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Astrocytic CREB in nucleus accumbens promotes susceptibility to chronic stress

BackgroundIncreasing evidence implicates astrocytes in stress and depression in both rodent models and human Major Depressive Disorder (MDD). Despite this, little is known about the transcriptional responses to stress of astrocytes within the nucleus accumbens (NAc), a key brain reward region, and their influence on behavioral outcomes. MethodsWe used whole cell sorting, RNA-sequencing, and bioinformatic analyses to investigate the NAc astrocyte transcriptome in male mice in response to chronic social defeat stress (CSDS). Immunohistochemistry was used to determine stress-induced changes in astrocytic CREB within the NAc. Finally, astrocytic regulation of depression-like behavior was investigated using viral-mediated manipulation of CREB in combination with CSDS. ResultsWe found a robust transcriptional response in NAc astrocytes to CSDS in stressed mice, with changes seen in both stress-susceptible and stress-resilient animals. Bioinformatic analysis revealed CREB, a transcription factor widely studied in neurons, as one of the top-predicted upstream regulators of the NAc astrocyte transcriptome, with opposite activation states seen in resilient versus susceptible mice. This bioinformatic result was confirmed at the protein level with immunohistochemistry. Viral overexpression of CREB selectively in NAc astrocytes promoted susceptibility to chronic stress. ConclusionsTogether, our data demonstrate that the astrocyte transcriptome responds robustly to CSDS and, for the first time, that transcriptional regulation in astrocytes contributes to depressive-like behaviors. A better understanding of transcriptional regulation in astrocytes may reveal unknown molecular mechanisms underlying neuropsychiatric disorders.

neuroscience↗

Critical analysis of polycyclic tetramate macrolactam biosynthetic cluster phylogeny and functional diversity

Polycyclic tetramate macrolactams (PTMs) are bioactive natural products commonly associated with certain actinobacterial and proteobacterial lineages. These molecules have been the subject of numerous structure-activity investigations since the 1970s. New members continue to be pursued in wild and engineered bacterial strains, and advances in PTM biosynthesis suggests their outwardly simplistic biosynthetic gene clusters (BGCs) belie unexpected product complexity. Towards addressing the origins of this complexity and understanding its influence on PTM discovery, we engaged in a combination of bioinformatics to systematically classify PTM BGCs, and PTM-targeted metabolomics to compare the products of select BGC types. By comparing groups of producers and BGC mutants, we exposed knowledge gaps that complicate bioinformatics-driven product predictions. In sum, we provide new insights into the evolution of PTM BGCs while systematically accounting for the PTMs discovered thus far. The combined computational and metabologenomic findings presented here should prove useful for guiding future discovery. IMPORTANCEPolycyclic tetramate macrolactam (PTM) pathways are frequently found within the genomes of biotechnologically-important bacteria, including Streptomyces and Lysobacter spp. Their molecular products are typically bioactive, having substantial agricultural and therapeutic interest. Leveraging bacterial genomics for the discovery of new related molecules is thus desirable, but drawing accurate structural predictions from bioinformatics alone remains challenging. This difficulty stems from a combination of previously underappreciated biosynthetic complexity and remaining knowledge gaps, compounded by a stream of yet-uncharacterized PTM biosynthetic loci gleaned from recently sequenced bacterial genomes. We engaged in the following study to create a useful framework for cataloging historic PTM clusters, identifying new cluster variations, and tracing evolutionary paths for these molecules. Our data suggests new PTM chemistry remains discoverable in nature. However, our metabolomic and mutational analyses emphasize practical limitations to genomics-based discovery by exposing hidden complexity.

microbiology↗

Unravelling the ancient fungal DNA from the Iceman's gut

Here, we explore the possible ancient fungal species in the gut of Otzi, the Iceman, a naturally mummified human found in the Tyrolean Alps (border between Italy and Austria). While ancient DNA (aDNA) has been extensively used to study human, animal, and plant evolution, this research focuses on ancient microbial diversity, specifically fungi. Fungal DNA is often underestimated in metagenomic samples, however here we hypothesise the possibility of retrieving ancient fungal sequences from Otzis gut. A robust bioinformatic pipeline has been developed to detect and authenticate fungal aDNA from stomach, small intestine, and large intestine samples. We revealed the presence of Pseudogymnoascus genus, with P. destructans and P. verrucosus as possible species, that were particularly abundant in the stomach and small intestine. We suggest that Otzi may have consumed these fungi accidentally, likely in association with other elements of his diet, and they thrived in his gut after his death due to their adaptability to harsh and cold environments. This research provides insight into the coexistence of ancient humans with specific fungal species and proposes and validates a conservative bioinformatic approach for detecting fungal aDNA in historical metagenomic samples. Significance statementDespite their essential interactions with all kingdoms of life, limited molecular studies have focused on ancient fungi. Here, we developed a thorough bioinformatic pipeline that allowed us to detect the presence of ancient DNA likely belonging to Pseudogymnoascus destructans and P. verrucosus in the gut of Otzi, a human naturally mummified over 3,000 years ago in the Tyrolean Alps. Both species can survive harsh environmental conditions, and P. destructans is known for its pathogenicity, suggesting that Otzi may have accidentally ingested them and providing valuable insights into how ancient humans coexisted with specific fungal species. We propose a highly reliable methodology for detecting ancient fungal DNA in metagenomic studies of historical samples that can have broader applications to understand ancient ecosystems and their interactions.

