Biology subjects

Ferrell, B. D.

Publications and source records attributed to Ferrell, B. D..

10 recordsLinked to original sources

bioRxiv2026

Single amino acid substitution in DNA Polymerase I dramatically alters infection dynamics of bacteriophage T7

Viruses constitute a significant proportion of Earths genetic diversity, yet most remain uncharacterized beyond their sequences in viral metagenomes. Linking viral genotypes to phenotypes--especially enzyme function to phage infection dynamics--is challenging due to the lack of cultured virus-host systems. DNA polymerase I (PolA), essential for genome replication in [~]25% of dsDNA phages, provides an opportunity to explore these connections. In phage T7, residue 526 is critical for nucleotide incorporation, with previous in vitro evidence indicating impacts on enzyme efficiency and fidelity. Previous analyses identified three substitutions at this position (Tyr/Y, Phe/F, Leu/L) linked with deeply rooted viral PolA clades. Mutation impacts at residue 526 were tested in vitro and in vivo. The Y526F protein exhibited a 50% reduction in specific activity, and when introduced via High Complexity Golden Gate Assembly into T7 demonstrated a 53% decrease in burst size and significantly longer latent period compared to wild type. The Y526L protein exhibited a 97% decrease in activity, and the Y526L phage was incapable of completing its lifecycle. These findings confirm historical biochemical data, provide in vivo context for these mutations in the T7-E. coli system, and offer experimental support for genotype-to-phenotype associations in viral PolA, informing viral metagenomics studies. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=162 SRC="FIGDIR/small/726624v1_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@14395e5org.highwire.dtl.DTLVardef@261504org.highwire.dtl.DTLVardef@2dc1e4org.highwire.dtl.DTLVardef@147a7f_HPS_FORMAT_FIGEXP M_FIG C_FIG Created in BioRender. Keown, R. (2026) https://BioRender.com/mhrmup3

microbiology↗

bioRxiv2026

Ribonucleotide reductases recapitulate biogeographic patterns within virioplankton according to ocean biogeochemistry

Many of the important biogeochemical implications of marine viruses focus on the activity of lytic viruses. It is generally thought that lytic viral activity contributes to maintaining bioavailable nutrient pools and controlling host community composition through specific infection and lysis processes. Among lytic viruses, genome replication is under stringent selection pressure as advantages in replication speed and fidelity can influence viral fitness. Deoxyribonucleotides (dNTPs) are the precursor substrate for DNA synthesis and ribonucleotide reductase (RNR) is the only known enzyme capable of reducing ribonucleotides into dNTPs. Thus, for a virus, encoding an RNR gene provides control over dNTP production. Overall, distributional patterns of the RNR-encoding virioplankton community mirrored that seen for total virioplankton throughout the global ocean. A majority of RNR-encoding virioplankton ([~]66%) carried the Class II enzyme and most of these carried the monomeric (NrdJm) gene. This was significant as NrdJm utilizes triphosphate ribonucleotides as opposed to the diphosphate ribonucleotides used by Class I and Class II dimeric RNRs. The distribution and frequency of Class I RNRs followed the concentration of required iron and manganese co-factors. Oceanic virioplankton encompassed a high diversity of Class I and II RNRs with the discovery of new phylogenetic clades not previously observed among viruses or within cellular life. Representing [~]10% of all virioplankton populations, RNR-encoding viral populations demonstrated surprising fidelity with the major biogeographical features defining oceanic ecosystems.

microbiology↗

bioRxiv2025

Comparative Analysis Reveals Host Species-Dependent Diversity Among 16 Virulent Bacteriophages Isolated Against Soybean Bradyrhizobium spp.

