中国农业科学Issue(4)：759-768,10.
基于DGGE和T-RFLP分析采食不同粗饲料梅花鹿瘤胃细菌区系
Analysis of Bacterial Diversity in Rumen of Sika Deer (Cervus nippon) fed Different Forages Using DGGE and T-RLFP
摘要
Abstract
[Objective]Bacterial communities play critical roles in the rumen fermentation of Sika deer (Cervus nippon), while the bacterial composition in the rumen of Sika deer is rarely reported. The objective of present study is to investigate the bacterial diversity in the rumen of Sika deer, which can provide a molecular basis for manipulation of rumen fermentation.[Method]Four two year old male rumen-cannulated Sika deers fed oak leaf (OL group, Sika deer A and B) and corn stover (CS group, Sika deer C and D) based diets were used in the present study. After 30 days of feeding, rumen contents including solid and liquid fractions were sampled, and the microbial genomic DNA was extracted. V3 region of ruminal bacterial 16S rRNA gene and 16S rRNA gene was amplified, which was used in the DGGE and T-RFLP analysis, respectively. The clustering analysis was applied to DGGE results. The dominant bands in DGGE profiles were obtained, and then used to clone sequencing in order to indentify the bacterial communities. The results of T-RFLP were also applied to clustering analysis, and the possible bacterial structure was speculated by Microbial Community AnalysisⅢ (MiCAⅢ) dataset. [Result]The clustering patterns of DGGE revealed that the similarity of bacterial diversity between CS group and OL group was lower than 65%, indicating that the bacterial diversity was affected by forage source. The similarity between Sika deer A and B in the OL group and between Sika deer C and D in the CS group was greater than 70%and 75%, respectively. In addition, the differences were found between animals in the same group. A total of 20 and 24 unique DGGE bands were obtained from the OL and CS groups, respectively. Sequences analysis of DGGE showed that the bacteria in the OL group were composed of Bacteroides, Firmicutes and Proteobacteria phyla, while they were composed of Bacteroides, Firmicutes, Proteobacteria and Synergistetes phyla in the CS group. Prevotella spp. was the dominant bacteria in the OL and CS groups, but the composition of genus Prevotella at species level was different in two groups. The dominant fibrolytic bacteria in two groups includes Clostridium spp. and Eubacterium spp.. The results of T-RFLP showed that the highest the highest richness, diversity and evenness indices, and the lowest dominance index were found in Sika deer D (CS group). The diversity indices in Sika deer A and B were comparative, but lower than that in Sika deer D, suggesting that forage (oak leaf) in the OL group affected the relative biomass of rumen microbiome. There were clear discrepancies in the diversity indices of Sika deer C and D, and the diversity indices in Sika deer C were lower than Sika deer A and B, indicating that the intra-individual variation. T-RFs representing 81, 214, 272 and 308 bp in OL group were dominant, 90, 95, 175, 273 and 274 bp were predominant in CS group, and 161, 259, 264, 266 and 284 bp were presented in all animals. According to the results of MiCAⅢ, the possible bacteria represented by these T-RFs could assign to the phyla Bacteroides, Firmicutes, Proteobacteria and Acidobacteria. The clustering analysis of T-RFs showed that two clusters were generated from the T-RFs in the two groups, and the T-RFs profiles in Sika deer A, B and C were similar, indicating that forage sources affected the profiles of T-RFs.[Conclusion]These results suggested that Prevotella spp. were the dominant bacteria in rumen of sika deer. The forage source affected the rumen bacterial communities.关键词
梅花鹿/细菌区系/普雷沃氏菌/单宁Key words
Sika deer/bacterial structure/Prevotella spp./tannins引用本文复制引用
李志鹏,姜娜,刘晗璐,崔学哲,荆祎,杨福合,李光玉..基于DGGE和T-RFLP分析采食不同粗饲料梅花鹿瘤胃细菌区系[J].中国农业科学,2014,(4):759-768,10.基金项目
国家科技支撑计划（2011BAI03B02）、中国农业科学院特产研究所创新工程专项经费 （）
