摩拉水牛及德宏水牛瘤胃产甲烷菌多样性比较

本试验旨在比较分析摩拉水牛及德宏水牛瘤胃液中产甲烷菌的多样性。选取健康雌性摩拉水牛及德宏水牛各3头,采用口腔导管法收集瘤胃液,酚—氯仿—异戊醇抽提法提取瘤胃液总DNA,用产甲烷菌特异性引物Met86F/Met1340R扩增产甲烷菌16SrDNA,构建16SrDNA基因克隆文库。摩拉水牛及德宏水牛各获得96个16SrDNA基因序列,均归类于Methanobacteriales目,其中德宏水牛有82个序列(18个OTUs)与已知菌的16SrDNA序列相似性≥97%,占总序列的85.4%,有14个序列(9个OTUs)与已知菌16SrDNA序列相似性为89%~97%;摩拉水牛有94个序列(13个OTUs)与已知菌16SrDNA序列相似性≥97%,占总序列的97.9%,仅有2个序列(1个OTUs)与已知菌16SrDNA序列相似性为94%。系统进化树分析表明,所有序列分别聚集于两大分支上,其中德宏水牛有13个OTUs代表序列和摩拉水牛7个OTUs代表序列聚集在进化树顶端的同一分支上,且在系统发育距离上与Methanobacteriales目中任何已知相似序列都相隔较远。德宏水牛瘤胃中的SGMT簇序列和RO簇序列所占总序列比列分别为83.3%、9.4%,摩拉水牛瘤胃中的SGMT簇序列和RO簇序列所占总序列的比列分别为51.0%、9.4%。由此可见,摩拉水牛及德宏水牛瘤胃产甲烷菌序列以Methanobacteriales目为主,其中德宏水牛拥有更多未知的产甲烷菌序列;德宏水牛瘤胃中的SGMT簇产甲烷菌序列比例要高于摩拉水牛。     

尼里-拉菲水牛及海子水牛瘤胃产甲烷菌多样性比较

试验旨在比较尼里-拉菲水牛及海子水牛瘤胃液中产甲烷菌的多样性,选取成年雌性尼里-拉菲水牛及海子水牛各3头作为试验动物,采用口腔导管法收集瘤胃液,提取瘤胃液总DNA,采用产甲烷菌特异性引物Met86F/Met1340R扩增16S rDNA,构建16S rDNA基因克隆文库。结果显示,从尼里-拉菲水牛和海子水牛中分别获得73、100条有效序列,均以Methanobrevibacter属为主。其中尼里-拉菲水牛有70条序列（17个OTUs）与已知菌的16S rDNA序列相似性 ≥ 97%,占总序列的95.89%,有3条序列（3个OTUs）与已知菌16S rDNA序列相似性处于93%~96%;海子水牛有70条序列（15个OTUs）与已知菌16S rDNA序列相似性 ≥ 97%,占总序列的70%,有30条序列（18个OTUs）与已培养菌16S rDNA序列相似性处于84%~96%。尼里-拉菲水牛和海子水牛瘤胃中的SGMT簇序列、RO簇序列所占总序列比例分别为76.71%、21.91%和76.00%、19.00%。系统进化树分析表明,所有序列分别聚集于两大分支上,其中海子水牛有16个OTUs代表序列和尼里-拉菲水牛8个OTUs代表序列聚集在进化树顶端的同一分支上,且系统发育距离与古菌中任何已知相似序列都相隔较远。综上所述,在本试验同等日粮条件下,尼里-拉菲水牛及海子水牛瘤胃内产甲烷菌序列均以Methanobrevibacter属为主,两水牛品种的SGMT簇序列比例基本一致,但尼里-拉菲水牛中的RO簇序列比例更高,海子水牛瘤胃内存在更多的未知产甲烷菌。     

