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Explore bioRxiv genomics preprints on sequencing, gene expression and genetic variation. Read source abstracts and verify methods in the manuscript; preprints are not peer reviewed.

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bioRxiv2017

Genomes from uncultivated prokaryotes: a comparison of metagenome-assembled and single-amplified genomes

BackgroundProkaryotes dominate the biosphere and regulate biogeochemical processes essential to all life. Yet, our knowledge about their biology is for the most part limited to the minority that has been successfully cultured. Molecular techniques now allow for obtaining genome sequences of uncultivated prokaryotic taxa, facilitating in-depth analyses that may ultimately improve our understanding of these key organisms.\n\nResultsWe compared results from two culture-independent strategies for recovering bacterial genomes: single-amplified genomes and metagenome-assembled genomes. Single-amplified genomes were obtained from samples collected at an offshore station in the Baltic Sea Proper and compared to previously obtained metagenome-assembled genomes from a time series at the same station. Among 16 single-amplified genomes analyzed, seven were found to match metagenome-assembled genomes, affiliated with a diverse set of taxa. Notably, genome pairs between the two approaches were nearly identical (>98.7% identity) across overlapping regions (30-80% of each genome). Within matching pairs, the single-amplified genomes were consistently smaller and less complete, whereas the genetic functional profiles were maintained. For the metagenome-assembled genomes, only on average 3.6% of the bases were estimated to be missing from the genomes due to wrongly binned contigs; the metagenome assembly was found to cause incompleteness to a higher degree than the binning procedure.\n\nConclusionsThe strong agreement between the single-amplified and metagenome-assembled genomes emphasizes that both methods generate accurate genome information from uncultivated bacteria. Importantly, this implies that the research questions and the available resources are allowed to determine the selection of genomics approach for microbiome studies.

bioinformatics↗
bioRxiv2021

Nuclear-cytoplasmic balance: whole genome duplications induce elevated organellar genome copy number

The plant genome is partitioned across three distinct subcellular compartments: the nucleus, mitochondria, and plastids. Successful coordination of gene expression among these organellar genomes and the nuclear genome is critical for plant function and fitness. Whole genome duplication events (WGDs) in the nucleus have played a major role in the diversification of land plants and are expected to perturb the relative copy number (stoichiometry) of nuclear, mitochondrial, and plastid genomes. Thus, elucidating the mechanisms whereby plant cells respond to the cytonuclear stoichiometric imbalance that follow WGDs represents an important yet underexplored question in understanding the evolutionary consequences of genome doubling. We used droplet digital PCR (ddPCR) to investigate the relationship between nuclear and organellar genome copy numbers in allopolyploids and their diploid progenitors in both wheat and Arabidopsis. Polyploids exhibit elevated organellar genome copy numbers per cell, largely preserving the cytonuclear stoichiometry observed in diploids despite the change in nuclear genome copy number. To investigate the timescale over which cytonuclear stoichiometry may respond to WGD, we also estimated organellar genome copy number in Arabidopsis synthetic autopolyploids and in a haploid-induced diploid line. We observed corresponding changes in organellar genome copy number in these laboratory-generated lines, indicating that at least some of the cellular response to cytonuclear stoichiometric imbalance is immediate following WGD. We conclude that increases in organellar genome copy numbers represent a common response to polyploidization, suggesting that maintenance of cytonuclear stoichiometry is an important component in establishing polyploid lineages. Significance StatementWhole genome duplications (WGD) have the potential to alter the stoichiometric balance between nuclear and organellar genomes. We used two separate diploid-polyploid complexes to show that plant cells with WGD exhibit elevated mitochondrial and plastid genome copy numbers, both immediately in lab-generated lines and in natural polyploids.

evolutionary biology↗
bioRxiv2021

mOTUpan: a robust Bayesian approach to leverage metagenome assembled genomes for core-genome estimation

