Effects Of Microbial Inoculum On Microbial Diversity During Composting Process Analyzed By PCR-DGGE

Livestock and poultry manure contains a large number of microorganisms, including pathogenic microorganisms. If livestock and poultry manure is discharged directly to the environment, it can cause the pollution to the environment. The best way to solve the intensive, large-scale processing method is the composting settlement with manure compost. Composting raw material contains a large number of lignocellulose which is difficult to degrade. A widely used method is compost with exogenous inoculum to solve the compost the refractory organics. In this study, we use the exogenous microbial inoculum which can degrade the lignocellulose to make composting piles. Research on how the exogenous microbial inoculum affected the physicochemical properties and degradation of lignocellulose during the composting process determined by the physicochemical index, rate of lignocellulosic degradation and lignocellulose enzyme activities. Moreover, bacteria, fungi and actinobacteria were tested by the method of polymerase chain reaction-denaturing gradient gel electrophoresis(PCR-DGGE) with the universal primers to investigate the effects of inoculum on variations of microbial community. The results of the study was as follows:1. Two settlements with exogenous microbial inoculum and without inoculum reached the thermophilic stage(>45℃) at the second day of composting process. Compost with inoculum reached the highest temperature of 72℃ at the third day, higher than that of blank control compost(55℃). While inoculated compost prolonged the thermophilic stage for 21 d( 2d~22d), blank control compost only keep 15 d. Besides, the average temperature in the inoculated compost was higher than the blank control compost during the thermophilic stage. The pH value of two compost maintained between 7.8 and 9.0,and pH value of inoculated compost was slightly lower than that of blank control compost. The results showed inoculum make pH value decrease. The moisture content continued decreasing during the whole composting process, and rate of moisture content decreased most quickly at the twelfth day. When compost was completely manured, the moisture content in inoculated and blank control compost was42%, 48%, respectively. C/N in different settlements was described with the “downward” trend. C/N of inoculated compost was 19.5 in the 31 th day which reached the maturity of composting standard,however, C/N of blank control compost reached below 20 in the 46 th day. Thus, compost with inoculum can improve high temperature and prolong the thermophilic stages, and also can promote the maturity of compost.2. Lignocellulose enzyme activities in inoculated compost was higher than blank control compostduring the whole composting process, especially the maximum value of carboxymethyl cellulose enzyme(CMCase), filter paper cellulase and hemicellulase were 1120 U·L-1, 268 U·L-1, 1681U·L-1,respectively. Compared with blank control compost with the enzyme maximum value of 959U·L-1,209 U·L-1, 1459 U·L-1. At the same time, rate of lignocellulose degradation varied with lignocellulose enzyme activities changes. Except lignin, the other two kinds of lignocellulose degradation rate and enzyme activity decreases with the increase of pH value during composting process. In the 12 th day, pH value(7.5) was the minimum value, and the rate of cellulose and hemicellulose get the maximum value(9.07%, 24%). Although rate of lignin degradation and lignin degrading enzyme(Laccase) has twice top value during the composting process, microbial inoculum has a strong effect on lignin during the lignocellulose degradation and improve the rate of lignocellulose degradation and lignocellulose enzyme activities totally.3. Varies of bacteria, fungi and actinobacteria which changed by microbial inoculum were analyzed by the PCR-DGGE technology. The results showed that the DGGE bands had a obvious dynamic change. A series of dominant bands emerged in the thermophilic stage and manure stage. UPGMA cluster analysis of DGGE profiles showed that the varies of microbial community structure and diversity is bacteria the best, actinobacteria the better, fungi the least. Similarity of bacterial, fungal and actinobacterial sample is 0.5~0.8, 0.14~0.84, 0.5~0.9, respectively. With the DGGE model analysis,composition analysis and dimensionality reduction to extract two factors showed that inoculated compost clustered differently from blank control compost during thermophilic stage which has significant values. Taking comprehensive consideration, temperature and inoculum plays a screening role in microbial succession. Especially, it was stronger in fungal community than in bacterial community, which is better than in actinobacterial community.The results of dominant bands sequences showed that exogenous microbial inoculumn can successfully colonized during the composting. In the thermophilic stage, the main advantage of bacterial strains can be divided into 2 categories: the genus Bacillus of Firmicutes and alpha-proteobacteria, beta Proteobacteria, gamma Proteobacteria of Proteobacteria; Fungi in composting mainly concentrated in 2 categories, including Ascomycota and Basidiomycota, during the thermophiclic stage the dominant fungal strains belong to Aspergillus and Trichocladium of Ascomycota, and the white rot fungi(Heterobasidion) of Basidiomycota.Actinobacterial communities can be divided into 3 categories: Nocardiopsis sp., Streptomyces sp. and Arthrobacter sp.. Detection of thermophilic stage of three kinds of microbes in the dominant strains for the majority of lignocellulose degrading bacteria, therefore, addition of inoculum can increase the number and types of lignocellulose degrading microbes.