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1.
Int J Mol Sci ; 22(16)2021 Aug 17.
Artigo em Inglês | MEDLINE | ID: mdl-34445529

RESUMO

The circulation of the H9N2 virus results in significant economic losses in the poultry industry, and its zoonotic transmission highlights the need for a highly sensitive and rapid diagnostic and detection system for this virus. In this study, the performance of lateral flow test strips for a fluorescent immunochromatographic test (FICT) was optimized for the diagnosis of H9N2 virus-infected animal samples. The novel monoclonal antibodies (McAbs) against influenza A H9 viruses were developed, and two categories of McAbs with linear and conformational epitopes were compared for the performance of rapid diagnostic performance in the presence of feces sample at different time points (2, 4, and 6 days) post-infection (dpi). The limit of detection (LOD) of FICT and Kd values were comparable between linear and conformational epitope McAbs. However, superior performance of linear epitope McAbs pairs were confirmed by two animal studies, showing the better diagnostic performance showing 100% relative sensitivity in fecal samples at 6 dpi although it showed less than 80% sensitivity in early infection. Our results imply that the comparable performance of the linear epitope McAbs can potentially improve the diagnostic performance of FICT for H9N2 detection in feces samples. This highly sensitive rapid diagnostic method can be utilized in field studies of broiler poultry and wild birds.


Assuntos
Fezes/virologia , Fluorescência , Vírus da Influenza A Subtipo H9N2/isolamento & purificação , Influenza Aviária/diagnóstico , Infecções por Orthomyxoviridae/diagnóstico , Doenças das Aves Domésticas/diagnóstico , Animais , Galinhas , Testes Diagnósticos de Rotina , Feminino , Vírus da Influenza A Subtipo H9N2/imunologia , Influenza Aviária/virologia , Limite de Detecção , Camundongos , Camundongos Endogâmicos BALB C , Infecções por Orthomyxoviridae/virologia , Doenças das Aves Domésticas/virologia
2.
J Med Virol ; 93(10): 5998-6007, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34232513

RESUMO

In the context of the coronavirus disease 2019 pandemic, we investigated the epidemiological and clinical characteristics of a young patient infected by avian influenza A (H5N6) virus in Anhui Province, East China, and analyzed genomic features of the pathogen in 2020. Through the cross-sectional investigation of external environment monitoring (December 29-31, 2020), 1909 samples were collected from Fuyang City. It was found that the positive rate of H5N6 was higher than other areas obviously in Tianma poultry market, where the case appeared. In addition, dual coinfections were detected with a 0.057% polymerase chain reaction positive rate the surveillance years. The virus was the clade 2.3.4.4, which was most likely formed by genetic reassortment between H5N6 and H9N2 viruses. This study found that the evolution rates of the hemagglutinin and neuraminidase genes of the virus were higher than those of common seasonal influenza viruses. The virus was still highly pathogenic to poultry and had a preference for avian receptor binding.


Assuntos
COVID-19/epidemiologia , Vírus da Influenza A/isolamento & purificação , Influenza Aviária/virologia , Influenza Humana/virologia , Animais , Pré-Escolar , China , Feminino , Genoma Viral/genética , Humanos , Vírus da Influenza A/classificação , Vírus da Influenza A/genética , Influenza Humana/diagnóstico , Mutação , Filogenia , Aves Domésticas/virologia , Vírus Reordenados/classificação , Vírus Reordenados/genética , Vírus Reordenados/isolamento & purificação , SARS-CoV-2 , Proteínas Virais/genética
3.
Viruses ; 13(6)2021 06 02.
Artigo em Inglês | MEDLINE | ID: mdl-34199456

RESUMO

Beginning in late 2017, highly pathogenic avian influenza (HPAI) H5N6 viruses caused outbreaks in wild birds and poultry in several European countries. H5N6 viruses were detected in 43 wild birds found dead throughout Denmark. Most of the Danish virus-positive dead birds were found in the period from February to April 2018. However, unlike the rest of Europe, sporadic HPAI H5N6-positive dead wild birds were detected in Denmark in July, August, September, and December 2018, with the last positive bird being found in January 2019. HPAI viruses were not detected in active surveillance of apparently healthy wild birds. In this study, we use full genome sequencing and phylogenetic analysis to investigate the wild bird HPAI H5N6 viruses found in Denmark. The Danish viruses were found to be closely related to those of contemporary HPAI H5N6 viruses detected in Europe. Their sequences formed two clusters indicating that at least two or more introductions of H5N6 into Denmark occurred. Notably, all viruses detected in the latter half of 2018 and in 2019 grouped into the same cluster. The H5N6 viruses appeared to have been maintained undetected in the autumn 2018.


