Virophages parasitize the replication of co-infecting giant viruses within eukaryotic cells, forming tripartite cell-virus-virophage (CVv) systems. Tripartite interactions are well-documented in protozoa, yet comparable systems in algae remain largely unexplored at the experimental level. Here, we report an experimentally validated CVv system involving the green, single-celled microalga Chlorella sp. DSL01, Dishui Lake large algal virus 1 (DSLLAV1), and multiple Dishui Lake virophages (DSLVs). Inoculation of Chlorella sp. DSL01 at low MOI established laboratory co-cultures in which time-series PCR detected DSLLAV1 early but not after Day 10, whereas all tested virophages persisted. Metagenomic profiling of the terminal supernatant (end-point sample) indicated a virophage-dominated assemblage with DSLV3 most represented. Droplet digital PCR at discrete time points (Days 5, 10, and 15) then provided absolute counts for DSLLAV1 and DSLV1/3/7, corroborating an early DSLLAV1 peak followed by collapse and/or a delayed rise of multiple virophages coincident with host growth recovery. Nested PCR on the algal pellet detected virophages DSLV1/3/7 but not DSLLAV1. Together, these results demonstrate that Chlorella sp. DSL01 supports co-infection by DSLLAV1 and multiple virophages, establishing an experimentally validated algal CVv system and revealing multi-virophage participation in freshwater algal virus-virophage-host dynamics.

We isolated a novel halophilic archaeon, strain DSL9, representing the proposed new species Haloliberatus hailidukes gen. nov., sp. nov., from Dishui Lake, China. Unlike most obligate halophiles, DSL9 survives in low salinity, even distilled water, without lysis. Genomic analysis revealed dual salinity adaptation strategies: salt-in and compatible solutes, including a complete trehalose biosynthesis pathway. The strain harbors multiple plasmids, notably a 111,311 bp large plasmid (pHdsl9-3) encoding replication (Orc1/Cdc6, SSB), transcription (TFIIB), transmission (T4SS cluster, ArdC-like protein), and recombination (XerA) modules. pHdsl9-3 provides auxiliary functions such as defense, genome diversification, ion detoxification, and suggests active horizontal gene transfer. Similar elements are widespread in Halobacteriales, highlighting their role in haloarchaeal genetic diversity and plasticity. The encoded XerA hinted at a function beyond DNA dimer resolution, suggesting it may have been adapted by other archaeal mobile genetic elements. These findings underscore the need to investigate plasmid-driven evolution and environmental adaptation mechanisms in haloarchaea.IMPORTANCEThis study reports the isolation and characterization of DSL9, a novel halophilic archaeon from a freshwater lake. Remarkably, DSL9 defies the typical obligate halophilic lifestyle by surviving in low-salinity environments, including distilled water, without cell lysis. A key discovery is the identification of a 111,311 bp large plasmid harboring essential modules for replication, transcription, transmission, and integration. Widespread distribution of similar elements across Halobacteriales suggests their crucial role in haloarchaeal genetic diversity and plasticity, warranting further study of plasmid-mediated evolution and adaptation strategies.

In the original publication [...].

The efficacy of immunotherapy in colorectal cancer (CRC) has long been considered to be closely associated with microsatellite instability (MSI) status. Patients with microsatellite stable (MSS) tumors typically exhibit poor responses to PD-1/PD-L1 inhibitors and a poor prognosis, often being categorized as immunologically 'cold' tumors. However, some MSS patients can still achieve favorable therapeutic responses, sometimes even surpassing those of certain MSI patients. Immune-cold and immune-hot tumor phenotypes are largely determined by the abundance, clonal expansion, and functional states of tumor-infiltrating T cells. This suggests that immunotherapy responses are driven by dynamic remodeling of T-cell clonality rather than by MSI status alone. To elucidate the underlying T cell clonal dynamics, integrated single-cell transcriptome (scRNA-seq) and T cell receptor sequencing (scTCR-seq) data analyses from 43 blood and tissue samples of MSI and MSS colorectal cancer patients before and after anti-PD-1 therapy were performed. Using our developed TCR reconstruction pipeline (TORBiT), we systematically analyzed the clonal architecture of the TCR repertoire, inter-tissue migration, and its association with T-cell functional state transitions. From a TCR clonal kinetic perspective, we revealed two distinct modes of immune Coercion that may further affect the immune response: a "high-fluctuation, deep-exhaustion" pattern in MSI tumors and a "high-baseline, strong-suppression" pattern in MSS tumors. These findings provide a novel theoretical foundation and research perspective for understanding the responsiveness and resistance mechanisms to immune checkpoint inhibitors.

