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Search indexed bioRxiv preprints in genomics, neuroscience, cell biology and bioinformatics. Read source abstracts and check manuscript versions; preprints are not peer reviewed.

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bioRxiv2024

Genome sizes of animal RNA viruses reflect phylogenetic constraints

Animal genomes are characterized by extensive variation in size. RNA viruses similarly exhibit substantial genomic diversity, with genome lengths ranging from 1.7 to 64 Kb. Despite the myriad of novel viruses discovered by metagenomics, we know little of the factors that shape the evolution of the genome size in RNA viruses. We analyzed the variation in genome sizes across orders and families of animal RNA viruses. We found that RNA viruses can have highly variable genome sizes within and among orders, with the Nidovirales (including the Coronaviridae) having both significantly larger genomes and a greater range of genome sizes than other orders. In the Bunyavirales, Amarillovirales, Nidovirales and Picornavirales the genome sizes of invertebrate-associated RNA viruses were significantly larger than those that infect vertebrates, in contrast to their animal hosts in which vertebrates commonly have larger genomes than invertebrates. However, in the Mononegavirales, vertebrate viruses were significantly larger than those viruses associated with invertebrates. There were similarly complex associations between genome size and patterns of genome segmentation. In the Bunyavirales, Reovirales, and Nidovirales, viruses with segmented genomes or that possessed a large number of segments, had significantly larger genome sizes that viruses with non-segmented genomes or a small number of segments, while in the Articulavirales there were no significant differences in genome size among viruses possessing any number of genome segments. More broadly, our analysis revealed that taxonomic position (i.e., RNA virus order) had a greater impact on genome size than whether viruses infected vertebrates or invertebrates or their pattern of genome segmentation. Hence, the phylogenetic constraints on genome size are of sufficient magnitude to shape other aspects of virus evolution.

microbiology↗
bioRxiv2026

Genome expansions and regulatory contact entanglement help preserve ancestral metazoan synteny

Chromosomes constitute deeply conserved evolutionary units in many metazoan genomes, with chromosomal fusions and fissions, accompanied by sub-chromosomal rearrangements, rewiring three dimensional genome architecture. How chromatin loops and compartments that define distal regulatory interactions within chromosomes impose functional constraints that affect this long-term evolutionary process (and vice-versa) is an emerging research topic. Genome expansions, especially through transposable element (TE) activity, test these constraints by increasing the genomic distances over which regulatory interactions must function and were thus suggested to be the drivers of chromosomal rearrangements. To study dynamics and stability of such distal interactions in the light of genome expansions, we focus on the cnidarian Hydra vulgaris, which, based on its simple and well-understood biology as well as one of the largest genomes among cnidarians, is particularly suited to test how chromatin loop and genome architecture respond to genome expansion. We investigate genome architecture using Micro-C, single-cell Hi-C (Dip-C), and DNA FISH, and perform comparative analysis using available genomic and epigenomic data. Contrary to prior expectations, our analysis of whole-genome data and particular loci (e.g., Wnt) suggests a scenario where genome expansion did not only result in chromatin loops often reaching several megabases in hydra, but also led to regulatory contact mixing and entanglement, introducing additional constraints to maintain ancestral genomic architecture. Generalizing these findings across hundreds of metazoan genomes, we show a new mechanistic role for genome expansion in yielding entangled long-range regulatory configurations that, in turn, decelerate chromosomal rearrangements, thus maintaining (and not breaking) ancestral regulatory states and synteny. Significance statementSome of the largest and most repetitive animal genomes retain a surprisingly high level of deeply conserved metazoan synteny. As repetitive regions are often associated with chromosomal rearrangements, it has been enigmatic why syntenic retention is so frequently observed. In this study, using hydra as a model system for both stem cell biology and an evolutionary history rich in transposable-element driven genome expansion, a multi-level conformational landscape dissection reveals a multitude of long-range regulatory states. We show that mixing of multiple such regulatory links accompanied by genomic expansion is associated with maintained ancestral synteny, thus pointing to the counter-intuitive role of genome expansions as "fossilization" agents across metazoan genomes.

evolutionary biology↗
bioRxiv2023

Deep convolutional and conditional neural networks for large-scale genomic data generation