尼里-拉菲水牛及海子水牛瘤胃产甲烷菌多样性比较

试验旨在比较尼里-拉菲水牛及海子水牛瘤胃液中产甲烷菌的多样性,选取成年雌性尼里-拉菲水牛及海子水牛各3头作为试验动物,采用口腔导管法收集瘤胃液,提取瘤胃液总DNA,采用产甲烷菌特异性引物Met86F/Met1340R扩增16S rDNA,构建16S rDNA基因克隆文库。结果显示,从尼里-拉菲水牛和海子水牛中分别获得73、100条有效序列,均以Methanobrevibacter属为主。其中尼里-拉菲水牛有70条序列(17个OTUs)与已知菌的16S rDNA序列相似性≥97%,占总序列的95.89%,有3条序列(3个OTUs)与已知菌16S rDNA序列相似性处于93%～96%;海子水牛有70条序列(15个OTUs)与已知菌16S rDNA序列相似性≥97%,占总序列的70%,有30条序列(18个OTUs)与已培养菌16S rDNA序列相似性处于84%～96%。尼里-拉菲水牛和海子水牛瘤胃中的SGMT簇序列、RO簇序列所占总序列比例分别为76.71%、21.91%和76.00%、19.00%。系统进化树分析表明,所有序列分别聚集于两大分支上,其中海子水牛有16个OTUs代表序列和尼里-拉菲水牛8个OTUs代表序列聚集在进化树顶端的同一分支上,且系统发育距离与古菌中任何已知相似序列都相隔较远。综上所述,在本试验同等日粮条件下,尼里-拉菲水牛及海子水牛瘤胃内产甲烷菌序列均以Methanobrevibacter属为主,两水牛品种的SGMT簇序列比例基本一致,但尼里-拉菲水牛中的RO簇序列比例更高,海子水牛瘤胃内存在更多的未知产甲烷菌。     

基于16S rRNA基因克隆文库技术分析广西富钟水牛瘤胃产甲烷菌组成及多样性

本研究旨在利用16S rRNA基因克隆库技术分析广西富钟水牛瘤胃产甲烷菌组成及多样性。选取3头体况基本一致的健康雌性富钟水牛作为试验动物,采用机械破壁法提取瘤胃内容物总DNA,采用产甲烷菌引物Met86F/Met1340R扩增16S rRNA基因,构建16S rRNA基因克隆文库。结果表明,本试验共获得93个非嵌合体16S rRNA序列,按照97%的相似性划分为39个分类操作单元（OTU）。其中,60个序列（15个OTU）与已培养细菌16S rRNA序列相似性≥97%,占总序列的64.5%;32个序列（23个OTU）与已培养菌16S rRNA序列相似性处于90%^<97%;仅有1个序列与Methanomassiliicoccus luminyensis相似性<90%。系统发育树分析表明,98.9%的序列均属于甲烷杆菌目（Methanobacteriales）,部分序列与Methanobacteriales中任何已知相似序列都相隔较远,它们可能代表Methanobacteriales中新的属或种。由上述结果可见,富钟水牛瘤胃产甲烷菌以Methanobacteriales为优势菌群,其中有许多未知的产甲烷菌需进一步分离培养并对其功能进行分析。     

基于16S rRNA基因序列分析梅花鹿瘤胃细菌多样性

本试验旨在对以柞树叶为主要粗饲料来源的梅花鹿瘤胃细菌多样性进行分析。提取瘤胃微生物基因组DNA,扩增细菌16S rRNA基因,构建16S rRNA基因克隆文库,分析梅花鹿瘤胃细菌区系组成。结果表明:1)试验共得到107个非嵌合体16S rRNA基因序列。按照97%序列相似性,划分为22个分类操作单元(OTUs)。其中91个序列(10个OTUs,85%总克隆序列)与已培养菌序列相似性≥97%,13个序列(10个OTUs,12.2%总克隆序列)与已培养菌序列相似性为90%~97%,其余序列相似性<90%。87.9%序列与普雷沃氏菌属(Prevotella spp.)相似。2)系统发育分析表明,梅花鹿瘤胃细菌由厚壁菌门和拟杆菌门组成。由结果可知,以富含单宁的柞树叶为主要粗饲料来源的梅花鹿的瘤胃中,普雷沃氏菌属是优势细菌,而牛、羊瘤胃中常见的纤维素降解菌未检测到,这可能与饲料中单宁含量高有关。     

