双孢蘑菇堆肥过程中细菌群落结构分析

Bacterial community structure analysis for mushroom(Agaricus bisporus) compost using PCR-DGGE technique

分别以玉米秸、牛粪和稻草、牛粪为主要原料,设计两种双孢蘑菇(Agaricus bisporus)堆肥配方并进行堆肥发酵,研究二者堆肥过程中细菌群落多样性.在建堆、一次发酵结束和二次发酵结束3个时期分别采集堆肥样品,提取总DNA,以大肠杆菌16S rDNA序列设计通用引物,进行PCR-DGGE扩增和序列分析.结果获得56条特异条带16S rDNA基因信息(GenBank登录号为KF630598-KF630653),分属于细菌的7个门、42个属.两种配方堆肥过程中主要类群属于变形菌门(Proteobacteria)、厚壁菌门(Firmicutes)和放线菌门(Actinobacteria),主要的优势种属包括Bacillus属、Paenibacillus属、Thermomonospora属、Thermasporomyces属、Pseudomonas属和Cellvibrio属.多样性指数分析显示,在堆肥过程中,稻草配方微生物多样性变化呈连续上升趋势,而玉米秸配方堆肥微生物多样性呈先升高、再降低的趋势.主成分分析(PCA)显示,玉米秸配方一次发酵结束的样品与二次发酵结束的样品聚为一类,说明玉米秸配方堆肥比预期提前腐熟.本研究表明,稻草配方微生物多样性高,所需腐熟时间长;玉米秸配方微生物多样性略低,但所需腐熟时间大大缩短.

This study aimed to investigate the bacterial communities in mushroom compost using straw, corn stalks, and cow dung as major compost matrix. Bacterial communities of samples at the beginning of composting, at the end of fermentation phase I and II were collected and analyzed using Polymerase Chain Reaction-Denaturing gradient gel electrophoresis（PCRDGGE） method based on 16 S rDNA universal primers from Escherichia coli. A total of 56 different clone sequences were obtained（GenBank accession numbers of KF630598-KF630653）. They were classified into 7 phyla and 43 genus. Dominant microflora during composting belonged to phyla Proteobacteria, Firmicutes, and Actinobacteria, with the dominant genus of Bacillus, Paenibacillus, Thermomonospora, Thermasporomyces, Pseudomonas, and Cellvibrio. Bacterial diversity（Shannon index） analysis showed that bacterial species in straw treatment continuously increased during composting, while those in corn stalks treatment firstly increased and then reduced. Principal component analysis showed that corn stalk compost at the end of fermentation phase I and phase II were clustered into one group, suggesting that corn stalk composts faster than anticipated. In general, straw compost has higher bacterial diversity but longer composting time, while corn stalk compost has lower bacterial diversity but shorter composting time.