Applications of generative models for genomic data have gained significant momentum in the past few years, with scopes ranging from data characterization to generation of genomic segments and functional sequences. In our previous study, we demonstrated that generative adversarial networks (GANs) and restricted Boltzmann machines (RBMs) can be used to create novel high-quality artificial genomes (AGs) which can preserve the complex characteristics of real genomes such as population structure, linkage disequilibrium and selection signals. However, a major drawback of these models is scalability, since the large feature space of genome-wide data increases computational complexity vastly. To address this issue, we implemented a novel convolutional Wasserstein GAN (WGAN) model along with a novel conditional RBM (CRBM) framework for generating AGs with high SNP number. These networks implicitly learn the varying landscape of haplotypic structure in order to capture complex correlation patterns along the genome and generate a wide diversity of plausible haplotypes. We performed comparative analyses to assess both the quality of these generated haplotypes and the amount of possible privacy leakage from the training data. As the importance of genetic privacy becomes more prevalent, the need for effective privacy protection measures for genomic data increases. We used generative neural networks to create large artificial genome segments which possess many characteristics of real genomes without substantial privacy leakage from the training dataset. In the near future with further improvements in haplotype quality and privacy preservation, large-scale artificial genome databases can be assembled to provide easily accessible surrogates of real databases, allowing researchers to conduct studies with diverse genomic data within a safe ethical framework in terms of donor privacy. Author summaryGenerative modelling has recently become a prominent research field in genomics, with applications ranging from functional sequence design to characterization of population structure. We previously used generative neural networks to create artificial genome segments which possess many characteristics of real genomes but these segments were short in size due to computational requirements. In this work, we present novel generative models for generating artificial genomes with larger sequence size. We test the generated artificial genomes with multiple summary statistics to assess the haplotype quality, overfitting and privacy leakage from the training dataset. Our findings suggest that although there is still room for improvement both in terms of genome quality and privacy preservation, convolutional architectures and conditional generation can be utilised for generating good quality, large-scale genomic data. In the near future with additional improvements, large-scale artificial genomes can be used for assembling surrogate biobanks as alternatives to real biobanks with access restrictions, increasing data accessibility to researchers around the globe.

genomics↗
bioRxiv2019

Genome compartmentalization predates species divergence in the plant pathogen genus Zymoseptoria

BackgroundAntagonistic co-evolution can drive rapid adaptation in pathogens and shape genome architecture. Comparative genome analyses of several fungal pathogens revealed highly variable genomes, for many species characterized by specific repeat-rich genome compartments with exceptionally high sequence variability. Dynamic genome architecture may enable fast adaptation to host genetics. The wheat pathogen Zymoseptoria tritici with its highly variable genome and has emerged as a model organism to study the genomic evolution of plant pathogens. Here, we compared genomes of Z. tritici isolates and genomes of sister species infecting wild grasses to address the evolution of genome composition and structure. ResultsUsing long-read technology, we sequenced and assembled genomes of Z. ardabiliae, Z. brevis, Z. pseudotritici and Z. passerinii, together with two isolates of Z. tritici. We report a high extent of genome collinearity among Zymoseptoria species and high conservation of genomic, transcriptomic and epigenomic signatures of compartmentalization. We identify high gene content variability both within and between species. In addition, such variability is mainly limited to the accessory chromosomes and accessory compartments. Despite strong host specificity and non-overlapping host-range between species, effectors are mainly shared among Zymoseptoria species, yet exhibiting a high level of presence-absence polymorphism within Z. tritici. Using in planta transcriptomic data from Z. tritici, we suggest different roles for the shared orthologs and for the accessory genes during infection of their hosts. ConclusionDespite previous reports of high genomic plasticity in Z. tritici, we describe here a high level of conservation in genomic, epigenomic and transcriptomic signatures of genome architecture and compartmentalization across the genus Zymoseptoria. The compartmentalized genome may reflect purifying selection to retain a functional core genome and relaxed selection on the accessory genome allowing a higher extent of polymorphism.

genomics↗
bioRxiv2026

Phylogenomics and comparative genomics of the genus Erwinia reveal taxonomic inconsistencies and evolutionary diversification

The genus Erwinia comprises a diverse group of bacteria associated with plants, insects, and the environment, including several economically important phytopathogens. The genus has been revised taxonomically many times, yet a thorough and genome-wide assessment of its evolutionary relationships and genomic diversity has been lacking. In this research, we carried out an extensive phylogenomic and comparative genomic analyses of the genus Erwinia using 104 genomes including historically important strains. Genome-wide analyses integrating average nucleotide identity (ANI), digital DNA-DNA hybridization (dDDH), core-genome phylogenomics, pan-genome analysis, and comparative genomics resolved evolutionary relationships across the genus and identified multiple taxonomic inconsistencies. The pan-genome analysis revealed a relatively small core genome alongside an extensive accessory genome, underscoring the substantial genomic plasticity and ongoing diversification within the genus. The comparative analyses further showed pronounced lineage-specific variation in secretion systems, exopolysaccharide biosynthetic loci, flagellar gene clusters, genomic islands, prophages, and iron acquisition systems, suggesting that virulence-associated determinants have evolved through differential gene gain, loss, and conservation across distinct lineages, thereby facilitating host and ecological niche adaptation. This lineage-specific variation indicates that pathogenicity in the genus is not driven by a single conserved set of virulence determinants but instead reflects distinct combinations of virulence-associated genes. These findings refine the genomic framework of the genus Erwinia, provide evidence for taxonomic revision of several lineages, and improve our understanding of the evolutionary relationships, genomic diversification, and lineage-specific adaptations associated with host interactions and ecological specialization. Impact StatementThis study provides the first comprehensive genome-wide phylogenomic framework for the genus Erwinia, integrating taxonomy, pan-genome diversity, virulence-associated determinants, and mobile genetic elements across all 18 currently recognized species. Analyses resolve evolutionary relationships, uncover multiple taxonomic inconsistencies, identify previously unrecognized species-level lineages, including a putative novel Erwinia species PL328 isolated from Cornus florida (dogwood), and reveal lineage-specific genomic features. These findings establish a valuable genomic foundation for future studies of Erwinia evolution, taxonomy, and plant-microbe interactions. Data SummaryGenomes sequenced in this study were submitted to the NCBI database under the accession numbers: JCBCPT000000000

