Control Of Avian Influenza Virus Research Articles

Effect of Nonpharmaceutical Interventions in Preventing COVID-19 on the Circulation of Avian Influenza Virus in Wuhan, Hubei Province, China

Background. In late 2019, several medical institutions in Wuhan, Hubei Province, China, reported cases of unexplained pneumonia. A novel coronavirus was isolated from human airway epithelial cells causing coronavirus disease-2019 (COVID-19). In recent years, many nonpharmaceutical interventions (NPIs) have been implemented to stop COVID-19 epidemic. This study aimed to explore the effect of NPIs on the circulation of avian influenza virus (AIV) in Wuhan. Materials and Methods. External environmental samples were collected and subjected to viral RNA extraction. Real-time quantitative polymerase chain reaction was used to detect the H5, H7, and H9 subtypes of AIV. Statistical analyses were performed using the chi-square test and binary logistic regression in SPSS 20.0 software. Results. A total of 2,451 external environmental samples were collected from seven districts from 2018 to 2022 in Wuhan, comprising 1,041 samples collected before COVID-19 and 1,410 samples after COVID-19. After COVID-19, the positive rate of AIV decreased significantly with the implementation of NPIs. The dominant subtype was the H9 subtype, followed by the H5 subtype. The positive rates of AIV in live poultry markets and poultry free-range sites were reduced significantly through the implementation of NPIs. Among the different sample types, higher positive rates of AIV were found in chopping boards, sewage, and cages. The positive rate of AIV was higher in trafficked source samples than that in autotrophic source samples. Conclusions. This study identified the characteristics of AIV in terms of different districts, surveillance sites, sample types, and bird sources in Wuhan. This study conducted a multifactorial analysis of the factors affecting AIV infection and provided a theoretical basis and guidance for the future prevention and control of AIV in Wuhan.

Epidemiology and molecular characterization of avian influenza A viruses H5N1 and H3N8 subtypes in poultry farms and live bird markets in Bangladesh

Avian influenza virus (AIV) remains a global threat, with waterfowl serving as the primary reservoir from which viruses spread to other hosts. Highly pathogenic avian influenza (HPAI) H5 viruses continue to be a devastating threat to the poultry industry and an incipient threat to humans. A cross-sectional study was conducted in seven districts of Bangladesh to estimate the prevalence and subtypes (H3, H5, and H9) of AIV in poultry and identify underlying risk factors and phylogenetic analysis of AIVs subtypes H5N1 and H3N8. Cloacal and oropharyngeal swab samples were collected from 500 birds in live bird markets (LBMs) and poultry farms. Each bird was sampled by cloacal and oropharyngeal swabbing, and swabs were pooled for further analysis. Pooled samples were analyzed for the influenza A virus (IAV) matrix (M) gene, followed by H5 and H9 molecular subtyping using real-time reverse transcription-polymerase chain reaction (rRT-PCR). Non-H5 and Non-H9 influenza A virus positive samples were sequenced to identify possible subtypes. Selected H5 positive samples were subjected to hemagglutinin (HA) and neuraminidase (NA) gene sequencing. Multivariable logistic regression was used for risk factor analysis. We found that IAV M gene prevalence was 40.20% (95% CI 35.98–44.57), with 52.38%, 46.96%, and 31.11% detected in chicken, waterfowl, and turkey, respectively. Prevalence of H5, H3, and H9 reached 22%, 3.4%, and 6.9%, respectively. Waterfowl had a higher risk of having AIV (AOR: 4.75), and H5 (AOR: 5.71) compared to chicken; more virus was detected in the winter season than in the summer season (AOR: 4.93); dead birds had a higher risk of AIVs and H5 detection than healthy birds, and the odds of H5 detection increased in LBM. All six H5N1 viruses sequenced were clade 2.3.2.1a-R1 viruses circulating since 2015 in poultry and wild birds in Bangladesh. The 12 H3N8 viruses in our study formed two genetic groups that had more similarity to influenza viruses from wild birds in Mongolia and China than to previous H3N8 viruses from Bangladesh. The findings of this study may be used to modify guidelines on AIV control and prevention to account for the identified risk factors that impact their spread.

