Molecular Study of Virulence Factors of Avian Isolates of Pasteurella multocida and Survey of Acute Isolates Lethality in Embryonic egg and Pullet

Present study was designed in order to molecular confirmation, capsular typing and detection of 12 important virulence factors include ompH, oma87, sodA, sodC, hgbA, hgbB, exBD-tonB, nanB, nanH, ptfA, pfhA and toxA in 18 Pasteurella multocida avian isolates from different areas of Iran with a strain of the organism was isolated from the epidemic pasteurellosis in the Northern region, and also survey of acute isolates lethality in poultry embryos and pullet. Molecular confirmation of tested isolates was performed using specific primers of kmt1. As well, capsular typing and detection of virulence factors of isolates were performed by application of multiplex PCR method and using specific primers of capsule biosynthesis genes and genes encoding virulence factors of organism. All isolates were confirmed molecularly but based on the results of capsular typing, 16 (88.8%) isolates with strain isolated from epidemic pasteurellosis were identified as type A and 2 (11.1%) as untypeable isolates. Detection of virulence genes showed that all studied isolates has six virulence genes of sodC, hgbA, hgbB, nanB, nanH and ptfA. The oma87, exBD-tonB, ompH, sodA and pfhA genes were respectively present in 94.4, 94.4, 83.3, 66.6 and 27.7% of the isolates, but there was not any toxA positive isolates. To determine the chicken embryo lethal dose 50 (CELD50) and lethal dose 50 (LD50) of the isolates, different dilutions of five acute isolates containing the majority of virulence genes as well as strain isolated from epidemic pasteurellosis were respectively inoculated to embryonated chicken eggs and lying pullets. Based on the results of CELD50, though all six isolates were able to waste the poultry embryos, but strain isolated from epidemic pasteurellosis was recognized as the most acute isolate. Results of LD50 test showed that none of the five acute isolates were able to waste the studied pullets while the LD50 of strain isolated from epidemic pasteurellosis was determined 5 × 101 colony forming unit per bird (cfu/case), and this strain was identified as the most acute understudy isolate. The findings of this study showed that, despite the presence of the majority of virulence genes in P. multocida avian isolates in Iran, and that the most acute isolates were able to waste the poultry embryos but they were not infectious to lying pullets, and virulence of strains isolated from birds with hyperacute form of the disease is far higher. Our data could be useful in the design of new and efficient vaccines that would be acceptable protection in animal populations.