不同粗纤维源日粮对梅花鹿瘤胃微生物区系的影响

为了研究不同粗纤维源日粮对梅花鹿瘤胃微生物区系的影响,选择6头健康公鹿分成A,B 2组,每组3头,分别以玉米秸秆和柞树叶作为粗纤维源,试验时期为2个月。瘤胃穿刺采集瘤胃液,通过焦磷酸高通量测序技术进行瘤胃微生物多样性分析。结果表明A组微生物多样性大于B组,2组间微生物群落结构存在明显差异:A组以拟杆菌门(Bacteroidetes)和厚壁菌门(Firmicutes)为优势菌门,2者约占瘤胃微生物总数量的80%以上;而B组的主要菌门为拟杆菌门(Bacteroidetes)、厚壁菌门(Firmicutes)和变形菌门(Proteobacteria),占瘤胃中微生物中总数量的90%以上。即不同粗纤维源日粮对瘤胃微生物区系产生明显影响。     

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。     

复合菌剂发酵玉米秸秆对绵羊瘤胃液细菌多样性的影响

旨在通过16SrDNA高通量测序技术研究发酵玉米秸秆对绵羊瘤胃液细菌多样性的影响。本研究选用装有瘤胃瘘管的南非肉用美利奴♂×东北细毛羊♀的F1代成年公羊12只,随机平均分为两组,分别饲喂青贮玉米秸秆和发酵玉米秸秆。分别在饲喂前1天、饲喂第7、21天晨饲后6h取瘤胃液。同一时间所采取的同一处理的样品混合均匀,分别命名为青贮组(CS:YD0、YD7、YD21)和发酵组(FCS:YS0、YS7、YS21),之后通过高通量测序进行分析。结果显示:1)绵羊瘤胃内细菌群落包括29个门,74个纲,135个目,215个科和428个属。2)青贮组以拟杆菌门(Bacteroidetes)为优势菌群,其次为黏胶球形菌门(Lentisphaerae)、厚壁菌门(Firmicutes)和纤维杆菌门(Fibrobacteres),发酵组优势菌亦为拟杆菌门,其次为厚壁菌门、黏胶球形菌门和纤维杆菌门。通过21d的饲喂试验可知,青贮组拟杆菌门和厚壁菌门所占比例较发酵组降低,两组黏胶球形菌门和纤维杆菌门菌群数量都有所增加,但青贮组增加幅度更大。3)拟杆菌门的普雷沃氏菌属(Prevotella)在发酵组饲喂7d后成为最优势菌属(丰度达15.56%),而在青贮组明显降低(丰度从10.48%降至3.17%);纤维杆菌属(Fibrobacter)相对丰度在饲喂21d后青贮组增加量大于发酵组。4)在97%相似性水平下,饲喂21d后发酵组的Shannon和Simpson指数更高,表明它拥有更高的瘤胃细菌多样性。复合菌剂发酵玉米秸秆提高了绵羊瘤胃细菌多样性。饲喂发酵玉米秸秆的绵羊瘤胃内拟杆菌门和厚壁菌门菌群的比例降低,而黏胶球形菌门和纤维杆菌门菌群的数量增加。     