genomics↗
bioRxiv2021

Workflows for rapid functional annotation of diverse arthropod genomes

Genome sequencing of a diverse array of arthropod genomes is already underway and these genomes will be used to study human health, agriculture, biodiversity and ecology. These new genomes are intended to serve as community resources and provide the foundational information that is required to apply omics technologies to a more diverse set of species. However, biologists require genome annotation to use these genomes and derive a better understanding of complex biological systems. Genome annotation incorporates two related but distinct processes: demarcating genes and other elements present in genome sequences (structural annotation); and associating function with genetic elements (functional annotation). While there are well established and freely available workflows for structural annotation of gene identification in newly assembled genomes, workflows for providing the functional annotation required to support functional genomics studies are less well understood. Genome-scale functional annotation is required for functional modeling (enrichment, networks, etc.) and a first-pass genome-wide functional annotation effort can rapidly identify under-represented gene sets for focused community annotation efforts. We present an open source, open access and containerized pipeline for genome-scale functional annotation of insect proteomes and apply it to a diverse range of arthropod species. We show that the performance of the predictions is consistent across a set of arthropod genomes with varying assembly and annotation quality.Complete instructions for running each component of the functional annotation pipeline on the command line, a high performance computing cluster and the CyVerse Discovery Environment can be found at the readthedocs site (https://agbase-docs.readthedocs.io/en/latest/agbase/workflow.html). Simple summaryGenomic technologies are accumulating information about genes at a faster rate than ever before, and sequencing initiatives like the Earth Biogenome Project, i5k and Ag100Pest are expected to increase this rate of acquisition. However, if genomic sequencing is to be used for improvement of human health, agriculture and our understanding of biological systems, it is necessary to identify genes and understand how they contribute to biological outcomes. While there are several well-established workflows for assembling genomic sequences and identifying genes, understanding gene function is essential to create actionable knowledge. Moreover this functional annotation process must be easily accessible and provide information at a genomic scale to keep up with new sequence data. We report a well defined workflow for rapid functional annotation of whole proteomes to produce Gene Ontology and pathways information. We test this workflow on a diverse set of arthropod genomes and compare it to common arthropod reference genomes. The workflow we described is freely and publicly available via a web interface on CyVerse or as biocontainers that can be deployed scalably on local computing systems.

bioinformatics↗
bioRxiv2022

Transposable element competition in shaping the deer mouse genome

The genomic landscape of transposable elements (TEs) varies dramatically across species, with some TEs demonstrating greater success in colonizing particular lineages than others. In mammals, LINE retrotransposons typically occupy more of the genome than any other TE and most LINE content is represented by a single family: L1. Here, we report an unusual genomic landscape of TEs in the deer mouse, Peromyscus maniculatus, a model for studying the genomic basis of adaptation. In contrast to other previously examined mammalian species, LTR elements occupy more of the deer mouse genome than LINEs (11% and 10% respectively). This pattern reflects a combination of relatively low LINE activity in addition to a massive invasion of lineage-specific endogenous retroviruses (ERVs). Deer mouse ERVs exhibit diverse origins spanning the retroviral phylogeny suggesting that these rodents have been host to a wide range of exogenous retroviruses. Notably, we were able to trace the origin of one ERV lineage, which arose within the last [~]11-18 million years, to a close relative of feline leukemia virus, revealing inter-ordinal horizontal transmission of these zoonotic viruses. Several lineage-specific ERV subfamilies have attained very high copy numbers, with the top five most abundant accounting for [~]2% of the genome. Concomitant to the expansive diversification of ERVs, we also observe a massive expansion of Kruppel-associated box domain-containing zinc finger genes (KZNFs), which likely control ERV activity and whose expansion may have been partially facilitated by ectopic recombination between ERVs. We also find evidence that ERVs directly impacted the evolutionary trajectory of LINEs by outcompeting them for genomic sites and frequently disrupting autonomous LINE copies. Together, our results illuminate the genomic ecology that shaped the deer mouse genomes TE landscape, opening up a range of opportunities to investigate the evolutionary processes that give rise to variation in mammalian genome structure. SummaryTransposable elements (TEs) are a highly diverse collection of genetic elements capable of mobilizing in genomes and function as important drivers of genome evolution. The landscape of TEs in a genome have been compared to a genomic ecosystem, with interactions between TEs and each other as well as TEs and their host, dictating the evolutionary success of TE lineages. While TE diversity and copy numbers can vary dramatically across taxa, the evolutionary reasons for this variation remain poorly understood. In mammals, long interspersed nuclear elements (LINEs) typically dominate, occupying more of the genome than any other TE. Here, we report a unique case in the deer mouse (Peromyscus maniculatus) in which long terminal repeat (LTR) retrotransposons occupy more of the genome than LINEs. We investigate the evolutionary origins and implications of the deer mouses distinct genomic landscape, revealing ecological processes that helped shape its evolution. Together, our results provide much-needed insight into the evolutionary processes that give rise to variation in mammalian genome structure.

