关键词: 噬菌体抗性, 枯草芽孢杆菌, gyrA基因, 流加发酵

Abstract: This work aims to screen anti-phage strain for solving the problem of phage infection in Bacillus subtilis production and increase fermentation of anti-phage strain. Double-layer-agar plating method was used to screen and purify phages with Bacillus subtilis as host from abnormal fermentation broth,and anti-phage strains were obtained by screening the Bacillus library in our laboratory using the purified anti-phages. Phylogenetic trees were constructed by 16S rDNA and gyrA gene sequences analysis to identify the anti-phages strain. Based on glucose content curve and growth curve of anti-phages strain batch fermentation,initial glucose concentration in the medium and glucose feeding strategy were optimized for improving the fermentation of the screened anti-phage strains. As results,phages S01 and S02 were screened from the abnormal fermentation broth,and a strain Bacillus sp. BS-2,which was not infected by the phages,was screened from the Bacilluslibrary in our laboratory. According to the 16S rDNA and the phylogenetic trees of gene gyrA,the strain BS-2 was characterized as Bacillus subtilis. The biomass was up to 2.43×10¹⁰ CFU/mL,while suitable initial concentration of glucose was 15 g/L,and glucose was fed into the broth at the speed of 2 g/L·h when the glucose concentration in the broth was lower than 5 g/L,and the total supplying quantity was 10 g/L. B. subtilis BS-2 shows strong ability of anti-phages and high titer under optimized fermentation conditions,which has promising potential for large-scale industrialization.

Key words: anti-phages, Bacillus subtilis, gene gyrA, fed-batch fermentation