Recent advances in sequencing and bioinformatics have expanded the tree of life by providing genomes for uncultured environmentally relevant clades, either through metagenome assembled genomes (MAGs) or single-cell assembled genomes (SAGs). While this expanded diversity can provide novel insights about microbial population structure, most tools available for core-genome estimation are overly sensitive to genome completeness. Consequently a major portion of the huge phylogenetic diversity uncovered by environmental genomics approaches remain excluded from such analyses. We present mOTUpan, a novel iterative Bayesian method for computing the core-genome for sets of genomes of highly diverse completeness range. The likelihood for each gene-cluster to belong to core- or accessory-genome is estimated by computing the probability of its presence/absence pattern in the target set of genomes. The core-genome prediction is computationally efficient and can be scaled up to thousands of genomes. It has shown comparable estimates as state-of-the-art tools Roary and PPanGGoLiN for high-quality genomes and outperforms them at lower completeness thresholds. mOTUpan wraps a bootstrapping procedure to estimate the quality of a specific core-genome prediction, as the accuracy of each run will depend on the specific completeness distribution and the number of genomes in the dataset under scrutiny. mOTUpan is implemented in the mOTUlizer software package, and available at github.com/moritzbuck/mOTUlizer, under GPL 3.0 license.

bioinformatics↗
bioRxiv2022

Population genetic analysis reveals the role of natural selection and phylogeography on genome-wide diversity in an extremely compact and reduced microsporidian genome

The determinants of variation in a species genome-wide nucleotide diversity include historical, environmental, and stochastic aspects. This diversity can inform us about the species past and present evolutionary dynamics. In parasites, the mode of transmission and the interactions with the host might supersede the effects of these aspects in shaping parasite genomic diversity. We used genomic samples from ten populations of the microsporidian parasite Ordospora colligata to investigate present genomic diversity and how it was shaped by evolutionary processes, specifically, the role of phylogeography, co-phylogeography (with the host), natural selection, and transmission mode. Although very closely related microsporidia cause diseases in humans, O. colligata is specific to the freshwater crustacean Daphnia magna and has one of the smallest known eukaryotic genomes. We found an overlapping phylogeography between O. colligata and its host highlighting the long-term, intimate relationship between them. The observed geographic distribution reflects previous findings that O. colligata exhibits adaptations to colder habitats, which differentiates it from other microsporidian gut parasites of D. magna predominantly found in warmer areas. The co-phylogeography allowed us to calibrate the O. colligata phylogeny and thus estimate its mutation rate. We found patterns of more efficient purifying selection in O. colligata relative to other microsporidia sharing the same host, which likely allowed this parasite to maintain its very compact genome. We also identified regions under potential selection related to coevolution including the ribosomal protein L24, a leucyl-tRNA synthetase, and a putative ABC-like lipid transport protein. Our whole-genome study provides insights into the evolution of one of the most reduced eukaryotic genomes and shows how different processes shape genomic diversity of an obligate parasite. Author summaryMicrosporidia are intracellular parasites that infect vertebrates, invertebrates, and even unicellular organisms. Due to their high variation in many aspects of life history and genomics, microsporidia have become a model clade for understanding evolutionary processes related to intracellular parasitism. However, the evolution of extreme genomic architectures in microsporidia and the coevolution with their hosts is still under-surveyed, especially given their role in human disease. Here, we study past and present evolutionary dynamics in a microsporidian species with one of the smallest known eukaryotic genomes, O. colligata. Close relatives of O. colligata cause death and disease in humans and agriculturally important animals. We show that purifying selection helped maintaining its reduced, compact genome and corroborate hypotheses about the evolution of different genome sizes in microsporidia. Importantly, we utilize the highly resolved phylogeny of its host to estimate the parasites mutation rate. This methodology allowed us to establish the first mutation rate estimate for a microsporidium, an estimate which is within the range of mutation rates estimated for phylogenetically related, non-parasitic fungi. Our study exemplifies how the combined knowledge about a species biology, ecology, and genomic diversity helps to resolve its evolutionary dynamics, in particular when phylogenomic information can be brought to bear for both host and parasite.

evolutionary biology↗
bioRxiv2023

Origin Matters: Using a Local Reference Genome Improves Measures in Population Genomics

Genome-level sequencing enables us to ask fundamental questions about the genetic basis of adaptation, population structure, and epigenetic mechanisms, but usually requires a suitable reference genome for mapping population-level re-sequencing data. In some model systems, multiple reference genomes are available, giving researchers the challenging task of determining which reference genome best suits their data. Here we compare the use of two different reference genomes for the three-spined stickleback (Gasterosteus aculeatus), one novel genome derived from a European gynogenetic individual and the published reference genome of a North American individual. Specifically, we investigate the impact of using a local reference versus one generated from a distinct lineage on several common population genomics analyses. Through mapping genome resequencing data of 60 sticklebacks from across Europe and North America, we demonstrate that genetic distance among samples and the reference impacts downstream analyses. Using a local reference genome increased mapping efficiency and genotyping accuracy, effectively retaining more and better data. Despite comparable distributions of the metrics generated across the genome using SNP data (i.e., {pi}, Tajimas D, and FST), window-based statistics using different references resulted in different outlier genes and enriched gene functions. A marker-based analysis of DNA methylation distributions had a comparably high overlap in outlier genes and functions, yet with distinct differences depending on the reference genome. Overall, our results highlight how using a local reference genome decreases reference bias to increase confidence in downstream analyses of the data. Such results have significant implications in all reference-genome-based population genomic analyses.