Phages play a role in shaping ecosystems by controlling host abundance via cell lysis, driving host evolution via horizontal gene transfer, and promoting nutrient cycling. The genus Bradyrhizobium includes bacteria able to symbiotically nodulate the roots of soybean (Glycine max), providing the plant with a direct source of biologically fixed nitrogen. Optimizing this symbiosis can minimize the use of nitrogen fertilizers and make soybean production more sustainable. Phages targeting Bradyrhizobium may modify their hosts genotype, alter phenotypic traits such as symbiotic effectiveness, and mediate competition among strains for nodulation sites. Sixteen phages were isolated against B. elkanii strains USDA94 and USDA31, and B. diazoefficiens strain USDA110. Comparative analyses revealed host species-dependent diversity in morphology, host range, and genome composition, leading to the identification of three previously undescribed phage species. Remarkably, all B. elkanii phages shared a siphophage morphology and formed a single species with >97% nucleotide identity, even when isolated from farms separated by up to [~]70 km, suggesting genomic stability across geographic scales. In contrast, phages isolated against B. diazoefficiens displayed podophage-like morphology, greater genetic diversity, and divided into two distinct species. Although no phages were recovered against B. japonicum strains or native Delaware Bradyrhizobium isolates tested, some Delaware isolates showed susceptibility during the host range assay. The phage genomes demonstrated features predicting phenotypes. Terminase genes predicted headful packaging among the phages which is critical for generalized transduction. The B. elkanii phages all carried tmRNA genes capable of recruiting stalled ribosomes and both phage groups carried DNA polymerase A indicating greater control of phage genome replication. State-of-the-art structural annotation revealed a tail fiber gene within a phage genome having the highest proportion (80.77%) of unknown genes. Together this work expands the limited knowledge available on soybean Bradyrhizobium phage ecology and genomics.

microbiology↗

bioRxiv2025

EXTENDING PROTEIN LANGUAGE MODELS TO A VIRAL GENOMIC SCALE USING BIOLOGICALLY INDUCED SPARSE ATTENTION

The transformer architecture in deep learning has revolutionized protein sequence analysis. Recent advancements in protein language models have paved the way for significant progress across various domains, including protein function and structure prediction, multiple sequence alignments and mutation effect prediction. A protein language model is commonly trained on individual proteins, ignoring the interdependencies between sequences within a genome. However, biological understanding reveals that protein-protein interactions span entire genomic regions, underscoring the limitations of focusing solely on individual proteins. To address these limitations, we propose a novel approach that extends the context size of transformer models across the entire viral genome. By training on large genomic fragments, our method captures long-range interprotein interactions and encodes protein sequences with integrated information from distant proteins within the same genome, offering substantial benefits in various tasks. Viruses, with their densely packed genomes, minimal intergenic regions, and protein annotation challenges, are ideal candidates for genome-wide learning. We introduce a long-context protein language model, trained on entire viral genomes, leveraging a sparse attention mechanism based on protein-protein interactions. Our semi-supervised approach supports long sequences of up to 61,000 amino acids (aa). Our evaluations demonstrate that the resulting embeddings significantly surpass those generated by single-protein models and outperform alternative large-context architectures that rely on static masking or non-transformer frameworks.

bioinformatics↗

bioRxiv2025

Fine-Tuning Protein Language Models Unlocks the Potential of Underrepresented Viral Proteomes

Protein language models (pLMs) have revolutionized computational biology by generating rich protein vector representations--or embeddings--enabling major advancements in de novo protein design, structure prediction, variant effect analysis, and evolutionary studies. Despite these breakthroughs, current pLMs often exhibit biases against proteins from underrepresented species, with viral proteins being particularly affected--frequently referred to as the "dark matter" of the biological world due to their vast diversity and ubiquity, yet sparse representation in training datasets. Here, we show that fine-tuning pre-trained pLMs on viral protein sequences, using diverse learning frameworks and parameter-efficient strategies, significantly enhances representation quality and improves performance on downstream tasks. To support further research, we provide source code for fine-tuning pLMs and benchmarking embedding quality. By enabling more accurate modeling of viral proteins, our approach advances tools for understanding viral biology, combating emerging infectious diseases, and driving biotechnological innovation.

bioinformatics↗

bioRxiv2023

B12-dependent virioplankton demonstrate interseasonal dynamics and associate with a diversity of pelagic bacterioplankton

