Community Structure And Diversity Of Anammox Bacteria In Cow Manure Compost

As a effective method to deal with waste, composting technology solved the problem that livestock manure directly discharged into the environment causing pollution to the environment, and achieved the purpose of resources rational utilization. The content of nitrogen was an important indicator of the quality of compost, so nitrogen transformation was a very important part in the composting process. Anaerobic ammonium oxidation is a process of transforming NH4+-N and NO2--N into N2 under anaerobic conditions, is of a part of the nitrogen transformation. Anaerobic ammonium oxidizing bacteria existed in a variety of environments. In theory, the existence of anaerobic ammonium oxidizing bacteria in the process of composting is possible, but there were few reports about the anammox bacteria in composting. This study used self-made compost fermentation tank to simulate the rapid composting process and analyzed anammox bacteria in cow manure and rice straw composting. The physicochemical data of the samples were test, and the clone library, polymerase chain reaction denaturing gradient gel electrophoresis(PCR-DGGE), quantitative PCR method were used to detect the changes of community structure, diversity and abundance of anaerobic ammonium oxidizing bacteria in composting process. This study will further enrich the theory of nitrogen cycle, and provide the theoretica l basis for the wide distribution of anaerobic ammonium oxidizing bacteria. The results were shown as follows:(1) The composting persisted for 30 days, in the 3th day the compost entered the thermophilic period(>45?), in the 5th day the compost reached the highest temperature of 5 8?, and the thermophilic period totally lasted 8 d(3 d-10 d). The p H value was between 8.0-9.0. The moisture content of compost was gradually decreased, especially the day of 4-7 and 13-23, the value were reduced to 59.7% at the day 29. The content of NH4+-N and NO3--N were shown the opposite trend, NH4+-N reached the peak in the thermophilic period, then decreased gradually with the composting process; NO3--N increased slowly in the early stage of compost, the rapid growth were found after thermophilic period. In the composting process the C/N showed a downward trend, in the day 29 the C/N of compost decreased to 18.3, which was regarded as the maturity standards of compost; the seed germination rate index(GI) of compost showed an upward trend with the increase of compost time, in the 29 th days(maturation period) the GI had reached more than 85%, which could be thought the compost material have been thoroughly decomposed.(2) Clone library method was applied to detect anammox bacteria in composting, 16 S r RNA gene and hzo gene clone libraries with high coverage rate and stable rarefaction curve indicated that these libraries could basically represent the anammox bacteria gene information under compost environment. 16 S r RNA gene and hzo gene clone libraries analysis showed that higher anammox bacteria diversity were found in thermophilic and cooling period, anammox bacteria phylogenetic analysis in compost showed that their sequences were similar to Candidatus Kuenenia, Candidatus Scalindua, Candidatus Brocadia. Cluster analysis showed that the community structure were more similar when the composting temperature or composting time of sample were closed, and the community structure of anammox bacteria in compost was constantly changing.(3) PCR-DGGE technology was applied to analyze the anammox bacteria 16 S r RNA gene in the process of composting. The results showed that the dominant bands were found in each sample. According to cluster analysis in UPGMA method that the DGGE bands for primers Pla46f/Amx368 r, anammox bacterial community similarity between 0.54-0.86 in each sample, while the anammox bacterial community similarity were between 0.35-0.84 with the application of the primers Amx368f/Amx820 r. The principal component analysis of DGGE bands showed that the community structure of anammox bacteria in compost were always changing. After the dominant bands recycled and sequenced, the phylogenetic evolution tree were constructed, the results were consistence with results of clone library, which were mainly similar to Candidatus Kuenenia, Candidatus Scalindua, Candidatus Brocadia three genera of anammox bacteria, in the thermophilic period the dominant anammox bacteria were similar to Candidatus Brocadia. A comprehensive comparison the diversity results of two pairs primers, the higher diversity were sample Day 7(thermophilic period) and Day 13(cooling period), the lower diversity was sample Day 29(maturation period).(4) The q PCR results of anammox bacteria 16 S r RNA gene showed that copy number between 2.13x105 to 1.15x106 copies per gram dry sample in each compost sample. The sample with maximum copy number was Day 23, while the sample with minimum number was Day 4.