Assuntos
Vírus da Influenza A/genética , Influenza Aviária/epidemiologia , Influenza Aviária/virologia , Animais , Animais Selvagens , Teorema de Bayes , Aves , Dinamarca/epidemiologia , Surtos de Doenças/veterinária , Evolução Molecular , Geografia Médica , História do Século XXI , Vírus da Influenza A/classificação , Vírus da Influenza A/isolamento & purificação , Influenza Aviária/história , Influenza Aviária/transmissão , Filogenia , Vigilância em Saúde Pública , RNA Viral
4.
J Gen Virol ; 102(6)2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-34061017

RESUMO

Avian H9N2 influenza viruses in East Asia are genetically diversified and multiple genotypes (A-W) have been established in poultry. Genotype S strains are currently the most prevalent strains, have caused many human infections and pose a public health threat. In this study, human adaptation mutations in the PB2 polymerase in genotype S strains were identified by database screening. Several PB2 double mutations were identified that acted cooperatively to produce higher genotype S virus polymerase activity and replication in human cells than in avian cells and to increase viral growth and virulence in mice. These mutations were chronologically and phylogenetically clustered in a new group within genotype S viruses. Most of the relevant human virus isolates carry the PB2-A588V mutation together with another PB2 mutation (i.e. K526R, E627V or E627K), indicating a host adaptation advantage for these double mutations. The prevalence of PB2 double mutations in human H9N2 virus isolates has also been found in genetically related human H7N9 and H10N8 viruses. These results suggested that PB2 double mutations in viruses in the field acted cooperatively to increase human adaptation of the currently prevalent H9N2 genotype S strains. This may have contributed to the recent surge of H9N2 infections and may be applicable to the human adaptation of several other avian influenza viruses. Our study provides a better understanding of the human adaptation pathways of genetically related H9N2, H7N9 and H10N8 viruses in nature.


Assuntos
Adaptação ao Hospedeiro , Vírus da Influenza A Subtipo H9N2/genética , Vírus da Influenza A Subtipo H9N2/fisiologia , Influenza Humana/virologia , RNA Polimerase Dependente de RNA/genética , RNA Polimerase Dependente de RNA/metabolismo , Proteínas Virais/genética , Proteínas Virais/metabolismo , Replicação Viral , Animais , Aves , Linhagem Celular , Genes Virais , Genótipo , Células HEK293 , Humanos , Vírus da Influenza A Subtipo H9N2/classificação , Vírus da Influenza A Subtipo H9N2/isolamento & purificação , Influenza Aviária/virologia , Camundongos , Camundongos Endogâmicos BALB C , Modelos Moleculares , Mutação , Infecções por Orthomyxoviridae/virologia , Filogenia , Aves Domésticas , RNA Polimerase Dependente de RNA/química , Vírus Reordenados/genética , Proteínas Virais/química , Zoonoses Virais , Virulência/genética
5.
Viruses ; 13(5)2021 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-34065033

RESUMO

Since late 2020, outbreaks of H5 highly pathogenic avian influenza (HPAI) viruses belonging to clade 2.3.4.4b have emerged in Europe. To investigate the evolutionary history of these viruses, we performed genetic characterization on the first HPAI viruses found in Denmark during the autumn of 2020. H5N8 viruses from 14 wild birds and poultry, as well as one H5N5 virus from a wild bird, were characterized by whole genome sequencing and phylogenetic analysis. The Danish H5N8 viruses were found to be genetically similar to each other and to contemporary European clade 2.3.4.4b H5N8 viruses, while the Danish H5N5 virus was shown to be a unique genotype from the H5N5 viruses that circulated at the same time in Russia, Germany, and Belgium. Genetic analyses of one of the H5N8 viruses revealed the presence of a substitution (PB2-M64T) that is highly conserved in human seasonal influenza A viruses. Our analyses showed that the late 2020 clade 2.3.4.4b HPAI H5N8 viruses were most likely new incursions introduced by migrating birds to overwintering sites in Europe, rather than the result of continued circulation of H5N8 viruses from previous introductions to Europe in 2016/2017 and early 2020.


