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1.
Int J Mol Sci ; 21(12)2020 Jun 26.
Artigo em Inglês | MEDLINE | ID: mdl-32604724

RESUMO

In the 21st century, three highly pathogenic betacoronaviruses have emerged, with an alarming rate of human morbidity and case fatality. Genomic information has been widely used to understand the pathogenesis, animal origin and mode of transmission of coronaviruses in the aftermath of the 2002-2003 severe acute respiratory syndrome (SARS) and 2012 Middle East respiratory syndrome (MERS) outbreaks. Furthermore, genome sequencing and bioinformatic analysis have had an unprecedented relevance in the battle against the 2019-2020 coronavirus disease 2019 (COVID-19) pandemic, the newest and most devastating outbreak caused by a coronavirus in the history of mankind. Here, we review how genomic information has been used to tackle outbreaks caused by emerging, highly pathogenic, betacoronavirus strains, emphasizing on SARS-CoV, MERS-CoV and SARS-CoV-2. We focus on shared genomic features of the betacoronaviruses and the application of genomic information to phylogenetic analysis, molecular epidemiology and the design of diagnostic systems, potential drugs and vaccine candidates.


Assuntos
Betacoronavirus/genética , Infecções por Coronavirus/virologia , Genoma Viral , Pandemias/prevenção & controle , Pneumonia Viral/virologia , Animais , Betacoronavirus/imunologia , Infecções por Coronavirus/diagnóstico , Infecções por Coronavirus/tratamento farmacológico , Desenho de Fármacos , Genes Virais , Humanos , Coronavírus da Síndrome Respiratória do Oriente Médio/genética , Epidemiologia Molecular , Filogenia , Pneumonia Viral/diagnóstico , Pneumonia Viral/tratamento farmacológico , Vírus da SARS/genética , Síndrome Respiratória Aguda Grave/virologia , Vacinas Virais/genética , Vacinas Virais/imunologia
2.
Virol J ; 17(1): 86, 2020 06 30.
Artigo em Inglês | MEDLINE | ID: mdl-32605577

RESUMO

The need for timely establishment of a complete diagnostic protocol of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is demanded worldwide. We selected 15 positive novel coronavirus disease 19 (COVID-19) patients with mild or no symptom. Initially, fecal samples were negative in the 67% (10/15) of the cases, while 33% (5/10) of the cases were positive. After serial virus RNA testing, 73% (11/15) of the cases resulted positive to fecal specimens. In particular, 15 days after the first positive respiratory specimens test, 6 fecal specimens became positive for SARS-CoV-2 RNA, while 13 respiratory test returned negative result. In conclusion, qRT-PCR assays of fecal specimens, is an important step to control infection, suggesting that samples remained positive for SARS-CoV-2 RNA longer time then respiratory tract samples. Our results enhance the recent knowledge on this emerging infectious disease and offer suggestions for a more complete diagnostic strategy.


Assuntos
Betacoronavirus/isolamento & purificação , Infecções por Coronavirus/diagnóstico , Fezes/virologia , Pneumonia Viral/diagnóstico , Betacoronavirus/genética , Infecções por Coronavirus/virologia , Feminino , Genes Virais/genética , Humanos , Masculino , Técnicas de Diagnóstico Molecular , Pandemias , Pneumonia Viral/virologia , RNA Viral/genética , RNA Viral/isolamento & purificação , Sistema Respiratório/virologia , Fatores de Tempo , Eliminação de Partículas Virais
3.
Theranostics ; 10(16): 7150-7162, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32641984

RESUMO

In December 2019, a new coronavirus disease (COVID-19) outbreak occurred in Wuhan, China. Severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2), which is the seventh coronavirus known to infect humans, is highly contagious and has rapidly expanded worldwide since its discovery. Quantitative nucleic acid testing has become the gold standard for diagnosis and guiding clinical decisions regarding the use of antiviral therapy. However, the RT-qPCR assays targeting SARS-CoV-2 have a number of challenges, especially in terms of primer design. Primers are the pivotal components of a RT-qPCR assay. Once virus mutation and recombination occur, it is difficult to effectively diagnose viral infection by existing RT-qPCR primers. Some primers and probes have also been made available on the WHO website for reference. However, no previous review has systematically compared the previously reported primers and probes and described how to design new primers in the event of a new coronavirus infection. This review focuses on how primers and probes can be designed methodically and rationally, and how the sensitivity and specificity of the detection process can be improved. This brief review will be useful for the accurate diagnosis and timely treatment of the new coronavirus pneumonia.