CRISPR, Quorum Sensing, and Arbitrium as strategic control measures for avian influenza virus. Proceeding of The First International Avian Influenza Summit, University of Arkansas- October 16-17, 2023”

Avian Influenza Virus (AIV) poses a significant threat to both avian populations and human health due to its potential for zoonotic transmission. The emergence of highly pathogenic strains necessitates the development of innovative control strategies. This abstract presents a comprehensive overview of the potential of CRISPR-based technologies, Quorum Sensing (QS) systems, and the application of the synthetic inducer molecule Arbitrium as strategic control measures for mitigating AIV outbreaks. CRISPR-based technologies have revolutionized genetic engineering and hold immense promise in combating AIV. By leveraging the adaptive immune system of prokaryotes, CRISPR offers precise and targeted gene editing capabilities. This enables the creation of AIV-resistant poultry strains by introducing specific antiviral sequences, thereby reducing viral transmission within avian populations. Quorum Sensing, a cell-to-cell communication mechanism utilized by bacteria, can be harnessed to modulate AIV infectivity. By engineering QS systems to detect AIV-specific molecular signatures, it becomes possible to trigger population-wide responses in avian hosts. This includes activating immune responses and initiating antiviral signaling cascades, effectively conferring enhanced resistance to AIV infection. The incorporation of Arbitrium, a synthetic signaling molecule, represents a novel approach to controlling AIV. By engineering avian hosts to recognize Arbitrium, it is possible to orchestrate precise temporal and spatial responses to viral challenges. This programmable control system empowers the host organism to mount robust antiviral defenses upon exposure to AIV, providing an additional layer of protection. Furthermore, a synergistic approach integrating CRISPR, QS, and Arbitrium technologies holds the potential to create a comprehensive and adaptable defense system against AIV. This multi-pronged strategy addresses viral challenges at multiple levels, from genetic modification to population-wide immune responses, thereby bolstering the overall resilience of avian populations to AIV outbreaks. In conclusion, integrating CRISPR-based technologies, Quorum Sensing, and Arbitrium signaling represents a promising avenue for developing strategic control measures against Avian Influenza Virus. The synergistic application of these technologies offers precise and targeted genetic modifications and enables dynamic, population-level responses to AIV challenges. This innovative approach holds significant potential for revolutionizing AIV control strategies and safeguarding both avian and human health.

Surveillance of avian influenza viruses in live bird markets of Shandong province from 2013 to 2019.

Avian influenza viruses (AIVs) seriously affect the poultry industry and pose a great threat to humans. Timely surveillance of AIVs is the basis for preparedness of the virus. This study reported the long-term surveillance of AIVs in the live bird market (LBM) of 16 cities in Shandong province from 2013 to 2019. A total of 29,895 samples were obtained and the overall positive rate of AIVs was 9.7%. The H9 was found to be the most predominant subtype in most of the time and contributed most to the monthly positve rate of AIVs as supported by the univariate and multivariate analysis, while H5 and H7 only circulated in some short periods. Then, the whole-genome sequences of 62 representative H9N2 viruses including one human isolate from a 7-year-old boy in were determined and they were genetically similar to each other with the median pairwise sequence identities ranging from 0.96 to 0.98 for all segments. The newly sequenced viruses were most similar to viruses isolated in chickens in mainland China, especially the provinces in Eastern China. Phylogenetic analysis showed that these newly sequenced H9N2 viruses belonged to the same clade for all segments except PB1. Nearly all of these viruses belonged to the G57 genotype which has dominated in China since 2010. Finally, several molecular markers associated with human adaptation, mammalian virulence, and drug resistance were identified in the newly sequenced H9N2 viruses. Overall, the study deepens our understanding of the epidemic and evolution of AIVs and provides a basis for effective control of AIVs in China.

Host diversity and behavior determine patterns of interspecies transmission and geographic diffusion of avian influenza A subtypes among North American wild reservoir species.

