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Bioinformatics (Oxford, England) May 2023As prevalent extrachromosomal replicons in many bacteria, plasmids play an essential role in their hosts' evolution and adaptation. The host range of a plasmid refers to...
MOTIVATION
As prevalent extrachromosomal replicons in many bacteria, plasmids play an essential role in their hosts' evolution and adaptation. The host range of a plasmid refers to the taxonomic range of bacteria in which it can replicate and thrive. Understanding host ranges of plasmids sheds light on studying the roles of plasmids in bacterial evolution and adaptation. Metagenomic sequencing has become a major means to obtain new plasmids and derive their hosts. However, host prediction for assembled plasmid contigs still needs to tackle several challenges: different sequence compositions and copy numbers between plasmids and the hosts, high diversity in plasmids, and limited plasmid annotations. Existing tools have not yet achieved an ideal tradeoff between sensitivity and precision on metagenomic assembled contigs.
RESULTS
In this work, we construct a hierarchical classification tool named HOTSPOT, whose backbone is a phylogenetic tree of the bacterial hosts from phylum to species. By incorporating the state-of-the-art language model, Transformer, in each node's taxon classifier, the top-down tree search achieves an accurate host taxonomy prediction for the input plasmid contigs. We rigorously tested HOTSPOT on multiple datasets, including RefSeq complete plasmids, artificial contigs, simulated metagenomic data, mock metagenomic data, the Hi-C dataset, and the CAMI2 marine dataset. All experiments show that HOTSPOT outperforms other popular methods.
AVAILABILITY AND IMPLEMENTATION
The source code of HOTSPOT is available via: https://github.com/Orin-beep/HOTSPOT.
Topics: Phylogeny; Plasmids; Software; Metagenome; Metagenomics; Bacteria
PubMed: 37086432
DOI: 10.1093/bioinformatics/btad283 -
MBio May 2021Plasmids play an important role in bacterial evolution by transferring niche-adaptive functional genes between lineages, thus driving genomic diversification. Bacterial...
Plasmids play an important role in bacterial evolution by transferring niche-adaptive functional genes between lineages, thus driving genomic diversification. Bacterial genomes commonly contain multiple, coexisting plasmid replicons, which could fuel adaptation by increasing the range of gene functions available to selection and allowing their recombination. However, plasmid coexistence is difficult to explain because the acquisition of plasmids typically incurs high fitness costs for the host cell. Here, we show that plasmid coexistence was stably maintained without positive selection for plasmid-borne gene functions and was associated with compensatory evolution to reduce fitness costs. In contrast, with positive selection, plasmid coexistence was unstable despite compensatory evolution. Positive selection discriminated between differential fitness benefits of functionally redundant plasmid replicons, retaining only the more beneficial plasmid. These data suggest that while the efficiency of negative selection against plasmid fitness costs declines over time due to compensatory evolution, positive selection to maximize plasmid-derived fitness benefits remains efficient. Our findings help to explain the forces structuring bacterial genomes: coexistence of multiple plasmids in a genome is likely to require either rare positive selection in nature or nonredundancy of accessory gene functions among the coexisting plasmids. Bacterial genomes often contain multiple coexisting plasmids that provide important functions like antibiotic resistance. Using lab experiments, we show that such plasmid coexistence within a genome is stable only in environments where the function they encode is useless but is unstable if the function is useful and beneficial for bacterial fitness. Where competing plasmids perform the same useful function, only the most beneficial plasmid is kept by the cell, a process that is similar to competitive exclusion in ecological communities. This process helps explain how bacterial genomes are structured: bacterial genomes expand in size by acquiring multiple plasmids when selection is relaxed but subsequently contract during periods of strong selection for the useful plasmid-encoded function.
Topics: Adaptation, Physiological; Bacteria; Genetic Fitness; Genome, Bacterial; Phenotype; Plasmids; Selection, Genetic
PubMed: 33975933
DOI: 10.1128/mBio.00558-21 -
Genes Nov 2023Plasmids mediate gene exchange across taxonomic barriers through conjugation, shaping bacterial evolution for billions of years. While plasmid mobility can be harnessed...
