稀释率对新型固液气分流式瘤胃模拟系统发酵效果的影响

本试验旨在研究稀释率对新型固液气分流式瘤胃模拟系统发酵效果的影响.采用单因子完全随机试验设计,共设4个处理,各处理稀释率分别为6％/h、8％/h、10％/h、12％/h,每个处理3个重复,每个重复1个发酵罐.试验期10 d.结果表明:随着稀释率的提高,发酵液pH极显著提高(P＜0.01),而氨态氮(NH3-N)浓度则极显著降低(P＜0.01).稀释率由6％/h上升到10％/h,总挥发性脂肪酸(TVFA)及戊酸浓度极显著下降(P＜0.01).稀释率从6％/h提高到8％/h时,原虫数量极显著提高了6.91％ (P ＜0.01),但当稀释率从8％/h提高到12％/h时,原虫数量却极显著下降了5.18％ (P ＜0.01).随着稀释率从6％/h增加到10％/h,木聚糖酶活性(P＜0.01)及微生物蛋白质(MCP)产量(P＜0.05)显著或极显著降低,粗蛋白质体外消失率极显著升高(P＜0.01).稀释率为10％/h和12％/h的2个处理,中性洗涤纤维体外消失率显著高于另外2个处理(P＜0.05).结果提示,稀释率能够影响新型固液气分流式瘤胃模拟系统的发酵情况,稀释率为8％/h时,原虫数量以及纤维素类酶活性、各瘤胃发酵参数更加有利于发酵,因此可推荐作为本系统最优运行条件.     

Effects of the ruminal comminution rate and microbial contamination of particles on accuracy of in situ estimates of ruminal degradability and intestinal digestibility of feedstuffs

Effects of considering the comminution rate (k_c) and the correction of microbial contamination (using ¹⁵N techniques) of particles in the rumen on estimates of ruminally undegraded fractions and their intestinal digestibility were examined generating composite samples (from rumen‐incubated residues) representative of the undegraded feed rumen outflow. The study used sunflower meal (SFM) and Italian ryegrass hay (RGH) and three rumen and duodenum cannulated wethers fed with a 40:60 RGH to concentrate diet (75 g DM/kgBW^0.75). Transit studies up to the duodenum with Yb‐SFM and Eu‐RGH marked samples showed higher k_c values (/h) in SFM than in RGH (0.577 vs. 0.0892, p = 0.034), whereas similar values occurred for the rumen passage rate (k_p). Estimates of ruminally undegraded and intestinal digestibility of all tested fractions decreased when k_c was considered and also applying microbial correction. Thus, microbial uncorrected k_p‐based proportions of intestinal digested undegraded crude protein overestimated those corrected and k_c?k_p‐based by 39% in SFM (0.146 vs. 0.105) and 761% in RGH (0.373 vs. 0.0433). Results show that both k_c and microbial contamination correction should be considered to obtain accurate in situ estimates in grasses, whereas in protein concentrates not considering k_c is an important source of error.     

Effects of replacing dietary starch with neutral detergent–soluble fibre on ruminal fermentation,microbial synthesis and populations of ruminal cellulolytic bacteria using the rumen simulation technique (RUSITEC)