Wild birds can carry avian influenza viruses (AIV), including those with pandemic or panzootic potential, long distances. Even though AIV has a broad host range, few studies account for host diversity when estimating AIV spread. We analyzed AIV genomic sequences from North American wild birds, including 303 newly sequenced isolates, to estimate interspecies and geographic viral transition patterns among multiple co-circulating subtypes. Our results show high transition rates within Anseriformes and Charadriiformes, but limited transitions between these orders. Patterns of transition between species were positively associated with breeding habitat range overlap, and negatively associated with host genetic distance. Distance between regions (negative correlation) and summer temperature at origin (positive correlation) were strong predictors of transition between locations. Taken together, this study demonstrates that host diversity and ecology can determine evolutionary processes that underlie AIV natural history and spread. Understanding these processes can provide important insights for effective control of AIV.

The Impact of the Closure of the Live Poultry Market due to COVID-19 on the Avian Influenza Virus in Nanchang, Jiangxi Province, China.

ABSTRACT.This article aims to understand the changes in the detection rates of H5, H7, and H9 subtypes of avian influenza viruses (AIVs) in the live poultry markets (LPMs) in Nanchang City, Jiangxi Province, before and after the outbreak of the COVID-19. From 2019 to 2020, we monitored the LPM and collected specimens, using real-time reverse transcription polymerase chain reaction technology to detect the nucleic acid of type A AIV in the samples. The H5, H7, and H9 subtypes of influenza viruses were further classified for positive results. We analyzed 1,959 samples before and after the outbreak and found that the positive rates of avian influenza A virus (39.69%) and H9 subtype (30.66%) after the outbreak were significantly higher than before the outbreak (26.84% and 20.90%, respectively; P < 0.001). In various LPMs, the positive rate of H9 subtypes has increased significantly (P ≤ 0.001). Positive rates of the H9 subtype in duck, fecal, daub, and sewage samples, but not chicken samples, have increased to varying degrees. This study shows that additional measures are needed to strengthen the control of AIVs now that LPMs have reopened after the relaxing of COVID-19–related restrictions.

Genetic characterization of highly pathogenic avian Influenza H5Nx clade 2.3.4.4b reveals independent introductions in nigeria.

Among recurrent sanitary emergencies able to spread rapidly worldwide, avian influenza is one of the main constraints for animal health and food security. In West Africa, Nigeria has been experiencing repeated outbreaks of different strains of avian influenza virus (AIV) since 2006 and is also recognized as a hot spot in the region for the introduction of emerging strains by migratory wild birds. Here, we generated complete genomes of 20 highly pathogenic avian influenza (HPAI) H5N8 viruses collected during active surveillance in Nigerian live bird markets (LBM) and from outbreaks reported in the country between 2016 and 2019. Phylogenetic analysis reveals that the Nigerian viruses cluster into four separate genetic groups within HPAI H5 clade 2.3.4.4b. The first group includes 2016-2017 Nigerian viruses with high genetic similarity to H5N8 viruses detected in Central African countries, while the second includes Nigerian viruses collected both in LBM and poultry farms (2018-2019), as well as in Cameroon, Egypt and Siberia. A natural reassortant strain identified in 2019 represents the third group: H5N8 viruses with the same gene constellation were identified in 2018 in South Africa. Finally, the fourth introduction represents the first detection in the African continent of the H5N6 subtype, which is related to European viruses. Bayesian phylogeographic analyses confirmed that the four introductions originated from different sources and provide evidence of the virus spread within Nigeria, as well as diffusion beyond its borders. The multiple epidemiological links between Nigeria, Central and Southern African countries highlight the need for harmonized and coordinated surveillance system to control AIV impact. Improved surveillance at the Wetlands, LBMs and early warning of outbreaks are crucial for prevention and control of AIV, which can be potentially zoonotic and be a threat to human health.

