Biology subjects

Conrad, R.

Publications and source records attributed to Conrad, R..

7 recordsLinked to original sources

bioRxiv2026

Reproducible Tools and Enhanced Computational Workflows for Batch Effect Evaluation of High-Throughput Data Using BatchQC

Batch effect correction is a common and often necessary step in data analysis to reduce bias due to technical and experimental factors when combining multiple batches of data. The severity of the batch effects dictates the correction strategy; therefore, a careful assessment of each datasets batch effects is necessary. BatchQC is an R package that provides reproducible tools and visualizations for quantitatively and qualitatively addressing batch effects across a broad range of data types. BatchQC integrates with standardized Bioconductor data structures and features an object-oriented design, enabling the application of workflows that can freely evaluate and process data within and outside the package tools. Common batch evaluation methods, along with novel quantitative metrics, help determine the benefits of batch correction for each dataset and enable direct comparisons between methods. Here, we present BatchQC as the first comprehensive batch-correction R package, with independent tools, reproducible workflows, visualization, and novel statistics.

bioinformatics↗

bioRxiv2025

Optimizing mouse metatranscriptome profiling by selective removal of redundant nucleic acid sequences

Metatranscriptome (MetaT) sequencing is a critical tool for profiling the dynamic metabolic functions of microbiomes. In addition to taxonomic information, MetaT also provides real-time gene expression data of both host and microbial populations, thus permitting authentic quantification of the functional (enzymatic) output of the microbiome and its host. The main challenge to effective and accurate MetaT analysis is the removal of highly abundant rRNA transcripts from these complex mixtures of microbes, which can number in the thousands of individual species. Regardless of methodology for rRNA depletion, the design of rRNA removal probes based solely upon taxonomic content of the microbiome typically requires very large numbers of individual probes, making this approach complex to commercially manufacture, costly, and frequently technically infeasible. In previous work [1], we designed a set of depletion probes for human stool samples using a design strategy based solely on sequence abundance, completely agnostic of the microbiomal species present. Here, we show that the human-based probes are less effective when used with mouse cecal samples. However, adapting additional rRNA depletion probes specifically to cecal content provides both greater efficiency and consistency for MetaT analysis of mouse samples. ImportanceSequencing total RNA from microbiome samples is seriously impaired by the overwhelming proportion of rRNA to mRNA content. As much as 99% of sequencing reads can be assigned to the rRNA content, thus removal of these abundant transcripts is critical to MetaT analysis. The use of Ribo Zero Plus rRNA depletion probes designed for human gut microbiomes proved to be less effective and more inconsistent across mouse cecal donor samples, a common experimental system for microbiome studies. In the present work, we have extended and refined a taxonomically-neutral probe design method for mouse cecal content. The additional probes were carefully chosen to limit the number needed for effective depletion to reduce both the cost and risk of introducing bias to MetaT analysis. Our results demonstrate this method as efficient and consistent for rRNA removal in mouse cecal samples, thus providing a significant increase in the number of mRNA-rich sequencing reads for MetaT analysis.

genomics↗

bioRxiv2024

Promiscuous and unbiased recombination underlies the sequence-discrete species of the SAR11 lineage in the deep ocean

Surveys of microbial communities (metagenomics) or isolate genomes have revealed sequence-discrete species. That is, members of the same species usually show >95% Average Nucleotide Identity (ANI) of shared genes among themselves vs. <83% ANI to members of other species while genome pairs showing between 83-95% ANI are comparatively rare. In these surveys, aquatic bacteria of the ubiquitous SAR11 clade (Class Alphaproteobacteria), which play a major role in carbon cycling, are an outlier and often do not exhibit discrete species boundaries, suggesting the potential for alternate modes of genetic differentiation. To explore evolution in SAR11, we analyzed high-quality, single-cell amplified genomes (SAGs) and companion metagenomes from an oxygen minimum zone (OMZ) in the Eastern Tropical Pacific Ocean, where the SAR11 make up [~]20% of the total microbial community. Our results show that SAR11 do form several sequence-discrete species, but their ANI range of discreteness is shifted to lower identities between 86-91%, with intra-species ANI ranging between 91-100%. Measuring recent gene exchange among these genomes based on a newly developed methodology revealed higher frequency of homologous recombination within compared to between species that affects sequence evolution at least twice as much as diversifying point mutation across the genome. Recombination in SAR-11 appears to be more promiscuous compared to other prokaryotic species and has facilitated the spreading of adaptive mutations within the species, further promoting the high intra-species diversity observed. These results implicate rampant, genome-wide homologous recombination as the mechanism that underlies the evolution of SAR11 into discrete species. SignificanceDistinguishing "species" is a pressing issue in microbiology, partly because the mechanisms that create and maintain clusters of genetically similar microbes are sparsely documented for natural populations. By leveraging high quality single-cell genomic data and a novel method for assessing homologous recombination, we show that rampant homologous recombination maintains species-level clusters of genomes for the most abundant order of marine bacteria, suggesting that these genomes may be evolving sexually to a much greater extent than previously thought. Therefore, our results identify a mechanism explaining the evolution of species in a major microbial group and have implications for understanding microbial diversity and the species concept more broadly.

microbiology↗

bioRxiv2024

Deep Learning Analysis on Images of iPSC-derived Motor Neurons Carrying fALS-genetics Reveals Disease-Relevant Phenotypes

