Isolation and Molecular Identification of Avibacterium Paragallinarum in Layers by Using Sybr Green Real Time PCR

Abstract

Infectious coryza is an infectious disease, caused by Avibacterium paragallinarum, causing economic losses to commercial poultry Industry. The samples were collected from 6 commercial Infectious Coryza suspected layer farms includes, nasal swabs(119), ocular swabs(119) and tracheal swabs(119) were pooled farm wise and organ wise. On o selective media the organism was produced satellite colonies after incubation of 48 hours at 37 c under anaerobic conditions. The organisms were showed positive test for sugar fermentation tests, negative for catalase test,Indole test and H2S production test. Further molecular confirmation was done by SYBR Green real time PCR by targeting the HPG2 gene. In this study the Ct value below 35 was consider as positive cutoff for Avibacterium paragallinarum. All samples were screened by targeting HPG2 gene and found 100 percent positive for Avibacterium paragallinarum. To control the economic losses associated with the Infectious coryza, an early ,accurate and rapid diagnosis is essential. So the SYBR Green real time PCR is a rapid and highly sensitive technique which can substitute conventional methods. The antibiogram pattern revealed that the isolated pathogens were showed the sensitivity to enrofloxacin,ceftriaxone and amikacin. The diagnosis can be done by isolation of the pathogen on selective media and identification by biochemical tests, but less sensitive and time consuming. So the molecular detection based PCR tests are the most advanced, among the PCR tests, SYBR Green real time PCR test is the rapid, accurate and most sensitive test in detection of pathogens. Hence the present study was aimed that isolation and molecular detection of Avibacterium paragallinarum in commercial layers by using SYBR Green real time PCR by targeting HPG2 gene.