山羊瘤胃产甲烷古菌多样性及与其他动物瘤胃的比较

旨在研究山羊瘤胃产甲烷古菌的多样性并与其他动物瘤胃进行比较。采用产甲烷菌特异性引物Met86F/Met1340R研究山羊瘤胃产甲烷古菌16SrRNA基因的多样性,随机挑选100个克隆进行序列分析。结果表明,所有克隆经限制性内切酶酶切后共获得16个OTU,绝大多数为甲烷短杆菌,其中与甲烷短杆菌菌株AK-87、ZA-10、OCP、SM9和30Y序列最相近的克隆数分别为58%、19%、7%、2%和2%,还有与Methanosphaera stadtmanae(1%)、Methanobacterium aarhusense(2%)、Aciduliprofundum boonei(3%)、Methanobrevibacter sp.AK-87(4%)和Methanobrevibactersp.1Y(2%)等相似的古菌序列。将研究结果与已经报道的关于牛和绵羊瘤胃产甲烷菌多样性比较,发现不同PCR引物可检测出不同菌群结构,而饲料类型、动物种类可影响瘤胃产甲烷菌的菌群结构。     

德宏水牛及其杂交后代染色体多态性分析

[目的]为德宏水牛进一步级进杂交方案的选择提供科学依据。[方法]采用外周血及耳组织细胞培养法与染色体G-带分析对20头德宏本地水牛、25头摩拉与德宏二元杂水牛以及35头尼里与摩拉和德宏三元杂水牛进行了染色体核型分析。[结果]表明:德宏水牛染色体数均为2n=48;二元杂水牛染色体数均为2n=49;三元杂水牛出现2n=49,2n=50两种类型。[结论]从细胞遗传学角度首次阐明了德宏水牛染色体核型特征,揭示了德宏水牛二元杂、三元杂后代染色体多态性遗传规律,并探讨了染色体多态性遗传机理及与繁殖性能的关系。     

沼泽型水牛微卫星DNA的遗传多样性分析

为探究中国沼泽型水牛品种或类群的遗传多样性,本研究采用了沼泽型水牛30个微卫星标记结合LabChip芯片检测法对德昌水牛、德宏水牛、温州水牛、贵州水牛、西林水牛、富钟水牛及两个引进品种摩拉水牛、尼里－拉菲水牛等40个样本进行了检测和遗传多样性分析。结果表明,在30个微卫星DNA标记上共发现332个等位基因,平均基因多样性和PIC值分别为0.7808和0.7554。聚类分析结果显示,德宏水牛和德昌水牛先聚为一类,随后与富钟水牛、贵州水牛、温州水牛、西林水牛聚为一类;摩拉水牛与尼里－拉菲水牛聚为一类。本研究证实了所选的30个沼泽型水牛微卫星DNA标记可作为有效的遗传标记用于水牛品种间的遗传多样性分析,同时也丰富了当前的水牛微卫星标记资源。     

