Study On Two Strains Of Pseudomonas Aeruginosa Source Forest Musk Deer Biological Characterisitcs And Extracellular Products Characterisitcs

Forest musk deer is a suppurative pneumonia by the Pseudomonas aeruginosa (Pseudomonas aeruginosa, PA) infection caused by difficulty in breathing, nostrils pus out as the main feature of chronic disease. This experiment was conducted to study the forest musk deer suffering from suppurative disease in vivo isolated two Pseudomonas aeruginosa (SP-N and SP-Q) alone and in 1:1 co-culture culture character case and its extracellular products the physical and chemical properties and biological properties, to provide a theoretical basis for the clinical diagnosis and treatment and prevention of disease.For SP-N and SP-Q for routine bacteriological identification and 16S rRNA identification, were found in two gram-negative bacteria, including SP-N as long bacteria, does not produce green pigment; SP-Q Brevibacterium, produce green pigment. By 16S rRNA sequencing and target fragments with NCBI comparison and found SP-N and SP-Q and P.Aeruginosa homology above 99%. Serotyping display, SP-N G type Pseudomonas aeruginosa, SP-Q type B P. aeruginosa.SP-N, SP-Q and SP-N and SP-Q (1:1) at 37 ℃ ordinary broth culture, its consistent growth curve. When cultured alone, SP-N highest number of viable cells at 14 h, in order to 5.6×10~10CFU/mL; SP-Q highest number of viable cells at 12 h, in order to 1.4×10~10 CFU/mL; 1:1 ratio of co-culture when, at 12h when the highest number of viable cells, was 1.0×10~10 CFU/mL. With 12 h of SP-N and SP-Q separate culture solution and a 1:1 co-cultured SP-N and SP-Q medium vaccinated mice, the median lethal dose was 5.05×106 CFU/mL,5.22 ×108 CFU/mL and 3.57x 108 CFU/mL, and the median lethal dose difference was significant (P<0.05), described the culture medium of mouse SP-N has a strong pathogenicity, but not co-culture enhanced pathogenicity.Flat glass sheet by extraction method of P. aeruginosa SP-N and SP-Q extracellular product (extracellular products, ECP), measured ECP SP-N protein concentration was 0.8mg/mL, ECP protein concentration of SP-Q of 0.36 mg/mL. Carried out by SDS-PAGE of ECP protein analysis we found that SP-N and SP-Q’s ECP protein bands are basically the same, there are seven protein bands whose molecular weights were 19 kDa,23 kDa,37 kDa,42 kDa,52 kDa,59 kDa and 66 kDa. Prepared with mouse anti-SP-N serum ECP 2 strains were blot hybridization showed that SP-N of ECP immunoreactive bands are mainly 2, molecular weight of about 42 kDa and 66 kDa. SP-Q immunoreactive bands of ECP mainly a molecular weight of about 42 kDa.Using the plate method for the analysis of protein activity ECP found SP-N and SP-Q have the ECP protease, gelatin urease activity and hemolytic toxicity; no lecithin, lipase, amylase activity and DNA. Through the solid and liquid culture observation shows, SP-N does not produce pyocyanin, SP-Q generates pyocyanin. The SP-N and SP-Q culture solution was extracted with chloroform, spectrophotometrically measured obtaining pyocyanin contents were 0 and 12.9 mg/ mL.ECP pathogenic test mice showed, SP-N and SP-Q for the ECP mouse LD50 are 2.12 mg/kg and 8.73 mg/kg.