Complete Genome Sequence of a Newcastle Disease Virus Isolated from Wild Peacock (Pavo cristatus) in India Sagar A. Khulape,a Satish S. Gaikwad,b Madhan Mohan Chellappa,a Bishnu Prasad Mishra,a Sohini Deya Division of Veterinary Biotechnology, Indian Veterinary Research Institute, Izatnagar, Uttar Pradesh, Indiaa; OIE Reference Laboratory for Newcastle Disease, Gyeonggi-do, South Koreab
We report here the complete genome sequence of a Newcastle disease virus (NDV) isolated from a wild peacock. Phylogenetic analysis showed that it belongs to genotype II, class II of NDV strains. This study helps to understand the ecology of NDV strains circulating in a wild avian host of this geographical region during the outbreak of 2012 in northwest India.
Citation Khulape SA, Gaikwad SS, Chellappa MM, Mishra BP, Dey S. 2014. Complete genome sequence of a Newcastle disease virus isolated from wild peacock (Pavo cristatus) in India. Genome Announc. 2(3):e00495-14. doi:10.1128/genomeA.00495-14. Copyright © 2014 Khulape et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 3.0 Unported license. Address correspondence to Sohini Dey, [email protected].
N
ewcastle disease (ND) is a listed disease of the World Organisation for Animal Health affecting various avian species, with wide geographical distribution. The disease is caused by avian paramyxovirus serotype 1 (APMV-1) or Newcastle disease virus (NDV), the prototype of theAvulavirusgenusoftheParamyxoviridaefamily.Wildbirdshaveconstantly been implicated in the spread of ND to domestic bird populations (1). In 2012, an NDV outbreak caused significant fatality in a wild peacockpopulationinthenorthwestregionofIndia(2).Thenaturalhabitats ofpeacocksarestreamsideandcultivatedareas.Thespeciesisalsoknown to forage around human habitats and villages for scraps. The movement ofthiswildspeciesincloseproximitytodomesticbirds,mostofwhichare reared as backyard poultry, always threatens to the point of becoming a disease outbreak. A necropsy sample comprised of brain tissue of a dead peacock suspected to have ND from the wild was received from Rewari of Haryana state, India. Virus identification was done by conventional diagnostic methods, such as virus isolation in 9-day-old specific-pathogen-free embryonated chicken egg hemagglutination test, which was further confirmed by a molecular diagnostic test, reverse transcription-PCR (RTPCR). Complete genome sequencing of the isolate was done using an overlapping RT-PCR strategy (3) to cover the full-length genome of the isolate(ABI3730DNAanalyzer;AppliedBiosystems).Thesequenceediting and assembly were done using the Serial Cloner program, and the isolate was referred to as NDV/Peacock/India/2012. The total length of the viral genomewasfoundtobe15,186nucleotides,similartothatofotherNDV strains from early genotypes found during 1930 to 1960 (1). Genome sequencing revealed the presence of a polybasic amino acid motif 111GRRQKRF117 at the fusion protein cleavage site (FPCS), a primary molecular determinant of virulence for NDV, characteristic of virulent viruses (4). The vulnerability of wild peacocks in this case may be due to the absence of neutralizing antibodies, as the country lacks a dedicated wild avian vaccination program. The phylogenetic relationship of NDV/Peacock/India/2012 with 54 previously characterized AMPV-1 strains from the Asian continent and other parts of the world was analyzed using the BEAST software package (5). The maximum clade credibility tree of the completed genome scale
May/June 2014 Volume 2 Issue 3 e00495-14
data placed the isolate in genotype II, class II of APMV type I viruses with the highest posterior probability value. The isolate was also placed as a different clade in genotype II viruses, as evidenced by the mean percent nucleotide sequence distance for the complete genome (5.7%), as determined by pairwise sequence comparison analysis (PASC) using the MEGA5 software (6). The elucidation of the complete genome of the viral isolate will aid in the development of a vaccine candidate against homologous genotypes that are in circulation in India. Nucleotide sequence accession number. The complete genome sequence of NDV/Peacock/India/2012 has been deposited in GenBank under the accession no. KJ769262. ACKNOWLEDGMENTS This work was supported by grants from National Fund for Basic and Strategic Research in Agriculture of ICAR (NFBSFARA/BS-3010) and the Department of Biotechnology, Government of India (BT/PR15373/AAQ/ 57/116/2011 and DBT-JRF/2012-13/107).