Dominant subtype switch in avian influenza viruses during 2016\u20132019 in China

We have surveyed avian influenza virus (AIV) genomes from live poultry markets within China since 2014. Here we present a total of 16,091 samples that were collected from May 2016 to February 2019 in 23 provinces and municipalities in China. We identify 2048 AIV-positive samples and perform next generation sequencing. AIV-positive rates (12.73%) from samples had decreased substantially since 2016, compared to that during 2014–2016 (26.90%). Additionally, H9N2 has replaced H5N6 and H7N9 as the dominant AIV subtype in both chickens and ducks. Notably, novel reassortants and variants continually emerged and disseminated in avian populations, including H7N3, H9N9, H9N6 and H5N6 variants. Importantly, almost all of the H9 AIVs and many H7N9 and H6N2 strains prefer human-type receptors, posing an increased risk for human infections. In summary, our nation-wide surveillance highlights substantial changes in the circulation of AIVs since 2016, which greatly impacts the prevention and control of AIVs in China and worldwide.

Controlling Avian Influenza Virus in Bangladesh: Challenges and Recommendations.

Avian influenza virus (AIV) remains a huge challenge for poultry production with negative repercussions for micro- and macro-economy and public health in Bangladesh. High (HP) H5N1 and low pathogenicity (LP) H9N2 AIV are currently endemic in poultry, and both have been reported to infect humans sporadically. Multiple virus introductions of different clades of HPAIV H5N1, reassorted genotypes, and on-going diversification of LPAIV H9N2 create a highly volatile virological environment which potentially implicates increased virulence, adaptation to new host species, and subsequent zoonotic transmission. Allotropy of poultry rearing systems and supply chains further increase the risk of virus spreading, which leads to human exposure and fosters the emergence of new potentially pre-pandemic virus strains. Here, we review the epidemiology, focusing on (i) risk factors for virus spreading, (ii) viral genetic evolution, and (iii) options for AIV control in Bangladesh. It is concluded that improved control strategies would profit from the integration of various intervention tools, including effective vaccination, enhanced biosecurity practice, and improved awareness of producers and traders, although widespread household poultry rearing significantly interferes with any such strategies. Nevertheless, continuous surveillance associated with rapid diagnosis and thorough virus characterization is the basis of such strategies.

The potential of putative zinc-binding motifs of haemagglutinin (HA) protein for categorization and prediction of pathogenicity of H5 subtypes of avian influenza virus

In the present study, we used the potential of bioinformatics and computational analysis to predict the existence and biological relevance of zinc finger (ZF) motifs in heamagglutinin (HA) protein of Avian Influenza (AI) virus. Sequence data of Avian Influenza (AI) viruses were retrieved from accessible databases (GenBank, GISAID, IRD) and analyzed for the existence, as well as functional prediction of the putative zinc finger or ‘‘zinc-binding’’ motif(s) of HA protein. It is hypothesized that the ZF motif(s) in HA of AI virus can be used as a ‘‘novel’’ biomarker for categorization of the virus and/or its virulence. As a model for analysis, we used the H5 subtypes of highly pathogenic, non-pathogenic and low pathogenic avian influenza (HPAI, NPAI and LPAI) viruses of H5N1 and H5N2 of avian and human origins. Interestingly, our method of characterization using the zinc-finger agrees with the existing classification in distinguishing between highly pathogenic and non-pathogenic or low pathogenic subtypes. The new method also clearly distinguished between low and non-pathogenic strains of H5N2 and H5N1 which are indistinguishable by the existing method that utilizes the sequence of the polybasic amino acids of the proteolytic cleavage site for pathogenicity. It is hypothesized that zinc through the activities of zinc-binding proteins modulates the virulence property of the viral subtypes. Our observation further revealed that only the HA protein among the eight encoded proteins of influenza viruses contain high numbers of Cys-His residues. It is expected that the information gathered from the analysis of the data will be useful to generate more research hypotheses/designs that will give further insight towards the identification and control of avian influenza virus through the molecular manipulation of zinc finger motifs present in viral HA protein.

Control of avian influenza in China: Strategies and lessons.