A rumen simulation technique (RUSITEC) apparatus with eight 800 ml fermenters was used to investigate the effects of replacing dietary starch with neutral detergent–soluble fibre (NDSF) by inclusion of sugar beet pulp in diets on ruminal fermentation, microbial synthesis and populations of ruminal cellulolytic bacteria. Experimental diets contained 12.7, 16.4, 20.1 or 23.8% NDSF substituted for starch on a dry matter basis. The experiment was conducted over two independent 15‐day incubation periods with the last 8 days used for data collection. There was a tendency that 16.4% NDSF in the diet increased the apparent disappearance of organic matter (OM) and neutral detergent fibre (NDF). Increasing dietary NDSF level increased carboxymethylcellulase and xylanase activity in the solid fraction and apparent disappearance of acid detergent fibre (ADF) but reduced the 16S rDNA copy numbers of Ruminococcus albus in both liquid and solid fractions and R. flavefaciens in the solid fraction. The apparent disappearance of dietary nitrogen (N) was reduced by 29.6% with increased dietary NDSF. Substituting NDSF for starch appeared to increase the ratios of acetate/propionate and methane/volatile fatty acids (VFA) (mol/mol). Replacing dietary starch with NDSF reduced the daily production of ammonia‐N and increased the growth of the solid‐associated microbial pellets (SAM). Total microbial N flow and efficiency of microbial synthesis (EMS), expressed as g microbial N/kg OM fermented, tended to increase with increased dietary NDSF, but the numerical increase did not continue as dietary NDSF exceeded 20.1% of diet DM. Results suggested that substituting NDSF for starch up to 16.4% of diet DM increased digestion of nutrients (except for N) and microbial synthesis, and further increases (from 16.4% to 23.8%) in dietary NDSF did not repress microbial synthesis but did significantly reduce digestion of dietary N.     

日粮不同NDF/NFC水平对周岁后荷斯坦奶牛生产性能、营养物质消化率、瘤胃发酵特征和甲烷排放的影响

本试验研究了日粮中不同中性洗涤纤维/非纤维性碳水化合物（NDF/NFC）水平对周岁后荷斯坦奶牛生产性能、营养物质消化率、瘤胃发酵特征及甲烷产量的影响，并在此基础上建立了甲烷排放预测模型，旨在获得我国生产模型下的甲烷排放规律和甲烷转化因子，为提高奶牛能量利用效率、建立国家或区域性温室气体排放清单和探索减排策略提供科学依据和支撑。将45头体况良好，平均为15月龄的荷斯坦后备奶牛随机分为3组，每组15头牛：低日粮NDF/NFC组（NDF/NFC=0.60）、中日粮NDF/NFC组（NDF/NFC=0.75）和高日粮NDF/NFC组（NDF/NFC=0.90），试验期为70 d，包括14 d的预饲期和56 d的正试期。结果表明：1）提高日粮NDF/NFC水平显著降低了奶牛的干物质采食量、有机物采食量、平均日增重、干物质和粗蛋白的表观消化率（P<0.05）；2）提高日粮NDF/NFC水平显著增加了瘤胃内总挥发性脂肪酸产量、乙酸的相对含量和乙酸/丙酸比例（P<0.05），显著降低了丙酸的相对含量（P<0.05）；3）随着日粮NDF/NFC水平的提高，瘤胃甲烷和甲烷能产量、甲烷/代谢体重、甲烷/干物质采食量、甲烷/有机物采食量、甲烷/中性洗涤纤维采食量显著提高（P<0.05）。甲烷转化因子也随着日粮NDF/NFC水平的增加而显著提高（P<0.05）；4）基于体重、采食量、营养物质含量和NDF/NFC分别建立了甲烷预测模型，其中基于干物质采食量和中性洗涤纤维采食量建立的预测模型的决定系数最高（R²=0.77）。因此，提高日粮中NDF/NFC水平可显著降低周岁后荷斯坦奶牛的生产性能、营养物质消化率和瘤胃内丙酸的相对含量，可显著提高瘤胃甲烷产量和甲烷转化因子。     

实时荧光定量PCR对瘤胃纤维分解菌定量方法的构建

试验构建了白色瘤胃球菌、黄色瘤胃球菌及产琥珀酸丝状杆菌等3种瘤胃纤维分解菌实时荧光绝对定量PCR的标准品及标准曲线,以用于纤维分解菌的定量测定。提取瘤胃微生物总DNA,以各菌特异性引物进行PCR扩增,回收纯化PCR产物,与pMD18-T Vector连接并转化到大肠杆菌。用氨苄青霉素培养基筛选阳性重组质粒,提取含目的片段质粒DNA,通过PCR及测序鉴定重组质粒。根据OD值确定浓度,将梯度稀释的质粒作为模板,进行荧光定量PCR反应做出标准曲线。结果表明:所构建的3条标准曲线具有很高的相关性(R2>0.999),并获得了高扩增效率产物(白色瘤胃球菌为101%、黄色瘤胃球菌为98.0%、产琥珀酸丝状杆菌为97.7%),说明成功构建了瘤胃纤维分解菌实时绝对定量PCR的标准品和标准曲线,为定量研究瘤胃纤维分解菌奠定基础。     

