奶牛瘤胃中一株纤维分解菌的筛选与鉴定

试验从奶牛瘤胃中分离得到1株纤维分解菌,研究其生化性质和种属分类.将富集的纤维分解菌群,通过厌氧平板分离培养得到纤维分解菌.利用全自动微生物分析系统的革兰氏阴性菌鉴定卡鉴定纤维分解菌的生化图谱,并对菌株16SrDNA进行PCR扩增并测序,与GenBank数据库中序列进行同源性比对,确定其种属分类地位.结果表明:通过体外培养,分离得到纤维分解菌CB11,纤维分解平均消失率达82%,革兰氏染色为阴性,绝对厌氧,能够产生β-半乳糖苷酶、β-葡萄糖苷酶、β-木糖苷酶等.通过建立系统发育树,CB11的16SrDNA序列与解肝素拟杆菌的相似度达99%.     

ISCA对瘤胃挥发性脂肪酸代谢及纤维素降解率的影响

通过5只装有瘤胃瘘管的绵羊，观察一种含铵的工业废液（ISCA）作为非蛋白氮饲料对瘤胃糖代谢的影响。采用自身对照，测定了添喂了ISCA前后瘤胃内挥发性脂肪酸（VFA）的浓度，PH值，温度及饲草中干物质（DM）和酸性洗涤纤维（ADF）的消失率等指标，并且进行了病理学检查，结果表明，每只羊日添喂ISCA 100ml，除饲后2h试验期乙酸／丙酸比例显著低于对照组（P＜0．05），外，上述各指标两期间均无显著差异，结果提示，ISCA对瘤胃糖类的降解与发酵过程及瘤胃内环境无不良影响，对消化道粘膜上皮和各主要实质脏器无毒副作用和直接损害。     

高产纤维降解酶牦牛瘤胃厌氧真菌分离株的筛选与鉴定

为获得高产纤维降解酶瘤胃厌氧真菌,从放牧饲养条件下随机选出10头屠宰后的青海大通牦牛瘤胃中采集食糜样本,通过亨盖特滚管技术筛选得到29株厌氧真菌菌株.以小麦秸秆为底物,测定这些菌株第6天培养液中乙酰酯酶、阿魏酸酯酶、木聚糖酶、羧甲基纤维素酶和微晶纤维素酶的活性.29株真菌之间5种酶活性差异显著,阿魏酸酯酶活性最高的菌株比活性最低的菌株活性高出7.9倍,差异最显著;而羧甲基纤维素仅高出2.5倍,差异最不显著.对各菌株5种酶活性进行相关分析,乙酰酯酶、阿魏酸酯酶、木聚糖酶和羧甲基纤维素酶相互之间均存在显著的正相关性(r>0.67,P<0.0001),而且它们之间的相关性高于其与微晶纤维素间的相关性(P>0.05).从中筛选得到1株高产纤维降解酶厌氧真菌YAK11,该菌株第6天培养液中5种酶活性分别达到165、10、1138、83和16 mU.通过18S rDNA基因序列分析该株菌,初步确定为Neocallimastix,frontalis.     

3种生理调节剂对山羊瘤胃液中脂肪酸组成的影响

研究3种生理调节剂对山羊瘤胃液中共轭亚油酸(CLA)前体反式油酸的积累与CLA含量的影响,选用8只装有永久性瘤胃瘘管的山羊,采用4×4拉丁方设计,试验设对照、葵花油、海南霉素和丙酮酸钙4个处理,每只山羊每天添喂剂量分别为0、0.3、0.05、0.05 g/kg体重。每两周采集瘤胃液样品,采用毛细管气相色谱法分析瘤胃液中反式油酸、CLA含量及其他脂肪酸组成,并对CLA与反式油酸含量进行了相关性分析。结果表明,3种生理调节剂都能在不同程度上提高C18∶1 trans-11(TVA)和CLA含量,其作用效果依次为葵花油>海南霉素>丙酮酸钙。瘤胃中TVA和CLA含量呈极显著正相关(P<0.01)。     

牛瘤胃臌胀的诊治

通过几年的临床实验,应用澄清石灰水、加入已炸开的花生油（菜油）、烟丝、大蒜、食用醋等药品结合瘤胃放气法治疗瘤胃臌胀,在东兴、思恩镇发现及治疗该病45头,治愈42头,治愈率达93．3％,是个简单、快速、高效的治疗方法。     

