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Precise genotyping of circular mobile elements from metagenomic data uncovers human-associated plasmids with recent common ancestors.

ABSTRACT: Mobile genetic elements with circular genomes play a key role in the evolution of microbial communities. Their circular genomes correspond to circular walks in metagenome graphs, and yet, assemblies derived from natural microbial communities produce graphs riddled with spurious cycles, complicating the accurate reconstruction of circular genomes. We present DomCycle, an algorithm that reconstructs likely circular genomes based on the identification of so-called "dominant" graph cycles. In the implementation, we leverage paired reads to bridge assembly gaps and scrutinize cycles through a nucleotide-level analysis, making the approach robust to misassembly artifacts. We validated the approach using simulated and real sequencing data. Application of DomCycle to 32 publicly available DNA shotgun sequence data sets from diverse natural environments led to the reconstruction of hundreds of circular mobile genomes. Clustering revealed 20 highly prevalent and cryptic plasmids that have clonal population structures with recent common ancestors. This method facilitates the study of microbial communities that evolve through horizontal gene transfer.

SUBMITTER: Shalon N

PROVIDER: S-EPMC9104695 | biostudies-literature | 2022 May

REPOSITORIES: biostudies-literature

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Precise genotyping of circular mobile elements from metagenomic data uncovers human-associated plasmids with recent common ancestors.

Shalon Nitan N Relman David A DA Yaffe Eitan E

Genome research 20220412 5

Mobile genetic elements with circular genomes play a key role in the evolution of microbial communities. Their circular genomes correspond to circular walks in metagenome graphs, and yet, assemblies derived from natural microbial communities produce graphs riddled with spurious cycles, complicating the accurate reconstruction of circular genomes. We present DomCycle, an algorithm that reconstructs likely circular genomes based on the identification of so-called "dominant" graph cycles. In the im ...[more]

PMID: 35414589

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Project description:Antimicrobial resistance genes (ARG) are commonly found on acquired mobile genetic elements (MGEs) such as plasmids or transposons. Understanding the spread of resistance genes associated with mobile elements (mARGs) across different hosts and environments requires linking ARGs to the existing mobile reservoir within bacterial communities. However, reconstructing mARGs in metagenomic data from diverse ecosystems poses computational challenges, including genome fragment reconstruction (assembly), high-throughput annotation of MGEs, and identification of their association with ARGs. Recently, several bioinformatics tools have been developed to identify assembled fragments of plasmids, phages, and insertion sequence (IS) elements in metagenomic data. These methods can help in understanding the dissemination of mARGs. To streamline the process of identifying mARGs in multiple samples, we combined these tools in an automated high-throughput open-source pipeline, MetaMobilePicker, that identifies ARGs associated with plasmids, IS elements and phages, starting from short metagenomic sequencing reads. This pipeline was used to identify these three elements on a simplified simulated metagenome dataset, comprising whole genome sequences from seven clinically relevant bacterial species containing 55 ARGs, nine plasmids and five phages. The results demonstrated moderate precision for the identification of plasmids (0.57) and phages (0.71), and moderate sensitivity of identification of IS elements (0.58) and ARGs (0.70). In this study, we aim to assess the main causes of this moderate performance of the MGE prediction tools in a comprehensive manner. We conducted a systematic benchmark, considering metagenomic read coverage, contig length cutoffs and investigating the performance of the classification algorithms. Our analysis revealed that the metagenomic assembly process is the primary bottleneck when linking ARGs to identified MGEs in short-read metagenomics sequencing experiments rather than ARGs and MGEs identification by the different tools.

Project description:qnrD is a plasmid mediated quinolone resistance gene from unknown origin, recently described in Enterobacteriaceae. It encodes a pentapeptide repeat protein 36-60% different from the other Qnr (A, B, C, S and VC). Since most qnrD-positive strains were described as strains belonging to Proteus or Providencia genera, we hypothesized that qnrD originated in Proteeae before disseminating to other enterobacterial species. We screened 317 strains of Proteeae for qnrD and its genetic support by PCR. For all the seven qnrD-positive strains (4 Proteus mirabilis, 1 Proteus vulgaris and 2 Providencia rettgeri) the gene was carried onto a small non-transmissible plasmid, contrarily to other qnr genes that are usually carried onto large multi-resistant plasmids. Nucleotide sequences of the qnrD-bearing plasmids were 96% identical. Plasmids contained 3 ORFs apart from qnrD and belonged to an undescribed incompatibility group. Only one plasmid, in P. vulgaris, was slightly different with a 1,568-bp insertion between qnrD and its promoter, leading to absence of quinolone resistance. We sought for similar plasmids in 15 reference strains of Proteeae, but which were tested negative for qnrD, and found a 48% identical plasmid (pVERM) in Providencia vermicola. In order to explain how qnrD could have been inserted into such native plasmid, we sought for gene mobilization structures. qnrD was found to be located within a mobile insertion cassette (mic) element which sequences are similar to one mic also found in pVERM. Our conclusions are that (i) the small non-transmissible qnrD-plasmids described here may result from the recombination between an as-yet-unknown progenitor of qnrD and pVERM, (ii) these plasmids are maintained in Proteeae being a qnrD reservoir (iii) the mic element may explain qnrD mobilization from non-transmissible plasmids to mobilizable or conjugative plasmids from other Enterobacteriaceae, (iv) they can recombined with larger multiresistant plasmids conjugated in Proteeae.

Dataset Information

Precise genotyping of circular mobile elements from metagenomic data uncovers human-associated plasmids with recent common ancestors.

Publications

Precise genotyping of circular mobile elements from metagenomic data uncovers human-associated plasmids with recent common ancestors.

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