RESUMO
In 2012, an avian influenza A H7N3 (A/Mexico/InDRE7218/2012; Mx/7218) virus was responsible for two confirmed cases of human infection and led to the death or culling of more than 22 million chickens in Jalisco, Mexico. Interestingly, this virus acquired an 8-amino acid (aa)-insertion (..PENPK-DRKSRHRR-TR/GLF) near the hemagglutinin (HA) cleavage site by nonhomologous recombination with host rRNA. It remains unclear which specific residues at the cleavage site contribute to the virulence of H7N3 viruses in mammals. Using loss-of-function approaches, we generated a series of cleavage site mutant viruses by reverse genetics and characterized the viruses in vitro and in vivo. We found that the 8-aa insertion and the arginine at position P4 of the Mx/7218 HA cleavage site are essential for intracellular HA cleavage in 293T cells, but have no effect on the pH of membrane fusion. However, we identified a role for the histidine residue at P5 position in viral fusion pH. In mice, the 8-aa insertion is required for Mx/7218 virus virulence; however, the basic residues upstream of the P4 position are dispensable for virulence. Overall, our study provides the first line of evidence that the insertion in the Mx/7218 virus HA cleavage site confers its intracellular cleavability, and consequently contributes to enhanced virulence in mice.
Assuntos
Glicoproteínas de Hemaglutininação de Vírus da Influenza/genética , Glicoproteínas de Hemaglutininação de Vírus da Influenza/metabolismo , Vírus da Influenza A Subtipo H7N3/genética , Vírus da Influenza A Subtipo H7N3/fisiologia , Mutagênese Insercional , Infecções por Orthomyxoviridae/virologia , Proteólise , Substituição de Aminoácidos , Animais , Linhagem Celular , Galinhas , Análise Mutacional de DNA , Modelos Animais de Doenças , Feminino , Humanos , Vírus da Influenza A Subtipo H7N3/isolamento & purificação , Influenza Aviária/virologia , México , Camundongos Endogâmicos BALB C , Infecções por Orthomyxoviridae/patologia , Genética Reversa , VirulênciaRESUMO
In June of 2012, an H7N3 highly pathogenic avian influenza (HPAI) virus was identified as the cause of a severe disease outbreak in commercial laying chicken farms in Mexico. The purpose of this study was to characterize the Mexican 2012 H7N3 HPAI virus (A/chicken/Jalisco/CPA1/2012) and determine the protection against the virus conferred by different H7 inactivated vaccines in chickens. Both adult and young chickens intranasally inoculated with the virus became infected and died at between 2 and 4 days postinoculation (p.i.). High virus titers and viral replication in many tissues were demonstrated at 2 days p.i. in infected birds. The virus from Jalisco, Mexico, had high sequence similarity of greater than 97% to the sequences of wild bird viruses from North America in all eight gene segments. The hemagglutinin gene of the virus contained a 24-nucleotide insert at the hemagglutinin cleavage site which had 100% sequence identity to chicken 28S rRNA, suggesting that the insert was the result of nonhomologous recombination with the host genome. For vaccine protection studies, both U.S. H7 low-pathogenic avian influenza (LPAI) viruses and a 2006 Mexican H7 LPAI virus were tested as antigens in experimental oil emulsion vaccines and injected into chickens 3 weeks prior to challenge. All H7 vaccines tested provided ≥90% protection against clinical disease after challenge and decreased the number of birds shedding virus and the titers of virus shed. This study demonstrates the pathological consequences of the infection of chickens with the 2012 Mexican lineage H7N3 HPAI virus and provides support for effective programs of vaccination against this virus in poultry.
