The Newcastle disease (ND) and avian influenza (AI) are two crucial viral ailments of poultry birds. In field conditions, these are controlled by vaccination and biosecurity measures. The cumbersome ...application process, potential for reversion to the wild pathogen, and financial load on farmers are only a few drawbacks of the currently available ND and AI vaccines. There was a pressing need to develop a stable, safer, bivalent vaccine for these two diseases. Hence, this study adopts the reverse genetic technology, thereby rescuing the recombinant LaSota strain of NDV, which expresses the haemagglutinin (HA) protein of indigenously isolated H9N2 virus (rNDV-H9). The rNDV-H9 was tested in terms of pathogenicity and growth kinetics. Subsequently, the day-old broiler chicks were immunised with rNDV-H9, followed by challenge infection by virulent NDV and H9N2 viruses. The results indicate that immunisation with rNDV-H9 has reduced the shedding of challenge viruses. The clinical progression of the diseases was slowed down, and all the birds were seroconverted. Results in this study indicate that rNDV-H9 is a promising candidate for immunisation in endemic areas of ND and AI, with minimum economic pressure on farmers and reduced vaccinal stress on poultry birds.
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•The vaccine has conferred optimum protection in chickens during pre-clinical trials.•It can differentiate infected from vaccinated animal & facilitates disease tracking.•The vaccine can reduce the economic pressure on farmer and vaccinal stress on birds.•The Seamless PCR cloning and RNA polymerase-II has improved the recovery of virus.
Ducks and wild waterfowl perpetuate all strains of influenza viruses in nature. In their natural host, influenza viruses typically cause asymptomatic infection and little pathology. Ducks are often ...resistant to influenza viruses capable of killing chickens. Here, we show that the influenza virus sensor, RIG-I, is present in ducks and plays a role in clearing an influenza infection. We show evidence suggesting that RIG-I may be absent in chickens, providing a plausible explanation for their increased susceptibility to influenza viruses compared with ducks. RIG-I detects RNA ligands derived from uncapped viral transcripts and initiates the IFN response. In this study, we show that the chicken embryonic fibroblast cell line, DF-1, cannot respond to a RIG-I ligand. However, transfection of duck RIG-I into DF-1 cells rescues the detection of ligand and induces IFN-β promoter activity. Additionally, DF-1 cells expressing duck RIG-I have an augmented IFN response resulting in decreased influenza replication after challenge with either low or highly pathogenic avian influenza virus. Implicating RIG-I in the antiviral response to an infection in vivo, we found that RIG-I expression is induced 200 fold, early in an innate immune response in ducks challenged with the H5N1 virus A/Vietnam/1203/04. Finding this natural disease resistance gene in ducks opens the possibility of increasing influenza resistance through creation of a transgenic chicken.
Summary Background Replication-competent virus vector vaccines might have advantages compared with non-replicating vector vaccines. We tested the safety and immunogenicity of an oral adenovirus ...serotype 4 vector vaccine candidate (Ad4-H5-Vtn) expressing the haemagglutinin from an avian influenza A H5N1 virus. Methods We did this phase 1 study at four sites in the USA. We used a computer-generated randomisation list (block size eight, stratified by site) to assign healthy volunteers aged 18–40 years to receive one of five doses of Ad4-H5-Vtn (107 viral particles VP, 108 VP, 109 VP, 1010 VP, 1011 VP) or placebo (3:1). Vaccine or placebo was given on three occasions, about 56 days apart. Participants, investigators, and study-site personnel were masked to assignment throughout the study. Subsequently, volunteers received a boost dose with 90 μg of an inactivated parenteral H5N1 vaccine. Primary immunogenicity endpoints were seroconversion by haemagglutination-inhibition (HAI), defined as a four-times rise compared with baseline titre, and HAI geometric mean titre (GMT). We solicited symptoms of reactogenicity daily for 7 days after each vaccination and recorded symptoms that persisted beyond 7 days as adverse events. Primary analysis was per protocol. This trial is registered with ClinicalTrials.gov , number NCT01006798. Findings We enrolled 166 participants (125 vaccine; 41 placebo) between Oct 19, 2009, and Sept 9, 2010. HAI responses were low: 13 of 123 vaccinees (11%, 95% CI 6–17) and three of 41 placebo recipients (7%, 2–20) seroconverted. HAI GMT was 6 (95% CI 5–7) for vaccinees, and 5 (5–6) for placebo recipients. However, when inactivated H5N1 vaccine became available, one H5N1 boost was offered to all participants. In this substudy, HAI seroconversion occurred in 19 of 19 participants in the 1011 VP cohort (100%; 95% CI 82–100) and eight of 22 placebo recipients (36%; 17–59); 17 of 19 participants in the 1011 VP cohort (89%; 67–99) achieved seroprotection compared with four of 22 placebo recipients (18%; 5–40); GMT was 135 (89–205) with 1011 VP, compared with 13 (7–21) with placebo. The cumulative frequency of abdominal pain, diarrhoea, and nasal congestion after all three vaccinations was significantly higher in vaccinees than placebo recipients (21 16·8% of 125 vs one 2·4% of 41, p=0·017; 24 19·2% of 125 vs two 4·9% of 41, p=0·027; 41 32·8% of 125 vs six 14·6% of 41, p=0·028; respectively). No serious treatment-related adverse events occurred. Interpretation Oral Ad4 vector priming might enhance the efficacy of poorly immunogenic vaccines such as H5N1. Funding Wellcome Trust Foundation, PaxVax.