利用MiSeq测序技术分析锦江牛瘤胃细菌多样性

本研究旨在利用MiSeq高通量测序技术揭示锦江牛瘤胃细菌组成及多样性。选用3头装有永久性瘤胃瘘管的锦江牛成年公牛[体重(400±20)kg],采集瘤胃液样品后提取细菌DNA,对瘤胃细菌的16S rDNA序列V4~V5区进行MiSeq测序,分析物种的丰度、分布及α-多样性。结果表明:3个样本共获得56 763条高质量16S rDNA序列,946个分类操作单元(OTU),锦江牛瘤胃细菌丰富度指数Chao指数、Ace指数分别为836、841,α-多样性指数Shannon指数、Simpson指数分别为4.96、0.021 5;经过物种注释,15个门、23个纲、26个科以及44个属在所有样品中被鉴定。在门水平,拟杆菌门(Bacteroidetes)(64.37%)、厚壁菌门(Firmicutes)(21.20%)和变形菌门(Proteobacteria)(7.59%)数量最多;在纲水平,主要包括拟杆菌纲(Bacteroidia)(52.88%)和梭菌纲(Clostridia)(17.03%);在科水平,主要包括普雷沃氏菌科(Prevotellaceae)(36.26%)、瘤胃菌科(Ruminococcaceae)(9.33%)和毛螺菌科(Lachnospiraceae)(4.89%);在属水平,优势细菌主要包括普雷沃氏菌属(Prevotella)(28.97%)、帕拉普氏菌属(Paraprevotella)(3.08%)、理研菌属(Rikenella)(2.28%)、梭菌_Ⅳ(ClostridiumⅣ)(1.77%)、解琥珀酸菌属(Succiniclastium)(1.62%)和瘤胃球菌属(Ruminococcus)(1.53%)。结果证明,锦江牛瘤胃细菌多样性相对较低;锦江牛瘤胃中最优势菌门是拟杆菌门,其次是厚壁菌门;普雷沃氏菌属是锦江牛瘤胃中最优势细菌属。     

和牛及其杂交牛瘤胃细菌多样性分析

为了揭示纯种和牛及杂交和牛瘤胃细菌组成及多样性,选取安装有永久性瘤胃瘘管的年龄、体重和体况接近的纯种和牛公牛(WW)、和牛♂×西门塔尔牛♀杂交F1代公牛(WS)及和牛♂×荷斯坦牛♀杂交F1代公牛(WH)各3头,采集试验牛瘤胃液,提取细菌DNA,对细菌16S rDNA V4～V5区(515F～907R)进行PCR扩增,扩增产物用Illumina HiSeq PE250测序平台进行高通量测序。结果表明:9个瘤胃液样本共获得604 174条有效16S rDNA序列,19 912个分类操作单元(OTU),物种丰富度指数(Chao指数)WW组极显著高于WH组(P0.01),WS组显著高于WH组(P0.05),Ace指数WW组与WS组均显著高于WH组(P0.05),但WW组与WS组相比差异不显著(P0.05)。在门水平上,3组相对含量最多的菌门为拟杆菌门(Bacteroidetes),其次是厚壁菌门(Firmicutes)和变形菌门(Proteobacteria),分别占测序总数的56.08%～66.29%、21.20%～27.01%和2.09%～5.45%;在科水平上,3组相对含量最多的菌属为普雷沃氏菌科(Prevotellaceae),占测序总量的37.12%～44.23%;在属水平上,3组相对含量最多的菌属为普雷沃氏菌属-1(Prevotella-1),占测序总量的23.98%～35.95%。在97%相似性水平下,3组共享OTU数为1 969个,占瘤胃液细菌总OTU数的59.83%,3组间瘤胃菌群结构存在一定差异,同一品种牛的瘤胃菌群结构表现出明显的聚集趋势。说明纯种和牛及杂交和牛瘤胃中最优势菌门均为拟杆菌门(Bacteroidetes),其次为厚壁菌门(Firmicutes)和变形菌门(Proteobacteria);最优势菌科为普雷沃氏菌科(Provotellaceae);最优势菌属均为普雷沃氏菌属-1(Prevotella-1),纯种和牛与杂交和牛在物种多样性指数上并无显著性差异。     

Methanogen diversity in the rumen of Indian Surti buffalo (Bubalus bubalis), assessed by 16S rDNA analysis

