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1.
Emerg Microbes Infect ; 9(1): 616-627, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32183621

RESUMO

Live bird markets (LBMs) in Korea have been recognized as a reservoir, amplifier, and source of avian influenza viruses (AIVs); however, little was known about the role of LBMs in the epidemiology of AIVs in Korea until recently. Through 10 years of surveillance (2006-2016) we have isolated and sequenced H9N2 viruses in Korean LBMs. To understand how H9N2 evolves and spreads in Korea, a statistical Bayesian phylogenetic model was used. Phylogenetic analysis suggests that three separate introductions of progenitor gene pools, Korean domestic duck-origin and two wild aquatic bird-origin AIVs, contributed to the generation of the five genotypes of H9N2 viruses in Korea. Phylogenetic reconstruction of ecological states infer that the LBMs are where chickens become infected with the virus, with domestic ducks playing a major role in the transmission and evolution of the H9N2 viruses. Three increases in the genetic diversity of H9N2 viruses were observed and coincided with transitions in host species and the locations (domestic farm, LBM, slaughterhouse, and wild aquatic bird habitat) where the viruses were isolated, accompanying genetic reassortment. Following the introduction of a wild aquatic bird-origin AIVs in 2008, six genes of the Korean lineage H9N2 virus were replaced with genes originating from wild aquatic birds, and viruses with this new genotype became predominant in Korean LBMs.


Assuntos
Vírus da Influenza A Subtipo H9N2/genética , Influenza Aviária/virologia , Animais , Biodiversidade , Aves , Variação Genética , Genótipo , Influenza Aviária/epidemiologia , Filogenia , República da Coreia
2.
Emerg Microbes Infect ; 8(1): 1465-1478, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31608791

RESUMO

The ANP32A is responsible for mammalian-restricted influenza virus polymerase activity. However, the mechanism of ANP32A modulation of polymerase activity remains poorly understood. Here, we report that chicken ANP32A (chANP32A) -X1 and -X2 stimulated mammalian-restricted PB2 627E polymerase activity in a dose-dependent manner. Distinct effects of ANP32A constructs suggested that the 180VK181 residues within chANP32A-X1 are necessary but not sufficient to stimulate PB2 627E polymerase activity. The PB2 N567D, T598V, A613V or F636L mutations promoted PB2 627E polymerase activity and chANP32A-X1 showed additive effects, providing further support that species-specific regulation of ANP32A might be only relevant with the PB2 E627K mutation. Rescue of cycloheximide-mediated inhibition showed that ANP32A is species-specific for modulation of vRNA but not mRNA and cRNA, demonstrating chANP32A-X1 compensated for defective cRNPs produced by PB2 627E virus in mammalian cells. The promoter mutations of cRNA enhanced the restriction of PB2 627E polymerase in mammalian cells, which could be restored by chANP32A-X1, indicating that ANP32A is likely to regulate the interaction of viral polymerase with RNA promoter. Coimmunoprecipitation showed that ANP32A did not affect the primary cRNPs assembly. We propose a model that chANP32A-X1 regulates PB2 627E polymerase for suitable interaction with cRNA promoter for vRNA replication.


Assuntos
Vírus da Influenza A Subtipo H1N1/enzimologia , Subtipo H7N9 do Vírus da Influenza A/enzimologia , Vírus da Influenza A Subtipo H9N2/enzimologia , Influenza Aviária/metabolismo , Influenza Humana/metabolismo , Doenças das Aves Domésticas/metabolismo , RNA Replicase/metabolismo , Proteínas de Ligação a RNA/metabolismo , Proteínas Virais/metabolismo , Sequência de Aminoácidos , Animais , Galinhas , Humanos , Vírus da Influenza A Subtipo H1N1/genética , Vírus da Influenza A Subtipo H1N1/fisiologia , Subtipo H7N9 do Vírus da Influenza A/genética , Subtipo H7N9 do Vírus da Influenza A/fisiologia , Vírus da Influenza A Subtipo H9N2/genética , Vírus da Influenza A Subtipo H9N2/fisiologia , Influenza Aviária/genética , Influenza Aviária/virologia , Influenza Humana/genética , Influenza Humana/virologia , Mutação , Doenças das Aves Domésticas/genética , Doenças das Aves Domésticas/virologia , Ligação Proteica , RNA Replicase/genética , RNA Viral/genética , RNA Viral/metabolismo , Proteínas de Ligação a RNA/química , Proteínas de Ligação a RNA/genética , Alinhamento de Sequência , Especificidade da Espécie , Proteínas Virais/genética , Replicação Viral
3.
Emerg Microbes Infect ; 8(1): 1370-1382, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31526249