bioinformatics↗
bioRxiv2023

Unveiling genome plasticity and a novel phage in Mycoplasma felis: Genomic investigations of four feline isolates

Mycoplasma felis has been isolated from diseased cats and horses, but to date only a single fully assembled genome of this species, of an isolate from a horse, has been characterised. This study aimed to characterise and compare the completely assembled genomes of four clinical isolates of M. felis from three domestic cats, assembled with the aid of short and long read sequencing methods. The completed genomes encoded a median of 759 open reading frames (min, 743, max 777) and had a median average nucleotide identity (ANI) of 98.2% with the genome of the available equid origin reference strain. Comparative genomic analysis revealed the occurrence of multiple horizontal gene transfer (HGT) events and significant genome reassortment. This had resulted in the acquisition or loss of numerous genes within the Australian felid isolate genomes, encoding putative proteins involved in DNA transfer, metabolism, DNA replication, host cell interaction, and restriction modification systems. Additionally, a novel mycoplasma phage was detected in one Australian felid M. felis isolate by genomic analysis and visualised using cryo-transmission electron microscopy. This study has highlighted the complex genomic dynamics in different host environments. Furthermore, the sequences obtained in this work will enable the development of new diagnostic tools, and identification of future infection control and treatment options for the respiratory disease complex in cats. Data summaryAll genome data for this study have been deposited in GenBank under BioProject PRJNA906261. Genome assemblies, as well as Illumina and Oxford Nanopore sequence reads for each isolate, can be found under their respective BioSamples: SAMN32182834 (isolate 047), SAMN32182835 (isolate 219), SAMN32182836 (isolate 329 and associated phage), and SAMN32182837 (isolate 632). The authors confirm all supporting data and protocols have been provided within the article. Impact statementMycoplasma felis is commonly associated with clinical cases of conjunctivitis and feline respiratory disease complex in cats, the leading cause of euthanasia in animal shelters. In the absence of vaccines, infection control is currently limited to the prolonged treatment with antimicrobials. Prior to this study there was only one complete genome assembly of an isolate of M. felis, which had been obtained from a horse. This study has provided the first high quality hybrid assembled genomes of M. felis isolates from cats. This work adds four new genomes from clinical cases, as well as the identification and validation of the presence of a novel phage that utilises the mycoplasma translation code. The genomic data presented here can assist future projects investigating improved diagnostics and development of new treatment options for this significant feline pathogen.

