School of Veterinary Medicine, Università di Bologna, Bologna, Italy.
J Dairy Sci. 2010 Jan;93(1):279-87. doi: 10.3168/jds.2009-2207.
The influence of pH dynamics on ruminal bacterial community composition was studied in 8 ruminally cannulated Holstein cows fitted with indwelling electrodes that recorded pH at 10-min intervals over a 54-h period. Cows were fed a silage-based total mixed ration supplemented with monensin. Ruminal samples were collected each day just before feeding and at 3 and 6h after feeding. Solid and liquid phases were separated at collection, and extracted DNA was subjected to PCR amplification followed by automated ribosomal intergenic spacer analysis (ARISA). Although cows displayed widely different pH profiles (mean pH=6.11 to 6.51, diurnal pH range=0.45 to 1.39), correspondence analysis of the ARISA profiles revealed that 6 of the 8 cows showed very similar bacterial community compositions. The 2 cows having substantially different community compositions had intermediate mean pH values (6.30 and 6.33) and intermediate diurnal pH ranges (averaging 0.89 and 0.81 pH units). Fortuitously, these 2 cows alone also displayed milk fat depression, along with markedly higher ruminal populations of 1 bacterial operational taxonomic unit (OTU) and reduced populations of another ARISA amplicon. Cloning and sequencing of the elevated OTU revealed phylogenetic similarity to Megasphaera elsdenii, a species reportedly associated with milk fat depression. The higher populations of both M. elsdenii and OTU246 in these 2 cows were confirmed using quantitative real-time PCR (qPCR) with species-specific primers, and the fraction of total bacterial rDNA copies contributed by these 2 taxa were very highly correlated within individual cows. By contrast, the fraction of total bacterial rDNA copies contributed by Streptococcus bovis and genus Ruminococcus, 2 taxa expected to respond to ruminal pH, did not differ among cows (mean= <0.01 and 10.6%, respectively, of rRNA gene copies, determined by qPCR). The data indicate that cows with widely differing pH profiles can have similar ruminal bacterial community compositions, and that milk fat depression can occur at intermediate ruminal pH. The results support recent reports that milk fat depression is associated with shifts in bacterial community composition in rumine and is specifically related to the relative abundance of Megasphaera elsdenii.
本研究旨在探讨 pH 动态对瘤胃细菌群落组成的影响。试验选择 8 头安装有内置电极的荷斯坦奶牛，这些奶牛每 10min 记录一次 pH 值，连续记录 54h。奶牛饲喂以青贮为基础的全混合日粮，日粮中添加莫能菌素。每天在饲喂前和饲喂后 3h 和 6h 采集瘤胃液样品。采集时固液分离，提取 DNA 后进行聚合酶链式反应（PCR）扩增，接着进行自动核糖体间隔区分析（ARISA）。尽管奶牛的 pH 值变化范围较大（平均 pH 值为 6.11-6.51，昼夜 pH 值范围为 0.45-1.39），但 ARISA 图谱的对应分析表明，8 头奶牛中有 6 头具有非常相似的细菌群落组成。2 头奶牛的细菌群落组成有显著差异，其平均 pH 值（6.30 和 6.33）和昼夜 pH 值范围（平均 0.89 和 0.81 pH 单位）处于中间水平。巧合的是，这 2 头奶牛同时还表现出乳脂率降低，并且瘤胃中 1 个细菌操作分类单元（OTU）的种群显著增加，另一个 ARISA 扩增子的种群减少。对升高的 OTU 进行克隆和测序，结果表明其与报道的与乳脂率降低有关的 Megasphaera elsdenii 具有系统发育相似性。这 2 头奶牛中 Megasphaera elsdenii 和 OTU246 的种群数量均使用针对特定物种的引物，通过实时荧光定量 PCR（qPCR）得到了证实，并且这 2 个分类单元在单个奶牛中的总细菌 rDNA 拷贝的比例高度相关。相比之下，奶牛之间的链球菌和瘤胃球菌（Ruminococcus）的总细菌 rDNA 拷贝比例没有差异（qPCR 检测的 rRNA 基因拷贝数分别为<0.01 和 10.6%），这 2 个分类单元预计会对瘤胃 pH 做出反应。研究结果表明，pH 值变化范围较大的奶牛可能具有相似的瘤胃细菌群落组成，并且乳脂率降低可能发生在中间的瘤胃 pH 值。结果支持了最近的报道，即乳脂率降低与瘤胃细菌群落组成的变化有关，并且与 Megasphaera elsdenii 的相对丰度有关。
