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1.
Biomed Res Int ; 2020: 4389089, 2020.
Article in English | MEDLINE | ID: mdl-32596311

ABSTRACT

The Coronavirus Disease 2019 (COVID-19) is a new viral infection caused by the severe acute respiratory coronavirus 2 (SARS-CoV-2). Genomic analyses have revealed that SARS-CoV-2 is related to Pangolin and Bat coronaviruses. In this report, a structural comparison between the Sars-CoV-2 Envelope and Membrane proteins from different human isolates with homologous proteins from closely related viruses is described. The analyses here reported show the high structural similarity of Envelope and Membrane proteins to the counterparts from Pangolin and Bat coronavirus isolates. However, the comparisons have also highlighted structural differences specific of Sars-CoV-2 proteins which may be correlated to the cross-species transmission and/or to the properties of the virus. Structural modelling has been applied to map the variant sites onto the predicted three-dimensional structure of the Envelope and Membrane proteins.


Subject(s)
Betacoronavirus/chemistry , Coronavirus Infections/virology , Pneumonia, Viral/virology , Viral Envelope Proteins/chemistry , Viral Matrix Proteins/chemistry , Alphacoronavirus/chemistry , Alphacoronavirus/classification , Alphacoronavirus/genetics , Amino Acid Sequence , Animals , Betacoronavirus/classification , Betacoronavirus/genetics , Chiroptera/virology , Coronaviridae/chemistry , Coronaviridae/classification , Coronaviridae/genetics , Eutheria/virology , Humans , Models, Molecular , Pandemics , Protein Conformation , Sequence Homology, Amino Acid , Species Specificity , Structural Homology, Protein , Viral Envelope Proteins/genetics , Viral Matrix Proteins/genetics
2.
Virulence ; 11(1): 707-718, 2020 01 01.
Article in English | MEDLINE | ID: mdl-32490723

ABSTRACT

With the outbreak of the recent severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in 2019, coronaviruses have become a global research hotspot in the field of virology. Coronaviruses mainly cause respiratory and digestive tract diseases, several coronaviruses are responsible for porcine diarrhea, such as porcine epidemic diarrhea virus (PEDV), porcine deltacoronavirus (PDCoV), and emerging swine acute diarrhea syndrome coronavirus (SADS-CoV). Those viruses have caused huge economic losses and are considered as potential public health threats. Porcine torovirus (PToV) and coronaviruses, sharing similar genomic structure and replication strategy, belong to the same order Nidovirales. Here, we developed a multiplex TaqMan-probe-based real-time PCR for the simultaneous detection of PEDV, PDCoV, PToV, and SADS-CoV for the first time. Specific primers and TaqMan fluorescent probes were designed targeting the ORF1a region of PDEV, PToV, and SADS-CoV and the ORF1b region of PDCoV. The method showed high sensitivity and specificity, with a detection limit of 1 × 102 copies/µL for each pathogen. A total of 101 clinical swine samples with signs of diarrhea were analyzed using this method, and the result showed good consistency with conventional reverse transcription PCR (RT-PCR). This method improves the efficiency for surveillance of these emerging and reemerging swine enteric viruses and can help reduce economic losses to the pig industry, which also benefits animal and public health.


Subject(s)
Communicable Diseases, Emerging/veterinary , Coronaviridae Infections/veterinary , Coronaviridae/isolation & purification , Polymerase Chain Reaction , Swine Diseases/diagnosis , Animals , Coinfection/diagnosis , Coinfection/veterinary , Communicable Diseases, Emerging/diagnosis , Coronaviridae/genetics , Coronaviridae Infections/diagnosis , Diarrhea/diagnosis , Diarrhea/veterinary , Open Reading Frames/genetics , Polymerase Chain Reaction/standards , RNA, Viral/genetics , Reproducibility of Results , Sensitivity and Specificity , Swine
3.
J Neurol Sci ; 414: 116884, 2020 07 15.
Article in English | MEDLINE | ID: mdl-32464367