genomics↗
bioRxiv2025

Replication rate-information storage trade-off shapes genome architecture across domains

Genome size varies widely among organisms, from compact genomes of bacteria to vast and complex genomes of eukaryotes. In this study, we theoretically identify the evolutionary pressures that may have driven this divergence in genome size. We use a parameter-free model to study genome size evolution under selection pressure to minimize replication time and maximize information storage capacity. We substantiate this choice by demonstrating a correlation between replication time and genome size using literature data. We show that bacteria tend to reduce genome size, constrained by a single replication origin, while eukaryotes expand their genomes by incorporating multiple replication origins. We propose a connection between genome size and cellular energetics, suggesting that endosymbiotic organelles, mitochondria and chloroplasts, evolutionarily regulate the number of replication origins, thereby influencing genome size in eukaryotes. We substantiate this claim by showing a correlation between organelle count and genome size data. We argue that nucleotide skews, present in nearly all genomes studied, and are used to identify replication origins, directly influence DNA unzipping kinetics, and hence its replication time. This connection enables us to derive nearly universal observations, such as Chargaffs second parity rule and replichore symmetrization, as adaptive consequences. We argue that high skews lead to faster replication, and substantiate it by showing a correlation between skews and replication speed in bacteria. The model reproduces more evolutionary genomic observations, such as a general preference for deletions over insertions across domains, and elongation and high variance of genome size under reduced selection pressure for replication rate, an integral component of C-value paradox. We highlight the possibility of regulation of the firing of latent replication origins in response to cues from the extracellular environment, leading to the regulation of cell cycle rates in multicellular eukaryotes. Significance StatementUnderstanding the forces shaping genome architecture is a long-standing challenge in evolutionary biology. Our study demonstrates that the balance between replication speed, influenced by nucleotide skews, and information storage, constrained by cellular energetics, drives the divergence in genome size between bacteria and eukaryotes. By quantifying selection pressure as the ratio of replication time to genomic information storage capacity, we show that this pressure enforces adaptive constraints, giving rise to observed features such as symmetric replichores and Chargaffs second parity rule. These insights not only help us resolve an enduring evolutionary puzzle, but also offer a unified framework linking genome organization, cellular specialization, and even potential mechanisms underlying carcinogenesis.

genomics↗
bioRxiv2019

The multiple population genetic and demographic routes to islands of genomic divergence

1. The way that organisms diverge into reproductively isolated species is a major question in biology. The recent accumulation of genomic data provides promising opportunities to understand the genomic landscape of divergence, which describes the distribution of differences across genomes. Genomic areas of unusually high differentiation have been called genomic islands of divergence. Their formation has been attributed to a variety of mechanisms, but a prominent hypothesis is that they result from divergent selection over a small portion of the genome, with surrounding areas homogenised by gene flow. Such islands have often been interpreted as being associated with divergence with gene flow. However other mechanisms related to genetic architecture and population history can also contribute to the formation of genomic islands of divergence.\n\n2. We currently lack a quantitative framework to examine the dynamics of genomic landscapes under the complex and nuanced conditions that are found in natural systems. Here, we develop an individual-based simulation to explore the dynamics of diverging genomes under various scenarios of gene flow, selection and genotype-phenotype maps.\n\n3. Our modelling results are consistent with empirical observations demonstrating the formation of genomic islands under genetic isolation. Importantly, we have quantified the range of conditions that produce genomic islands. We demonstrate that the initial level of genetic diversity, drift, time since divergence, linkage disequilibrium, strength of selection and gene flow are all important factors that can influence the formation of genomic islands. Because the accumulation of genomic differentiation over time tends to erode the signal of genomic islands, genomic islands are more likely to be observed in recently divergent taxa, although not all recently diverged taxa will necessarily exhibit islands of genomic divergence. Gene flow primarily slows the swamping of islands of divergence with time.\n\n4. By using this framework, further studies may explore the relative influence of particular suites of events that contribute to the emergence of genomic islands under sympatric, parapatric and allopatric conditions. This approach represents a novel tool to explore quantitative expectations of the speciation process, and should prove useful in elucidating past and projecting future genomic evolution of any taxa.

genomics↗
bioRxiv2026

Quantitative comparison of fungal genome assembly strategies using short and long-reads from simulated and empirical sequencing data