Through infection and lysis of their coexisting bacterial hosts, viruses impact the biogeochemical cycles sustaining globally significant pelagic oceanic ecosystems. Currently, little is known of the ecological interactions between lytic viruses and their bacterial hosts underlying these biogeochemical impacts at ecosystem scales. This study focused on populations of lytic viruses carrying the B12- dependent Class II monomeric ribonucleotide reductase (RNR) gene, ribonucleotide triphosphate reductase (RTPR), documenting seasonal changes in pelagic virioplankton and bacterioplankton using amplicon sequences of RTPR and the 16S rRNA gene, respectively. Amplicon sequence libraries were analyzed using compositional data analysis tools that account for the compositional nature of these data. Both virio- and bacterioplankton communities responded to environmental changes typically seen across seasonal cycles as well as shorter term upwelling-downwelling events. Defining RTPR-carrying viral populations according to major phylogenetic clades proved a more robust means of exploring virioplankton ecology than operational taxonomic units defined by percent sequence homology. Virioplankton RTPR populations showed positive associations with a broad phylogenetic diversity of bacterioplankton including dominant taxa within pelagic oceanic ecosystems such as Prochlorococcus and SAR11. Temporal changes in RTPR-virioplankton, occurring as both free viruses and within infected cells, indicated possible viral-host pairs undergoing sustained infection and lysis cycles throughout the seasonal study. Phylogenetic relationships inferred from RTPR sequences mirrored ecological patterns in virio- and bacterioplankton populations demonstrating possible genome to phenome associations for an essential viral replication gene.

ecology↗

bioRxiv2022

Spontaneously produced lysogenic phages are an important component of the soybean Bradyrhizobium mobilome

The ability to nodulate and fix atmospheric nitrogen in soybean root nodules makes soybean Bradyrhizobium spp. (SB) critical in supplying humanitys nutritional needs. The intricacies of SB-plant interactions have been studied extensively; however, bradyrhizobial ecology as influenced by phages has received somewhat less attention even though these interactions may significantly impact soybean yield. In batch culture four SB strains, S06B (B. japonicum, S06B-Bj), S10J (B. japonicum, S10J-Bj), USDA 122 (B. diazoefficiens, USDA 122-Bd), and USDA 76T (B. elkanii, USDA 76-Be), spontaneously (without apparent exogenous chemical or physical induction) produced phages throughout the growth cycle; for three strains, phage concentrations exceeded cell numbers by ca. 3-fold after 48 h incubation. Observed spontaneously produced phages (SPP) were tailed. Phage terminase large-subunit protein phylogeny revealed possible differences in phage packaging and replication mechanisms. Bioinformatic analyses predicted multiple prophage regions within each SB genome preventing accurate identification of SPP genomes. A DNA sequencing approach was developed that accurately delineated the boundaries of four SPP genomes within three of the SB chromosomes. Read mapping suggested that the SPP are capable of transduction. In addition to the phages, bacterial strains S06B-Bj and USDA 76-Be were rich in mobile elements consisting of insertion sequences (IS) and large, conjugable, broad host range plasmids. The prevalence of SPP along with IS and plasmids indicate that horizontal gene transfer likely plays an outsized role in SB ecology and may subsequently impact soybean productivity. ImportancePrevious studies have shown that IS and plasmids mediate horizontal gene transfer (HGT) of symbiotic nodulation (nod) genes in SB; however, these events require close cell to cell contact which could be limited in soil environments. Bacteriophage assisted gene transduction through spontaneously produced prophages could provide stable means of HGT not limited by the constraints of proximal cell to cell contact. Phage mediated HGT events could be important in SB population ecology with concomitant impacts on soybean agriculture.