microbiology↗
bioRxiv2026

DipSkmer: Reference-free population genomics with diploid genome skims

Ecologists and conservation biologists rely on genetic diversity as a key essential biodiversity variable (EBV) used to track population health and dynamics, and utilize the population parameter{theta} (estimated by the average pairwise genomic distance) as a key metric of diversity. While whole-genome-sequencing (wgs) is increasingly affordable, it will be considerable time before the full diversity of life is represented by high-quality assembled genomes; even then, constant monitoring will still require repeated sampling of populations. In contrast, genome skimming (low-coverage, short-read wgs) is highly cost-effective but challenging to analyze because the coverage is too low for assembly and reliable error correction. Mature methods, such as Mash, exist for estimating pairwise genomic distances based on the Jaccard similarity of k-mer sets computed using sketching techniques. Some, such as Skmer, additionally model the impacts of low coverage. These methods have been successfully applied to assembly-free species identification and phylogenetics; however, their use in population genetics has been limited. This is because these methods implicitly treat genomes as haploid and heterozygosity confounds true estimates of genomic distance for diploid organisms. In this paper, we address this problem through a number of technical advances. First, we use coalescent theory to mathematically derive how the Jaccard index between two diploid samples changes with the scaled population size parameter ({theta}). Next, we derive an estimator that computes{theta} from the Jaccard index, in addition to several auxiliary variables, which we also estimate from the genome skims. The resulting method, DipSkmer, enables more accurate estimates of coverage, sequencing error, and pairwise nucleotide distance for diploid samples. Analyses of both simulated and empirical datasets show that for diploids and low distances (e.g., < 2%), Dip-Skmer produces the most accurate pairwise distance estimates, outperforming existing alignment-free methods such as Mash and Skmer, and closely approximates ANGSD, a reference and alignment-based tool. AvailabilityThe code for DipSkmer is available at https://github.com/echarvel3/ReSkmer/tree/DipSkmer-REFACTOR. Simulation scripts and environments are available at https://github.com/echarvel3/dipskmer_scripts. Author SummaryThe process of obtaining full-genome population genomic measurements for biodiversity monitoring remains expensive due to the need for high-coverage sequencing and reference assemblies. Genome skimming has been shown to be a viable, low-coverage alternative for obtaining genomic distances, and alignment- and assembly-free methods exist for analyzing nuclear data from skimming data to estimate the distance between samples. However, existing methods fail to model within-sample heterozygosity, expected for diploid organisms. Given the dominance of diploidy among species of interest to ecologists, the implications of these simplifying assumptions warrant further study. Here, we present a mathematical model of the k-mer sets sampled from two diploid genomes from a Wright-Fisher population. We use the model to develop DipSkmer, a k-mer-based, reference-free method for estimating nucleotide diversity and population divergence that, unlike its predecessors, models within-sample heterozygosity. Benchmarking shows more accurate genomic diversity estimates compared to existing reference-free, genome-skimming methods and comparable performance to the popular high-coverage, reference-based method, ANGSD. Thus, DipSkmer enables accessible, less expensive population monitoring through genetic diversity estimates.

bioinformatics↗
bioRxiv2020

Biodiversity genomics of small metazoans: high quality de novo genomes from single specimens of field-collected and ethanol-preserved springtails

Genome sequencing of all known eukaryotes on Earth promises unprecedented advances in evolutionary sciences, ecology, systematics and in biodiversity-related applied fields such as environmental management and natural product research. Advances in DNA sequencing technologies make genome sequencing feasible for many non-genetic model species. However, genome sequencing today relies on large quantities of high quality, high molecular weight (HMW) DNA which is mostly obtained from fresh tissues. This is problematic for biodiversity genomics of Metazoa as most species are small and yield minute amounts of DNA. Furthermore, briging living specimens to the lab bench not realistic for the majority of species. Here we overcome those difficulties by sequencing two species of springtails (Collembola) from single specimens preserved in ethanol. We used a newly developed, genome-wide amplification-based protocol to generate PacBio libraries for HiFi long-read sequencing. The assembled genomes were highly continuous. They can be considered complete as we recovered over 95% of BUSCOs. Genome-wide amplification does not seem to bias genome recovery. Presence of almost complete copies of the mitochondrial genome in the nuclear genome were pitfalls for automatic assemblers. The genomes fit well into an existing phylogeny of springtails. A neotype is designated for one of the species, blending genome sequencing and creation of taxonomic references. Our study shows that it is possible to obtain high quality genomes from small, field-preserved sub-millimeter metazoans, thus making their vast diversity accessible to the fields of genomics.

genomics↗
bioRxiv2023

The revised leopard gecko (Eublepharis macularius) reference genome provides insight into the considerations of genome phasing and assembly

Genomic resources across squamate reptiles (lizards and snakes) have lagged behind other vertebrate systems and high-quality reference genomes remain scarce. Of the 23 chromosome-scale reference genomes across the order, only 12 of the ~60 squamate families are represented. Within geckos (infraorder Gekkota), a species-rich clade of lizards, chromosome-level genomes are exceptionally sparse representing only two of the seven extant families. Using the latest advances in genome sequencing and assembly methods, we generated one of the highest quality squamate genomes to date for the leopard gecko, Eublepharis macularius (Eublepharidae). We compared this assembly to the previous, short-read only, E. macularius reference genome published in 2016 and examined potential factors within the assembly influencing contiguity of genome assemblies using PacBio HiFi data. Briefly, the read N50 of the PacBio HiFi reads generated for this study was equal to the contig N50 of the previous E. macularius reference genome at 20.4 kilobases. The HiFi reads were assembled into a total of 132 contigs, which was further scaffolded using HiC data into 75 total sequences representing all 19 chromosomes. We identified that 9 of the 19 chromosomes were assembled as single contigs, while the other 10 chromosomes were each scaffolded together from two or more contigs. We qualitatively identified that percent repeat content within a chromosome broadly affects its assembly contiguity prior to scaffolding. This genome assembly signifies a new age for squamate genomics where high-quality reference genomes rivaling some of the best vertebrate genome assemblies can be generated for a fraction previous cost estimates. This new E. macularius reference assembly is available on NCBI at JAOPLA010000000. The genome version and its associated annotations are also available via this Figshare repository https://doi.org/10.6084/m9.figshare.20069273.

