4. Isolation, identification and molecular characterization of virulent avian infectious bronchitis virus in Khyber Pakhtunkhwa, Pakistan
Abstract
Infectious bronchitis virus (IBV) of the Corona viridae family is a single stranded RNA virus; a causative agent of respiratory and urogenital tract diseases in chickens. This pathogen causes huge economic losses throughout the world where chickens are produced for commercial purposes. Embryonated egg inoculation was used for detection and amplification of IBV RNA, (specific pathogen free) SPF eggs (n=50, 9-11 days old) were inoculated with IBV suspected samples. After incubating the eggs for 72-144hours, the infectious allantoic fluids were harvested when signs of curling, stunting, dwarfing and death of the embryos appeared after inoculation. Viral RNA used in the RT-PCR was extracted from virus containing allantoic fluid and reverse transcribed to cDNA for detection of IBV and their serotypes. PCR products were electrophoresed to visualize the results. A high percentage (36/50eggs) of the eggs (P<0.05) were found infected with the virus in embryonic cells. The main necrotic lesions caused by the IBV in eggs were curling, stunting, dwarfing and mortality. First round PCR showed 75% samples were positive for IBV, whereas second round PCR confirmed 77.77% samples were positive for the Massachusetts serotype. In summary, despite of vaccination against IBV, this viral pathogen still circulates in Khyber Pukhtunkhwa. In addition, Massachusetts serotype is emerging with new variants leading to vaccine failure.
Keywords: Curling; Embryonated eggs; Infectious bronchitis disease; Massachusetts; Poultry; RT-PCR