Assuntos
Betacoronavirus/genética , Infecções por Coronavirus/diagnóstico , Pneumonia Viral/diagnóstico , RNA Viral/genética , RNA/genética , Reação em Cadeia da Polimerase em Tempo Real/métodos , Sequência de Bases , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/virologia , Desenho de Fármacos , Genes Virais , Humanos , Conformação de Ácido Nucleico , Pandemias , Pneumonia Viral/epidemiologia , Pneumonia Viral/virologia , RNA/química , Sondas RNA/genética , RNA Viral/química , Reação em Cadeia da Polimerase em Tempo Real/estatística & dados numéricos , Sensibilidade e Especificidade , Nanomedicina Teranóstica
4.
In Vivo ; 34(3 Suppl): 1629-1632, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32503821

RESUMO

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a positive-sense single-stranded RNA virus. It is contagious in humans and is the cause of the coronavirus disease 2019 (COVID-19) pandemic. In the current analysis, we searched for SARS-CoV-2 sequences within the human genome. To compare the SARS-CoV-2 genome to the human genome, we used the blast-like alignment tool (BLAT) of the University of California, Santa Cruz Genome Browser. BLAT can align a user sequence of 25 bases or more to the genome. BLAT search results revealed a 117-base pair SARS-CoV-2 sequence in the human genome with 94.6% identity. The sequence was in chromosome 1p within an intronic region of the netrin G1 (NTNG1) gene. The sequence matched a sequence in the SARS-CoV-2 orf1b (open reading frames) gene. The SARS-CoV-2 human sequence lies within non-structural proteins 14 and 15 (NSP14 and NSP15), and is quite close to the viral spike sequence, separated only by NSP16, a 904-base pair sequence. The mechanism for SARS-CoV-2 infection is the binding of the virus spike protein to the membrane-bound form of angiotensin-converting enzyme 2 and internalization of the complex by the host cell. It is probably no accident that a sequence from the SARS-CoV-2 orf1b gene is found in the human NTNG1 gene, implicated in schizophrenia, and that haloperidol, used to treat schizophrenia, may also be a treatment for COVID-19. We suggest, therefore, that it is important to investigate other haloperidol analogs. Among them are benperidol, bromperidol, bromperidol decanoate, droperidol, seperidol hydrochloride, and trifluperidol. These analogs might be valuable in the treatment of COVID-19 and other coronavirus infections.


Assuntos
Betacoronavirus/genética , Cromossomos Humanos Par 1/genética , Exorribonucleases/genética , Genes Virais , Netrina-1/genética , Proteínas não Estruturais Virais/genética , Proteínas Virais/genética , Animais , Antivirais/farmacologia , Antivirais/uso terapêutico , Sequência de Bases , Infecções por Coronavirus/tratamento farmacológico , DNA Complementar/genética , Endorribonucleases/genética , Haloperidol/análogos & derivados , Haloperidol/farmacologia , Haloperidol/uso terapêutico , Humanos , Íntrons/genética , Pan troglodytes/genética , Pandemias , Pneumonia Viral/tratamento farmacológico , RNA Viral/genética , Esquizofrenia/tratamento farmacológico , Esquizofrenia/genética , Alinhamento de Sequência , Homologia de Sequência do Ácido Nucleico , Especificidade da Espécie
5.
Biosens Bioelectron ; 164: 112316, 2020 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-32553350