genomics↗
bioRxiv2023

Remarkably high repeat content in the genomes of sparrows: the importance of genome assembly completeness for transposable element discovery.

Transposable elements (TE) play critical roles in shaping genome evolution. However, the highly repetitive sequence content of TEs is a major source of assembly gaps. This makes it difficult to decipher the impact of these elements on the dynamics of genome evolution. The increased capacity of long-read sequencing technologies to span highly repetitive regions of the genome should provide novel insights into patterns of TE diversity. Here we report the generation of highly contiguous reference genomes using PacBio long read and Omni-C technologies for three species of sparrows in the family Passerellidae. To assess the influence of sequencing technology on TE annotation, we compared these assemblies to three chromosome-level sparrow assemblies recently generated by the Vertebrate Genomes Project and nine other sparrow species generated using a variety of short- and long-read technologies. All long-read based assemblies were longer in length (range: 1.12-1.41 Gb) than short-read assemblies (0.91-1.08 Gb). Assembly length was strongly correlated with the amount of repeat content, with longer genomes showing much higher levels of repeat content than typically reported for the avian order Passeriformes. Repeat content for the Bells sparrow (31.2% of genome) was the highest level reported to date for a songbird genome assembly and was more in line with woodpecker (order Piciformes) genomes. CR1 LINE elements retained from an expansion that occurred 25-30 million years ago were the most abundant TEs in the song sparrow genome. Although the other five sparrow species also exhibit evidence for a spike in CR1 LINE activity at 25-30 million years ago, LTR elements stemming from more recent expansions were the most abundant elements in these species. LTRs were uniquely abundant in the Bells sparrow genome deriving from two recent peaks of activity. Higher levels of repeat content (79.2-93.7%) were found on the W chromosome relative to the Z (20.7-26.5) or autosomes (16.1-30.9%). These patterns support a dynamic model of transposable element expansion and contraction underpinning the seemingly constrained and small sized genomes of birds. Our work highlights how the resolution of difficult-to-assemble regions of the genome with new sequencing technologies promises to transform our understanding of avian genome evolution.

genomics↗
bioRxiv2019

Quality control of large genome datasets using genome fingerprints

The 1000 Genomes Project (TGP) is a foundational resource which serves the biomedical community as a standard reference cohort for human genetic variation. There are now seven public versions of these genomes. The TGP Consortium produced the first by mapping its final data release against human reference sequence GRCh37, then "lifted over these genomes to the improved reference sequence (GRCh38) when it was released, and remapped the original data to GRCh38 with two similar pipelines. As best practice quality validation, the pipelines that generated these versions were benchmarked against the Genome In A Bottle Consortiums platinum quality genome (NA12878). The New York Genome Center recently released the results of independently resequencing the cohort at greater depth (30X), a phased version informed by the inclusion of related individuals, and independently remapped the original variant calls to GRCh38. We evaluated all seven versions using genome fingerprinting, which supports ultrafast genome comparison even across reference versions. We noted multiple issues including discrepancies in cohort membership, disagreement on the overall level of variation, evidence of substandard pipeline performance on specific genomes and in specific regions of the genome, cryptic relationships between individuals, inconsistent phasing, and annotation distortions caused by the history of the reference genome itself. We therefore recommend global quality assessment by rapid genome comparisons, using genome fingerprints and other metrics, alongside benchmarking as part of best practice quality assessment of large genome datasets. Our observations also help inform the decision of which version to use, to support analyses by individual researchers.

