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Zoonotic viruses are widely seen as the primary threat for future pandemics. Bats are the most diverse group of mammals, with more than 1400 species distributed across most habitats on Earth. So far, 31 known virus families were associated with bats, although the understanding of most viruses were insufficient. Continuous efforts to discover, understand and monitor these bats viruses, is thereby an area of public health interest. This systematic review was designed to catalogue publications reporting novel bat virus discoveries within PubMed, SCOPUS, and Web of Science databases, within a 5-year period from 2018 to 2022. Various experimental parameters, including sampling locations, methodology, bat species diversity, similarity to known viruses, species demarcation of new viruses, and genomic sequencing strategies, were extracted from 41 publications and analyzed. In total, 72 novel viruses from 19 virus families were identified between 2018 and 2022, particularly from Genomoviridae (DNA viruses) and Coronaviridae (RNA viruses). That said, only a limited number of bat families featured extensively despite noticeable shift towards next generation sequencing methods and metagenomics pipeline for virus identification across different sampling methods. This review aims to provide a comprehensive analysis of the global efforts made over the past five years to identify and characterize emerging viruses in bat species, and to provide a detailed overview of the current technologies and methodologies used in these studies., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

The potato leafhopper ( Empoasca fabae , PLH) is a serious pest that feeds on a wide range of agricultural crops and is found throughout the United States but is not known to be a vector for plant-infecting viruses. We probed the diversity of virus sequences in field populations of PLH collected from four Midwestern states: Illinois, Indiana, Iowa, and Minnesota. High-throughput sequencing data from total RNAs extracted from PLH were used to assemble sequences of fifteen positive-stranded RNA viruses, two negative-stranded RNA viruses, and one DNA virus. These sequences included ten previously described plant viruses and eight putative insect-infecting viruses. All but one of the insect-specific viruses were novel and included three solemoviruses, one iflavirus, one phenuivirus, one lispivirus, and one ambidensovirus. Detailed analyses of the novel genome sequences and their evolutionary relationships with related family members were conducted. Our study revealed a diverse group of plant viruses circulating in the PLH population and discovered novel insect viruses, expanding knowledge on the untapped virus diversity in economically important crop pests. Our findings also highlight the importance of monitoring the emergence and circulation of plant-infecting viruses in agriculturally important arthropod pests.

Bats are natural hosts of multiple viruses, many of which have clear zoonotic potential. The search for emerging viruses has been aided by the implementation of metagenomic tools, which have also enabled the detection of unprecedented viral diversity. Currently, this search is mainly focused on RNA viruses, which are largely over-represented in databases. To compensate for this research bias, we analyzed fecal samples from 189 Spanish bats belonging to 22 different species using viral metagenomics. This allowed us to identify 52 complete or near-complete viral genomes belonging to the families Adenoviridae , Circoviridae , Genomoviridae , Papillomaviridae , Parvoviridae, Polyomaviridae and Smacoviridae . Of these, 30 could constitute new species, doubling the number of viruses currently described in Europe. These findings open the door to a more thorough analysis of bat DNA viruses and their zoonotic potential., Importance: Metagenomics has become a fundamental tool to characterize the global virosphere, allowing us not only to understand the existing viral diversity and its ecological implications but also to identify new and emerging viruses. RNA viruses have a higher zoonotic potential, but this risk is also present for some DNA virus families. In our study, we analyzed the DNA fraction of fecal samples from 22 Spanish bat species, identifying 52 complete or near-complete genomes of different viral families with zoonotic potential. This doubles the number of genomes currently described in Europe. Metagenomic data often produce partial genomes that can be difficult to analyze. Our work, however, has characterized a large number of complete genomes, thus facilitating their taxonomic classification and enabling different analyses to be carried out to evaluate their zoonotic potential. For example, recombination studies are relevant since this phenomenon could play a major role in cross-species transmission., Competing Interests: The authors declare no conflict of interest.

Acute encephalitis syndrome (AES) causes significant morbidity and mortality worldwide. In Nepal, Japanese encephalitis virus (JEV) accounts for ~5-20% of AES cases, but ~75% of AES cases are of unknown etiology. We identified a gemykibivirus in CSF collected in 2020 from an 8-year-old male patient with AES using metagenomic next-generation sequencing. Gemykibiviruses are single stranded, circular DNA viruses in the family Genomoviridae . The complete genome of 2,211 nucleotides was sequenced, which shared 98.69% nucleotide identity to its closest relative, Human associated gemykibivirus 2 isolate SAfia-449D. Two real-time PCR assays were designed, and screening of 337 cerebrospinal fluid (CSF) and 164 serum samples from AES patients in Nepal collected in 2020 and 2022 yielded 11 CSF and 1 serum sample that were positive in both PCR assays. Complete genomes of seven of the positives were sequenced. These results identify a potential candidate etiologic agent of encephalitis in Nepal., Importance: Viral encephalitis is a devastating disease, but unfortunately, worldwide, the causative virus in many cases is unknown. Therefore, it is important to identify viruses that could be responsible for cases of human encephalitis. Here, using metagenomic sequencing of CSF, we identified a gemykibivirus in a male child from Nepal with acute encephalitis syndrome (AES). We subsequently detected gemykibivirus DNA in CSF or serum of 12 more encephalitis patients by real-time PCR. The virus genomes we identified are highly similar to gemykibiviruses previously detected in CSF of three encephalitis patients from Sri Lanka. These results raise the possibility that gemykibivirus could be an underrecognized human pathogen., Competing Interests: The authors declare no conflict of interest.

Background: Cutavirus (CuV) is associated with mycosis fungoides; however, the CuV status in parapsoriasis en plaques (PP), a premalignant inflammatory condition of mycosis fungoides, has not been fully delineated., Methods: Fifty-five Japanese patients with chronic inflammatory skin diseases, including 13 patients with PP, were studied., Results: CuV DNA was detected significantly more frequently in biopsies of the lesional skin from patients with PP (38%; 4 of 13) than in those from patients with other inflammatory skin diseases (2%; 1 of 42; P = .009). All CuV-positive PP cases were of the large-plaque parapsoriasis (LPP) subtype. The viral loads ranged from 83 450 to 2 164 170 copies/103 cells. We recovered near-full-length CuV sequences from the CuV-positive LPP biopsies, all of which were of the Japanese/Asian genotype. The CuV genome appeared to be present within lymphoid cells infiltrating the epidermis and dermis. CuV NS1 and VP1 gene transcripts were also detected in the affected tissues., Conclusions: The detection of high levels of CuV DNA with the expression of viral mRNA suggests a potential role for CuV in the pathogenesis of LPP, making it necessary to study further the impact of CuV, especially regarding the viral genotype, on the outcomes of patients with CuV-positive LPP., Competing Interests: Potential conflicts of interest. All authors: No reported conflicts. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed., (© The Author(s) 2023. Published by Oxford University Press on behalf of Infectious Diseases Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Background: Haematological patients exhibit immune system abnormalities that make them susceptible to viral infections. Understanding the relationship between the virome in the blood plasma of haematological patients and their clinical characteristic is crucial for disease management. We aimed to explore the presence of viral pathogens and identify close associations between viral infections and various clinical features., Results: A total of 21 DNA viruses and 6 RNA viruses from 12 virus families were identified from 1383 patients. Patients with haematological diseases exhibited significantly higher diversity, prevalence, and co-detection rates of viral pathogens. During fever episodes, pathogen detection was notably higher, with Epstein-Barr virus (EBV) and Mucorales infections being the most probable culprits for fever symptoms in non-haematological patients. The detection rate of torque teno virus (TTV) significantly increases in haematological patients after transplantation and during primary lung infections. Additionally, TTV-positive patients demonstrate significantly higher absolute neutrophil counts, while C-reactive protein and procalcitonin levels are notably lower. Furthermore, TTV, cytomegalovirus, and parvovirus B19 (B19V) were found to be more prevalent in non-neutropenic patients, while non-viral pathogenic infections, such as Gram-negative bacteria and Mucorales, were more common in neutropenic patients. Pegivirus C (HPgV-C) infection often occurred post-transplantation, regardless of neutropenia. Additionally, some viruses such as TTV, B19V, EBV, and HPgV-C showed preferences for age and seasonal infections., Conclusions: Analysis of the plasma virome revealed the susceptibility of haematological patients to plasma viral infections at specific disease stages, along with the occurrence of mixed infections with non-viral pathogens. Close associations were observed between the plasma virome and various clinical characteristics, as well as clinical detection parameters. Understanding plasma virome aids in auxiliary clinical diagnosis and treatment, enabling early prevention to reduce infection rates in patients and improve their quality of life. Video Abstract., (© 2024. The Author(s).)