不同生理阶段荷斯坦奶牛瘤胃细菌多样性研究

本试验旨在探究不同生理阶段荷斯坦奶牛瘤胃细菌多样性。选取处于围产前期、泌乳前期、泌乳中期和泌乳后期的经产荷斯坦奶牛各4头,共16头,采集瘤胃液用于测定瘤胃发酵参数和提取微生物DNA,采用Illumina Miseq PE300平台测定瘤胃细菌组成。结果表明:1)围产前期奶牛瘤胃微生物蛋白、乙酸、丁酸、总挥发性脂肪酸浓度与乙酸/丙酸显著高于泌乳期(P0.05)。围产前期奶牛瘤胃丙酸浓度显著低于泌乳期(P0.05)。2)在门的水平上,围产前期奶牛瘤胃内的优势菌门为拟杆菌门和厚壁菌门,泌乳期奶牛瘤胃内的优势菌门则为拟杆菌门、变形菌门和厚壁菌门。围产前期奶牛瘤胃内的拟杆菌门、SR1细菌、蓝菌门、柔膜菌门、TM7细菌和纤维杆菌门的相对丰度显著高于泌乳期(P0.05),而泌乳期各阶段之间差异不显著(P0.05);围产前期奶牛瘤胃内的变形菌门的相对丰度显著低于泌乳期(P0.05)。3)在科的水平上,围产前期奶牛瘤胃内的优势菌科为普雷沃氏菌科,而泌乳期奶牛瘤胃内的优势菌科为琥珀酸弧菌科和普雷沃氏菌科。围产前期奶牛瘤胃内的普雷沃氏菌科、瘤胃菌科、理研菌科、BS11细菌、RF16细菌、SR1细菌、Gastranaerophilales、克里斯滕森菌科、TM7细菌、RF9细菌和纤维杆菌科的相对丰度显著高于泌乳期(P0.05);围产前期奶牛瘤胃内的琥珀酸弧菌科、毛螺菌科和韦荣球菌科则显著低于泌乳期(P0.05)。综上得出,奶牛从围产前期进入到泌乳期,瘤胃细菌多样性显著降低;泌乳期各阶段瘤胃细菌组成差异较小。     

利用16S rRNA测序、PCR-DGGE和qRT-PCR方法分析肉牛瘤胃上皮细菌群落特征

目前对于附着于瘤胃上皮的细菌群落及其功能了解较少,本研究目的是利用16S rRNA测序、PCR-DGGE和qRT-PCR方法比较分析肉牛瘤胃上皮与瘤胃内容物细菌群落的多样性差异.从6个16S rDNA(～1.4 kb)文库中随机选取2785条序列,结果表明3个瘤胃内容物文库与3个瘤胃上皮文库的细菌群落存在差异.以97％的序列相似度进行计算,结果发现瘤胃内容物菌群文库的多样性指数(4.12/4.42/4.88)比瘤胃上皮组织菌群文库的多样性指数(2.90/2.73/3.23)高.厚壁菌门和拟杆菌门分别是瘤胃上皮组织和内容物的优势菌群,纤维杆菌门、浮霉菌门和疣唯菌门仅在瘤胃内容物中存在.通过PCR-DGGE对22头阉牛瘤胃内容物和瘤胃上皮的细菌群落分析,进一步证实瘤胃上皮栖息着一类以厚壁菌为主的独特菌群,这类菌不仅具有消化饲料的作用,还有其他未知功能.     

健康鲁西黄牛消化道和阴道的细菌多样性研究

为了研究健康鲁西黄牛口腔、瘤胃、肠道和阴道的细菌组成及其相似性,分别提取各部位样本细菌的总DNA,特异扩增16Sr DNA的V3~V4高变区,扩增产物进行测序,对测序结果进行Chao1和Shannon Index多样性指数分析、OTUs交叠分析、聚类分析、物种分布波动性分析。结果发现,阴道和肠道的菌群多样性更高且比较相似,而口腔和瘤胃的则相对较低且比较相近;各样本中厚壁菌门、拟杆菌门所占的比例均较高;口腔和阴道的变形菌门较多,瘤胃的纤维杆菌门较多;阴道和肠道厚壁菌门占优势。结果表明:健康鲁西黄牛阴道和肠道细菌相似性很高并且含有相同的优势菌为厚壁菌门。阴道与口腔和瘤胃细菌的相似性很低,口腔、瘤胃和肠道的细菌相似性也很低。     