High-quality fungal reference genomes are essential for comparative, functional, and evolutionary studies, yet fungal genome features such as repeats, structural rearrangements, accessory chromosomes, and intron-rich genes can complicate genome assembly and the selection of cost-effective sequencing strategies. Here, we benchmark fungal genome assembly performance using simulated and empirical short- and long-read datasets to evaluate how sequencing depth, assembler choice, and genome characteristics influence contiguity, completeness, accuracy, and computational requirements. Using simulated reads from complete fungal genomes spanning diverse sizes and compositions, we evaluated short-read, long-read, hybrid, and polished long-read assemblies across sequencing depths from 10X to 100X. Key trends were validated using empirical sequencing data from 10 fungal isolates assembled with multiple strategies, including different Flye assembler parameter sensitivity and short-read polishing. Across datasets, long reads produced the largest improvements in contiguity, with most gains achieved at [~]20-40X coverage and diminishing returns beyond moderate depth. Short-read polishing substantially improved base-level accuracy at relatively low cost, with [~]10-20X coverage often sufficient to approach maximal error reduction. Hybrid assemblers showed strong algorithmic variability, with trade-offs between contiguity, error rates, and computational demand. Genome architecture also influenced outcomes, as larger and more feature-dense genomes benefited more from long-read data while GC content had limited impact. Overall, our results suggest that moderate long-read coverage ([~]30-40X) combined with modest short-read polishing ([~]10-20X), particularly using Flye plus Polypolish, provides a strong balance of contiguity, completeness, accuracy, and resource efficiency for generating high-quality fungal genome assemblies. Impact statementFungal genome sequencing is expanding rapidly across ecology, plant pathology, biotechnology, and clinical and veterinary microbiology, yet experimental design decisions regarding sequencing depth, assembler selection, and hybrid workflows are still largely guided by bacterial benchmarking studies or limited single-species comparisons. Because fungal genomes vary widely in size, repeat content, and gene architecture, these assumptions can lead to inefficient sequencing strategies, increased computational costs, and suboptimal assemblies. Here, we develop a reproducible assembly benchmarking framework that combines large-scale simulations from 66 complete fungal genomes spanning plant, animal, and human-associated taxa with newly generated short- and long-read sequencing data from 10 field-collected isolates. This approach enables evaluation of assembler performance across diverse genome architectures and tests whether patterns identified in simulations translate to real biological datasets. Across both simulated and empirical datasets, we show that reliable fungal genome reconstruction can be achieved without excessive sequencing depth by identifying consistent performance thresholds. Assembly contiguity and completeness stabilize at moderate long-read coverage, after which improvements depend more strongly on assembler choice and genome structure than on additional data volume. Hybrid workflows show trade-offs in accuracy, contiguity, and computational demand, whereas targeted short-read polishing provides an efficient strategy for improving base-level accuracy. These findings offer practical guidance for fungal genome assembly and support more robust downstream genomic analyses across non-model microbial systems, including ecologically, agriculturally, and clinically important fungi. Data summaryThe reference fungal genomes used for simulation are available in the NCBI Assembly database under the accession numbers listed in Supplementary Table S1. Empirical raw sequencing data generated for this study are deposited in the NCBI Sequence Read Archive (SRA) under accession PRJNA1474061. All scripts used for read simulation, assembly, polishing, benchmarking, and statistical analyses are available in the project GitHub repository (github.com/bielasilva/fungi_assembly_benchmarking). Software versions, parameters, and workflow configurations are provided within the repository and detailed in the Methods.

genomics↗
bioRxiv2020

Bacterial genomic GC content drifts slowly downward due to a biased mutation rate

In every kingdom of life, GC->AT transitions occur more frequently than any other type of mutation due to the spontaneous deamination of cytidine. In eukaryotic genomes, this slow loss of GC base pairs is counteracted by biased gene conversion which increases genomic GC content as part of the recombination process. However, this type of biased gene conversion has not been observed in bacterial genomes so we hypothesized that GC->AT transitions cause a reduction of genomic GC content in prokaryotic genomes on an evolutionary time scale. To test this hypothesis, we used a phylogenetic approach to analyze triplets of closely related genomes representing a wide range of the bacterial kingdom. The resulting data indicate that genomic GC content is slowly declining in bacterial genomes where GC base pairs comprise 40% or more of the total genome. In contrast, genomes containing less than 40% GC base pairs have fewer opportunities for GC->AT transitions to occur so genomic GC content is relatively stable or actually increasing at a slow rate. It should be noted that this observed change in genomic GC content is the net change in shared parts of the genome and does not apply to parts of the genome that have been lost or acquired since the genomes being compared shared common ancestor. However, a more detailed analysis of two Caulobacter genomes revealed that the acquisition of mobile elements by the two genomes actually reduced the total genome content as well.

microbiology↗
bioRxiv2021

A non-adaptive demographic mechanism for genome expansion in Streptomyces

The evolution of microbial genome size is driven by gene acquisition and loss events that occur at scales from individual genomes to entire pangenomes. The equilibrium between gene gain and loss is shaped by evolutionary forces, including selection and drift, which are in turn influenced by population demographics. There is a well-known bias towards deletion in microbial genomes, which promotes genome streamlining. Less well described are mechanisms that promote genome expansion, giving rise to the many microbes, such as Streptomyces, that have unusually large genomes. We find evidence of genome expansion in Streptomyces sister-taxa, and we hypothesize that a recent demographic range expansion drove increases in genome size through a non-adaptive mechanism. These Streptomyces sister-taxa, NDR (northern-derived) and SDR (southern-derived), represent recently diverged lineages that occupy distinct geographic ranges. Relative to SDR genomes, NDR genomes are larger, have more genes, and their genomes are enriched in intermediate frequency genes. We also find evidence of relaxed selection in NDR genomes relative to SDR genomes. We hypothesize that geographic range expansion, coupled with relaxed selection, facilitated the introgression of non-adaptive horizontally acquired genes, which accumulated at intermediate frequencies through a mechanism known as genome surfing. We show that similar patterns of pangenome structure and genome expansion occur in a simulation that models the effects of population expansion on genome dynamics. We show that non-adaptive evolutionary phenomena can explain expansion of microbial genome size, and suggest that this mechanism might explain why some bacteria with large genomes can be found in soil.

