Project description:Avian influenza virus subtype H9N2 is endemic in commercial poultry in Tunisia. This subtype affects poultry and wild birds in Tunisia and poses a potential zoonotic risk. Tunisian H9N2 strains carry, in their hemagglutinins, the human-like marker 226 L that is most influential in avian-to-human viral transmission. For a better understanding of how ecological aspects of the H9N2 virus and its circulation in poultry, migratory birds and environment shapes the spread of the dissemination of H9N2 in Tunisia, herein, we investigate the epidemiological, evolutionary and zoonotic potential of seven H9N2 poultry isolates and sequence their whole genome. Phylogeographic and phylodymanic analysis were used to examine viral spread within and among wild birds, poultry and environment at geographical scales. Genetic evolution results showed that the eight gene sequences of Tunisian H9N2 AIV were characterized by molecular markers involved with virulence and mammalian infections. The geographical distribution of avian influenza virus appears as a network interconnecting countries in Europe, Asia, North Africa and West Africa. The spatiotemporal dynamics analysis showed that the H9N2 virus was transmitted from Tunisia to neighboring countries notably Libya and Algeria. Interestingly, this study also revealed, for the first time, that there was a virus transmission between Tunisia and Morocco. Bayesian analysis showed exchanges between H9N2 strains of Tunisia and those of the Middle Eastern countries, analysis of host traits showed that duck, wild birds and environment were ancestry related to chicken. The subtypes phylodynamic showed that PB1 segment was under multiple inter-subtype reassortment events with H10N7, H12N5, H5N2 and H6N1 and that PB2 was also a subject of inter-subtype reassortment with H10N4.
Project description:Almost the full range of 16 haemagglutinin (HA) and nine neuraminidase subtypes of avian influenza viruses (AIVs) has been detected either in waterfowl, land-based poultry or in the environment in Bangladesh. AIV infections in Bangladesh affected a wide range of host species of terrestrial poultry. The highly pathogenic avian influenza (AI) H5N1 and low pathogenic AI H9N2 were found to co-circulate and be well entrenched in the poultry population, which has caused serious damage to the poultry industry since 2007. By reviewing the available scientific literature, the overall situation of AIVs in Bangladesh is discussed. All Bangladeshi (BD) H5N1 and H9N2 AIV sequences available at GenBank were downloaded along with other representative sequences to analyse the genetic diversity among the circulating AIVs in Bangladesh and to compare with the global situation. Three different H5N1 clades, 2.2.2, 2.3.2.1 and 2.3.4.2, have been detected in Bangladesh. Only 2.3.2.1a is still present. The BD LP H9N2 viruses mostly belonged to the H9 G1 lineage but segregated into many branches, and some of these shared internal genes with HP viruses of subtypes H7N3 and H5N1. However, these reassortment events might have taken place before introduction to Bangladesh. Currently, H9N2 viruses continue to evolve their HA cleavage, receptor binding and glycosylation sites. Multiple mutations in the HA gene associated with adaptation to mammalian hosts were also observed. Strict biosecurity at farms and gradual phasing out of live-bird markets could be the key measures to better control AIVs, whereas stamping out is not a practicable option in Bangladesh. Vaccination also could be an additional tool, which however, requires careful planning. Continuous monitoring of AIVs through systematic surveillance and genetic characterisation of the viruses remains a hallmark of AI control.
Project description:Low pathogenic H9N2 avian influenza (LPAI H9N2) is considered one of the most important diseases found in poultry (broiler, laying hens, breeding chickens, and turkeys). This infection causes considerable economic losses. The objective of this work was to monitor and assess the presence of avian influenza virus (AIV) H9N2 in eight different regions of Morocco using real-time RT-PCR, and to assess the phylogenetic and molecular evolution of the H9N2 viruses between 2016 and 2019. Field samples were collected from 108 farms suspected of being infected with LPAI H9N2 virus. Samples were analyzed using H9N2-specific real-time RT-PCR. Highly positive samples were subjected to virus isolation and seven isolates were fully sequenced. Low pathogenic H9N2 avian influenza virus was introduced in Morocco in 2016. We show that in 2018-2019, the virus was still present irrespective of vaccination status. Phylogenetic and molecular analyses showed mutations related to virulence, although our viruses were related to 2016 Moroccan viruses and grouped in the G1 lineage. Specific amino acid substitutions were identified in Moroccan H9N2 viruses that are believed to lead to increased resistance to antiviral drugs.