The methanogenic communities in buffalo rumen were characterized using a culture-independent approach of a pooled sample of rumen fluid from three adult Surti buffaloes. Buffalo rumen is likely to include species of various methanogens, so 16S rDNA sequences were amplified and cloned from the sample. A total of 171 clones were sequenced to examine 16S rDNA sequence similarity. About 52.63% sequences (90 clones) had ≥ 90% similarity, whereas, 46.78% of the sequences (81 clones) were 75-89% similar to 16S rDNA database sequences, respectively. Phylogenetic analyses were also used to infer the makeup of methanogenic communities in the rumen of Surti buffalo. As a result, we distinguished 23 operational taxonomic units (OTUs) based on unique 16S rDNA sequences: 12 OTUs (52.17%) affiliated to Methanomicrobiales order, 10 OTUs (43.47%) of the order Methanobacteriales and one OTU (4.34%) of Methanosarcina barkeri like clone, respectively. In addition, the population of Methanomicrobiales and Methabacteriales orders were also observed, accounting 4% and 2.17% of total archea. This study has revealed the largest assortment of hydrogenotrophic methanogens phylotypes ever identified from rumen of Surti buffaloes.     

广西的尼里—拉菲水牛和摩拉水牛mtDNA D-loop区遗传多样性研究

[目的]探究引入广西南宁的尼里-拉菲水牛和摩拉水牛的mtDNA D-loop区遗传多样性与母系起源。[方法]采用PCR扩增、测序及生物信息学方法。[结果]对从广西南宁采集的52个江河型水牛（尼里-拉菲水牛25个,摩拉水牛27个）mtDNA D-loop序列与GenBank下载的20条尼里-拉菲水牛和23条摩拉水牛序列进行联合分析,共检测到112个变异位点,定义42个单倍型,发现摩拉水牛（Hd: 0.934±0.027）与尼里-拉菲水牛的遗传多样性（Hd: 0.929±0.017）都很丰富。NJ系统发育树显示尼里-拉菲水牛和摩拉水牛含有江河型和沼泽型水牛mtDNA支系,表明尼里-拉菲水牛和摩拉水牛引入中国后均与沼泽型水牛进行了杂交,在外貌上很难区分。[结论]引入广西南宁的尼里-拉菲水牛和摩拉水牛与沼泽型水牛存在广泛的血缘混杂现象。     

摩拉水牛和尼里-拉菲水牛PRKAA2基因SNPs检测及遗传多样性分析

为研究水牛蛋白激酶AMP活化的催化亚基α2（protein kinase AMP-activated catalytic subunit alpha 2,PRKAA2）基因多态性,本试验以摩拉水牛和尼里-拉菲水牛基因组DNA为模板,扩增PRKAA2基因外显子4及内含子3部分序列,通过常规测序法检测其SNP并进行遗传多样性分析。结果发现,PRKAA2基因外显子4内存在1个SNP位点（c.462 G>A）,PRKAA2基因内含子3部分序列存在3个SNPs位点（IVS3.557 T>C、IVS3.560 C>T和IVS3.565 G>A）。经遗传多样性分析表明,在c.462 G>A位点的野生纯合型和杂合型比突变纯合型更有优势,IVS3.557 T>C和IVS3.560 C>T位点的突变纯合型为非优势基因型,IVS3.565 G>A位点杂合型为优势基因型。IVS3.565 G>A位点在摩拉水牛群体中处于Hardy-Weinberg非平衡状态;c.462 G>A位点在尼里-拉菲水牛群体中处于Hardy-Weinberg非平衡状态。4个SNPs位点在摩拉水牛群体中均为中度多态;c.462 G>A、IVS3.557 T>C位点在尼里-拉菲水牛群体中为低度多态,IVS3.560 C>T、IVS3.565 G>A位点为中度多态。IVS3.557 T>C位点在两个水牛群体中杂合度较低。说明摩拉水牛IVS3.565 G>A位点和尼里-拉菲水牛c.462 G>A位点的基因型频率和基因频率遗传状态不平衡,尼里-拉菲水牛群体中IVS3.557 T>C位点遗传变异小,选择潜力不高。4个多态位点可以构建5种单倍型,其中T-C-G-G是摩拉水牛群体和尼里-拉菲水牛群体的优势单倍型。综上,本研究检测的摩拉水牛和尼里-拉菲水牛PRKAA2基因上4个SNPs位点可为水牛标记辅助选择育种提供参考。     