RESUMO

Egypt is a hotspot for avian influenza virus (AIV) due to the endemicity of H5N1 and H9N2 viruses. AIVs were isolated from 329 samples collected in 2016-2018; 48% were H9N2, 37.1% were H5N8, 7.6% were H5N1, and 7.3% were co-infections with 2 of the 3 subtypes. The 32 hemagglutinin (HA) sequences of the H5N1 viruses formed a well-defined lineage within clade 2.2.1.2. The 10 HA sequences of the H5N8 viruses belonged to a subclade within 2.3.4.4. The 11 HA of H9N2 isolates showed high sequence homology with other Egyptian G1-like H9N2 viruses. The prevalence of H5N8 viruses in ducks (2.4%) was higher than in chickens (0.94%). Genetic reassortment was detected in H9N2 viruses. Antigenic analysis showed that H9N2 viruses are homogenous, antigenic drift was detected among H5N1 viruses. AI H5N8 showed higher replication rate followed by H9N2 and H5N1, respectively. H5N8 was more common in Southern Egypt, H9N2 in the Nile Delta, and H5N1 in both areas. Ducks and chickens played a significant role in transmission of H5N1 viruses. The endemicity and co-circulation of H5N1, H5N8, and H9N2 AIV coupled with the lack of a clear control strategy continues to provide avenues for further virus evolution in Egypt.


Assuntos
Coinfecção/veterinária , Monitoramento Epidemiológico/veterinária , Evolução Molecular , Virus da Influenza A Subtipo H5N1/genética , Vírus da Influenza A Subtipo H9N2/genética , Vírus Reordenados , Animais , Galinhas , Coinfecção/epidemiologia , Coinfecção/virologia , Patos , Egito/epidemiologia , Virus da Influenza A Subtipo H5N1/isolamento & purificação , Vírus da Influenza A Subtipo H9N2/isolamento & purificação , Influenza Aviária/epidemiologia , Influenza Aviária/transmissão , Doenças das Aves Domésticas/epidemiologia , Doenças das Aves Domésticas/virologia , Homologia de Sequência , Proteínas Virais/genética
4.
PLoS Comput Biol ; 15(8): e1007189, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31386651

RESUMO

Model-based phylodynamic approaches recently employed generalized linear models (GLMs) to uncover potential predictors of viral spread. Very recently some of these models have allowed both the predictors and their coefficients to be time-dependent. However, these studies mainly focused on predictors that are assumed to be constant through time. Here we inferred the phylodynamics of avian influenza A virus H9N2 isolated in 12 Asian countries and regions under both discrete trait analysis (DTA) and structured coalescent (MASCOT) approaches. Using MASCOT we applied a new time-dependent GLM to uncover the underlying factors behind H9N2 spread. We curated a rich set of time-series predictors including annual international live poultry trade and national poultry production figures. This time-dependent phylodynamic prediction model was compared to commonly employed time-independent alternatives. Additionally the time-dependent MASCOT model allowed for the estimation of viral effective sub-population sizes and their changes through time, and these effective population dynamics within each country were predicted by a GLM. International annual poultry trade is a strongly supported predictor of virus migration rates. There was also strong support for geographic proximity as a predictor of migration rate in all GLMs investigated. In time-dependent MASCOT models, national poultry production was also identified as a predictor of virus genetic diversity through time and this signal was obvious in mainland China. Our application of a recently introduced time-dependent GLM predictors integrated rich time-series data in Bayesian phylodynamic prediction. We demonstrated the contribution of poultry trade and geographic proximity (potentially unheralded wild bird movements) to avian influenza spread in Asia. To gain a better understanding of the drivers of H9N2 spread, we suggest increased surveillance of the H9N2 virus in countries that are currently under-sampled as well as in wild bird populations in the most affected countries.


Assuntos
Vírus da Influenza A Subtipo H9N2 , Influenza Aviária/transmissão , Modelos Biológicos , Migração Animal , Animais , Animais Selvagens/virologia , Ásia/epidemiologia , Teorema de Bayes , Aves/virologia , Comércio , Biologia Computacional , Monitoramento Ambiental , Vírus da Influenza A Subtipo H9N2/classificação , Vírus da Influenza A Subtipo H9N2/genética , Influenza Aviária/epidemiologia , Influenza Aviária/virologia , Modelos Lineares , Filogeografia/estatística & dados numéricos , Dinâmica Populacional , Aves Domésticas/virologia , Análise Espaço-Temporal
5.
Vet Microbiol ; 235: 234-242, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31383307

RESUMO

During 2012-2015, six H5N1 avian influenza viruses were isolated from domestic birds and the environment around Qinghai Lake. Phylogenetic analysis of HA genes revealed that A/chicken/Gansu/XG2/2012 (CK/GS/XG2/12) belonged to clade 2.3.2.1a, while A/environment/Qinghai/1/2013 (EN/QH/1/13), A/chicken/Qinghai/QH1/2015 (CK/QH/QH1/15), A/chicken/Qinghai/QH2/2015 (CK/QH/QH2/15), A/chicken/Qinghai/QH3/2015 (CK/QH/QH3/15), and A/goose/Qinghai/QH6/2015 (GS/QH/QH6/15) belonged to clade 2.3.2.1c. Further analysis of the internal genes of the isolates found that the PB2 gene of EN/QH/1/13 had 99.6% nucleotide identity with that of A/tiger/Jiangsu/1/2013 (H5N1), which clustered into an independent branch with PB2 from multiple subtypes. PB2, PB1, and M genes of CK/QH/QH3/15 were from H9N2, suggesting it was a reassortant of H5N1 and H9N2. Animal studies of three selected viruses revealed that CK/GS/XG2/12, EN/QH/1/13, and CK/QH/QH3/15 were highly lethal to chickens, with intravenous pathogenicity indexes (IVPIs) of 2.97, 2.81, and 3.00, respectively, and systemically replicated in chickens. In a mouse study, three selected H5N1 viruses were highly pathogenic to mice and readily replicated in the lungs, nasal turbinates, kidneys, spleens, and brains. Therefore, isolates in this study appear to be novel reassortants that were circulating at the interface of wild and domestic birds around Qinghai Lake and are lethal to chickens and mice. These data suggest that more extensive surveillance should be implemented, and matched vaccines should be chosen for the domestic birds in this area.