REFERENCES 1. Alexander DJ. 1998. Newcastle disease virus and other avian paramyxoviruses, p 56 –163. In Swayne DE (ed), A laboratory manual for the isolation and identification of avian pathogens, 3rd ed. American Association of Avian Pathologists, Kennett Square, PA. 2. Kumar A, Maan S, Mahajan NK, Rana VP, Jindal N, Batra K, Ghosh A, Mishra SK, Kapoor S, Maan NS. 2013. Detection and molecular characterization of Newcastle disease virus in peafowl (Pavo cristatus) in Haryana state, India. Indian J. Virol. 24:380–385. http://dx.doi.org/10.1007/s13337-013-0169-5. 3. Chellappa MM, Dey S, Gaikwad S, Kataria JM, Vakharia VN. 2012. Complete genome sequence of Newcastle disease virus mesogenic vaccine strain R2B from India. J. Virol. 86:13814–13815. http://dx.doi.org/10.1128/JVI.02552-12. 4. Glickman RL, Syddall RJ, Iorio RM, Sheehan JP, Bratt MA. 1988. Quantitative basic residue requirements in the cleavage-activation site of the fusion glycoprotein as a determinant of virulence for Newcastle disease virus. J. Virol. 62:354 –356. 5. Drummond AJ, Suchard MA, Xie D, Rambaut A. 2012. Bayesian phylogenetics with BEAUti and the BEAST 1.7. Mol. Biol. Evol. 29:1969-1973. http://dx.doi.org/10.1093/molbev/mss075. 6. Tamura K, Peterson D, Peterson N, Stecher G, Nei M, Kumar S. 2011. MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Mol. Biol. Evol. 28:2731–2739. http://dx.doi.org/10.1093/molbev/msr121.
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Received 3 May 2014 Accepted 14 May 2014 Published 5 June 2014
We report here the complete genome sequence of a Newcastle disease virus (NDV) isolated from a wild peacock. Phylogenetic analysis showed that it belongs to genotype II, class II of NDV strains. This study helps to understand the ecology of NDV strains circulating in a wild avian host of this geographical region during the outbreak of 2012 in northwest India.
Citation Khulape SA, Gaikwad SS, Chellappa MM, Mishra BP, Dey S. 2014. Complete genome sequence of a Newcastle disease virus isolated from wild peacock (Pavo cristatus) in India. Genome Announc. 2(3):e00495-14. doi:10.1128/genomeA.00495-14. Copyright © 2014 Khulape et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 3.0 Unported license. Address correspondence to Sohini Dey, [email protected].