Effects of formic acid and corn flour supplementation of banana pseudostem silages on nutritional quality of silage,growth, digestion,rumen fermentation and cellulolytic bacterial community of Nubian black goats

The objective of this study was to evaluate the effects of adding formic acid and corn flour supplementation to banana pseudostem silages on the nutritional quality of these silages, growth, digestion, rumen fermentation and cellulolytic bacterial community of Nubian black goats fed these silages. Banana pseudostem silage was prepared either conventionally without any additives(CON) or mixed with 0.6% formic acid(F), 10% corn flour(C), or both(F+C). Four experimental diets containing 40% of the corresponding silages were designed with roughage to concentrate ratio of 50:50(dry matter(DM) basis). A total of 48 Nubian black castrated goats(body weight(BW),(22.64±1.82) kg; 4-mon-old) were randomized into one of the four treatment groups with 12 replicates of one castrated goat per replicate for each treatment in a completely randomized design. Each group was fed on one of the four experimental diets for 40 days. A factorial arrangement of treatments of 2(formic acid levels: 0 and 0.6%)×2(corn flour: 0 and 10%) was adopted. Formic acid supplementation increased(P0.05) average daily gain, as well as lactic acid, propionate and butyric acid and water-soluble carbohydrate concentrations, but decreased(P0.05) the feed conversion rate, p H value, acetate/propionate ratio, and butyric acid concentration relative to the CON group. Corn flour supplementation increased(P0.05) the apparent digestibility of crude protein, neutral detergent fiber, and non-fibrous carbohydrate and Fibrobacter succinogenes, Ruminococcus albus, and Butyrivibrio fibrisolvens populations, but decreased(P0.05) the Ruminococcus flavefaciens population relative to the CON group. There were no F×C treatment interactions(P0.05) for any of the other indices except for the apparent digestibility of non-fibrous carbohydrate(NFC)(P0.05). The results demonstrated that adding 0.6% formic acid and 10% corn flour supplementation to banana pseudostem silages improved the nutritional quality of these silages and enhanced the growth performance of Nubian black castrated goats by improving apparent nutrient digestibility, and regulating ruminal fermentation and bacteria populations.     

不同水分TMR对绵羊瘤胃纤维分解酶活力与微生物生态的影响

为探讨不同水分的全混合日粮(TMR)对绵羊瘤胃纤维分解酶活力和微生物生态的影响,以延边半细毛羊为为试材,手术安装瘤胃瘘管以接取瘤胃液进行试验,喂不另加水分的TMR日粮(A组)、加35%水分的TMR日粮(B组)、加50%水分的TMR日粮(C组)共3组,检测瘤胃pH值、瘤胃内纤毛虫数、瘤胃内酶活力。结果表明,瘤胃pH值A组略低于B、C组但各组之间差异不显著,瘤胃纤毛虫数和瘤胃内酶活力的变化与pH值的变化呈一致性,即不同水分的TMR直接影响瘤胃内pH值的变化,利于维持瘤胃内pH的平衡;间接影响瘤胃内纤毛虫数和酶活力,这有助于维持瘤胃内微生物生态环境的稳定。     

Impacts of the unsaturation degree of long-chain fatty acids on the volatile fatty acid profiles of rumen microbial fermentation in goats in vitro