Plasmids mediate gene exchange across taxonomic barriers through conjugation, shaping bacterial evolution for billions of years. While plasmid mobility can be harnessed for genetic engineering and drug-delivery applications, rapid plasmid-mediated spread of resistance genes has rendered most clinical antibiotics useless. To solve this urgent and growing problem, we must understand how plasmids spread across bacterial communities. Here, we applied machine-learning models to identify features that are important for extending the plasmid host range. We assembled an up-to-date dataset of more than thirty thousand bacterial plasmids, separated them into 1125 clusters, and assigned each cluster a distribution possibility score, taking into account the host distribution of each taxonomic rank and the sampling bias of the existing sequencing data. Using this score and an optimized plasmid feature pool, we built a model stack consisting of DecisionTreeRegressor, EvoTreeRegressor, and LGBMRegressor as base models and LinearRegressor as a meta-learner. Our mathematical modeling revealed that sequence brevity is the most important determinant for plasmid spread, followed by P-loop NTPases, mobility factors, and β-lactamases. Ours and other recent results suggest that small plasmids may broaden their range by evading host defenses and using alternative modes of transfer instead of autonomous conjugation.
Topics: Host Specificity; Plasmids; Bacteria; beta-Lactamases; Machine Learning
PubMed: 38002987
DOI: 10.3390/genes14112044 -
Journal of Medical Microbiology Dec 2023. Various plasmid-mediated resistance genes have been reported in , but little is known about their global distribution features, evolution pattern and spread.. The...
. Various plasmid-mediated resistance genes have been reported in , but little is known about their global distribution features, evolution pattern and spread.. The potential mobilization mechanisms of resistance plasmids in have been poorly explored.. The aim of the study was to investigate the prevalence and diversity of plasmid-mediated resistance genes among isolates, and focus on the analysis of the features of the resistance plasmids from .. The plasmids tested were sequenced using the Illumina HiSeq platform in conjunction with PCR and inverted PCR. The susceptibility of the host strains was determined by broth microdilution. The transfer of plasmids tested was conducted by electroporation. The sequence data were compared using bioinformatics tools and the data from our laboratory and the National Center for Biotechnology Information (NCBI) database.. Nineteen plasmids were identified from our laboratory and these resistance plasmids were functional and transferable. Moreover, we clustered five types of genetic backbones of plasmids from and revealed the global distribution features of the plasmid-mediated resistance genes.. This is the first report of the coexistence of (H)-bearing type I plasmid and (C)-bearing type II plasmid in one clinical isolate. In addition, this study provides the first view of the global distribution of plasmid-mediated resistance genes and classifies the plasmids in according to their backbone regions.
Topics: Plasmids; Haemophilus parasuis; Base Sequence
PubMed: 38112519
DOI: 10.1099/jmm.0.001767 -
Current Microbiology Jan 2022Caulobacter is a well-studied bacterial genus, but little is known about the plasmids that are found in some wild Caulobacter isolates. We used bioinformatic approaches...
Caulobacter is a well-studied bacterial genus, but little is known about the plasmids that are found in some wild Caulobacter isolates. We used bioinformatic approaches to identify nine plasmids from seven different Caulobacter strains and grouped them based on their size and the similarity of their repABC, parAB, and mobAB genes. Protein pathway analysis of the genes on the K31p1 and K31p2 plasmids showed many metabolic pathways that would enhance the metabolic versatility of the host strain. In contrast, the CB4 plasmid contained 21 heavy metal resistance genes with the majority coding for proteins that enhance copper resistance. Growth assays of C. henricii CB4 demonstrated increased copper resistance and quantitative PCR showed an increase in the expression of eight heavy metal genes when induced with copper.
Topics: Bacteria; Caulobacter; Copper; Metals, Heavy; Plasmids
PubMed: 34982248
DOI: 10.1007/s00284-021-02742-z -
MicrobiologyOpen Oct 2022Recent research demonstrated that some Listeria monocytogenes plasmids contribute to stress survival. However, only a few studies have analyzed gene expression patterns...