The Phylum Cressdnaviricota consists of a large number of circular Rep-encoding single-stranded (CRESS)-DNA viruses. Recently, metagenomic analyzes revealed their ubiquitous distribution in a diverse range of eukaryotes. Data relating to CRESS-DNA viruses in humans remains scarce. Our study investigated the presence and genetic diversity of CRESS-DNA viruses in human vaginal secretions. Vaginal swabs were collected from 28 women between 29 and 43 years old attending a fertility clinic in New York City. An exploratory metagenomic analysis was performed and detection of CRESS-DNA viruses was confirmed through analysis of near full-length sequences of the viral isolates. A phylogenetic tree was based on the REP open reading frame sequences of the CRESS-DNA virus genome. Eleven nearly complete CRESS-DNA viral genomes were identified in 16 (57.1%) women. There were no associations between the presence of these viruses and any demographic or clinical parameters. Phylogenetic analysis indicated that one of the sequences belonged to the genus Gemycircularvirus within the Genomoviridae family, while ten sequences represented previously unclassified species of CRESS-DNA viruses. Novel species of CRESS-DNA viruses are present in the vaginal tract of adult women. Although they be transient commensal agents, the potential clinical implications for their presence at this site cannot be dismissed., (© 2024 Wiley Periodicals LLC.)

As part of a sea turtle health monitoring program on the central east coast of Queensland, Australia, stranded and sick green sea turtles ( Chelonia mydas ) were subjected to necropsy and histopathology. A subset of these turtles had myocarditis of varying severity, which could not be attributed to parasitism by spirorchid flukes or bacterial infections. We, therefore, undertook an investigation to determine whether virus infections might be part of the pathogenesis. Deep sequencing revealed abundant DNA virus contigs in the heart tissue, of which CRESS and circoviruses appeared to be the most consistently present. Further analysis revealed the homology of some of the circoviruses to the beak and feather disease virus. While a causative link to myocarditis could not be established, the presence of these viruses may play a contributing role by affecting the immune system and overall health of animals exposed to pollutants, higher water temperatures, and decreasing nutrition.

Apis mellifera filamentous virus (AmFV) is a double-stranded DNA virus that infects Apis mellifera bees. To our knowledge, this is the first comprehensive study aiming to detect and analyse the genetic diversity and prevalence of AmFV in Korean honeybee colonies. Phylogenetic analysis based on baculovirus repeat open reading frame-N gene (Bro-N) sequences revealed that AmFV isolates from the Republic of Korea (ROK) fell into two distinct lineages, with genetic origins in Switzerland and China, with nucleotide similarities of 98.3% and 98.2%, respectively. Our prevalence analysis demonstrated a noteworthy infection rate of AmFV in 545 honeybee colonies, reaching 33.09% in 2022 and increasing to 44.90% by 2023. Intriguingly, we also detected AmFV in Varroa destructor mites, highlighting their potential role as vectors and carriers of AmFV. The presence of AmFV was correlated with an increased infection rate of sacbrood virus, deformed wing virus, Lake Sinai virus 2, black queen cell virus, and Nosema ceranae in honeybee colonies. These findings provide valuable insight into the prevalence and potential transmission mechanisms of AmFV in honeybee colonies in the ROK. The results of this study may be instrumental in the effective management of viral infections in honeybee apiaries., (© 2024. The Author(s).)

The FTA card has emerged as a promising alternative for nucleic acid extraction. The FTA card is a filter paper impregnated with chemicals that preserve and stabilize the genetic material present in the sample, allowing for its storage and transport at room temperature. The aim of this study was to test the card for the detection of RNA and DNA nucleic acids. Two RNA viruses (Senecavirus A and classical swine fever virus) and two DNA viruses (African swine fever virus and suid alphaherpesvirus 1) were tested, and in all cases, there was a decrease in sensitivity. The methods exhibited good repeatability and demonstrated a rapid and practical use for sample transport and nucleic acid extraction., (© 2024. The Author(s) under exclusive licence to Sociedade Brasileira de Microbiologia.)

DNA viruses that produce persistent infections have been proposed as potential causes for the extinction of Neanderthals, and, therefore, the identification of viral genome remnants in Neanderthal sequence reads is an initial step to address this hypothesis. Here, as proof of concept, we searched for viral remnants in sequence reads of Neanderthal genome data by mapping to adenovirus, herpesvirus and papillomavirus, which are double-stranded DNA viruses that may establish lifelong latency and can produce persistent infections. The reconstructed ancient viral genomes of adenovirus, herpesvirus and papillomavirus revealed conserved segments, with nucleotide identity to extant viral genomes and variable regions in coding regions with substantial divergence to extant close relatives. Sequence reads mapped to extant viral genomes showed deamination patterns of ancient DNA, and these ancient viral genomes showed divergence consistent with the age of these samples (≈50,000 years) and viral evolutionary rates (10 -5 to 10 -8 substitutions/site/year). Analysis of random effects showed that the Neanderthal mapping to genomes of extant persistent viruses is above what is expected by random similarities of short reads. Also, negative control with a nonpersistent DNA virus does not yield statistically significant assemblies. This work demonstrates the feasibility of identifying viral genome remnants in archaeological samples with signal-to-noise assessment.

The phylum Cressdnaviricota comprises viruses with single-stranded, circular DNA genomes that encode an HUH-type endonuclease (known as Rep). The phylum includes two classes, eight orders, and 11 families. Here, we report the creation of a twelfth family in the order Mulpavirales, class Arfiviricetes of the phylum Cressdnaviricota. The family Amesuviridae comprises viruses that infect plants and is divided into two genera: Temfrudevirus, including the species Temfrudevirus temperatum (with temperate fruit decay-associated virus as a member), and Yermavirus, including the species Yermavirus ilicis (with yerba mate-associated circular DNA virus as a member). Both viruses encode Rep proteins with HUH endonuclease and SH3 superfamily helicase domains. Phylogenetic analysis indicates that the replicative module of amesuviruses constitutes a well-supported monophyletic clade related to Rep proteins from viruses in the order Mulpavirales. Furthermore, both viruses encode a single capsid protein (CP) related to geminivirus CPs. Phylogenetic incongruence between the replicative and structural modules of amesuviruses suggests a chimeric origin resulting from remote recombination events between ancestral mulpavirales and geminivirids. The creation of the family Amesuviridae has been ratified by the International Committee on Taxonomy of Viruses (ICTV)., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Austria, part of Springer Nature.)

