Biology subjects

Dutilh, B. E.

Publications and source records attributed to Dutilh, B. E..

At least 19 recordsLinked to original sources

bioRxiv2026

High-throughput genomic feature extraction reveals environmental adaptations of prokaryotes

Understanding the adaptations of microorganisms to their environment is key to predicting the stability and dynamics of microbial communities. To uncover molecular mechanisms of environmental response, we extracted genomic features from 13,554 prokaryotic isolates, and trained machine learning models to identify which ones are most strongly associated with the microbial salinity, temperature, oxygen, and pH preferences. To extract these features in high throughput, including gene families, non-coding RNAs (ncRNAs), oligonucleotides, and amino acid usage, we built FxTractor, a scalable and adjustable pipeline available at: https://github.com/MGXlab/FxTractor. We validated the performance of our models with experimental data from a newly isolated deep-sea extremophile belonging to the genus Limnochorda that is not well-represented among the ML training sets, showing strong agreement between predictions and the conditions used to isolate this strain. Our analysis revealed specific gene and ncRNA families associated with each of the four environmental parameters, uncovering both established and potentially new molecular mechanisms. Examples include the bacterial large Signaling Recognition Particle in isolates that are able to grow at high temperatures ([≥]55{degrees}C), suggesting a role in translational pausing and structural stability under thermal stress. We also found the anti-hemB ncRNA to be associated with low-salinity (<0.7% NaCl), indicating a conserved antisense mechanism regulating the energetic costs of heme biosynthesis. Together, these findings provide new insights into microbe-environment interactions, and show how FxTractor enables high throughput discovery of genomic associations.

bioinformatics↗

bioRxiv2026

Museum specimens reveal evolutionary drivers and functions of oral microbiome in mammals

The microbiomes of wild animals, despite being integral to host survival, remain largely unexplored, particularly beyond the gut. Here, using > 450 museum-preserved dental calculus samples from 34 ecologically and phylogenetically diverse mammalian species, most of them previously unstudied, we determine the main drivers of oral microbiome evolution. We show that, similar to the gut, the oral microbiome is shaped by host ecology, particularly diet, and to a lesser extent host phylogenetic relationships. It may contribute to host adaptation, by synthesising essential micronutrients and degrading potentially harmful compounds. We also find that some oral microorganisms consistently associated with mammals throughout over evolutionary time scales and provide evidence for a likely oral origin of some rumen taxa. Together our findings demonstrate the importance of this mostly overlooked microbial community for mammalian biology.

evolutionary biology↗

bioRxiv2026

Computational prediction resolves thousands of homooligomeric phage protein structures

Bacteriophages (phages) play essential roles in microbial systems, yet most phage proteins remain poorly characterised. Protein tertiary and quaternary structure information contributes valuable information about protein function. As many phage proteins function as homooligomers, complexes that consist of multiple identical subunits, there is great interest in computationally predicting their configurations. Here we present a computational framework, the Phage Homomer Level Estimate and Generation Method (PHLEGM) for inferring homooligomeric states directly from the protein sequence by combining AlphaFold-Multimer modelling with inter-subunit interface quality assessment. We proceeded to experimentally validate two out of nine predicted homooligomers using size exclusion chromatography and complementary hydrodynamic techniques. These efforts confirmed our predictions for a dimer and a trimer, highlighting the value of experimentally benchmarked computational predictions and showing the challenges of heterologous phage protein production. Applied to >22,000 phage protein sequences in the PHROGs database, our approach revealed extensive diversity in phage homooligomeric protein complexes. Benchmarking against protein language model-based predictors on a curated reference set of known phage homooligomers demonstrated superior accuracy of our structure-based method, achieving robust performance in classifying protein homooligomeric states, with the highest accuracy observed for trimers and higher-order complexes. These results highlight the value of computational predictions to decipher the complexities of the vast viral sequence space. All predicted complex structures and functional inferences are made publicly available to support structural and functional studies of phage proteins.

