Isolation And Identification Of Haemophilus Parasuis In Parts Of Sichuan Province

Haemophilus parasuis(HPs) is the pathogenic bacteria of Glasser’s Disease, which mainly causes serious fibrinous polyserositis, arthritis and meningitis of weaned pigs and leads to heavy economic losses. In this study,55 clinical samples were collected from 13 counties of 8 districts of Sichuan Province, then researches on isolation and identification as well as biological characteristics of HPs were done. The main results were as follows:1. Bacterial isolation and identification of HPsFrom June to November in 2013,55 suspected clinical samples were collected from 13 counties of 8 districts of Sichuan Province. A total of 14 strains of HPs were successfully isolated, among which 6 from Nanchong,4 from Guangan,2 from Yaan and 2 from Leshan, all the isolates reacted positively in Gram staining, biochemical identification, PCR identification. All isolates were pleomorphic after Gram staining and showed consistent biochemical reaction, no hemolysis and required NAD for growth. Gel diffusion test (GD) was used to serotyp these isolates,3 isolates were serovar 5 and 1 isolate was serovar 13. It was showed that the 14 isolates were quite sensitive to florfenicol and doxycycline in drug sensitivity test.2. The biological characteristics research on HPsIn this research, culture medium of bacteria isolation was screened and optimized, it was found that TSA solid culture medium was the most suitable one for HPs growth, on which HPs could be grown up 20h after inoculation. Concentration of NAD and calf serum for growth effect of HPs was optimized as well, it was shown that 10μg/ml NAD and 6% calf serum in TSB could make HPs grow best. It was shown in determination of growth curve and viable count that the peak growth of HPs was detected after 12h culturing in TSB medium. Different isolates with same serotype were inoculated to Kunming mice and the lethal dose(LD50) was determined in order to compare the virulence of these isolates, the result showed that the virulence of the strains isolated from the same farm had no difference and the strains isolated from different regions had certain differences.