microbiology↗

Uncovering the class II-bacteriocin predatiome in salivarius streptococci

Facing the surge of antibiotic resistance, the medical field has a critical need for alternatives to treat bacterial infections. Among these, the use of bacteriocins, ribosomally-synthesized antimicrobial peptides produced by bacteria, is considered to be a promising route. In the human commensal Streptococcus salivarius, the production of unmodified class II bacteriocins is directly controlled at the transcriptional level by the quorum-sensing ComRS system. Here, we used an integrated approach combining bioinformatics and synthetic biology to identify novel bacteriocins from salivarius streptococci active against human pathogens. We developed a bioinformatic pipeline that combines conservation of DNA motifs for genetic regulation and features of bacteriocin primary sequences to uncover cryptic class II bacteriocins. Notably, we discovered more than 50 novel bacteriocin candidates clustered into 21 groups from 100 genomes of S. salivarius. Strain-based analysis of bacteriocin cocktails revealed an important diversity restricted to seven distinct loci, probably resulting from bacteriocin intra- and inter- species exchanges. Using in vitro or in vivo production and synthetic biology tools, we showed that most of them are active against a panel of Gram-positive bacteria, including clinically- relevant pathogens. Overall, this work provides a new search-to-test generic pipeline for the discovery of novel bacteriocins in Gram-positive bacteria that could be used in cocktails for broad applications in the food and biomedical fields. IMPORTANCETo survive in highly challenging environments, streptococci have evolved a competence- predation coupling mechanism for genome plasticity. This developmental process is highly regulated at the transcriptional level, masking the predation killing effects in usual laboratory conditions. Here, we present a general strategy that combines bioinformatics and synthetic biology to unveil class II bacteriocins in streptococci. Its implementation to the beneficial species Streptococcus salivarius revealed around 40 class II salivaricin cocktails explained by the plasticity of seven independent loci. Notably, the salivaricin predatiome includes a subtle blend of fratricins, sobrinicins, and broad-spectrum bacteriocins with overlapping activities against a wide spectrum of low-GC Gram-positive bacteria, including notorious pathogens. Furthermore, most of those bacteriocins are predicted to be variants of a common -hairpin structure, indicating that their mode of action evolved convergently. Finally, the discovery of ca. 50 novel bacteriocins offers perspectives for the rational assembly of potent cocktails active against pathogenic staphylococci, streptococci or enterococci.

microbiology↗

Evaluation of false positive and false negative errors in targeted next generation sequencing

BackgroundAlthough next generation sequencing (NGS) has been adopted as an essential diagnostic tool in various diseases, NGS errors have been the most serious problem in clinical implementation. Especially in cancers, low level mutations have not been easy to analyze, due to the contaminating normal cells and tumor heterozygosity. ResultsIn targeted NGS (T-NGS) analyses for reference-standard samples containing mixtures of homozygote H. mole DNA with blood genomic DNA at various ratios from four certified NGS service providers, large differences in the lower detection limit of variants (16.3 times, 1.51[~]24.66%) and the false positive (FP) error rate (4280 times, 5.814 x 10-4 [~]1.359 x 10-7) were found. Employment of the commercially available Dragen system for bioinformatic analyses reduced FP errors in the results from companies BB and CC, but the errors originating from the NGS raw data persisted. Bioinformatic conditional adjustment to increase sensitivity (less than 2 times) led to a much higher FP error rate (610[~]8200 times). In addition, problems such as biased preferential reference base calls during bioinformatic analysis and high-rate FN errors in HLA regions were found in the NGS analysis. ConclusionT-NGS results from certified NGS service providers can be quite various in their sensitivity and FP error rate, suggesting the necessity of further quality controls for clinical implementation of T-NGS. The present study also suggests that mixtures of homozygote and heterozygote DNAs can be easily employed as excellent reference-standard materials for quality control of T-NGS.

genomics↗

Evaluation of long-read sequencing for Ostreid herpesvirus type 1 genome characterization from Magallana gigas infected tissues