CrAssphage and related viruses of the order Crassvirales (hereafter referred to as crassviruses) were originally discovered by cross-assembly of metagenomic sequences. They are the most abundant viruses in the human gut, are found in the majority of individual gut viromes, and account for up to 95% of the viral sequences in some individuals 1-4 . Crassviruses are likely to have major roles in shaping the composition and functionality of the human microbiome, but the structures and roles of most of the virally encoded proteins are unknown, with only generic predictions resulting from bioinformatic analyses 4,5 . Here we present a cryo-electron microscopy reconstruction of Bacteroides intestinalis virus ΦcrAss001 6 , providing the structural basis for the functional assignment of most of its virion proteins. The muzzle protein forms an assembly about 1 MDa in size at the end of the tail and exhibits a previously unknown fold that we designate the 'crass fold', that is likely to serve as a gatekeeper that controls the ejection of cargos. In addition to packing the approximately 103 kb of virus DNA, the ΦcrAss001 virion has extensive storage space for virally encoded cargo proteins in the capsid and, unusually, within the tail. One of the cargo proteins is present in both the capsid and the tail, suggesting a general mechanism for protein ejection, which involves partial unfolding of proteins during their extrusion through the tail. These findings provide a structural basis for understanding the mechanisms of assembly and infection of these highly abundant crassviruses., (© 2023. The Author(s).)

The South Island robin (Petroica australis) is a small passerine bird endemic to New Zealand (Aotearoa). Although its population has declined recently and it is considered 'at risk,' little research has been done to identify viruses in this species. This study aimed to survey the diversity of single-stranded DNA viruses associated with South Island robins in a small, isolated population on Nukuwaiata Island. In total, 108 DNA viruses were identified from pooled fecal samples collected from 38 individual robins sampled. These viruses belong to the Circoviridae (n = 10), Genomoviridae (n = 12), and Microviridae (n = 73) families. A number of genomes that belong to the phylum Cressdnaviricota but are otherwise unclassified (n = 13) were also identified. These results greatly expand the known viral diversity associated with South Island robins, and we identify a novel group of viruses most closely related genomoviruses., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Bovine serum has been widely used as a universal supplement in culture media and other applications, including the manufacture of biological products and the production of synthetic meat. Currently, commercial bovine serum is tested for possible viral contaminants following regional guidelines. Regulatory agencies' established tests focused on detecting selected animal origin viruses and are based on virus isolation, immunofluorescence, and hemadsorption assays. However, these tests may fail to detect new or emerging viruses in biological products. High-throughput sequencing is a powerful option since no prior knowledge of the viral targets is required. In the present study, we evaluate the virome of seven commercial batches of bovine serum from Mexico (one batch), New Zealand (two batches), and the United States (four batches) using a specific preparation and enrichment method for pooled samples and sequencing using an Illumina platform. A variety of circular replicase-encoding single-stranded (CRESS) DNA families ( Genomoviridae , Circoviridae , and Smacoviridae ) was identified. Additionally, CrAssphage, a recently discovered group of bacteriophage correlated with fecal contamination, was identified in 85% of the tested batches. Furthermore, sequences representing viral families with single-stranded DNA ( Parvoviridae ), double-stranded DNA ( Polyomaviridae and Adenoviridae ), single-stranded RNA ( Flaviviridae , Picornaviridae , and Retroviridae ), and double-stranded RNA ( Reoviridae ) were identified. These results support that high-throughput sequencing associated with viral enrichment is a robust tool and should be considered an additional layer of safety when testing pooled biologicals to detect viral contaminants overlooked by the current testing protocols.

Viral metagenomic studies have enabled the discovery of many unknown viruses and revealed that viral communities are much more diverse and ubiquitous than previously thought. Some viruses have multiple genome components that are encapsidated either in separate virions (multipartite viruses) or in the same virion (segmented viruses). In this study, we identify what is possibly a novel bipartite plant-associated circular single-stranded DNA virus in a wild prickly pear cactus, Opuntia discolor , that is endemic to the Chaco ecoregion in South America. Two ~1.8 kb virus-like circular DNA components were recovered, one encoding a replication-associated protein (Rep) and the other a capsid protein (CP). Both of the inferred protein sequences of the Rep and CP are homologous to those encoded by members of the family Geminiviridae . These two putatively cognate components each have a nonanucleotide sequence within a likely hairpin structure that is homologous to the origins of rolling-circle replication (RCR), found in diverse circular single-stranded DNA viruses. In addition, the two components share similar putative replication-associated iterative sequences (iterons), which in circular single-stranded DNA viruses are important for Rep binding during the initiation of RCR. Such molecular features provide support for the possible bipartite nature of this virus, which we named utkilio virus (common name of the Opuntia discolor in South America) components A and B. In the infectivity assays conducted in Nicotiana benthamiana plants, only the A component of utkilio virus, which encodes the Rep protein, was found to move and replicate systemically in N. benthamiana . This was not true for component B, for which we did not detect replication, which may have been due to this being a defective molecule or because of the model plants ( N. benthamiana ) used for the infection assays. Future experiments need to be conducted with other plants, including O. discolor , to understand more about the biology of these viral components.

It is widely recognized that pathogens can be transmitted across the placenta from mother to foetus. Recent re-evaluation of metagenomic studies indicates that the placenta has no unique microbiome of commensal bacteria. However, viral transmission across the placenta, including transmission of DNA viruses such as the human herpesviruses, is possible. A fuller understanding of which DNA virus sequence can be found in the placenta is required. We employed a metagenomic analysis to identify viral DNA sequences in placental metagenomes from full-term births (20 births), pre-term births (13 births), births from pregnancies associated with antenatal infections (12 births) or pre-term births with antenatal infections (three births). Our analysis found only a small number of DNA sequences corresponding to the genomes of human herpesviruses in four of the 48 metagenomes analysed. Therefore, our data suggest that DNA virus infection of the placenta is rare and support the concept that the placenta is largely free of pathogen infection.

Recently, the culture of American eels (Anguilla rostrate) in China has been impacted by emergence of a disease with signs of haemorrhagic gill necrosis. The gills of diseased eels are covered with petecchia and they bleed when the operculum is pressed. In this study, a novel American eel adomavirus (AEAdoV) was isolated from the diseased eels using the eel ovary cell line (EO). The virus proliferated in the EO cells with a maximum TCID 50 /ml of 10 6.29 ± 0.23 at 6 days post-infection. The virions were non-enveloped with a diameter of 75-85 nm and shown to be a DNA virus upon 5-iodo-2'-deoxyuridine (IDU) treatment. PCR assays showed that AEAdoV encodes a superfamily 3 helicases (S3H) replicase and shared high similarities with Anguilla marmorata adomavirus (MEAdoV). Although no clinical signs or mortality was observed among the eels injected with AEAdoV, the virus was reisolated from livers, kidneys and gills of injected eels at 35 days post-injection. Our results suggested that AEAdoV exhibited a latent infection in A. rostrata. The pathogenicity of the AEAdoV needs to be confirmed further., (© 2021 John Wiley & Sons Ltd.)

Metagenomic next-generation sequencing has transformed the discovery and diagnosis of infectious disease, with the power to characterise the complete 'infectome' (bacteria, viruses, fungi, parasites) of an individual host organism. However, the identification of novel pathogens has been complicated by widespread microbial contamination in commonly used laboratory reagents. Using total RNA sequencing ("metatranscriptomics") we documented the presence of contaminant viral sequences in multiple 'blank' negative control sequencing libraries that comprise a sterile water and reagent mix. Accordingly, we identified 14 viral sequences in 7 negative control sequencing libraries. As in previous studies, several circular replication-associated protein encoding (CRESS) DNA virus-like sequences were recovered in the blank control libraries, as well as contaminating sequences from the Totiviridae , Tombusviridae and Lentiviridae families of RNA virus. These data suggest that viral contamination of common laboratory reagents is likely commonplace and can comprise a wide variety of viruses.