Amyotrophic lateral sclerosis (ALS) is a devastating condition with very limited treatment options. It is a heterogeneous disease with complex genetics and unclear etiology, making the discovery of disease-modifying interventions very challenging. To discover novel mechanisms underlying ALS, we leverage a unique platform that combines isogenic, induced pluripotent stem cell (iPSC)-derived models of disease-causing mutations with rich phenotyping via high-content imaging and deep learning models. We introduced eight mutations that cause familial ALS (fALS) into multiple donor iPSC lines, and differentiated them into motor neurons to create multiple isogenic pairs of healthy (wild-type) and sick (mutant) motor neurons. We collected extensive high-content imaging data and used machine learning (ML) to process the images, segment the cells, and learn phenotypes. Self-supervised ML was used to create a concise embedding that captured significant, ALS-relevant biological information in these images. We demonstrate that ML models trained on core cell morphology alone can accurately predict TDP-43 mislocalization, a known phenotypic feature related to ALS. In addition, we were able to impute RNA expression from these image embeddings, in a way that elucidates molecular differences between mutants and wild-type cells. Finally, predictors leveraging these embeddings are able to distinguish between mutant and wild-type both within and across donors, defining cellular, ML-derived disease models for diverse fALS mutations. These disease models are the foundation for a novel screening approach to discover disease-modifying targets for familial ALS.

neuroscience↗

bioRxiv2023

How formate dehydrogenase-lacking acetogen Clostridium bovifaecis utilize formate as the sole carbon source for its acetogenic growth?

Little is known about the growth of formate dehydrogenase-lacking acetogen on formate as sole carbon source. Here, we analyzed formate metabolism in Clostridium bovifaecis strain BXX using different concentrations of formate. The results show that C. bovifaecis converted formate (11.5-96 mM) into acetate with molar ratio of 2.0:1[~]2.6:1 by using L-cysteine in the anaerobic medium as electron source according to the stoichiometry of acetogenesis. Genome analysis of C. bovifaecis revealed genes encoding anaerobic ribonucleoside triphosphate reductase (nrdD and nrdG) catalyzing the oxidation of formate to CO2 while ATP is being reduced to the desoxy form. The existence of nrdD was verified by PCR, reverse transcription-PCR analysis and acetogenesis from formate. The process mode of acetogenesis from formate in C. bovifaecis provides insight into the unique metabolic feature of an FDH-lacking acetogen. IMPORTANCEWood-Ljungdahl pathway (WLP) lacking formate dehydrogenase (FDH) which catalyzes CO2 reduction to formate has been reported to occur acetogenesis only in the presence of formate and exogenous CO2, which seems to result from the formate-rich habitats adaptation of gastrointestinal acetogens. Here, we found FDH-lacking Clostridium bovifaecis strain BXX converted formate (11.5-96 mM) into acetate with molar ratio of 2.0:1[~]2.6:1 fitting the stoichiometry of acetogenesis when using formate as the sole carbon source. CO2 needed in the carbonyl branch of WLP was from the oxidation of formate to CO2 catalyzed by anaerobic ribonucleoside triphosphate reductase while ATP is being reduced to the desoxy form. L-cysteine in the anaerobic medium was the electron source of WLP. The process mode of acetogenesis from formate in C. bovifaecis provides insight into how an FDH-lacking acetogen can make a living from the simplest resources as carbon source, which has both ecological and biotechnological significance.

microbiology↗

bioRxiv2020

Reconstructing genomes of carbon monoxide oxidisers in volcanic deposits including members of the class Ktedonobacteria

Microorganisms can potentially colonize volcanic rocks using the chemical energy in reduced gases such as methane, hydrogen (H2) and carbon monoxide (CO). In this study, we analysed soil metagenomes from Chilean volcanic soils, representing three different successional stages with ages of 380, 269 and 63 years, respectively. A total of 19 metagenome-assembled genomes (MAGs) were retrieved from all stages with a higher number observed in the youngest soil (1640: 2 MAGs, 1751: 1 MAG, 1957: 16 MAGs). Genomic similarity indices showed that several MAGs had amino-acid identity (AAI) values >50% to the phyla Actinobacteria, Acidobacteria, Gemmatimonadetes, Proteobacteria and Chloroflexi. Three MAGs from the youngest site (1957) belonged to the class Ktedonobacteria (Chloroflexi). Complete cellular functions of all the MAGs were characterised, including carbon fixation, terpenoid backbone biosynthesis, formate oxidation and CO oxidation. All 19 environmental genomes contained at least one gene encoding a putative carbon monoxide dehydrogenase (CODH). Three MAGs had form I coxL operon (encoding the large subunit CO-dehydrogenase). One of these MAGs (MAG-1957-2.1, Ktedonobacterales) was highly abundant in the youngest soil. MAG-1957-2.1 also contained genes encoding a [NiFe]-hydrogenase and hyp genes encoding accessory enzymes and proteins. Little is known about the Ktedonobacterales through cultivated isolates, but some species can utilize H2 and CO for growth. Our results strongly suggest that the remote volcanic sites in Chile represent a natural habitat for Ktedonobacteria and they may use reduced gases for growth.

microbiology↗

bioRxiv2020

Cooperation between CRISPR-Cas types enables adaptation in an RNA-targeting system

CRISPR-Cas immune systems adapt to new threats by acquiring spacers from invading nucleic acids such as phage genomes. However, some CRISPR-Cas loci lack genes necessary for spacer acquisition, despite apparent variation in spacer content between strains. It has been suggested that such loci may use acquisition machinery from co-occurring CRISPR-Cas systems. Here, using a lytic dsDNA phage, we observe spacer acquisition in the native host Flavobacterium columnare that carries an acquisition-deficient subtype VI-B locus and a complete subtype II-C locus. We characterize acquisition events in both loci and show that the RNA-targeting VI-B locus acquires spacers in trans using acquisition machinery from the DNA-targeting II-C locus. Our observations reinforce the concept of modularity in CRISPR-Cas systems and raise further questions regarding plasticity of adaptation modules.

microbiology↗