Assuntos
Genótipo , Vírus da Influenza A Subtipo H5N8/genética , Vírus da Influenza A/genética , Influenza Humana/epidemiologia , Influenza Humana/virologia , Animais , Dinamarca/epidemiologia , Surtos de Doenças , Genes Virais , Genoma Viral , Geografia Médica , Humanos , Vírus da Influenza A Subtipo H5N8/classificação , Vírus da Influenza A/classificação , Influenza Aviária/epidemiologia , Influenza Aviária/virologia , Filogenia , Vigilância da População , Aves Domésticas/virologia
6.
Viruses ; 13(5)2021 05 07.
Artigo em Inglês | MEDLINE | ID: mdl-34067187

RESUMO

Low-pathogenicity avian influenza viruses (LPAIV) introduced by migratory birds circulate in wild birds and can be transmitted to poultry. These viruses can mutate to become highly pathogenic avian influenza viruses causing severe disease and death in poultry. In March 2019, an H7N3 avian influenza virus-A/Spot-billed duck/South Korea/WKU2019-1/2019 (H7N3)-was isolated from spot-billed ducks in South Korea. This study aimed to evaluate the phylogenetic and mutational analysis of this isolate. Molecular analysis revealed that the genes for HA (hemagglutinin) and NA (neuraminidase) of this strain belonged to the Central Asian lineage, whereas genes for other internal proteins such as polymerase basic protein 1 (PB1), PB2, nucleoprotein, polymerase acidic protein, matrix protein, and non-structural protein belonged to that of the Korean lineage. In addition, a monobasic amino acid (PQIEPR/GLF) at the HA cleavage site, and the non-deletion of the stalk region in the NA gene indicated that this isolate was a typical LPAIV. Nucleotide sequence similarity analysis of HA revealed that the highest homology (99.51%) of this isolate is to that of A/common teal/Shanghai/CM1216/2017 (H7N7), and amino acid sequence of NA (99.48%) was closely related to that of A/teal/Egypt/MB-D-487OP/2016 (H7N3). An in vitro propagation of the A/Spot-billed duck/South Korea/WKU2019-1/2019 (H7N3) virus showed highest (7.38 Log10 TCID50/mL) virus titer at 60 h post-infection, and in experimental mouse lungs, the virus was detected at six days' post-infection. Our study characterizes genetic mutations, as well as pathogenesis in both in vitro and in vivo model of a new Korea H7N3 viruses in 2019, carrying multiple potential mutations to become highly pathogenic and develop an ability to infect humans; thus, emphasizing the need for routine surveillance of avian influenza viruses in wild birds.


Assuntos
Patos/virologia , Vírus da Influenza A Subtipo H7N3/classificação , Vírus da Influenza A Subtipo H7N3/genética , Influenza Aviária/epidemiologia , Influenza Aviária/virologia , Animais , Animais Selvagens/virologia , Células Cultivadas , Feminino , Genes Virais , Genoma Viral , Genômica/métodos , História do Século XXI , Especificidade de Hospedeiro , Vírus da Influenza A Subtipo H7N3/isolamento & purificação , Influenza Aviária/história , Camundongos , Infecções por Orthomyxoviridae/patologia , Infecções por Orthomyxoviridae/virologia , Filogenia , Vigilância em Saúde Pública , Vírus Reordenados , República da Coreia/epidemiologia , Replicação Viral
7.
Viruses ; 13(5)2021 05 10.
Artigo em Inglês | MEDLINE | ID: mdl-34068495

RESUMO

Avian influenza virus A (H7N9), after circulating in avian hosts for decades, was identified as a human pathogen in 2013. Herein, amino acid substitutions possibly essential for human adaptation were identified by comparing the 4706 aligned overlapping nonamer position sequences (1-9, 2-10, etc.) of the reported 2014 and 2017 avian and human H7N9 datasets. The initial set of virus sequences (as of year 2014) exhibited a total of 109 avian-to-human (A2H) signature amino acid substitutions. Each represented the most prevalent substitution at a given avian virus nonamer position that was selectively adapted as the corresponding index (most prevalent sequence) of the human viruses. The majority of these avian substitutions were long-standing in the evolution of H7N9, and only 17 were first detected in 2013 as possibly essential for the initial human adaptation. Strikingly, continued evolution of the avian H7N9 virus has resulted in avian and human protein sequences that are almost identical. This rapid and continued adaptation of the avian H7N9 virus to the human host, with near identity of the avian and human viruses, is associated with increased human infection and a predicted greater risk of human-to-human transmission.