2Since the 1990s, the Pacific oyster Magallana gigas has faced significant mortality, which has been associated with the detection of the Ostreid Herpesvirus type 1 (OsHV-1). Due to the complex genomic architecture and the presence of multiple genomic isomers, short-read sequencing using Illumina method struggles to accurately assemble tandem and repeat regions and to identify and characterize large structural variations in the OsHV-1 genome. Third-generation sequencing technologies, as long-read real-time nanopore sequencing from Oxford Nanopore Technologies (ONT), offer new possibilities for OsHV-1 whole genome analysis. Identification of the best method for extraction of high molecular weight (HMW) DNA and development of accurate bioinformatic pipelines for its characterization are now required. To this end, we evaluated and compared six HWM methods and one conventional DNA extraction kit for their ability to extract OsHV-1 DNA from M. gigas- infected tissues. We then evaluated the ability of ONT sequencing to produce an accurate OsHV-1 genome from both whole genome and "adaptive sampling" (AS) sequencing approaches. Finally, we evaluated the efficiency of bioinformatics tools for de novo assembly and consensus calling to generate accurate OsHV-1 genomes. The HMW DNA extraction kit coupled with ONT sequencing and dedicated bioinformatics tools allowed us to produce accurate OsHV-1 genomes compared to those assembled using Illumina technology. The AS approach allowed up to 60% enrichment for viral data, and the long reads generated by ONT allowed the characterization of OsHV-1 isomers. Together with its portability, this sequencing shows great promise as a diagnostic tool for the characterization of unculturable aquatic viruses directly from host tissues. 3 ImportanceMany aquatic viruses threaten commercially valuable species and cause significant economic losses during outbreaks. To improve our understanding of the origin, transmission patterns and spread of these viruses, additional genomic data are essential. However, genomic characterization of unculturable large DNA viruses is a major challenge. In the present study, we have successfully evaluated the ability of ONT sequencing and adaptive sequencing (AS) to sequence and assemble the complete OsHV-1 genome. Our results show that it is now possible to sequence the whole genome of large DNA viruses directly from infected host tissue, without the need for prior in vitro propagation or prior laboratory steps for virus enrichment.

genomics↗

The challenge of chromatin model comparison and validation a project from the first international 4D Nucleome Hackathon

The computational modeling of chromatin structure is highly complex and challenging due to the hierarchical organization of chromatin, which reflects its diverse biophysical principles, as well as inherent dynamism, which underlies its complexity. The variety of methods for chromatin structure modeling, which are based on different approaches, assumptions and scales of modeling, suggests that there is a necessity for a comprehensive benchmark. This inspired us to conduct a project at the NIH-funded 4D Nucleome Hackathon on March 18-21, 2024 at The University of Washington in Seattle, USA. The hackathon provided an amazing opportunity to gather an international, multi-institutional and unbiased group of experts to discuss, understand and undertake the challenges of chromatin model comparison and validation. These challenges seem straightforward in theory, however in practice, they are challenging and ambiguous. To address them, we developed a bioinformatics workflow for chromatin model comparison and validation, in which we use distance matrices to represent chromatin models, and we calculate Spearman correlation coefficients between pairs of matrices to estimate correlations between models, as well as between models and experimental data. During the 4-day hackathon, we tested our workflow on several distinct software packages for chromatin structure modeling and we discovered several challenges that include: 1) different aspects of chromatin biophysics and scales complicate model comparisons, 2) expertise in biology, bioinformatics, and physics is necessary to conduct a comprehensive research on chromatin structure, 3) bioinformatic software, which is often developed in academic settings, is characterized by insufficient support and documentation. Therefore, our work constitutes a way to advance the modeling of the 3D organization of the human genome, while emphasizing the importance of establishing guidelines for software development and standardization.

genomics↗

Connectivity Map and perturbation-based sensitivity analysis identifies MEK inhibitors as senolytics in human lung fibroblasts

Recently, the elimination of the disease-associated accumulation of senescent cells using senolytics has been shown to exert health benefits in animal studies. However, due to the heterogeneity of cell senescence and its unrecognized master regulators, drug development faces a complexity that must be handled. Bioinformatic elucidation of genes and pathways involved in senolysis and prediction of senolytic activity of compounds can cut costs and facilitate faster achievements in the field. In the present investigation, after obtaining the consensus gene signature of senescent fibroblasts of lung origin and deriving its anti-apoptotic module, we utilized Connectivity Map (CMap) alongside small molecule and genetic perturbation sensitivity data in cancer cell lines to identify drugs and genetic interventions that might induce apoptosis or sensitize senescent cells to apoptosis. Through bioinformatic evaluations, we speculate that activation of early stages of autophagy which contributes to the formation of autophagosomes, concurrent with the activation of waste protein concealment system by the mean of p62 and chaperoning system alongside an increase in JUNB gene expression can secure the survival of the senescent cells even when homeostasis of different cellular processes is disrupted. Moreover, our bioinformatic evaluation proposed selumetinib, a MEK inhibitor, as a senolytic against senescent lung fibroblasts. The senolytic activity of a variety of MEK inhibitors in senescent lung fibroblasts was confirmed using human lung fibroblasts in vitro.

