| Digitaria sanguinalis is a devastating weed widely distributed in the world. So far, the control of D. in the field was mostly used chemical herbicides. However, the environmental pollution and drug resistance are rising with the application of chemical herbicides. Because of the advantages of environment-friendly and objective to the weeds specially, the biological control has become a research hotspot. The biocontrol strain Col-68(Colletotrichum hanaui) was isolated from the diseased leaves of D. The preliminary studies have shown that Col-68has a strong pathogenicity to D, and meanwhile it was no harm on the main crops and lawn grass, such as rice, cotton, corn, soybeans, wheat, peanut and rye grass. Biological characteristics of Col-68have been investigated before, and its characteristics genetic improvement has also been carried out. A major constraint in the development of this biocontrol strain to bio-herbicide is the inefficiency of inoculum production. The objective of this study was to explore submerged fermentation for mass production of Col-68.Optimization of four parameters, including initial total sugar concentration, carbon source, nitrogen source and carbon-nitrogen ratio (C/N), was surveyed by using single factor experiment for the optimal production of sporulation of biocontrol strain Col-68and the increasing activity of fermentation filtrate against to D. root growth. The optimal fermentation medium was as follows:The optimal carbon source was surcose, the optimal nitrogen source was nitrate nitrogen, carbon-nitrogen ratio was2.5:1, and an initial total sugar concentration between of30～50g·L-1. The suitable fermentation conditions were as follows:initial pH7.0, temperature28℃, medium volume120mL/300mL, inoculum volume5%(v/v), for10day. In such optimal nutrition and environmental conditions, the maximum sporulation and herbicidal activity of fermentation filtrate of were obtained. The results also showed that the inhibition of fermentation filtrate of the strain Col-68to D. root growth was significantly higher than the sprout.Nowadays the chemical herbicides cannot be a substitute for bio-herbicides to control weeds directly. However it is very meaningful to find several pathogens mixed with chemical herbicides which can enhance for control of weeds. Some study found that mixtures with lower doses of chemical herbicides can be effectively weaken weed defense responses, with way also can reduce the use of herbicides. In our study we selecting different concentrations of several chemical herbicides include of atrazine, haloxyfop, and Paraquat, and mix with strains of Col-68concentration of106cfu·mL-1. We found that lower concentrations of atrazine and haloxyfop could enhance the growth of strain and increase spore germination of Col-68. And the growth inhibition of strains gradually decreased with the reducing of the herbicide concentration. When the concentrations of haloxyfop lower than15mg·mL-1and atrazine concentration is lower than100mg·mL-1, the inhibition of strains growth was almost disappeared. But the inhibition of Paraquat to the growth of strain and spore germination of Col-68is higher than the other two herbicides. Col-68conidia (106cfu·mL-1) mixed with160mg·mL-1atrazine or12mg·mL-1haloxyfop, the weed control effect was significantly higher than each one, respectivelly.This study led to the development of a protocol for production of Col-68spores using submerged fermentation. Inoculum produced with this protocol can be used for laboratory, greenhouse, and field trials in development of the bio-herbicide. |
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