Assuntos
Animais Domésticos/virologia , Virus da Influenza A Subtipo H5N1/genética , Influenza Aviária/epidemiologia , Lagos/virologia , Células A549 , Animais , Galinhas/virologia , China/epidemiologia , Cães , Patos/virologia , Evolução Molecular , Feminino , Humanos , Virus da Influenza A Subtipo H5N1/patogenicidade , Vírus da Influenza A Subtipo H9N2/genética , Vírus da Influenza A Subtipo H9N2/patogenicidade , Influenza Aviária/mortalidade , Influenza Aviária/virologia , Células Madin Darby de Rim Canino , Camundongos , Camundongos Endogâmicos BALB C , Filogenia , Vírus Reordenados/genética , Vírus Reordenados/patogenicidade , Replicação Viral
6.
mSphere ; 4(4)2019 08 21.
Artigo em Inglês | MEDLINE | ID: mdl-31434749

RESUMO

Highly transmissible influenza viruses (IV) must remain stable and infectious under a wide range of environmental conditions following release from the respiratory tract into the air. Understanding how expelled IV persist in the environment is critical to limiting the spread of these viruses. Little is known about how the stability of different IV in expelled aerosols is impacted by exposure to environmental stressors, such as relative humidity (RH). Given that not all IV are equally capable of efficient airborne transmission in people, we anticipated that not all IV would respond uniformly to ambient RH. Therefore, we have examined the stability of human-pathogenic seasonal and avian IV in suspended aerosols and stationary droplets under a range of RH conditions. H3N2 and influenza B virus (IBV) isolates are resistant to RH-dependent decay in aerosols in the presence of human airway surface liquid, but we observed strain-dependent variations in the longevities of H1N1, H3N2, and IBV in droplets. Surprisingly, low-pathogenicity avian influenza H6N1 and H9N2 viruses, which cause sporadic infections in humans but are unable to transmit person to person, demonstrated a trend toward increased sensitivity at midrange to high-range RH. Taken together, our observations suggest that the levels of vulnerability to decay at midrange RH differ with virus type and host origin.IMPORTANCE The rapid spread of influenza viruses (IV) from person to person during seasonal epidemics causes acute respiratory infections that can lead to hospitalizations and life-threatening illness. Atmospheric conditions such as relative humidity (RH) can impact the viability of IV released into the air. To understand how different IV are affected by their environment, we compared the levels of stability of human-pathogenic seasonal and avian IV under a range of RH conditions and found that highly transmissible seasonal IV were less sensitive to decay under midrange RH conditions in droplets. We observed that certain RH conditions can support the persistence of infectious viruses on surfaces and in the air for extended periods of time. Together, our findings will facilitate understanding of factors affecting the persistence and spread of IV in our environment.


Assuntos
Microbiologia Ambiental , Interações Hospedeiro-Patógeno , Viabilidade Microbiana , Orthomyxoviridae/fisiologia , Aerossóis , Animais , Aves , Humanos , Umidade , Vírus da Influenza A Subtipo H1N1/fisiologia , Vírus da Influenza A Subtipo H3N2/fisiologia , Vírus da Influenza A Subtipo H9N2/fisiologia , Influenza Aviária/virologia , Influenza Humana/transmissão , Influenza Humana/virologia , Orthomyxoviridae/classificação , Temperatura Ambiente
7.
Int Immunopharmacol ; 74: 105737, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31288152

RESUMO

Influenza A virus usually leads to economic loss to breeding farms and pose a serious threat to human health. Virus infecting tissues directly and influenza virus-induced excessive production of inflammatory factors play the key role in pathogenesis of the disease, but the mechanism is not well clarified. Here, the role of autophagy was investigated in H9N2 influenza virus-triggered inflammation. The results showed that autophagy was induced by H9N2 virus in A549 cells and in mice. Inhibiting autophagy by an autophagy inhibitor (3-methyladenine, 3-MA) or knockdown of Atg5(autophagy-related gene) by Atg5 siRNA significantly suppressed H9N2 virus replication, H9N2 virus-triggered inflammatory cytokines and chemokines, including IL-1ß, TNF-α, IL-8, and CCL5 in vitro and in vivo, and suppressed H9N2 virus-triggered acute lung injury as indicated as accumulative mortality of mice, inflammatory cellular infiltrate and interstitial edema, thickening of the alveolar walls in mice lung tissues, increased inflammatory cytokines and chemokines, increased W/D ratio in mice. Moreover, autophagy mediated inflammatory responses through Akt-mTOR, NF-κB and MAPKs signaling pathways. Our data showed that autophagy was essential in H9N2 influenza virus-triggered inflammatory responses, and autophagy could be target to treat influenza virus-caused lung inflammation.


