The Analysis Of App Composting Microbial Community Structure And Species Characteristics By PCR-DGGE

With the rapid development of intensive farming, but also caused serious pollution from manure、sewage and dead livestock debris. With respect to animal manure pollution of the environment, dead livestock due to carry large amounts of pathogenic microorganisms, if not promptly treated, humans and animals will be more dangerous. According to statistics, livestock and poultry of the farms in the mortality rate between 5% ~10%, while the mortality rate of pigs between 8% ~12%, in case of major outbreaks, the mortality rate will rise. Therefore, the treatment and disposal of the dead livestock is the solution to environmental pollution, the only way to achieve healthy development of animal husbandry.Composting is a piled biodegradable organic matter through aerobic microbial decomposition into a stable, soil-like humus process. The compost of dead pigs compared to conventional processing methods such as landfill, incineration, high temperature boiling, etc., with more safety and environmental advantages. The material transformation of composting due to the metabolic activity of microbial composting system to complete. Therefore, to fully understand the dynamic change and flora species of App composting system, tap the advantaged bacterium of App compost system, lay the foundation for develop the microbial agents of App composting, promoting composting process and shorten the composting cycle.This study was carried out natural fermentation test of the cycle of six months for the compost of corpse of dead pigs, detected the different stages of the composting process physical and chemical parameters, using PCR-DGGE technique analyzes the different types of microbial of the composting process stages and constructed phylogenetic tree. Separated and selected five dominant bacteria that can live with high temperature, and by physiological and biochemical experiments and 16 s r RNA sequencing of these five dominant bacteria were preliminarily identified. The main findings are as follows:(1)Real-time monitoring of the temperature changes of different stages of different heights App compost, the nitrogen content 、moisture content, p H、organic matter content、the effective number of viable cells、 roundworm mortality、heavy metal content and the number of Escherichia coli to be measured. The changes of pile temperature in the composting experienced three stages that is typical warming period, the high temperature period, cooling period, and the temperature of the pile with Upper layer、Middle and a scowl is basically the same, the first three months is the organic matter degradation stage, microbial activity is frequent, middle temperature near 70 ℃; p H changes follow the first drop- and then rise- and then the process of decline trend, The all soft tissue of the bodies of dead pigs is degraded, the stacking is mature, the appearance is black particles and like humus, can see the bones and teeth, but these refractory materials are brittle and easily crumbled. The Nitrogen content is about 5.3%, the organic matter content is 53%, E. coli values below 100 / g, the kill rate of ascaris close to 100%, the effective number of viable cells close to 0.35 million / g. around the pile live with no maggot pupae and newly emerged adult flies, achieved the regulations of 《stool sanitary standards(GB7959-87)》regulations.(2)Extracted microbial genomic DNA, completed the amplification of 16 s r RNA V6 ~ V8 region fragment. By denaturing gradient gel electrophoresis, obtained the DGGE profiles of 18 samples. Through analysis 18 samples by Quantity One software, the richness of various periods is relatively high. The distribution of bacteria in first three months is messy, The distribution of the bacteria in last three months is stabilized. The pathogens exist In first three months and they are killed in last three months. After Gel Extraction of the 45 bands have been sequenced, and compared with the known sequence in the NCBI, get the detailed results of these 45 sequences and completed their phylogenetic analysis. From the results, the composting process exist a lot of unknown bacteria and their role in the composting process has yet to be explored, Bacillus subtilis、Bacillus lincheniformis 、Bacillus thuringiensis and Virgibacillus play a key role in the compost.(3)Traditional culture combined with high temperature treatment, isolated the five dominant bacteria. By physiological and biochemical identification, microscopic morphology, 16 s r RNA sequencing. The results show, The similarity beteen J1 that is unknown bacteria and Uncultured bacterium clone(JF155193) is 98%, J2, J3 and J5 is Bacillus subtillis strain, J4 is Bacillus thuringiensis strain. Amylase activity from strong to weak was J4, J1, J2, J3, J5, protease activity from strong to weak was J1, J5, J2, J4, J3, lipase activity from strong to weak was J3, J2, J5, J1 and J4 don’t produce lipase.