microbiology↗

bioRxiv2026

Cosmopolitan gut bacteriophages expand the phenotype of health-related bacteria

Metagenomics has vastly expanded our knowledge of the human gut virome, yet the focus on dominant taxa has missed low-abundant but highly prevalent bacteriophages. Here, high-sensitivity taxonomic profiling of 53,976 samples from 42 countries, including non-industrialised populations and ancient DNA from five archaeological sites, reveals that only 0.3% of 64,238 phage genera are cosmopolitan, occurring in >20% of all subjects and on six continents. The most prevalent of these is Mushuvirus, with two distinct species detected in 60% of individuals, previously missed due to artefacts in reference genomes and their consistently low relative abundance (<0.8%). Population genomics shows that while one species is globally ubiquitous, the other predominates in individuals with West Eurasian ancestry. Together with 12 additional novel genera, we propose to classify these viruses into a new family Mushuviridae, found in [~]89% of studied humans. These transposable phages integrate into diverse hosts from two taxonomic orders of short-chain fatty acid-producing bacteria. Multi-omics experiments with Faecalibacterium demonstrate that hosts produce and secrete Mushuvirus-encoded receptor-binding proteins, which are actively diversified in lysogenic hosts. Together, these findings illuminate a globally distributed, active phage lineage that has persisted for millennia and continues to reshape the functional repertoire of bacteria central to human gut health.

microbiology↗

bioRxiv2025

Widespread synchronization of codon usage in functionally related genes

The usage of synonymous codons varies along the genome, with strong biases in conserved and highly expressed genes that are optimized for efficient translation. The extent to which selection shapes codon usage in other genes, as well as the associations between gene function, gene expression, and codon usage, remains an important open question. We developed and optimized a novel approach to detect synchronized changes in codon usage patterns and applied it to 15,005 species-representative bacterial genomes spanning the 15 most represented phyla. We show that codon usage is extensively shaped by selection in both highly and lowly expressed genes, with at least [~]20-46% of gene families showing synchronized codon usage evolution across genomes. We reveal that gene pairs with parallel codon usage adaptation are co-expressed, co-regulated, metabolically connected, and functionally associated. By identifying synchronized codon usage evolution between gene pairs, we have generated a genome-wide set of functional associations reflecting correlated expression across species. This underappreciated layer of coordinated codon usage adaptation has important implications for function discovery and engineering.

genomics↗

bioRxiv2025

Scalable and systematic hierarchical virus taxonomy with vConTACT3

Viruses are key players in diverse ecosystems, but studying their impacts is technically and taxonomically challenging. Taxonomic complexities derive from undersampling, diverse DNA and RNA genomes with multiple evolutionary origins, and lack of a universal barcode gene. While virus ecogenomics has expanded access to and understanding of the virosphere, available classification tools poorly scale to modern discovery-based datasets, lack taxonomic resolution, and/or are unable to classify novel sequence space. Here we develop, benchmark, and release vConTACT3, a machine learning-based tool that improves scalability and accuracy, adds extensive user-requested features, expands classification to both eukaryote and prokaryote viruses for 4/6 officially recognized realms, and establishes accurate hierarchical taxonomy from genus to order. Application to 48,069 public virus genomes provided new taxonomy assignments for thousands of taxa, revealed support for fewer taxonomic ranks than currently available, and systematically identified taxonomically problematic areas across the virosphere.

bioinformatics↗

bioRxiv2025

Barbell Resolves Demultiplexing and Trimming Issues in Nanopore Data

BackgroundOxford Nanopore sequencing enables long-read sequencing across diverse applications, yet the experimental artifacts introduced by Nanopore barcoding are not well characterized. These artifacts can affect demultiplexing accuracy and downstream analyses. ResultsWe performed a rapid barcoding experiment on 66 diagnostic samples and found that 83% of reads carried the expected single-barcode pattern, while 17% contained multiple barcodes or other artifacts. Current demultiplexers, including the widely used Dorado, fail to correctly handle these complex cases, leaving approximately 7% of reads partially trimmed and contaminated with adapter fragments. Additional issues include the presence of two barcodes at the same read end--either identical, originating from the same sample, or different, introduced after pooling. The latter can lead to barcode bleeding when the outer barcode is incorrectly selected. To address these challenges, we developed Barbell, a pattern-aware demultiplexer that detects all barcode configurations. Barbell reduces trimming errors by three orders of magnitude, minimizes barcode bleeding, and supports custom experimental setups such as shorter barcodes, dual-end barcodes, and custom flank sequences. ConclusionsOur results highlight the impact of complex barcode attachments in Nanopore sequencing and demonstrate that Barbell drastically reduces their effects on downstream analyses. Barbell is open source and available at https://github.com/rickbeeloo/barbell.