Assuntos
Surtos de Doenças , Vírus da Influenza A Subtipo H7N3/patogenicidade , Vacinas contra Influenza/imunologia , Influenza Aviária/epidemiologia , Influenza Aviária/virologia , Estruturas Animais/virologia , Animais , Animais Domésticos , Aves , Galinhas , Análise por Conglomerados , Glicoproteínas de Hemaglutininação de Vírus da Influenza/genética , Vírus da Influenza A Subtipo H7N3/imunologia , Vírus da Influenza A Subtipo H7N3/isolamento & purificação , Vacinas contra Influenza/administração & dosagem , Vacinas contra Influenza/isolamento & purificação , Influenza Aviária/patologia , Influenza Aviária/prevenção & controle , México/epidemiologia , Filogenia , RNA Ribossômico 28S/genética , RNA Viral/genética , Recombinação Genética , Homologia de Sequência , Análise de Sobrevida , Vacinas de Produtos Inativados/administração & dosagem , Vacinas de Produtos Inativados/imunologia , Vacinas de Produtos Inativados/isolamento & purificação , Carga Viral , Eliminação de Partículas ViraisRESUMO
BACKGROUND: A characteristic difference between highly and non-highly pathogenic avian influenza strains is the presence of an extended, often multibasic, cleavage motif insertion in the hemagglutinin protein. Such motif is found in H7N3 strains from chicken farm outbreaks in 2012 in Mexico. METHODS: Through phylogenetic, sequence and structural analysis, we try to shed light on the role, prevalence, likelihood of appearance and origin of the inserted cleavage motifs in these H7N3 avian influenza strains. RESULTS: The H7N3 avian influenza strain which caused outbreaks in chicken farms in June/July 2012 in Mexico has a new extended cleavage site which is the likely reason for its high pathogenicity in these birds. This cleavage site appears to have been naturally acquired and was not present in the closest low pathogenic precursors. Structural modeling shows that insertion of a productive cleavage site is quite flexible to accept insertions of different length and with sequences from different possible origins. Different from recent cleavage site insertions, the origin of the insert here is not from the viral genome but from host 28S ribosomal RNA (rRNA) instead. This is a novelty for a natural acquisition as a similar insertion has so far only been observed in a laboratory strain before. Given the abundance of viral and host RNA in infected cells, the acquisition of a pathogenicity-enhancing extended cleavage site through a similar route by other low-pathogenic avian strains in future does not seem unlikely. Important for surveillance of these H7N3 strains, the structural sites known to enhance mammalian airborne transmission are dominated by the characteristic avian residues and the risk of human to human transmission should currently be low but should be monitored for future changes accordingly. CONCLUSIONS: This highly pathogenic H7N3 avian influenza strain acquired a novel extended cleavage site which likely originated from recombination with 28S rRNA from the avian host. Notably, this new virus can infect humans but currently lacks critical host receptor adaptations that would facilitate human to human transmission.
Assuntos
Glicoproteínas de Hemaglutininação de Vírus da Influenza/genética , Vírus da Influenza A Subtipo H7N3/genética , Vírus da Influenza A Subtipo H7N3/isolamento & purificação , Influenza Aviária/virologia , RNA Ribossômico 28S/genética , RNA Viral/genética , Recombinação Genética , Animais , Galinhas , México , Filogenia , Análise de Sequência de DNARESUMO
H7 subtype influenza A viruses, responsible for numerous outbreaks in land-based poultry in Europe and the Americas, have caused over 100 cases of confirmed or presumed human infection over the last decade. The emergence of a highly pathogenic avian influenza H7N3 virus in poultry throughout the state of Jalisco, Mexico, resulting in two cases of human infection, prompted us to examine the virulence of this virus (A/Mexico/InDRE7218/2012 [MX/7218]) and related avian H7 subtype viruses in mouse and ferret models. Several high- and low-pathogenicity H7N3 and H7N9 viruses replicated efficiently in the respiratory tract of mice without prior adaptation following intranasal inoculation, but only MX/7218 virus caused lethal disease in this species. H7N3 and H7N9 viruses were also detected in the mouse eye following ocular inoculation. Virus from both H7N3 and H7N9 subtypes replicated efficiently in the upper and lower respiratory tracts of ferrets; however, only MX/7218 virus infection caused clinical signs and symptoms and was capable of transmission to naive ferrets in a direct-contact model. Similar to other highly pathogenic H7 viruses, MX/7218 replicated to high titers in human bronchial epithelial cells, yet it downregulated numerous genes related to NF-κB-mediated signaling transduction. These findings indicate that the recently isolated North American lineage H7 subtype virus associated with human conjunctivitis is capable of causing severe disease in mice and spreading to naive-contact ferrets, while concurrently retaining the ability to replicate within ocular tissue and allowing the eye to serve as a portal of entry.