ABSTRACT

A comprehensive review of the neurological disorders reported during the current COVID-19 pandemic demonstrates that infection with SARS-CoV-2 affects the central nervous system (CNS), the peripheral nervous system (PNS) and the muscle. CNS manifestations include: headache and decreased responsiveness considered initial indicators of potential neurological involvement; anosmia, hyposmia, hypogeusia, and dysgeusia are frequent early symptoms of coronavirus infection. Respiratory failure, the lethal manifestation of COVID-19, responsible for 264,679 deaths worldwide, is probably neurogenic in origin and may result from the viral invasion of cranial nerve I, progressing into rhinencephalon and brainstem respiratory centers. Cerebrovascular disease, in particular large-vessel ischemic strokes, and less frequently cerebral venous thrombosis, intracerebral hemorrhage and subarachnoid hemorrhage, usually occur as part of a thrombotic state induced by viral attachment to ACE2 receptors in endothelium causing widespread endotheliitis, coagulopathy, arterial and venous thromboses. Acute hemorrhagic necrotizing encephalopathy is associated to the cytokine storm. A frontal hypoperfusion syndrome has been identified. There are isolated reports of seizures, encephalopathy, meningitis, encephalitis, and myelitis. The neurological diseases affecting the PNS and muscle in COVID-19 are less frequent and include Guillain-Barré syndrome; Miller Fisher syndrome; polyneuritis cranialis; and rare instances of viral myopathy with rhabdomyolysis. The main conclusion of this review is the pressing need to define the neurology of COVID-19, its frequency, manifestations, neuropathology and pathogenesis. On behalf of the World Federation of Neurology we invite national and regional neurological associations to create local databases to report cases with neurological manifestations observed during the on-going pandemic. International neuroepidemiological collaboration may help define the natural history of this worldwide problem.


Subject(s)
Betacoronavirus , Cerebrovascular Disorders/etiology , Coronavirus Infections/complications , Nervous System Diseases/etiology , Neuromuscular Diseases/etiology , Pandemics , Pneumonia, Viral/complications , Registries , Adult , Animals , Cerebrovascular Disorders/physiopathology , Communicable Diseases, Emerging/epidemiology , Communicable Diseases, Emerging/virology , Coronaviridae/pathogenicity , Coronaviridae/physiology , Coronaviridae/ultrastructure , Coronavirus Infections/epidemiology , Coronavirus Infections/physiopathology , Coronavirus Infections/veterinary , Coronavirus Infections/virology , Cytokine Release Syndrome/etiology , Cytokine Release Syndrome/physiopathology , Endothelium, Vascular/pathology , Endothelium, Vascular/virology , Humans , Models, Animal , Nervous System Diseases/physiopathology , Neuromuscular Diseases/physiopathology , Organ Specificity , Peptidyl-Dipeptidase A/physiology , Pneumonia, Viral/physiopathology , Thrombophilia/etiology , Thrombophilia/physiopathology , Viral Tropism
4.
PLoS Pathog ; 16(5): e1008421, 2020 05.
Article in English | MEDLINE | ID: mdl-32407364

ABSTRACT

The outbreak of a novel corona Virus Disease 2019 (COVID-19) in the city of Wuhan, China has resulted in more than 1.7 million laboratory confirmed cases all over the world. Recent studies showed that SARS-CoV-2 was likely originated from bats, but its intermediate hosts are still largely unknown. In this study, we assembled the complete genome of a coronavirus identified in 3 sick Malayan pangolins. The molecular and phylogenetic analyses showed that this pangolin coronavirus (pangolin-CoV-2020) is genetically related to the SARS-CoV-2 as well as a group of bat coronaviruses but do not support the SARS-CoV-2 emerged directly from the pangolin-CoV-2020. Our study suggests that pangolins are natural hosts of Betacoronaviruses. Large surveillance of coronaviruses in pangolins could improve our understanding of the spectrum of coronaviruses in pangolins. In addition to conservation of wildlife, minimizing the exposures of humans to wildlife will be important to reduce the spillover risks of coronaviruses from wild animals to humans.