Virophages are small double-stranded DNA viruses that parasitize giant viruses, modulating virus–host interactions and influencing microbial community dynamics. Despite their ecological significance, virophages and giant viruses remain poorly studied in extreme environments. Here, we present the first metagenomic survey of virophage and giant virus diversity in Lake Namtso, a high-altitude saline lake on the Tibetan Plateau. Metagenomic assembly and phylogenomic analyses uncovered 93 virophage major capsid protein sequences spanning seven established families, alongside numerous unclassified lineages. Two nearly complete virophage genomes were reconstructed: Namtso Virophage 1 (NMV1), which encodes both replication- and integration-associated genes and likely represents a novel family, and Namtso Virophage 2 (NMV2), affiliated with Omnilimnoviroviridae but distinguished by duplicated protease genes and dual DNA methyltransferases. Parallel analyses identified over 18,000 giant virus marker genes, with DNA PolB affiliated to Imitervirales, Pimascovirales, Asfuvirales, Algavirales, and Chitovirales, as well as divergent lineages representing potential novel Nucleocytoplasmic large DNA viruses (NCLDVs). Homologous protein analysis and tetranucleotide clustering suggest extensive host–virus–virophage interactions. These findings significantly expand the known diversity and genomic repertoire of virophages and giant viruses, highlight their ecological roles in sustaining microbial resilience, and provide new insights into viral evolution and adaptation in extreme high-altitude ecosystems.

Mitochondria not only generate ATP and metabolites essential for nuclear and cytoplasmic processes but also actively shape nuclear epigenetic regulation. Conversely, the nucleus encodes most of the proteins required for mitochondrial functions, and intriguingly, certain nuclear-encoded epigenetic factors—such as DNA and histone modifiers—also localize to mitochondria, where they modulate mitochondria genome stability, gene expression, metabolic flux, and organelle integrity. This reciprocal interplay defines mitochondria as both a source and a target of epigenetic regulation, integrating energy metabolism with gene expression and cellular homeostasis. This review highlights emerging mechanisms that link mitochondrial metabolism to chromatin remodeling, DNA and histone modifications, and transcriptional control, as well as how nuclear epigenetic enzymes translocate into mitochondria and regulates their functions. We also briefly introduce recent methodological advances that enable spatially selective depletion of mitochondrial proteins, offering new tools to dissect this bidirectional communication. Together, these insights underscore mitochondria’s central role as an energetic and epigenetic hub coordinating nuclear function, development, and disease.

The escalating challenge of antibiotic resistance poses a global health emergency. Edible and medicinal plants (EMPs) provide a rich source of safe, effective and affordable phytochemical alternatives. Hence, a thorough inquiry into this domain is critical to meet the urgent demand of enhancement of existing antibacterial pharmacopeia and development of innovative approaches or novel antimicrobials to tackle the burgeoning issue of multi-drug resistant foodborne pathogens. In this review, we consolidate the recent discovery of antibacterial activity of extracts from diverse species of EMPs, new antibacterial compounds derived from these plants and their structural details of relevant antibacterial function, as well as recent advances in understanding how these compounds prevent prolification of foodborne pathogens. Finally, we discuss how we can utilize antibacterial phytochemicals in food package materials, preservers, and disinfectants in food industry.

