Construction Of The Compound Inoculants And Study Of Fermentation Mechanism For Cotton Stalk Silage

The abundance and nutrient value of cotton stalks in Xinjiang make them a feasible feedstock for ruminant animals.However,the cotton stalks are poor taste and fewer utilized due to the high content of lignocellulose,as well as the presence of free gossypol.To solve the aforementioned problems,the lactic acid bacteria,cellulolytic bacteria and gossypol-degradation bacteria were screened,and the fermentation processes were also optimized.Furthermore,the mechanisms of cotton silage degradation was analyzed by illumina sequencing of 16S rRNA gene.9 lactic acid bacteria,4 cellulolytic bacteria and 2 gossypol-degrading bacteria were obtained by culturable method.The lactic acid bacteria were Lactobacillus sp.,Enterococcus sp.and Pediococcus sp..Lactobacillus L2 produced acid fastest,the pH reached 3.35 after 15 hours of growth.The cellulolytic bacteria included Bacillus sp..B.acillus C3 possessed the highest CMCase of 27.89 U/mL.The gossypol-degrading bacteria were Candida sp.G3 and Saccharomyces G9.G3 could degrade 69.5%of gossypol in the cotton stalk,which was higher than G9.Bacillus C3,Enterococcus L16 and Lactobacillus L2 could also degrade free gossypol efficiently.In order to improve the synergy effect of mixed inoculants,the combination of strains and fermentation technology were improved according to pH,crude protein,dry matter,degredation rate of cellulose and free gossypol.The results showed that the best compound inoculants were Lactobacillus L2 combined with Bacillus C3?L2:C3=2:1?.Further research showed that 0.1%cellulase added with L2 and C3 could accelerate the ensiling of cotton stalk.The optimum fermentation parameters were obtained by response surface method?RSM?.The best parameters were as follows:0.5%inoculation,63%water content,7%molasses,anaerobic fermentation for 60 days.Based on this process,the pH of the silage was 4.12,the content of dry matter and crued protein were 37%and 7.16%respectively,the degradation rate of cellulose reached 21.5%.The palatability of cotton silage was improved greatly.The results of high throughput sequencing showed that Lactobacillus,Pediococcus,Enterobacteriaceae?g_unclassified?,Escherichia-Shigella,Enterobacter and Clostridium_sensu_stricto₁2 were the key genus that influence the quality of cotton stalk silage.After inoculating L2 and C3,Lactobacillus became the dominant bacteria quickly,the growth of Enterococcaceae was inhibited observably.Lactobacillus showed a negative correlation to pH and positive correlation to crude protein,while Enterococcaceae showed opposed performance to Lactobacillus.In addition,Lactobacillus and Pediococcus were proved as the most important beneficial bacteria for cotton stalk silage,while the presence of Enterococcaceae and Clostridium_sensu_stricto₁2 would cause the spoilage of feeds.After adding cellulase to the inoculants,the diversity of dominant bacterial community during ensiling were changed weakly.But the degradation rate of hemicellulose,cellulose and lignin were improved significantly and Lactobacillus reached to the maximum abundance more quickly,which might contribute to the acceleration of fermentation process by adding cellulase.