Assuntos
Lesão Pulmonar Aguda/imunologia , Proteína 5 Relacionada à Autofagia/metabolismo , Autofagia/genética , Vírus da Influenza A Subtipo H9N2/fisiologia , Influenza Humana/imunologia , Infecções por Orthomyxoviridae/imunologia , Células A549 , Animais , Proteína 5 Relacionada à Autofagia/genética , Citocinas/metabolismo , Humanos , Mediadores da Inflamação/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , RNA Interferente Pequeno/genética , Transdução de Sinais
8.
Transbound Emerg Dis ; 66(6): 2507-2516, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31357255

RESUMO

Vietnamese poultry are host to co-circulating subtypes of avian influenza viruses, including H5N1 and H9N2, which pose a great risk to poultry productivity and to human health. AIVs circulate throughout the poultry trade network in Vietnam, with live bird markets being an integral component to this network. Traders at LBMs exhibit a variety of trading practices, which may influence the transmission of AIVs. We identified trading practices that impacted on AIV prevalence in chickens marketed in northern Vietnamese LBMs. We generated sequencing data for 31 H9N2 and two H5N6 viruses. Viruses isolated in the same LBM or from chickens sourced from the same province were genetically closer than viruses isolated in different LBMs or from chickens sourced in different provinces. The position of a vendor in the trading network impacted on their odds of having AIV-infected chickens. Being a retailer and purchasing chickens from middlemen was associated with increased odds of infection, whereas odds decreased if vendors purchased chickens directly from large farms. Odds of infection were also higher for vendors having a greater volume of ducks unsold per day. These results indicate how the spread of AIVs is influenced by the structure of the live poultry trading network.


Assuntos
Comércio , Influenza Aviária/epidemiologia , Produtos Avícolas , Aves Domésticas , Animais , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Virus da Influenza A Subtipo H5N1/genética , Virus da Influenza A Subtipo H5N1/isolamento & purificação , Vírus da Influenza A Subtipo H9N2/genética , Vírus da Influenza A Subtipo H9N2/isolamento & purificação , Reação em Cadeia da Polimerase , Prevalência , RNA Viral/análise , Fatores de Risco , Vietnã/epidemiologia
9.
Comp Immunol Microbiol Infect Dis ; 65: 165-175, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31300109

RESUMO

Avian influenza vaccines are commonly used in the poultry industry, and some medicinal plants can increase the efficacy of such vaccines. The objective of this study was to evaluate the effect of Immulant® (IMU) (a commercial product based on Echinacea and Nigella sativa) on stress induced by dexamethasone (DEX) in chickens vaccinated (VAC) against the H9N2 avian influenza virus (AIV-H9N2). Seven experimental groups were included: the negative control, VAC, DEX, VAC + DEX, VAC + DEX + IMU, VAC + IMU and IMU groups. The vaccinated chickens (at 10 days of age) were injected daily with DEX for three days pre-vaccination and for three days pre-challenge and orally administered 1% IMU for 6 weeks post-vaccination (PV). The chickens were then challenged intranasally with AIV-H9N2 at 28 days PV. Serum, blood, tracheal and cloacal swabs and tissue samples were collected in the 1st and 4th weeks PV and at different time points post-challenge. The results showed significant changes (P ≤ 0.05) in oxidative stress and antioxidant biomarkers (malondialdehyde, nitric oxide and reduced glutathione), haematological and immunological parameters, final live weights, relative organ weights and histopathological lesions between the VAC+DEX group and the VAC group. Moreover, IMU significantly increased protection rates post-challenge, HI antibody titers and heterophil phagocytic activity and decreased DEX-induced stress and virus shedding titers. In conclusion, oral administration of 1% IMU for six weeks can enhance the immune response after AI-H9N2 vaccination and reduce the pathogenicity of infection in stressed chickens.


Assuntos
Adjuvantes Imunológicos/administração & dosagem , Galinhas/imunologia , Echinacea/química , Vírus da Influenza A Subtipo H9N2/patogenicidade , Vacinas contra Influenza/imunologia , Nigella sativa/química , Adjuvantes Imunológicos/química , Animais , Anticorpos Antivirais/sangue , Dexametasona/administração & dosagem , Imunossupressão , Influenza Aviária/prevenção & controle , Influenza Aviária/virologia , Aves Domésticas , Estresse Fisiológico , Virulência , Eliminação de Partículas Virais
10.
Comp Immunol Microbiol Infect Dis ; 65: 219-225, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31300117