genomics↗
bioRxiv2026

The Vertebrate Genomes Project Phase I: A global reference genome resource

The Vertebrate Genomes Project (VGP) aims to produce complete and near-error-free reference genomes for all [~]70,000 extant vertebrate species1. Organized in four phases, it progressively targets all vertebrate orders, families, genera, and eventually all species. Here we present the completion of VGP Phase I, delivering reference genomes for [~]95% of vertebrate orders, along with additional lineages within those orders, totaling 816 species and 1.6 trillion base pairs of main haplotype sequence. These genomes were assembled and annotated over an 8-year period (2018-2026) of rapid advances in genome sequencing, assembly, and annotation methods2-4, alongside the growth of associated consortium initiatives and international collaborations5-9. They represent some of the highest-quality vertebrate genomes currently available, and most have become the primary reference for their respective species in public databases. Comparative analyses across a subset of 579 species when we reached a threshold of 85% of orders allowed us to reconstruct the genome of the last common ancestor of all vertebrates 500 million years ago, identify diverse modes of sex chromosome evolution, reveal clade-specific three-dimensional genome architecture, discover methylated epigenetic landscapes across vertebrates, and provide a framework for studying gene and pseudogene evolution, immune loci, cancer-associated genes, and other trait-associated loci. Approximately a quarter of this subset are listed as Vulnerable to Critically Endangered by the IUCN Red List of Threatened Species, and have enabled more advanced genomic investigations of extinction risk. VGP Phase I delivers a reference backbone for vertebrate genomics, enabling discoveries that would otherwise remain out of reach across evolution, conservation, and medicine. Talking pointsO_LIThe flagship paper of VGP Phase I C_LIO_LIThe highest quality collection of genomes within the eukaryotic domain of life C_LIO_LIEvolution of genome sequencing technology quality throughout VGP Phase I C_LIO_LIA driver project that has been a model for multiple large-scale, high-quality reference genome projects C_LIO_LIReleases all currently unpublished genomes in Phase I from scientific study embargoes C_LIO_LIMultiple biological discoveries across the vertebrate tree of life C_LI

genomics↗
bioRxiv2023

Pan-genome analysis of six complete Paracoccus type strain genomes from hybrid next generation sequencing

The genus Paracoccus capable of inhabiting a variety of different ecological niches both, marine and terrestrial is globally distributed. In addition, Paracoccus is taxonomically, metabolically and regarding lifestyle highly diverse. Until now, little is known on how Paracoccus can adapt to such a range of different ecological niches and lifestyles. In the present study, the genus Paracoccus was phylogenomically analyzed (n = 160) and revisited, allowing species level classification of 16 so far unclassified Paracoccus sp. strains and detection of five misclassifications. Moreover, we performed pan-genome analysis of Paracoccus-type strains, isolated from a variety of ecological niches, including different soils, tidal flat sediment, host association such as the bluespotted cornetfish, Bugula plumosa, and the reef-building coral Stylophora pistillata to elucidate either i) the importance of lifestyle and adaptation potential, and ii) the role of the genomic equipment and niche adaptation potential. Six complete genomes were de novo hybrid assembled using a combination of short and long-read technologies. These Paracoccus genomes increase the number of completely closed high-quality genomes of type strains from 15 to 21. Pan-genome analysis revealed an open pan-genome composed of 13,819 genes with a minimal chromosomal core (8.84 %) highlighting the genomic adaptation potential and the huge impact of extra-chromosomal elements. All genomes are shaped by the acquisition of various mobile genetic elements including genomic islands, prophages, transposases, and insertion sequences emphasizing their genomic plasticity. In terms of lifestyle, each mobile genetic elements should be evaluated separately with respect to the ecological context. Free- living genomes, in contrast to host-associated, tend to comprise (1) larger genomes, or the highest number of extra-chromosomal elements, (2) higher number of genomic islands and insertion sequence elements, and (3) a lower number of intact prophage regions. Due to the vast number of adaptive genes, Paracoccus can quickly adapt to changing environmental conditions.

microbiology↗
bioRxiv2024

Intraspecific genome size variation in Rorippa indica reveals a tropical adaptation by genomic enlargement