Calf diarrhoea has been a major cause of economic losses in the global dairy industry. Many factors, including multiple pathogen infections, can directly or indirectly cause calf diarrhoea. This study compared the faecal virome between 15 healthy calves and 15 calves with diarrhoea. Significantly lower diversity of viruses was found in samples from animals with diarrhoea than those in the healthy ones, and this feature may also be related to the age of the calves. Viruses belonging to the families Astroviridae and Caliciviridae that may cause diarrhoea in dairy calves have been characterized, which revealed that reads of caliciviruses and astroviruses in diarrhoea calves were much higher than those in healthy calves. Five complete genomic sequences closely related to Smacoviridae have been identified, which may participate in the regulation of the gut virus community ecology of healthy hosts together with bacteriophages. This research provides a theoretical basis for further understanding of known or potential enteric pathogens related to calf diarrhoea.

The spread, emergence, and adaptation of pathogens causing marine disease has been problematic to fisheries and aquaculture industries for the last several decades creating the need for strategic management and biosecurity practices. The Pacific oyster (Crassostrea gigas), a highly productive species globally, has been a target of disease and mortality caused by a viral pathogen, the Ostreid herpesvirus 1 (OsHV-1) and its microvariants (OsHV-1 µvars). During routine surveillance to establish health history at a shellfish aquaculture nursery system in San Diego, California, the presence of OsHV-1 in Pacific oyster juveniles was detected. Quantification of OsHV-1 in tissues of oysters revealed OsHV-1 viral loads > 10 6 copies/mg. We characterized and identified the OsHV-1 variant by sequencing of ORFs 4 (C2/C6) and 43 (IA1/IA2), which demonstrated that this variant is a novel OsHV-1 microvariant: OsHV-1 µvar SD. A pilot transmission study indicates that OsHV-1 µvar SD is infectious with high viral loads ~ 7.57 × 10 6 copies/mg detected in dead individuals. The detection of OsHV-1 µvar SD in a large port mirrors previous studies conducted in Australia where aquaculture farms and feral populations near port locations may be at a higher risk of OsHV-1 emergence. Further research is needed to understand the impacts of OsHV-1 µvar SD, such as transmission studies focusing on potential vectors and characterization of virulence as compared to other OsHV-1 µvars. To increase biosecurity of the global aquaculture industry, active and passive surveillance may be necessary to reduce spread of pathogens and make appropriate management decisions., (Copyright © 2021. Published by Elsevier Inc.)

Soil is known to harbor viruses, but the majority are uncharacterized and their responses to environmental changes are unknown. Here, we used a multi-omics approach (metagenomics, metatranscriptomics and metaproteomics) to detect active DNA viruses and RNA viruses in a native prairie soil and to determine their responses to extremes in soil moisture. The majority of transcribed DNA viruses were bacteriophage, but some were assigned to eukaryotic hosts, mainly insects. We also demonstrated that higher soil moisture increased transcription of a subset of DNA viruses. Metaproteome data validated that the specific viral transcripts were translated into proteins, including chaperonins known to be essential for virion replication and assembly. The soil viral chaperonins were phylogenetically distinct from previously described marine viral chaperonins. The soil also had a high abundance of RNA viruses, with highest representation of Reoviridae. Leviviridae were the most diverse RNA viruses in the samples, with higher amounts in wet soil. This study demonstrates that extreme shifts in soil moisture have dramatic impacts on the composition, activity and potential functions of both DNA and RNA soil viruses., (© 2021. The Author(s).)

Diarrhoea and poor growth among growing pigs is responsible for significant economic losses in pig herds globally and can have a wide range of possible aetiologies. Next generation sequencing (NGS) technologies are useful for the detection and characterisation of diverse groups of viruses and bacteria and can thereby provide a better understanding of complex interactions among microorganisms potentially causing clinical disease. Here, we used a metagenomics approach to identify and characterise the possible pathogens in colon and lung samples from pigs with diarrhoea and poor growth in an Australian pig herd. We identified and characterized a wide diversity of porcine viruses including RNA viruses, in particular several picornaviruses-porcine sapelovirus (PSV), enterovirus G (EV-G), and porcine teschovirus (PTV), and a porcine astrovirus (PAstV). Single stranded DNA viruses were also detected and included parvoviruses like porcine bocavirus (PBoV) and porcine parvovirus 2 (PPV2), porcine parvovirus 7 (PPV7), porcine bufa virus (PBuV), and porcine adeno-associated virus (AAV). We also detected single stranded circular DNA viruses such as porcine circovirus type 2 (PCV2) at very low abundance and torque teno sus viruses (TTSuVk2a and TTSuVk2b). Some of the viruses detected here may have had an evolutionary past including recombination events, which may be of importance and potential involvement in clinical disease in the pigs. In addition, our metagenomics data found evidence of the presence of the bacteria Lawsonia intracellularis , Brachyspira spp., and Campylobacter spp. that may, together with these viruses, have contributed to the development of clinical disease and poor growth.

Eukaryotic circular single-stranded DNA (ssDNA) viruses were known only to infect plants and vertebrates until the discovery of the isolated DNA mycovirus from the fungus Sclerotinia sclerotiorum . Similar viral sequences were reported from several other sources and classified in ten genera within the Genomoviridae family. The current study reports two circular ssDNA mycoviruses isolated from the phytopathogen Botrytis cinerea , and their assignment to a newly created genus tentatively named Gemydayirivirus . The mycoviruses, tentatively named botrytis gemydayirivirus 1 (BGDaV1) and BGDaV2, are 1701 and 1693 nt long and encode three and two open reading frames (ORFs), respectively. Of the predicted ORFs, only ORF I, which codes for a replication initiation protein (Rep), shared identity with other proteins in GenBank. BGDaV1 is infective as cell-free purified particles and confers hypovirulence on its natural host. Investigation revealed that BGDaV1 is a target for RNA silencing and genomic DNA methylation, keeping the virus at very low titre. The discovery of BGDaV1 expands our knowledge of the diversity of genomoviruses and their interaction with fungal hosts.

French commercial hatcheries are massively producing Crassostrea gigas selected for their higher resistance to OsHV-1, and soon should also implement selection for increasing resistance to Vibrio aestuarianus. The first objective of this study was to optimize the breeding programs for dual resistance to OsHV-1 and V. aestuarianus to determine the earliest life stage for which oysters are able to develop disease resistance. Wild stocks and selected families were tested using experimental infections by both pathogens at the larval, spat and juvenile stages. Oyster families could be evaluated for OsHV-1 as soon as the larval stage by a bath method, but this only highlighted the most resistant families; those that showed the highest resistance to V. aestuarianus could be determined using the cohabitation method at the juvenile stage. The second objective of this study was to determine if selection to increase/decrease the resistance to OsHV-1 and V. aestuarianus could have an impact on other major pathogens currently detected in hatchery at the larval stage, and in nursery and field at the spat/juveniles stages (V. coralliilyticus, V. crassostreae, V. tasmaniensis, V. neptunius, V. europaeus, V. harveyi, V. chagasi). No relationship was found between mortality caused by V. aestuarianus/OsHV-1 and the mortality caused by the other virulent bacterial strains tested regardless the stages, except between OsHV-1 and V. tasmaniensis at the juvenile stage. Finally, miscellaneous findings were evidenced such as (1) bath for bacterial challenges was not adapted for spat, (2) the main pathogens at the larval stage were OsHV-1 and V. coralliilyticus using bath, while it was V. coralliilyticus, V. europaeus, and V. neptunius at the juvenile stage by injection, and (4) variation in mortality was observed among families/wild controls for all pathogens at larval and juvenile stages, except for V. harveyi for larvae., (Copyright © 2021 Elsevier Inc. All rights reserved.)