亚麻籽油及丙酸前体物质对延边黄牛瘤胃发酵特性及细菌区系多样性的影响

本试验通过调控日粮结构,研究其对延边黄牛瘤胃发酵特性及微生物区系的影响。选择8头年龄、体重(510±5.8)kg的安装有永久性瘤胃瘘管的延边黄牛,采用4×4拉丁方设计。试验分为CON组(基础日粮)、LO组(基础日粮+4%亚麻籽油)、LOM组(LO组+2%苹果酸)、LOF组(LO组+2%延胡索酸)。使用焦磷酸测序法对瘤胃内微生物基因组进行分析。结果表明:日粮中添加苹果酸和延胡索酸,可以增加瘤胃内的p H值、总挥发性脂肪酸的浓度及丙酸的摩尔比例;LOM组和LOF组,拟杆菌门(Bacteroidetes)与普氏菌属(Prevotella)在瘤胃细菌菌群中占主导地位;CON组和LO组,厚壁菌门(Firmicutes)和瘤胃球菌属(Ruminococcus)为主要的优势菌。由此可知,日粮中添加适当水平的亚麻籽油和丙酸前体物质,可以改善延边黄牛瘤胃发酵模式,并对瘤胃内微生物群落结构有较大的影响。     

添加菊粉和单宁对湖羊瘤胃挥发性脂肪酸和微生物区系的影响

研究旨在日粮中添加菊粉和单宁对湖羊瘤胃挥发性脂肪酸和瘤胃微生物区系的影响。试验采用单因素试验设计，选用体重相近、健康无病的断奶湖羊公羔24只，随机分为4组，每组6只。对照组饲喂基础饲粮，试验组Ⅰ、Ⅱ、Ⅲ组分别在基础饲粮中添加0.1%菊粉、0.1%菊粉+2%单宁、0.1%菊粉+2%单宁+4%聚乙二醇（polyethylene glycol,PEG）。饲养75 d后屠宰，采集瘤胃内容物用于测定瘤胃挥发性脂肪酸和微生物多样性。结果表明：试验Ⅱ组中乙酸/丙酸值显著提高（P<0.05）；试验Ⅲ组中异丁酸比例显著降低（P<0.05）。试验Ⅰ、Ⅱ、Ⅲ组中OTU数量、Chao1指数显著降低（P<0.05）。在门水平上，拟杆菌门、厚壁菌门和变形菌门的相对丰度占总菌门的95%以上。与对照组相比，试验Ⅰ、Ⅱ、Ⅲ组显著提高了拟杆菌门、变形菌门的相对丰度（P<0.05），显著降低了厚壁菌门、螺旋体门和纤维杆菌门的相对丰度（P<0.05）。在属水平上，与对照组相比，试验Ⅱ、Ⅲ组中理研菌科RC9菌属的相对丰度显著增加（P<0.05），而瘤胃球菌科NK4A214菌属的相对丰度显著降...     

短时高温好氧固态发酵对猪粪细菌群落结构的影响

为了研究畜禽粪便固态发酵前后微生物的变化,采用高通量测序技术,通过可操作分类单元分析、α多样性分析和物种组成分析,比较了猪粪短时高温好氧固态发酵前后的细菌群落组成,结果表明,发酵前后可操作分类单元总数分别为151个和126个,发酵后细菌群落多样性指数降低;发酵前相对丰度最高的分别是变形菌门59.48%、拟杆菌门23.97%和厚壁菌门13.79%,发酵后变形菌门和拟杆菌门分别减少至0.66%和1.00%,而厚壁菌门增加至99.01%,是发酵猪粪的优势菌群。本研究表明发酵后猪粪细菌群落多样性下降,抗生素抗性基因潜在携带者相对丰度降低,耐高温的有机质降解菌成为优势菌属,该研究对猪粪的资源化处理具有理论指导意义。     