In recent decades, multiple subtypes (i.e. H9N2, H5N1 and H7N9) of avian influenza virus (AIV) have become widespread in China, which has caused enormous economic losses and posed considerable threats to public health. In this review, with the aim to provide insights into and guidelines for the control of AIV spread in China and globally in the future, we analysed the reasons why AIV has persisted in China based on socio-economic features, including poultry biosecurity, live bird markets, live bird transportation, wild birds, poultry waterfowl, poultry density, poultry population and infected birds. We also described the present status of the AIV subtypes H9, H5 and H7 in China to elucidate the effectiveness of the strategies currently employed in China (i.e. culling, mass vaccination and biosecurity improvement) to control the disease based on a literature review and our unpublished surveillance data collected over a 12-year period from 2007 to 2018. We then summarized the lessons to be learned from the control experience in China, including whether culling of infected birds is of limited value for disease control and whether improved biosecurity is a better option than culling and vaccination for the long-term control of AIV, and when the vaccine strain should be updated.

MODELLING THE DYNAMICS OF AVIAN INFLUENZA WITH NONLINEAR RECOVERY RATE AND PSYCHOLOGICAL EFFECT

In this paper, a SI-SEIR type avian influenza epidemic model with psychological effect, nonlinear recovery rate and saturation inhibition effect is formulated to study the transmission and control of avian influenza virus. By setting the basic reproductive number as the threshold parameter and constructing Lyapunov function, Dulac function and using the Li-Muldowney's geometry approach, we prove the local and global stability of disease-free equilibria and endemic equilibrium. Theoretical analysis are carried out to show the role of the saturation inhibition effect, psychological effect and effective medical resources in this model, and numerical simulations are also given to verify the results.

Herbal Medicine Additives as Powerful Agents to Control and Prevent Avian Influenza Virus in Poultry – A Review

Abstract The complicated epidemiological situation of avian influenza viruses (AIV) caused by continuous emergence of new subtypes with failure of eradication, monitoring and vaccination strategies opens the door to alternative solutions to save the status quo and prevent new disasters for the poultry industry. Using of synthetic antiviral drugs such as neuraminidase and hemagglutinin inhibitors has been limited due to development of drug resistance and expensive commercial application. One of the most promising alternatives is herbal products and botanicals. This review presents a comprehensive and specialized view of in vivo studies of herbal plants in poultry species. Many herbal extracts as Nigella sativa oil, Astragalus, Cochinchina momordica and Sargassum pallidum polysaccharides proved very effective as adjuvants for AIV vaccines. Another beneficial role of herbs is enhancement of host response to vaccination with further better prevention of infection and easier control. For enumeration not inventory, this is best achieved with the use of virgin coconut oil, Echinacea purpurea, Ginseng stem-and-leaf saponins (GSLS), Astragalus polysaccharides (APS), Myrtus communis oil, Garlic powder, Turmeric, Thyme and Curcumin. This review aimed to evaluate most of the in vivo studies performed on poultry species as a step and a guide for scientists and field practitioners in establishment of new effective herbal-based drugs for prevention and control of AIV in poultry.

Protective efficacy in farmed ducks of a duck enteritis virus-vectored vaccine against H5N1, H5N6, and H5N8 avian influenza viruses

Ducks play a key role in the maintenance and spread of avian influenza viruses (AIVs) in nature, and control of AIVs in ducks has important implications for AIV eradication from poultry. We previously constructed a recombinant duck enteritis virus (DEV), rDEVus78HA, that expresses the HA gene of an H5N1 AIV and showed that rDEVus78HA immunization provides complete protection against both DEV and H5N1 AIV challenge in specific-pathogen-free ducks. In this study, we performed a 60-week clinical trial and found that this rDEVus78HA vaccine can function as a bivalent vaccine in farmed ducks against lethal challenge with DEV and H5N1 virus. Moreover, we found that rDEVus78HA-vaccinated ducks were efficiently protected against challenges with recently isolated heterologous H5N6 and H5N8 viruses. Our results demonstrate that rDEVus78HA could be extremely valuable for the control of DEV and H5 AIVs in ducks.

