The COVID-19 pandemic underscored the significance of omics technology and Wastewater-Based Epidemiology for epidemic preparedness. This study investigates the virosphere in wastewater samples from Natal (Brazil), aiming to understand its structure, relationships, and potential. Metaviromic analysis was used on DNA and RNA from weekly samples collected over a year (June/2021 to May/2022) from three wastewater treatment plants. The virosphere showed stability, particularly in viruses infecting microorganisms and plants. However, an alternation of representatives of viruses that infect animals has been observed. Among the most abundant viruses infecting microorganisms are genera associated with the bacterial genera Escherichia, Pseudomonas, and Caulobacte. Regarding the viruses infecting plants, Sobemovirus and Tobamovirus are the most abundant genera. Odontoglossum ringspot virus was identified as a possible RNA virus biomarker. Among DNA viruses infecting animals, genera Bocaparvovirus and Mastadenovirus are the most prevalent. Intriguingly, some Poxviridae family members were observed in the samples. Co-occurrence network analysis identified potential biomarkers like Volepox virus, Anatid herpesvirus 1, and Caviid herpesvirus 2. Among RNA viruses affecting animals, Mamastrovirus, Rotavirus, and Norovirus genera were the most abundant pathogens. Furthermore, members of the Coronaviridae family exhibited a high degree of centrality values in the co-occurrence network, even connecting with unclassified viruses. The study emphasizes the importance of research in understanding the roles of unclassified viruses. In addition, we observed an association between Coronaviridae reads, rainfall, and the number of reported COVID-19 cases. Our study highlights the diversity and complexity of the viral community in wastewater and the need for research to understand better the ecological roles unclassified viruses play. Such advances will significantly contribute to our preparedness and response to future viral threats. Furthermore, our study contributes to knowledge of virosphere dynamics, offering insights that can contribute to the direction of future public health policies and interventions.

Genetic variations are instrumental for unraveling phage evolution and deciphering their functional implications. Here, we explore the underlying fine-scale genetic variations in the gut phageome, especially structural variations (SVs). By using virome-enriched long-read metagenomic sequencing across 91 individuals, we identified a total of 14,438 nonredundant phage SVs and revealed their prevalence within the human gut phageome. These SVs are mainly enriched in genes involved in recombination, DNA methylation, and antibiotic resistance. Notably, a substantial fraction of phage SV sequences share close homology with bacterial fragments, with most SVs enriched for horizontal gene transfer (HGT) mechanism. Further investigations showed that these SV sequences were genetic exchanged between specific phage-bacteria pairs, particularly between phages and their respective bacterial hosts. Temperate phages exhibit a higher frequency of genetic exchange with bacterial chromosomes and then virulent phages. Collectively, our findings provide insights into the genetic landscape of the human gut phageome.

Immune-mediated gastrointestinal (GI) diseases, including achalasia, celiac disease, and inflammatory bowel diseases, pose significant challenges in diagnosis and management due to their complex etiology and diverse clinical manifestations. While genetic predispositions and environmental factors have been extensively studied in the context of these conditions, the role of viral infections and virome dysbiosis remains a subject of growing interest. This review aims to elucidate the involvement of viral infections in the pathogenesis of immune-mediated GI diseases, focusing on achalasia and celiac disease, as well as the virome dysbiosis in IBD. Recent evidence suggests that viral pathogens, ranging from common respiratory viruses to enteroviruses and herpesviruses, may trigger or exacerbate achalasia and celiac disease by disrupting immune homeostasis in the GI tract. Furthermore, alterations in the microbiota and, specifically, in the virome composition and viral-host interactions have been implicated in perpetuating chronic intestinal inflammation in IBD. By synthesizing current knowledge on viral contributions to immune-mediated GI diseases, this review aims to provide insights into the complex interplay between viral infections, host genetics, and virome dysbiosis, shedding light on novel therapeutic strategies aimed at mitigating the burden of these debilitating conditions on patients\' health and quality of life.