RESUMO

Recent research suggests that SARS-CoV-2-infected individuals can be highly infectious while asymptomatic or pre-symptomatic, and that an infected person may infect 5.6 other individuals on average. This situation highlights the need for rapid, sensitive SARS-CoV-2 diagnostic assays capable of high-throughput operation that can preferably utilize existing equipment to facilitate broad, large-scale screening efforts. We have developed a CRISPR-based assay that can meet all these criteria. This assay utilizes a custom CRISPR Cas12a/gRNA complex and a fluorescent probe to detect target amplicons produced by standard RT-PCR or isothermal recombinase polymerase amplification (RPA), to allow sensitive detection at sites not equipped with real-time PCR systems required for qPCR diagnostics. We found this approach allowed sensitive and robust detection of SARS-CoV-2 positive samples, with a sample-to-answer time of ~50 min, and a limit of detection of 2 copies per sample. CRISPR assay diagnostic results obtained nasal swab samples of individuals with suspected COVID-19 cases were comparable to paired results from a CDC-approved quantitative RT-PCR (RT-qPCR) assay performed in a state testing lab, and superior to those produced by same assay in a clinical lab, where the RT-qPCR assay exhibited multiple invalid or inconclusive results. Our assay also demonstrated greater analytical sensitivity and more robust diagnostic performance than other recently reported CRISPR-based assays. Based on these findings, we believe that a CRISPR-based fluorescent application has potential to improve current COVID-19 screening efforts.


Assuntos
Betacoronavirus/isolamento & purificação , Sistemas CRISPR-Cas , Infecções por Coronavirus/diagnóstico , Pneumonia Viral/diagnóstico , Sequência de Bases , Betacoronavirus/genética , Técnicas Biossensoriais/métodos , Técnicas Biossensoriais/estatística & dados numéricos , Infecções por Coronavirus/virologia , Corantes Fluorescentes , Genes Virais , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Sequenciamento de Nucleotídeos em Larga Escala/estatística & dados numéricos , Humanos , Técnicas de Amplificação de Ácido Nucleico/métodos , Técnicas de Amplificação de Ácido Nucleico/estatística & dados numéricos , Pandemias , Pneumonia Viral/virologia , Valor Preditivo dos Testes , RNA Viral/análise , RNA Viral/genética , Reprodutibilidade dos Testes , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Reação em Cadeia da Polimerase Via Transcriptase Reversa/estatística & dados numéricos , Sensibilidade e Especificidade
6.
Arch Virol ; 165(8): 1827-1835, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32507978

RESUMO

Human cytomegalovirus (HCMV) infection causes high morbidity and mortality among immunocompromised patients and can remain in a latent state in host cells. Expression of the immediate-early (IE) genes sustains HCMV replication and reactivation. As a novel genome-editing tool, the clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated protein 9 (Cas9) system has been extensively utilized to modify and edit genomic DNA. In the present study, the CRISPR/Cas9 system was used to target the IE region of the HCMV genome via specific single-guide RNAs (sgRNAs). Infection with CRISPR/Cas9/sgRNA lentiviral constructs significantly reduced viral gene expression and virion production in HFF primary fibroblasts and inhibited viral DNA production and reactivation in the THP-1 monocytic cell line. Thus, the CRISPR/Cas9/sgRNA system can accurately and efficiently target HCMV replication and reactivation and represents a novel therapeutic strategy against latent HCMV infection.


Assuntos
Sistemas CRISPR-Cas/genética , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas/genética , Citomegalovirus/genética , Endonucleases/genética , Genes Virais/genética , Replicação Viral/genética , Linhagem Celular , Infecções por Citomegalovirus/virologia , DNA Viral/genética , Fibroblastos/virologia , Edição de Genes/métodos , Expressão Gênica/genética , Células HEK293 , Humanos , RNA Guia/genética , Células THP-1
7.
Emerg Infect Dis ; 26(7): 1610-1612, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32568058

RESUMO

We characterized novel coronaviruses detected in US bottlenose dolphins (BdCoVs) with diarrhea. These viruses are closely related to the other 2 known cetacean coronaviruses, Hong Kong BdCoV and beluga whale CoV. A deletion in the spike gene and insertions in the membrane gene and untranslated regions were found in US BdCoVs (unrelated to severe acute respiratory syndrome coronavirus 2).