The ability of T-cell receptors (TCRs) to recognize neoantigens is fundamental to the initiation and maintenance of adaptive immune responses. In TCR-based immunotherapies, elucidating the recognition patterns of TCRs for peptides and accurately identifying therapeutically relevant TCR-peptide pairs remain critical challenges. Here, we present a novel dual-pathway network model, ProTCR, which integrates the protein language model ProtT5 with deep learning methods. By incorporating both global and local feature extraction mechanisms, ProTCR enables efficient representation of amino acid sequences, thereby enhancing the model's generalizability across diverse data distributions and improving its biological interpretability. ProTCR demonstrates robust performance and broad applicability across various datasets, including neoantigens, previously unseen peptides, and MHC class II-restricted epitopes, overcoming the reliance on known peptide-TCR pairs observed in previous studies. It also offers new insights for predicting diverse classes of antigenic peptides. We applied ProTCR to several clinically relevant scenarios, including immunotherapeutic target identification in acute myeloid leukemia, neoantigen-targeted immunotherapy in solid tumours, and antigen-specific T cell recognition against pathogens such as influenza and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Across these complex settings, ProTCR consistently maintained high accuracy and stability, demonstrating strong cross-task adaptability and broad potential for clinical application. This work not only provides a powerful tool for elucidating immune response mechanisms but also offers a solid computational foundation for the design of neoantigen or TCR based precision immunotherapy strategies.

RNA viruses are highly abundant and diverse in marine ecosystems, yet the currently discovered viral species represent only a minute fraction of the total diversity, with viral communities in port ecosystems particularly understudied. To bridge this gap, in summer, approximately 100 L of surface water was collected from Yangshan Port. Viral particles were concentrated using tandem filtration and 50-kDa tangential-flow ultrafiltration, followed by metatranscriptomic sequencing of extracted RNA. This approach yielded 74 full-length RNA-dependent RNA polymerase (RdRp) sequences, which clustered into 72 distinct genus-level (or higher) taxonomic units—significantly expanding the known diversity of seaport RNA viruses. Phylogenetic and EFI-EST network analyses showed that 93

Neoantigen-based vaccines represent a promising approach in cancer immunotherapy, with the key to their effective clinical application lying in the precise identification of immunogenic neoantigens. Existing methods primarily focus on genomic variations, lacking integration of multi-omics data and essential filtering steps, which limits comprehensive assessment of immunogenicity and results in only a small subset of neoantigens capable of eliciting effective immune responses. Moreover, the complexity and poor portability further hinder the clinical applicability. To address these limitations, we developed OmniNeo, an automated multi-omics-based neoantigen discovery framework. 1) OmniNeo integrates whole-genome/exome sequencing (WGS/WES), transcriptomic, and proteomics data to simultaneously identify neoantigenic epitopes derived from SNVs/Indels, frameshift mutations, gene fusions, and non-coding region variations; 2) The pipeline incorporates a convolutional neural network-based model, OmniNeo-CNN along with multiple filtering mechanisms to quantify the immunogenicity and T-cell receptor (TCR) recognition potential of predicted neoantigen candidates through multiple features; 3) The workflow is built on nextflow, offering a one-stop, scalable, and portable solution for rapid and efficient neoantigen prediction. Finally, we demonstrated the practical application procedures of this workflow in potential tumor immunotherapy through case study analyses of liver cancer samples. The tool is freely accessible as an open-source resource via https://github.com/linfengxu/OmniNeo, https://zenodo.org/records/15340824.

