Biology subjects

Holmes, E. C.

Publications and source records attributed to Holmes, E. C..

At least 91 records · Page 5Linked to original sources

bioRxiv2021

RNA virome diversity and Wolbachia infection in individual Drosophila simulans flies

The endosymbiont bacterium Wolbachia is associated with multiple mutualistic effects on insect biology, including nutritional and antiviral properties. Wolbachia naturally occurs in Drosophila fly species, providing an operational model host to study how virome composition may be impacted by its presence. Drosophila simulans populations can carry a variety of Wolbachia strains. In particular, the wAu strain of Wolbachia has been associated with strong antiviral protection under experimental conditions. We used D. simulans sampled from the Perth Hills, Western Australia, to investigate the potential virus protective effect of the wAu strain on individual wild-caught flies. Our data revealed no appreciable variation in virus composition and abundance between Wolbachia infected/uninfected individuals associated with the presence/absence of wAu. However, it remains unclear whether wAu might impact viral infection and host survival by increasing tolerance rather than inducing complete resistance. These data also provide new insights into the natural virome diversity of D. simulans. Despite the small number of individuals sampled, we identified a repertoire of RNA viruses, including Nora virus, Galbut virus, Chaq virus, Thika virus and La Jolla virus, that have been identified in other Drosophila species. In addition, we identified five novel viruses from the families Reoviridae, Tombusviridae, Mitoviridae and Bunyaviridae. Overall, this study highlights the complex interaction between Wolbachia and RNA virus infections and provides a baseline description of the natural virome of D. simulans.

microbiology↗

bioRxiv2021

Revealing RNA virus diversity and evolution in unicellular algae transcriptomes

Remarkably little is known about the diversity and evolution of RNA viruses in unicellular eukaryotes. We screened a total of 570 transcriptomes from the Marine Microbial Eukaryote Transcriptome Sequencing Project (MMETSP) project that encompasses a wide diversity of microbial eukaryotes, including most major photosynthetic lineages (i.e. the microalgae). From this, we identified 30 new and divergent RNA virus species, occupying a range of phylogenetic positions within the overall diversity of RNA viruses. Approximately one-third of the newly described viruses comprised single-stranded positive-sense RNA viruses from the order Lenarviricota associated with fungi, plants and protists, while another third were related to the order Ghabrivirales, including members of the protist and fungi-associated Totiviridae. Other viral species showed sequence similarity to positive-sense RNA viruses from the algae-associated Marnaviridae, the double-stranded RNA Partitiviridae, as well as a single negative-sense RNA virus related to the Qinviridae. Importantly, we were able to identify divergent RNA viruses from distant host taxa, revealing the ancestry of these viral families and greatly extending our knowledge of the RNA viromes of microalgal cultures. Both the limited number of viruses detected per sample and the low sequence identity to known RNA viruses imply that additional microalgal viruses exist that could not be detected at the current sequencing depth or were too divergent to be identified using sequence similarity. Together, these results highlight the need for further investigation of algal-associated RNA viruses as well as the development of new tools to identify RNA viruses that exhibit very high levels of sequence divergence.

microbiology↗

bioRxiv2021

Diagnosis and analysis of unexplained cases of childhood encephalitis in Australia using metagenomic next-generation sequencing

