Reserch On The Infectivity Of Bacillus Licheniformis’ Phage And Mutagenesis Of Anti-phage Strain

Bacillus licheniformis is one of the common probiotics in feed. The phenomenon of phage infection is very common in bacillus licheniformis fermentation production. In this paper, the bacillus licheniformis phage traits and mutation screening of phage resistant strain has been studied. The results are as follows:(1)Through separation and purification,two kinds of phage are got from Bacillus licheniformis bacteria powder, named DB01and DB02. The results show that both kinds of phage cannot infect bacillus subtilis. Some bacillus licheniformis is more sensitive on these two kinds of phage, the phage concentration in the liquid cultivated solution is up to109pfu/mL. Infection ability of two kinds of phage on the bacillus licheniformis of YCB,ZCS,TC is weak and specificity is strong.(2)Then the distribution and the killing method of the production environment of phage are studied. The average content of soil, plant air, dust in the Bacillus subtilis phage respectively reached1.7×105pfu/g,2.9×103pfu/g,6×104pfu/g, and species are rich. Heat treatment at100℃for5minutes cannot completely kill the plaque, so extending killing time is necessary.1.4%bleaching powder can completely kill phage in laboratory, and the factory uses2%bleach disinfection, a few phage can still survive.(3)Effect of temperature of the phage on Bacillus licheniformis is studied. Bacteria grow better, number of phage is fewer. When phage pollution is serious, the number of bacteria can only be maintained at around5billion. From March to July, the average temperature of Beihai is from19℃to34℃, and the average temperature of28℃is the critical point, count above the critical point can be maintained more than100billion.(4)The UV lamp methylene guanidine compound mutation conditions are stidied. When UV:20w UV lamp irradiation distance,40cm,3min, put the newspaper over stored11h, lethal rate at87%; methylene guanidine:20ug/mL,20min, lethal rate at98%, in4℃refrigerator overnight. After mutagenesis, Bacillus licheniformis strain resistance was 3.75.(5)According to the characteristics of the resistant strains which are prone to degradation, method which phage with resistant strains are cultivated to maintain its resistance is studied. Using103’75pfu/mL phage concentration, bacillus licheniformis grows relatively slowly, the growth cycle extended1～2h. The test uses10325pfu/mL phage concentration antagonistic strains to preserve and expand the subculture, then in the condition of solid-state fermentation under the coexistence of phage, bacterium number reached44billion/g. When comparison with resistant strains in the absence of phage preservation and subculture and phage coexistence solid state culture, count is4.8billion/g only, showing that resistant strains with phage preservation, activation, and expand training have a positive role in remaining resistant to bacterial strain.