Assuntos
Golfinho Nariz-de-Garrafa/virologia , Infecções por Coronavirus/veterinária , Diarreia/veterinária , Gammacoronavirus/classificação , Gammacoronavirus/genética , Animais , Infecções por Coronavirus/virologia , Diarreia/virologia , Gammacoronavirus/isolamento & purificação , Gammacoronavirus/fisiologia , Genes Virais , Genoma Viral , Mutação , Filogenia , Deleção de Sequência , Glicoproteína da Espícula de Coronavírus/genética , Proteínas da Matriz Viral/genética
8.
Viruses ; 12(5)2020 04 30.
Artigo em Inglês | MEDLINE | ID: mdl-32366025

RESUMO

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which first occurred in Wuhan (China) in December of 2019, causes a severe acute respiratory illness with a high mortality rate, and has spread around the world. To gain an understanding of the evolution of the newly emerging SARS-CoV-2, we herein analyzed the codon usage pattern of SARS-CoV-2. For this purpose, we compared the codon usage of SARS-CoV-2 with that of other viruses belonging to the subfamily of Orthocoronavirinae. We found that SARS-CoV-2 has a high AU content that strongly influences its codon usage, which appears to be better adapted to the human host. We also studied the evolutionary pressures that influence the codon usage of five conserved coronavirus genes encoding the viral replicase, spike, envelope, membrane and nucleocapsid proteins. We found different patterns of both mutational bias and natural selection that affect the codon usage of these genes. Moreover, we show here that the two integral membrane proteins (matrix and envelope) tend to evolve slowly by accumulating nucleotide mutations on their corresponding genes. Conversely, genes encoding nucleocapsid (N), viral replicase and spike proteins (S), although they are regarded as are important targets for the development of vaccines and antiviral drugs, tend to evolve faster in comparison to the two genes mentioned above. Overall, our results suggest that the higher divergence observed for the latter three genes could represent a significant barrier in the development of antiviral therapeutics against SARS-CoV-2.


Assuntos
Betacoronavirus/genética , Códon , Coronavirus/genética , Genoma Viral , Composição de Bases , Betacoronavirus/química , Betacoronavirus/fisiologia , Evolução Biológica , Coronavirus/classificação , Genes Virais , Especificidade de Hospedeiro , Mutação , Filogenia
9.
Environ Microbiol ; 22(6): 2001-2006, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32367648

RESUMO

The origin of the SARS-CoV-2 virus remains enigmatic. It is likely to be a continuum resulting from inevitable mutations and recombination events. These genetic changes keep developing in the present epidemic. Mutations tending to deplete the genome in its cytosine content will progressively lead to attenuation as a consequence of Muller's ratchet, but this is counteracted by recombination when different mutants co-infect the same host, in particular, in clusters of infection. Monitoring as a function of time the genome sequences in closely related cases is critical to anticipate the future of SARS-CoV-2 and hence of COVID-19.


Assuntos
Infecções por Coronavirus/imunologia , Infecções por Coronavirus/virologia , Pneumonia Viral/imunologia , Pneumonia Viral/virologia , Sequência de Bases , Betacoronavirus/genética , Betacoronavirus/imunologia , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/patologia , Infecções por Coronavirus/prevenção & controle , Evolução Molecular , Genes Virais/genética , Humanos , Mutação , Pandemias , Pneumonia Viral/epidemiologia , Pneumonia Viral/patologia , Recombinação Genética , Vacinas Virais/imunologia
10.
Anal Chem ; 92(14): 9699-9705, 2020 07 21.
Artigo em Inglês | MEDLINE | ID: mdl-32441935

RESUMO

A novel coronavirus (SARS-CoV-2) was recently identified in patients with acute respiratory disease and spread quickly worldwide. A specific and rapid diagnostic method is important for early identification. The reverse-transcription recombinase-aided amplification (RT-RAA) assay is a rapid detection method for several pathogens. Assays were performed within 5-15 min as a one-step single tube reaction at 39 °C. In this study, we established two RT-RAA assays for the S and orf1ab gene of SARS-CoV-2 using clinical specimens for validation. The analytical sensitivity of the RT-RAA assay was 10 copies for the S and one copy for the orf1ab gene per reaction. Cross-reactions were not observed with any of the other respiratory pathogens. A 100% agreement between the RT-RAA and real-time PCR assays was accomplished after testing 120 respiratory specimens. These results demonstrate that the proposed RT-RAA assay will be beneficial as it is a faster, more sensitive, and more specific tool for the detection of SARS-CoV-2.