Encephalitis is most often caused by a variety of infectious agents, the identity of which is commonly determined through diagnostic tests utilising cerebrospinal fluid (CSF). Immune-mediated disorders are also a differential in encephalitis cases. We investigated the clinical characteristics and potential aetiological agents of unexplained encephalitis through metagenomic next-generation sequencing of residual clinical samples of multiple tissue types and independent clinical review. A total of 43 specimens, from both sterile and non-sterile sites, were collected from 18 encephalitis cases with no cause identified by the Australian Childhood Encephalitis study. Samples were subjected to total RNA sequencing to determine the presence and abundance of potential pathogens, to reveal mixed infections, pathogen genotypes, and epidemiological origins, and to describe the possible aetiologies of unexplained encephalitis. From this, we identified five RNA and two DNA viruses associated with human infection from both non-sterile (nasopharyngeal aspirates, nose/throat swabs, urine, stool rectal swab) and sterile (cerebrospinal fluid, blood) sites. These comprised two human rhinoviruses, two human seasonal coronaviruses, two polyomaviruses and one picobirnavirus. With the exception of picobirnavirus all have been previously associated with respiratory disease. Human rhinovirus and seasonal coronaviruses may be responsible for five of the encephalitis cases reported here. Immune-mediated encephalitis was considered clinically likely in six cases and RNA sequencing did not identify a possible pathogen in these cases. The aetiology remained unknown in nine cases. Our study emphasises the importance of respiratory viruses in the aetiology of unexplained child encephalitis and suggests that the routine inclusion of non-CNS sampling in encephalitis clinical guidelines/protocols could improve the diagnostic yield. Author SummaryEncephalitis is caused by both infectious agents and auto-immune disorders. However, the aetiological agents, including viruses, remain unknown in around half the cases of encephalitis in many cohorts. Importantly, diagnostic tests are usually based on the analysis of cerebrospinal fluid which may limit their utility. We used a combination of meta-transcriptomic sequencing and independent clinical review to identify the potential causative pathogens in cases of unexplained childhood encephalitis. Accordingly, we identified seven viruses associated with both sterile and non-sterile sampling sites. Human rhinovirus and seasonal coronaviruses were considered as most likely responsible for five of the 18 encephalitis cases studied, while immune-mediated encephalitis was considered the cause in six cases, and we were unable to determine the aetiology in nine cases. Overall, we demonstrate the role of respiratory viruses as a cause of unexplained encephalitis and that sampling sites other than cerebrospinal fluid is of diagnostic value.

microbiology↗

bioRxiv2021

Slippery when wet: cross-species transmission of divergent coronaviruses in bony and jawless fish and the evolutionary history of the Coronaviridae

The Nidovirales comprise a genetically diverse group of positive-sense single-stranded RNA virus families that infect a range of invertebrate and vertebrate hosts. Recent metagenomic studies have identified nido-like virus sequences, particularly those related to the Coronaviridae, in a range of aquatic hosts including fish, amphibians and reptiles. We sought to identify additional members of the Coronaviridae in both bony and jawless fish through a combination of total RNA sequencing (meta-transcriptomics) and data mining of published RNA sequencing data, and from this reveal more of the long-term patterns and processes of coronavirus evolution. Accordingly, we identified a number of divergent viruses that fell within the Letovirinae subfamily of the Coronaviridae, including those in a jawless fish - the pouched lamprey. By mining fish transcriptome data we identified additional virus transcripts matching these viruses in bony fish from both marine and freshwater environments. These new viruses retained sequence conservation in the RNA-dependant RNA polymerase across the Coronaviridae, but formed a distinct and diverse phylogenetic group. Although there are broad-scale topological similarities between the phylogenies of the major groups of coronaviruses and their vertebrate hosts, the evolutionary relationships of viruses within the Letovirinae does not mirror that of their hosts. For example, the coronavirus found in the pouched lamprey fell within the phylogenetic diversity of bony fish letoviruses, indicative of past host switching events. Hence, despite possessing a phylogenetic history that likely spans the entire history of the vertebrates, coronavirus evolution has been characterised by relatively frequent cross-species transmission, particularly in hosts that reside in aquatic habitats.

microbiology↗

bioRxiv2021

Identification of novel bat coronaviruses sheds light on the evolutionary origins of SARS-CoV-2 and related viruses

Although a variety of SARS-CoV-2 related coronaviruses have been identified, the evolutionary origins of this virus remain elusive. We describe a meta-transcriptomic study of 411 samples collected from 23 bat species in a small (~1100 hectare) region in Yunnan province, China, from May 2019 to November 2020. We identified coronavirus contigs in 40 of 100 sequencing libraries, including seven representing SARS-CoV-2-like contigs. From these data we obtained 24 full-length coronavirus genomes, including four novel SARS-CoV-2 related and three SARS-CoV related genomes. Of these viruses, RpYN06 exhibited 94.5% sequence identity to SARS-CoV-2 across the whole genome and was the closest relative of SARS-CoV-2 in the ORF1ab, ORF7a, ORF8, N, and ORF10 genes. The other three SARS-CoV-2 related coronaviruses were nearly identical in sequence and clustered closely with a virus previously identified in pangolins from Guangxi, China, although with a genetically distinct spike gene sequence. We also identified 17 alphacoronavirus genomes, including those closely related to swine acute diarrhea syndrome virus and porcine epidemic diarrhea virus. Ecological modeling predicted the co-existence of up to 23 Rhinolophus bat species in Southeast Asia and southern China, with the largest contiguous hotspots extending from South Lao and Vietnam to southern China. Our study highlights both the remarkable diversity of bat viruses at the local scale and that relatives of SARS-CoV-2 and SARS-CoV circulate in wildlife species in a broad geographic region of Southeast Asia and southern China. These data will help guide surveillance efforts to determine the origins of SARS-CoV-2 and other pathogenic coronaviruses.

