尿液嘌呤衍生物法估测瘤胃微生物蛋白质产量及其评价

近年来,以尿液嘌呤衍生物为标记物估测流入十二指肠瘤胃微生物蛋白质产量的非损害(non-inva-sive)方法得到较快发展。该方法的依据是反刍动物尿液中嘌呤衍生物主要来源于瘤胃内微生物体嘌呤的降解,尿液嘌呤衍生物与瘤胃微生物产量成正比。该方法的主要优点是不需要瘘管动物,但对瘤胃微生物蛋白质产量的估测值较十二指肠嘌呤法和15N法估测值低。许多研究者提出了不同的估算公式,适用于牛或羊的估算公式是不同的。     

用尿中嘌呤衍生物估测瘤胃微生物蛋白产量的研究进展

反刍动物尿中嘌呤衍生物包括尿囊素、尿酸、黄嘌呤和次黄嘌呤,尿中嘌呤衍生物是一种很好的估测瘤胃微生物蛋白合成量的标记物。本研究就动物内源嘌呤排泄、尿中嘌呤衍生物排泄量与嘌呤吸收量关系模型建立、全收尿法和点尿法计算微生物蛋白供给以及最近对不同动物(包括绵羊、黄牛、山羊、水牛、牦牛、骆驼、驼羊)嘌呤衍生物排泄的研究等方面,阐述用尿中嘌呤代谢物估测瘤胃微生物蛋白产量的方法。     

尿液嘌呤法估测瘤胃微生物蛋白研究

瘤胃微生物蛋白质（MCP）是反刍家畜的重要蛋白质来源。近年来以尿液嘌呤衍生物估测反刍动物瘤胃微生物蛋白产量的方法正逐步取代传统的标记法，其优点是不用瘘管，无损害，操作简单；缺点是其值为相对值，尚需进一步标准化。     

用嘌呤衍生物估测牛羊小肠消化吸收的微生物蛋白产量(综述)

综述了一种测定微生物蛋白（ＭＣＰ）的简单方法——嘌呤衍生物法。其测值可用作估测代谢蛋白的ＭＣＰ部分，对建立和完善反刍动物蛋白质新体系具有重要意义。     

瘤胃微生物蛋白质合成量估测标记物的研究

确定瘤胃微生物细胞净合成量对研究反刍动物营养需要至关重要。目前所采用的估测微生物蛋白合成量的方法主要是借助外科造瘘手术,利用瘤胃微生物自身固有的物质或通过外源性同位素标记物标记来估测微生物蛋白合成量。近年来,以尿嘌呤衍生物作为标记物估测流入十二指肠的瘤胃微生物蛋白产量的非损害方法也得到较快发展。笔者认为内外源标记结合的方法最合适。     

用嘌呤衍生物估测牛关小肠消化吸收的微生物蛋白产量（综述）

综述了一种测定微生物蛋白（ＭＣＰ）的简单方法－－嘌呤衍生物法。其测值可用作估测是白的ＭＣＰ部分，对建立和完善反刍动物蛋白质新体系具有重要意义。     

Response of urinary purine derivatives excretion to veast RNA infusion levels in the yak abomasum

本研究对3头牦牛施以皱胃瘘管手术,以酵母RNA为嘌呤碱基供体,连续注射4期,以期测定牦牛吸收嘌呤的回收率,为尿嘌呤衍生物估测牦牛瘤胃微生物氮产量的模型积累数据.结果表明,牦牛皱胃连续注射酵母RNA可使尿囊素(564～1 426 μmol/kg BW0.75)、总嘌呤衍生物(purine derivative,PD)排出量(629～1 507 μmol/kgBW0.75)及尿囊素占总PD比例线性提高(0.90～0.95)(P＜0.01)；嘌呤碱基注射水平对尿酸、肌酐及尿氮排出量影响不显著(P＞0.05).回归分析发现,牦牛皱胃嘌呤注射量(X,mmol/d)与尿嘌呤衍生物排出量(Y,mmol/d)间存在线性关系:Y=0.85X+33.02 (R2 =0.96,P＜0.001),牦牛吸收嘌吟在尿中的回收率为85％.     