RESUMO

The interaction between a low pathogenic avian influenza virus (A/CK/TUN/145/2012), a H9N2 Tunisian isolate, and a vaccine strain (H120) of avian infectious bronchitis, administered simultaneously or sequentially three days apart to chicks during 20 days, was evaluated using ELISA antibody levels, quantitative reverse transcription-polymerase chain reaction (qRT-PCR) analyses and histopathology examination. First, the in vivo replication interference of avian influenza virus (AIV) and infectious bronchitis virus (IBV) was evaluated using qRT-PCR to detect accurately either AIV or IBV genomes or viral copy numbers during dual infections. Second, we have determined the amount of specific antibodies in sera of chick's infected with AIV alone, IBV alone, mixed AIV + IBV, IBV then AIV or AIV IBV 3 days later using an ELISA test. Finally, histopathological analyses of internal organs from inoculated chicks were realized. Quantitative results of AIV and IBV co-infection showed that interferences between the two viruses yielded decreased viral growth. However, in the case of super-infection, the second virus, either AIV or IBV, induced a decrease in the growth of the first inoculated virus. According to our results, vaccine application was safe and do not interfere with AIV H9N2 infection, and does not enhance such infection. In conclusion, co-infection of chicks with AIV and IBV, simultaneously or sequentially, affected the clinical signs, the virus replication dynamics as well as the internal organ integrity. The results proposed that infection with heterologous virus may result in temporary competition for cell receptors or competent cells for replication, most likely interferon-mediated.


Assuntos
Coinfecção/veterinária , Vírus da Bronquite Infecciosa/crescimento & desenvolvimento , Vírus da Influenza A Subtipo H9N2/crescimento & desenvolvimento , Interferência Viral , Replicação Viral , Animais , Anticorpos Antivirais/sangue , Galinhas/imunologia , Galinhas/virologia , Coinfecção/virologia , Influenza Aviária/virologia , Doenças das Aves Domésticas/virologia , RNA Viral/análise , Vacinas Virais/imunologia
11.
PLoS Pathog ; 15(7): e1007919, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31265471

RESUMO

Avian influenza virus H9N2 has been endemic in birds in the Middle East, in particular in Egypt with multiple cases of human infections since 1998. Despite concerns about the pandemic threat posed by H9N2, little is known about the biological properties of H9N2 in this epicentre of infection. Here, we investigated the evolutionary dynamics of H9N2 in the Middle East and identified phylogeny-associated PB2 mutations that acted cooperatively to increase H9N2 replication/transcription in human cells. The accumulation of PB2 mutations also correlated with an increase in H9N2 virus growth in the upper and lower airways of mice and in virulence. These mutations clustered on a solvent-exposed region in the PB2-627 domain in proximity to potential interfaces with host factors. These PB2 mutations have been found at high prevalence during evolution of H9N2 in the field, indicating that they have provided a selective advantage for viral adaptation to infect poultry. Therefore, continuous prevalence of H9N2 virus in the Middle East has generated a far more fit or optimized replication phenotype, leading to an expanded viral host range, including to mammals, which may pose public health risks beyond the current outbreaks.


Assuntos
Vírus da Influenza A Subtipo H9N2/genética , Vírus da Influenza A Subtipo H9N2/patogenicidade , Influenza Humana/virologia , Mutação , RNA Replicase/genética , Proteínas Virais/genética , Animais , Evolução Molecular , Feminino , Células HEK293 , Especificidade de Hospedeiro/genética , Humanos , Vírus da Influenza A Subtipo H9N2/fisiologia , Influenza Humana/epidemiologia , Mamíferos/virologia , Camundongos , Camundongos Endogâmicos BALB C , Oriente Médio/epidemiologia , Modelos Moleculares , Infecções por Orthomyxoviridae/virologia , Filogenia , RNA Replicase/química , RNA Replicase/metabolismo , Vírus Reordenados/genética , Vírus Reordenados/patogenicidade , Vírus Reordenados/fisiologia , Proteínas Virais/química , Proteínas Virais/metabolismo , Virulência/genética , Replicação Viral/genética , Zoonoses/virologia
12.
J Biol Regul Homeost Agents ; 33(3): 929-933, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31184098

RESUMO

Newcastle disease (ND) and avian influenza (AI) are globally considered as a serious threat to the chicken and other avian species. The paramyxovirus type 1 and orthomyxovirus type A are RNA viruses, which cause ND and AI infection, respectively.


Assuntos
Coinfecção/veterinária , Influenza Aviária/patologia , Doença de Newcastle/patologia , Tropismo Viral , Animais , Galinhas , Coinfecção/virologia , Vírus da Influenza A Subtipo H9N2 , Vírus da Doença de Newcastle
13.
Vet Microbiol ; 234: 77-82, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31213275

RESUMO

Control of currently circulating re-assorted low-pathogenicity avian influenza (LPAI) H9N2 is a major concern for both animal and human health. Thus, an improved LPAI H9N2 vaccination strategy is needed to induce complete immunity in chickens against LPAI H9N2 virus strains. Cytokines play a crucial role in mounting both the type and extent of an immune response generated following infection with a pathogen or after vaccination. To improve the efficacy of inactivated LPAI H9N2 vaccine, prokaryotic expression recombination chicken interferon-α (rchIFN-α) was used as vaccine adjuvant.In this study chIFN-α was used as adjuvant in inactivated AI H9N2 vaccine, modulated the immune response of chickens against the vaccine antigen through enhanced humoral and Th1-biased cell-mediated immunity, compared to chickens that received single AI H9N2 vaccine. To further test the protective efficacy of this improved vaccination regimen, immunized chickens were challenged with a high dose of LPAI H9N2 virus. Combined administration rchIFN-α showed markedly enhanced protection compared to single administration of the vaccine, as determined by mortality, clinical severity, and feed and water intake. This enhancement of protective immunity was further confirmed by reduced rectal shedding and replication of AIV H9N2 in challenged chickens. Our results indicate the value of combined administration of rchIFN-α to generate an effective immunization strategy in chickens against LPAI H9N2.