This study investigated the impacts of the degree of unsaturation(unsaturity) of long-chain fatty acids on volatile fatty acid(VFA) profiles of rumen fermentation in vitro. Six types of long-chain fatty acids, including stearic acid(C18:0, control group), oleic acid(C18:1, n-9), linoleic acid(C18:2, n-6), α-linolenic acid(C18:3, n-3), arachidonic acid(C20:4, n-6) and eicosapentaenoic acid(C20:5, n-3), were tested. Rumen fluid from three goats fitted with ruminal fistulae was used as inoculum and the inclusion rate of long-chain fatty acid was at 3%(w/w) of substrate. Samples were taken for VFA analysis at 0, 3, 6, 9, 12, 18 and 24 h of incubation, respectively. The analysis showed that there were significant differences in the total VFA among treatments, sampling time points, and treatment×time point interactions(P0.01). α-Linolenic acid had the highest total VFA(P0.01) among different long-chain fatty acids tested. The molar proportion of acetate in total VFA significantly differed among treatments(P0.01) and sampling time points(P0.01), but not treatment×time point interactions(P0.05). In contrast, the molar proportion of propionate did not differ among treatments during the whole incubation(P0.05). However, for butyrate molar proportions, significant differences were found not only among sampling time points but also among treatments and treatment×time point interactions(P0.01), with eicosapentaenoic acid having the highest value(P0.01). Additionally, no statistically significant differences were found in the acetate to propionate ratios among treatments groups(P0.05), even the treatments stearic acid and α-linolenic acid were numerically higher than the others. The inclusion of 3% long-chain unsaturated fatty acids differing in the degree of unsaturation brought out a significant quadratic regression relation between the total VFA concentration and the double bond number of fatty acid. In conclusion, the α-linolenic acid with 3 double bonds appeared better for improving rumen microbial fermentation and the total VFA concentration.     

相同亚油酸水平日粮对绵羊瘤胃液和血液脂肪酸组成的影响

为提高反刍动物产品中CLA含量寻找植物油的有效添加模式,研究以植物油调控的相同亚油酸水平日粮对绵羊瘤胃液脂肪酸动态变化和血浆脂肪酸组成的影响.选取4只雄性、健康安装有永久性瘤胃瘘管的成年绵羊(东北半细毛羊×陶赛特羊),体重(40±1 kg),采用4×4拉丁方试验设计,研究不添加油脂组(CK组)、添加4％花生油(PO组)...     

奶牛瘤胃上皮细胞分离培养与鉴定

在体外分离培养奶牛瘤胃上皮细胞,可为奶牛瘤胃生理功能和吸收机制研究和永生化奶牛瘤胃上皮细胞奠定基础。通过胰蛋白酶消化法从奶牛瘤胃组织中分离出瘤胃上皮细胞,并通过免疫细胞化学的方法检测细胞角蛋白,CCK-8检测细胞生长曲线,β-半乳糖苷酶染色鉴定细胞的衰老以及细胞染色体核型分析来鉴定瘤胃上皮细胞。结果表明:1)利用胰蛋白酶消化法可以稳定的培养出瘤胃上皮细胞并且能够在体外传代大约5代;2)通过在显微镜下进行细胞形态的观察呈单层"岛屿状";3)在荧光显微镜下细胞角蛋白抗原能够发出绿色荧光;4)奶牛瘤胃上皮细胞传至第5代,β-半乳糖苷酶被染成蓝色,同时,细胞出现衰老和停滞生长;5)染色体核型分析表明奶牛瘤胃上皮细胞的染色体数为60条。研究发现通过酶消化奶牛瘤胃组织能够获得奶牛瘤胃上皮细胞,但只能传至5代。     

富硒益生菌对山羊瘤胃发酵和瘤胃液脂肪酸含量的影响

为研究日粮中添加富硒益生菌(SP)对山羊瘤胃发酵和瘤胃液脂肪酸含量变化的影响,以8头装有永久性瘘管的山羊为对象,采用4×4拉丁方设计,进行4期试验,每期15 d。日粮中分别添加0(对照)、8(LSP)、16(MSP)和32(HSP)mL.d-1的SP。结果表明:日粮添加SP能显著影响山羊瘤胃pH值(P<0.01)、总挥发性脂肪酸(P<0.05)、微生物蛋白(P<0.05)和trans11-C18∶1浓度(P<0.05);并有升高瘤胃乙酸比例(P=0.08)、α-C18∶3浓度(P=0.07)以及降低瘤胃丁酸比例(P=0.05)、氨氮浓度(P=0.08)的趋势。说明日粮添加SP对缓解瘤胃pH值的下降,促进瘤胃总挥发性脂肪酸的产生、微生物蛋白的合成和对氨氮的利用,以及提高瘤胃液α-C18∶3和共轭亚油酸前体trans11-C18∶1的含量有积极作用。     

In vitro investigation of the effect of dairy propionibacteria on rumen pH,lactic acid and volatile fatty acids