Project description:Avian influenza virus subtype H9N2 has been circulating in the Middle East since the 1990s. For uncertain reasons, H9N2 was not detected in Egyptian farms until the end of 2010. Circulation of H9N2 viruses in Egyptian poultry in the presence of the enzootic highly pathogenic H5N1 subtype adds a huge risk factor to the Egyptian poultry industry. In this study, 22 H9N2 viruses collected from 2011 to 2013 in Egypt were isolated and sequenced. The genomic signatures and protein sequences of these isolates were analyzed. Multiple mammalian-host-associated mutations were detected that favor transmission from avian to mammalian hosts. Other mutations related to virulence were also identified. Phylogenetic data showed that Egyptian H9N2 viruses were closely related to viruses isolated from neighboring Middle Eastern countries, and their HA gene resembled those of viruses of the G1-like lineage. No reassortment was detected with H5N1 subtypes. Serological analysis of H9N2 virus revealed antigenic conservation among Egyptian isolates. Accordingly, continuous surveillance that results in genetic and antigenic characterization of H9N2 in Egypt is warranted.
Project description:H9N2 avian influenza virus is a zoonotic agent with a broad host range that can contribute genetic information to H5 or H7N9 subtype viruses, which are significant threats to both humans and birds. Thus, there is a great need for a vaccine to control H9N2 avian influenza. Three mutant viruses of an H9N2 virus A/chicken/Taixing/10/2010 (rTX-NS1-73, rTX-NS1-100, and rTX-NS1-128) were constructed with different NS1 gene truncations and confirmed by western blot analysis. The genetic stability, pathogenicity, transmissibility, and host immune responses toward these mutants were evaluated. The mutant virus rTX-NS1-128 exhibited the most attenuated phenotype and lost transmissibility. The expression levels of interleukin 12 in the nasal and tracheal tissues from chickens immunized with rTX-NS1-128 were significantly upregulated on day 3 post-immunization and the IgA and IgG antibody levels were significantly increased on days 7, 14, and 21 post-immunization when compared to chickens that received an inactivated vaccine. rTX-NS1-128 also protected chickens from challenge by homologous and heterologous H9N2 avian influenza viruses. The results indicate that rTX-NS1-128 can be used as a potential live-attenuated vaccine against H9N2 avian influenza.
Project description:H9N2 subtype avian influenza viruses (AIVs) have been isolated from various species of wild birds and domestic poultry in the world, and occasionally transmitted to humans. Although H9N2 AIVs are seldom isolated from ostriches, seven such strains were isolated from sick ostriches in China between 2013 and 2014. Sequence analysis showed several amino acid changes relating to viral adaptation in mammals were identified. The phylogenetic analyses indicated that these isolates were quadruple reassortant viruses, which are different from the early ostrich isolates from South Africa or Israel. Most of the ostrich virus carried a human-type receptor-binding property. The chicken experiments showed the ostrich strains displayed low pathogenicity, while they could cause mild to severe symptoms in chicken. Theses strains could efficiently transmit among chickens, and one strain showed higher transmissibility. The virus could not kill mice, and merely replicated in the lung of mice. The ostrich strains could not efficiently transmit between guinea pigs in the direct contact model. These results suggested we should pay attention to the interface between ostrich and other domestic fowl, and keep an eye on this population when monitoring of influenza virus.
Project description:Human infection with avian influenza A(H9N2) virus was identified in Bangladesh in 2011. Surveillance for influenza viruses in apparently healthy poultry in live-bird markets in Bangladesh during 2008-2011 showed that subtype H9N2 viruses are isolated year-round, whereas highly pathogenic subtype H5N1 viruses are co-isolated with subtype H9N2 primarily during the winter months. Phylogenetic analysis of the subtype H9N2 viruses showed that they are reassortants possessing 3 gene segments related to subtype H7N3; the remaining gene segments were from the subtype H9N2 G1 clade. We detected no reassortment with subtype H5N1 viruses. Serologic analyses of subtype H9N2 viruses from chickens revealed antigenic conservation, whereas analyses of viruses from quail showed antigenic drift. Molecular analysis showed that multiple mammalian-specific mutations have become fixed in the subtype H9N2 viruses, including changes in the hemagglutinin, matrix, and polymerase proteins. Our results indicate that these viruses could mutate to be transmissible from birds to mammals, including humans.