bioinformatics↗

bioRxiv2025

Bacterial community adaptation after freshwater and seawater coalescence

Microbial community coalescence, the merging of entire microbial communities, is common across ecosystems, particularly in estuaries where freshwater and seawater mix. The complexity of these habitats makes the in situ study of community dynamics after coalescence challenging, highlighting the need for controlled experiments to unravel the factors influencing the coalescence in the estuary. To study these processes, we combined natural freshwater and seawater bacterial communities at five different mixing ratios and incubated them in parallel microcosms containing freshwater or seawater incubation media. Forty mixed communities were tracked over six passages using Nanopore full-length 16S rRNA gene sequencing. In the original field samples, freshwater hosted more diverse communities than seawater. While the communities were structurally distinct, shared bacterial families accounted for approximately 95% of total reads. Many low-abundance taxa were lost upon laboratory incubation, while potentially faster-growing ones were enriched. We found that the coalescence outcome was strongly shaped by the incubation media, whereas the mixing ratio had a minor influence. Mixed communities converged toward the source community native to the incubation media, with increasing similarity at a higher source community proportion. In freshwater, a 25% inoculum of the freshwater community was sufficient to re-establish a near-native freshwater community, whereas in seawater, similarity to the native seawater community depended on the inoculation ratio. Network analysis showed a tightly connected module of seawater families, reflecting their shared habitat preference or cooperation, whereas freshwater families were more loosely connected. We also observed that most families were unaffected by mixing ratios or temporal dynamics, with only a few showing mixing ratio dependence. For instance, the freshwater family Comamonadaceae and seawater families Marinomonadaceae and Pseudoalteromonadaceae were dominant in their respective native environments, and increased in mixed communities in proportion to their initial source proportion. Overall, environmental filtering had a stronger impact than the mixing ratio of source communities on coalesced communities, with habitat-specific taxa further modulating the outcome. These findings advanced our understanding of microbial responses to coalescence and provided insights into microbial community assembly in dynamic estuarine systems. HighlightsO_LIEnvironmental filtering outweighs the microbial source community ratio in shaping coalescence outcomes. C_LIO_LIAsymmetric resilience: freshwater communities require a lower inoculum to re-establish than seawater communities. C_LIO_LIModularity of the seawater source community was observed during coalescence. C_LIO_LIFull-length 16S rRNA gene profiling with a custom dual-barcoding Nanopore protocol enables cost-effective bacterial community tracking. C_LIO_LIControlled coalescence experiments offer mechanistic insights into estuarine microbial community transitions. C_LI

ecology↗

bioRxiv2025

Chemistry-based vectors map the chemical space of natural biomes from untargeted mass spectrometry data

Untargeted metabolomics can comprehensively map the chemical space of a biome, but is limited by low annotation rates (<10%). We used chemistry-based vectors, consisting of molecular fingerprints or chemical compound classes, predicted from mass spectrometry data, to characterize compounds and samples. These chemical characteristics vectors (CCVs) estimate the fraction of compounds with specific chemical properties in a sample. Unlike the aligned MS1 data with intensity information, CCVs incorporate actual chemical properties of compounds, offering deeper insights into sample comparisons. Thus, we identified key compound classes differentiating biomes, such as ethers which are enriched in environmental biomes, while steroids enriched in animal host-related biomes. In biomes with greater variability, CCVs revealed key clustering compound classes, such as organonitrogen compounds in animal distal gut and lipids in animal secretions. CCVs thus enhance the interpretation of untargeted metabolomic data, providing a quantifiable and generalizable understanding of the chemical space of natural biomes.