pharmacology and toxicology↗

CLCNet: a contrastive learning and chromosome-aware network for genomic prediction in plants

Genomic selection (GS) leverages genome-wide markers and phenotypes to predict breeding values, with its effectiveness largely dependent on the accuracy of genomic prediction (GP) models. However, GP methods often struggle to capture inter-individual variability and are limited by the curse of dimensionality, where the number of SNPs far exceeds the sample size. To address these challenges, we present CLCNet (Contrastive Learning and Chromosome-aware Network), a novel deep learning framework that integrates contrastive learning and chromosome-aware feature modeling. CLCNet comprises two key components: (i) a contrastive learning module that enhances the models ability to capture fine-grained, genotype-dependent phenotypic differences among individuals, and (ii) a chromosome-aware module that captures structured feature selection at both chromosome and genome levels, thereby distilling the most informative SNPs. We evaluated CLCNet across four crop species, covering ten agronomically important traits, and compared it with a diverse set of classical linear, machine learning, and deep learning models. CLCNet achieved superior prediction performance, with statistically significant improvements in Pearson correlation coefficient (PCC), ranging from 0.34% to 12.19% over baseline, together with reduced mean squared error (MSE). Performance gains were more pronounced for traits with moderate linkage disequilibrium (LD; r2 = 0.21-0.36) and high heritability (h2 > 0.66), such as those in maize, rapeseed, and soybean. For cotton traits characterized by high LD (r2 = 0.74) and lower heritability (h2 < 0.50), CLCNet maintained robust performance without degradation. Overall, these results demonstrate that CLCNet is an effective framework for improving genomic prediction accuracy and holds strong potential for practical applications in plant breeding. Short abstractCLCNet is a novel deep learning framework for genomic prediction that integrates contrastive learning with chromosome-aware feature selection. By jointly modeling inter-individual genotype-phenotype variation and chromosomal genomic structure, CLCNet improves prediction accuracy under high-dimensional, low-sample-size conditions. Across four crop species and ten agronomic traits, CLCNet consistently outperformed classical statistical, machine learning, and existing deep learning models. The framework also identified biologically relevant SNPs and candidate genes, demonstrating its potential for practical applications in genomic selection and computational plant breeding. Key pointsO_LIWe propose CLCNet, a multi-task deep learning framework that integrates contrastive learning with chromosome-aware feature selection for genomic prediction, under high-dimensional, low-sample-size conditions. C_LIO_LIThe chromosome-aware module explicitly exploits genomic structural information to select representative and informative SNPs across chromosomes. C_LIO_LIContrastive learning improves model robustness by stabilizing representation learning and reducing the influence of random effects across samples. C_LIO_LIBy complementing GWAS analyses, CLCNet provides additional insights into genotype-phenotype relationships with potential relevance for gene discovery. C_LI Biographical NoteJiangwei Huang is a PhD candidate at the Institute of Genetics and Developmental Biology, Chinese Academy of Sciences. His research focuses on genomic prediction, deep learning, and computational plant breeding. Zhihan Yang is a PhD candidate at the Institute of Genetics and Developmental Biology, Chinese Academy of Sciences. Her research interests include genomic prediction and bioinformatics. Rongcheng Han is an associate professor at the Institute of Genetics and Developmental Biology, Chinese Academy of Sciences. His research focuses on bioinformatics and plant phenomics. Yuqiang Jiang is a professor at the Institute of Genetics and Developmental Biology, Chinese Academy of Sciences. His research interests include plant genomics, plant phenomics and genetic improvement. Organization descriptionThe Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, is a leading research institute focusing on genetics, genomics, molecular breeding, bioinformatics, and systems biology in plants and animals.

genomics↗

Microbiome-oriented data mining of operational monitoring of anaerobic digestion reactor during steady operation period, failure, and restoration.