The ostreid herpes virus (OsHV-1), associated with massive mortalities in the bivalve Crassostrea gigas, was detected for the first time in the cephalopod Octopus vulgaris. Wild adult animals from a natural breeding area in Spain showed an overall prevalence of detection of 87.5% between 2010 and 2015 suggesting an environmental source of viral material uptake. Overall positive PCR detections were significantly higher in adult animals (p = 0.031) compared to newly hatched paralarvae (62%). Prevalence in embryos reached 65%. Sequencing of positive amplicons revealed a match with the variant OsHV-1 µVar showing the genomic features that distinguish this variant in the ORF4. Gill tissues from adult animals were also processed for in situ hybridization and revealed positive labelling. Experimental exposure trials in octopus paralarvae were carried out by cohabitation with virus injected oysters and by immersion in viral suspension observing a significant decrease in paralarval survival in both experiments. An increase in the number of OsHV-1 positive animals was detected in dead paralarvae after cohabitation with virus injected oysters. No signs of viral replication were observed based on lack of viral gene expression or visualization of viral structures by transmission electron microscopy. The octopus response against OsHV-1 was evaluated by gene expression of previously reported transcripts involved in immune response in C. gigas suggesting that immune defences in octopus are also activated after exposure to OsHV-1., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Members of the Delphinidae family are widely distributed across the world's oceans. We used a viral metagenomic approach to identify viruses in orca (Orcinus orca) and short-finned pilot whale (Globicephala macrorhynchus) muscle, kidney, and liver samples from deceased animals. From orca tissue samples (muscle, kidney, and liver), we identified a novel polyomavirus (Polyomaviridae), three cressdnaviruses, and two genomoviruses (Genomoviridae). In the short-finned pilot whale we were able to identify one genomovirus in a kidney sample. The presence of unclassified cressdnavirus within two samples (muscle and kidney) of the same animal supports the possibility these viruses might be widespread within the animal. The orca polyomavirus identified here is the first of its species and is not closely related to the only other dolphin polyomavirus previously discovered. The identification and verification of these viruses expands the current knowledge of viruses that are associated with the Delphinidae family., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Beekeeping is a widespread activity in Argentina, mainly producing honey that has gained both national and international recognition. There are more than 3,000,000 hives in the country, mainly concentrated in Buenos Aires Province (approximately 1,000,000 hives). In recent decades, worrying rates of hive loss have been observed in many countries around the world. In Latin America, the estimated loss of hives is between 13% (Peru and Ecuador) and 53% (Chile). Argentina had annual losses of 34% for the period of October 1, 2016 to October 1, 2017. The causes of these losses are not clear but probably involve multiple stressors that can act simultaneously. One of the main causes of loss of bee colonies worldwide is infestation by the ectoparasitic mite Varroa destructor in combination with viral infections. To date, 10 viruses have been detected that affect honey bees (Apis mellifera) in Argentina. Of these, deformed wing virus, sacbrood virus, acute bee paralysis virus, chronic bee paralysis virus, and Israeli acute bee paralysis can be transmitted by mites. Deformed wing virus and the AIK complex are the viruses most often associated with loss of hives worldwide. Considering that bee viruses have been detected in Argentina in several hymenopteran and non-hymenopteran insects, these hosts could act as important natural reservoirs for viruses and play an important role in their dispersal in the environment. Further studies to investigate the different mechanisms by which viruses spread in the environment will enable us to develop various strategies for the control of infected colonies and the spread of viruses in the habitat where they are found.

Viral infections are major causes of morbidity and mortality in solid-organ and hematopoietic stem cell transplant recipients. This study evaluated the performance of the Galileo Pathogen Solution metagenomics Next-Generation sequencing assay to detect and quantify 11 DNA viruses (cytomegalovirus, Epstein-Barr virus, BK virus, human adenovirus, JC virus, herpes simplex virus 1 and 2, varicella zoster virus, human herpesvirus 6A and 6B, and parvovirus B19) and to qualitatively detect torque teno virus. DNA extracted from 47 plasma samples of viremic transplant recipients were subjected to DNA library preparation with pathogen enrichment/human background depletion, sequencing, and automated data analysis. The viral loads were determined with the Galileo assay using a standard curve generated from a calibration panel. All of the samples tested had a 100% agreement with the real-time quantitative PCR (qPCR) assays in detecting the primary virus targets and the majority of the quantified samples had a viral load difference within 0.46 log 10 IU/mL or copies/mL. The mean difference for cytomegalovirus between the Galileo and qPCR assays was 0.21 log 10 IU/mL (SD, ±0.43 log 10 IU/mL). The mean difference for BK virus between the Galileo and qPCR assays was 0.17 log 10 cp/mL (SD, ±0.67 log 10 cp/mL). Additionally, 75 co-infections were detected in 31 samples by the Galileo assay. The study findings show that the Galileo assay can simultaneously detect and quantify multiple viruses in transplant recipients with results that are comparable with standard-of-care qPCR assays., (Copyright © 2021 Association for Molecular Pathology and American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

Microvariant genotypes of Ostreid herpesvirus 1 (OsHV-1) are associated with mass mortality events of Pacific oysters in many countries. The OsHV-1 microvariant (µVar) emerged in France 2008 and caused significant economic losses as it became endemic and displaced the previously dominant OsHV-1 reference genotype. Recently, considerable genotypic variation has been described for OsHV-1 microvariants, however, less is known about variation in viral phenotype. This study used an in vivo laboratory infection model to assess differences in total cumulative mortality, peak viral load, transmissibility, and dose-response for three OsHV-1 isolates obtained between 2011 and 2015 from endemic waterways in Australia. This followed field observations of apparent reductions in the severity of mass mortalities over this time. Significantly higher hazard of death and cumulative mortality were observed for an isolate obtained in 2011 compared to isolates from 2014-2015. In keeping with other studies, the hazard of death was higher in oysters challenged by injection compared to challenge by cohabitation and the mortality was higher when the initial dose was 1 × 10 4 OsHV-1 DNA copies per oyster injection compared to 1 × 10 2 DNA copies. There was no difference in the quantity of OsHV-1 DNA at time of death that could be related to isolate or dose, suggesting similar pathogenetic processes in the individual oysters that succumbed to end-stage disease. While the isolates examined in this study were biased towards pathogenic types of OsHV-1, as they were collected during disease outbreaks, the variation in virulence that was observed, when combined with prior data on subclinical infections, suggests that surveillance for low virulence genotypes of OsHV-1 would be rewarding. This may lead to new approaches to disease management which utilize controlled exposure to attenuated strains of OsHV-1.