N
ewcastle disease (ND) is a listed disease of the World Organisation for Animal Health affecting various avian species, with wide geographical distribution. The disease is caused by avian paramyxovirus serotype 1 (APMV-1) or Newcastle disease virus (NDV), the prototype of theAvulavirusgenusoftheParamyxoviridaefamily.Wildbirdshaveconstantly been implicated in the spread of ND to domestic bird populations (1). In 2012, an NDV outbreak caused significant fatality in a wild peacockpopulationinthenorthwestregionofIndia(2).Thenaturalhabitats ofpeacocksarestreamsideandcultivatedareas.Thespeciesisalsoknown to forage around human habitats and villages for scraps. The movement ofthiswildspeciesincloseproximitytodomesticbirds,mostofwhichare reared as backyard poultry, always threatens to the point of becoming a disease outbreak. A necropsy sample comprised of brain tissue of a dead peacock suspected to have ND from the wild was received from Rewari of Haryana state, India. Virus identification was done by conventional diagnostic methods, such as virus isolation in 9-day-old specific-pathogen-free embryonated chicken egg hemagglutination test, which was further confirmed by a molecular diagnostic test, reverse transcription-PCR (RTPCR). Complete genome sequencing of the isolate was done using an overlapping RT-PCR strategy (3) to cover the full-length genome of the isolate(ABI3730DNAanalyzer;AppliedBiosystems).Thesequenceediting and assembly were done using the Serial Cloner program, and the isolate was referred to as NDV/Peacock/India/2012. The total length of the viral genomewasfoundtobe15,186nucleotides,similartothatofotherNDV strains from early genotypes found during 1930 to 1960 (1). Genome sequencing revealed the presence of a polybasic amino acid motif 111GRRQKRF117 at the fusion protein cleavage site (FPCS), a primary molecular determinant of virulence for NDV, characteristic of virulent viruses (4). The vulnerability of wild peacocks in this case may be due to the absence of neutralizing antibodies, as the country lacks a dedicated wild avian vaccination program. The phylogenetic relationship of NDV/Peacock/India/2012 with 54 previously characterized AMPV-1 strains from the Asian continent and other parts of the world was analyzed using the BEAST software package (5). The maximum clade credibility tree of the completed genome scale
May/June 2014 Volume 2 Issue 3 e00495-14
data placed the isolate in genotype II, class II of APMV type I viruses with the highest posterior probability value. The isolate was also placed as a different clade in genotype II viruses, as evidenced by the mean percent nucleotide sequence distance for the complete genome (5.7%), as determined by pairwise sequence comparison analysis (PASC) using the MEGA5 software (6). The elucidation of the complete genome of the viral isolate will aid in the development of a vaccine candidate against homologous genotypes that are in circulation in India. Nucleotide sequence accession number. The complete genome sequence of NDV/Peacock/India/2012 has been deposited in GenBank under the accession no. KJ769262. ACKNOWLEDGMENTS This work was supported by grants from National Fund for Basic and Strategic Research in Agriculture of ICAR (NFBSFARA/BS-3010) and the Department of Biotechnology, Government of India (BT/PR15373/AAQ/ 57/116/2011 and DBT-JRF/2012-13/107).
REFERENCES 1. Alexander DJ. 1998. Newcastle disease virus and other avian paramyxoviruses, p 56 –163. In Swayne DE (ed), A laboratory manual for the isolation and identification of avian pathogens, 3rd ed. American Association of Avian Pathologists, Kennett Square, PA. 2. Kumar A, Maan S, Mahajan NK, Rana VP, Jindal N, Batra K, Ghosh A, Mishra SK, Kapoor S, Maan NS. 2013. Detection and molecular characterization of Newcastle disease virus in peafowl (Pavo cristatus) in Haryana state, India. Indian J. Virol. 24:380–385. http://dx.doi.org/10.1007/s13337-013-0169-5. 3. Chellappa MM, Dey S, Gaikwad S, Kataria JM, Vakharia VN. 2012. Complete genome sequence of Newcastle disease virus mesogenic vaccine strain R2B from India. J. Virol. 86:13814–13815. http://dx.doi.org/10.1128/JVI.02552-12. 4. Glickman RL, Syddall RJ, Iorio RM, Sheehan JP, Bratt MA. 1988. Quantitative basic residue requirements in the cleavage-activation site of the fusion glycoprotein as a determinant of virulence for Newcastle disease virus. J. Virol. 62:354 –356. 5. Drummond AJ, Suchard MA, Xie D, Rambaut A. 2012. Bayesian phylogenetics with BEAUti and the BEAST 1.7. Mol. Biol. Evol. 29:1969-1973. http://dx.doi.org/10.1093/molbev/mss075. 6. Tamura K, Peterson D, Peterson N, Stecher G, Nei M, Kumar S. 2011. MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Mol. Biol. Evol. 28:2731–2739. http://dx.doi.org/10.1093/molbev/msr121.
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Downloaded from http://genomea.asm.org/ on October 6, 2015 by guest
Received 3 May 2014 Accepted 14 May 2014 Published 5 June 2014