The International Committee on Taxonomy of Viruses (ICTV) holds a ratification vote annually following the review of newly proposed taxa by ICTV Study Groups and members of the virology community. This article reports changes to the taxonomy of viruses infecting archaea that were approved and ratified by the ICTV in March 2025. Six new families of head-tailed viruses expanded the order Caudoviricetes (realm Duplodnaviria); one new family of filamentous viruses was added to the order Ligamenvirales (realm Adnaviria); one new family of viruses with pleomorphic virions was included within a new phylum, new order and new class in the kingdom Trapavirae (realm Monodnaviria); finally, three new families were created for spindle-shaped viruses that remain unassigned to higher level taxa. The 25 new species represent viruses infecting a broad range of archaea, including members of the classes Archaeoglobi, Bathyarchaeia, Methanobacteria, Methanomicrobia, Nitrososphaeria and Poseidoniia. Most of these viruses have been discovered by metagenomics in samples derived from diverse environments, including ambient and extreme marine ecosystems, the gastrointestinal tract of humans and animals, anaerobic digesters and terrestrial hot springs. Following this taxonomic update, archaeal viruses are officially classified into a total of 163 virus species in 94 genera within 62 families.

Background/Objectives: Cell–cell communication (CCC) is a critical process within the tumor microenvironment, governing regulatory interactions between cancer cells and other cellular subpopulations. Aiming to improve the accuracy and completeness of intercellular gene-regulatory network inference, we constructed a novel spatial-resolved gene-regulatory network framework (spGRN). Methods: Firstly, the spatial multi-omics data of colorectal cancer (CRC) patients were analyzed. We precisely located the tumor boundaries and then systematically constructed the spGRN framework to study the network regulation. Subsequently, the key signaling molecules obtained by the spGRN were identified and further validated by the spatial-proteomics dataset. Results: Through the constructed spatial gene regulatory network, we found that in the communication with malignant cells, the highly expressed ligands LIF and LGALS3BP and receptors IL6ST and ITGB1 in fibroblasts can promote tumor proliferation, and the highly expressed ligands S100A8/S100A9 in plasma cells play an important role in regulating inflammatory responses. Further, validation of the key signaling molecules by the spatial-proteomics dataset highlighted the role of these genes in mediating the regulation of boundary-related cells. Furthermore, we applied the spGRN to publicly available single-cell and spatial-transcriptomics datasets from three other cancer types. The results demonstrate that ITGB1 and its target genes FOS/JUN were commonly expressed in all four cancer types, indicating their potential as pan-cancer therapeutic targets. Conclusion: the spGRN was proven to be a useful tool to select signal molecules as potential biomarkers or valuable therapeutic targets.

Klebsiella oxytoca can cause gastrointestinal infections in humans. Within the instestinal tract, the bacterium encounters diverse host-derived signals (most notably bile salts, BS) and secondary metabolites (such as short-chain fatty acid salts, SCFAs) produced by microbial community. However, the mechanism of K. oxytoca response to BS and SCFAs stresses has been rarely reported. Recently, we found K. oxytoca in 14 species of aquatic food animals, of which K. oxytoca 7-7-27 of mussel origin had strong biofilm formation characteristics and many putative virulence-associated genes. Herein, we investigated the impact of normal physiological concentrations of BS (0.2

Along with the widespread application of plastics, the release of polystyrene microplastics (PS-MPs) into the environment raises concerns about the risks of PS-MPs to human health.

Bacteria in the genus Vibrio, including at least 152 species, thrive in marine and estuarine environments and are frequently detected in aquatic products worldwide. Of these, 12 species have been implicated in human infectious diseases, such as the life-threatening pandemic cholera, acute gastroenteritis, and severe sepsis. Nevertheless, molecular mechanisms of their pathogenesis are not fully uncovered yet. Prophages are found prevalent in Vibrio spp. genomes, carrying a number of genes with various functions. In this review, we deciphered the evolutionary relationship between prophages and Vibrio species and highlighted the impact of prophages on the bacterial pathogenicity, environmental fitness, and genome evolution, based on 149 newly discovered intact prophages located in the genomes of 82 Vibrio spp., which we searched and collected from Web of Science Core Collection in the most recent 5 years. The effects of prophages on resistance to superinfection, strain competition, and their regulation were also discussed. This review underscored crucial roles of prophages in shaping Vibrio spp. genomes and their implications for food safety and public health.

