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Tare D.S.,National Institute of Virology Microbial Containment Complex | Pawar S.D.,National Institute of Virology Microbial Containment Complex
Journal of Virological Methods | Year: 2015

Avian influenza (AI) H9N2 viruses are endemic in many bird species, and human infections of H9N2 viruses have been reported. Oseltamivir phosphate (Tamiflu®) is the available antiviral drug for the treatment and prophylaxis of influenza. There are no reports of use of embryonated chicken egg as a model to study susceptibility of AI viruses to oseltamivir carboxylate (OC), the active metabolite. The present study was undertaken to explore the use of embryonated chicken eggs as a model for testing OC against the AI H9N2 viruses. A total of three AI H9N2 viruses, isolated in poultry in India, were used. Various virus dilutions were tested against 14μg/ml of OC. Three methods, namely (1) the in vitro virus-drug treatment, (2) drug delivery and virus challenge by allantoic route, and (3) drug delivery by albumen route and virus challenge by allantoic route were explored. The viruses were also tested using the fluorescence-based neuraminidase inhibitor (NAI) assay. There was significant inhibition (p<0.05) of the H9N2 viruses in presence of OC. The infectious virus titers as well as hemagglutination titers were significantly lower in presence of OC as compared to controls. The in vitro treatment of virus and drug; and drug and virus delivery at the same time by allantoic route showed significantly higher inhibition (p<0.05) of virus growth than that by the albumen route. In the NAI assay, the half maximal inhibitory concentration (IC50) values of the H9N2 viruses were within the standard range for known susceptible reference virus. In conclusion, the H9N2 viruses used in the study were susceptible to OC. Embryonated chicken egg could be used as a model to study susceptibility of AI viruses to antiviral drugs. © 2015 Elsevier B.V. Source


Pawar S.D.,National Institute of Virology Microbial Containment Complex | Murtadak V.B.,National Institute of Virology Microbial Containment Complex | Kale S.D.,National Institute of Virology Microbial Containment Complex | Shinde P.V.,National Institute of Virology Microbial Containment Complex | Parkhi S.S.,National Institute of Virology Microbial Containment Complex
Journal of Virological Methods | Year: 2015

In view of the emerging avian influenza (AI) viruses, it is important to study the susceptibility of AI viruses to inactivating agents for preparation of antigens and inactivated vaccines. The available information on susceptibility of both the high and low pathogenic AI viruses to different inactivating agents is inadequate and ambiguous. It has been shown that different subtypes of influenza viruses require different physical and chemical conditions for inactivation of infectivity. The present study was undertaken to evaluate the use of beta-propiolactone (BPL), formalin and ether for inactivation and its impact on antigenicity of AI viruses.A total of nine high and low pathogenic AI viruses belonging to four influenza A subtypes were included in the study. The H5N1 viruses were from the clades 2.2, 2.3.2.1 and 2.3.4. The H9N2 virus included in the study was of the G1 genotype, while the H11N1 and H4N6 viruses were from the Eurasian lineage. The viruses were treated with BPL, formalin and with ether. The confirmation of virus inactivation was performed by two serial passages of inactivated viruses in embryonated chicken eggs.The infectivity of all tested AI viruses was eliminated using 0.1% BPL and 0.1% formalin. Ether eliminated infectivity of all tested low pathogenic AI viruses; however, ether with 0.2% or 0.5% Tween-20 was required for inactivation of the highly pathogenic AI H5N1 viruses. Treatment with BPL, ether and formalin retained virus hemagglutination (HA) titers. Interestingly ether treatment resulted in significant rise in HA titers (. P<. 0.05) of all tested AI viruses. This data demonstrated the utility of BPL, formalin and ether for the inactivation of infectivity of AI viruses used in the study for the preparation of inactivated virus antigens for research and diagnosis of AI. © 2015. Source


Shinde P.V.,National Institute of Virology Microbial Containment Complex | Koratkar S.S.,National Institute of Virology Microbial Containment Complex | Pawar S.D.,National Institute of Virology Microbial Containment Complex | Kale S.D.,National Institute of Virology Microbial Containment Complex | And 2 more authors.
Avian Diseases | Year: 2012

An avian influenza (AI) surveillance was undertaken in Maharashtra state, India during the period 2010-2011. There are no reports of AI surveillance in emus from India. A total of 202 blood samples and 467 tracheal and cloacal swabs were collected from eight emu farms. A hemagglutination inhibition (HI) assay was performed for detection of antibodies against AI H5N1, H7N1, H9N2, and avian paramyxovirus type 1 (APMV-1) viruses. A microneutralization (MN) assay was performed to confirm the presence of neutralizing antibodies against AI H9N2 and to compare with HI assays. A total of 28.2% and 28.7% of samples were positive for antibodies against AI H9N2 by HI and MN assays, respectively, using ≥1:40 as a cut-off titer; 15.3% samples were positive for APMV-1 by HI assay using a ≥1:10 cut-off titer. Seropositivity of AI H9N2 was nil in the grower (<1 yr) age group and highest (78%) in the breeder (23 yr) age group, whereas seropositivity against APMV-1 was observed in all age groups. Performance of both HI and MN assays was similar, suggesting the utility of using the MN assay along with HI assay for surveillance studies. This is the first report of the seroprevalence of AI H9N2 and APMV-1 in emus in India. © 2012 American Association of Avian Pathologists. Source

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