bioinformatics↗

bioRxiv2025

Deletion of the moe A gene in Flavobacterium IR1 drives structural color shift from green to blue and alters polysaccharide metabolism

Structural color (SC), generated by light interacting with nanostructured materials, are responsible for the brightest and most vivid coloration in nature. Despite being widespread within the tree of life, there is little knowledge of the genes involved. Partial exceptions are some colonies of Flavobacteriia in which genes involved in a number of pathways, including gliding motility and polysaccharide metabolism, have been linked to SC. A previous genomic analysis of SC and non-SC bacteria suggested that the pterin pathway is involved in the organization of bacteria to form SC. Thus here, we focus on the moeA molybdopterin molybdenum transferase. When this gene was deleted from Flavobacterium IR1, the knock- out mutant showed a strong blue shift in SC of the colony, different from the green SC of the wild-type. The moeA mutant showed a particularly strong blue shift when grown on kappa- carrageenan and was upregulated for starch degradation. To further analyze the molecular changes, proteomic analysis was performed, showing the upregulation of various polysaccharide utilization loci, which supported the link between moeA and polysaccharide metabolism in SC. Overall, we demonstrated that single-gene mutations could change the optical properties of bacterial SC, which is unprecedented when compared to multicellular organisms where structural color is the result of several genes and can not yet be addressed genetically.

microbiology↗

bioRxiv2024

Jaeger: an accurate and fast deep-learning tool to detect bacteriophage sequences

Viruses are integral to every biome on Earth, yet we still need a more comprehensive picture of their identity and global distribution. Global metagenomics sequencing efforts revealed the genomic content of tens of thousands of environmental samples, however identifying the viral sequences in these datasets remains challenging due to their vast genomic diversity. Here, we address identifying bacteriophage sequences in unlabeled sequencing data. In a recent benchmarking paper, we observed that existing deep-learning tools show a high true positive rate, but may also produce many false positives when confronted with divergent sequences. To tackle this challenge, we introduce Jaeger, a novel deep-learning method designed specifically for identifying bacteriophage genome fragments. Extensive benchmarking on the IMG/VR database and real-world metagenomes reveals Jaegers consistent high sensitivity (0.87) and precision (0.92). Applying Jaeger to over 16,000 metagenomic assemblies from the MGnify database yielded over five million putative phage contigs. On average, Jaeger is around 20 times faster than the other state-of-the-art methods. Jaeger is available at https://github.com/MGXlab/Jaeger.

bioinformatics↗

bioRxiv2024

Tunturi virus isolates and metagenome-assembled viral genomes provide insights into the virome of Acidobacteriota in Arctic tundra soils

BackgroundArctic soils are climate-critical areas, where microorganisms play crucial roles in nutrient cycling processes. Acidobacteriota are phylogenetically and physiologically diverse bacteria that are abundant and active in Arctic tundra soils. Still, surprisingly little is known about acidobacterial viruses in general and those residing in the Arctic in particular. Here, we applied both culture-dependent and -independent methods to study the virome of Acidobacteriota in Arctic soils. ResultsFive virus isolates, Tunturi 1-5, were obtained from Arctic tundra soils, Kilpisjarvi, Finland (69{degrees}N), using Tunturibacter spp. strains originating from the same area as hosts. The new virus isolates have tailed particles with podo-(Tunturi 1, 2, 3), sipho-(Tunturi 4), or myovirus-like (Tunturi 5) morphologies. The dsDNA genomes of the viral isolates are 63-98 kbp long, except Tunturi 5, which is a jumbo phage with a 309-kbp genome. Tunturi 1 and Tunturi 2 share 88% overall nucleotide identity, while the other three are not related to one another. For over half of the open reading frames in Tunturi genomes, no functions could be predicted. To further assess the Acidobacteriota-associated viral diversity in Kilpisjarvi soils, bulk metagenomes from the same soils were explored and a total of 1881 viral operational taxonomic units (vOTUs) were bioinformatically predicted. Almost all vOTUs (98%) were assigned to the class Caudoviricetes. For 125 vOTUs, including five (near-)complete ones, Acidobacteriota hosts were predicted. Acidobacteriota-linked vOTUs were abundant across sites, especially in fens. Terriglobia-associated proviruses were observed in Kilpisjarvi soils, being related to proviruses from distant soils and other biomes. Approximately genus- or higher-level similarities were found between Tunturi viruses, Kilpisjarvi vOTUs, and other soil vOTUs, suggesting some shared groups of Acidobacteriota viruses across soils. ConclusionsThis study provides acidobacterial virus isolates as laboratory models for future research and adds insights into the diversity of viral communities associated with Acidobacteriota in tundra soils. Predicted virus-host links and viral gene functions suggest various interactions between viruses and their host microorganisms. Largely unknown sequences in the isolates and metagenome-assembled viral genomes highlight a need for more extensive sampling of Arctic soils to better understand viral functions and contributions to ecosystem-wide cycling processes in the Arctic.

