| Cellulase can eliminate soluble non-starch polysaccharides,hydrolyze them into polysaccharide small molecules,reduce the viscosity of intestinal nutrients,improve the digestibility and absorption rate of feed,and then improve the production performance of livestock and poultry.Therefore,in this study,a strain of Aspergillus niger with high cellulase activity was screened from fermented feed,its cellulase production conditions were optimized,and the culture of Aspergillus niger obtained by optimal fermentation parameters was added to the base diet of Kazakh sheep(♂)in the fattening stage to study the feed value of the culture.The main findings are as follows:(1)Experiment 1 Isolation and identification of feed Aspergillus niger and its cellulase activity detection.In this study,a fungal strain with high cellulase activity was isolated from fermented feed,and the strain was identified by colony morphology and ITS sequencing.Then the growth curve,qualitative analysis of enzyme production and the effects of different carbon sources on cellulase activity and xylanase activity were determined.The results showed that the strain was identified as Aspergillus niger,named Aspergillus niger 0729,and the sequencing data was uploaded to the National Microbiology Science Data Center.The optimal inoculation time of Aspergillus niger 0729 in PDB medium was 14～38 h.Enzyme-producing qualitative studies have shown that this strain produces proteases,amylase,pectinase,xylanase,lipase and phytase,and glucose oxidase.Different carbon sources,Aspergillus niger has different enzyme production capacity.When xylose was used as carbon source,the endoglucanase activity and filter paper enzyme activity of Aspergillus niger were the highest on the second day of culture,reaching 18.70 and 15.26 U / mL,respectively.As a carbon source,filter paper had the highest xylanase activity induced by Aspergillus niger on the first day of culture,reaching 19.10 U / mL.(2)Experiment 2 Optimization of solid-state fermentation conditions for cellulase production by Aspergillus niger.Through single factor experiments(carbon source,nitrogen source,ammonium sulfate addition amount,dipotassium hydrogen phosphate addition amount,magnesium sulfate addition amount,culture time,initial moisture of medium and inoculation amount),three factors that significantly affect the enzyme activity were found.Then,the response surface method was used to systematically optimize the cellulase production conditions of solid-state fermentation in order to obtain the best enzyme production conditions of Aspergillus niger on solid-state medium.Finally,Aspergillus niger culture was obtained through the best fermentation conditions.The results showed that the single factor test results showed that the best carbon source was sugar beet granule meal,the best nitrogen source was ammonium sulfate and the best addition amount was 6 %,the best addition amount of dipotassium hydrogen phosphate was3 %,the addition amount of magnesium sulfate was 0.3 %,the initial moisture of the best medium was 50 %,the best culture time was 3 d,and the best inoculation amount was 15 %.PB test showed that ammonium sulfate,culture time and initial moisture of medium had great influence on cellulase activity.The results of response surface methodology showed that the optimum amount of ammonium sulfate was 2 %,the optimum culture time was 2 d,and the optimum initial moisture of the medium was 50 %.The optimized endoglucanase activity and filter paper activity increased by 189 % and 176 %,respectively.(3)Experiment 3 Effects of Aspergillus niger culture on nutrient apparent digestibility and rumen pH of Kazakh sheep(♂)during fattening periodThe changes of nutrients before and after fermentation of Aspergillus niger culture were determined,and it was used as an additive to feed fattening Kazakh sheep to study the effect of Aspergillus niger culture on apparent nutrient digestibility and rumen pH,laying a foundation for its application in mutton sheep diet.The results showed that the dry matter content of the raw material group was significantly higher than that of the blank fermentation group and the fermentation group(P< 0.05).The crude protein content of the fermentation group was significantly higher than that of the blank fermentation group and the raw material group(P < 0.05).The true protein content of the blank fermentation group was significantly higher than that of the fermentation group and the raw material group(P < 0.05).The crude fat content of the blank fermentation group was significantly higher than that of the fermentation group and the raw material group(P < 0.05).The pH value of the blank fermentation group was significantly lower than that of the fermentation group and the raw material group(P <0.05).The crude fiber content of the fermentation group and the blank fermentation group was significantly lower than that of the raw material group(P < 0.05).The content of neutral detergent fiber in fermentation group was significantly lower than that in blank fermentation group and raw material group(P < 0.05).The content of acid detergent fiber in fermentation group was not significantly different from that in blank fermentation group,but significantly lower than that in raw material group(P < 0.05).The crude ash content of the fermentation group was significantly higher than that of the blank fermentation group and the raw material group(P < 0.05).Compared with the control group,the apparent digestibility of dry matter,crude protein,starch,neutral detergent fiber and acid detergent fiber in the experimental group were significantly increased(P < 0.05).Aspergillus niger culture can stabilize rumen pH.In summary,this study isolated a strain of Aspergillus niger 0729 with high cellulase activity.The solid-state fermentation conditions for cellulase production were optimized,the endoglucanase activity and filter paper activity were increased by 189 % and 176 %,respectively.The nutritional quality of the culture fermented by Aspergillus niger was improved and as an additive,it could improve the nutrient apparent digestibility and stabilize the rumen pH of Kazakh sheep during fattening period. |
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