Assuntos
Adaptação Biológica , Interações Hospedeiro-Patógeno , Subtipo H7N9 do Vírus da Influenza A/fisiologia , Influenza Aviária/virologia , Influenza Humana/virologia , Infecções por Orthomyxoviridae/virologia , Substituição de Aminoácidos , Animais , Aves , Variação Genética , Humanos , RNA Viral , Especificidade da Espécie
8.
BMC Vet Res ; 17(1): 200, 2021 May 28.
Artigo em Inglês | MEDLINE | ID: mdl-34049549

RESUMO

BACKGROUND: Highly pathogenic avian influenza viruses (HPAIVs) of H5 subtype pose a great threat to the poultry industry and human health. In recent years, H5N6 subtype has rapidly replaced H5N1 as the most predominate HPAIV subtype circulating in domestic poultry in China. In this study, we describe the genetic and phylogenetic characteristics of a prevalent H5N6 strain in Guangdong, China. RESULTS: Nucleotide sequencing identified a H5N6 subtype HPAIV, designated as A/chicken/Dongguan/1101/2019 (DG/19), with a multibasic cleavage site in the hemagglutinin (HA). Phylogenetic analysis revealed DG/19 was a reassortant of H5N1, H5N2, H5N8, and H6N6 subtypes of avian influenza viruses. A number of mammalian adaptive markers such as D36N in the HA were identified. CONCLUSIONS: Our results showed that HPAIV H5N6 strains still emerge in well-managed groups of chicken farms. Considering the increasing prevalence of H5N6 HPAIV, and the fact that H5N6 HPAIVs are well adapted to migratory birds, an enhanced surveillance for the East Asian-Australasian flyway should be undertaken to prevent potential threats to the poultry industry and human health.


Assuntos
Galinhas/virologia , Vírus da Influenza A/classificação , Vírus da Influenza A/genética , Influenza Aviária/virologia , Animais , China , Genes Virais , Vírus da Influenza A/isolamento & purificação , Filogenia , Vírus Reordenados/classificação , Vírus Reordenados/genética , Vírus Reordenados/isolamento & purificação
10.
Viruses ; 13(3)2021 03 19.
Artigo em Inglês | MEDLINE | ID: mdl-33808674

RESUMO

Reassortment among co-infecting influenza A viruses (IAVs) is an important source of viral diversity and can facilitate expansion into novel host species. Indeed, reassortment played a key role in the evolution of the last three pandemic IAVs. Observed patterns of reassortment within a coinfected host are likely to be shaped by several factors, including viral load, the extent of viral mixing within the host and the stringency of selection. These factors in turn are expected to vary among the diverse host species that IAV infects. To investigate host differences in IAV reassortment, here we examined reassortment of two distinct avian IAVs within their natural host (mallards) and a mammalian model system (guinea pigs). Animals were co-inoculated with A/wildbird/California/187718-36/2008 (H3N8) and A/mallard/Colorado/P66F1-5/2008 (H4N6) viruses. Longitudinal samples were collected from the cloaca of mallards or the nasal tract of guinea pigs and viral genetic exchange was monitored by genotyping clonal isolates from these samples. Relative to those in guinea pigs, viral populations in mallards showed higher frequencies of reassortant genotypes and were characterized by higher genotype richness and diversity. In line with these observations, analysis of pairwise segment combinations revealed lower linkage disequilibrium in mallards as compared to guinea pigs. No clear longitudinal patterns in richness, diversity or linkage disequilibrium were present in either host. Our results reveal mallards to be a highly permissive host for IAV reassortment and suggest that reduced viral mixing limits avian IAV reassortment in a mammalian host.