microbiology↗

bioRxiv2021

Metagenomic sequencing reveals a lack of virus exchange between native and invasive freshwater fish across the Murray-Darling Basin, Australia

Biological invasions are among the biggest threats to freshwater biodiversity. This is increasingly relevant in the Murray-Darling Basin, Australia, particularly since the introduction of the common carp (Cyprinus carpio). This invasive species now occupies up to 90% of fish biomass, with hugely detrimental impacts on native fauna and flora. To address the ongoing impacts of carp, cyprinid herpesvirus 3 (CyHV-3) has been proposed as a potentially effective biological control. Crucially, however, it is unknown whether CyHV-3 and other cyprinid herpesviruses already exist in the Murray-Darling. Further, little is known about those viruses that naturally occur in wild freshwater fauna, and the frequency with which these viruses jump species boundaries. To document the evolution and diversity of freshwater fish viromes and better understand the ecological context to the proposed introduction of CyHV-3, we performed a meta-transcriptomic viral survey of invasive and native fish across the Murray-Darling Basin, covering over 2,200 km of the river system. Across a total of 36 RNA libraries representing 10 species, we failed to detect CyHV-3 nor any closely related viruses. Rather, meta-transcriptomic analysis identified 18 vertebrate-associated viruses that could be assigned to the Arenaviridae, Astroviridae, Bornaviridae, Caliciviridae, Coronaviridae, Chuviridae, Flaviviridae, Hantaviridae, Hepeviridae, Paramyxoviridae, Picornaviridae, Poxviridae, Reoviridae and Rhabdoviridae families, and a further 27 that were deemed to be associated with non-vertebrate hosts. Notably, we revealed a marked lack of viruses that are shared among invasive and native fishes sampled here, suggesting that there is little virus transmission from common carp to native fish species. Overall, this study provides the first data on the viruses naturally circulating in a major river system and supports the notion that fish harbour a large diversity of viruses with often deep evolutionary histories. Author SummaryThe ongoing invasion of the common carp in the Murray-Darling Basin, Australia, has wreaked havoc on native freshwater ecosystems. This has stimulated research into the possible biological control of invasive carp through the deliberate release of the virus cyprinid herpesvirus 3 (CyHV-3). However, little is known on the diversity of viruses that naturally circulate in wild freshwater fauna, whether these viruses are transmitted between invasive and native species, nor if CyHV-3 or other cyprinid herpesviruses are already present in the basin. To address these fundamental questions we employed meta-transcriptomic next-generation sequencing to characterise the total assemblage of viruses (i.e. the viromes) in three invasive and seven native fish species cohabiting at 10 sites across 2,200 km of the river system. From this analysis we identified 18 vertebrate-associated viruses across 14 viral families, yet a marked lack of virus transmission between invasive and native species. Importantly, no CyHV-3 was detected. This study shows that freshwater fish harbour a high diversity and abundance of viruses, that viruses have likely been associated with fish for millennia, and that there is likely little direct virus transmission between introduced and native species.

microbiology↗

bioRxiv2021

Frequent intergenotypic recombination between the non-structural and structural genes is a major driver of epidemiological fitness in caliciviruses