不同营养水平对4～6月龄荷斯坦犊牛尿中嘌呤衍生物排出量的影响

本试验采用3头(120±5)日龄、体重为(72±2)kg的荷斯坦公犊牛,通过调整日粮营养水平来观察其尿中嘌呤衍生物(PD)的排出规律,比较其与成年反刍动物之间的差异,考察犊牛期能否用尿嘌呤衍生物法来估测瘤胃微生物蛋白。试验设计3种营养水平日粮,采用随机区组试验设计。结果发现,犊牛尿中嘌呤衍生物中尿酸大约为50%～75%,而在成年反刍动物尿酸不到15%。PD的排出量随可消化有机物(DOM)和可消化粗蛋白(DCP)进食量升高而上升,存在弱的相关性(R2=0.76,n=9)。其排出水平为0.12～0.24mmol/(BW0.75·d)。作为估测成年反刍动物微生物蛋白合成量的另一个指标[PD]×BW0.75/[C](其中[PD]为嘌呤衍生物浓度,[C]为尿中肌酐酸浓度,BW0.75为代谢体重,简称PDC),与DOM和DCP进食量存在负相关,这与成年反刍动物的研究结果是相反的。     

尿液嘌呤衍生物法估测瘤胃微生物蛋白产量的研究进展

一种非伤害性方法-尿液嘌呤衍生物(PD)作为标记物估测瘤胃微生物蛋白产量近年来得到较快发展.本文就这种方法的原理依据,发展及应用进行了综述,分析了这种方法存在的一些问题,展望了将来的研究和发展方向.     

微生物氮测定方法比较

定量测定瘤胃微生物细胞净合成量对研究反刍动物营养需要至关重要。目前所采用的估测微生物蛋白合成量的方法主要是借助外科造瘘手术，利用瘤胃微生物自身固有的物质或通过外源性同位素标记物标记来估测微生物蛋白合成量。近年来，以尿液嘌呤衍生物作为标记物估测瘤胃微生物蛋白产量的非损害方法也得到较快发展。笔者认为内外源标记结合的方法最合适。     

Technical note: A system for continuous recording of ruminal pH in cattle

Continuous recording of ruminal pH in cannulated cattle has been practiced to study rumen metabolism. However, most systems reported did not permit animal mobility during pH recording. Therefore, the objective of this study was to develop a continuous rumen pH data acquisition system that permitted animal mobility during data acquisition. A further objective was to compare the pH readings obtained using the continuous recording system to readings obtained at the same time using spot sampling. The continuous recording system was composed of a heavy-duty electrode and a data logger. The electrode was attached to a 0.5-kg weight to help maintain the electrode in the ventral sac of the rumen. The electrode was connected via a 0.5-m cable to a lightweight data logger that was mounted on the animal's back using a belt wrapped around the girth. The data logger was battery powered and could hold over 13,000 pH data values. A personal digital assistant was used to configure and download data from the data logger during the experiment. Ruminal pH was continuously recorded (every 10 s) using a dry Holstein cow fed alfalfa hay ad libitum in a 3-d experiment to compare the performance of the continuous system to spot samples taken from the ventral sac of the rumen, the same location as the continuous electrode. The spot samples were collected 3 times per d for 3 d. At every sampling time, 3 replicate samples were collected, pH was determined immediately using a handheld pH meter, and readings were averaged (n = 3) and compared with the average of the 3 pH readings recorded using the continuous system at the same time. The pH recorded by spot sampling (6.63 +/- 0.04) was greater (P = 0.009) than that of the continuous system (6.56 +/- 0.03), with a correlation of r = 0.88 (P = 0.002). The continuous recording system has the potential to facilitate measurement of ruminal pH in free-roaming cattle.     

Research note: a method for studying local differences in ruminal fermentation in dairy cattle.