Ruminal acidosis is a prevalent disorder in ruminants such as dairy cows and feedlot beef cattle, caused primarily by the inclusion of a high percentage of readily fermentable concentrates in the diet. The disorder presents as an accumulation of lactic acid, a decrease of p H in the rumen and a subsequent imbalance of the rumen fermentation process with detrimental impacts on the animal's health and productivity. Dairy propionibacteria, a group of bacteria characterised by utilization of lactic acid as the favoured carbon source, with propionic acid produced as a by-product, were evaluated in this study as potential direct-fed microbials for use in controlling ruminal acidosis. Acidosis was simulated by introduction of high concentrations of lactic acid into rumen fluid samples and a multi-strain in vitro analysis was conducted, whereby changes in p H and lactic acid metabolism were compared in identical acidified rumen samples, following inoculation with various propionibacteria. This was followed by a study to evaluate the effect of bacterial inoculation dosage on acid metabolism. The results indicated that lactic acid levels in the rumen fluid were significantly reduced, and propionic acid and acetic acid concentrations both significantly increased, following addition of propionibacteria. Significant ‘between strains' differences were observed, with Propionibacterium acidopropionici 341, Propionibacterium freudenreichii CSCC 2207, Propionibacterium jensenii NCFB 572 and P. jensenii 702 each producing more rapid reduction of lactic acid concentration than P. freudenreichii CSCC 2206, P. acidopropionici ATCC 25562 and Propionibacterium thoenii ATCC 4874. Furthermore, the efficacy of this application was dosage related, with the rates of reduction in lactic acid levels and production of propionic acid, both significantly greater for the higher(1010 cfu mL~(-1)) compared with lower(105 cfu mL~(-1)) dosage inoculation. The results confirmed that the introduction of propionibacteria could promote more rapid reduction of lactic acid levels than would occur without their addition, demonstrating their potential in controlling ruminal acidosis.     

不同植物激素对原始小球藻生长及油脂含量的影响

研究了不同植物激素(IBA、NAA、6-BA、2,4-D、TDA)对原始小球藻(Chlorella protothecoides)的生长及脂肪酸组成的影响。使用三角瓶进行静置培养,在培养基中添加0.5 mg/L的不同植物激素,结果表明:培养基中添加不同的植物激素对原始小球藻的生长有不同的影响,以2,4-D的促进效应最为明显,最终干重为1.18 g/L,是空白对照的1.2倍;不同植物激素对粗油脂含量的影响不同,其中添加2,4-D的培养液中所收获的藻粉油脂含量最高,为藻粉干重的19.74%,比空白对照高26.21%;对粗油脂进行GC-MS分析,结果显示不同植物激素的添加对于小球藻脂肪酸成分的影响不大,但略微提升了C16和C18脂肪酸的相对含量,使之更加适合作为生物柴油的来源。结合总脂收获量的高低及脂肪酸成分的不同,本试验最终确定2,4-D为小球藻培养基最理想的植物激素类添加物。     

The effect of lactic acid bacteria inoculums on in vitro rumen fermentation,methane production,ruminal cellulolytic bacteria populations and cellulase activities of corn stover silage

The objective of this study was to investigate the effect of lactic acid bacteria(LAB) inoculums on fermentation quality and in vitro digestibility of corn stover silage. Corn stover was ensiled without(control) or with Lactobacillus plantarum(LP), Enterococcus faecalis(EF), and Enterococcus mundtii(EM) for 45 days. The fermentation characteristics were assessed, and subsequent in vitro dry matter digestibility(DM-D), neutral detergent fiber digestibility(NDF-D), volatile fatty acids(VFA), methane(CH_4) production, cellulolytic bacteria proportions and their activities per corn stover silage were also determined. There was no significant difference(P0.05) among the silage pH, lactic acid, crude protein(CP), water soluble carbohydrates(WSC) and lignocelluloses contents of different treatments. The relative proportions of Ruminococcus flavefaciens and Fibrobacter succinogenes, carboxymethyl-ocellulose and β-glycosidase activities, DM-D, NDF-D, and VFA production of in vitro incubation was higher(P0.05) for silages inoculated with LP and EF than those of the control silage. Silage inoculated with LP showed the lowest(P0.05) CH_4 production per unit yield of VFA, which was positively corresponded to the lowest(P0.05) ratio of acetate to propionate. In summary, the ensiling fermentation quality and subsequent utilization of corn stover silage were efficiently improved by inoculated with L. plantarum.     