Genome size exhibits substantial variation across organisms, but the underlying causes and ecological consequences remain unclear. While interspecific comparisons have suggested selective pressures against large genomes, intraspecific variation has been less explored. Here, we investigate genome size variation within the hexaploid yellowcress herb, Rorippa indica, by integrating flow cytometry, plastomic phylogeography, genomic repeat profiling, and reciprocal common garden experiments. We analyzed 192 accessions from 83 natural populations, revealing a 128 Mb range in genome size. Plastome haplotype analysis identified a haplogroup that shifted niches to tropics and evolved larger genomes. A strong correlation was found between larger genomes and tropical habitats characterized by higher temperatures and lower seasonality. Genomic repeat content, particularly 45S rDNA and Ty1-copia transposable elements, was associated with larger genomes. Reciprocal transplantation experiments confirmed the adaptive nature of large genomes in tropical environments, with individuals exhibiting lower growth rates but higher fecundity. Our findings support the "large genome selection" hypothesis, suggesting that genome size enlargement, driven by genomic repetitive elements, can be an adaptive response to high temperatures of tropics. As global warming continues, plants with larger genomes may exhibit slower growth but increased reproductive output, potentially impacting ecosystem dynamics and agricultural productivity.

evolutionary biology↗
bioRxiv2018

Genome assembly of the A-group Wolbachia in Nasonia oneida and phylogenomic analysis of Wolbachia strains revealed genome evolution and lateral gene transfer

Wolbachia are obligate intracellular bacteria which commonly infect various nematode and arthropod species. Based on depth differences, we assembled the genome of Wolbachia in the parasitoid jewel wasp species Nasonia oneida (wOne), using 10X Genomics Chromium linked-read technology. The final draft assembly consists of 1,293,406 bp in 47 scaffolds with 1,114 coding genes and 97.01% genome completeness assessed by checkM. wOne is the A1 strain previously reported in N. oneida, and pyrosequencing confirms that the wasp strain lacks A2 and B types, which were likely lost during laboratory culturing. Polymorphisms identified in the wOneA1 genome have elevated read depths, indicating recent gene duplications rather that strain variation. These polymorphisms are enriched in nonsynonymous changes in 27 coding genes, including phase baseplate assembly proteins and transporter activity related genes. wOneA1 is more closely grouped with A-Wolbachia in the Drosophila simulans (wHa) than A-Wolbachia found in wasps. Genome variation was next evaluated in 34 published Wolbachia genomes for 211 single ortholog genes, and revealed six supergroup discordant trees, indicating recombination events not only between A and B supergroups, but also between A and E supergroups. Comparisons of strain divergence using the five genes of the Multi Locus Strain Typing (MLST) system show a high correlation (rho=0.98) between MLST and whole genome divergences, indicating that MLST is a reliable method for identifying related strains when whole genome data are not available. Assembling bacterial genomes from host genome projects can provide an effective method for sequencing Wolbachia genomes and characterizing their diversity.\n\nAuthor SummaryMore than half of the arthropod species are infected by the obligated intracellular bacteria Wolbachia. As one of the most widespread parasitic microbes, Wolbachia mediate important biological processes such as cytoplasmic incompatibility and lateral gene transfer in insects. Their evolutionary relationship has been characterize using five protein-coding and 16S rRNA genes. In this work, we identified 211 conserved single copies genes in 34 genome sequenced Wolbachia strains, and we discovered that they maintain the supergroup relationship classified previously based on selected genes. We constructed phylogenetic trees for individual genes and found only six genes display discordant tree structure between supergroups, due to lateral gene transfer and homologous recombination events. But these events are not common (3%) in Wolbachia genomes, at least in these conserved single copy genes. In addition to known lateral gene transfer events between A and B supergroups, we identified transfers between A and E supergroups for the first time. Selective maintenance of such transfers suggests possible roles in Wolbachia infection related functions. We also found enriched nonsynonymous polymorphisms in Nasonia oneida Wobachia genome, and their differences are more likely to result from gene duplications within the strain, rather than strain variation within the parasitoid.

evolutionary biology↗
bioRxiv2019

Within-population genome size variation is mediated by multiple genomic elements that segregate independently during meiosis

Within-species variation in genome size has been documented in many animals and plants. Despite its importance for understanding eukaryotic genome diversity, there is only sparse knowledge about how individual-level processes mediate genome size variation in populations. Here we study a natural population of the rotifer Brachionus asplanchnoidis whose members differ up to 1.9-fold in genome size, but were still able to interbreed and produce viable offspring. We show that genome size is highly heritable and can be artificially selected up or down, but not beyond a minimum diploid genome size. Analyses of segregation patterns in haploid males reveal that large genomic elements (several megabases in size) provide the substrate of genome size variation. These elements, and their segregation patterns, explain the generation of new genome size variants, the short-term evolutionary potential of genome size change in populations, and some seemingly paradoxical patterns, like an increase in genome size variation among highly inbred lines. Our study suggests that a conceptual model involving only two variables, (1) a minimum genome size of the population, and (2) a vector containing information on additional elements that may increase genome size in this population (size, number, and meiotic segregation behaviour), can effectively address most scenarios of short-term evolutionary change of genome size in a population.