Assuntos
Vírus da Influenza A Subtipo H3N8/fisiologia , Influenza Aviária , Infecções por Orthomyxoviridae , Animais , Cães , Patos , Feminino , Cobaias , Influenza Aviária/epidemiologia , Influenza Aviária/virologia , Estudos Longitudinais , Células Madin Darby de Rim Canino , Infecções por Orthomyxoviridae/epidemiologia , Infecções por Orthomyxoviridae/veterinária , Infecções por Orthomyxoviridae/virologia , Vírus Reordenados
11.
Viruses ; 13(3)2021 03 22.
Artigo em Inglês | MEDLINE | ID: mdl-33810083

RESUMO

The threat of a new influenza pandemic is real. With past pandemics claiming millions of lives, finding new ways to combat this virus is essential. Host cells have developed a multi-modular system to detect incoming pathogens, a phenomenon called sensing. The signaling cascade triggered by sensing subsequently induces protection for themselves and their surrounding neighbors, termed interferon (IFN) response. This response induces the upregulation of hundreds of interferon-stimulated genes (ISGs), including antiviral effectors, establishing an antiviral state. As well as the antiviral proteins induced through the IFN system, cells also possess a so-called intrinsic immunity, constituted of antiviral proteins that are constitutively expressed, creating a first barrier preceding the induction of the interferon system. All these combined antiviral effectors inhibit the virus at various stages of the viral lifecycle, using a wide array of mechanisms. Here, we provide a review of mammalian and avian influenza A restriction factors, detailing their mechanism of action and in vivo relevance, when known. Understanding their mode of action might help pave the way for the development of new influenza treatments, which are absolutely required if we want to be prepared to face a new pandemic.


Assuntos
Vírus da Influenza A , Influenza Aviária , Influenza Humana , Animais , Aves , Interações Hospedeiro-Patógeno , Humanos , Imunidade Inata , Vírus da Influenza A/imunologia , Vírus da Influenza A/fisiologia , Influenza Aviária/imunologia , Influenza Aviária/virologia , Influenza Humana/imunologia , Influenza Humana/virologia , Aves Domésticas , Replicação Viral
12.
PLoS Pathog ; 17(4): e1009561, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33905456

RESUMO

The H7N9 avian influenza virus (AIV) that emerged in China have caused five waves of human infection. Further human cases have been successfully prevented since September 2017 through the use of an H7N9 vaccine in poultry. However, the H7N9 AIV has not been eradicated from poultry in China, and its evolution remains largely unexplored. In this study, we isolated 19 H7N9 AIVs during surveillance and diagnosis from February 2018 to December 2019, and genetic analysis showed that these viruses have formed two different genotypes. Animal studies indicated that the H7N9 viruses are highly lethal to chicken, cause mild infection in ducks, but have distinct pathotypes in mice. The viruses bound to avian-type receptors with high affinity, but gradually lost their ability to bind to human-type receptors. Importantly, we found that H7N9 AIVs isolated in 2019 were antigenically different from the H7N9 vaccine strain that was used for H7N9 influenza control in poultry, and that replication of these viruses cannot, therefore, be completely prevented in vaccinated chickens. We further revealed that two amino acid mutations at positions 135 and 160 in the HA protein added two glycosylation sites and facilitated the escape of the H7N9 viruses from the vaccine-induced immunity. Our study provides important insights into H7N9 virus evolution and control.


Assuntos
Subtipo H7N9 do Vírus da Influenza A/genética , Subtipo H7N9 do Vírus da Influenza A/isolamento & purificação , Vacinas contra Influenza/uso terapêutico , Influenza Aviária/prevenção & controle , Doenças das Aves Domésticas/virologia , Animais , Animais de Zoológico/virologia , Galinhas/virologia , China/epidemiologia , Patos/virologia , Controle de Infecções/métodos , Subtipo H7N9 do Vírus da Influenza A/classificação , Subtipo H7N9 do Vírus da Influenza A/fisiologia , Influenza Aviária/epidemiologia , Influenza Aviária/virologia , Camundongos , Filogenia , Vigilância da População , Aves Domésticas , Doenças das Aves Domésticas/epidemiologia , Doenças das Aves Domésticas/prevenção & controle
13.
Emerg Microbes Infect ; 10(1): 822-832, 2021 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-33866955