Subject(s)
Betacoronavirus/classification , Betacoronavirus/genetics , Coronavirus Infections/virology , Disease Reservoirs/virology , Eutheria/virology , Pneumonia, Viral/virology , Animals , Coronaviridae/classification , Coronaviridae/genetics , Host Specificity , Humans , Pandemics , Phylogeny , Sequence Homology, Nucleic Acid , Zoonoses/prevention & control , Zoonoses/virology
5.
Biochem Biophys Res Commun ; 527(4): 993-999, 2020 07 05.
Article in English | MEDLINE | ID: mdl-32446559

ABSTRACT

Most viruses inhibit the innate immune system and/or the RNA degradation processes of host cells to construct an advantageous intracellular environment for their survival. Characteristic RNA sequences within RNA virus genomes or RNAs transcribed from DNA virus genomes contribute toward this inhibition. In this study, we developed a method called "Fate-seq" to comprehensively identify the RNA sequences derived from RNA and DNA viruses, contributing RNA stability in the cells. We examined the stabilization activity of 5,924 RNA fragments derived from 26 different viruses (16 RNA viruses and 10 DNA viruses) using next-generation sequencing of these RNAs fused 3' downstream of GFP reporter RNA. With the Fate-seq approach, we detected multiple virus-derived RNA sequences that stabilized GFP reporter RNA, including sequences derived from severe acute respiratory syndrome-related coronavirus (SARS-CoV). Comparative genomic analysis revealed that these RNA sequences and their predicted secondary structures are highly conserved between SARS-CoV and the novel coronavirus, SARS-CoV-2, which is responsible for the global outbreak of the coronavirus-associated disease that emerged in December 2019 (COVID-19). These sequences have the potential to enhance the stability of viral RNA genomes, thereby augmenting viral replication efficiency and virulence.


Subject(s)
Betacoronavirus/genetics , Coronavirus Infections/virology , Pneumonia, Viral/virology , RNA Stability , RNA, Viral/chemistry , SARS Virus/genetics , Base Sequence , Betacoronavirus/chemistry , Conserved Sequence , Coronaviridae/genetics , Genome, Viral , HeLa Cells , Humans , Nucleic Acid Conformation , Pandemics , SARS Virus/chemistry , Sequence Analysis, RNA
6.
Front Biosci (Landmark Ed) ; 25: 1894-1900, 2020 06 01.
Article in English | MEDLINE | ID: mdl-32472763

ABSTRACT

We analyzed the nucleocapsid and surface proteins from several Coronaviridae viruses using an alignment-free computer program. Three isolates of novel, human coronavirus (SARS0CoV-2) (2019) that are responsible for the current pandemic and older SARS strains of human and animal coronaviruses were examined. The nucleocapsid and glycoprotein sequences are identical for the three novel 2019 human isolates and they are closely related to these sequences in six bat and human SARS coronaviruses. This strongly supports the bat origin of the pandemic, novel coronavirus. One surface glycoprotein fragment of 111 amino acids is the largest, conserved, common permutation in the examined bat SARS-like and human SARS viruses, including the Covid-19 virus. BLAST analysis confirmed that this fragment is conserved only in the human and bat SARS strains. This fragment likely is involved in infectivity and is of interest for vaccine development. Surface glycoprotein and nucleocapsid protein sequence homologies of 58.9% and 82.5%, respectively, between the novel SARS0CoV-2 strains and the human SARS (2018) virus suggest that existing anti-SARS vaccines may provide some protection against the novel coronavirus.