microbiology↗

bioRxiv2024

SpeSpeNet: An interactive and user-friendly tool to create and explore microbial correlation networks

Correlation networks are commonly used to explore microbiome data. In these networks, nodes are taxa and edges represent correlations between their abundance patterns across samples. As clusters of correlating taxa (co-abundance clusters) often indicate a shared response to environmental drivers, network visualization contributes to system understanding. Currently, most tools for creating and visualizing co-abundance networks from microbiome data either require the researcher to have coding skills, or they are not user-friendly, with high time expenditure and limited customizability. Furthermore, existing tools lack focus on the relationship between environmental drivers and the structure of the microbiome, even though many edges in correlation networks can be understood through a shared relationship of two taxa with the environment. For these reasons we developed SpeSpeNet (Species-Species Network, https://tbb.bio.uu.nl/SpeSpeNet), a practical and user-friendly R-shiny tool to construct and visualize correlation networks from taxonomic abundance tables. The details of data preprocessing, network construction, and visualization are automated, require no programming ability for the web version, and are highly customizable, including associations with user-provided environmental data. Here, we present the details of SpeSpeNet and demonstrate its utility using three case studies.

ecology↗

bioRxiv2024

Ultrafast and accurate sequence alignment and clustering of viral genomes

Viromics produces millions of viral genomes and fragments annually, overwhelming traditional sequence comparison methods. We introduce Vclust, a novel approach that determines average nucleotide identity by Lempel-Ziv parsing and clusters viral genomes with thresholds endorsed by authoritative viral genomics and taxonomy consortia. Vclust demonstrates superior accuracy and efficiency compared to existing tools, clustering millions of virus genomes in a few hours on a mid-range workstation.

bioinformatics↗

bioRxiv2024

High-resolution quantification of the rhizosphere effect along a soil-to-root gradient shows selection-driven convergence of rhizosphere microbiomes

Plants secrete a complex array of organic compounds, constituting about a third of their photosynthetic products, into the surrounding soil. As a result, concentration gradients are established from the roots into the bulk soil, known as the rhizosphere. Soil microbes benefit from these root exudates for their survival and propagation, and consequently, the composition of the rhizosphere microbial community follows the gradient of available compounds, a phenomenon oftentimes referred to as the rhizosphere effect. However, the fine-grained changes in the microbial community along this soil-root gradient have not been well described. Yet such insights would enable us to underpin the ecological rules underlying root microbial community assembly. Therefore, here we harvested the roots of individual Arabidopsis thaliana plants grown in three different natural soils at high-resolution, such that we could interrogate community assembly and predict microbial growth rate across consecutive, fine-grained, rhizosphere compartments. We found that the strength of the rhizosphere effect depends on root proximity and that microbial communities closer to the roots harbour related microbes. Closer to the roots, microbial community assembly became less random and more driven by selection-based processes. Intriguingly, we observed priority effects, where related microbes that arrive first are more likely to establish, and that microbes might use different ecological growth strategies to colonise the rhizosphere. All effects appeared to be independent from starting conditions as microbial community composition converged on the root despite different soil microbial seed banks. Together, our results provide a high-resolution view of the microbiome changes across the soil-root gradient.

