacterial host.
That matters because better tracking can reveal resistance before it spreads unnoticed.
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