Cell-virus-virophage (CVv) systems involve virophages parasitizing giant viruses within eukaryotic hosts, forming unique virus-virus interactions with complex ecological implications. However, quantitative tools for studying such systems-particularly in freshwater algae-remain limited. In this study, we developed and optimized a duplex droplet digital PCR (ddPCR) assay to simultaneously detect and quantify Dishui Lake Large Algal Virus 1 (DSLLAV1), a Mimiviridae-like algal giant virus, and its associated Dishui Lake virophage 8 (DSLV8) in the Dishui Lake ecosystem. Target-specific primers and TaqMan probes were designed based on viral genomic sequences, and assay conditions were optimized for annealing temperature, primer/probe concentrations, and droplet separation. The established assay demonstrated high specificity and sensitivity, with detection limits of 0.13 and 0.16 copies/µL for DSLLAV1 and DSLV8, respectively. The method outperformed qPCR in sensitivity and maintained stability across environmental and infection-derived samples. This ddPCR method provides a robust platform for monitoring virus-virophage dynamics and offers new opportunities for investigating the ecological and evolutionary roles of CVv systems in aquatic environments.

Klebsiella oxytoca originating from shellfish Scapharca subcrenata contains a number of virulence-related genes. In this study, we investigated its pathogenicity using a murine intestinal infection model and predicted its antibacterial compounds and targets via molecular docking analysis. The results revealed that the intake of K. oxytoca 8-2-11 strain (109 CFU/day) via oral gavage for 7 days reduced the average body weight of the mice. The bacterium was present in fecal samples but absent from blood, lung, and liver samples from the mice. The intake of K. oxytoca 8-2-11 significantly altered colon bacteriota, with reduced abundance of Firmicutes, Lachnospiraceae, Lactobacillaceae, Lactobacillus, and Lactobacillus murinus, and increased in Bacteroidota, Muribaculaceae, and Alistipes (p < 0.05). Forty-four bioactive compounds in Scutellaria baicalensis and Forsythia suspensa were screened for docking with 117 potential virulence factors (VFs) in K. oxytoca 8-2-11. The compound baicalin displayed higher binding affinity toward these VFs, with the lowest mean binding energy (−8.4 kcal/mol). Baicalin was able to bind to key VFs in biofilm formation and adherence/motility (e.g., Mrks and EcpA) via forming stable hydrogen bonds, π-stacking, and π-cation interaction. In vitro, baicalin inhibited the bacterial growth and biofilm formation. This study establishes the first murine infection model using aquatic animal-derived K. oxytoca, and it provides candidate antibacterial compounds and targets for control of K. oxytoca infections.

Following a 3.5-year enrichment cultivation period, a novel ammonia-oxidising archaeon (AOA), designated strain CL1T, was isolated from Yangshan Harbour (East China Sea). Strain CL1T demonstrates a maximum ammonia tolerance of up to 10 mM. Its optimal growth conditions include a pH range of 7-8, a salinity of 2%-3%, and a temperature range of 20°C-25°C. Under these conditions, strain CL1T achieved a maximum specific growth rate of 0.87 d-1, with cell yields estimated at 3.92 × 106 cells mL-1 μM ammonia-1. Genomic sequencing revealed that strain CL1T possesses a genome size of 1.63 megabases with a high completeness of 99.95%. Phylogenetic analysis based on the 16S rRNA gene and whole-genome data placed strain CL1T within the genus Nitrosarchaeum. The average nucleotide identity (ANI) between the genome of strain CL1T and its closest relative was 92.01%, confirming that strain CL1T represents a novel species within Nitrosarchaeum. Metabolic pathway analysis demonstrated that strain CL1T encodes key enzymes for ammonia oxidation, including ammonia monooxygenase (amoA, amoB, amoC) and copper oxidase, indicating its capacity for ammonia oxidation. Additionally, strain CL1T likely assimilates ammonia through the GS-GOGAT and GDH pathways. Consistent with the observation of extracellular vesicles (EVs) in strain CL1T via electron microscopy, genome annotation identified core genes associated with EVs function, such as vps4 and FtsZ. The isolation of strain CL1T provides a valuable model system for investigating its ammonia metabolism and exploring its ecological interactions with other AOA, ammonia-oxidising bacteria (AOB) and nitrite-oxidising bacteria (NOB), thereby contributing to a deeper understanding of nitrogen cycling mechanisms in aquatic environments.