plant biology↗

bioRxiv2024

Bacterial family-specific enrichment and functions of secretion systems in the rhizosphere

The plant rhizosphere is a highly selective environment where bacteria have developed traits to establish themselves or outcompete other microbes. These traits include bacterial secretion systems (SSs) that range from Type I (T1SS) to Type IX (T9SS) and can play diverse roles. The best known functions are to secrete various proteins or other compounds into the extracellular space or into neighbouring cells, including toxins to attack other microbes or effectors to suppress plant host immune responses. Here, we aimed to determine which bacterial SSs were associated with the plant rhizosphere. We utilised paired metagenomic datasets of rhizosphere and bulk soil samples from five different plant species grown in a wide variety of soil types, amounting to ten different studies. The T3SS and T6SS were generally enriched in the rhizosphere, as observed in studies of individual plant-associated genera. We also identified additional SSs that have received less attention thus far, such as the T2SS, T5SS and Bacteroidetes-specific T6SSiii and T9SS. The predicted secreted proteins of some of these systems (T3SS, T5SS and T6SS) could be linked to functions such as toxin secretion, adhesion to the host and facilitation of plant-host interactions (such as root penetration). The most prominent bacterial taxa with rhizosphere- or soil-enriched SSs included Xanthomonadaceae, Oxalobacteraceae, Comamonadaceae, Caulobacteraceae, and Chitinophagaceae, broadening the scope of known plant-associated taxa that use these systems. We anticipate that the SSs and taxa identified in this study may be utilised for the optimisation of bioinoculants to improve plant productivity.

microbiology↗

bioRxiv2023

Graphite: painting genomes using a colored De Bruijn graph

The recent growth of microbial sequence data allows comparisons at unprecedented scales, enabling tracking of strains, mobile genetic elements, or genes. Querying a genome against a large reference database can easily yield thousands of matches that are tedious to interpret and pose computational challenges. We developed Graphite that uses a colored De Bruijn graph (cDBG) to paint query genomes, selecting the local best matches along the full query length. By focusing on the closest genomic match of each query region, Graphite reduces the number of matches while providing promising leads for genomic forensics. When applied to hundreds of Campylobacter genomes we found extensive gene sharing, including a previously undetected C. coli plasmid that matched a C. jejuni chromosome. Together, genome painting using cDBGs as enabled by Graphite, can reveal new biological phenomena by mitigating computational hurdles. Graphite is implemented in Julia, available at https://github.com/MGXlab/Graphite.

bioinformatics↗

bioRxiv2023

Phylogeny and disease links of a widespread and ancient gut phage lineage

Viruses are a core component of the human microbiome, impacting health and disease through interactions with gut bacteria1 and the immune system2. Most viruses in the human microbiome are bacteriophages, which exclusively infect bacteria. Individual gut bacteriophages can affect bacterial bile acid deconjugation3, and can alter their infection strategy based on dietary content4. Up to recently, most studies of the gut virome have focused on low taxonomic scales (e.g., viral operational taxonomic units), hampering population-level analyses. We previously identified the expansive and widespread Candidatus Heliusviridae bacteriophage family in a cohort with inhabitants of Amsterdam, the Netherlands. Here, we study their biodiversity and evolution in a wide variety of human populations. With a detailed phylogeny based on sequences from six viral genome databases, we now propose the Candidatus order Heliusvirales to accommodate these viruses. We identify Ca. Heliusvirales viruses in 80% of 5,441 individuals across 39 studies, and also in nine out of thirteen analyzed metagenomes from ancient humans that lived in Europe and North America between 1,000 and 5,000 years ago. We show that a large Ca. Heliusvirales lineage has diversified starting at the appearance of Homo sapiens some 200,000-300,000 years ago. Ancient peoples and modern hunter-gatherers further have distinct Ca. Heliusvirales populations that are characterized by lower richness than modern urbanized people. Within urbanized people, those suffering from type 1 and type 2 diabetes, as well as inflammatory bowel disease, have higher Ca. Heliusvirales richness than healthy controls. We thus conclude that these ancient core members of the human gut virome have thrived with increasingly westernized lifestyles of the human population.

microbiology↗