Subject(s)
Betacoronavirus/genetics , Coronaviridae/genetics , Coronavirus Infections , Nucleocapsid Proteins/genetics , Pandemics , Pneumonia, Viral , Spike Glycoprotein, Coronavirus/genetics , Algorithms , Amino Acid Sequence , Animals , Chiroptera/virology , Coronaviridae/classification , Genome, Viral/genetics , Humans , Software , Species Specificity , Viral Envelope Proteins/genetics
7.
Vet Microbiol ; 244: 108693, 2020 May.
Article in English | MEDLINE | ID: mdl-32402329

ABSTRACT

The recent pandemic caused by the novel human coronavirus, referrred to as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), not only is having a great impact on the health care systems and economies in all continents but it is also causing radical changes of common habits and life styles. The novel coronavirus (CoV) recognises, with high probability, a zoonotic origin but the role of animals in the SARS-CoV-2 epidemiology is still largely unknown. However, CoVs have been known in animals since several decades, so that veterinary coronavirologists have a great expertise on how to face CoV infections in animals, which could represent a model for SARS-CoV-2 infection in humans. In the present paper, we provide an up-to-date review of the literature currently available on animal CoVs, focusing on the molecular mechanisms that are responsible for the emergence of novel CoV strains with different antigenic, biologic and/or pathogenetic features. A full comprehension of the mechanisms driving the evolution of animal CoVs will help better understand the emergence, spreading, and evolution of SARS-CoV-2.


Subject(s)
Coronaviridae/classification , Coronavirus Infections/veterinary , Pneumonia, Viral/epidemiology , Pneumonia, Viral/virology , Animals , Betacoronavirus/classification , Betacoronavirus/genetics , Betacoronavirus/isolation & purification , Coronaviridae/genetics , Coronavirus Infections/epidemiology , Coronavirus Infections/transmission , Coronavirus Infections/virology , Disease Models, Animal , Evolution, Molecular , Humans , Pandemics , Pneumonia, Viral/transmission , Zoonoses/epidemiology , Zoonoses/transmission , Zoonoses/virology
10.
Infect Genet Evol ; 81: 104272, 2020 07.
Article in English | MEDLINE | ID: mdl-32142938

ABSTRACT

The seventh novel human infecting Betacoronavirus that causes pneumonia (2019 novel coronavirus, 2019-nCoV) originated in Wuhan, China. The evolutionary relationship between 2019-nCoV and the other human respiratory illness-causing coronavirus is not closely related. We sought to characterize the relationship of the translated proteins of 2019-nCoV with other species of Orthocoronavirinae. A phylogenetic tree was constructed from the genome sequences. A cluster tree was developed from the profiles retrieved from the presence and absence of homologs of ten 2019-nCoV proteins. The combined data were used to characterize the relationship of the translated proteins of 2019-nCoV to other species of Orthocoronavirinae. Our analysis reliably suggests that 2019-nCoV is most closely related to BatCoV RaTG13 and belongs to subgenus Sarbecovirus of Betacoronavirus, together with SARS coronavirus and Bat-SARS-like coronavirus. The phylogenetic profiling cluster of homolog proteins of one annotated 2019-nCoV protein against other genome sequences revealed two clades of ten 2019-nCoV proteins. Clade 1 consisted of a group of conserved proteins in Orthocoronavirinae comprising Orf1ab polyprotein, Nucleocapsid protein, Spike glycoprotein, and Membrane protein. Clade 2 comprised six proteins exclusive to Sarbecovirus and Hibecovirus. Two of six Clade 2 nonstructural proteins, NS7b and NS8, were exclusively conserved among 2019-nCoV, BetaCoV_RaTG, and BatSARS-like Cov. NS7b and NS8 have previously been shown to affect immune response signaling in the SARS-CoV experimental model. Thus, we speculated that knowledge of the functional changes in the NS7b and NS8 proteins during evolution may provide important information to explore the human infective property of 2019-nCoV.