基于16S rRNA高通量测序技术分析中国西门塔尔牛瘤胃微生物多样性和功能预测的研究

为系统探讨中国西门塔尔牛瘤胃内的微生物多样性及其功能,本试验利用16S rRNA基因高通量测序技术检测分析西门塔尔牛(20月龄,平均体重约为550 kg)瘤胃液样本菌群结构并进行PICRUSt功能预测。结果显示,两组样本通过Illumina Miseq测序平台共获得66 712条优质序列,聚类分析得到1 797个操作分类单元(OTU),经分类学鉴定分属13个门20个纲22个目33个科及59个属;厚壁菌门(Firmicutes)和拟杆菌门(Bacteroidetes)为优势菌群,所占比例分别为62.46%和22.45%;基于属的组成,依次为粪杆菌真核菌群([Eubacterium]_coprostanoligenes_group,9.16%)、瘤胃球菌属_2 (Ruminococcus_2,7.90%)、未知属f型拟杆菌目RF16菌群(norank_f__Bacteroidales_RF16_group,6.23%)、未知属f型瘤胃菌科(norank_f__Ruminococcaceae,4.79%)、理研菌科_RC9菌群(Rikenellaceae_RC9_gut_group,4.72%)、未知属f型拟杆菌目BS11菌群(norank_f_Bacteroidales_BS11_gut_group,4.49%)、瘤胃球菌属_1(Ruminococcus_1,4.43%)等;16S rRNA基因组的PICRUSt功能预测发现,瘤胃菌群功能主要集中在碳水化合物转运及代谢上,可能含有大量的纤维素和木质素降解酶基因。综上,基于16S rRNA基因的高通量测序技术全面揭示了西门塔尔牛瘤胃菌群的多样性,并预测其含有丰富的蛋白质分解、木质纤维素降解酶系,为探索西门塔尔牛瘤胃微生物奠定基础,也为进一步挖掘与某些重要营养生理功能密切相关的瘤胃微生物功能基因提供参考。     

Diversity Analysis of Rumen Microbes in Yunnan Gayals (Bos frontalis)

In order to study the rumen microbial diversity in Yunnan gayals (Bos frontalis), we analyzed the diversity of microbes using largely assembled sequence data obtained by metagenomic sequencing DNA isolated from rumen samples of eight adult galyals. The genome primary sequence (total metagenome size was >3 Gb) was assembled with short reads.Based on the gayal rumen microbial metagenomic sequencing, we received a total of 13 546 196 EGTs. Because of the relatively large amount, 10 387 566 EGTs (76.68% of all EGTs ) were randomly selected to analyze microbial communities by BLAST search of the NCBI-NR database. Microbial species annotations analysis revealed that microbial communities from gayals rumen were mainly composed of the bacterial domain, eukarya domain, archaea domain and unknown microbial composition. The majority microbes mainly belonged to ten phylums, and 94% EGTs belonged to Bacteroidetes, Firmicutes, Proteobacteria and Actinobacteria, of which the highest abundance phylums were Bacteroidete and Firmicutes, accounting for 48% and 42%, respectively. In addition, the main cellulolytic bacteria in rumen of gayals were Ruminococcus flavefacie, Butyrivibrio fibrisolvens, Ruminococcus albus and Fibrobacter succinogenes. They accounted for 1.60%, 1.20%, 0.86% and 0.52% of the total bacteria, respectively. These results further indicated that EGTs data could be used to analyze and explain environmental microbial community structure and function. There was great significance to study the composition of microorganisms in special environments.     

林蛙养殖场地土壤菌群结构及潜在病原菌分析

试验旨在了解林蛙养殖场地土壤菌群组成和结构特征、喷洒益生菌后的土壤菌群变化和潜在致病菌的种类与数量。试验设置3个组：养殖场地组、益生菌喷洒组和半人工养殖组,用高通量测序技术进行检测。3个试验组共获得191 588条有效序列,1 369个操作分类单元（operational taxonomic units,OTU）,经分类学鉴定分属于438个属,29个门。3组样本菌群数量较大,多样性水平也相对较高。优势菌门为变形菌门（Proteobacteria）、拟杆菌门（Bacteroidetes）、蓝细菌门（Cyanobacteria）、芽单胞菌门（Gemmatimonadetes）、绿弯菌门（Chloroflexi）、放线菌门（Actinobacteria）、酸杆菌门（Acidobacteria）、厚壁菌门（Firmicutes）;优势菌属为黄杆菌属（Flavobacterium）、热单胞菌属（Thermomonas）、Sphingopyxis、紫杆菌属（Porphyrobacter）、蓝细菌纲中未定种类（norank_c__Cyanobacteria）。各分组之间菌群多样性差异不显著,各组菌群结构相似度高。土壤中潜在的红腿病致病菌在属的水平有气单胞菌属（Aeromonas）、柠檬酸杆菌属（Citrobacter）、金黄杆菌属（Chryseobacterium）、假单胞菌属（Pseudomonas）。喷洒益生菌后土壤菌群变化不明显。本研究为全面了解东北林蛙养殖场地土壤微生物组成及变化、发掘有益微生物种类和东北林蛙疾病的预防和治疗提供理论依据。     

