Molecular Detection and Sequencing for S1 Glycoprotein Gene of Bronchitis Virus of 2016 Epidemic from Sindh and Punjab  

作　　者：Ahmad Umer Sultan Muhammad Danish Mehmood Rameez Hassan Huma Anwar Sana Noreen Faisal Amin Sajjad Hussain 

机构地区：[1]Ottoman Pharma (Immuno Division), Lahore, Pakistan [2]Institute of Molecular Biology and Biotechnology, University of Lahore, Lahore, Pakistan [3]Grand Parent Diagnostic Laboratory, Lahore, Pakistan [4]Veterinary Research Institute, Lahore, Pakistan

出　　处：《Advances in Bioscience and Biotechnology》2018年第12期649-660,共12页生命科学与技术进展（英文）

摘　　要：Infectious Bronchitis (IB) is highly contagious disease of commercial poultry causing substantial economic loses by producing poor quality meat in broilers and effecting production in breeder birds. The causative agent has been reported as most hazardous pathogen among other infectious agent even after being immunized with multi-variant strain vaccine. Currently, different strain such as H-120, 4/91 and D274 have been used extensively for immunoprophylaxis against velogenic strain across Pakistan with minimal protection reported. In current study PCR analysis was used to investigate the molecular nature of IB isolates from Punjab and Sind province of Pakistan in 2016 epidemics. Total of 100 tracheal samples were considered for virus inoculation in 10 days old chicken embryonated eggs. The IBV infected amniotic fluid was neutralized with monoclonal antisera of H-120, 4/91 and D274 strains. The IBV screened samples were subjected for RNA extraction and subsequent to PCR using type specific primer of each strain. The amplified product of 840 bp was sequenced through Sanger sequencing. On the basis of PCR results, four similar amplified products from both regions were obtained showing similarities in agarose gel electrophoresis, but they differ from each other on the basis of nucleotides sequence. Phylogenetic analysis revealed that nucleotide sequences of isolates from Karachi were similar to the IBV H-120, Mass-41 and Connecticut 46 reference strains. Whereas, isolates from the Punjab province are analogous to the Mans-2, Mans-3, 9/41(UK) but did not show significant similarity with other reference strain. Therefore, it is recommended that use of M-41 and H-120 in vaccine production could be effective measure against velogenic infectious agent in Sindh particularly in Karachi, whereas, it would be better to incorporate either of the variant GQ281656.1, AY279533.1 in vaccine because of their highest level of resemblance with genetically sequenced isolates from Lahore and its surroundings.Infectious Bronchitis (IB) is highly contagious disease of commercial poultry causing substantial economic loses by producing poor quality meat in broilers and effecting production in breeder birds. The causative agent has been reported as most hazardous pathogen among other infectious agent even after being immunized with multi-variant strain vaccine. Currently, different strain such as H-120, 4/91 and D274 have been used extensively for immunoprophylaxis against velogenic strain across Pakistan with minimal protection reported. In current study PCR analysis was used to investigate the molecular nature of IB isolates from Punjab and Sind province of Pakistan in 2016 epidemics. Total of 100 tracheal samples were considered for virus inoculation in 10 days old chicken embryonated eggs. The IBV infected amniotic fluid was neutralized with monoclonal antisera of H-120, 4/91 and D274 strains. The IBV screened samples were subjected for RNA extraction and subsequent to PCR using type specific primer of each strain. The amplified product of 840 bp was sequenced through Sanger sequencing. On the basis of PCR results, four similar amplified products from both regions were obtained showing similarities in agarose gel electrophoresis, but they differ from each other on the basis of nucleotides sequence. Phylogenetic analysis revealed that nucleotide sequences of isolates from Karachi were similar to the IBV H-120, Mass-41 and Connecticut 46 reference strains. Whereas, isolates from the Punjab province are analogous to the Mans-2, Mans-3, 9/41(UK) but did not show significant similarity with other reference strain. Therefore, it is recommended that use of M-41 and H-120 in vaccine production could be effective measure against velogenic infectious agent in Sindh particularly in Karachi, whereas, it would be better to incorporate either of the variant GQ281656.1, AY279533.1 in vaccine because of their highest level of resemblance with genetically sequenced isolates from Lahore and its surroundings.

关 键 词：Infectious BRONCHITIS VIRUS S1 GLYCOPROTEIN Polymerase Chain Reaction VIRUS NEUTRALIZATION Test Molecular Evolutionary Genetics Analysis 

分 类 号：R73[医药卫生—肿瘤]