The laboratory mouse strain C57BL/6 is widely used as an animal model for various applications. It is becoming increasingly clear that the bacterial enteric community highly influences the phenotype. Eukaryotic viruses represent a sparsely investigated member of the enteric microbiome that might also affect the phenotype. We here investigated the presence of enteric eukaryotic DNA viruses (EDVs) in specific pathogen-free (SPF) C57BL/6N mice purchased from three vendors upon arrival and after being fed a low-fat diet (LFD) or high-fat diet (HFD). We detected genetic fragments of EDVs belonging to the viral families of Herpes-, Mimi-, Baculo- and Phycodnaviridae represented by two genera; Chlorovirus and Prasinovirus. The EDVs were detected in the mice upon arrival and persisted for 13 weeks. However, these signals of EDVs were only detected at notable levels in mice fed LFD from 2 out of 3 vendors, which suggested that the enteric composition of these EDVs were affected by both vendor (p < 0.003) and different dietary regimes (p < 0.013). This highlights the need of additional studies assessing the potential function of these EDVs that may influence the mouse phenotype and the reproducibility of animal studies using this C57BL/6N substrain., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

In this work, we determined the diversity and dynamics of the gut virome of infants during the first year of life. Fecal samples were collected monthly, from birth to one year of age, from three healthy children living in a semi-rural village in Mexico. Most of the viral reads were classified into six families of bacteriophages including five dsDNA virus families of the order Caudovirales, with Siphoviridae and Podoviridae being the most abundant. Eukaryotic viruses were detected as early as two weeks after birth and remained present all along the first year of life. Thirty-four different eukaryotic virus families were found, where eight of these families accounted for 98% of all eukaryotic viral reads: Anelloviridae, Astroviridae, Caliciviridae, Genomoviridae, Parvoviridae, Picornaviridae, Reoviridae and the plant-infecting viruses of the Virgaviridae family. Some viruses in these families are known human pathogens, and it is surprising that they were found during the first year of life in infants without gastrointestinal symptoms. The eukaryotic virus species richness found in this work was higher than that observed in previous studies; on average between 7 and 24 virus species were identified per sample. The richness and abundance of the eukaryotic virome significantly increased during the second semester of life, probably because of an increased environmental exposure of infants with age. Our findings suggest an early and permanent contact of infants with a diverse array of bacteriophages and eukaryotic viruses, whose composition changes over time. The bacteriophages and eukaryotic viruses found in these children could represent a metastable virome, whose potential influence on the development of the infant's immune system or on the health of the infants later in life, remains to be investigated., Competing Interests: The authors have declared that no competing interests exist.

Background: Severe acute respiratory infection (SARI) results in a tremendous disease burden worldwide. Available research on active surveillance among hospitalized adult patients suffering from SARI in China is limited. This pilot study aimed to identify associated etiologies and describe the demographic, epidemiological and clinical profiles of hospitalized SARI patients aged over 16 years in Jinshan, Shanghai., Methods: Active surveillance was conducted at 1 sentinel hospital in Jinshan district, Shanghai, from April 2017 to March 2018. Hospitalized SARI patients aged over 16 years old were enrolled, and nasopharyngeal swabs were collected within 24 hours of admission and tested for multiple respiratory viruses (including 18 common viruses) and Mycoplasma pneumoniae with real-time polymerase chain reaction. Demographic, epidemiological and clinical information was obtained from case report forms., Results: In total, 397 SARI patients were enrolled; the median age was 68 years, and 194 (48.9%) patients were male. A total of 278 (70.0%) patients had at least one underlying chronic medical condition. The most frequent symptoms were cough (99.2%) and sputum production (88.4%). The median duration of hospitalization was 10 days. A total of 250 infection patients (63.0%) were positive for at least one pathogen, of whom 198 (49.9%) were positive for a single pathogen and 52 (13.1%) were positive for multiple pathogens. The pathogens identified most frequently were M. pneumoniae (23.9%, 95/397), followed by adenovirus (AdV) (11.6%, 46/397), influenza virus A/H3N2 (Flu A/H3N2) (11.1%, 44/397), human rhinovirus (HRhV) (8.1%, 32/397), influenza virus B/Yamagata (Flu B/Yamagata) (6.3%, 25/397), pandemic influenza virus A/H1N1 (Flu A/pH1N1) (4.0%, 16/397), parainfluenza virus (PIV) type 1 (2.0%, 8/397), human coronavirus (HCoV) type NL63 (2.0%, 8/397), HCoV 229E (1.5%, 6/397), HCoV HKU1 (1.5%, 6/397), PIV 3 (1.5%, 6/397), human metapneumovirus (HMPV) (1.5%, 6/397), PIV 4 (1.3%, 5/397), HCoV OC43 (1.0%, 4/397), influenza virus B/Victoria (Flu B/Victoria) (0.5%, 2/397), respiratory syncytial virus (RSV) type B (0.5%, 2/397), and human bocavirus (HBoV) (0.3%, 1/397). The seasonality of pathogen-confirmed SARI patients had a bimodal distribution, with the first peak in the summer and the second peak in the winter. Statistically significant differences were observed with respect to the rates of dyspnea, radiographically diagnosed pneumonia and the presence of at least one comorbidity in patients who were infected with only M. pneumoniae, AdV, HRhV, Flu A/H3N2, Flu A /pH1N1 or Flu B/Yamagata. The differences in the positivity rates of the above 6 pathogens among the different age groups were nonsignificant., Conclusions: M. pneumoniae, AdV and Flu A/H3N2 were the main pathogens detected in hospitalized SARI patients aged over 16 years old in Jinshan district, Shanghai. Our findings highlight the importance of sustained multipathogen surveillance among SARI patients in sentinel hospitals, which can provide useful information on SARI etiologies, epidemiology, and clinical characteristics., Competing Interests: The authors have declared that no competing interests exist.

Aeromonas hydrophila is an opportunistic pathogen that infects fish, amphibians, mammals, and humans. This study isolated a myophage, vB&#95;AhyM&#95;Ahp2 (Ahp2), that lytically infects A. hydrophila . We observed that 96% of the Ahp2 particles adsorbed to A. hydrophila within 18 min. Ahp2 also showed a latent period of 15 min with a burst size of 142 PFU/cell. This phage has a linear double-stranded DNA genome of 47,331 bp with a GC content of 57%. At least 20 Ahp2 proteins were detected by SDS-polyacrylamide gel electrophoresis; among them, a 40-kDa protein was predicted as the major capsid protein. Sequence analysis showed that Ahp2 has a genome organization closely related to a group of Aeromonas phages (13AhydR10RR, 14AhydR10RR, 85AhydR10RR, phage 3, 32 Asp37, 59.1), which infect Aeromonas hydrophila and Aeromonas salmonicida . The tail module encompassing ORF27-29 in the Ahp2 genome was present in all Aeromonas phages analyzed in this study and likely determines the host range of the virus. This study found that Ahp2 completely lyses A. hydrophila AH300206 in 3.5 h at a MOI of 0.0001 and does not lysogenize its host. Altogether, these findings show that Ahp2 is a lytic Aeromonas phage and could be a candidate for therapeutic phage cocktails.

Bioconversion of organic materials is the foundation of many applications in chemical engineering, microbiology and biochemistry. Herein, we introduce a new methodology to quantitatively determine conversion of biomass in viral infections while simultaneously imaging morphological changes of the host cell. As proof of concept, the viral replication of an unidentified giant DNA virus and the cellular response of an amoebal host are studied using soft X-ray microscopy, titration dilution measurements and thermal gravimetric analysis. We find that virions produced inside the cell are visible from 18 h post infection and their numbers increase gradually to a burst size of 280-660 virions. Due to the large size of the virion and its strong X-ray absorption contrast, we estimate that the burst size corresponds to a conversion of 6-12% of carbonaceous biomass from amoebal host to virus. The occurrence of virion production correlates with the appearance of a possible viral factory and morphological changes in the phagosomes and contractile vacuole complex of the amoeba, whereas the nucleus and nucleolus appear unaffected throughout most of the replication cycle.

