Molecular Characterization of Indigenous Isolates of Avibacterium paragallinarum and Media Optimization of its Growth for Vaccinal Seed Production
Molecular Characterization of Indigenous Isolates of Avibacterium paragallinarum and Media Optimization of its Growth for Vaccinal Seed Production
Aatif Masood Ahmad Khan1, Masood Rabbani1*, Arfan Ahmad1, Muhammad Wasim2 and Sohail Raza1
ABSTRACT
The current study was conducted for isolation, identification, and rapid detection of Avibacterium paragallinarum, the causal agent of Infectious Coryza (IC), from layer chickens in various farms in Pakistan. A total of 46 isolates were obtained from a total of 244 clinical samples from 122 sick and moribund or dead layer chickens showing signs of IC like slight swelling of infra orbital sinuses and nostril mucus membranes. Two sampling sites were chosen, squeezing the nostrils for live birds and infra orbital sinus for moribund killed with later providing more live bacterial isolates. Isolation, biochemical identification, species-specific PCR tests and classical serotyping were performed. Two types of swab samples were used, one for direct PCR detection and other for the conventional diagnosis procedure. All isolates were identified as Avibacterium paragallinarum and required nicotinamide adenine dinucleotide phosphate (NAD) for their growth. The isolates were typical for Av. paragallinarum as they were not able to ferment neither galactose nor trehalose. Performing PCR of direct swab samples provided higher detection level as compared to the PCR from isolated colonies following conventional diagnostic procedure. In the current study, page serotyping demonstrated that all isolated Av. paragallinarum belonged to page serovars B. Among the media evaluated for growth of the isolated strain BHI/SN appeared to be the optimum media supporting the maximum growth rate and providing the maximum cfu count and the highest value of specific growth rate during the exponential phase of bacterial growth curve were obtained at 8 h. Thus the current study demonstrates that diagnostic approach using direct swab samples for the detection of Av. paragallinarum was found to be more accurate, rapid and sensitive than routine, culture-based PCR analysis and isolated Av. paragallinarum can be grown in BHI/SN to achieve maximum biomass, a trait required for vaccine production, using brain heart infusion broth media.
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