BACKGROUND: Pneumonia is typically caused by a variety of pathogenic microorganisms. Traditional research often focuses on the infection of a few microorganisms, whereas metagenomic studies focus on the impact of the bacteriome and mycobiome on respiratory diseases. Reports on the virome characteristics of pediatric pneumonia remain relatively scarce.
METHODS: We employed de novo assembly and combined homology- and feature-based methods to characterize the respiratory virome in whole-genome DNA sequencing samples from oropharynx (OP) swabs, nasopharynx (NP) swabs, and bronchoalveolar lavage fluids (BALF) of children with pneumonia.
RESULTS: Significant differences were observed in the alpha and beta diversity indexes, as well as in the composition of the oropharyngeal virome, between pneumonia cases and controls. We identified 1137 viral operational taxonomic units (vOTUs) with significant differences, indicating a preference of pneumonia-reduced vOTUs for infecting Prevotella, Neisseria, and Veillonella, whereas pneumonia-enriched vOTUs included polyomavirus, human adenovirus, and phages targeting Staphylococcus, Streptococcus, Granulicatella, and Actinomyces. Comparative analysis revealed higher relative abundances and prevalence rates of pneumonia-enriched OP vOTUs in NP and BALF samples compared to pneumonia-reduced vOTUs. Additionally, virome analysis identified six pediatric patients with severe human adenovirus or polyomavirus infections, five of whom might have been undetected by targeted polymerase chain reaction (PCR)-based testing.
CONCLUSIONS: This study offers insights into pediatric pneumonia respiratory viromes, highlighting frequent transmission of potentially pathogenic viruses and demonstrating virome analysis as a valuable adjunct for pathogen detection.

Shrews being insectivores, serve as natural reservoirs for a wide array of zoonotic viruses, including the recently discovered Langya henipavirus (LayV) in China in 2018. It is crucial to understand the shrew-associated virome, viral diversity, and new viruses. In the current study, we conducted high-throughput sequencing on lung samples obtained from 398 shrews captured along the eastern coast of China, and characterized the high-depth virome of 6 common shrew species (Anourosorex squamipes, Crocidura lasiura, Crocidura shantungensis, Crocidura tanakae, Sorex caecutiens, and Suncus murinus). Our analysis revealed numerous shrew-associated viruses comprising 54 known viruses and 72 new viruses that significantly enhance our understanding of mammalian viruses. Notably, 34 identified viruses possess spillover-risk potential and six were human pathogenic viruses: LayV, influenza A virus (H5N6), rotavirus A, rabies virus, avian paramyxovirus 1, and rat hepatitis E virus. Moreover, ten previously unreported viruses in China were discovered, six among them have spillover-risk potential. Additionally, all 54 known viruses and 12 new viruses had the ability to cross species boundaries. Our data underscore the diversity of shrew-associated viruses and provide a foundation for further studies into tracing and predicting emerging infectious diseases originated from shrews.

Viruses as the prevailing biological entities are poorly understood in underground realms. Here, we establish the first metagenomic Groundwater Virome Catalogue (GWVC) comprising 280,420 viral species ( ≥ 5 kb) detected from 607 monitored wells in seven geo-environmental zones throughout China. In expanding ~10-fold the global portfolio of known groundwater viruses, we uncover over 99% novel viruses and about 95% novel viral clusters. By linking viruses to hosts from 119 prokaryotic phyla, we double the number of microbial phyla known to be virus-infected in groundwater. As keystone ultrasmall symbionts in aquifers, CPR bacteria and DPANN archaea are susceptible to virulent viruses. Certain complete CPR viruses even likely infect non-CPR bacteria, while partial CPR/DPANN viruses harbor cell-surface modification genes that assist symbiont cell adhesion to free-living microbes. This study reveals the unknown viral world and auxiliary metabolism associated with methane, nitrogen, sulfur, and phosphorus cycling in groundwater, and highlights the importance of subsurface virosphere in viral ecology.

While the viromes and immune systems of bats and rodents have been extensively studied, comprehensive data are lacking for insectivores (order Eulipotyphla) despite their wide geographic distribution. Anthropogenic land use and outdoor recreational activities, as well as changes in the range of shrews, may lead to an expansion of the human-shrew interface with the risk of spillover infections, as reported for Borna disease virus 1. We investigated the virome of 45 individuals of 4 white-toothed shrew species present in Europe, using metagenomic RNA sequencing of tissue and intestine pools. Moderate to high abundances of sequences related to the families Paramyxoviridae, Nairoviridae, Hepeviridae and Bornaviridae were detected. Whole genomes were determined for novel orthoparamyxoviruses (n=3), orthonairoviruses (n=2) and an orthohepevirus. The novel paramyxovirus, tentatively named Hasua virus, was phylogenetically related to the zoonotic Langya virus and Mòjiāng virus. The novel orthonairoviruses, along with the potentially zoonotic Erve virus, fall within the shrew-borne Thiafora virus genogroup. The highest viral RNA loads of orthoparamyxoviruses were detected in the kidneys, in well-perfused organs for orthonairoviruses and in the liver and intestine for orthohepevirus, indicating potential transmission routes. Notably, several shrews were found to be coinfected with viruses from different families. Our study highlights the virus diversity present in shrews, not only in biodiversity-rich regions but also in areas influenced by human activity. This study warrants further research to characterize and assess the clinical implications and risk of these viruses and the importance of shrews as reservoirs in European ecosystems.