microbiology↗
bioRxiv2021

GMSECT: Genome-Wide Massive Sequence Exhaustive Com-parison Tool for structural and copy number variations

GMSECT is a parallel robust Application Interface that efficiently handles the large genomic sequences for rapid and efficient processing. It is a message passing interface based parallel computing Tool that can be operated on a cluster for Massive Sequences Exhaustive Comparison, to identify matches such as the structural variants. The GMSECT algorithm can be implemented using other parallel application programming interfaces as well such as Posix-Threads or can even be implemented in a serial submission fashion. There is complete flexibility to the choice of comparison tool that can be deployed and with the optional parameters as of the choice of comparison tools to suit the speed, sensitivity and specificity of pair wise alignment. The algorithm is simple and robust, and can be applied to compare multiple genomes, chromosomes or large sequences, of different individuals for personalized genome comparison and works good for homologous as well as distant species. The tool can even be applied to smaller genomes like the microbial genome such as the Escherichia coli or algae such as Chlamydomonas reihardtti or yeast Saccharomyces cerevisiae to quickly conduct comparisons, and thus finds its application to the pharmaceuticals and microbial product based firms for research and development. The application Interface can efficiently and rapidly compare massive sequences to detect for the presence of numerous types of DNA variation existing in the genome ranging from single nucleotide polymorphism (SNPs) to larger structural alterations, such as copy-number variants (CNVs) and inversions. The new algorithm has been tested for comparing the chromosome 21 of Celeras R27c compilation with all the 48 chromosomes of Celeras R27c compilation and with all the 48 chromosomes of the human Build 35 reference sequence, which took just 2 Hours and 10 minutes using the pair wise blast algorithm choice and with 110 processors each with 2.2 GHz capacity and 2 GB memory. GMSECT facilitates rapid scanning and interpretation in personalized sequencing project. The application interface with the above resources and alignment choice is estimated to do exhaustive comparison of the human genome with itself in just 2.35 days. An exhaustive comparison of an individuals genome with a reference genome would comprise of a two self genome comparison and a non-self genome comparison which is estimated to take about 9.4 days with the above resources. With the advent of personalized genome sequencing project, it would be desirable to compare 100s of individuals genome with a reference genome. This would involve a non-self genome and a self genome comparison for each genome, and would take around 7 days for each individuals genome using GMSECT and the above mentioned resources.

bioinformatics↗
bioRxiv2018

Resolving the complex Bordetella pertussis genome using barcoded nanopore sequencing

The genome of Bordetella pertussis is complex, with high GC content and many repeats, each longer than 1,000 bp. Short-read DNA sequencing is unable to resolve the structure of the genome; however, long-read sequencing offers the opportunity to produce single-contig B. pertussis assemblies using sequencing reads which are longer than the repetitive sections. We used an R9.4 MinION flow cell and barcoding to sequence five B. pertussis strains in a single sequencing run. We then trialled combinations of the many nanopore-user-community-built long-read analysis tools to establish the current optimal assembly pipeline for B. pertussis genome sequences. Our best long-read-only assemblies were produced by Canu read correction followed by assembly with Flye and polishing with Nanopolish, whilst the best hybrids (using nanopore and Illumina reads together) were produced by Canu correction followed by Unicycler. This pipeline produced closed genome sequences for four strains, revealing inter-strain genomic rearrangement. However, read mapping to the Tohama I reference genome suggests that the remaining strain contains an ultra-long duplicated region (over 100 kbp), which was not resolved by our pipeline. We have therefore demonstrated the ability to resolve the structure of several B. pertussis strains per single barcoded nanopore flow cell, but the genomes with highest complexity (e.g. very large duplicated regions) remain only partially resolved using the standard library preparation and will require an alternative library preparation method. For full strain characterisation, we recommend hybrid assembly of long and short reads together; for comparison of genome arrangement, assembly using long reads alone is sufficient.\n\nDATA SUMMARYO_LIFinal sequence read files (fastq) for all 5 strains have been deposited in the SRA, BioProject PRJNA478201, accession numbers SAMN09500966, SAMN09500967, SAMN09500968, SAMN09500969, SAMN09500970\nC_LIO_LIA full list of accession numbers for Illumina sequence reads is available in Table S1\nC_LIO_LIAssembly tests, basecalled read sets and reference materials are available from figshare: https://figshare.com/projects/Resolving_the_complex_Bordetella_pertussis_genome_using_barcoded_nanopore_sequencing/31313\nC_LIO_LIGenome sequences for B. pertussis strains UK36, UK38, UK39, UK48 and UK76 have been deposited in GenBank; accession numbers: CP031289, CP031112, CP031113, QRAX00000000, CP031114\nC_LIO_LISource code and full commands used are available from Github: https://github.com/nataliering/Resolving-the-complex-Bordetella-pertussis-genome-using-barcoded-nanopore-sequencing\nC_LI\n\nIMPACT STATEMENTOver the past two decades, whole genome sequencing has allowed us to understand microbial pathogenicity and evolution on an unprecedented level. However, repetitive regions, like those found throughout the B. pertussis genome, have confounded our ability to resolve complex genomes using short-read sequencing technologies alone. To produce closed B. pertussis genome sequences it is necessary to use a sequencing technology which can generate reads longer than these problematic genomic regions. Using barcoded nanopore sequencing, we show that multiple B. pertussis genomes can be resolved per flow cell. Use of our assembly pipeline to resolve further B. pertussis genomes will advance understanding of how genome-level differences affect the phenotypes of strains which appear monomorphic at nucleotide-level.\n\nThis work expands the recently emergent theme that even the most complex genomes can be resolved with sufficiently long sequencing reads. Additionally, we utilise a more widely accessible alternative sequencing platform to the Pacific Biosciences platform already used by large research centres such as the CDC. Our optimisation process, moreover, shows that the analysis tools favoured by the sequencing community do not necessarily produce the most accurate assemblies for all organisms; pipeline optimisation may therefore be beneficial in studies of unusually complex genomes.