Assuntos
Betacoronavirus/química , Betacoronavirus/genética , Infecções por Coronavirus/diagnóstico , Técnicas de Amplificação de Ácido Nucleico/métodos , Pneumonia Viral/diagnóstico , Reação em Cadeia da Polimerase/métodos , Bactérias/química , Bactérias/genética , Reações Cruzadas , Sondas de DNA , Genes Virais , Humanos , Pandemias , Plasmídeos , Infecções Respiratórias/microbiologia , Infecções Respiratórias/virologia , Sensibilidade e Especificidade , Proteínas Virais/genética , Vírus/química , Vírus/genética
12.
J Infect Dis ; 222(2): 223-233, 2020 06 29.
Artigo em Inglês | MEDLINE | ID: mdl-32433742

RESUMO

Severe acute respiratory syndrome coronavirus (SARS-CoV) was discovered as a novel pathogen in the 2002-2003 SARS epidemic. The emergence and disappearance of this pathogen have brought questions regarding its source and evolution. Within the genome sequences of 281 SARS-CoVs, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and SARS-related CoVs (SARSr-CoVs), a ~430 bp genomic region (from 27 701 bp to 28 131 bp in AY390556.1) with regular variations was investigated. This ~430 bp region overlaps with the ORF8 gene and is prone to deletions and nucleotide substitutions. Its complexity suggested the need for a new genotyping method for coronaviruses related to SARS-similar coronaviruses (SARS-CoV, SARSr-CoV, and SARS-CoV-2). Bat SARSr-CoV presented 3 genotypes, of which type 0 is only seen in bat SARSr-CoV, type I is present in SARS in the early phase, and type II is found in all SARS-CoV-2. This genotyping also shows potential usage in distinguishing the SARS-similar coronaviruses from different hosts and geographic areas. This genomic region has important implications for predicting the epidemic trend and studying the evolution of coronavirus.


Assuntos
Betacoronavirus/genética , Genoma Viral , Vírus da SARS/genética , Proteínas da Matriz Viral/genética , Animais , Sequência de Bases , Quirópteros/virologia , Eutérios/virologia , Evolução Molecular , Genes Virais , Variação Genética , Humanos , Fases de Leitura Aberta , Filogenia , Alinhamento de Sequência , Deleção de Sequência , Glicoproteína da Espícula de Coronavírus/genética , Viverridae/virologia
13.
ACS Nano ; 14(6): 7617-7627, 2020 06 23.
Artigo em Inglês | MEDLINE | ID: mdl-32437124

RESUMO

The current outbreak of the pandemic coronavirus disease 2019 (COVID-19) caused by the severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) demands its rapid, convenient, and large-scale diagnosis to downregulate its spread within as well as across the communities. But the reliability, reproducibility, and selectivity of majority of such diagnostic tests fail when they are tested either to a viral load at its early representation or to a viral gene mutated during its current spread. In this regard, a selective "naked-eye" detection of SARS-CoV-2 is highly desirable, which can be tested without accessing any advanced instrumental techniques. We herein report the development of a colorimetric assay based on gold nanoparticles (AuNPs), when capped with suitably designed thiol-modified antisense oligonucleotides (ASOs) specific for N-gene (nucleocapsid phosphoprotein) of SARS-CoV-2, could be used for diagnosing positive COVID-19 cases within 10 min from the isolated RNA samples. The thiol-modified ASO-capped AuNPs agglomerate selectively in the presence of its target RNA sequence of SARS-CoV-2 and demonstrate a change in its surface plasmon resonance. Further, the addition of RNaseH cleaves the RNA strand from the RNA-DNA hybrid leading to a visually detectable precipitate from the solution mediated by the additional agglomeration among the AuNPs. The selectivity of the assay has been monitored in the presence of MERS-CoV viral RNA with a limit of detection of 0.18 ng/µL of RNA having SARS-CoV-2 viral load. Thus, the current study reports a selective and visual "naked-eye" detection of COVID-19 causative virus, SARS-CoV-2, without the requirement of any sophisticated instrumental techniques.