Analysis of Rumen Microbial Diversity and Functional Prediction of Chinese Simmental Cattle Based on 16S rRNA High-throughput Sequencing Technology

To systematically explore the microbial diversity and function in the rumen of Chinese Simmental cattle,this experiment used 16S rRNA gene high-throughput sequencing technology to detect and analyze the rumen fluid samples of Simmental cattle (about 20 months old,the average weight was 550 kg),and its function was predicted by PICRUSt.The results showed that a total of 66 712 high-quality sequences were obtained from the Illumina Miseq sequencing platform,and 1 797 operational classification units (OTU) were obtained by cluster analysis.The taxonomic identification was divided into 13 phylums,20 classes,22 orders,33 families and 59 genuses;Firmicutes and Bacteroidetes were dominant bacteria,accounting for 62.46% and 22.45%,respectively;Based on the genus composition,[Eubacterium]_coprostanoligenes_group (9.16%),Ruminococcus_2 (7.90%),norank_f__Bacteroidales_RF16_group(6.23%),norank_f__Ruminococcaceae(4.79%),Rikenellaceae_RC9_gut_group(4.72%),norank_f_Bacteroidales_BS11_gut_group(4.49%),Ruminococcus_1(4.43%),etc.The PICRUSt function of the 16S rRNA genome predicted that the rumen flora function was mainly concentrated in carbohydrate transport and metabolism,and might contain a large amount of cellulose and lignin degrading enzyme genes.In summary,the high-throughput sequencing technology based on 16S rRNA gene fully revealed the diversity of rumen bacteria in Simmental cattle,and predicted that it was rich in protein decomposition and lignocellulose degrading enzymes.The basis of microbial cognition provided a reference for further research on rumen microbial functional genes closely related to some important nutritional and physiological functions.     

基于16S rRNA高通量测序技术分析草原红牛瘤胃微生物多样性和功能预测的研究

为系统探讨草原红牛瘤胃内的微生物多样性及其功能,本试验利用16S rRNA基因高通量测序技术检测分析草原红牛(20月龄左右,均重为577.5 kg)瘤胃液样本菌群结构并进行PICRUSt功能预测。结果显示:通过Illumina Miseq测序平台共获得35 848条优质序列,聚类分析得到387个操作分类单元(OTU),经分类学鉴定分属15个门、20个纲、25个目、41个科及110个属;厚壁菌门(Firmicutes)和拟杆菌门(Bacteroidetes)为优势菌群,所占比例分别为50.09%和41.11%;基于属的组成,依次为普雷沃菌属(Prevotella)15.20%、未知属f型拟杆菌目(norankfBacteroidales)BS11菌群8.66%、瘤胃菌科(Ruminococcaceae)NK4A214菌群6.96%、理研菌科(Rikenellaceae)RC9菌群5.56%、未知属(Christensenellaceae)R-7菌群4.01%、瘤胃球菌属2(Ruminococcus2)3.33%等;16S rRNA基因组的PICRUSt功能预测结果显示,瘤胃内菌群功能主要集中在碳水化合物转运及代谢,表面草原红牛体内含有大量的纤维素和木质素降解酶基因。综上,基于16S rRNA基因的高通量测序技术全面揭示了草原红牛瘤胃菌群的多样性,且预测其含有丰富的蛋白质分解、木质纤维素降解酶系,为探索草原红牛瘤胃微生物的认知提供了基础,也为挖掘其他重要营养生理功能相关的瘤胃微生物功能基因提供了参考。     

Characterization of bovine ruminal epithelial bacterial communities using 16S rRNA sequencing, PCR-DGGE, and qRT-PCR analysis