Diagnosis of acute respiratory infections (ARIs) is challenging due to the broad diversity of potential microbial causes. We used metagenomic next-generation sequencing (mNGS) to analyze the nasopharyngeal virome of ARI patients, who had undergone testing with a clinical multiplex PCR panel (Amplisens ARVI-screen-FRT). We collected nasopharyngeal swabs from 49 outpatient adults, 32 of whom had ARI symptoms and were PCR-positive, and 4 asymptomatic controls in Kazakhstan during Spring 2021. We assessed the biodiversity of the mNGS-derived virome and concordance with PCR results. PCR identified common ARI viruses in 65% of the symptomatic cases. mNGS revealed viral taxa consisting of human, non-human eukaryotic and bacteriophage groups, comprising 15, 11 and 28 genera, respectively. Notable ARI-associated human viruses included rhinovirus (16.3%), betaherpesvirus 7 (14.3%) and Epstein-Barr virus (8.16%). The primary phage hosts were Streptococcus spp. (32.7%), Pseudomonas aeruginosa (24.5%) and Burkholderia spp. (20.4%). In total, 47% of ARIs were linked solely to bacterial pathogens, a third to viral-bacterial co-infections, and less than 10% to only viral infections by mNGS. PCR showed low concordance with mNGS, except for rhinovirus. These results underscore the importance of broad diagnostic methods and question the effectiveness of commonly used PCR panels in ARI diagnosis.

Co-composting with exogenous microbial inoculant, presents an effective approach for the harmless utilization of livestock manure and agroforestry wastes. However, the impact of inoculant application on the variations of viral and antibiotic resistance genes (ARGs) remains poorly understood, particularly under varying manure quantity (low 10 % vs. high 20 % w/w). Thus, employing virome and metagenomic sequencing, we examined the influence of Streptomyces-Bacillus Inoculants (SBI) on viral communities, phytopathogen, ARGs, mobile genetic elements, and their interrelations. Our results indicate that SBI shifted dominant bacterial species from Phenylobacterium to thermotropic Bordetella, and the quantity of manure mediates the effect of SBI on whole bacterial community. Major ARGs and genetic elements experienced substantial changes with SBI addition. There was a higher ARGs elimination rate in the composts with low (∼76 %) than those with high manure (∼70 %) application. Virus emerged as a critical factor influencing ARG dynamics. We observed a significant variation in virus community, transitioning from Gemycircularvirus- (∼95 %) to Chlamydiamicrovirus-dominance. RDA analysis revealed that Gemycircularvirus was the most influential taxon in shaping ARGs, with its abundance decreased approximately 80 % after composting. Collectively, these findings underscore the role of microbial inoculants in modulating virus communities and ARGs during biowaste co-composting.

Viruses are core components of the human microbiome, impacting health through interactions with gut bacteria and the immune system. Most human microbiome viruses are bacteriophages, which exclusively infect bacteria. Until recently, most gut virome studies focused on low taxonomic resolution (e.g., viral operational taxonomic units), hampering population-level analyses. We previously identified an expansive and widespread bacteriophage lineage in inhabitants of Amsterdam, the Netherlands. Here, we study their biodiversity and evolution in various human populations. Based on a phylogeny using sequences from six viral genome databases, we propose the Candidatus order Heliusvirales. We identify heliusviruses in 82% of 5441 individuals across 39 studies, and in nine metagenomes from humans that lived in Europe and North America between 1000 and 5000 years ago. We show that a large lineage started to diversify when Homo sapiens first appeared some 300,000 years ago. Ancient peoples and modern hunter-gatherers have distinct Ca. Heliusvirales populations with lower richness than modern urbanized people. Urbanized people suffering from type 1 and type 2 diabetes, as well as inflammatory bowel disease, have higher Ca. Heliusvirales richness than healthy controls. We thus conclude that these ancient core members of the human gut virome have thrived with increasingly westernized lifestyles.