RESUMO

The enzootic and zoonotic nature of H9N2 avian influenza viruses poses a persistent threat to the global poultry industry and public health. In particular, the emerging sublineage h9.4.2.5 of H9N2 viruses has drawn great attention. In this study, we determined the effects of the flexibility at residues 226 and 227 in the hemagglutinin on the receptor avidity and immune evasion of H9N2 viruses. The solid-phase direct binding assay showed that residue 226 plays a core role in the receptor preference of H9N2 viruses, while residue 227 affects the preference of the virus for a receptor. Consequently, each of these two successive residues can modulate the receptor avidity of H9N2 viruses and influence their potential of zoonotic infection. The antigenic map based on the cross-hemagglutination inhibition (HI) titers revealed that amino acid substitutions at positions 226 or 227 appear to be involved in antigenic drift, potentially resulting in the emergence of H9N2 immune evasion mutants. Further analysis suggested that increased receptor avidity facilitated by residue 226Q or 227M resulted in a reduction in the HI titer. Among the four naturally-occurring amino acid combinations comprising QQ, MQ, LQ, and LM, the number of viruses with LM accounted for 79.64% of the sublineage h9.4.2.5 and the rescued virus with LM exhibited absolute advantages of in vitro and in vivo replication and transmission. Collectively, these data demonstrate that residues 226 and 227 are under selective pressure and their synergistic regulation of receptor avidity and antigenicity is related to the evolution of circulating H9N2 viruses.


Assuntos
Glicoproteínas de Hemaglutininação de Vírus da Influenza/química , Glicoproteínas de Hemaglutininação de Vírus da Influenza/genética , Vírus da Influenza A Subtipo H9N2/imunologia , Influenza Aviária/virologia , Doenças das Aves Domésticas/virologia , Motivos de Aminoácidos , Substituição de Aminoácidos , Animais , Galinhas , Testes de Inibição da Hemaglutinação , Glicoproteínas de Hemaglutininação de Vírus da Influenza/imunologia , Humanos , Evasão da Resposta Imune , Vírus da Influenza A Subtipo H9N2/química , Vírus da Influenza A Subtipo H9N2/genética , Influenza Aviária/genética , Influenza Aviária/imunologia , Doenças das Aves Domésticas/genética , Doenças das Aves Domésticas/imunologia , Domínios Proteicos , Receptores Virais/genética , Receptores Virais/imunologia
14.
Viruses ; 13(4)2021 04 02.
Artigo em Inglês | MEDLINE | ID: mdl-33918166

RESUMO

Genetic analysis of circulating avian influenza viruses (AIVs) in wild birds at different geographical regions during the same period could improve our knowledge about virus transmission dynamics in natural hosts, virus evolution as well as zoonotic potential. Here, we report the genetic and molecular characterization of H6N2 influenza viruses isolated from migratory birds in Turkey, Egypt, and Uganda during 2017-2018. The Egyptian and Turkish isolates were genetically closer to each other than they were to the virus isolated from Uganda. Our results also suggest that multiple reassortment events were involved in the genesis of the isolated viruses. All viruses contained molecular markers previously associated with increased replication and/or pathogenicity in mammals. The results of this study indicate that H6N2 viruses carried by migratory birds on the West Asian/East African and Mediterranean/Black Sea flyways have the potential to transmit to mammals including humans. Additionally, adaptation markers in these viruses indicate the potential risk for poultry, which also increases the possibility of human exposure to these viruses.


Assuntos
Vírus da Influenza A/classificação , Vírus da Influenza A/genética , Influenza Aviária/virologia , Filogenia , Vírus Reordenados/genética , Migração Animal , Animais , Animais Selvagens/virologia , Galinhas/virologia , Egito , Genoma Viral , Humanos , Vírus da Influenza A/isolamento & purificação , Influenza Aviária/transmissão , Aves Domésticas/virologia , Turquia , Uganda
15.
Life Sci Alliance ; 4(6)2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-33820827

RESUMO

Infection of certain influenza viruses is triggered when its HA is cleaved by host cell proteases such as proprotein convertases and type II transmembrane serine proteases (TTSP). HA with a monobasic motif is cleaved by trypsin-like proteases, including TMPRSS2 and HAT, whereas the multibasic motif found in high pathogenicity avian influenza HA is cleaved by furin, PC5/6, or MSPL. MSPL belongs to the TMPRSS family and preferentially cleaves [R/K]-K-K-R↓ sequences. Here, we solved the crystal structure of the extracellular region of human MSPL in complex with an irreversible substrate-analog inhibitor. The structure revealed three domains clustered around the C-terminal α-helix of the SPD. The inhibitor structure and its putative model show that the P1-Arg inserts into the S1 pocket, whereas the P2-Lys and P4-Arg interacts with the Asp/Glu-rich 99-loop that is unique to MSPL. Based on the structure of MSPL, we also constructed a homology model of TMPRSS2, which is essential for the activation of the SARS-CoV-2 spike protein and infection. The model may provide the structural insight for the drug development for COVID-19.