不同生育期串叶松香草和聚合草氨基酸瘤胃降解特性研究

[目的]研究不同生育期串叶松香草和聚合草氨基酸的瘤胃降解特性,为牧草适宜收获期的确定及其资源利用提供依据.[方法]以6头装有永久瘤胃瘘管的小尾寒羊为试验动物,用尼龙袋法测定不同生育期串叶松香草和聚合草在绵羊瘤胃降解前后氨基酸含量的变化,研究氨基酸的瘤胃降解特性.[结果](1)随生育期的推进,串叶松香草和聚合草中氨基酸含...     

日粮添加亚油酸和二十二碳六烯酸对奶牛瘤胃原虫奇数碳链脂肪酸和支链脂肪酸的影响

试验选取4头安装有永久性瘤胃、十二指肠和回肠瘘管的荷斯坦泌乳奶牛,采用4×4拉丁方试验设计,研究日粮中添加亚油酸(LA)和二十二碳六烯酸(DHA)对瘤胃原虫中奇数碳链脂肪酸和支链脂肪酸的影响.试验结果表明:奇数碳链脂肪酸和支链脂肪酸(OBCFA)是原虫脂肪酸的重要组成部分,与对照组相比,日粮中单独添加LA或DHA对瘤胃原虫中主要OBCFA的含量无显著影响,而当LA和DHA协同添加时C15∶0和C17∶0的含量显著降低(P0.05),但对异构酸和反异构酸的含量无显著影响.     

黄河滩区2种禾本科牧草不同生育期氨基酸瘤胃降解特性研究

【目的】研究多年生黑麦草和紫羊茅不同生育期氨基酸的瘤胃降解特性。【方法】以6头装有永久瘤胃瘘管的小尾寒羊为试验动物,用尼龙袋法测定不同生育期多年生黑麦草和紫羊茅在绵羊瘤胃降解前后粗蛋白、总氨基酸及各种氨基酸含量的变化。【结果】1)除开花期外,多年生黑麦草在各个生育期的粗蛋白及总氨基酸含量均高于紫羊茅;2种牧草都富含脯氨酸、谷氨酸、天冬氨酸、亮氨酸和缬氨酸,而含硫氨基酸含量较低;粗蛋白和总氨基酸含量均随生育期的延长呈下降趋势,即在分蘖期最高,在成熟期最低。2)2种牧草各氨基酸在分蘖期易降解,而在成熟期不易降解。3)2种牧草中的甘氨酸、脯氨酸、酪氨酸、精氨酸、苏氨酸、赖氨酸和苯丙氨酸在瘤胃中较易降解(P<0.05);多年生黑麦草中的天冬氨酸、丝氨酸、谷氨酸、缬氨酸、异亮氨酸和亮氨酸在瘤胃中不易降解(P<0.05),紫羊茅中的丝氨酸、异亮氨酸和亮氨酸在瘤胃中不易降解(P<0.05)。【结论】2种牧草的粗蛋白和氨基酸含量受品种和生育期影响较大,氨基酸在瘤胃中的降解程度与牧草生育期有关。     

苏子油对瘤胃体外发酵模式、脂肪酸组成及甲烷生成的影响

为探讨苏子油对瘤胃体外发酵的影响,采用人工瘤胃技术分析不同添加水平的苏子油对瘤胃发酵模式、发酵参数和瘤胃脂肪酸代谢的影响.结果表明:添加不同剂量苏子油对发酵液pH值和氨氮(NH3-N)含量无显著影响(P0.05);与对照相比较,随着苏子油添加量的增加干物质降解率、中性洗涤纤维降解率、酸性洗涤纤维降解率、发酵底物单位产气量显著降低(P0.05),甲烷产量呈下降趋势(P0.05);对TVFA无显著影响(P0.05),丙酸含量显著增加(P0.05),乙酸含量和乙酸丙酸比例显著降低(P0.05);C18:1、c9,t11-CLA和总的长链脂肪酸浓度均明显升高(P0.01).表明在粮中添加一定量的苏子油,可改变瘤胃发酵类型、降低甲烷产量、调节瘤胃液脂肪酸组成.