Molecular characteristic analysis of neuraminidase genes of avian influenza virus H9N2 in environments in Weining, Guizhou Province during 2015-2017

Objective To understand the genetic variations of neuraminidase (NA) genes of avian influenza virus H9N2 in Weining, Guizhou Province, and to provide the scientific evidence for the prevention and control of avian influenza virus. Methods Ribonucleic acids (RNA) were extracted and NA genes were amplified and sequenced from 13 randomly selected H9N2 positive samples from the live poultry market (LPM) environments in north of Weining Yi and Hui and Miao autonomous county (Weining), Guizhou Province during 2015 to 2017. Then the homology, genetic evolution, and sites of stalk deletion areas, potential N-glycosylation, receptor binding regions and drug resistance of H9N2 subtype avian influenza viruses were analyzed by a series of bioinformation software. Results Homology analysis revealed that there were 93.0%-100.0% and 92.1%-100.0% similarity among 13 strains H9N2 avian influenza viruses in nucleotide and amino acid of the NA gene, respectively. All strains belonged to DK/HK/Y280/97 sub-lineage, but their genetic sources were complex and diverse. Thirteen strains had a stalk deletion of 3 amino acid residues TEI at positions 63-65 and 3 isolates had mutation QN to QK at positions 39-40. The potential N-glycosylation sites at amino acid residues 86, 146, 200, and 234 of the NA protein of all strains were highly conserved, while other N-glycosylation sites had quantity and site mutations. There were different mutation types at the three sialic acid binding site areas, especially at 399-404 area. All NA protease activity sites and key sites of the 13 strains had no mutations associated with resistance to the neuraminidase inhibitor drugs. Conclusions All 13 strains H9N2 viruses belongs to DK/HK/Y280/97 sub-lineage in Weining, Guizhou Province during 2015-2017, and their genetic sources are complex and diverse. The mutations on sites of stalk areas, potential N-glycosylation and sialic acid binding site areas are presented at different degrees. Hence, enhancing surveillance and controlling H9N2 avian influenza virus is necessary. Key words: Influenza A virus, H9N2 subtype; Neuraminidase gene; Molecular characteristic

The second-generation thiazolide haloxanide is a potent inhibitor of avian influenza virus replication

The emergence of new avian influenza virus (AIV) strains able to infect humans represents a serious threat to global human health. In addition to surveillance and vaccine development, antiviral therapy remains crucial for AIV control; however, the increase in drug-resistant AIV strains underscores the need for novel approaches to anti-influenza chemotherapy. We have previously shown that the thiazolide anti-infective nitazoxanide (NTZ) inhibits influenza A/PuertoRico/8/1934(H1N1) virus replication, and this effect was associated with inhibition of viral hemagglutinin (HA) maturation. Herein we investigated the activity of the second-generation thiazolide haloxanide (HLN) against H5N9, H7N1 and H1N1 AIV infection in vitro, and explored the mechanism of the antiviral action. Using the A/chicken/Italy/9097/1997(H5N9) AIV as a model, we show that HLN and its precursor p-haloxanide are more effective than NTZ against AIV, with IC50 ranging from 0.03 to 0.1 μg/ml, and SI ranging from 200 to >700, depending on the multiplicity of infection. Haloxanide did not affect AIV entry into target cells and did not cause a general inhibition of viral protein expression, whereas it acted at post-translational level by inhibiting HA maturation at a stage preceding resistance to endoglycosidase-H digestion. Importantly, this effect was independent of the AIV-HA subtype and the host cell. Immunomicroscopy and receptor-binding studies confirmed that HLN-induced alterations impair AIV-HA trafficking to the host cell plasma membrane, a key step for viral morphogenesis. The results indicate that haloxanide could provide a new tool for treatment of avian influenza virus infections.

Stability analysis and optimal control of avian influenza virus A with time delays

In this research article we have developed a mathematical model to describe the spread of avian influenza A (H7N9) virus from the birds to human. We also consider the incubation periods of avian influenza A (H7N9) virus in this model with different time delay in the infective avian and human populations. By analyzing behaviour of the model, we calculate the basic reproduction number and investigate the local and global stability of equilibria of the system. Here we also set up an optimal control problem and used a quadratic control to reduce the spread of the disease and the cost of treatment. Computer simulations are carried out to explain the analytical findings.

Recombinant Hemagglutinin of Avian Influenza Virus H5 Expressed in the Chloroplast of Chlamydomonas reinhardtii and Evaluation of Its Immunogenicity in Chickens.