The diversity of lagoviruses (Caliciviridae) in Australia has increased considerably. By the end of 2017, five variants from three viral genotypes were present in populations of Australian rabbits, while prior to 2014 only two variants were known. To understand the interactions between these lagovirus variants we monitored their geographical distribution and relative incidence over time through a landscape-scale competition study, and from this, revealed potential drivers of epidemiological fitness. Within three years of the arrival of GI.1bP-GI.2 (RHDV2) into Australia, we observed the emergence of two novel recombinant lagovirus variants, GI.4eP-GI.2 (4e-recombinant) in New South Wales and GI.4cP-GI.2 (4c-recombinant) in Victoria. Although both novel recombinants contain the non-structural genes from benign, rabbit-specific, enterotropic viruses, these variants were recovered from the livers of both rabbits and hares that had died acutely. This suggests that determinants of host and tissue tropism for lagoviruses are associated with the structural genes, and that tropism is intricately connected with pathogenicity. Phylogenetic analyses demonstrated that the 4c-recombinant emerged independently on multiple occasions, with five distinct lineages observed. Both new recombinant variants replaced the previous dominant parental RHDV2 in their respective geographical areas, despite sharing an identical or near-identical (i.e., single amino acid change) major capsid protein with the parental virus. This suggests that epidemiological fitness of these recombinants was not driven by antigenic variation in the capsid, implicating the non-structural genes as key drivers of epidemiological fitness. Molecular clock estimates place the GI4.e recombination event in early to mid-2015, while the five GI.4c recombination events occurred from late 2015 through to early 2017. The emergence of at least six viable recombinant variants within a two-year period highlights an unprecedented frequency of these events, detectable only due to intensive surveillance, and demonstrates the importance of recombination in lagovirus evolution.

microbiology↗

bioRxiv2021

A novel rubi-like virus in the Pacific electric ray (Tetronarce californica) reveals the complex evolutionary history of the Matonaviridae

Rubella virus (RuV) is the causative agent of rubella ("German measles") and remains a global health concern. Until recently, RuV was the only known member of the genus Rubivirus and the only virus species classified within the Matonaviridae family of positive-sense RNA viruses. Other matonaviruses, including two new rubella-like viruses, Rustrela virus and Ruhugu virus, have been identified in several mammalian species, along with more divergent viruses in fish and reptiles. To screen for the presence of additional novel rubella-like viruses we mined published transcriptome data using genome sequences from Rubella, Rustrela, and Ruhugu viruses as baits. From this, we identified a novel rubella-like virus in a transcriptome of Tetronarce californica (Pacific electric ray) that is more closely related to mammalian Rustrela virus than to the divergent fish matonavirus and indicative of a complex pattern of cross-species virus transmission. Analysis of host reads confirmed that the sample analysed was indeed from a Pacific electric ray, and two other viruses identified in this animal, from the Arenaviridae and Reoviridae, grouped with other fish viruses. These findings indicate that the evolutionary history of the Matonaviridae is more complex than previously thought and highlights the vast number of viruses still to be discovered.

microbiology↗

bioRxiv2020

Meta-transcriptomic identification of divergent Amnoonviridae in fish

Tilapia lake virus (TiLV) has caused mass mortalities in farmed and wild tilapia with serious economic and ecological consequences. Until recently, this virus was the sole member of the Amnoonviridae, a family within the order Articulavirales comprising segmented negative-sense RNA viruses. We sought to identify additional viruses within the Amnoonviridae through total RNA sequencing (meta-transcriptomics) and data mining of published transcriptomes. Accordingly, we sampled marine fish species from both Australia and China and discovered two new viruses within the Amnoonviridae, tentatively called Flavolineata virus and Piscibus virus, respectively. In addition, by mining vertebrate transcriptome data we identified nine additional virus transcripts matching to multiple genomic segments of TiLV in both marine and freshwater fish. These new viruses retained sequence conservation with the distantly related Orthomyxoviridae in the RdRp subunit PB1, but formed a distinct and diverse phylogenetic group. These data suggest that the Amnoonviridae have a broad host range within fish and that greater animal sampling will identify additional divergent members of the Articulavirales.

microbiology↗

bioRxiv2020

Divergent influenza-like viruses of amphibians and fish support an ancient evolutionary association