evolutionary biology↗
bioRxiv2019

G-OnRamp: Generating genome browsers to facilitate undergraduate-driven collaborative genome annotation

Scientists are sequencing new genomes at an increasing rate with the goal of associating genome contents with phenotypic traits. After a new genome is sequenced and assembled, structural gene annotation is often the first step in analysis. Despite advances in computational gene prediction algorithms, most eukaryotic genomes still benefit from manual gene annotation. Undergraduates can become skilled annotators, and in the process learn both about genes/genomes and about how to utilize large datasets. Data visualizations provided by a genome browser are essential for manual gene annotation, enabling annotators to quickly evaluate multiple lines of evidence (e.g., sequence similarity, RNA-Seq, gene predictions, repeats). However, creating genome browsers requires extensive computational skills; lack of the expertise required remains a major barrier for many biomedical researchers and educators.\n\nTo address these challenges, the Genomics Education Partnership (GEP; https://gep.wustl.edu/) has partnered with the Galaxy Project (https://galaxyproject.org) to develop G-OnRamp (http://g-onramp.org), a web-based platform for creating UCSC Assembly Hubs and JBrowse genome browsers. G-OnRamp can also convert a JBrowse instance into an Apollo instance for collaborative genome annotations in research and educational settings. G-OnRamp enables researchers to easily visualize their experimental results, educators to create Course-based Undergraduate Research Experiences (CUREs) centered on genome annotation, and students to participate in genomics research.\n\nDevelopment of G-OnRamp was guided by extensive user feedback from in-person workshops. Sixty-five researchers and educators from over 40 institutions participated in these workshops, which produced over 20 genome browsers now available for research and education. For example, genome browsers for four parasitoid wasp species were used in a CURE engaging 142 students taught by 13 faculty members -- producing a total of 192 gene models. G-OnRamp can be deployed on a personal computer or on cloud computing platforms, and the genome browsers produced can be transferred to the CyVerse Data Store for long-term access.

scientific communication and education↗
bioRxiv2026

Comparative genomics reveals extensive genomic conservation and limited microdiversification among Xenorhabdus bovienii isolates recovered from a single Steinernema feltiae isolation event.

Xenorhabdus bovienii is a symbiotic bacterium associated with entomopathogenic nematodes of the genus Steinernema. Comparative genomic analyses of closely related isolates provide an opportunity to investigate fine-scale diversification, genome plasticity, and the evolutionary processes shaping symbiotic bacterial populations. Here, we analyzed four X. bovienii isolates (XenUTI4.1-XenUTI4.4) recovered from a single Steinernema feltiae isolation event using comparative genomics approaches integrating average nucleotide identity (ANI), single-nucleotide polymorphism (SNP) analyses, pangenome reconstruction, biosynthetic gene cluster (BGC) prediction, and mobile element-associated annotation screening. Whole-genome comparisons revealed extremely high genomic similarity among isolates, with ANI values exceeding 99.84%. Read-based SNP analyses identified only 23-36 annotated variants relative to the XenUTI4.1 reference genome, indicating limited sequence divergence despite detectable microvariation. Functional annotation of these variants showed that most corresponded to missense or synonymous substitutions affecting a small number of coding sequences. Pangenome analysis identified 4,712 orthologous gene clusters, including a highly conserved core genome of 4,256 clusters (90.3%) shared by all isolates and a relatively small accessory genome comprising 456 clusters. antiSMASH analyses revealed broadly conserved secondary metabolite biosynthetic potential across the four genomes, whereas screening of genome annotations identified abundant phage-related, transposase-associated, and recombination-associated genes consistent with ongoing genome plasticity. Collectively, these results demonstrate that the analyzed X. bovienii isolates represent a highly conserved population exhibiting limited but detectable genomic microdiversification. The coexistence of a large core genome, a modest accessory gene complement, and numerous mobile element-associated functions suggests that localized sequence variation and mobile genetic elements contribute to genomic diversification within S. feltiae-associated X. bovienii populations.

genetics↗