Anaerobic digestion (AD) is an essential biotechnology for sustainable waste management and bioenergy production, driven by a complex microbial ecosystem. However, the stability and efficiency of AD reactors are frequently challenged by microbial shifts resulting from operational disturbances. This study employs microbiome-oriented data mining and bioinformatics approaches to analyze microbial community dynamics in an anaerobic digestion reactor under steady-state operation, failure, and restoration phases. Data were extracted from a biogas plant and used to evaluate a system failure, providing crucial insights into microbial shifts associated with operational instability. A custom-built microbiome-oriented database, 2Gas, was developed to integrate sequencing data with physicochemical monitoring results, enabling structured data storage, retrieval, and correlation analysis. This database was instrumental in identifying key microbial taxa responsible for reactor destabilization and in predicting failure events based on microbial and physicochemical trends. Through high-throughput 16S rRNA gene sequencing and advanced bioinformatics analysis, microbial shifts were correlated with key operational parameters such as pH, volatile fatty acids (VFAs), and ammonia concentrations. Results indicate that methanogenic archaea, particularly Methanosarcina, experienced a sharp decline during reactor failure, followed by a gradual recovery, demonstrating their sensitivity to environmental stressors. Additionally, the balance between Firmicutes and Actinobacteria was identified as a crucial determinant of reactor stability, with Firmicutes recovering in tandem with system restoration. Feedstock composition was found to play a significant role in microbial shifts, with seasonal variability influencing community composition and reactor performance. Machine learning models applied to the database suggested the potential for predictive analytics in anticipating system failures. By leveraging microbial abundance patterns, these models were able to identify early warning signs of instability. Restoration efforts, including feedstock adjustments and operational parameter optimization, successfully reinstated core microbial communities essential for biogas production. This study underscores the critical role of microbiome monitoring in anaerobic digestion and highlights the potential of bioinformatics tools for enhancing reactor resilience. Future research should focus on real-time microbiome tracking, machine learning-driven predictive maintenance, and adaptive operational strategies to further improve AD efficiency and biogas yield.

microbiology↗

FlowAgent: A Modular Agent-Based System for Automated Workflow Management and Data Interpretation

Reproducibility, automation, and flexibility remain persistent challenges in bioinformatics, where complex workflows require integration of diverse computational tools, rigorous quality control, and dynamic adaptability to evolving datasets. Existing workflow managers often trade off usability for flexibility, limiting accessibility for non-specialists and enforcing rigid execution pipelines. We present FlowAgent, an adaptive agent-based framework that embeds computational heuristics and intelligent decision-making into workflow management. Unlike static pipeline managers, FlowAgent leverages agentic systems for context-aware automation, enhancing error detection, correction, and real-time optimisation while minimising manual intervention. By integrating bioinformatics tools with adaptive control, FlowAgent ensures robust quality control and enables workflow execution through an intuitive natural language interface. This approach democratises advanced computational methods for biologists while allowing bioinformaticians to focus on high-level analysis. FlowAgent redefines workflow automation, making bioinformatics pipelines more intelligent, efficient, and accessible, ultimately accelerating scientific discovery across diverse omics data modalities. Availability and implementationFlowAgent is available with GLPv3 license at https://github.com/EnteloBio/flowagent

genomics↗

Investigating pancreatic beta cell membrane epitopes using unbiased cell-based Fab-phage display

The phenotypic and functional changes of cells in response to physiological and pathological conditions are strongly influenced by the roles of plasma membrane proteins. Recombinant Fab antibody-based phage display for an unbiased antigen-driven affinity selection is a suitable approach for identifying novel membrane proteins. Alterations in the function and distribution of cell membrane proteins in pancreatic {beta} cells have been observed in pathological conditions like diabetes. In this study, we integrated an unbiased cell-based Fab-phage display screening method with bioinformatics tools to identify and characterize Fabs that selectively bind to pancreatic {beta} cells in conditions simulated by a hyperglycemic environment. We isolated three Fab-phages, namely Fab_53, 538, and 54.68, that have binding properties matching specific epitopes on the MIN6 membrane. These Fabs are part of the immunoglobulin G groups that contain Kappa light chains. Bioinformatics analysis of the variable domains of their light and heavy chains (VL and VH) revealed that the potential epitope binding sites on the {beta} cell membrane are associated with pathways involved in insulin activity. Through FACS and IF analysis, we found that, of the three Fabs, Fab_538 exhibited the strongest binding to MIN6 cells. The use of InterProScan software resulted in the generation of 344 potential Fab_538 binding epitopes, and from these, AF2Complex predicted 10 interacting antigens. Our goal in combining Fab-phage display with bioinformatic tools is to develop a more effective, specific, and streamlined method for identifying disease-modifying membrane epitopes for monoclonal antibodies (mAbs).