Assuntos
Influenza Aviária/virologia , Proteínas de Membrana/química , Orthomyxoviridae/patogenicidade , Serina Endopeptidases/química , Animais , Aves , Cristalografia por Raios X , Humanos , Conformação Proteica
16.
Res Vet Sci ; 136: 540-549, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-33887563

RESUMO

Despite the low pathogenicity of the H9N2 avian influenza viruses, they can induce severe economic losses in various poultry sectors in conjunction with other factors. In Egypt, low-pathogenic avian influenza (LPAI) H9N2 became endemic in 2011 and has undergone continuous genetic evolution since then. The regular monitoring of the evolution of the virus is necessary to control its spread. During 2017-2020, there were 44 positive samples isolated, and these viruses were genetically sequenced to determine the hemagglutinin (HA) gene circulating in Egypt. The molecular analysis revealed at least nine changes in amino acid residues in comparison with the reference Egyptian strain from the original introduction in 2011 (A/qu/Egypt/113413v/2011), with a similarity of 95%-96%. Amino acid residues 180 and 216 are the most important residues in terms of positive selection pressure. Phylogenetically, the new Egyptian H9N2 viruses in 2017-2020 belonged to a new subcluster related to the strains that had been circulating since 2015. Comparative analysis of the HA gene of LPAI H9N2 viruses in Egypt from 2011 to 2020 supports a continuous evolution through the years with persistent markers.


Assuntos
Galinhas/virologia , Evolução Molecular , Glicoproteínas de Hemaglutininação de Vírus da Influenza/genética , Vírus da Influenza A Subtipo H9N2/genética , Influenza Aviária/virologia , Doenças das Aves Domésticas/virologia , Animais , Egito/epidemiologia , Influenza Aviária/epidemiologia , Filogenia
17.
PLoS Pathog ; 17(4): e1009490, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33891662

RESUMO

Repeated outbreaks due to H3N1 low pathogenicity avian influenza viruses (LPAIV) in Belgium were associated with unusually high mortality in chicken in 2019. Those events caused considerable economic losses and prompted restriction measures normally implemented for eradicating high pathogenicity avian influenza viruses (HPAIV). Initial pathology investigations and infection studies suggested this virus to be able to replicate systemically, being very atypical for H3 LPAIV. Here, we investigate the pathogenesis of this H3N1 virus and propose a mechanism explaining its unusual systemic replication capability. By intravenous and intracerebral inoculation in chicken, we demonstrate systemic spread of this virus, extending to the central nervous system. Endoproteolytic viral hemagglutinin (HA) protein activation by either tissue-restricted serine peptidases or ubiquitous subtilisin-like proteases is the functional hallmark distinguishing (H5 or H7) LPAIV from HPAIV. However, luciferase reporter assays show that HA cleavage in case of the H3N1 strain in contrast to the HPAIV is not processed by intracellular proteases. Yet the H3N1 virus replicates efficiently in cell culture without trypsin, unlike LPAIVs. Moreover, this trypsin-independent virus replication is inhibited by 6-aminohexanoic acid, a plasmin inhibitor. Correspondingly, in silico analysis indicates that plasminogen is recruitable by the viral neuraminidase for proteolytic activation due to the loss of a strongly conserved N-glycosylation site at position 130. This mutation was shown responsible for plasminogen recruitment and neurovirulence of the mouse brain-passaged laboratory strain A/WSN/33 (H1N1). In conclusion, our findings provide good evidence in natural chicken strains for N1 neuraminidase-operated recruitment of plasminogen, enabling systemic replication leading to an unusual high pathogenicity phenotype. Such a gain of function in naturally occurring AIVs representing an established human influenza HA-subtype raises concerns over potential zoonotic threats.


Assuntos
Surtos de Doenças/veterinária , Vírus da Influenza A/patogenicidade , Influenza Aviária/virologia , Neuraminidase/metabolismo , Plasminogênio/metabolismo , Doenças das Aves Domésticas/virologia , Animais , Galinhas , Glicosilação , Vírus da Influenza A/enzimologia , Vírus da Influenza A/fisiologia , Neuraminidase/genética , Replicação Viral
18.
Vet Res ; 52(1): 36, 2021 Mar 03.
Artigo em Inglês | MEDLINE | ID: mdl-33658079