Subject(s)
Betacoronavirus/classification , Betacoronavirus/genetics , Evolution, Molecular , Phylogeny , Viral Nonstructural Proteins/genetics , Coronaviridae/classification , Coronaviridae/genetics
12.
São Paulo; AMHB; mar. 3, 2020. 21 p.
Monography in Portuguese | LILACS, HomeoIndex Homeopathy, MOSAICO - Integrative health | ID: biblio-1087382

ABSTRACT

Estamos no meio da pandemia do Covid-19, em franco desenvolvimento no Brasil. No momento há perspectivas terapêuticas medicamentosa sem fases iniciais de teste, especialmente para o tratamento dos casos graves, aqueles que implicam em internação ou encaminhamento para Unidades de Terapia Intensiva. Pouco tem sido abordado sobre tentativas de tratamento de pacientes com síndrome respiratória leve. Não há escolha terapêutica efetiva para o início do quadro e, desta forma, a doença cursa de acordo com a resposta imune ou suscetibilidade individual do acometido, e há muito pouco de efetivo e específico que altere a história natural dessa enfermidade. (AU)


Subject(s)
Arsenicum Album/therapeutic use , China officinalis/therapeutic use , Chininum Arsenicosum/therapeutic use , Epidemic Genius , Coronaviridae , Coronaviridae Infections , Bryonia , Pandemics , Homeopathy , Brazil/epidemiology
14.
Mem Inst Oswaldo Cruz ; 114: e190198, 2019.
Article in English | MEDLINE | ID: mdl-31596309

ABSTRACT

BACKGROUND: In Brazil the implementation of the Sentinel Surveillance System of Influenza began in 2000. Central public health laboratories use reverse transcription-quantitative polymerase chain reaction (RT-qPCR) for diagnosis of respiratory viruses, but this protocol identifies only specific targets, resulted in inconclusive diagnosis for many samples. Thus, high-throughput sequencing (HTS) would be complementary method in the identification of pathogens in inconclusive samples for RT-qPCR or other specific detection protocols. OBJECTIVES: This study aimed to detect unidentified viruses using HTS approach in negative samples of nasopharynx/tracheal secretions by the standard RT-qPCR collected in the Federal District, Brazil. METHODS: Nucleic acids were extracted from samples collected in winter period of 2016 and subjected to HTS. The results were confirmed by the multiplex PR21 RT-qPCR, which identifies 21 respiratory pathogens. FINDINGS: The main viruses identified by HTS were of families Herpesviridae, Coronaviridae, Parvoviridae and Picornaviridae, with the emphasis on rhinoviruses. The presence of respiratory viruses in the samples was confirmed by the PR21 multiplex RT-qPCR. Coronavirus, enterovirus, bocavirus and rhinovirus were found by multiplex RT-qPCR as well as by HTS analyses. MAIN CONCLUSIONS: Wide virus diversity was found by different methodologies and high frequency of rhinovirus occurrence was confirmed in population in winter, showing its relevance for public health.


Subject(s)
Coronaviridae/isolation & purification , Herpesviridae/isolation & purification , Nasopharynx/virology , Parvoviridae/isolation & purification , Picornaviridae/isolation & purification , Trachea/virology , Coronaviridae/classification , Coronaviridae/genetics , DNA, Viral/genetics , Herpesviridae/classification , Herpesviridae/genetics , Humans , Parvoviridae/classification , Parvoviridae/genetics , Picornaviridae/classification , Picornaviridae/genetics , RNA, Viral/genetics , Real-Time Polymerase Chain Reaction , Reverse Transcriptase Polymerase Chain Reaction
15.
J Microbiol Biotechnol ; 29(11): 1817-1829, 2019 Nov 28.
Article in English | MEDLINE | ID: mdl-31546302