Assuntos
Imunogenicidade da Vacina , Vacinas contra Influenza/imunologia , Influenza Aviária/prevenção & controle , Interferon-alfa/genética , Adjuvantes Imunológicos , Animais , Anticorpos Antivirais/sangue , Galinhas , Imunidade Celular , Imunidade Humoral , Vírus da Influenza A Subtipo H9N2/imunologia , Vacinas contra Influenza/genética , Influenza Aviária/imunologia , Interferon-alfa/imunologia , Organismos Livres de Patógenos Específicos , Vacinas de Produtos Inativados/genética , Vacinas de Produtos Inativados/imunologia , Vacinas Sintéticas/administração & dosagem , Vacinas Sintéticas/imunologia , Eliminação de Partículas Virais
14.
Sheng Wu Gong Cheng Xue Bao ; 35(6): 1029-1040, 2019 Jun 25.
Artigo em Chinês | MEDLINE | ID: mdl-31231999

RESUMO

To evaluate the optimal administration frequency for interferon-α (IFN-α) and the effect of its combined use with inactive virus on chicken flocks, the prokaryotic expression plasmid pET-22b-ChIFN-α was constructed and transferred into Escherichia coli BL21(DE3) host bacteria to induce the expression of chicken IFN-α and to harvest recombinant proteins inclusion bodies. The expression of recombinant chicken IFN-α was confirmed by SDS-PAGE, and the results demonstrated that the chicken IFN-α (20 kDa) was highly expressed using the prokaryotic expression vector with a concentration of 0.2 mg/mL in the medium. Chicken IFN-α was diluted to 2.5×104 U/fowls and administered to immunized specific-pathogen-free chickens orally in combination with inactivated H9N2 subtype influenza virus. Chicken that received chicken IFN-α were safe after three repeated immunizations (96 h). In addition, chicken IFN-α could induce higher levels of antiviral-related inducible genes in peripheral blood, spleen, and thymus of chicken flocks. The results of a challenge assay revealed that the lowest detoxification rates of chicken IFN-α ranged from three to five days, suggesting a higher capacity to resist H9N2 subtype avian influenza virus. The present study obtained the optimal immune frequency and immunization period for chicken IFN-α to provide theoretical support for the optimal clinical application of IFN-α.


Assuntos
Vírus da Influenza A Subtipo H9N2 , Administração Oral , Animais , Galinhas , Humanos , Interferon-alfa , Replicação Viral
15.
Vet Microbiol ; 233: 1-4, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31176393

RESUMO

Domestic ducks are considered as the interface between wild aquatic birds and terrestrial poultry and play an important role in the transmission and evolution of avian influenza viruses (AIVs). However, the infectivity of H9N2 AIVs in different domestic duck species has not been systematically evaluated. Here we investigated the infectivity of various genotypes of chicken H9N2 AIVs in Pekin duck (Anas Platyrhynchos), Mallard duck (Anas Platyrhynchos) and Muscovy duck (Cairina Moschata) through intranasal inoculation. We found that Pekin ducks and Mallard ducks were generally resistant to chicken H9N2 virus infection, while Muscovy ducks were relatively susceptible to H9N2 AIVs. All the tested viruses were isolated from oropharynx, trachea and lung tissues of Muscovy ducks. Additionally, genotype 57 (G57) H9N2 AIVs, which was predominant in chickens since 2010, showed increased virus replication in this duck species, indicating an improved interspecies transmission ability of recent H9N2 viruses from chickens to ducks. Our results demonstrated the role of Muscovy ducks in the ecology of H9N2 AIVs. More attentions should be paid to this host during viral surveillances. Additionally, inactivated H9N2 vaccine may be unnecessarily used in Pekin and Mallard ducks.


Assuntos
Patos/virologia , Influenza Aviária/transmissão , Doenças das Aves Domésticas/virologia , Replicação Viral , Animais , Galinhas/virologia , Suscetibilidade a Doenças , Vírus da Influenza A Subtipo H9N2/genética , Vírus da Influenza A Subtipo H9N2/fisiologia , Pulmão/virologia , Orofaringe/virologia , Traqueia/virologia
16.
Artigo em Inglês | MEDLINE | ID: mdl-31174705