molecular biology↗

Tracking polyhydroxyalkanoate biosynthesis in thermophilic microorganisms

Polyhydroxyalkanoates are biopolyesters synthesized and stored in intracellular granules by diverse prokaryotes. Despite intense research efforts and prior evidence of a rather widespread phylogenetic occurrence of the related genetic machinery, reports on extreme thermophilic and hyperthermophilic polyhydroxyalkanoates producers remain scarce. However, thermophilic cell factories for bioplastic production would serve as an excellent example of Next-Generation Industrial Biotechnology. In this study, we aim to address this research gap by establishing a bioinformatics pipeline to mine genomes of extremely and moderately thermophilic microorganisms for signatures of potential polyhydroxyalkanoate production. Based on a collection of verified protein sequences of polyhydroxyalkanoate polymerase PhaC, the key biosynthetic enzyme, carefully curated sets of thermophilic bacterial and archaeal genomes were screened. This revealed that although PhaC-encoding genes are prevalent in diverse moderately thermophilic bacteria, they are absent in the considered extreme thermophilic bacteria. In contrast, a few limited examples of extreme thermophilic archaea were found to encode putative phaC genes embedded within a typical polyhydroxyalkanoate synthesis operon in their genomes, namely within the genera Ferroglobus, Geoglobus and Archaeoglobus, while no hits were found in extreme thermophilic bacteria. The latter included Thermus thermophilus, which was previously reported as a polyhydroxyalkanoates producer. This was refuted in our bioinformatics analysis and moreover, the predicted absence of polyhydroxyalkanoates synthesis in T. thermophilus was experimentally confirmed by employing various extraction and analytical methods. Based on the findings in this study, we conclude that polyhydroxyalkanoate production is very scarce in extreme thermophiles and hyperthermophiles, for reasons that remain to be elucidated. HighlightsO_LIA bioinformatics pipeline was constructed to screen thermophilic genomes for PhaC. C_LIO_LIPHA production is widespread in moderate thermophiles but rare in extreme thermophiles. C_LIO_LIExtreme thermophilic archaea belonging to specific genera exceptionally harbor PHA synthesis genes. C_LIO_LINo PHA synthesis genes were found in extreme thermophilic bacteria like Thermus spp. C_LIO_LIExperimental work confirmed the absence of PHAs in Thermus thermophilus. C_LI Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=113 SRC="FIGDIR/small/652502v1_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@11fafe3org.highwire.dtl.DTLVardef@1548075org.highwire.dtl.DTLVardef@bd9ebaorg.highwire.dtl.DTLVardef@fff3e0_HPS_FORMAT_FIGEXP M_FIG C_FIG

microbiology↗

ERSAtool: A User-Friendly R/Shiny Comprehensive Transcriptomic Analysis Interface Suitable for Education

RNA sequencing (RNA-seq) has become an essential technology for assessing gene expression profiles in biomedical research. However, the coding complexity of RNA-seq data analysis remains a significant barrier for students and researchers without extensive bioinformatics expertise. We present the Educational RNA-Seq Analysis tool (ERSAtool), a comprehensive R/Shiny interface that provides an intuitive graphical visualization of the complete RNA-seq analysis workflow. The application is built on established Bioconductor packages and upholds high standards in analyses while significantly reducing the technical expertise required to conduct sophisticated transcriptomic analyses. ERSAtool supports various input formats, such as raw count matrices and STAR alignment outputs. It generates sample information metadata through direct integration with the Gene Expression Omnibus (GEO) provided by the National Center for Biotechnology Information (NCBI). The application guides users through normalization, data visualization, differential expression analysis, and functional interpretation using Gene Ontology (GO) and Gene Set Enrichment Analysis (GSEA). All results can be compiled into comprehensive, downloadable reports that enhance reproducibility and knowledge sharing. The design includes targeted features that facilitate educational use, making it especially useful for teaching transcriptomics in undergraduate to graduate-level bioinformatics courses. By connecting command-line bioinformatics tools with accessible graphical interfaces, ERSAtool improves accessibility to advanced transcriptomic analysis capabilities, potentially accelerating discoveries across various biological fields. The source code for the ERSAtool package is available at https://github.com/SuzukiLabTAMU/ERSAtool and is released under the GNU General Public License v3.0 (GPL-3.0).

genetics↗

Large-scale classification of metagenomic samples: a comparative analysis of classical machine learning techniques vs a novel brain-inspired hyperdimensional computing approach