► The hemagglutinin (HA) of influenza viruses mediates receptor binding. ► Receptor binding specificity is a major determinant of host range restriction. ► Avian influenza viruses do not transmit ...efficiently from human to human. ► Changes in binding specificity are required for cross-species transfer.
Influenza A virus infection begins with the binding of the hemagglutinin (HA) glycoprotein to sialic acid-containing receptors on the surface of the target cell. Avian influenza viruses, including avian H5N1, H7, and H9N2 viruses, can occasionally cross the species barrier and infect humans; however, these viruses do not spread efficiently from person to person, perhaps, partly, owing to differences in the receptor-binding specificities of human and avian influenza viruses. The HAs of avian influenza viruses must adapt to receptors in humans to acquire efficient human-to-human transmissibility. In this review, we discuss the receptor binding specificity of influenza A viruses and its role in interspecies transmission.
Statistical methods for molecular dating of viral origins have been used extensively to infer the time of most common recent ancestor for many rapidly evolving pathogens. However, there are a number ...of cases, in which epidemiological, historical, or genomic evidence suggests much older viral origins than those obtained via molecular dating. We demonstrate how pervasive purifying selection can mask the ancient origins of recently sampled pathogens, in part due to the inability of nucleotide-based substitution models to properly account for complex patterns of spatial and temporal variability in selective pressures. We use codon-based substitution models to infer the length of branches in viral phylogenies; these models produce estimates that are often considerably longer than those obtained with traditional nucleotide-based substitution models. Correcting the apparent underestimation of branch lengths suggests substantially older origins for measles, Ebola, and avian influenza viruses. This work helps to reconcile some of the inconsistencies between molecular dating and other types of evidence concerning the age of viral lineages.
Abstract Human influenza causes substantial morbidity and mortality. Currently, licensed influenza vaccines offer satisfactory protection if they match the infecting strain, but they come with ...significant drawbacks. These vaccines are derived from prototype viruses, containing the hemagglutinin of influenza viruses that are likely to cause the next epidemic. Their usefulness against a future pandemic, however, remains problematic. A vaccine based on the ectodomain of influenza matrix protein 2 (M2e) could overcome these drawbacks. M2e is highly conserved in both human and avian influenza A viruses. The low immunogenicity against natural M2e can be overcome by fusing M2e to an appropriate carrier such as Hepatitis B virus-derived virus-like particles. Such chimeric particles can be produced in a simple and safe bacterial expression system, requiring minimal biocontainment, and can be obtained in a pure form. Experiments in animal models have demonstrated that M2e-based vaccines induce protection against a lethal challenge with various influenza A virus subtypes. Furthermore, the production and use of an effective M2e-vaccine could be implemented at any time regardless of seasonality, both in an epidemic as well as in a pandemic preparedness program. In animal models, M2e-vaccines administered parenterally or intranasally protect against disease and mortality following challenge with various influenza A strains. Adjuvants suitable for human use improve protection, which correlates with higher anti-M2e antibody responses of defined subtypes. Recently, Phase I clinical studies with M2e-vaccines have been completed, indicating their safety and immunogenicity. Further clinical development of this universal influenza A vaccine candidate is being pursued in order to validate its protective efficacy in humans.
This book contains comprehensive and updated information on how to perform Avian Influenza and Newcastle Disease diagnosis from the suspicion in the field to the characterisation of isolates. It ...provides guidelines to outbreak management, field investigation, necropsy techniques, sampling methods and complete laboratory diagnosis, including molecular methods. The outstanding images collected from field outbreaks, including clinical and pathological findings, and the selection of laboratory protocols make this publication unique. It will therefore be an invaluable tool for all veterinarians, scientists, animal health professionals, and public health officials involved in the management of these infections.