Assuntos
Betacoronavirus/genética , Técnicas Biossensoriais/métodos , Infecções por Coronavirus/diagnóstico , Nanopartículas Metálicas , Proteínas do Nucleocapsídeo/genética , Oligonucleotídeos Antissenso/genética , Pneumonia Viral/diagnóstico , Sequência de Bases , Betacoronavirus/isolamento & purificação , Colorimetria/métodos , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/virologia , Genes Virais , Ouro , Humanos , Nanopartículas Metálicas/ultraestrutura , Microscopia Eletrônica de Transmissão , Nanotecnologia/métodos , Pandemias , Pneumonia Viral/epidemiologia , Pneumonia Viral/virologia , Capuzes de RNA/genética , RNA Viral/genética , Ressonância de Plasmônio de Superfície/métodos
14.
Virology ; 546: 51-66, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32452417

RESUMO

Overlapping genes originate by a mechanism of overprinting, in which nucleotide substitutions in a pre-existing frame induce the expression of a de novo protein from an alternative frame. In this study, I assembled a dataset of 319 viral overlapping genes, which included 82 overlaps whose expression is experimentally known and the respective 237 homologs. Principal component analysis revealed that overlapping genes have a common pattern of nucleotide and amino acid composition. Discriminant analysis separated overlapping from non-overlapping genes with an accuracy of 97%. When applied to overlapping genes with known genealogy, it separated ancestral from de novo frames with an accuracy close to 100%. This high discriminant power was crucial to computationally design variants of de novo viral proteins known to possess selective anticancer toxicity (apoptin) or protection against neurodegeneration (X protein), as well as to detect two new potential overlapping genes in the genome of the new coronavirus SARS-CoV-2.


Assuntos
Betacoronavirus/genética , Evolução Molecular , Homologia de Genes , Genes Virais , Algoritmos , Sequência de Aminoácidos , Sequência de Bases , Biologia Computacional , Simulação por Computador , Análise Discriminante , Análise dos Mínimos Quadrados , Análise de Componente Principal
15.
Nat Microbiol ; 5(5): 668-674, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32341570

RESUMO

Virus taxonomy emerged as a discipline in the middle of the twentieth century. Traditionally, classification by virus taxonomists has been focussed on the grouping of relatively closely related viruses. However, during the past few years, the International Committee on Taxonomy of Viruses (ICTV) has recognized that the taxonomy it develops can be usefully extended to include the basal evolutionary relationships among distantly related viruses. Consequently, the ICTV has changed its Code to allow a 15-rank classification hierarchy that closely aligns with the Linnaean taxonomic system and may accommodate the entire spectrum of genetic divergence in the virosphere. The current taxonomies of three human pathogens, Ebola virus, severe acute respiratory syndrome coronavirus and herpes simplex virus 1 are used to illustrate the impact of the expanded rank structure. This new rank hierarchy of virus taxonomy will stimulate further research on virus origins and evolution, and vice versa, and could promote crosstalk with the taxonomies of cellular organisms.


Assuntos
Classificação , Vírus/classificação , Vírus/genética , Doenças Transmissíveis/virologia , Ebolavirus/classificação , Ecologia , Evolução Molecular , Genes Virais , Herpesvirus Humano 1/classificação , Humanos , Vírus da SARS/classificação
18.
Arch Virol ; 165(6): 1357-1366, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32285202

RESUMO

Since the incursion of avian influenza virus subtype H5N8 in Egypt in late 2016, it has spread rapidly, causing severe losses in poultry production. Multiple introductions of different reassorted strains were observed in 2017. In this study, a genetic characterization of the HA gene was carried out with 31 isolates selected from different governorates and sectors. Fifteen isolates were selected for NA gene sequence analysis. The HA and NA genes were divided into two subgroups (I and II) with positive selection pressure identified at positions 174 and 29, respectively. The HA gene contained two novel mutations in the antigenic sites, A and E. The HA nucleotide sequence identity ranged from 77 to 90% with different vaccine seeds. Full-genome sequence analysis was carried out for eight viruses, representing different governorates and sectors, to identify the predominant reassorted strain in Egypt. All viruses were similar to a reassorted strain of clade 2.3.4.4b that has been identified in Germany, among other countries. Analysis of these viruses revealed mutations specific to Egyptian strains and not the original virus characterized in 2017 (A/duck/Egypt/F446/2017), with a novel antiviral resistance marker, V27A, indicating resistance to amantadine in the M2 protein of two strains. The results indicate increased variability of circulating H5N8 viruses compared to earlier viruses sequenced in 2016 and 2017. The predominant reassorted virus circulating in 2017 and 2018 originated from an early 2017 strain. It is important to continue this surveillance of avian influenza viruses to monitor the evolution of circulating viruses.