In this study, we detected a circular replication-associated protein (Rep)-encoding single-stranded (CRESS) DNA virus (named Po-Circo-like (PCL) virus) in intestinal tissue samples of pigs, and the complete genome sequences of three strains (named PCL viruses GX14, GX15 and GX19) were obtained. Unlike PCL virus strains 21 and 22, whose genome sequences have 3,912 and 3,923 nucleotides (nt), respectively, the strains revealed in this study have a circular genome with 3,944 nt and five major open reading frames (ORFs). Among these ORFs, ORF1 encodes the Rep and not the typical capsid protein encoded in PCV. Furthermore, the strains in this study share 79.2%-96.0% nucleic acid identity and 83.0%-98.1% amino acid identity with ORF1 of the reference strains. Moreover, the Rep of the PCL virus in this study shared 19.9%-22.2% (<30%) identity of its amino acid sequence with PCV but shared 34.9%-94.8% (>30%) identity of its amino acid sequence with sequences of five proteins that are expressed by the family Kirkoviridae. [Correction added on 24 December 2020 after first online publication: The preceding sentence has been corrected in this version.] Interestingly, the stem loop of the PCL virus has one nucleotide substitution, T1328G. The Bo-Circo-like CH strain shares high nucleic acid and amino acid similarity (>80%) with the PCL virus. Moreover, Bo-Circo-like CH and GX-19 had similar stem-loop sequences. The PCL virus might therefore be transmitted to non-porcine hosts by cross-species transmission routes. Phylogenetic analysis classified the PCL virus into the new family Kirkoviridae and indicated its close relationship with the Bo-Circo-like virus. A phylogenetic divergence analysis based on the rep gene classified all PCL virus strains into two genotypes (PCLa and PCLb). In conclusion, the present study is the first detailed report of the PCL virus, which is a potential new virus in pigs that might be involved in cross-species transmission. Further investigation is needed to determine the pathogenesis of this virus and its epidemiologic impact., (© 2020 Blackwell Verlag GmbH.)

The Pacific oyster ( Crassostreae gigas ) has been introduced from Asia to numerous countries around the world during the 20th century. C. gigas is the main oyster species farmed worldwide and represents more than 98% of oyster production. The severity of disease outbreaks that affect C. gigas , which primarily impact juvenile oysters, has increased dramatically since 2008. The most prevalent disease, Pacific oyster mortality syndrome (POMS), has become panzootic and represents a threat to the oyster industry. Recently, major steps towards understanding POMS have been achieved through integrative molecular approaches. These studies demonstrated that infection by Ostreid herpesvirus type 1 µVar (OsHV-1 µvar) is the first critical step in the infectious process and leads to an immunocompromised state by altering hemocyte physiology. This is followed by dysbiosis of the microbiota, which leads to a secondary colonization by opportunistic bacterial pathogens, which in turn results in oyster death. Host and environmental factors ( e.g. oyster genetics and age, temperature, food availability, and microbiota) have been shown to influence POMS permissiveness. However, we still do not understand the mechanisms by which these different factors control disease expression. The present review discusses current knowledge of this polymicrobial and multifactorial disease process and explores the research avenues that must be investigated to fully elucidate the complexity of POMS. These discoveries will help in decision-making and will facilitate the development of tools and applied innovations for the sustainable and integrated management of oyster aquaculture., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Petton, Destoumieux-Garzón, Pernet, Toulza, de Lorgeril, Degremont and Mitta.)

The endoscopic features between herpes simplex virus (HSV) and cytomegalovirus (CMV) esophagitis overlap significantly, and hence the differential diagnosis between HSV and CMV esophagitis is sometimes difficult. Therefore, we developed a machine-learning-based classifier to discriminate between CMV and HSV esophagitis. We analyzed 87 patients with HSV esophagitis and 63 patients with CMV esophagitis and developed a machine-learning-based artificial intelligence (AI) system using a total of 666 endoscopic images with HSV esophagitis and 416 endoscopic images with CMV esophagitis. In the five repeated five-fold cross-validations based on the hue-saturation-brightness color model, logistic regression with a least absolute shrinkage and selection operation showed the best performance (sensitivity, specificity, positive predictive value, negative predictive value, accuracy, and area under the receiver operating characteristic curve: 100%, 100%, 100%, 100%, 100%, and 1.0, respectively). Previous history of transplantation was included in classifiers as a clinical factor; the lower the performance of these classifiers, the greater the effect of including this clinical factor. Our machine-learning-based AI system for differential diagnosis between HSV and CMV esophagitis showed high accuracy, which could help clinicians with diagnoses.

Fur seal feces-associated circular DNA virus (FSfaCV) is an unclassified circular replication-associated protein (Rep)-encoding single-stranded (CRESS) DNA virus that has been detected in mammals (fur seals and pigs). The biology and epidemiology of the virus remain largely unknown. To investigate the virus diversity among pigs in Anhui Province, China, we pooled 600 nasal samples in 2017 and detected viruses using viral metagenomic methods. From the assembled contigs, 12 showed notably high nucleotide acid sequence similarities to the genome sequences of FSfaCVs. Based on these sequences, a full-length genome sequence of the virus was then obtained using overlapping PCR and sequencing, and the virus was designated as FSfaCV-CHN (GenBank No. MK462122). This virus shared 91.3% and 90.9% genome-wide nucleotide sequence similarities with the New Zealand fur seal strain FSfaCV-as50 and the Japanese pig strain FSfaCV-JPN1, respectively. It also clustered with the two previously identified FSfaCVs in a unique branch in the phylogenetic tree based on the open reading frame 2 (ORF2), Rep-coding gene, and the genome of the reference CRESS DNA viruses. Further epidemiological investigation using samples collected in 2018 showed that the overall positive rate for the virus was 56.4% (111/197) in Anhui Province. This is the first report of FSfaCVs identified in pigs in China, and further epidemiological studies are warranted to evaluate the influence of the virus on pigs.

Over that last decade, coupling multiple strand displacement approaches with high throughput sequencing have resulted in the identification of genomes of diverse groups of small circular DNA viruses. Using a similar approach but with recovery of complete genomes by PCR, we identified a diverse group of single-stranded viruses in yellow-bellied marmot (Marmota flaviventer) fecal samples. From 13 fecal samples we identified viruses in the family Genomoviridae (n = 7) and Anelloviridae (n = 1), and several others that ware part of the larger Cressdnaviricota phylum but not within established families (n = 19). There were also circular DNA molecules identified (n = 4) that appear to encode one viral-like gene and have genomes of <1545 nts. This study gives a snapshot of viruses associated with marmots based on fecal sampling., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Kaumoebavirus infects the amoeba Vermamoeba vermiformis and has recently been described as a distant relative of the African swine fever virus. To characterize the diversity and evolution of this novel viral genus, we report here on the isolation and genome sequencing of a second strain of Kaumoebavirus , namely LCC10. Detailed analysis of the sequencing data suggested that its 362-Kb genome is linear with covalently closed hairpin termini, so that DNA forms a single continuous polynucleotide chain. Comparative genomic analysis indicated that although the two sequenced Kaumoebavirus strains share extensive gene collinearity, 180 predicted genes were either gained or lost in only one genome. As already observed in another distant relative, i.e., Faustovirus , which infects the same host, the center and extremities of the Kaumoebavirus genome exhibited a higher rate of sequence divergence and the major capsid protein gene was colonized by type-I introns. A possible role of the Vermamoeba host in the genesis of these evolutionary traits is hypothesized. The Kaumoebavirus genome exhibited a significant gene strand bias over the two-third of genome length, a feature not seen in the other members of the "extended Asfarviridae " clade. We suggest that this gene strand bias was induced by a putative single origin of DNA replication located near the genome extremity that imparted a selective force favoring the genes positioned on the leading strand.

