Rheumatoid arthritis-associated interstitial lung disease (RA-ILD) is a serious and common extra-articular disease manifestation. Patients with RA-ILD experience reduced bacterial diversity and gut bacteriome alterations. However, the gut mycobiome and virome in these patients have been largely neglected. In this study, we performed whole-metagenome shotgun sequencing on fecal samples from 30 patients with RA-ILD, and 30 with RA-non-ILD, and 40 matched healthy controls. The gut bacteriome and mycobiome were explored using a reference-based approach, while the gut virome was profiled based on a nonredundant viral operational taxonomic unit (vOTU) catalog. The results revealed significant alterations in the gut microbiomes of both RA-ILD and RA-non-ILD groups compared with healthy controls. These alterations encompassed changes in the relative abundances of 351 bacterial species, 65 fungal species, and 4,367 vOTUs. Bacteria such as Bifidobacterium longum, Dorea formicigenerans, and Collinsella aerofaciens were enriched in both patient groups. Ruminococcus gnavus (RA-ILD), Gemmiger formicilis, and Ruminococcus bromii (RA-non-ILD) were uniquely enriched. Conversely, Faecalibacterium prausnitzii, Bacteroides spp., and Roseburia inulinivorans showed depletion in both patient groups. Mycobiome analysis revealed depletion of certain fungi, including Saccharomyces cerevisiae and Candida albicans, in patients with RA compared with healthy subjects. Notably, gut virome alterations were characterized by an increase in Siphoviridae and a decrease in Myoviridae, Microviridae, and Autographiviridae in both patient groups. Hence, multikingdom gut microbial signatures showed promise as diagnostic indicators for both RA-ILD and RA-non-ILD. Overall, this study provides comprehensive insights into the fecal virome, bacteriome, and mycobiome landscapes of RA-ILD and RA-non-ILD gut microbiota, thereby offering potential biomarkers for further mechanistic and clinical research.

The blood virome is dominated by the Anelloviridae family, which emerges early in life; the anellome, which represents the variety of anelloviruses within an individual, stabilizes by adulthood [...].

BACKGROUND: Respiratory viruses significantly impact global morbidity and mortality, causing more disease in humans than any other infectious agent. Beyond pathogens, various viruses and bacteria colonize the respiratory tract without causing disease, potentially influencing respiratory diseases\' pathogenesis. Nevertheless, our understanding of respiratory microbiota is limited by technical constraints, predominantly focusing on bacteria and neglecting crucial populations like viruses. Despite recent efforts to improve our understanding of viral diversity in the human body, our knowledge of viral diversity associated with the human respiratory tract remains limited.
METHODS: Following a comprehensive search in bibliographic and sequencing data repositories using keyword terms, we retrieved shotgun metagenomic data from public repositories (n = 85). After manual curation, sequencing data files from 43 studies were analyzed using EVEREST (pipEline for Viral assEmbly and chaRactEriSaTion). Complete and high-quality contigs were further assessed for genomic and taxonomic characterization.
RESULTS: Viral contigs were obtained from 194 out of the 868 FASTQ files processed through EVEREST. Of the 1842 contigs that were quality assessed, 8% (n = 146) were classified as complete/high-quality genomes. Most of the identified viral contigs were taxonomically classified as bacteriophages, with taxonomic resolution ranging from the superkingdom level down to the species level. Captured contigs were spread across 25 putative families and varied between RNA and DNA viruses, including previously uncharacterized viral genomes. Of note, airway samples also contained virus(es) characteristic of the human gastrointestinal tract, which have not been previously described as part of the lung virome. Additionally, by performing a meta-analysis of the integrated datasets, ecological trends within viral populations linked to human disease states and their biogeographical distribution along the respiratory tract were observed.
CONCLUSIONS: By leveraging publicly available repositories of shotgun metagenomic data, the present study provides new insights into viral genomes associated with specimens from the human respiratory tract across different disease spectra. Further studies are required to validate our findings and evaluate the potential impact of these viral communities on respiratory tract physiology.

The advancement of bioinformatics and sequencing technology has resulted in the identification of an increasing number of new RNA viruses. This study systematically identified the RNA virome of the willow-carrot aphid, Cavariella aegopodii (Hemiptera: Aphididae), using metagenomic sequencing and rapid amplification of cDNA ends (RACE) approaches. C. aegopodii is a sap-sucking insect widely distributed in Europe, Asia, North America, and Australia. The deleterious effects of C. aegopodii on crop growth primarily stem from its feeding activities and its role as a vector for transmitting plant viruses. The virome includes Cavariella aegopodii virga-like virus 1 (CAVLV1) and Cavariella aegopodii iflavirus 1 (CAIV1). Furthermore, the complete genome sequence of CAVLV1 was obtained. Phylogenetically, CAVLV1 is associated with an unclassified branch of the Virgaviridae family and is susceptible to host antiviral RNA interference (RNAi), resulting in the accumulation of a significant number of 22nt virus-derived small interfering RNAs (vsiRNAs). CAIV1, on the other hand, belongs to the Iflaviridae family, with vsiRNAs ranging from 18 to 22 nt. Our findings present a comprehensive analysis of the RNA virome of C. aegopodii for the first time, offering insights that could potentially aid in the future control of the willow-carrot aphid.