Recent research demonstrated that some Listeria monocytogenes plasmids contribute to stress survival. However, only a few studies have analyzed gene expression patterns of L. monocytogenes plasmids. In this study, we identified four previously published stress-response-associated transcriptomic data sets which studied plasmid-harboring L. monocytogenes strains but did not include an analysis of the plasmid transcriptomes. The four transcriptome data sets encompass three distinct plasmids from three different L. monocytogenes strains. Differential gene expression analysis of these plasmids revealed that the number of differentially expressed (DE) L. monocytogenes plasmid genes ranged from 30 to 45 with log fold changes of -2.2 to 6.8, depending on the plasmid. Genes often found to be DE included the cadmium resistance genes cadA and cadC, a gene encoding a putative NADH peroxidase, the putative ultraviolet resistance gene uvrX, and several uncharacterized noncoding RNAs (ncRNAs). Plasmid-encoded ncRNAs were consistently among the highest expressed genes. In addition, one of the data sets utilized the same experimental conditions for two different strains harboring distinct plasmids. We found that the gene expression patterns of these two L. monocytogenes plasmids were highly divergent despite the identical treatments. These data suggest plasmid-specific gene expression responses to environmental stimuli and differential plasmid regulation mechanisms between L. monocytogenes strains. Our findings further our understanding of the dynamic expression of L. monocytogenes plasmid-encoded genes in diverse environmental conditions and highlight the need to expand the study of L. monocytogenes plasmid genes' functions.
Topics: Listeria monocytogenes; Transcriptome; Plasmids; Cadmium
PubMed: 36314750
DOI: 10.1002/mbo3.1315 -
Microbial Genomics Oct 2020Bacterial plasmids play a large role in allowing bacteria to adapt to changing environments and can pose a significant risk to human health if they confer virulence and...
Bacterial plasmids play a large role in allowing bacteria to adapt to changing environments and can pose a significant risk to human health if they confer virulence and antimicrobial resistance (AMR). Plasmids differ significantly in the taxonomic breadth of host bacteria in which they can successfully replicate, this is commonly referred to as 'host range' and is usually described in qualitative terms of 'narrow' or 'broad'. Understanding the host range potential of plasmids is of great interest due to their ability to disseminate traits such as AMR through bacterial populations and into human pathogens. We developed the MOB-suite to facilitate characterization of plasmids and introduced a whole-sequence-based classification system based on clustering complete plasmid sequences using Mash distances (https://github.com/phac-nml/mob-suite). We updated the MOB-suite database from 12 091 to 23 671 complete sequences, representing 17 779 unique plasmids. With advances in new algorithms for rapidly calculating average nucleotide identity (ANI), we compared clustering characteristics using two different distance measures - Mash and ANI - and three clustering algorithms on the unique set of plasmids. The plasmid nomenclature is designed to group highly similar plasmids together that are unlikely to have multiple representatives within a single cell. Based on our results, we determined that clusters generated using Mash and complete-linkage clustering at a Mash distance of 0.06 resulted in highly homogeneous clusters while maintaining cluster size. The taxonomic distribution of plasmid biomarker sequences for replication and relaxase typing, in combination with MOB-suite whole-sequence-based clusters have been examined in detail for all high-quality publicly available plasmid sequences. We have incorporated prediction of plasmid replication host range into the MOB-suite based on observed distributions of these sequence features in combination with known plasmid hosts from the literature. Host range is reported as the highest taxonomic rank that covers all of the plasmids which share replicon or relaxase biomarkers or belong to the same MOB-suite cluster code. Reporting host range based on these criteria allows for comparisons of host range between studies and provides information for plasmid surveillance.
Topics: Bacteria; Conjugation, Genetic; Databases, Genetic; Host Specificity; Humans; Molecular Typing; Plasmids
PubMed: 32969786
DOI: 10.1099/mgen.0.000435 -
Microbiology Spectrum Oct 2014Toxin-antitoxin (TA) systems are small genetic modules formed by a stable toxin and an unstable antitoxin that are widely present in plasmids and in chromosomes of... (Review)
Review
Toxin-antitoxin (TA) systems are small genetic modules formed by a stable toxin and an unstable antitoxin that are widely present in plasmids and in chromosomes of Bacteria and Archaea. Toxins can interfere with cell growth or viability, targeting a variety of key processes. Antitoxin inhibits expression of the toxin, interacts with it, and neutralizes its effect. In a plasmid context, toxins are kept silent by the continuous synthesis of the unstable antitoxins; in plasmid-free cells (segregants), toxins can be activated owing to the faster decay of the antitoxin, and this results in the elimination of these cells from the population (postsegregational killing [PSK]) and in an increase of plasmid-containing cells in a growing culture. Chromosomal TA systems can also be activated in particular circumstances, and the interference with cell growth and viability that ensues contributes in different ways to the physiology of the cell. In this article, we review the conditional activation of TAs in selected plasmidic and chromosomal TA pairs and the implications of this activation. On the whole, the analysis underscores TA interactions involved in PSK and points to the effective contribution of TA systems to the physiology of the cell.