Influenza viruses (family Orthomyxoviridae) infect a variety of vertebrates, including birds, humans, and other mammals. Recent metatranscriptomic studies have uncovered divergent influenza viruses in amphibians, fish and jawless vertebrates, suggesting that these viruses may be widely distributed. We sought to identify additional vertebrate influenza-like viruses through the analysis of publicly available RNA sequencing data. Accordingly, by data mining, we identified the complete coding segments of five divergent vertebrate influenza-like viruses. Three fell as sister lineages to influenza B virus: salamander influenza-like virus in Mexican walking fish (Ambystoma mexicanum) and plateau tiger salamander (Ambystoma velasci), siamese algae-eater influenza-like virus in siamese algae-eater fish (Gyrinocheilus aymonieri) and chum salmon influenza-like virus in chum salmon (Oncorhynchus keta). Similarly, we identified two influenza-like viruses of amphibians that fell as sister lineages to influenza D virus: cane toad influenza-like virus and the ornate chorus frog influenza-like virus, in the cane toad (Rhinella marina) and ornate chorus frog (Microhyla fissipes), respectively. Despite their divergent phylogenetic positions, these viruses retained segment conservation and splicing consistent with transcriptional regulation in influenza B and influenza D viruses, and were detected in respiratory tissues. These data suggest that influenza viruses have been associated with vertebrates for their entire evolutionary history.

microbiology↗

bioRxiv2020

Meta-transcriptomic discovery of a divergent circovirus and a chaphamaparvovirus in captive reptiles with proliferative respiratory syndrome

Viral pathogens are being increasingly described in association with mass morbidity and mortality events in reptiles. However, our knowledge of reptile viruses and their role in population health remains limited. Herein, we describe a meta-transcriptomic investigation of a mass morbidity and mortality event in a colony of central bearded dragons (Pogona vitticeps) in 2014. Severe, extensive proliferation of the respiratory epithelium was consistently found in affected dragons. Similar proliferative lung lesions were identified in bearded dragons from the same colony in 2020 in association with increased intermittent mortality. Total RNA sequencing of bearded dragon tissue identified two divergent DNA viruses: a reptile-infecting circovirus, denoted bearded dragon circovirus (BDCV), and the first exogeneous reptilian chaphamaparvovirus - bearded dragon chaphamaparvovirus (BDchPV). Phylogenetic analysis revealed that BDCV was most closely related to bat-associated circoviruses, exhibiting 70% amino acid sequence identity. In contrast, the newly discovered BDchPV showed approximately 35-40% identity in the non-structural (NS) protein to parvoviruses obtained from tilapia fish and crocodiles in China. Subsequent specific PCR assays detected BDCV exclusively and comprehensively within animals with proliferative pulmonary lesions and respiratory disease. This study expands our understanding of viral diversity in the context of diseased reptiles in captivity.

microbiology↗

bioRxiv2020

How accurately can we assess zoonotic risk?

The authors have withdrawn their manuscript as a website associated with a zoonotic risk prediction tool mentioned here was not public. Therefore, the authors do not wish this work to be cited as reference for the project. If you have any questions, please contact the corresponding author

microbiology↗

bioRxiv2020

Extensive Genetic Diversity and Host Range of Rodent-borne Coronaviruses

To better understand the genetic diversity, host association and evolution of coronaviruses (CoVs) in China we analyzed a total of 696 rodents encompassing 16 different species sampled from Zhejiang and Yunnan provinces. Based on the reverse transcriptase PCR-based CoV screening CoVs of fecal samples and subsequent sequence analysis of the RdRp gene, we identified CoVs in diverse rodent species, comprising Apodemus agrarius, Apodemus latronum, Bandicota indica, Eothenomys miletus, E. eleusis, Rattus andamanesis, Rattus norvegicus, and R. tanezumi. Apodemus chevrieri was a particularly rich host, harboring 25 rodent CoVs. Genetic and phylogenetic analysis revealed the presence of three groups of CoVs carried by a range of rodents that were closely related to the Lucheng Rn rat coronavirus (LRNV), China Rattus coronavirus HKU24 (ChRCoV_HKU24) and Longquan Rl rat coronavirus (LRLV) identified previously. One newly identified A. chevrieri-associated virus closely related to LRNV lacked an NS2 gene. This virus had a similar genetic organization to AcCoV-JC34, recently discovered in the same rodent species in Yunnan, suggesting that it represents a new viral subtype. Notably, additional variants of LRNV were identified that contained putative nonstructural NS2b genes located downstream of the NS2 gene that were likely derived from the host genome. Recombination events were also identified in the ORF1a gene of Lijiang-71. In sum, these data reveal the substantial genetic diversity and genomic complexity of rodent-borne CoVs, and greatly extend our knowledge of these major wildlife virus reservoirs.

genetics↗

bioRxiv2020

Pathogenicity, tissue tropism and potential vertical transmission of SARSr-CoV-2 in Malayan pangolins