热应激对摩拉和尼里-拉菲水牛血液生化指标的影响

【目的】 试验旨在探究热应激条件下摩拉和尼里-拉菲水牛的血液生化指标变化规律。【方法】 选取体况相近的健康摩拉和尼里-拉菲水牛各11头。试验期划分为热应激期和非应激期, 并于各试验期分别进行4次采血, 每次间隔1周。测定血清激素浓度、抗氧化指标和离子浓度等生化指标。【结果】 摩拉水牛热应激期血清超氧化物歧化酶(SOD)活性及总胆固醇(CHOL)、丙二醛(MDA)、热休克蛋白70(HSP70)、钾离子(K⁺)、钠离子(Na⁺)、氯离子(Cl^-)含量均显著高于非应激期(P<0.05), 谷丙转氨酶(ALT)、谷草转氨酶(AST)、谷胱甘肽过氧化物酶(GSH-Px)活性及总抗氧化能力(T-AOC)、胰岛素样生长因子1(IGF-1)、皮质醇(CORT)、瘦素(LEP)、钙离子(Ca²⁺)含量均显著低于非应激期(P<0.05);尼里-拉菲水牛热应激期血清白蛋白(ALB)、CHOL水平及谷氨酰基转移酶(GGT)活性均显著高于非应激期(P<0.05), 球蛋白(GLOB)含量显著低于非应激期(P<0.05)。在热应激期, 摩拉水牛ALT活性及总蛋白(TP)、GLOB、CHOL、尿素氮(BUN)、甘油三酯(TG)含量均显著高于尼里-拉菲水牛(P<0.05), ALB、四碘甲腺原氨酸(T₄)、K⁺含量均显著低于尼里-拉菲水牛(P<0.05);在非应激期, 摩拉水牛ALT活性、BUN、TG、葡萄糖(GLU)含量及T-AOC均显著高于尼里-拉菲水牛(P<0.05)。【结论】 热应激降低了摩拉和尼里-拉菲水牛的内分泌激素水平和抗氧化能力并影响了机体电解质平衡, 尼里-拉菲水牛的免疫功能也有一定下降; 在热应激条件下, 摩拉水牛表现出更强的清除自由基能力, 尼里-拉菲水牛表现出更好的代谢、体温调节能力以及电解质代偿能力。     

Twinning rate in buffaloes: A case report

The information of twin pregnancy frequencies in buffaloes is scarce in the literature. Evidence even indicates that twin pregnancies are not carried to term, but it does not seem to be the case. In this case report, the event of twin pregnancy in buffaloes is reported as well as its frequency. Twin birth is the presence of two or more embryos in a single gestation. Multiple births are uncommon in buffaloes since the species is uniparous. We studied a population of Murrah buffaloes in which slightly more than 13,000 births were observed. In this study, the frequency of occurrence of twin births in Murrah buffaloes was 0.14%. This low incidence impaired the estimation of genetic parameters. Twins can have advantages and disadvantages depending on the production system, which are discussed in this case report.     

高产与低产德宏奶水牛耐热性能差异的试验研究

基于DHI数据,选择高产与低产的德宏奶水牛各15头作为实验动物。在平均温湿指数（THI）在72以上的饲养环境中进行为期27d的试验,对奶水牛的泌乳量、乳品质、热应激相关激素的浓度和生理参数进行了测试分析。相比高产奶水牛,低产奶水牛的体细胞数量较高,非脂乳固体含量却相对较低（P〈0．05）,其他乳品质参数在两组间无显著性差异。在热应激条件下,低产组奶水牛的促肾上腺皮质激素（adrenocomcotropic hormone,ACTH）浓度相对较低,而皮质醇（cortisol,Cor）浓度却相对较高,反映代谢水平的胰岛素（insulin,INS）和T3浓度相对较高。从热应激相关激素浓度和乳成分来看,     

Comparison of voluntary feed intake, rumen passage and degradation kinetics between crossbred Brahmam cattle (Bos indicus) and swamp buffaloes (Bubalus bubalis) fed a fattening diet based on corn silage