Classical machine learning techniques have revolutionized bioinformatics, enabling researchers to extract knowledge from complex biological data. However, these techniques often struggle with high-dimensional data, where the increasing number of features leads to decreased performance, also affecting models accuracy. To address this problem, we explore hyperdimensional computing (HDC), an emerging brain-inspired computational paradigm that leverages high-dimensional vectors and simple arithmetic operations to represent and manipulate complex patterns, as an alternative approach in the context of supervised machine learning. In this work, we present a comprehensive comparative analysis of HDC against established machine learning techniques across a range of classification tasks. As a representative use case, we focus on classifying heterogeneous metagenomic samples based on their quantitative microbial profiles, using publicly available microbiome datasets. Our results demonstrate that HDC achieves comparable, and in some cases, superior classification accuracy to classical methods. Furthermore, our findings highlight the potential of HDC for improved computational efficiency, particularly when dealing with large-scale datasets, suggesting the HDC-based classifier as a promising tool for bioinformatics research, particularly in areas characterized by high-dimensional data. We also offer a Galaxy powered toolset to analyze your own datasets and generate reproducible workflows and adopt these methods in your own research with ease. Our investigation into the application of a HDC-based supervised machine learning technique for classifying microbial profiles in metagenomic samples yielded promising results, demonstrating the potential of this novel computational paradigm to complement and, in some cases, surpass the performances of well established machine learning techniques. ImportanceThe growing complexity and dimensionality of biological data require more efficient and scalable machine learning approaches. HDC offers a novel alternative to conventional methods, showing resilience to high-dimensionality while maintaining competitive accuracy. This study demonstrates the effectiveness of HDC in classifying metagenomic samples based on their microbial composition. Our results suggest that HDC not only matches, but sometimes exceeds the performance of well-established methods. We make this approach accessible to the broader bioinformatics community with an open-source tool fully integrated into the Galaxy platform, facilitating its adoption and reproducibility, with the aim of integrating HDC into mainstream biological data analysis pipelines, especially for complex, high-dimensional tasks in microbiome research.

microbiology↗

A Globally Representative Immunopeptidomics Approach to Identify Population-Wide Vaccine Candidates

Vaccine development has historically relied on preclinical testing using one or two cell lines, typically of Caucasian descent, contributing to vaccines that may fail to provide adequate protection across genetically diverse populations. A major factor in these failures is the lack of consideration for human leukocyte antigen (HLA) diversity, with over 30,000 HLA alleles worldwide exhibiting distinct frequencies across ethnic groups and geographic regions. Here, we present a globally representative immunopeptidomics approach that addresses HLA diversity at the earliest stages of vaccine antigen discovery. We established a panel of 30 cell lines from the 1000 Genomes Project that represent 75% of the worlds most common HLA alleles. Using a combined computational and experimental approach, we identified 18 common binding motifs (7 MHC class I and 11 MHC class II) shared across 24 cell lines from the panel. We developed an accessible bioinformatic tool that predicts globally presentable antigenic regions from any pathogen proteome by identifying peptides matching these common binding motifs. Validation with Salmonella enterica serovar Typhimurium and SARS-CoV-2 Spike protein demonstrated that our method successfully identifies known immunogenic regions, with experimentally detected epitopes showing substantial overlap with predicted regions of interest. Notably, only three cell lines were required to validate highly immunogenic S. enterica targets, including OmpC--a porin with known 100% protective efficacy--detected across all tested lines. Our approach enables researchers to rapidly screen pathogen proteomes using freely available bioinformatic tools, with optional experimental validation requiring minimal resources. By identifying antigens with broad population coverage before clinical trials begin, this method has the potential to increase vaccine success rates while ensuring equitable protection across diverse populations. The cell line panel, common binding motifs, and bioinformatic workflow are publicly available, providing an accessible pathway for developing vaccines that truly serve global populations.

immunology↗

Novel mutations associated with autosomal dominant congenital cataract are identified in Chinese families

PurposeAs the leading cause of the impairment of vision of children, congenital cataract is considered as a hereditary disease, especially autosomal dominant congenital cataract (ADCC). The purpose of this study is to identify the genetic defect of six Chinese families with ADCC.\n\nSubjects and MethodsSix Chinese families with ADCC were recruited in the study. (103 members in total, 96 members alive, 27 patients in total) Genomic DNA samples extracting from probands peripheral blood cells were captured the mutations using a specific eye disease enrichment panel with next generation sequencing. After initial pathogenicity prediction, sites with specific pathogenicity were screened for further validation. Sanger sequencing was conducted in the other individuals in the families and other 100 normal controls. Mutations definitely related with ADCC will then be analyzed by bioinformatics analysis. The pathogenic effect of the amino acid changes and structural and functional changes of the proteins were finally analyzed by bioinformatics analysis.\n\nResultsSeven mutations in six candidate genes associated with ADCC of six families were detected (MYH9 c.4150G>C, CRYBA4 c.169T>C, RPGRRIP1 c.2669G>A, WFS1 c.1235T>C, CRYBA4 c.26C>T, EPHA2 c.2663+1G>A, and PAX6 c.11-2A>G). All the seven mutations were only detected on affected individuals in the families. Among them there are three novel mutations (MYH9 c.4150G>C, CRYBA4 c.169T>C, RPGRRIP1 c.2669G>A) and four that have been reported (WFS1 c.1235T>C, CRYBA4 c.26C>T, EPHA2 c.2663+1G>A, and PAX6 c.11-2A>G). RPGRIP1 (c.2669G>A) mutation and CRYBA4 (c.26C>T) mutation are predicted to be benign according to bioinformatics analysis while the other five mutations (EPHA2, PAX6, MYH9, CRYBA4 c.169T>C, WFS1) are thought to be pathogenic.\n\nConclusionWe report two novel heterozygous mutations (MYH9 c.4150G>C and CRYBA4 c.169T>C) in six Chinese families supporting their vital roles in causing ADCC.