This study characterized a single-stranded circular DNA virus in Botrytis cinerea-namely, Botrytis cinerea genomovirus 1 (BcGV1). The genome of BcGV1 was 1710 nucleotides (nts) long, possessing two ORFs, encoding a putative replication initiation protein (Rep) and a hypothetical protein. The Rep contained seven conserved motifs. The two ORFs were separated by two intergenic regions; the large intergenic region (LIR) contained 259 nts while the small intergenic region (SIR) contained 95 nts. A nonanucleotide, TAACAGTAC, in the LIR was predicted to be associated with the initiation of viral replication. Based on the phylogenetic tree constructed by Reps, BcGV1 belongs to the family Genomoviridae, forming an independent branch, indicating that BcGV1 may belong to a new genus. BcGV1 could be detected in 6.7% of tested B. cinerea strains, suggesting that BcGV1 may be widely distributed in the Chinese B. cinerea population., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Marseilleviridae is a family of large double-stranded DNA viruses that is currently divided into five subgroups, lineages A-E. Hokutovirus and kashiwazakivirus, both of which belong to lineage B, have been reported to induce host acanthamoeba cells to form aggregations called "bunches". This putatively results in increased opportunities to infect acanthamoeba cells, in contrast to lineage A, which has been reported to not form "bunches". In the present study, we isolated 14 virus strains of the family Marseilleviridae from several Japanese water samples, 11 of which were identified as lineage B viruses. All 11 lineage B strains caused infected amoeba cells to form bunches. We then investigated the involvement of monosaccharides in bunch formation by amoeba cells infected with hokutovirus. Galactose inhibited bunch formation, thereby allowing amoeba cells to delay the process, whereas mannose and glucose did not. A kinetic image analysis of hokutovirus-infected amoeba cells confirmed the inhibition of bunch formation by galactose. The number of hokutovirus-infected amoeba cells increased more rapidly than that of tokyovirus-infected cells, which belongs to lineage A. This result suggests that bunch formation by infected amoeba cells is advantageous for lineage B viruses.

Genomoviruses (family Genomoviridae) are circular single-stranded DNA viruses that have been mainly identified through metagenomics studies in a wide variety of samples from various environments. Here, we describe 98 genomes of genomoviruses found associated with members of 19 plant families from Australia, Brazil, France, South Africa and the USA. These 98 genomoviruses represent 29 species, 26 of which are new, in the genera Gemykolovirus (n = 37), Gemyduguivirus (n = 9), Gemygorvirus (n = 8), Gemykroznavirus (n = 6), Gemycircularvirus (n = 21) and Gemykibivirus (n = 17).

Since August 2014, the University of Minnesota Veterinary Diagnostic Laboratory has received cases of turkey enteritis that are clinically different from previously described cases of poult enteritis syndrome and light turkey syndrome. The birds develop dark green and extremely foul-smelling diarrhea starting at 8-10 wk of age, which may last up to 15-16 wk of age. The affected turkey flocks show poor uniformity, and feed conversion and market weights are reduced. Multiple-age farms are affected more often than the single-age farms. Morbidity varies from flock to flock and in some cases reaches 100%. At necropsy, undigested feed with increased mucus is observed in the intestines along with prominent mucosal congestion and/or hemorrhage. Microscopically, lymphocytic infiltrates expand the villi in duodenum and jejunum to form lymphoid follicles, which are often accompanied by heterophils. Next generation sequencing (Illumina Miseq) on a pool of feces from affected birds identified genetic sequences of viruses belonging to Astroviridae, Reoviridae, Picornaviridae, Picobirnaviridae, and Adenoviridae. On testing pools of fecal samples from apparently healthy (16 pools) and affected birds (30 pools), there was a higher viral load in the feces of affected birds. Picobirnavirus was detected only in the affected birds; 20 of 30 pools (66.7%) were positive. These results indicate that a high viral load of turkey picobirnavirus alone, or in association with novel picornaviruses, may be a cause of this new type of turkey enteritis.

In this study, we present an analysis of metagenome sequences obtained from a filtrate of a siphon tissue homogenate of otter clams (Lutraria rhynchaena) with swollen-siphon disease. The viral signal was mined from the metagenomic data, and a novel circular ssDNA virus was identified. Genomic features and phylogenetic analysis showed that the virus belongs to the phylum Cressdnaviricota, which consists of viruses with circular, single-stranded DNA (ssDNA) genomes. Members of this phylum have been identified in various species and in environmental samples. The newly found virus is distantly related to the currently known members of the phylum Cressdnaviricota.

Common or European hedgehogs can be found throughout Western Europe. They are known carriers of a variety of parasitic and bacterial pathogens, and have also been shown to carry several viruses, including morbilli-like paramyxoviruses, although the pathogenic and zoonotic potential of some of these viruses has yet to be determined. We report here the discovery of a novel paramyxovirus in Belgian hedgehogs, named Belerina virus. The virus was detected by nanopore sequencing of RNA isolated from hedgehog tissue. Out of 147 animals screened in this study, 57 tested positive for Belerina virus (39%), indicating a high prevalence of this virus in the Belgian hedgehog population. Based on its divergence from other known paramyxovirus species, Belerina virus is thought to represent a new species in the family Paramyxoviridae. Phylogenetic analysis groups Belerina virus together with the bat-borne Shaan virus within the genus Jeilongvirus, although expanding the tree with partial genomes shows Belerina virus forming a separate subclade within this genus, alongside a yet-unnamed paramyxovirus isolated from a greater tube-nosed bat. In summary, we discuss the complete genome sequence of Belerina virus, a putative new paramyxovirus species commonly found in Belgian hedgehogs.

Hosts and viruses are constantly evolving in response to each other: as a host attempts to suppress a virus, the virus attempts to evade and suppress the host's immune system. Here, we describe the recurrent evolution of a virulent strain of a DNA virus, which infects multiple Drosophila species. Specifically, we identified two distinct viral types that differ 100-fold in viral titer in infected individuals, with similar differences observed in multiple species. Our analysis suggests that one of the viral types recurrently evolved at least four times in the past ~30,000 years, three times in Arizona and once in another geographically distinct species. This recurrent evolution may be facilitated by an effective mutation rate which increases as each prior mutation increases viral titer and effective population size. The higher titer viral type suppresses the host-immune system and an increased virulence compared to the low viral titer type., Competing Interests: TH, RU No competing interests declared, (© 2020, Hill and Unckless.)

Recognition of the non-self signature of invading pathogens is a crucial step for the initiation of the innate immune mechanisms of the host. The host response to viral and bacterial infection involves sets of pattern recognition receptors (PRRs), which bind evolutionarily conserved pathogen structures, known as pathogen-associated molecular patterns (PAMPs). Recent advances in the identification of different types of PRRs in teleost fish revealed a number of cytosolic sensors for recognition of viral and bacterial nucleic acids. These are DExD/H-box RNA helicases including a group of well-characterized retinoic acid inducible gene I (RIG-I)-like receptors (RLRs) and non-RLR DExD/H-box RNA helicases (e.g., DDX1, DDX3, DHX9, DDX21, DHX36 and DDX41) both involved in recognition of viral RNAs. Another group of PRRs includes cytosolic DNA sensors (CDSs), such as cGAS and LSm14A involved in recognition of viral and intracellular bacterial dsDNAs. Moreover, dsRNA-sensing protein kinase R (PKR), which has a role in antiviral immune responses in higher vertebrates, has been identified in fish. Additionally, fish possess a novel PKR-like protein kinase containing Z-DNA binding domain, known as PKZ. Here, we review the current knowledge concerning cytosolic sensors for recognition of viral and bacterial nucleic acids in teleosts.