Three crossbred tropical cattle and three crossbred buffaloes, fitted with rumen cannulas, were used to investigate the differences in feed intake, digestibility and parameters with rumen degradation kinetics between cattle and buffalo fed a fattening diet in the Philippines. The animals were fed a diet consisting of 50% corn silage (CS), 30% brewer's grain (BG) and 20% concentrate mixture (CM) on a dry matter (DM) basis, at a level of 3% bodyweight (BW) as DM. Total DM intake (DMI) and rumen fill were greater (P < 0.05) for buffaloes than for cattle, but significant differences were not detected between the animal species when those were expressed as percentage of BW. The DMI per metabolic body size for BG and CM were greater for buffaloes than for cattle (P < 0.05), whereas that of CS did not differ between the species (P > 0.05). The weight proportion of consumed feedstuffs differed between cattle and buffaloes (P < 0.05), although nutrient intake per total DMI did not differ significantly (P > 0.05) between the animal species. Nutrients digestibility was higher (P < 0.05) for buffaloes than for cattle. Ruminal passage rate constant of feed particles and rumen fluid did not differ (P > 0.05) between the animal species. Higher DM degradation rate constant and effective degradability with CS were prominent for buffaloes than for cattle (P < 0.05). Degradation parameters did not differ (P > 0.05) between cattle and buffaloes, neither with BG nor CM. The results indicate that the greater digestibility with DM and energy for buffaloes are ascribable to greater effective degradability of CS fed ad libitum level.     

水牛分离精子与不同来源卵母细胞体外受精的效果研究

本研究的目的是探讨水牛分离精子与不同来源(活体采卵或屠宰场卵巢采卵)卵母细胞体外受精的效果。活体采卵是选用20头空怀河流型母水牛(其中摩拉母牛12头,尼里-拉菲母水牛8头)每间隔3 d采卵1次,连续采卵5~6周,活体采集卵母细胞;屠宰场卵巢采卵是收集屠宰场水牛卵巢,用10 mL注射器连接18 G针头吸取水牛卵巢上可视的卵泡来收集卵母细胞。将收集的AB级水牛卵母细胞在相同的条件下进行体外成熟、然后用分离或未分离精子进行体外受精以及体外培养至囊胚。结果发现:活体采卵组和屠宰场收集的水牛卵母细胞组用分离精子受精分裂率和囊胚率没有差异(P>0.05);分离精子和未分离精子的体外受精分裂率和囊胚率也没有差异(P>0.05)。由此说明,水牛分离精子可以用于体外生产性控胚胎。     

The community structure and microbial linkage of rumen protozoa and methanogens in response to the addition of tea seed saponins in the diet of beef cattle

Background: This study investigated changes in rumen protozoal and methanogenic communities, along with the correlations among microbial taxa and methane(CH_4) production of six Belmont Red Composite beef steers fed tea seed saponins(TSS). Animals were fed in three consecutive feeding periods, a high-grain basal diet for 14 d(BD period) then a period of progressive addition of TSS to the basal diet up to 30 g/d for 20 d(TSS period), followed by the basal diet for 13 d without TSS(BDP post-control period).Results: The study found that TSS supplementation decreased the amount of the protozoal genus Entodinium and increased Polyplastron and Eudiplodinium genera. During BDP period, the protozoa community of steers did not return to the protozoal profiles observed in BD period, with higher proportions of Metadinium and Eudiplodinium and lower Isotricha. The addition of TSS was found to change the structure of methanogen community at the subgenus level by decreasing the abundance of methanogens in the SGMT clade and increasing the abundance of methanogens in the RO clade. The correlation analysis indicated that the abundance of SGMT clade methanogens were positively correlated with Isotricha, and Isotricha genus and SGMT clade methanogens were positively correlated with CH_4 production. While RO clade were positively correlated with the proportion of Metadinium genus, which was negatively correlated with CH_4 emission.Conclusions: These results suggest that different genera of rumen protozoa ciliates appear to be selectively inhibited by TSS, and the change in methanogen community at the subgenus level may be due to the mutualistic relationships between methanogens and rumen ciliates.