Viral diseases pose a significant threat to tomato crops (Solanum lycopersicum L.), one of the world\'s most economically important vegetable crops. The limited genetic diversity of cultivated tomatoes contributes to their high susceptibility to viral infections. To address this challenge, tomato breeding programs must harness the genetic resources found in native populations and wild relatives. Breeding efforts may aim to develop broad-spectrum resistance against the virome. To identify the viruses naturally infecting 19 advanced lines, derived from native tomatoes, high-throughput sequencing (HTS) of small RNAs and confirmation with PCR and RT-PCR were used. Single and mixed infections with tomato mosaic virus (ToMV), tomato golden mosaic virus (ToGMoV), and pepper huasteco yellow vein virus (PHYVV) were detected. The complete consensus genomes of three variants of Mexican ToMV isolates were reconstructed, potentially forming a new ToMV clade with a distinct 3\' UTR. The absence of reported mutations associated with resistance-breaking to ToMV suggests that the Tm-1, Tm-2, and Tm-22 genes could theoretically be used to confer resistance. However, the high mutation rates and a 63 nucleotide insertion in the 3\' UTR, as well as amino acid mutations in the ORFs encoding 126 KDa, 183 KDa, and MP of Mexican ToMV isolates, suggest that it is necessary to evaluate the capacity of these variants to overcome Tm-1, Tm-2, and Tm-22 resistance genes. This evaluation, along with the characterization of advanced lines using molecular markers linked to these resistant genes, will be addressed in future studies as part of the breeding strategy. This study emphasizes the importance of using HTS for accurate identification and characterization of plant viruses that naturally infect tomato germplasm based on the consensus genome sequences. This study provides crucial insights to select appropriate disease management strategies and resistance genes and guide breeding efforts toward the development of virus-resistant tomato varieties.

Perinereis species are essential benthonic animals in coastal ecosystems and have significant roles as live feed in aquaculture, owing to their high-protein and low-fat nutritional profile. Despite their ecological importance, the viral communities associated with these organisms need to be better understood. In this study, we generated 2.6 × 108 reads using meta-transcriptomic sequencing and de novo assembled 5.3 × 103 virus-associated contigs. We identified 12 novel RNA viruses from two species, Perinereis aibuhitensis and P. wilsoni, which were classified into four major viral groups: Picobirnaviridae, Marnaviridae, unclassified Picornavirales, and unclassified Bunyavirales. Our findings revealed the hidden diversity of viruses and genome structures in Perinereis, enriching the RNA virosphere and expanding the host range of Picobirnaviridae, Marnaviridae, and Bunyavirales. This study also highlighted the potential biosecurity risk of the novel viruses carried by Perinereis to aquaculture.

BACKGROUND: Virome studies on birds, including chickens are relatively scarce, particularly from the African continent. Despite the continuous evolution of RNA viruses and severe losses recorded in poultry from seasonal viral outbreaks, the information on RNA virome composition is even scantier as a result of their highly unstable nature, genetic diversity, and difficulties associated with characterization. Also, information on factors that may modulate the occurrence of some viruses in birds is limited, particularly for domesticated birds. Viral metagenomics through advancements in sequencing technologies, has enabled the characterization of the entire virome of diverse host species using various samples.
METHODS: The complex RNA viral constituents present in 27 faecal samples of asymptomatic chickens from a South African farm collected at 3-time points from two independent seasons were determined, and the impact of the chicken\'s age and collection season on viral abundance and diversity was further investigated. The study utilized the non-invasive faecal sampling method, mRNA viral targeted enrichment steps, a whole transcriptome amplification strategy, Illumina sequencing, and bioinformatics tools.
RESULTS: The results obtained revealed a total of 48 viral species spanning across 11 orders, 15 families and 21 genera. Viral RNA families such as Coronaviridae, Picornaviridae, Reoviridae, Astroviridae, Caliciviridae, Picorbirnaviridae and Retroviridae were abundant, among which picornaviruses, demonstrated a 100% prevalence across the three age groups (2, 4 and 7 weeks) and two seasons (summer and winter) of the 27 faecal samples investigated. A further probe into the extent of variation between the different chicken groups investigated indicated that viral diversity and abundance were significantly influenced by age (P = 0.01099) and season (P = 0.00099) between chicken groups, while there was no effect on viral shedding within samples in a group (alpha diversity) for age (P = 0.146) and season (P = 0.242).
CONCLUSIONS: The presence of an exceedingly varied chicken RNA virome, encompassing avian, mammalian, fungal, and dietary-associated viruses, underscores the complexities inherent in comprehending the causation, dynamics, and interspecies transmission of RNA viruses within the investigated chicken population. Hence, chickens, even in the absence of discernible symptoms, can harbour viruses that may exhibit opportunistic, commensal, or pathogenic characteristics.