genomics↗

Comparative genomics confirms a rare melioidosis human-to-human transmission event and reveals incorrect phylogenomic reconstruction due to polyclonality

Human-to-human transmission of the melioidosis bacterium, Burkholderia pseudomallei, is exceedingly rare, with only a handful of suspected cases documented to date. Here, we used whole-genome sequencing (WGS) to characterise one such unusual B. pseudomallei transmission event, which occurred between a breastfeeding mother with mastitis and her child. Two strains corresponding to multilocus sequence types (STs) 259 and 261 were identified in the mothers sputum from both the primary culture sweep and in purified colonies, confirming an unusual polyclonal infection in this patient. In contrast, primary culture sweeps of the mothers breast milk and the childs cerebrospinal fluid and blood samples contained only ST-259, indicating monoclonal transmission to the child. Analysis of purified ST-259 isolates showed no genetic variation between mother and baby isolates, providing the strongest possible evidence of B. pseudomallei transmission, probably via breastfeeding. Next, phylogenomic analysis of all isolates, including the mothers mixed ST-259/261 sputum sample was performed to investigate the effects of mixtures on phylogenetic inference. Inclusion of this mixture caused a dramatic reduction in the number of informative SNPs, resulting in branch collapse of ST-259 and ST-261 isolates, and several instances of incorrect topology in a global B. pseudomallei phylogeny, resulting in phylogenetic incongruence. Although phylogenomics can provide clues about the presence of mixtures within WGS datasets, our results demonstrate that this methodology can lead to phylogenetic misinterpretation if mixed genomes are not correctly identified and omitted. Using current bioinformatic tools, we demonstrate a robust method for bacterial mixture identification and strain parsing that avoids these pitfalls.\n\nImpact StatementBurkholderia pseudomallei is the causative agent of melioidosis, a tropical disease of high mortality. B. pseudomallei infections occur almost exclusively through contact with contaminated soil and water. Using whole-genome sequencing (WGS), we investigated a rare case of suspected B. pseudomallei transmission from mother to child. The mothers sputum, breast milk and the babys blood and cerebrospinal fluid (CSF) specimens were collected, and DNA was extracted from both pure colonies and primary culture sweeps to capture potential strain mixtures. In-depth analysis of genetic variants identified two strains in the mothers sputum belonging to multilocus sequence types ST-259 and ST-261, whereas the child was infected with only ST-259. Comparative genomics revealed no genetic differences between mother and child ST-259 isolates, providing the strongest possible evidence of transmission to the child via breast milk. The sputum strain mixture was subsequently used to develop a bioinformatic method for identification and quantification of mixtures from WGS data. Using this method, we found ST-259 and ST-261 at an 87%:13% ratio, respectively. Finally, we demonstrate the negative impact that even a single strain mixture event can have on both within-ST and global phylogenomic inferences. Our findings highlight the need for bioinformatic quality control to avoid unintended consequences of phylogenomic incongruence and branch collapse.\n\nData SummaryO_LIWhole-genome sequencing data have been deposited in the NCBI Sequence Read Archive (SRA) and GenBank under BioProject accession number PRJNA559002.\nC_LIO_LIThe GenBank accession number for MSHR0643 assembly is VXLH00000000.1.\nC_LIO_LIThe SRA accession numbers for all raw sequence data are listed in Table 1.\nC_LI\n\nO_TBL View this table:\norg.highwire.dtl.DTLVardef@c3b580org.highwire.dtl.DTLVardef@c11407org.highwire.dtl.DTLVardef@9e443org.highwire.dtl.DTLVardef@169014borg.highwire.dtl.DTLVardef@d4026c_HPS_FORMAT_FIGEXP M_TBL O_FLOATNOTable 1.C_FLOATNO O_TABLECAPTIONSummary of ST-259 and ST-261 Burkholderia pseudomallei isolates.\n\nC_TABLECAPTION C_TBL RepositoriesAll sequencing data generated as part of this study can be found under the NCBI BioProject PRJNA559002 with accession numbers listed in Table 1.

microbiology↗