Assuntos
Vírus da Influenza A Subtipo H5N8/genética , Influenza Aviária/epidemiologia , Influenza Aviária/virologia , Doenças das Aves Domésticas/epidemiologia , Doenças das Aves Domésticas/virologia , Vírus Reordenados , Animais , Aves/virologia , Surtos de Doenças , Egito/epidemiologia , Genes Virais , Genótipo , Geografia Médica , Vírus da Influenza A Subtipo H5N8/classificação , Filogenia , Aves Domésticas/virologia , RNA Viral
19.
Arch Virol ; 165(6): 1419-1431, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32307603

RESUMO

Sheeppox and goatpox are important transboundary animal viral diseases of sheep and goats caused by sheeppox virus (SPPV) and goatpox virus (GTPV), respectively, of the genus Capripoxvirus, family Poxviridae. Among the proteins encoded by the capripoxvirus (CaPV) genome, ORF095 (vaccinia virus A4L homolog) is an immunodominant virion core protein that plays a pivotal role in virus assembly and morphogenesis. In the present study, sequence analysis of the ORF095 genes of 27 SPPV and GTPV isolates or field samples from different geographical regions of India was performed, and structure was prediction was done by homology modeling. A multiple sequence alignment of different CaPV isolates revealed that CaPV-A4L is highly conserved, with several species-specific signature residues, namely A93, A216, A315, G136 and G146 in GTPV, G47, A63, A168 and A276 in SPPV, and G48 and C98 in lumpy skin disease virus (LSDV). Phylogenetically, the CaPV isolates were separated into three major clusters, GTPV, SPPV and LSDV, based on the complete coding sequence of the CaPV-A4L gene. Genus-specific clustering of poxviruses was observed in phylogenetic analysis based on A4L protein homologs of chordopoxviruses. A secondary structure prediction showed the presence of six α-helices and one ß-sheet as well as some coils. The signature residues identified here are potentially useful for genotyping, and the predicted characteristics of the CaPV-A4L protein make it an ideal candidate for use as an immunogenic or diagnostic antigen for the development of immunoassays in  the sero-evaluation of CaPV in target hosts.


Assuntos
Capripoxvirus/genética , DNA Viral/genética , Genes Virais , Infecções por Poxviridae/veterinária , Animais , Doenças das Cabras/virologia , Cabras/virologia , Índia , Fases de Leitura Aberta , Filogenia , Reação em Cadeia da Polimerase , Alinhamento de Sequência , Análise de Sequência de DNA , Ovinos/virologia , Doenças dos Ovinos/virologia , Especificidade da Espécie
20.
Zhonghua Jie He He Hu Xi Za Zhi ; 43(4): 337-339, 2020 Apr 12.
Artigo em Chinês | MEDLINE | ID: mdl-32294816

RESUMO

The case reports 2 cases of novel coronavirus pneumonia diagnosed by concurrent bronchoalveolar lavage in our hospital, 1 case had a history of epidemiology, clinical symptoms and high imaging suspicion, but repeated negative throat swabs. One patient was diagnosed 2019-nCoV. Before the patient was discharged, the clinical symptoms disappeared, the chest CT showed significant improvement, and the pharynx swab was twice negative, reaching the discharge standard.We detected the ORF 1ab gene, the N gene and the nucleic acid of the new coronavirus in the broncho-alveolar lavage fluid of 2 patients. The results showed that the positive rate of bronchoalveolar lavage for detection of new coronavirus nucleic acid was high, and bronchoalveolar lavage for suspected or confirmed new coronavirus pneumonia patients with negative detection of nucleic acid in pharynx swabs but still residual lung lesions was helpful for early diagnosis, treatment and prognosis.


Assuntos
Líquido da Lavagem Broncoalveolar/virologia , Infecções por Coronavirus/diagnóstico , Pneumonia Viral/diagnóstico , Betacoronavirus , Lavagem Broncoalveolar , Técnicas de Laboratório Clínico , Genes Virais , Humanos , Pandemias , Faringe/virologia
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