Topics: Archaea; Bacteria; Biological Transport; Cell Division; Extrachromosomal Inheritance; Microbial Viability; Plasmids; Toxins, Biological
PubMed: 26104348
DOI: 10.1128/microbiolspec.PLAS-0009-2013 -
Complete sequence verification of plasmid DNA using the Oxford Nanopore Technologies' MinION device.BMC Bioinformatics Mar 2023Sequence verification is essential for plasmids used as critical reagents or therapeutic products. Typically, high-quality plasmid sequence is achieved through...
BACKGROUND
Sequence verification is essential for plasmids used as critical reagents or therapeutic products. Typically, high-quality plasmid sequence is achieved through capillary-based Sanger sequencing, requiring customized sets of primers for each plasmid. This process can become expensive, particularly for applications where the validated sequence needs to be produced within a regulated and quality-controlled environment for downstream clinical research applications.
RESULTS
Here, we describe a cost-effective and accurate plasmid sequencing and consensus generation procedure using the Oxford Nanopore Technologies' MinION device as an alternative to capillary-based plasmid sequencing options. This procedure can verify the identity of a pure population of plasmid, either confirming it matches the known and expected sequence, or identifying mutations present in the plasmid if any exist. We use a full MinION flow cell per plasmid, maximizing available data and allowing for stringent quality filters. Pseudopairing reads for consensus base calling reduces read error rates from 5.3 to 0.53%, and our pileup consensus approach provides per-base counts and confidence scores, allowing for interpretation of the certainty of the resulting consensus sequences. For pure plasmid samples, we demonstrate 100% accuracy in the resulting consensus sequence, and the sensitivity to detect small mutations such as insertions, deletions, and single nucleotide variants. In test cases where the sequenced pool of plasmids contains subclonal templates, detection sensitivity is similar to that of traditional capillary sequencing.
CONCLUSIONS
Our pipeline can provide significant cost savings compared to outsourcing clinical-grade sequencing of plasmids, making generation of high-quality plasmid sequence for clinical sequence verification more accessible. While other long-read-based methods offer higher-throughput and less cost, our pipeline produces complete and accurate sequence verification for cases where absolute sequence accuracy is required.
Topics: Sequence Analysis, DNA; Nanopores; Plasmids; DNA; High-Throughput Nucleotide Sequencing
PubMed: 36964503
DOI: 10.1186/s12859-023-05226-y -
Plasmid 2022Two phylogenetically distantly-related IncF plasmids, F and pED208, serve as important models for mechanistic and structural studies of F-like type IV secretion systems...
Two phylogenetically distantly-related IncF plasmids, F and pED208, serve as important models for mechanistic and structural studies of F-like type IV secretion systems (T4SSs) and F pili. Here, we present the pED208 sequence and compare it to F and pUMNF18, the closest match to pED208 in the NCBI database. As expected, gene content of the three cargo regions varies extensively, although the maintenance/leading regions (MLRs) and transfer (Tra) regions also carry novel genes or motifs with predicted modulatory effects on plasmid stability, dissemination and host range. By use of a Cre recombinase assay for translocation (CRAfT), we recently reported that pED208-carrying donors translocate several products of the MLR (ParA, ParB1, ParB2, SSB, PsiB, PsiA) intercellularly through the T4SS. Here, we extend these findings by reporting that pED208-carrying donors translocate 10 additional MLR proteins during conjugation. In contrast, two F plasmid-encoded toxin components of toxin-antitoxin (TA) modules, CcdB and SrnB, were not translocated at detectable levels through the T4SS. Remarkably, most or all of the pED208-encoded MLR proteins and CcdB and SrnB were translocated through heterologous T4SSs encoded by IncN and IncP plasmids pKM101 and RP4, respectively. Together, our sequence analyses underscore the genomic diversity of the F plasmid superfamily, and our experimental data demonstrate the promiscuous nature of conjugation machines for protein translocation. Our findings raise intriguing questions about the nature of T4SS translocation signals and of the biological and evolutionary consequences of conjugative protein transfer.
Topics: Type IV Secretion Systems; Plasmids; Escherichia coli; F Factor; Sequence Analysis; Conjugation, Genetic; Bacterial Proteins
PubMed: 36228885
DOI: 10.1016/j.plasmid.2022.102652