宏基因组法研究不同堆肥处理下猪粪微生物动态结构特征

Study on the Characteristics of Microbial Diversity in Swine Manure of Different Treatments During Fermentation by Metagemomics Sequencing

试验旨在应用宏基因组测序技术探究两种堆肥处理方式下猪粪中微生物的结构组成与分布特征。试验采集同一粪池中的新鲜猪粪,随机分成添加有效微生物(EM)菌的F组和未添加EM菌的K组,每组粪堆各1堆,堆成圆锥形粪堆,粪堆高1.2 m,底部直径2 m;采集堆肥第0、7、14和21天的粪便样本并提取粪便总DNA,运用宏基因组高通量测序技术,比较F组和K组的微生物多样性(组成及结构)差异,并对功能基因进行注释。结果表明,猪粪便中的细菌共测序获得1083607个有效开放阅读框(ORFs);共鉴定出147个门、118个纲、258个目、593个科、2343个属和13193个种;在门水平上,相对丰度前十的细菌界优势菌门有:变形菌门(Proteobacteria)、拟杆菌门(Bacteroidetes)、放线菌门(Actinobacteria)、厚壁菌门(Firmicutes)、疣微菌门(Verrucomicrobia)、绿弯菌门(Chloroflexi)、柔壁菌门(Tenericutes)、浮霉菌门(Planctomycetes)、螺旋体门(Spirochaetes)和酸杆菌门(Acidobacteria);在属水平上,两组平均丰度值排名前三十五的属分别属于变形菌门、拟杆菌门、厚壁菌门、放线菌门和疣微菌门;通过KEGG功能预测分析,新陈代谢通路是最丰富的,氨基酸代谢通路和碳水化合物代谢通路是包含功能基因数量最多的二级通路。两组的菌群结构多样性差异表明添加EM菌后会促进堆肥前期的菌群变化。

To study the characteristic and composition of swine manure of two treatments during composting using metagenomic technology.Fecal samples were collected from the same pig tank and randomly divided into two treatments after stirring,such as F group which was added to EM bacteria and K group without EM bacteria.Each treatment was made into a conical pile with a height of 1.2 m and a diameter of 2 m at the bottom.The samples were collected at 0,7,14 and 21 days during composting of two treatments and used to extract genomic DNA.Fecal microbial diversity was investigated by metagenomic sequencing technology.The results showed that in total 1083607 ORFs were obtained after sequencing,and the bacteria in the swine manure included 147 phyla,118 classes,258 orders,593 families,2343 genera,and 13193 species.At phyla level,the relative abundance of the top 10 bacterias include that Proteobacteria,Bacteroidetes,Actinobacteria,Firmicutes,Verrucomicrobia,Chloroflexi,Tenericutes,Planctomycetes,Spirochaetes,and Acidobacteria.The top 35 bacterias of the two treatments at genera level with the relative abundance were classified into 5 phylas,respectively belong to Proteobacteria,Bacteroidetes,Firmicutes,Actinobacteria and Verrucomicrobia.The KEGG functional prediction showed that an abundance of functional genes such as those involved in amino acid transport and carbohydrate metabolism pathway.The differences in the structure of two groups showed that the addition of EM bacteria would promote the changes of the flora at the early stage of composting.