BACKGROUND: Pan-virus detection, and virome investigation in general, can be challenging, mainly due to the lack of universally conserved genetic elements in viruses. Metagenomic next-generation sequencing can offer a promising solution to this problem by providing an unbiased overview of the microbial community, enabling detection of any viruses without prior target selection. However, a major challenge in utilising metagenomic next-generation sequencing for virome investigation is that data analysis can be highly complex, involving numerous data processing steps.
RESULTS: Here, we present Entourage to address this challenge. Entourage enables short-read sequence assembly, viral sequence search with or without reference virus targets using contig-based approaches, and intrasample sequence variation quantification. Several workflows are implemented in Entourage to facilitate end-to-end virus sequence detection analysis through a single command line, from read cleaning, sequence assembly, to virus sequence searching. The results generated are comprehensive, allowing for thorough quality control, reliability assessment, and interpretation. We illustrate Entourage\'s utility as a streamlined workflow for virus detection by employing it to comprehensively search for target virus sequences and beyond in raw sequence read data generated from HeLa cell culture samples spiked with viruses. Furthermore, we showcase its flexibility and performance on a real-world dataset by analysing a preassembled Tara Oceans dataset. Overall, our results show that Entourage performs well even with low virus sequencing depth in single digits, and it can be used to discover novel viruses effectively. Additionally, by using sequence data generated from a patient with chronic SARS-CoV-2 infection, we demonstrate Entourage\'s capability to quantify virus intrasample genetic variations, and generate publication-quality figures illustrating the results.
CONCLUSIONS: Entourage is an all-in-one, versatile, and streamlined bioinformatics software for virome investigation, developed with a focus on ease of use. Entourage is available at https://codeberg.org/CENMIG/Entourage under the MIT license.

Historically neglected by microbial ecologists, soil viruses are now thought to be critical to global biogeochemical cycles. However, our understanding of their global distribution, activities and interactions with the soil microbiome remains limited. Here we present the Global Soil Virus Atlas, a comprehensive dataset compiled from 2,953 previously sequenced soil metagenomes and composed of 616,935 uncultivated viral genomes and 38,508 unique viral operational taxonomic units. Rarefaction curves from the Global Soil Virus Atlas indicate that most soil viral diversity remains unexplored, further underscored by high spatial turnover and low rates of shared viral operational taxonomic units across samples. By examining genes associated with biogeochemical functions, we also demonstrate the viral potential to impact soil carbon and nutrient cycling. This study represents an extensive characterization of soil viral diversity and provides a foundation for developing testable hypotheses regarding the role of the virosphere in the soil microbiome and global biogeochemistry.

Wastewater biotreatment systems harbor a rich diversity of microorganisms, and the effectiveness of biotreatment systems largely depends on the activity of these microorganisms. Specifically, viruses play a crucial role in altering microbial behavior and metabolic processes throughout their infection phases, an aspect that has recently attracted considerable interest. Two metagenomic approaches, viral-like particle-concentrated (VPC, representing free viral-like particles) and non-concentrated (NC, representing the cellular fraction), were employed to assess their efficacy in revealing virome characteristics, including taxonomy, diversity, host interactions, lifestyle, dynamics, and functional genes across processing units of three wastewater treatment plants (WWTPs). Our findings indicate that each approach offers unique insights into the viral community and functional composition. Their combined use proved effective in elucidating WWTP viromes. We identified nearly 50,000 viral contigs, with Cressdnaviricota and Uroviricota being the predominant phyla in the VPC and NC fractions, respectively. Notably, two pathogenic viral families, Asfarviridae and Adenoviridae, were commonly found in these WWTPs. We also observed significant differences in the viromes of WWTPs processing different types of wastewater. Additionally, various phage-derived auxiliary metabolic genes (AMGs) were active at the RNA level, contributing to the metabolism of the microbial community, particularly in carbon, sulfur, and phosphorus cycling. Moreover, we identified 29 virus-carried antibiotic resistance genes (ARGs) with potential for host transfer, highlighting the role of viruses in spreading ARGs in the environment. Overall, this study provides a detailed and integrated view of the virosphere in three WWTPs through the application of VPC and NC metagenomic approaches. Our findings enhance the understanding of viral communities, offering valuable insights for optimizing the operation and regulation of wastewater treatment systems.