Effects of Antidesma thwaitesianum Muell. Arg. pomace as a source of plant secondary compounds on digestibility,rumen environment,hematology, and milk production in dairy cows

Mao pomace meal (MPM) contains condensed tannins and saponins at 92 and 98 g/kg, respectively, and these substances can be used to manipulate ruminal fermentation in ruminant. Four multiparous lactating Holstein cows with 45 ± 5 days in milk were randomly assigned according to a 4 × 4 Latin square design to receive four different levels of MPM supplementation at 0, 100, 200, and 300 g/head/day, respectively. Cows were fed with concentrate diets at 1:1.5 of concentrate to milk yield ratio and urea‐treated (3%) rice straw was fed ad libitum. The results revealed that feed intake, nutrient digestibility, blood urea nitrogen, and hematological parameters were not affected by MPM supplementation (p > 0.05). However, ruminal pH and propionate were increased quadratically (p < 0.05) in cows receiving MPM whereas acetate, acetate to propionate ratio and estimate methane production were decreased (p < 0.05). Supplementation of MPM linearly decreased ruminal ammonia nitrogen and protozoal population at 4 hr postfeeding (p < 0.05). Milk production and milk composition were similar among treatments (p > 0.05). In conclusion, supplementation of MPM at 200 g/head/day could modify ruminal fermentation and reduce methane production without adverse effect on feed intake, digestibility, hematological parameters, and milk production in dairy cows.     

Influence of storage time and processing on chemical composition and in vitro ruminal fermentation of olive cake

Olive oil extraction generates olive cake (OC) that could be used in ruminant feeding. However, the chemical composition of OC is affected by multiple factors, being therefore highly variable. The objective of this study was to analyse the influence of storage time and further processing: crude, exhausted (subjected to a second oil extraction) and cyclone (obtained from a cyclone separator) on nutritive value of OC samples. Twelve samples (six crude and six exhausted) were obtained monthly from the same pond from 1 to 6 storage months, and nine samples (three crude, three exhausted and three cyclone) were obtained monthly from a different pond from 6 to 9 months storage. Chemical composition was analysed, and OC samples were fermented in vitro with sheep rumen fluid. Increasing storage time up to 6 months decreased sugars and total soluble polyphenols content but increased fibre content in OC. Dry matter effective degradability (DMED) decreased linearly (p < 0.001) by 35.9 and 45.5% as storage time augmented from 1 to 6 months for crude and exhausted OC, respectively. Crude OC had lower DMED values than exhausted OC (averaged values 0.255 and 0.294 g/g, respectively). Both potential production and rate of gas production were lower (p ≤ 0.018) in crude compared with exhausted OC, which was attributed to the high fat content of crude OC (≥86 g/kg dry matter). For samples stored longer than 6 months, cyclone had greater (p < 0.05) DMED than crude and exhausted OC (averaged values 0.207, 0.164 and 0.164 g/g, respectively). The results indicate that ruminal degradability of OC is reduced with advancing storage time, but only subtle changes were observed during the first two months. Cyclone showed greater degradability than crude and exhausted OC, but differences between crude and exhausted OC became negligible after five storage months.     