Edible and medicinal plants provide a treasure trove of natural phytochemicals for mining the next generation of green food preservatives. Herein, we evaluated antibacterial activities of 55–95% ethanol extracts from the edible rhizome of Rumex madaio (RmEEs). The 75% ethanol extract displayed the strongest antibacterial activity, and its purified fraction 2 (RmEE-F2) blocked the proliferation of common pathogens Staphylococcus aureus and Vibrio cholerae, with minimum inhibitory concentrations (MICs) of 391 μg/mL. RmEE-F2 (1 × MIC) altered the bacterial cell surface biophysical parameters and impaired cell structure, resulting in intracellular nucleic acid and protein leakage. It manifested bacteriostatic rates of 88.21–91.17% against S. aureus and V. cholerae in spiked fish (Carassius auratus) and shrimp (Penaeus vannamei) during storage at 4 °C for 24 h. Meanwhile, RmEE-F2 effectively rendered the pH rising and reduced lipid oxidation and protein degradation of C. auratus and P. vannamei meat samples at 4 °C for 6 days. Additionally, RmEE-F2 (< 781 µg/mL) showed non-cytotoxicity to human colon Caco-2, liver HepG-2, and lung A549 cell lines, and rescued V. cholerae and S. aureus-infected Caco-2 cellcells with enhanced viability of 14.31–16.60% (1 × MIC). Comparative transcriptomic analysis revealed down-regulated protein synthesis, cell wall and cell membrane synthesis, and or DNA replication and repair in the tested bacteria triggered by RmEE-F2. The major antibacterial compounds in RmEE-F2 included melibiose (9.86%), 3-(N, N-dimethylaminomethyl) indole (7.12%), and citric acid (6.07%). Overall, this study underscores the promising potential of RmEE-F2 for aquatic product green preservation.

ABSTRACT Archaea play a significant role in natural ecosystems and the human body. Archaeal viruses exert a considerable influence on the structure and composition of archaeal communities and their associated ecological environments. The present study revealed the complete genomes of 38 archaeal head-tailed proviruses through comprehensive data mining. The hosts of these proviruses were identified as belonging to the following four dominant phyla: Halobacteriota, Thermoplasmatota, Thermoproteota, and Nanoarchaeota. In addition to the 14 proviruses of halophilic archaea related to the Graaviviridae family, the remaining proviruses exhibited limited genetic similarities to known (pro)viruses, suggesting the existence of 14 potential novel families. Of the 38 archaeal proviruses, 30 have the potential to lyse host cells. Eleven proviruses contain genes linked to antiviral defense mechanisms, including those involved in restriction modification (RM), clustered regularly interspaced short palindromic repeat (CRISPR)-associated (CRISPR–Cas) nucleases, defense island system associated with restriction-modification (DISARM), and DNA degradation (Dnd). Moreover, auxiliary metabolic genes were identified in the proviruses of Bathyarchaeia and Halobacteriota archaea, including those involved in carbohydrate and amino acid metabolism. Our findings indicate the diversity of archaeal viruses, their interactions with archaeal hosts, and their roles in the adaptation of the host.IMPORTANCEThe field of archaeal virology has seen a rapid expansion through the use of metagenomics, yet the diversity of these viruses remains largely uncharted. In this study, the complete genomes of 38 novel archaeal proviruses were identified for the following four dominant phyla: Halobacteriota, Thermoplasmatota, Thermoproteota, and Nanoarchaeota. Two families and six genera of Archaea were the first to be identified as hosts for viruses. The proviruses were found to contain diverse genes that were involved in distinct adaptation strategies of viruses to hosts. Our findings contribute to the expansion of the lineages of archaeal viruses and highlight their intricate interactions and essential roles in enabling host survival and adaptation to diverse environmental conditions.