ABSTRACT

Porcine deltacoronavirus (PDCoV) is an emerging swine enteric coronavirus that causes diarrhea in neonatal piglets. Like other coronaviruses, PDCoV encodes at least three accessory or species-specific proteins; however, the biological roles of these proteins in PDCoV replication remain undetermined. As a first step toward understanding the biology of the PDCoV accessory proteins, we established a stable porcine cell line constitutively expressing the PDCoV NS7 protein in order to investigate the functional characteristics of NS7 for viral replication. Confocal microscopy and subcellular fractionation revealed that the NS7 protein was extensively distributed in the mitochondria. Proteomic analysis was then conducted to assess the expression dynamics of the host proteins in the PDCoV NS7-expressing cells. Highresolution two-dimensional gel electrophoresis initially identified 48 protein spots which were differentially expressed in the presence of NS7. Seven of these spots, including two upregulated and five down-regulated protein spots, showed statistically significant alterations, and were selected for subsequent protein identification. The affected cellular proteins identified in this study were classified into functional groups involved in various cellular processes such as cytoskeleton networks and cell communication, metabolism, and protein biosynthesis. A substantial down-regulation of α-actinin-4 was confirmed in NS7-expressing and PDCoV-infected cells. These proteomic data will provide insights into the understanding of specific cellular responses to the accessory protein during PDCoV infection.


Subject(s)
Coronaviridae Infections/veterinary , Coronaviridae/physiology , Swine Diseases/virology , Viral Regulatory and Accessory Proteins/metabolism , Actinin/metabolism , Animals , Cell Line , Coronaviridae/genetics , Coronaviridae/metabolism , Coronaviridae Infections/virology , Host-Pathogen Interactions , Mitochondria/metabolism , Proteomics , Swine , Viral Regulatory and Accessory Proteins/genetics
16.
Mol Cell Probes ; 47: 101435, 2019 10.
Article in English | MEDLINE | ID: mdl-31415867

ABSTRACT

Currently in China, porcine epidemic diarrhea virus (PEDV), transmissible gastroenteritis virus (TGEV), porcine rotavirus (PoRV), and porcine deltacoronavirus (PDCoV) are the major causes of porcine viral diarrhea, and mixed infections in clinics are common, resulting in significant economic losses in pig industry. Here, a dual priming oligonucleotide (DPO)-based multiplex real-time SYBR Green RT-PCR assay were developed for accurately differentiating PEDV, TGEV, PoRV, and PDCoV in clinical specimens targeting the N gene of TGEV, PEDV, and PDCoV, and the VP7 gene of PoRV. Results showed that the DPO primer allowed a wider annealing temperature range (40-65 °C) and had a higher priming specificity compared to conventional primer, in which more than 3 nucleotides in the 3'- or 5'-segment of DPO primer mismatched with DNA template, PCR amplification efficiency would decrease substantially or extension would not proceed. DPO-based multiplex real-time RT-PCR method had analytical detection limit of 8.63 × 102 copies/µL, 1.92 × 102 copies/µL, 1.74 × 102 copies/µL, and 1.76 × 102 copies/µL for PEDV, TGEV, PoRV, and PDCoV in clinical specimens, respectively. A total of 672 clinical specimens of piglets with diarrheal symptoms were collected in Northeastern China from 2017 to 2018 followed by analysis using the assay, and epidemiological investigation results showed that PEDV, TGEV, PoRV, and PDCoV prevalence was 19.05%, 5.21%, 4.32%, and 3.87%, respectively. The assay developed in this study showed higher detection accuracy than conventional RT-PCR method, suggesting a useful tool for the accurate differentiation of the four major viruses causing porcine viral diarrhea in practice.


Subject(s)
Coronaviridae/classification , DNA Primers/genetics , Diarrhea/veterinary , Real-Time Polymerase Chain Reaction/methods , Swine Diseases/virology , Animals , Coronaviridae/genetics , Coronaviridae/isolation & purification , Coronavirus/genetics , Coronavirus/isolation & purification , Diarrhea/virology , Porcine epidemic diarrhea virus/genetics , Porcine epidemic diarrhea virus/isolation & purification , RNA, Viral/genetics , Rotavirus/genetics , Rotavirus/isolation & purification , Species Specificity , Swine , Transmissible gastroenteritis virus/genetics , Transmissible gastroenteritis virus/isolation & purification
17.
J Wildl Dis ; 55(1): 105-112, 2019 01.
Article in English | MEDLINE | ID: mdl-30216128