RESUMO

This study was conducted to perform the comparative molecular characterization of avian influenza virus (AIV) H9N2, pathogenicity and seroprevalence in commercial and backyard poultry flocks. Fifty commercial poultry flocks were investigated between 2012 and 2015. Eighteen flocks (36%) out of 50 were positive HA. Seven (38.9%) out of 18 were positive by chromatographic strip test for AI common antigen. By Real-time RT-PCR, only two flocks were positive H9. The molecular characterization of two different AI-H9N2 viruses, one isolated from a broiler flock (A/chicken/Egypt/Mansoura-18/2013) and the other from a layer flock (A/chicken/Egypt/Mansoura-36/2015) was conducted on HA gene. Moreover, a higher seroprevalence, using the broiler strain as a known antigen, was shown in backyard chicken flocks 15/26 (57.7%) than duck flocks 9/74 (12.2%). Interestingly, the pathogenicity index (PI) of the H9N2 broiler strain in inoculated experimental chickens ranged from 1.2 (oculonasal route) to 1.9 (Intravenous route). The PI indicated a highly pathogenic effect, with high mortality (up to 100%) in the inoculated chickens correlated with the high mortality (80%) in the flock where the virus was isolated. The firstly recorded clinical signs, including cyanosis in the combs and wattles and subcutaneous haemorrhages in the leg shanks and lesions, as well as histopathology and immunohistochemistry, revealed a systemic infection of the high pathogenicity with the H9N2 virus. Conversely, the H9N2 layer strain showed a low pathogenicity. In conclusion, as a first report, the molecular analysis and pathogenicity of the tested strains confirmed the presence of a high pathogenicity AIV-H9N2 with systemic infections.


Assuntos
Vírus da Influenza A Subtipo H9N2/genética , Vírus da Influenza A Subtipo H9N2/patogenicidade , Influenza Aviária/virologia , Doenças das Aves Domésticas/virologia , Aves Domésticas/virologia , Animais , Galinhas/virologia , Cianose/virologia , Patos/virologia , Egito/epidemiologia , Influenza Aviária/mortalidade , Doenças das Aves Domésticas/mortalidade , Reação em Cadeia da Polimerase em Tempo Real , Estudos Soroepidemiológicos , Perus/virologia , Virulência
17.
Br Poult Sci ; 60(5): 493-498, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31116018

RESUMO

1. Infectious bursal disease virus (IBDV) causes immunosuppression in chickens, increasing their susceptibility to other infectious diseases and resulting in vaccination failure. Here, we investigated the immune-depressing effect of IBDV on H9N2 avian influenza viral infection in the broiler chickens. 2. For this purpose, chickens were divided into four groups. In group A, chickens were inoculated with IBDV at 21 days of age and H9N2 avian influenza virus (AIV) 5 days later. Groups B and C only received AIV at 26 days of age and IBDV at 21 days, respectively. The control group (D) were inoculated with normal saline at the same times. Tissue samples from different organs were collected on the days 1, 3, 6, 9, and 12 after H9N2 infection. 3. Macroscopic observation showed IBD lesions in groups A and C, including swollen bursa with the presence of gelatinous exudates, haemorrhages in the thigh muscle, edema, and nephritis. 4. Reverse Transcription-PCR was used to study H9N2 AIV dissemination, and qRT-PCR to determine viral genome copy number in different organs. A considerable titre of AIV was found in the trachea, lungs, cecal tonsils, spleens, and feces of infected chickens. The genome copy number of the virus in the trachea and lungs of group A was significantly higher than that in group B on the first day after inoculation. But in the other days post inoculation, RT-PCR did not detect the AIV genome in group A. Although there might have been some immunosuppression in group A, IBDV could interfere with AIV replication in the chickens of this group. 5. In conclusion, we propose that pre-exposure to IBDV at 3 weeks of age reduces the replication and shedding of H9N2 in broiler chicken.


Assuntos
Infecções por Birnaviridae/veterinária , Galinhas , Vírus da Doença Infecciosa da Bursa/fisiologia , Vírus da Influenza A Subtipo H9N2/fisiologia , Influenza Aviária/virologia , Doenças das Aves Domésticas , Eliminação de Partículas Virais/fisiologia , Animais , Infecções por Birnaviridae/virologia , Coinfecção/veterinária , Coinfecção/virologia , Distribuição Aleatória
18.
Avian Pathol ; 48(5): 470-476, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31142135

RESUMO

Between November 2017 and February 2018, Ghanaian poultry producers reported to animal health authorities a dramatic increase in mortality rate and a relevant drop in egg production in several layer hen farms. Laboratory investigations revealed that the farms had been infected by the H9N2 influenza subtype. Virological and molecular characterization of the viruses identified in Ghana is described here for the first time. Whole genome analysis showed that the viruses belong to the G1-lineage and cluster with viruses identified in North and West Africa. The low pathogenicity of the virus was confirmed by the intravenous pathogenicity index assay. Further investigations revealed co-infection with infectious bronchitis virus of the GI-19 lineage, which very likely explained the severity of the disease observed during the outbreaks. The H9N2 outbreaks in Ghana highlight the importance of performing a differential diagnosis and an in-depth characterization of emerging viruses. In addition, the detection of a potentially zoonotic subtype, such as the H9N2, in a region where highly pathogenic avian influenza H5Nx is currently circulating highlights the urgency of implementing enhanced monitoring strategies and supporting improved investments in regional diagnostic technologies. RESEARCH HIGHLIGHTS Influenza A H9N2 subtype was detected in layer hens in Ghana in 2017-2018 Whole genome characterization of seven H9N2 viruses was performed Phylogenetic trees revealed that the H9N2 viruses belong to the G1 lineage The HA protein possesses the amino acid mutations 226L and 155T Co-infection with infectious bronchitis virus of the GI-19 lineage was identified.