Globally, avian influenza (AI) is a serious problem in poultry farming. Despite vaccination, the prevalence of AI in México highlights the need for new approaches to control AI and to reduce the economic losses associated with its occurrence in susceptible birds. Recombinant proteins from avian influenza virus (AIV) have been expressed in different organisms, such as plants. The present study investigated the feasibility of designing and expressing the HA protein of AIV in the transplastomic microalga Chlamydomonas reinhardtii as a novel approach for AIV control and taking advantage of culture conditions, its reproductive range, and safe use in consideration of the generally regarded as safe food ingredient regulatory classification. The results showed that the HA protein of AIV in C. reinhardtii presents antigenic activity by western blot test and through its application in chickens, demonstrating its feasibility as a recombinant antigen against AIV.

The role of IRF7 and NF-κB pathways in the induction of antiviral responses in chicken tracheal epithelial cells following exposure to TLR3 and 4 ligands

Abstract With recent outbreaks of avian influenza viruses (AIV), there is a need for better understanding of host responses in the respiratory system to AIV in order to develop and improve prophylactic procedures for control of AIV. Toll-like receptors (TLRs) are molecules in the immune system that rapidly induce antiviral responses in the host. Activation of TLRs by their ligands leads to a variety of cellular responses, including the production of interferons (IFNs). However, there is little information about induced antiviral responses in chicken trachea following TLR ligand treatment. We cultured primary chicken tracheal epithelial cells (cTECs) to determine antiviral responses induced by TLR3 and 4 ligands. These cells were treated with polyI:C and LPS from Escherichia coli 026:B6, TLR3 and 4 ligands respectively, before AIV infection. We also used BX795 and celastrol, inhibitors of IRF7 and NF-κB pathways, respectively, in order to gain insight into the mechanisms involved in the induced antiviral responses. Our results showed that treatment of cTECs with TLR ligands reduced the replication of AIV in cTECs. Treatment of cTECs with polyI:C resulted in highest reduction of viral replication in cTECs followed by treatment of cTECs with LPS. BX795 increased AIV replication in polyI:C and LPS treated cells while celastrol increased AIV replication in polyI:C treated cells. Overall, these results provided evidence that cTECs are able to mount antiviral responses against AIV after TLR ligand stimulation. Moreover, we determined the role of IRF7 and NF-κB signalling pathways in the induction of antiviral responses. Studies are underway to examine the ability of cTECs to produce type I IFNs and the effect of other cells, such as macrophages, on cTECs.

Phylogenetic Analysis and Pathogenicity Assessment of Two Strains of Avian Influenza Virus Subtype H9N2 Isolated from Migratory Birds: High Homology of Internal Genes with Human H10N8 Virus.

Two human-infecting avian influenza viruses (AIVs), H7N9 and H10N8, have emerged in China, which further indicate that the H9N2 subtype of AIVs, as an internal gene donor, may have an important role in the generation of new viruses with cross-species transmissibility and pathogenicity. H9N2 viruses that contain such internal genes widely exist in poultry but are rarely reported in migratory birds. In this study, two strains of the H9N2 virus were isolated from fecal samples of migratory birds in 2014: one strain from Caizi Lake in Anhui Province and one from Chen Lake in Hubei Province of China. Nucleotide sequence analysis revealed high homology of all six internal genes of these two strains with the internal genes of the human H10N8 virus in Jiangxi Province, as well as with the human H7N9 virus. Phylogenetic analysis indicated a possible origin of these two strains from poultry in South China. Both of the two viruses tested could replicated in respiratory organs of infective mice without adaption, by both strains of the H9N2 AIVs from wild birds, suggesting their potential capacity for directly infecting mammals. Our findings indicate the existence of H9N2 viruses that contain internal genes highly homologous with human H10N8 or H7N9 viruses. Wild birds can contribute to the spread of the H9N2 virus that contains the “harmful” internal gene complex, leading to gene rearrangement with other influenza viruses and to the generation of new pathogenic viruses. Therefore, strengthening AIV surveillance in wild birds can promote an understanding of the presence and prevalence of viruses and provide scientific evidence for the prevention and control of AIVs and human-infecting AIVs.