ABSTRACT

We identified two RNA (paramyxovirus and coronavirus) and two DNA (adenovirus and herpesvirus) viruses in a common aquatic bird, the Neotropic Cormorant ( Phalacrocorax brasilianus), and determined their phylogenetic relationships to other global circulating variants. We analyzed 104 cloacal swabs from individuals collected at locations in Central Chile. Sequences were obtained from amplicons using consensus primers targeting conserved genes of the virus families Paramyxoviridae, Coronaviridae, Adenoviridae, and Herpesviridae. A total of 20.2% of the samples was positive for coronavirus, 8.7% for adenovirus, and 3.8% for herpesvirus. No paramyxoviruses were detected. All coronaviruses were identified as viruses of the Gammacoronavirus genus, closely related to the infectious bronchitis virus clade (bootstrap clade support=75%). All adenovirus samples were identified as Aviadenovirus, related to a gull and falcon adenovirus (Bayesian posterior probability=0.86). The herpesviruses identified were related to the infectious laryngotracheitis virus ( Gallid herpesvirus 1) of the genus Iltovirus (bootstrap clade support=99%). We provide information about the diversity of viruses circulating among apparently healthy Neotropic Cormorants.


Subject(s)
Adenoviridae/genetics , Bird Diseases/virology , Birds/virology , Coronaviridae/genetics , Herpesviridae/genetics , Adenoviridae/isolation & purification , Adenoviridae Infections/epidemiology , Adenoviridae Infections/veterinary , Adenoviridae Infections/virology , Animals , Bird Diseases/epidemiology , Chile/epidemiology , Coronaviridae/isolation & purification , Coronavirus Infections/epidemiology , Coronavirus Infections/veterinary , Coronavirus Infections/virology , Herpesviridae/isolation & purification , Herpesviridae Infections/epidemiology , Herpesviridae Infections/veterinary , Herpesviridae Infections/virology , Phylogeny
18.
Virology ; 524: 160-171, 2018 11.
Article in English | MEDLINE | ID: mdl-30199753

ABSTRACT

Transcriptomics has the potential to discover new RNA virus genomes by sequencing total intracellular RNA pools. In this study, we have searched publicly available transcriptomes for sequences similar to viruses of the Nidovirales order. We report two potential nidovirus genomes, a highly divergent 35.9 kb likely complete genome from the California sea hare Aplysia californica, which we assign to a nidovirus named Aplysia abyssovirus 1 (AAbV), and a coronavirus-like 22.3 kb partial genome from the ornamented pygmy frog Microhyla fissipes, which we assign to a nidovirus named Microhyla alphaletovirus 1 (MLeV). AAbV was shown to encode a functional main proteinase, and a translational readthrough signal. Phylogenetic analysis suggested that AAbV represents a new family, proposed here as Abyssoviridae. MLeV represents a sister group to the other known coronaviruses. The importance of MLeV and AAbV for understanding nidovirus evolution, and the origin of terrestrial nidoviruses are discussed.


Subject(s)
Coronaviridae/classification , Genome, Viral/genetics , Nidovirales Infections/virology , Nidovirales/classification , Transcriptome , California , Computational Biology , Coronaviridae/genetics , Coronaviridae/isolation & purification , Evolution, Molecular , Nidovirales/genetics , Nidovirales/isolation & purification , Peptide Chain Termination, Translational/genetics , Peptide Hydrolases/genetics , Phylogeny , Viral Proteins/genetics , Virion
19.
J Gen Virol ; 99(9): 1253-1260, 2018 09.
Article in English | MEDLINE | ID: mdl-30058998