SARS-CoV-2 is having severe impact on public health at a global scale. Malayan pangolin SARS-CoV-2-related coronavirus (SARSr-CoV-2) is closely related to SARS-CoV-2. We show that CT scans of virus-positive pangolins reveal bilateral ground-glass opacities in lungs in similar manner to COVID-19 patients. The virus infected multiple organs in pangolins, with the lungs being the major target. Histological expression showed that ACE2 and TMPRSS2 are co-expressed with viral RNA. Transcriptome analysis revealed an inadequate interferon response, with different dysregulated chemokines and cytokines responses in pregnant and non-pregnant adults and fetuses. Viral RNA and protein were detected in three fetuses providing evidence for vertical virus transmission. In sum, our study identifies the biological framework of SARSr-CoV-2 in pangolins, revealing striking similarities to COVID-19 in humans.

microbiology↗

bioRxiv2020

Red fox viromes across an urban-rural gradient

The Red fox (Vulpes vulpes) has established large populations in Australias urban and rural areas since its introduction following European settlement. Foxes cryptic and highly adaptable nature allows them to invade cities and live among humans while remaining largely unnoticed. Urban living and access to anthropogenic food resources also influences fox ecology. Urban foxes grow larger, live at higher densities and are more social than their rural counterparts. These ecological changes in urban red foxes are likely to impact the pathogens that they harbour, and foxes could pose a disease risk to humans and other species that share these urban spaces. To assess this possibility, we used a meta-transcriptomic approach to characterise the viromes of urban and rural foxes across the Greater Sydney region in Australia. Urban and rural foxes differed significantly in virome composition, with rural foxes harbouring a greater abundance of viruses compared to their urban counterparts. In contrast, urban fox viromes comprised a greater diversity of viruses compared to rural foxes. We identified nine potentially novel vertebrate-associated viruses in both urban and rural foxes, some of which are related to viruses associated with disease in domestic species and humans. These included members of the Astroviridae, Picobirnaviridae, Hepeviridae and Picornaviridae as well as rabbit haemorrhagic disease virus-2 (RHDV2). This study sheds light on the viruses carried by urban and rural foxes and emphasises the need for greater genomic surveillance of foxes and other invasive species at the human-wildlife interface. ImportanceUrbanisation of wild environments is increasing as human populations continue to expand. Remnant pockets of natural environments and other green spaces in urban landscapes provide invasive wildlife such as red foxes with refuges within urban areas, where they thrive on the food resources provisioned by humans. Close contact between humans, domestic species and foxes likely increases the risk of novel pathogen emergence. Indeed, the vast majority of emerging infectious diseases in humans originate in wild animals. Here, we explored potential differences in viromes between urban fox invaders and their rural counterparts. Viromes of foxes and their ectoparasites comprise a diversity of viruses including those from the Astroviridae, Picobirnaviridae, Hepeviridae, Caliciviridae and Picornaviridae. Microbial surveillance in foxes and other urban wildlife is vital for monitoring viral emergence and for the prevention of infectious diseases.

microbiology↗

bioRxiv2020

Meta-transcriptomic detection of diverse and divergent RNA viruses in green and chlorarachniophyte algae