microbiology↗
bioRxiv2020

Draft Genome Assemblies and Annotations of Agrypnia vestita Walker, and Hesperophylaxmagnus Banks Reveal Substantial Repetitive Element Expansion in Tube Case-making Caddisflies (Insecta: Trichoptera)

Trichoptera (caddisflies) play an essential role in freshwater ecosystems; for instance, larvae process organic material from the water and are food for a variety of predators. Knowledge on the genomic diversity of caddisflies can facilitate comparative and phylogenetic studies thereby allowing scientists to better understand the evolutionary history of caddisflies. While Trichoptera are the most diverse aquatic insect order, they remain poorly represented in terms of genomic resources. To date, all long-read based genomes have been sequenced from individuals in the retreat-making suborder, Annulipalpia, leaving [~]275 Ma of evolution without high-quality genomic resources. Here, we report the first long-read based de novo genome assemblies of two tube case-making Trichoptera from the suborder Integripalpia, Agrypnia vestita Walker and Hesperophylax magnus Banks. We find that these tube case-making caddisflies have genome sizes that are at least three-fold larger than those of currently sequenced annulipalpian genomes and that this pattern is at least partly driven by major expansion of repetitive elements. In H. magnus, long interspersed nuclear elements (LINEs) alone exceed the entire genome size of some annulipalpian counterparts suggesting that caddisflies have high potential as a model for understanding genome size evolution in diverse insect lineages. SignificanceThere is a lack of genomic resources for aquatic insects. So far, only three high-quality genomes have been assembled, all from individuals in the retreat-making suborder Annulipalpia. In this article, we report the first high-quality genomes of two case-making species from the suborder Integripalpia, which are essential for studying genomic diversity across this ecologically diverse insect order. Our research reveals larger genome sizes in the tube case-makers (suborder Integripalpia, infraorder Phryganides), accompanied by a disproportionate increase of repetitive DNA. This suggests that genome size is at least partly driven by a major expansion of repetitive elements. Our work shows that caddisflies have high potential as a model for understanding how genomic diversity might be linked to functional diversification and forms the basis for detailed studies on genome size evolution in caddisflies. Data depositionThis project has been deposited at NCBI under the Bioproject ID: PRJNA668166

genomics↗
bioRxiv2026

Australian giant kelp genome assemblies show distinct Southern Hemisphere genetics

Giant kelp, Macrocystis pyrifera, occurs across northern and southern Hemisphere temperate coasts and is at high risk from ocean warming. Few giant kelp forests remain across the Southeast Australian shelf, while a handful of forests are actively being restored. Genomic resources can greatly aid in the conservation of remnant populations and enhance restoration efforts. Reference genomes are a fundamental resource as they are a prerequisite to, or enhance, many analyses used in conservation genomics. A single reference genome is available for giant kelp, assembled from a Californian haploid specimen. However, increasing evidence of genetic divergence between Northern and Southern Hemisphere populations highlights the need for regionally representative reference genomes. Here, we present two genome assemblies from the diploid vegetative tissue of Australian giant kelp specimens. We performed de novo genome assembly using long-read sequencing (PacBio HiFi and ONT R10.4 Simplex) and scaffolded the assemblies with the ONT reads, assembling 98-99% of the genomes into 35 pseudo-chromosomes. Genome sizes ranged from 528-534 Mbp, with BUSCO completeness scores of 97-98% and QV scores of 51-52. Genome annotation identified 17,565-17,800 genes in the Australian genomes. Genomic divergence between the Australian and Californian giant kelp genomes was seven-fold greater than between Australian genomes (1.5% vs 0.2%), supporting a Northern-Southern Hemisphere genetic divergence. Functional divergence was also observed between Australian and Californian genomes, reflected by differing patterns of enrichment in gene ontologies linked to energy metabolism, proteostasis and stress responses. These two new genome assemblies will serve as valuable resources for ongoing research into Southern Hemisphere giant kelp genetics, while providing the basis for genomic-guided conservation and restoration of remnant giant kelp forests in Australia.

