肉鸭养殖过程中发酵床垫料菌群结构变化的研究

Succession of bacterial community in duck bio-bed over time

摘要:

【目的】探究发酵床使用时间和肉鸭粪便微生物对发酵床垫料菌群结构、总菌和大肠杆菌数量的影响。【方法】采集江苏某肉鸭发酵床养殖场内刚制作完成的发酵床垫料样品，及其饲养4批次、8批次后的垫料样品，同时采集各批次34日龄肉鸭粪便样品，采用变性梯度凝胶电泳技术(Denaturing gradient gel electrophoresis，DGGE)、16S rRNA基因序列分析和实时荧光定量PCR (Quantitative real-time PCR)技术对发酵床使用过程中垫料菌群结构进行定性和定量研究。【结果】0批次(D0)与4批次(D4)、8批次(D8)垫料菌群相似性分别为68.81%、70.82%，而4批次与8批次间垫料菌群的相似性则达81.93%，显著高于D4、D8与D0间相似性(P<0.05)。条带6、8 (最相似菌分别为Leqionella tunisiensis、Pedobacte bauzanensis)在3个时间点垫料菌群中均表现优势，且含量较为稳定；条带10 (最相似菌为Rummeliibacillus suwonesis)仅在2个重复使用垫料菌群中表现优势；条带12、13 (最相似菌分别是Psychrobacter sp. PRwf-1、Iamia majanohamensis)共同存在于垫料样和粪便样。肉鸭粪便中大肠杆菌的数量显著高于4批次、8批次垫料中的数量(P<0.05)，与0批次垫料间差异不显著(P>0.05)。【结论】使用时间和肉鸭粪便微生物共同影响了发酵床垫料菌群结构和数量，菌群结构随使用时间的延长而趋于稳定。

Abstract:

[Objective] This study aimed to character the succession of bacterial community in the litter during the fermentation of duck bio-bed. [Methods] The experiment was conducted in one meat duck bio-bed farm of Jiangsu province. The new litter (D0), used litter from 4th (D4), 8th (D8) meat duck flock and feces from duck at 34 d age were sampled. Denaturing gradient gel electrophoresis (DGGE), 16S rRNA gene sequencing and quantitative real-time PCR were used to monitor the composition changes of microbiota in the bio-bed. [Results] DGGE profiles showed that, the similarity between D4 and D8 was 81.93%, which was significantly higher than the similarity between D4, D8 and D0 (68.81% and 70.82%, respectively) (P<0.05). Bands 6 and 8 (the closest relatives: Leqionella tunisiensis and Pedobacter bauzanensis, respectively) were dominant in all litter samples. Band 10 (the closest relative: Rummeliibacillus suwonesis) was only dominant in used litters. Bands 12 and 13 (the closest relatives: Psychrobacter sp. PRwf-1, Iamia majanohamensis, respectively) commonly presented in all litters and feces. Real-time PCR results showed that the number of Escherichia coli in duck feces was significantly higher than that in D4 and D8 litters (P<0.05), while no significant difference was observed between duck feces and D0 litter (P>0.05). [Conclusion] Both the application time of bio-bed and duck feces origin microorganisms can affect the bacterial community and the number of E. coli in the bio-bed, and the bacterial community structure tends to be stable in line with the application time of the bio-bed.