Screening And Co-culture Of Mixed Strains For Synergistic Decomposing Of Lignocellulose From Compost

In this thesis, the study was focus on the lignocellulolytic microorganisms in the composting. First, the cellulolytic strains were isolated from the composting, and then these strains were cocultured with lignolytic strains also isolated from the composting in agar plate. The better couples selected from agar plates were tested by the mixed culture in solid-state fermentation on straw. The compatible strains with Phanerochaete chrysosporium for biodegradation of straw were screened by adjusting cellulose agar with the addition of the extract from solid-state fermentation of Phanerochaete chrysosporium on straw. The enzyme, the degradation of lignocellulose and the microbial community structure were studied during the mixed culture.Four better cellulolytic strains were isolated from the composting by cellulose agar, cellulose-congo red agar and solid-state fermentation, they were signed D, X, P and T. The strain X was considered as the best one for the steady enzyme production and the better decomposing of semicellulose and cellulose. The peak value of its endoglucanase was 37.07 IU/g.Some combined strains for degradation of straw were screened from 6 cellulolytic and 8 ligninolytic strains stored by us. First, binary combinations of thirteen strains prepared for studying were inoculated into the nutrient agar plates by parallel streaking. By observing the colonial morphology of these strains, nine couple of strains was found that they could grow well together. Then, these couples were examined by solid-state fermentation with straw powder. After fermentation, the hemicelluloses, cellulose and lignin contents of residues were measured respectively. The results indicated the combined strains AF93252+M5 and XP+M5 were the ideal couples because these couples had higher efficiencies in mixed culture than that in pure culture. The hemicellulose content of XP+M5 was reduced to 20.6% which is 2.67 and 3.79 percent less than XP and M5 respectively. This simple and high efficiency method about screening combined strains was proved to be useful for the studying about improving the efficiency of biodegradation of straw.The compatible strains with Phanerochaete chrysosporium for biodegradation of straw were screened by adjusting cellulose agar with the addition of the extract from solid-state fermentation of Phanerochaete chrysosporium on straw. Six stably growing strains were selected by the compatibility experiments on the PDA and MEA mediums. Then we studied the enzyme and the degradation of lignocellulose with the mixed culture of the six strains and Phanerochaete chrysosporium. Three couples of fungi were considered as the optimized ones for the synergisms in producing cellulase and decomposing cellulose. The improved decomposition efficiency of lignin maybe resulted from the impact of some extracellular enzymes and some active factors of low molecular weight in the fermentation.The enzymes productions, the decomposing of lignocellulose and the microbial community were studied during the mixed culture of Phanerochaete chrysosporium and Penicillium steckii.The value and the dynamics of the extracellular lignocellulolytic enzymes productions were changed by mixed culture in the solid-state fermentation. The enzyme production was steadier in mixed culture than that in pure culture, and the peak could maintain longer. The peak value of Lac is 1.28 IU/mL and appears in the mixed culture. In the mixed culture, the LiP is 52 IU/mL which is about two times higher than that in the pure culture at the end of the fermentation.The decomposing of lignocellulose was best in the mixed culture at the end of the fermentation; the decomposing rates of hemicellulose, cellulose and lignin were 53.1%, 52.7% and 54.9%, respectively. The mixed culture may improve the decomposing by three ways, the first is improving the amount and the stability of the lignocellulolytic enzymes, the second is enriching some extracellular low-molecular-weight biochemical agents, and the third is cooperation of the two factors mentioned above.The community-level physiological profiles obtained with Biolog microplates and phospholipid fatty acid (PLFA) patterns were proper to describe the dynamic changes of microbial community. These methods could provide some useful information for analyzing the changes of enzyme production and lignocellulose decomposing. The analysis of average well color development (AWCD), Richness and Shannon-diversity indicate the microbial community has been improved in the mixed culture than that in the pure culture. The Principal components analysis (PCA) suggest the microbial community of the mixed culture is similar with that of the Phanerochaete chrysosporium at the 7^th day, whereas this community is similar with that of the Penicillium steckii at the end of fermentation. The succession of the microbial community may result in the steady enzyme production in the mixed culture.In conclusion, the decomposing rate of lignocellulose can be raised by the proper mixed strains which compatible with each other. Analyzing the interactions of mixed strains by agar plate and adding the extract from solid-state fermentation of some strain into cellulose agar can simplify the screening the compatible strains for mixed culture. These methods would be helpful for solving the problem of subjectivity about strains selecting, they could provide useful information for the researching of the complex compost inoculum based on the lignocellulolytic microorganisms. It is necessary to study the dynamic changes of microbial community for testing the feasibility of mixed culture.