Currently, knowledge regarding the ecology and function of bacteria attached to the epithelial tissue of the rumen wall is limited. In this study, the diversity of the bacterial community attached to the rumen epithelial tissue was compared to the rumen content bacterial community using 16S rRNA gene sequencing, PCR-DGGE, and qRT-PCR analysis. Sequence analysis of 2785 randomly selected clones from six 16S rDNA (～1.4kb) libraries showed that the community structures of three rumen content libraries clustered together and were separated from the rumen tissue libraries. The diversity index of each library revealed that ruminal content bacterial communities (4.12/4.42/4.88) were higher than ruminal tissue communities (2.90/2.73/3.23), based on 97% similarity. The phylum Firmicutes was predominant in the ruminal tissue communities, while the phylum Bacteroidetes was predominant in the ruminal content communities. The phyla Fibrobacteres, Planctomycetes, and Verrucomicrobia were only detected in the ruminal content communities. PCR-DGGE analysis of the bacterial profiles of the rumen content and ruminal epithelial tissue samples from 22 steers further confirmed that there is a distinct bacterial community that inhibits the rumen epithelium. The distinctive epimural bacterial communities suggest that Firmicutes, together with other epithelial-specific species, may have additional functions other than food digestion.     

日粮能量水平对湖羊瘤胃发酵特性和微生物组成的影响

本试验旨在探究不同能量水平对湖羊瘤胃发酵特性和微生物组成的影响。选取湖羊公羊48只（体重为（17.77±1.15） kg）随机分为2组：1）对照组（C组）,代谢能ME=8.83 MJ·kg^-1;2）高能组（H组）,ME=10.84 MJ·kg^-1。每组6个重复,每个重复4只羊。试验期105 d,其中预试期15 d,正试期90 d。试验结束后,采集瘤胃液测定瘤胃发酵参数,同时对微生物细菌V3-V4区进行16S rRNA高通量测序以检测瘤胃微生物组成。结果显示：1） H组和C组在瘤胃pH、总挥发性脂肪酸、乙酸、丙酸、异丁酸、丁酸、异戊酸、戊酸及乙丙比、氨态氮和微生物菌体蛋白浓度差异均不显著（P>0.05）;2）物种多样性结果显示,H组的Chao 1、Observed species、PD whole tree和Shannon指数均显著低于C组（P<0.05）;3）物种注释发现,拟杆菌门（Bacteroidetes）和厚壁菌门（Firmicutes）占比最高,但两者的相对丰度在H组和C组间差异均不显著（P>0.05）,H组的瘤胃球菌属（Ruminococcus）和厌氧弧菌属（Anaerovibrio）相对丰度均显著高于C组（P<0.05）。在本试验条件下,提高日粮能量水平对瘤胃发酵特性影响不显著,但是会降低瘤胃微生物的丰富度和均匀度,并改变部分菌属的相对丰度。     

The impact of rumen cannulation on the microbial community of goat rumens as measured using 16S rRNA high‐throughput sequencing

The main objective of this study was to determine the impact of rumen cannulation on the microbial community of goat rumens using 16S rRNA high‐throughput sequencing. Twelve Boer crossbred goats were used in the experiment: six goats were surgically fitted with rumen cannula, and the other six were used as controls. All goats were fed the same diet for 20 days, after which their rumen digesta were sampled once per week for three consecutive weeks. Total microbial DNA was extracted from the collected rumen fluid and was used as a template to amplify the V4 hypervariable region of the 16S rRNA gene. High‐throughput sequencing was performed using an Illumina MiSeq platform, and the sequences were analyzed primarily using the Quantitative Insights into Microbial Ecology pipeline software. The results showed that the Chao 1 index, the observed species index and the Shannon‐Wiener index were not significantly different (p > 0.05) between the two groups. Bacteroidetes, Firmicutes, Tenericutes were the predominant phylum in both groups, and their relative abundance was 60.63%, 29.48%, 2.24% (n = 6, CT group) and 61.17%, 26.92%, 1.66% (n = 6, RC group) respectively. At the phylum level, the relative abundance of Proteobacteria was significantly higher (p < 0.001) in the microbial communities of RC goats, and Planctomycetes and Chloroflexi were significantly lower (p = 0.02). The abundances of other phyla were not significantly different between treatments. A total of 19 lower‐level taxa also exhibited significant differences (p < 0.05) in relative abundance between the groups. In addition, there were 18 genera shared within the control group, 26 shared within the rumen‐cannulated group, and 16 shared by both groups. Prevotella was the most abundant shared genus, although its abundance was not significantly different (p > 0.05) between the groups. In conclusion, although the most abundant microbes kept stable, rumen cannulation had the potential to significantly change rumen microbial communities in goats.     