A method was developed for studying local differences in ruminal fermentation. The developed sampler consisted of an acrylic glass container (460 cm3) with an aperture for digesta sampling, which could be opened and closed by the scaled "T" rod. The scale was a reference for defined rumen layers: top, middle, 5 to 10 cm and 25 to 35 cm beneath the top of particles mat, respectively, and bottom 5 to 10 cm above the rumen floor. The repeatability of the method was proved in two rumen cannulated cows. Particle/fluid ratio, pH and sample amount were measured 2 to 2 1/2 h after morning feeding in four replicates each day (over 5 days), rumen layer and animal. No significant differences between replicates were observed. The coefficients of variation (CV) of the particle/fluid ratio varied between 8.7% and 13.6%. Top layer had higher CV than middle and bottom layer. CV of pH ranged between 0.59% and 1.27%. The developed method of sampling showed satisfactory repeatability for investigation of digesta properties and fermentation in different rumen layers.     

不同蛋白质组成的日粮对瘤胃发酵及微生物蛋白合成的影响

本试验选用4只体况良好、体重(30.0±2.5) kg的山羊, 安装瘤胃瘘管,按4×4拉丁方试验设计,研究4种不同蛋白质豆粕（A组）、棉粕（B组）、菜粕（C组）和DDGS（D组）组成的日粮对山羊瘤胃发酵及微生物蛋白合成的影响。研究结果表明,各组瘤胃液pH无显著差异（P>0.05）;A组的氨氮（NH₃-N）浓度显著高于D组（P<0.05）;A组的微生物蛋白（MCP）浓度显著高于B、C组（P<0.05）,极显著高于D组（P<0.01）;各组乙酸、丙酸、丁酸浓度差异显著,总VFA浓度A、B组显著高于C、D组（P<0.05）     

A technique for sampling total rumen contents in sheep

The paper describes the manufacture of an easily removable large rumen fistula closure for sheep and a restraining cradle which is used when sampling total rumen contents. Eight sheep have been fitted with the fistula and have been used regularly for nine months. The technique is rapid and simple and the closure is more reliable than other types which are often removed accidentally.     

The ruminal bacterial community in lactating dairy cows has limited variation on a day-to-day basis

Dairy cows rely on a complex ruminal microbiota to digest their host-indigestible feed. Our ability to characterize this microbiota has advanced significantly due to developments in next-generation sequencing. However, efforts to sample the rumen, which typical y involves removing digesta directly from the rumen via a cannula, intubation, or rumenocentesis, is costly and labor intensive. As a result, the majority of studies characterizing the rumen microbiota are conducted on samples col ected at a single time point. Currently, it is unknown whether there is significant day-to-day variation in the rumen microbiota, a factor that could strongly influence conclusion drawn from studies that sample at a single time point. To address this, we examined day-to-day changes in the ruminal microbiota of lactating dairy cows using next-generation sequencing to determine if single-day sampling is representative of sampling across 3 consecutive days. We sequenced single-day solid and liquid fractions of ruminal digesta col ected over 3 consecutive days from 12 cannulated dairy cows during the early, middle, and late stages of a single lactation cycle using the V4 region of the bacterial 16 S r RNA gene. We then generated 97% similarity operational taxonomic units(OTUs) from these sequences and showed that any of the individual samples from a given 3-day sampling period is equivalent to the mean OTUs determined from the combined 3-d data set. This finding was consistent for both solid and liquid fractions of the rumen,and we thus conclude that there is limited day-to-day variability in the rumen microbiota.     

反刍动物过瘤胃蛋白的保护措施

过瘤胃蛋白就是将一些蛋白质经过处理，避免在瘤胃内被发酵、降解，而直接进入小肠后再被消化吸收，从而达到提高饲料蛋白质利用率的目的。保护饲料蛋白免遭微生物过量降解，提高蛋白质的总体利用率的方法有许多，主要有物理方法、化学方法、生物学调控、瘤胃外流速度的调控、利用食管沟反射提高过瘤胃蛋白等。本文就目前蛋白质的各种过瘤胃保护途径和效果进行了综述。     

丙酸镁对西门塔尔牛尿嘌呤衍生物和微生物蛋白的影响

选用4头年龄3.5岁,体况良好,体重500kg的中国西门塔尔牛阉牛。采用4×4拉丁方设计,研究日粮添加丙酸镁(100g/d)、丙酸(87g/d)和氧化镁(24g/d)对尿嘌呤衍生物(PD)和微生物蛋白(MCP)的影响。结果表明:日粮中添加丙酸镁、丙酸和氧化镁后,丙酸镁组和丙酸组尿囊素、PD、微生物氮(MN)、MCP较对照组和氧化镁组显著提高(P<0.05),但丙酸镁组和丙酸组差异不显著(P>0.05)。日粮添加丙酸镁、丙酸和氧化镁对尿酸(uric acid)含量无显著影响(P>0.05)。日粮中添加丙酸镁和丙酸提高了尿囊素含量和PD、MN和MCP产量,但处理对尿酸没有影响。     

