title: Comparison of a nucleoprotein gene based RT-PCR with real time RT-PCR for diagnosis of avian influenza in clinical specimens.
abstract: A nucleoprotein (NP) gene based reverse transcription polymerase chain reaction (npRT-PCR) assay was developed in our laboratory which could detect 35.09% of the experimental samples negative for virus isolation in first passage but positive by third passage. Reducing the reaction volume to 12.5 μl did not alter the test sensitivity and the results did not vary when duplicate samples were run in a different thermal cycler. The positive and negative agreements of this test in clinical specimens were compared with a matrix gene based real time RT-PCR with virus isolation as standard. A total of 516 clinical specimens including tissues, swabs and feces submitted from various States of India as part of active surveillance for avian influenza were tested by npRT-PCR, RRT-PCR and virus isolation in 9-11 day old embryonated specific pathogen free chicken eggs. The positive and negative agreements of npRT-PCR with virus isolation were found to be 0.909±0.022 and 0.980±0.004 respectively and that of RRT-PCR with virus isolation were 0.902±0.023 and 0.977±0.005 respectively. Since the positive and negative agreements of both npRT-PCR and RRT-PCR tests were similar, we suggest that this test can be used by peripheral veterinary laboratories that do not have real time PCR facility for active surveillance of AIV.
authors: High Security Animal Disease Laboratory, Indian Veterinary Research Institute, Anand Nagar, Bhopal 462 021, Madhya Pradesh, India. nagavetbio@gmail.com
authors: S
authors: S
authors: Nagarajan
authors: H V
authors: HV
authors: Murugkar
authors: C
authors: C
authors: Tosh
authors: P
authors: P
authors: Behera
authors: R
authors: R
authors: Khandia
authors: R
authors: R
authors: Jain
authors: M
authors: M
authors: Katare
authors: Z
authors: Z
authors: Syed
authors: S
authors: S
authors: Tripati
authors: S C
authors: SC
authors: Dubey
mesh_terms: D000818:Animals; D000835:Animals, Wild / Q000821:virology; D001717:Birds / Q000821:virology; D005814:Genes, Viral / Q000235:genetics*; D053124:Influenza A Virus, H5N1 Subtype / Q000235:genetics*; D057920:Influenza A Virus, H7N3 Subtype / Q000235:genetics*; D053127:Influenza A Virus, H9N2 Subtype / Q000235:genetics*; D005585:Influenza in Birds / Q000175:diagnosis*; D009698:Nucleoproteins / Q000235:genetics*; D011159:Population Surveillance / Q000379:methods; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary*; D014764:Viral Proteins / Q000235:genetics
chemical_list: D009698:Nucleoproteins; D014764:Viral Proteins