不同蛋白源日粮添加乳酸链球菌素对育肥湖羊瘤胃发酵及微生物菌群结构的影响

本试验旨在探究不同蛋白源日粮条件下添加乳酸链球菌素（nisin）对育肥湖羊瘤胃发酵及微生物菌群结构的影响。试验采用2×2因子设计,两因子分别为蛋白源[豆粕（SBM）、干玉米酒精糟及其可溶物（DDGS）]和nisin （添加水平为0或30.5 mg·kg^-1 DM）,两两组合,配制4种等氮等能日粮。选取32只体重（23±2） kg的断奶湖羊公羔,按照随机区组设计,根据体重分为2个区组（低体重组,16只;高体重组,16只）,每个区组的湖羊随机分配到4个组并分别饲喂对应日粮,单栏饲喂。试验预饲期1周,正式期9周,试验期结束时每组从高、低体重区组中各随机选取3只湖羊（共24只）进行屠宰,采集瘤胃内容物,提取微生物基因组DNA,利用Illumina MiSeq测序和Real-time qPCR方法分析瘤胃微生物菌群结构。研究结果表明,除Ruminococcaceae NK4A214 group和Unclassified Bacteroidetes相对丰度外,日粮蛋白源与nisin对其他所有测定指标（瘤胃发酵参数、瘤胃菌群数量、瘤胃细菌多样性及相对丰度等）均不存在交互作用（P>0.05）。日粮添加nisin对所有测定指标均无显著影响（P>0.05）。与饲喂豆粕湖羊相比,饲喂DDGS湖羊瘤胃乙酸、氨态氮浓度及总支链脂肪酸（BCVFA）浓度显著降低（P<0.05）。qPCR结果显示,饲喂不同蛋白源日粮湖羊瘤胃总菌、真菌及甲烷菌数量均无显著差异（P≥0.053）,但相对于饲喂豆粕湖羊而言,饲喂DDGS湖羊瘤胃内原虫和嗜氨梭菌（C.aminophilum）数量显著降低（P<0.05）。Illumina-MiSeq测序结果表明,饲喂DDGS湖羊瘤胃内细菌Chao1指数和ACE指数显著升高（P<0.05）。不同日粮处理组在门水平上的优势菌门均为Bacteroidetes及Firmicutes;在属水平各处理组的主要优势菌属为Prevotella 1、Christensenellaceae R-7 group和Ruminococcaceae NK4A214 group。饲喂不同蛋白源湖羊瘤胃细菌在门水平上并未产生显著影响（P≥0.14）;在属水平上,饲喂DDGS湖羊瘤胃Pseudobutyrivibrio和Roseburia丰度均显著高于饲喂豆粕湖羊（P<0.05）,而Butyrivibrio 2以及Ruminococcaceae UCG-005的相对丰度显著低于饲喂豆粕湖羊（P<0.05）。综上所述,使用DDGS作为日粮蛋白源改变了湖羊瘤胃发酵参数及微生物菌群结构,原虫和C.aminophilum数量的降低可能是导致瘤胃氨态氮浓度显著降低的主要原因;日粮添加30.5 mg·kg^-1 DM的nisin对育肥期湖羊瘤胃发酵参数及微生物菌群结构均无显著影响。     

Evaluating of the Association Between Early-stage Weight and Gut Microbiota in Duroc Pigs Using 16S rRNA Sequencing

