Mutation Breeding And Fermentation Technology Of Spinosyns-Producing Strain

Spinosyns are novel macrolide insecticides produced by fermentation of Saccharopolyspora spinosa. The most active components of the spinosyn family are spinosyn A and spinosyn D, and others are significantly less active. Spinosad, a mixture of spinosyn A and spinosyn D, is highly effective against insect pests, such as lepidopterans and dipterans. The spinosad possesses a low toxicity for no-target insects, mammals and plants, as well as an excellent compatibility for environment. Therefore, the development of spinosad will give rise to a stimulatory effect on economic and social benefit. The aim of this study is to investigate the strain improvement by mutation breeding and optimize the spinosyn fermentation process.Ultraviolet, a mutagen, combined with antibiotics was applied to screen resistant mutants. The results showed that strain AE8-24 was obtained by this method. The AE8-24 possessed the double resistance of apramycin and erythromycin. In addition, the titer of AE8-24 was 130.7μg/ml, which was increased by 47% compared with that of the strain SP-29. According to the process of spinosyn biosynthesis, the rational selection of AE8-24 was performed to improve productive strains. The optimal strain AERD1-7 with the multi-resistance of apramycin, erythromycin, rhamnose and 2-deoxy-D-glucose, was selected. The titer of AERD1-7 reached 171.4μg/ml, 1.92 times compared with that of parental strain SP-29. Subculture test indicated that the hereditary character of AERD1-7 was stable.Then, the medium composition and fermentation conditions of AERD1-7 were investigated. The effects of carbon sources and nitrogen sources on spinosyn fermentation were studied. Uniform design method was applied to optimize the seed and fermentation medium, and the effect of precursors on spinosyn biosynthesis was evaluated. In addition, the conditions of shaking flasks' fermentation were optimized. The result indicated that the productivity of spinosad on the optimal fermentation condition reached 567.1μg/ml, which was 2.98 times as much as that on the original condition.