genomics↗
bioRxiv2026

The Dynamics of Long Terminal Repeat Proliferation in the Hesperis matronalis Genome

Genome sizes vary across four orders of magnitude in flowering plants, with consequences for evolution. Much of this variation is due to differences in transposable element content, particularly in long terminal repeat (LTR) retrotransposons. Despite their importance in plant genome evolution, LTRs have long been challenging to characterize because of their repetitive nature, but recent advances in sequencing allow more detailed explorations of their behavior. They are now known to occupy distinct genomic niches, and to evolve and proliferate over time as they escape host controls. In this study, we present a new genome sequence of the largest Brassicaceae genome, Hesperis matronalis, and describe its transposable element complement and how LTRs contributed to its genome expansion. We find evidence for both early proliferation of Ty3 elements and rapid recent expansion of Ty1-copia elements. In addition, we place the H. matronalis LTRs in a broader context of LTR evolution in the Brassicaceae, showing that the dominant copia families are part of an evolutionary radiation endemic to Hesperis. Finally, we describe differences in LTR age, proximity to genes, and apparent removal rate which suggest consistent genomic niches over the lifespan of a TE family. These results shed light on how LTRs evolve dynamically with host genomes, and have contributed to the expansion of the largest genome in the Brassicaceae. Significance StatementOrganisms differ in many ways, including the size of their genomes. Genome size, in turn, can affect many aspects of evolution, such as what kinds of mutations are available to natural selection and how effectively natural selection can act. The "complexity" of an organism does not predict its genome size; rather, much of this variation is explained by the amount of transposable elements, or "jumping genes," that inhabit a genome. This study describes the evolutionary history of the transposable elements in dames rocket (Hesperis matronalis), which has the largest genome of any plant in the mustard family (Brassicaceae). We find that the genome inflation in Hesperis is due to transposable elements that are not found elsewhere in the Brassicaceae, and likely diversified within Hesperis. Characterizing the evolution and behavior of transposable elements in this genome offers a better understanding of the forces that determine genome size and ultimately affect the evolution of life on earth.

genomics↗
bioRxiv2020

Mycobacterium tuberculosis complex lineage 5 exhibits high levels of within-lineage genomic diversity

Pathogens of the Mycobacterium tuberculosis complex (MTBC) are considered monomorphic, with little gene content variation between strains. Nevertheless, several genotypic and phenotypic factors separate the different MTBC lineages (L), especially L5 and L6 (traditionally termed Mycobacterium africanum), from each other. However, genome variability and gene content especially of L5 and L6 strains have not been fully explored and may be potentially important for pathobiology and current approaches for genomic analysis of MTBC isolates, including transmission studies. We compared the genomes of 358 L5 clinical isolates (including 3 completed genomes and 355 Illumina WGS (whole genome sequenced) isolates) to the L5 complete genomes and H37Rv, and identified multiple genes differentially present or absent between H37Rv and L5 strains. Additionally, considerable gene content variability was found across L5 strains, including a split in the L5.3 sublineage into L5.3.1 and L5.3.2. These gene content differences had a small knock on effect on transmission cluster estimation, with clustering rates influenced by the selection of reference genome, and with potential over-estimation of recent transmission when using H37Rv as the reference genome. Our data show that the use of H37Rv as reference genome results in missing SNPs in genes unique for L5 strains. This potentially leads to an underestimation of the diversity present in the genome of L5 strains and in turn affects the transmission clustering rates. As such, a full capture of the gene diversity, especially for high resolution outbreak analysis, requires a variation of the single H37Rv-centric reference genome mapping approach currently used in most WGS data analysis pipelines. Moreover, the high within-lineage gene content variability suggests that the pan-genome of M. tuberculosis is at least several kilobases larger than previously thought, implying a concatenated or reference-free genome assembly (de novo) approach may be needed for particular questions. Data summarySequence data for the Illumina dataset are available at European Genome-phenome Archive (EGA; https://www.ebi.ac.uk/ega/) under the study accession numbers PRJEB38317 and PRJEB38656. Individual runs accession numbers are indicated in Table S8. PacBio raw reads for the L5 Benin genome are available on the ENA accession SAME3170744. The assembled L5 Benin genome is available on NCBI with accession PRJNA641267. To ensure naming conventions of the genes in the three L5 genomes can be followed, we have uploaded these annotated GFF files to figshare at https://doi.org/10.6084/m9.figshare.12911849.v1. Custom python scripts used in this analysis can be found at https://github.com/conmeehan/pathophy.

genetics↗