RESUMO

Exosomes are membrane vesicles containing proteins, lipids, DNA, mRNA, and micro RNA (miRNA). Exosomal miRNA from donor cells can regulate the gene expression of recipient cells. Here, Ri chickens were divided into resistant (Mx/A; BF2/B21) and susceptible (Mx/G; BF2/B13) trait by genotyping of Mx and BF2 genes. Then, Ri chickens were infected with H5N1, a highly pathogenic avian influenza virus (HPAIV). Exosomes were purified from blood serum of resistant chickens for small RNA sequencing. Sequencing data were analysed using FastQCv0.11.7, Cutadapt 1.16, miRBase v21, non-coding RNA database, RNAcentral 10.0, and miRDeep2. Differentially expressed miRNAs were determined using statistical methods, including fold-change, exactTest using edgeR, and hierarchical clustering. Target genes were predicted using miRDB. Gene ontology analysis was performed using gProfiler. Twenty miRNAs showed significantly different expression patterns between resistant control and infected chickens. Nine miRNAs were up-regulated and 11 miRNAs were down-regulated in the infected chickens compared with that in the control chickens. In target gene analysis, various immune-related genes, such as cytokines, chemokines, and signalling molecules, were detected. In particular, mitogen-activated protein kinase (MAPK) pathway molecules were highly controlled by differentially expressed miRNAs. The result of qRT-PCR for miRNAs was identical with sequencing data and miRNA expression level was higher in resistant than susceptible chickens. This study will help to better understand the host immune response, particularly exosomal miRNA expression against HPAIV H5N1 and could help to determine biomarkers for disease resistance.


Assuntos
Galinhas , Exossomos/genética , Influenza Aviária/virologia , MicroRNAs/genética , Doenças das Aves Domésticas/virologia , Animais , Virus da Influenza A Subtipo H5N1/fisiologia
19.
J Vet Sci ; 22(2): e21, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-33774937

RESUMO

In this study, we describe the isolation and characterization of previously unreported Y280-lineage H9N2 viruses from two live bird markets in Korea in June 2020. Genetic analysis revealed that they were distinct from previous H9N2 viruses circulating in Korea and had highest homology to A/chicken/Shandong/1844/2019(H9N2) viruses. Their genetic constellation showed they belonged to genotype S, which is the predominant genotype in China since 2010, where genotype S viruses have infected humans and acted as internal gene donors to H5 and H7 zoonotic influenza viruses. Active surveillance and control measures need to be enhanced to protect the poultry industry and public health.


Assuntos
Galinhas , Genótipo , Vírus da Influenza A Subtipo H9N2/genética , Influenza Aviária/virologia , Doenças das Aves Domésticas/virologia , Animais , Vírus da Influenza A Subtipo H9N2/classificação , Filogenia , República da Coreia
20.
Emerg Microbes Infect ; 10(1): 753-761, 2021 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-33754959

RESUMO

Sub-Saharan Africa was historically considered an animal influenza cold spot, with only sporadic highly pathogenic H5 outbreaks detected over the last 20 years. However, in 2017, low pathogenic avian influenza A(H9N2) viruses were detected in poultry in Sub-Saharan Africa. Molecular, phylogenetic, and antigenic characterization of isolates from Benin, Togo, and Uganda showed that they belonged to the G1 lineage. Isolates from Benin and Togo clustered with viruses previously described in Western Africa, whereas viruses from Uganda were genetically distant and clustered with viruses from the Middle East. Viruses from Benin exhibited decreased cross-reactivity with those from Togo and Uganda, suggesting antigenic drift associated with reduced replication in Calu-3 cells. The viruses exhibited mammalian adaptation markers similar to those of the human strain A/Senegal/0243/2019 (H9N2). Therefore, viral genetic and antigenic surveillance in Africa is of paramount importance to detect further evolution or emergence of new zoonotic strains.


Assuntos
Vírus da Influenza A Subtipo H9N2/genética , Vírus da Influenza A Subtipo H9N2/imunologia , Influenza Aviária/virologia , Doenças das Aves Domésticas/virologia , África ao Sul do Saara , Animais , Anticorpos Antivirais/imunologia , Variação Antigênica , Galinhas/virologia , Reações Cruzadas , Evolução Molecular , Humanos , Vírus da Influenza A Subtipo H9N2/patogenicidade , Vírus da Influenza A Subtipo H9N2/fisiologia , Influenza Humana/virologia , Filogenia , Virulência , Replicação Viral
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