ABSTRACT

White bream virus (WBV), a poorly characterized plus-strand RNA virus infecting freshwater fish of the Cyprinidae family, is the prototype species of the genus Bafinivirus in the subfamily Torovirinae (family Coronaviridae, order Nidovirales). In common with other nidoviruses featuring >20 kilobase genomes, bafiniviruses have been predicted to encode an exoribonuclease (ExoN) in their replicase gene. Here, we used information on the substrate specificity of bafinivirus 3C-like proteases to express WBV ExoN in an active form in Escherichia coli. The 374-residue protein displayed robust 3'-to-5' exoribonuclease activity in the presence of Mg2+ ions and, unlike its coronavirus homologues, did not require a protein cofactor for activity. Characterization of mutant forms of ExoN provided support for predictions on putative active-site and conserved zinc-binding residues. WBV ExoN was revealed to be most active on double-stranded RNA substrates containing one or two non-paired 3'-terminal nucleotides, supporting its presumed role in increasing the fidelity of the bafinivirus RNA-dependent RNA polymerase.


Subject(s)
Coronaviridae/enzymology , Exoribonucleases/metabolism , Viral Proteins/metabolism , Coronaviridae/metabolism , Exoribonucleases/genetics , Gene Expression Regulation, Enzymologic , Gene Expression Regulation, Viral , RNA, Double-Stranded , RNA, Viral , Substrate Specificity , Virus Replication
20.
Virol J ; 15(1): 95, 2018 05 25.
Article in English | MEDLINE | ID: mdl-29801460

ABSTRACT

BACKGROUND: Emergence and re-emergence of porcine epidemic diarrhea virus (PEDV) in North America, Asia and Europe has caused severe economic loss to the global swine industry. However, the virome of PEDV infected pigs and its effect on disease severity remains unknown. The advancements of sequencing technology have made it possible to characterize the entire microbiome of different body sites for any host. METHODS: The objective of this study was to characterize the RNA virome in PEDV-positive pigs using the hypothesis-free metagenomics approach based on next-generation sequencing. Specifically, 217 PEDV-positive swine fecal swab samples collected from diarrheic piglets over 17 US states during 2015-2016 were analyzed. RESULTS: A Kraken algorithm-based bioinformatics analysis revealed the presence of up to 9 different RNA genera besides PEDV (Alphacoronavirus genus), including Mamastrovirus (52%, 113/217), Enterovirus (39%, 85/217), Sapelovirus (31%, 67/217), Posavirus (30%, 66/217), Kobuvirus (23%, 49/217), Sapovirus (13%, 28/217), Teschovirus (10%, 22/217), Pasivirus (9%, 20/217), and Deltacoronavirus (3%, 6/217). There were 58 out of 217 piglets (27%) have PEDV infection alone whereas the remaining 159 (73%) shed 2 up to 9 different viruses. CONCLUSION: These findings demonstrated that PEDV infected diarrheic pigs had an extensive RNA viral flora consisting of four different families: Astroviridae, Picornaviridae, Caliciviridae, and Coronaviridae.


Subject(s)
Astroviridae/genetics , Caliciviridae/genetics , Coronaviridae/genetics , Coronavirus Infections/veterinary , Picornaviridae/genetics , Porcine epidemic diarrhea virus/genetics , Swine Diseases/epidemiology , Algorithms , Amino Acid Sequence , Animals , Astroviridae/classification , Astroviridae/isolation & purification , Caliciviridae/classification , Caliciviridae/isolation & purification , Coinfection , Computational Biology , Coronaviridae/classification , Coronaviridae/isolation & purification , Coronavirus Infections/epidemiology , Coronavirus Infections/virology , Farms , Feces/virology , High-Throughput Nucleotide Sequencing , Metagenomics/methods , Phylogeny , Picornaviridae/classification , Picornaviridae/isolation & purification , Porcine epidemic diarrhea virus/classification , Porcine epidemic diarrhea virus/isolation & purification , RNA, Viral/genetics , Sequence Alignment , Sequence Analysis, DNA , Sequence Homology, Amino Acid , Swine , Swine Diseases/virology , United States/epidemiology
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