The purpose of this study was to explore the possible association of the intestinal microbiome with early-stage weight in Duroc pigs.80 days old weaned piglets with Lighter body weight (LBW,n=9) and heavier body weight (HBW,n=9) were selected from 91 individuals based on their body weights ranked.Fecal samples from piglets in the early grower phase were performed for microbial diversity,composition and predicted functionality by sequencing the V3-V4 region of 16S rRNA gene.The results showed that there was no significant differences in microbial Alpha diversity between HBW and LBW groups(P>0.05).The two dominant phyla detected in both groups were Firmicutes (65.62% and 66.57% in HBW and LBW groups) and Bacteroidetes (30.87% and 29.80% in HBW and LBW groups).Four dominant genera were Prevotella (23.82% and 20.84% in HBW and LBW groups),Lactobacillus (9.11% and 16.55% in HBW and LBW groups),norank_f_Ruminococcaceae (10.32% and 9.98% in HBW and LBW groups),and norank_f_Lachnospiraceae (6.33% and 4.53% in HBW and LBW groups).PCoA analysis showed that there were small differences in gut microbes among Duroc piglets,whereas,some weight-associated microbial compositional differences were revealed between HBW and LBW groups.At the genus level,the abundance of several genera,such as g_Kitasatospora,g_norank_o_Tremblayales,g_norank_f_Erysipelotrichaceae,g_unclassified_f_[Paraprevotellaceae] and g_Coprobacillus,were significantly higher in HBW group than in LBW group (P<0.05).The abundance of Corynebacterium in LBW group was significantly higher than that in HBW group (P<0.05).Moreover,the KEGG pathways analysis showed that the sesquiterpenoid biosynthesis,insulin signaling pathway,ascorbate and aldarate metabolism,and biosynthesis of ansamycins were significantly enriched in HBW group (P<0.05),however,the secretion system pathway was significantly enriched in LBW group (P<0.05).These results might provide insights into host-microbe interactions occurring in the early stage piglets and would be useful in contributing to healthy pig breeding.     

巨菌草内生细菌多样性及其促生特性

为探究不同生境巨菌草（Pennisetum sp.）内生细菌群落组成多样性,明确促生菌株的促生特性,本试验采用菌株16S rDNA全序列分析法,对5个不同生境的巨菌草中内生细菌的遗传多样性进行系统发育分析,并以溶磷、固氮、产吲哚乙酸（Indoleacetic Acid,IAA）、产铁载体和抑菌活性等为筛选标准,对初筛菌株进行多项促生活性测定。本研究中,从巨菌草植株中共分离得到187株内生菌株,其中86株具有分泌IAA的能力,占全部菌株的46%,菌株XJ-4产IAA的能力最强,分泌的IAA浓度为30.13 mg·L^-1;47株具有溶磷能力,占全部菌株的25%,菌株FJ-23,FJ-15溶磷能力最高,分别为7.10 mg·L^-1和7.35 mg·L^-1;128株可以在Ashby无氮培养基中正常生长,表明64%的巨菌草内生细菌都具有潜在的固氮能力;34株可以在产铁载体的培养基中正常生长,表明18%的菌株产铁载体;试验中,80%以上的内生细菌对1种或者多种指示菌具有抑制作用,其中菌株SX-21,SX-30,SX-1,NY-10表现出对3种指示菌的广谱性抑菌活性,菌株SX-30对玉米大斑病菌的抑制效果最好,抑菌圈达到30 mm。供试菌株共产生了23种遗传图谱类型,通过对每种图谱类型的代表性菌株进行16S rDNA全序列分析,23株菌株分别归属于芽孢杆菌属（Bacillus）、根瘤菌属（Rhizobium）、寡养单胞菌属（Stenotrophomonas）、假单胞菌属（Pseudomonas）、不动杆菌属（Acinetobacter）和肠杆菌属（Enterobacter）。巨菌草中具促生特性的内生细菌表现出丰富的多样性,可为进一步构建巨菌草促生菌菌群提供良好的种质资源。