Our knowledge of the diversity and evolution of the virosphere will likely increase dramatically with the study of microbial eukaryotes, including the microalgae in few RNA viruses have been documented to date. By combining meta-transcriptomic approaches with sequence and structural-based homology detection, followed by PCR confirmation, we identified 18 novel RNA viruses in two major groups of microbial algae - the chlorophytes and the chlorarachniophytes. Most of the RNA viruses identified in the green algae class Ulvophyceae were related to those from the families Tombusviridae and Amalgaviridae that have previously been associated with plants, suggesting that these viruses have an evolutionary history that extends to when their host groups shared a common ancestor. In contrast, seven ulvophyte associated viruses exhibited clear similarity with the mitoviruses that are most commonly found in fungi. This is compatible with horizontal virus transfer between algae and fungi, although mitoviruses have recently been documented in plants. We also document, for the first time, RNA viruses in the chlorarachniophytes, including the first observation of a negative-sense (bunya-like) RNA virus in microalgae. The other virus-like sequence detected in chlorarachniophytes is distantly related to those from the plant virus family Virgaviridae, suggesting that they may have been inherited from the secondary chloroplast endosymbiosis event that marked the origin of the chlorarachniophytes. More broadly, this work suggests that the scarcity of RNA viruses in algae most likely results from limited investigation rather than their absence. Greater effort is needed to characterize the RNA viromes of unicellular eukaryotes, including through structure-based methods that are able to detect distant homologies, and with the inclusion of a wider range of eukaryotic microorganisms. Author summaryRNA viruses are expected to infect all living organisms on Earth. Despite recent developments in and the deployment of large-scale sequencing technologies, our understanding of the RNA virosphere remains anthropocentric and largely restricted to human, livestock, cultivated plants and vectors for viral disease. However, a broader investigation of the diversity of RNA viruses, especially in protists, is expected to answer fundamental questions about their origin and long-term evolution. This study first investigates the RNA virus diversity in unicellular algae taxa from the phylogenetically distinct ulvophytes and chlorarachniophytes taxa. Despite very high levels of sequence divergence, we were able to identify 18 new RNA viruses, largely related to plant and fungi viruses, and likely illustrating a past history of horizontal transfer events that have occurred during RNA virus evolution. We also hypothesise that the sequence similarity between a chlorarachniophyte-associated virga-like virus and members of Virgaviridae associated with plants may represent inheritance from a secondary endosymbiosis event. A promising approach to detect the signals of distant virus homologies through the analysis of protein structures was also utilised, enabling us to identify potential highly divergent algal RNA viruses.

microbiology↗

bioRxiv2020

A divergent Articulavirus in an Australian gecko identified using meta-transcriptomics and protein structure comparisons

The discovery of highly divergent RNA viruses is compromised by their limited sequence similarity to known viruses. Evolutionary information obtained from protein structural modelling offers a powerful approach to detect distantly related viruses based on the conservation of tertiary structures in key proteins such as the viral RNA-dependent RNA polymerase (RdRp). We utilised a template-based approach for protein structure prediction from amino acid sequences to identify distant evolutionary relationships among viruses detected in meta-transcriptomic sequencing data from Australian wildlife. The best predicted protein structural model was compared with the results of similarity searches against protein databases based on amino acid sequence data. Using this combination of meta-transcriptomics and protein structure prediction we identified the RdRp (PB1) gene segment of a divergent negative-sense RNA virus in a native Australian gecko (Geyra lauta) that was confirmed by PCR and Sanger sequencing. Phylogenetic analysis identified the Gecko articulavirus (GECV) as a newly described genus within the family Amnoonviridae, order Articulavirales, that is most closely related to the fish virus Tilapia tilapinevirus (TiLV). These findings provide important insights into the evolution of negative-sense RNA viruses and structural conservation of the viral replicase among members of the order Articulavirales.

microbiology↗

bioRxiv2020

Metagenomic identification of diverse animal hepaciviruses and pegiviruses

The RNA virus family Flaviviridae harbours several important pathogens of humans and other animals, including Zika virus, dengue virus and hepatitis C virus. The Flaviviridae are currently divided into four genera - Hepacivirus, Pegivirus, Pestivirus and Flavivirus - each of which have a diverse host range. Members of the genus Hepacivirus are associated with a diverse array of animal species, including humans and non-human primates, other mammalian species, as well as birds and fish, while the closely related pegiviruses have been identified in a variety of mammalian taxa including humans. Using a combination of meta-transcriptomic and whole genome sequencing we identified four novel hepaciviruses and one novel variant of a known virus, in five species of native Australian wildlife, expanding our knowledge of the diversity in this important group of RNA viruses. The infected hosts comprised native Australian marsupials and birds, as well as a native gecko (Gehyra lauta). The addition of these novel viruses led to the identification of a distinct marsupial clade within the hepacivirus phylogeny that also included an engorged Ixodes holocyclus tick collected while feeding on Australian long-nosed bandicoots (Perameles nasuta). Gecko and avian associated hepacivirus lineages were also identified. In addition, by mining the short-read archive (SRA) database we identified another five novel members of Flaviviridae, comprising three new hepaciviruses from avian and primate hosts, as well as two primate-associated pegiviruses. The large-scale phylogenetic analysis of these novel hepacivirus and pegivirus genomes provides additional support for virus-host co-divergence over evolutionary time-scales.

microbiology↗