Assuntos
Conjuntivite/virologia , Vírus da Influenza A Subtipo H7N3/patogenicidade , Infecções por Orthomyxoviridae/virologia , Tropismo Viral , Animais , Modelos Animais de Doenças , Células Epiteliais/virologia , Feminino , Furões , Humanos , Vírus da Influenza A Subtipo H7N3/isolamento & purificação , Masculino , México , Camundongos , Camundongos Endogâmicos BALB C , Infecções por Orthomyxoviridae/transmissão , Sistema Respiratório/virologiaRESUMO
During June-August 2012, Mexico's National Service for Health, Safety, and Food Quality reported outbreaks of highly pathogenic avian influenza (HPAI) A (H7N3) virus in poultry on farms throughout the state of Jalisco. This report describes two cases of conjunctivitis without fever or respiratory symptoms caused by HPAI A (H7N3) virus infection in humans associated with exposure to infected poultry.
Assuntos
Conjuntivite Viral , Vírus da Influenza A Subtipo H7N3/isolamento & purificação , Influenza Humana , Exposição Ocupacional , Adulto , Animais , Conjuntivite Viral/diagnóstico , Conjuntivite Viral/tratamento farmacológico , Conjuntivite Viral/patologia , Conjuntivite Viral/transmissão , Feminino , Humanos , Influenza Aviária/transmissão , Influenza Humana/diagnóstico , Influenza Humana/tratamento farmacológico , Influenza Humana/patologia , Influenza Humana/transmissão , Masculino , México , Pessoa de Meia-Idade , Aves Domésticas , ZoonosesRESUMO
Two different wild duck species common in Chile and neighboring countries, Chiloe wigeon (Anas sibilatrix) and cinnamon teal (Anas cyanoptera), were intranasally inoculated with 10(6) mean embryo infective dose (EID50) of the H7N3 low pathogenicity (LP) avian influenza virus (AIV) (A/chicken/Chile/176822/02) or high pathogenicity (HP) AIV (A/chicken/Chile/ 184240-1/02), in order to study the infectivity and pathobiology of these viruses. None of the virus-inoculated ducks had clinical signs or died, but most seroconverted by 14 days postinoculation (DPI), indicating a productive virus infection. Both LPAIV and HPAIV were isolated from oral swabs from two of six Chiloe wigeons and from oral and/or cloacal swabs from all five of the cinnamon teal at 2 DPI. Both LPAIV and HPAIV were efficiently transmitted to cinnamon teal contacts but not to Chiloe wigeon contacts. This study demonstrates that the cinnamon teal and Chiloe wigeons were susceptible to infection with both Chilean H7N3 LPAIV and HPAIV, but only the cinnamon teal showed contact transmission of the virus between birds, suggesting that the cinnamon teal has the potential to be a reservoir for these viruses, especially the LPAIV, as was demonstrated in 2001 with isolation of a genetically related H7N3 LPAIV strain in a cinnamon teal in Bolivia. However, the definitive source of the H7N3 Chilean LPAIV still remains unknown.
Assuntos
Patos , Vírus da Influenza A Subtipo H7N3/genética , Vírus da Influenza A Subtipo H7N3/patogenicidade , Influenza Aviária/virologia , Animais , Anticorpos Antivirais/análise , Cloaca/virologia , Suscetibilidade a Doenças/veterinária , Ensaio de Imunoadsorção Enzimática , Vírus da Influenza A Subtipo H7N3/classificação , Vírus da Influenza A Subtipo H7N3/isolamento & purificação , Influenza Aviária/imunologia , Influenza Aviária/patologia , Influenza Aviária/transmissão , Doenças Pulmonares Intersticiais/fisiopatologia , Doenças Pulmonares Intersticiais/veterinária , Orofaringe/virologia , Sistema Respiratório/virologia , Especificidade da EspécieRESUMO
Wild birds serve as natural reservoirs and sometimes harbor low-pathogenic avian influenza viruses. However, mutation of the virus can result in highly pathogenic strains, often more common among H5 and H7 genotypes. We report the isolation of a low-pathogenic H7N3 avian influenza in a Peruvian wetland.