Project description:Human outbreaks with avian influenza have been, so far, constrained by poor viral adaptation to non-avian hosts. This could be overcome via co-infection, whereby two strains share genetic material, allowing new hybrid strains to emerge. Identifying areas where co-infection is most likely can help target spaces for increased surveillance. Ecological niche modeling using remotely-sensed data can be used for this purpose. H5N1 and H9N2 influenza subtypes are endemic in Egyptian poultry. From 2006 to 2015, over 20,000 poultry and wild birds were tested at farms and live bird markets. Using ecological niche modeling we identified environmental, behavioral, and population characteristics of H5N1 and H9N2 niches within Egypt. Niches differed markedly by subtype. The subtype niches were combined to model co-infection potential with known occurrences used for validation. The distance to live bird markets was a strong predictor of co-infection. Using only single-subtype influenza outbreaks and publicly available ecological data, we identified areas of co-infection potential with high accuracy (area under the receiver operating characteristic (ROC) curve (AUC) 0.991).
Project description:Avian influenza A(H9N2) viruses are an increasing threat to global poultry production and, through zoonotic infection, to human health where they are considered viruses with pandemic potential. Vaccination of poultry is a key element of disease control in endemic countries, but vaccine effectiveness is persistently challenged by the emergence of antigenic variants. Here we employed a combination of techniques to investigate the genetic basis of H9N2 antigenic variability and evaluate the role of different molecular mechanisms of immune escape. We systematically tested the influence of published H9N2 monoclonal antibody escape mutants on chicken antisera binding, determining that many have no significant effect. Substitutions introducing additional glycosylation sites were a notable exception, though these are relatively rare among circulating viruses. To identify substitutions responsible for antigenic variation in circulating viruses, we performed an integrated meta-analysis of all published H9 haemagglutinin sequences and antigenic data. We validated this statistical analysis experimentally and allocated several new residues to H9N2 antigenic sites, providing molecular markers that will help explain vaccine breakdown in the field and inform vaccine selection decisions. We find evidence for the importance of alternative mechanisms of immune escape, beyond simple modulation of epitope structure, with substitutions increasing glycosylation or receptor-binding avidity, exhibiting the largest impacts on chicken antisera binding. Of these, meta-analysis indicates avidity regulation to be more relevant to the evolution of circulating viruses, suggesting that a specific focus on avidity regulation is required to fully understand the molecular basis of immune escape by influenza, and potentially other viruses.
Project description:H9N2 avian influenza viruses are continuously monitored by the World Health Organization because they are endemic; they continually reassort with H5N1, H7N9 and H10N8 viruses; and they periodically cause human infections. We characterized H9N2 influenza viruses carrying internal genes from highly pathogenic H7N3 viruses, which were isolated from chickens or quail from live-bird markets in Bangladesh between 2010 and 2013. All of the H9N2 viruses used in this study carried mammalian host-specific mutations. We studied their replication kinetics in normal human bronchoepithelial cells and swine tracheal and lung explants, which exhibit many features of the mammalian airway epithelium and serve as a mammalian host model. All H9N2 viruses replicated to moderate-to-high titers in the normal human bronchoepithelial cells and swine lung explants, but replication was limited in the swine tracheal explants. In Balb/c mice, the H9N2 viruses were nonlethal, replicated to moderately high titers and the infection was confined to the lungs. In the ferret model of human influenza infection and transmission, H9N2 viruses possessing the Q226L substitution in hemagglutinin replicated well without clinical signs and spread via direct contact but not by aerosol. None of the H9N2 viruses tested were resistant to the neuraminidase inhibitors. Our study shows that the Bangladeshi H9N2 viruses have the potential to infect humans and highlights the importance of monitoring and characterizing this influenza subtype to better understand the potential risk these viruses pose to humans.