Recent cases of avian influenza H5N1 and the swine-origin 2009 H1N1 have caused a great concern that a global disaster like the 1918 influenza pandemic may occur again. Viral transmission begins with ...a critical interaction between hemagglutinin (HA) glycoprotein, which is on the viral coat of influenza, and sialic acid (SA) containing glycans, which are on the host cell surface. To elucidate the role of HA glycosylation in this important interaction, various defined HA glycoforms were prepared, and their binding affinity and specificity were studied by using a synthetic SA microarray. Truncation of the N-glycan structures on HA increased SA binding affinities while decreasing specificity toward disparate SA ligands. The contribution of each monosaccharide and sulfate group within SA ligand structures to HA binding energy was quantitatively dissected. It was found that the sulfate group adds nearly 100-fold (2.04 kcal/mol) in binding energy to fully glycosylated HA, and so does the biantennary glycan to the monoglycosylated HA glycoform. Antibodies raised against HA protein bearing only a single N-linked GlcNAc at each glycosylation site showed better binding affinity and neutralization activity against influenza subtypes than the fully glycosylated HAs elicited. Thus, removal of structurally nonessential glycans on viral surface glycoproteins may be a very effective and general approach for vaccine design against influenza and other human viruses.
Understanding the transmission dynamics and persistence of avian influenza viruses (AIVs) in the wild is an important scientific and public health challenge because this system represents both a ...reservoir for recombination and a source of novel, potentially human-pathogenic strains. The current paradigm locates all important transmission events on the nearly direct fecal/oral bird-to-bird pathway. In this article, on the basis of overlooked evidence, we propose that an environmental virus reservoir gives rise to indirect transmission. This transmission mode could play an important epidemiological role. Using a stochastic model, we demonstrate how neglecting environmentally generated transmission chains could underestimate the explosiveness and duration of AIV epidemics. We show the important pathogen invasion implications of this phenomenon: the nonnegligible probability of outbreak even when direct transmission is absent, the long-term infectivity of locations of prior outbreaks, and the role of environmental heterogeneity in risk.
► 22 Flavonoids were examined for activity against H5N1 influenza A viruses. ► Biochanin A and baicalein exerted the highest potency. ► Biochanin A and baicalein interfered with H5N1 replication. ► ...Biochanin A and baicalein interfered with virus-induced cytokine expression. ► Biochanin A and baicalein differ in their molecular antiviral mechanisms.
From a panel of 22 flavonoids, we identified six compounds (apigenin, baicalein, biochanin A, kaempferol, luteolin, naringenin) that inhibited influenza A nucleoprotein production in human lung epithelial (A549) cells infected with the highly pathogenic avian influenza H5N1 virus strain A/Thailand/Kan-1/04 in non-toxic concentrations. Baicalein (IC50: 18.79±1.17μM, selectivity index 5.82) and biochanin A (IC50 8.92±1.87μM, selectivity index 5.60) were selected for further experiments. Both compounds reduced H5N1 infectious titres (baicalein 40μM: 29-fold reduction, biochanin A 40μM: 55-fold reduction after infection at MOI 0.01), virus-induced caspase 3 cleavage, nuclear export of viral RNP complexes, and enhanced the effects of the neuraminidase inhibitor zanamivir. Biochanin A and baicalein also inhibited the replication of the H5N1 strain A/Vietnam/1203/04. Time of addition experiments indicated that both compounds interfere with H5N1 replication after the adsorption period. Further mechanistic investigations revealed clear differences between these two flavonoids. Only baicalein interfered with the viral neuraminidase activity (39±7% inhibition at 100μM, the maximum concentration tested). In contrast to baicalein, biochanin A affected cellular signalling pathways resulting in reduced virus-induced activation of AKT, ERK 1/2, and NF-kB. Moreover, biochanin A inhibited the virus-induced production of IL-6, IL-8, and IP-10 while baicalein inhibited IL-6 and IL-8 production without affecting IP-10 levels. In primary human monocyte-derived macrophages, only baicalein but not biochanin A impaired H5N1 virus replication. Both flavonoids interfered with the H5N1-induced production of IL-6, IP-10, and TNF-α but not of IL-8 in macrophages. These findings indicate that closely related flavonoids can exert anti-H5N1 effects by different molecular mechanisms.
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