脾酪氨酸激酶参与酿酒酵母甘露聚糖诱导绵羊瘤胃上皮细胞β-防御素-1(SBD-1)的表达

为了探索脾酪氨酸激酶(Syk)是否参与酿酒酵母甘露聚糖(S.c M)诱导绵羊瘤胃上皮细胞(ORECs)β-防御素-1(SBD-1)表达的过程,首先利用免疫组化、RT-PCR和免疫荧光等方法检测Syk在ORECs内的表达情况;然后采用qPCR和Western blot方法检测S.c M刺激ORECs后Syk的表达变化,同时用Western blot方法检测Syk的磷酸化水平;接着用3条Syk特异性siRNAs(#1、#2和#3)转染ORECs 24 h后,qPCR检测Syk mRNA的表达变化,筛选出干扰效果最佳的Syk siRNA;最后用效果最佳的siRNA和Syk特异性抑制剂R406分别处理ORECs后,采用qPCR和ELISA检测SBD-1的表达变化,以确定Syk在S.c M诱导SBD-1表达过程中的作用。结果显示:Syk在ORECs内表达;且S.c M刺激ORECs后Syk的mRNA和蛋白表达水平显著高于未刺激组(P<0.01或P<0.05),S.c M刺激ORECs不同时间(5、15、30、45和60 min)均能使Syk发生磷酸化,且刺激15 min后磷酸化水平达到最大(P<0.01);此外,Syk的3条特异性siRNAs转染ORECs后Syk的表达均降低,且Syk siRNA#2的抑制效果最明显(P<0.01);同时Syk siRNA#2和R406均能极显著降低S.c M诱导ORECs SBD-1的表达(P<0.01)。上述结果表明,Syk参与S.c M诱导ORECs SBD-1的表达。     

Effects of different forage combinations in total mixed rations on in vitro gas production kinetics,ruminal and milk fatty acid profiles of lactating cows

This study aimed to determine the effects of different forage combinations on in vitro gas production (GP) kinetics, ruminal and milk fatty acid profiles. Forty‐five lactating cows were randomly arranged into three groups and fed three total mixed rations (TMRs) with different forage combinations: TMR1, 23% alfalfa hay, 7% Chinese wild ryegrass hay and 15% whole corn silage; TMR2, 30% corn stover plus 15% whole corn silage; TMR3, 30% rice straw plus 15% whole corn silage. In vitro dry matter disappearance ranked: TMR1 > TMR2 > TMR3, and highest cumulative GP and asymptotic GP occurred in TMR1 while no difference occurred between TMR2 and TMR3. The average GP rate ranked: TMR1 > TMR2 > TMR3. TMR1 in comparison with TMR2 and TMR3 presented lower rumen contents of acetate and butyrate and greater rumen contents of propionate, valerate, C13:0, C14:0, C15:0, C18:1cis‐9, C18:2n‐6, C18:3n‐3, C20:0 and C22:0 as well as milk C18:2n‐6 and C18:3n‐3 proportions. Transfer efficiencies of C18:2n‐6 and C18:3n‐3 from diet to milk ranked: TMR1 > TMR2 > TMR3. The findings suggest TMRs containing alfalfa hay and Chinese wild ryegrass hay in comparison with corn stover or rice straw improve rumen fermentation and transfer efficiency of C18:2n‐6 and C18:3n‐3.     

3-硝基酯-1-丙醇抑制反刍动物瘤胃甲烷排放的机理及对动物的影响

反刍动物甲烷的排放既造成饲料能量的浪费,也会加剧全球变暖作用。在反刍动物瘤胃中,产甲烷菌主要利用二氧化碳转化产生甲烷。产甲烷菌转化二氧化碳的最后一步反应需要甲基辅酶M还原酶参与,3-硝基酯-1-丙醇(3-nitrooxypropanol,3-NOP)是一种甲基辅酶M类似物,能与辅酶B结合,从而减少甲基辅酶M与辅酶B结合生成甲烷,因此3-NOP能有效地降低瘤胃甲烷的产生。本文旨在阐明3-NOP抑制反刍动物瘤胃甲烷产生的机制以及对反刍动物生产的影响。