Assuntos
Infecções por Coronavirus/veterinária , Surtos de Doenças/veterinária , Genoma Viral/genética , Vírus da Bronquite Infecciosa/isolamento & purificação , Vírus da Influenza A Subtipo H9N2/isolamento & purificação , Influenza Aviária/epidemiologia , Animais , Galinhas , Coinfecção/veterinária , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/virologia , Gana/epidemiologia , Vírus da Bronquite Infecciosa/genética , Vírus da Bronquite Infecciosa/patogenicidade , Vírus da Influenza A Subtipo H9N2/genética , Vírus da Influenza A Subtipo H9N2/patogenicidade , Influenza Aviária/virologia , Filogenia
19.
Virus Genes ; 55(4): 440-447, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31025287

RESUMO

The untranslated regions within viral segments are the essential promoter elements required for the initiation of viral replication and transcription. The end of the UTR sequence and part of the ORF sequence constitute the packaging signal for progeny viruses. To explore the influence of single-point and multi-site joint mutations in the UTR of the NA gene on the viral expression, we select clones with upregulated expression of the reporter gene and analyze their sequence characteristics. Bioinformatics methods were used to analyze polymorphisms in the untranslated region (UTR) of the neuraminidase gene of the H9N2 influenza A virus. Using the RNA polymerase I reporting system with enhanced green fluorescence protein (EGFP) gene as the reporter gene, libraries containing random mutations at sites within the N2 UTR were constructed using random mutagenesis. The mutants were selected from the randomized mutagenesis libraries for the N2-UTR. The N2-UTR-RNA polymerase I fluorescence reporter system was identified by sequencing and transfected into infected MDCK cells. The expression of the reporter EGFP was observed using fluorescence microscopy, and the relative fluorescence intensity was measured using a multifunctional microplate reader to analyze the expression of the reporter gene (EGFP) qualitatively and quantitatively. Herein, an RNA polymerase reporter system was constructed to rescue the mutated viruses and measure their tissue culture infective dose (TCID50). The results showed that the U13 → C13 mutation in the 3'end of the NA gene promoted the expression of viral RNA and protein, and mutation of other sites within the UTR could differentially regulate viral genomic transcription and translation. These data showed that the U13 → C13 mutation within the variable region of the 3'UTR of the NA gene in the H9N2 influenza virus promotes viral genomic expression and infection.


Assuntos
Vírus da Influenza A Subtipo H9N2/genética , Neuraminidase/genética , Proteínas Virais/genética , Replicação Viral/genética , Regiões 3' não Traduzidas , Animais , Cães , Regulação Viral da Expressão Gênica , Genes Reporter , Proteínas de Fluorescência Verde/genética , Vírus da Influenza A Subtipo H9N2/fisiologia , Células Madin Darby de Rim Canino , Mutagênese , RNA Viral
20.
Vet Microbiol ; 232: 151-155, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31030840

RESUMO

Influenza virus-like sequences of H17N10 and H18N11 were identified in bats, despite there has been no live virus isolated. The genetic analysis indicated that they have distinct but relatively close evolutionary relationships to known influenza A viruses. However, the infectivity and adaptation of bat influenza viruses in avian species remain unclear. In this study, two modified bat influenza viruses cH9cN2/H17 and cH9cN2/H18 containing HA and NA coding regions replaced with those of H9N2 influenza A virus were generated in the background of the H17N10 or H18N11 viruses. These two modified viruses replicated less efficiently than wild type H9N2 virus in cultured chicken cells. The mini-genome assay showed that viral ribonucleoproteins (vRNPs) of H9N2 has significantly higher polymerase activity than that of bat influenza viruses in avian cells. In chicken study, compared with H9N2 virus, which replicated and transmitted efficiently in chickens, the cH9cN2/H17 and cH9cN2/H18 viruses only replicated in chicken tracheas with lower titers. Pathological examination showed that the H9N2 caused severer lesions in lung and trachea than the modified bat influenza viruses. Notably, the cH9cN2/H18 transmitted among chickens, but not cH9cN2/H17, and chicken IFN-ß antagonism results showed that H18N11 NS1 protein inhibited chicken IFN-ß response more efficiently than H17N10 NS1 protein in avian cells. Taken together, our data indicated that the internal genes of bat influenza viruses adapted poorly to chickens, while the internal genes of H18N11 seemed to adapt to chickens better than H17N10.


Assuntos
Adaptação Fisiológica/genética , Quimera/genética , Quirópteros/virologia , Vírus da Influenza A Subtipo H9N2/genética , Orthomyxoviridae/genética , Animais , Sequência de Bases , Galinhas/virologia , Hemaglutininas Virais/genética , Influenza Aviária/virologia , Neuraminidase/genética , Doenças das Aves Domésticas/virologia , Ribonucleoproteínas
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