瘤胃微生物定量方法研究进展

瘤胃微生物的定量有传统计数和现代分子定量2大类方法。其中，传统方法包括经典的亨氏滚管法和最大可能法，但它们测定瘤胃微生物总数的结果可能偏低。运用分子方法定量一些重要的微生物不仅能很好地解释它们在瘤胃代谢中的作用，而且有助于描述更多的微生物特性。因此，作者综述了瘤胃微生物定量方法的发展历程及研究进展，重点描述了运用分子定量方法的研究情况。     

低淀粉饲粮条件下不同瘤胃降解淀粉水平对体外瘤胃发酵的影响

本试验旨在研究玉米作为淀粉来源的低淀粉饲粮条件下不同瘤胃降解淀粉(RDS)水平对体外瘤胃发酵的影响。以3头安装有永久性瘤胃瘘管的健康荷斯坦奶牛作为瘤胃液供体,各组分别以不同RDS水平饲粮作为发酵底物,体外产气法测定培养48 h时产气量和瘤胃发酵参数以及24 h时瘤胃微生物区系变化。结果表明:1)随着饲粮RDS水平的提高,体外培养48 h时产气量、潜在产气部分和产气速率呈线性升高(P0.05),快速发酵部分产气量呈线性下降(P0.05),干物质消失率呈线性升高(P0.05);2)随着饲粮RDS水平的提高,体外培养48 h时发酵液微生物蛋白、乙酸、丙酸、丁酸和总挥发性脂肪酸浓度呈线性升高(P0.05),pH和氨态氮浓度没有显著变化(P0.05);3)随着饲粮RDS水平的提高,体外培养24 h时发酵液中白色瘤胃球菌和嗜淀粉瘤胃杆菌的相对数量呈线性升高(P0.05),黄色瘤胃球菌、琥珀酸丝状杆菌、溶纤维丁酸弧菌、牛链球菌和溶淀粉琥珀酸单胞菌的相对数量没有显著变化(P0.05)。综合考虑,低淀粉饲粮条件下提高RDS水平有利于瘤胃发酵。     

Comparative characterization of reticular and duodenal digesta and possibilities of estimating microbial outflow from the rumen based on reticular sampling in dairy cows

The objective of this experiment was to investigate the possibility of estimating the outflow of nutrients and microbial protein from the rumen based on sampling reticular contents as an alternative to duodenal sampling. Microbial protein flow estimates were also compared with a third method based on sampling of ruminal contents. Reticular and duodenal digesta and ruminal contents were recovered from 4 cows used in a 4 x 4 Latin square design experiment, in which the ruminal effects of 4 exogenous enzyme preparations were studied. Large and small particulate and fluid markers were used to estimate digesta flow in a triple-marker model; 15N was used as a microbial marker. Reticular and duodenal digesta were segregated into small and large particles (SP and LP, respectively) and a fluid phase, and ruminal digesta was segregated into particulate and fluid phases. Compared with digesta recovered at the duodenum, reticular digesta had lower OM and greater NDF contents. The proportion of microbial N was notably greater in the fluid phase of reticular digesta. Ruminal outflow of DM and OM was greater (by 17 and 28%) and that of NDF was lower (by 14%) when estimated from duodenal compared with reticular samples. There was no difference in the estimated flow of starch and nonammonia and microbial N between the reticular and duodenal techniques. Microbial N flow estimated based on ruminal sampling was similar to those based on duodenal and reticular sampling. The ruminal method, however, grossly overestimated flow of DM, OM, and NDF. This study supports the concept that microbial protein outflow from the rumen can be measured based on sampling of ruminal or reticular digesta. The reticular sampling technique can also provide reliable estimates for ruminal digestibility of OM, N, and fiber fractions. These findings need to be confirmed in experiments with basal diets varying in structure and forage-to-concentrate ratios.