microbiology↗

bioRxiv2021

PASV: Automatic protein partitioning and validation using conserved residues

BackgroundIncreasingly, researchers use protein-coding genes from targeted PCR amplification or direct metagenomic sequencing in community and population ecology. Analysis of protein-coding genes presents different challenges from those encountered in traditional SSU rRNA studies. Most protein-coding sequences are annotated based on homology to other computationally-annotated sequences, which can lead to inaccurate annotations. Therefore, the results of sensitive homology searches must be validated to remove false-positives and assess functionality. Multiple lines of in silico evidence can be gathered by examining conserved domains and residues identified through biochemical investigations. However, manually validating sequences in this way can be time consuming and error prone, especially in large environmental studies. ResultsAn automated pipeline for protein active site validation (PASV) was developed to improve validation and partitioning accuracy for protein-coding sequences, combining multiple sequence alignment with expert domain knowledge. PASV was tested using commonly misannotated proteins: ribonucleotide reductase (RNR), alternative oxidase (AOX), and plastid terminal oxidase (PTOX). PASV partitioned 9,906 putative Class I alpha and Class II RNR sequences from bycatch in a global viral metagenomic investigation with >99% true positive and true negative rates. PASV predicted the class of 2,579 RNR sequences in >98% agreement with manual annotations. PASV correctly partitioned all 336 tested AOX and PTOX sequences. ConclusionsPASV provides an automated and accurate way to address post-homology search validation and partitioning of protein-coding marker genes. Source code is released under the MIT license and is found with documentation and usage examples on GitHub at https://github.com/mooreryan/pasv.

bioinformatics↗

bioRxiv2018

CRISPR spacers indicate preferential matching of specific virioplankton genes

Viral infection exerts selection pressure on marine microbes as viral-induced cell lysis causes 20 to 50% of cell mortality resulting in fluxes of biomass into oceanic dissolved organic matter. Archaeal and bacterial populations can defend against viral infection using the CRISPR-Cas system which relies on specific matching between a spacer sequence and a viral gene. If a CRISPR spacer match to any gene within a viral genome is equally effective in preventing lysis, then no viral genes should be preferentially matched by CRISPR spacers. However, if there are differences in effectiveness then certain viral genes may demonstrate a greater frequency of CRISPR spacer matches. Indeed, homology search analyses of bacterioplankton CRISPR spacer sequences against virioplankton sequences revealed preferential matching of replication proteins, nucleic acid binding proteins, and viral structural proteins. Positive selection pressure for effective viral defense is one parsimonious explanation for these observations. CRISPR spacers from virioplankton metagenomes preferentially matched methyltransferase and phage integrase genes within virioplankton sequences. These viriolankton CRISPR spacers may assist infected host cells in defending against competing phage. Analyses also revealed that half of the spacer-matched viral genes were unknown and that some genes matched several spacers and some spacers matched multiple genes, a many-to-many relationship. Thus, CRISPR spacer matching may be an evolutionary algorithm, agnostically identifying those genes under stringent selection pressure for sustaining viral infection and lysis. Investigating this subset of viral genes could reveal those genetic mechanisms essential to viral-host interactions and provide new technologies for optimizing CRISPR defense in beneficial microbes.

microbiology↗

bioRxiv2018

Fast and sensitive protein sequence homology searches using hierarchical cluster BLAST

The throughput of DNA sequencing continues to increase, allowing researchers to analyze genomes of interest at greater depths. An unintended consequence of this data deluge is the increased cost of analyzing these datasets. As a result, genome and metagenome annotation pipelines are left with a few options: (i) search against smaller reference databases, (ii) use faster, but less sensitive, algorithms to assess sequence similarities, or (iii) invest in computing hardware specifically designed to improve BLAST searches such as GPGPU systems and/or large CPU-rich clusters.\n\nWe present a pipeline that improves the speed of amino acid sequence homology searches with a minimal decrease in sensitivity and specificity by searching against hierarchical clusters. Briefly, the pipeline requires two homology searches: the first search is against a clustered version of the database and the second is against sequences belonging to clusters with a hit from the first search. We tested this method using two assembled viral metagenomes and three databases (Swiss-Prot, Metagenomes Online, and UniRef100). Hierarchical cluster homology searching proved to be 12-times faster than BLASTp and produced alignments that were nearly identical to BLASTp (precision=0.99; recall=0.97). This approach is ideal when searching large collections of sequences against large databases.

bioinformatics↗