Validation and application of real-time polymerase chain reaction assays for representative rumen bacteria

Real‐time polymerase chain reaction (PCR) assays for 11 representative rumen bacterial species were validated. The sensitivity was tested by using the serially diluted target 16S rDNA from respective bacterial species. The recovery of the target DNA and the assay reproducibility were determined using DNA from rumen fluid spiked with different quantities of the target. Minimum detection levels for the target were 10–100 copies in pure culture. The recovery of the added target ranged from 82.4 to 116.6%. The intra‐ and inter‐assay variations of each assay were <9.4 and <12.6%, respectively. Therefore, the real‐time PCR assays evaluated in the present study are considered to be sufficiently reliable for monitoring all 11 bacterial species in the rumen. The assays were then applied to the monitoring of the bacterial species attached to ruminally incubated rice straw. Among the monitored fibrolytic species, Fibrobacter succinogenes was found to be the most dominant, accounting for 2.61% of total bacteria after 24 h incubation. Selenomonas ruminantium and Streptococcus bovis, non‐fibrolytics, were detected on the rice straw at 8.96% and 1.16% of total bacteria, respectively. Such high levels of non‐fibrolytics on the plant fiber suggest a synergistic relationship between fibrolytics and non‐fibrolytics.     

Cellulase Supplementation Does Not Improve the Digestibility of a High-Forage Diet in Horses

Six mature Arabian geldings were used in a two-period crossover study to investigate the effects of cellulase supplementation on fiber digestion. Horses were randomly assigned to either a control (CO; n = 3) or a cellulase (CE; n = 3) treatment for the first period and then treatments were switched for period 2. Each period consisted of a 10-day diet adaptation followed by a 3-day total fecal collection. The enzyme mixture contained 40,000 cellulase units/g and was fed at a rate of 3 g/day split evenly between two feedings. During the diet adaptation period, horses had ad libitum access to timothy hay and were also fed 165 g whole oats as a carrier for the supplement. When eating the CO treatment, horses consumed 16% more hay than when on the CE treatment (P = .004). Fecal output also tended to be greater when horses consumed the CO treatment as compared with CE treatment (P = .07). No differences were found between treatments for fecal percent dry matter (DM%), fecal neutral detergent fiber (NDF), fecal acid detergent fiber (ADF), fecal nitrogen (N), or fecal gross energy (GE). There was a trend for horses consuming the CO treatment to digest more NDF than when consuming the CE treatment (34.6% ± 1.5 vs 31% ± 1.5; P = .07). Horses also digested a greater %ADF, %N, and Mcal of energy when consuming the CO treatment than when consuming the CE treatment (P < .05). Cellulase addition to a hay-based horse diet decreased digestion of fiber components.     

3种不同精粗比日粮条件下空怀小尾寒羊母羊的消化与代谢

本试验选取18只体重约42 kg的空怀小尾寒羊母羊,按体重分为3组(n=6),分别喂给以玉米秸秆为粗饲料的精粗比为20∶80(日粮1)、30∶70(日粮2)和40∶60(日粮3)的日粮,以研究不同精粗比日粮对空怀小尾寒羊母羊消化代谢的影响。结果表明,3组羊只的有机物采食量分别为970.1±95.1、1137.3±144.8和1279.2±138.7 g/d.只;玉米秸秆的采食量分别为834.3±75.6、856.1±110.2和822.0±121.7 g/d.只;有机物表观消化率分别为52.53%±5.16%、55.73%±2.64%和59.68%±1.95%;粗蛋白质表观消化率分别为58.73%±7.60%、61.05%±2.61%和62.84%±2.33%;纤维素表观消化率分别为57.42%±6.04%、56.12%±1.06%和53.42%±3.83%;氮保留量分别为0.62±1.17、3.15±2.22和5.87±1.56 g/d.只;钙保留量分别为0.39±0.05、0.59±0.10和0.79±0.06 g/d.只;磷保留量分别为0.40±0.04、0.59±0.06和0.82±0.05 g/d.只;所获得的可消化能分别为9.40±1.37、12.31±2.05和15.09±1.83 M J/d.只;试验期体增重分别为-12.4±46.1、101.2±60.0和157.1±96.5 g/d.只。由此得出结论,在以玉米秸秆为粗饲料的条件下,当空怀小尾寒羊母羊日粮中的精料量为20%时,体重难以维持,并且氮保留率也低(3.29%);增加日粮中的精料量到30%或40%时,可提高日增重,并且使氮保留率达到13.82%或22.09%。增加日粮的精料比例可不同程度地提高有机物、能量、粗蛋白质和钙磷的表观消化率,但对纤维素类物质的消化有一定影响。增加动物的可消化能量供应可大幅度改善氮的利用率,也增加钙磷的保留量。提高日粮的精粗料比例,并不影响小尾寒羊对玉米秸秆的采食量。     

Differential ability of alcohol fermentation between the seeds of flooding-tolerant and flooding-susceptible varieties of Echinochloa crus-galli (L.) Beauv.

Seed germination and respiratory metabolism under aerobic and anaerobic conditions were studied in the flooding-tolerant and flooding-susceptible varieties of Echinochloa crus-galli with an identical genome and common ancestry . In the flooding-tolerant E. crus-galli var. formosensis , the seeds imbibed under nitrogen could germinate and exhibited an I/N quotient > 0.6. They accelerated glycolysis and concomitantly produced large and equimolar quantities of carbon dioxide and ethanol, suggesting that the seeds of this variety showed operation of the Pasteur effect and respired through alcohol fermentation under anaerobic conditions. The seeds excreted most of the toxic fermentation product. In contrast, the seeds of the flooding-susceptible E. crus-galli var. praticola were capable of germinating only under aerobic conditions through the conventional aerobic respiration and were unable to anaerobically respire to germinate through alcohol fermentation regardless of the presence of both sufficient alcohol dehydrogenase activity and a high redox charge of the pyridine nucleotides.     

Innovative liquid formulation of digestates for producing a biofertilizer based on Bacillus siamensis: Field testing on sweet pepper

A biofertilizer (BF) based on the plant growth promoting rhizobacterium (PGPR) Bacillus siamensis was produced using anaerobic digestate (AD) as the main ingredient of the growth medium, alongside a carbon source from residual origin. The use of residues for the growth of PGPR reduces the production costs of biofertilizers, but makes an assessment of the possible toxicity of residues for the bacteria or plants necessary. Therefore, the growth medium of PGPR was first optimized using the response surface methodology (RSM), followed by phytotoxicity tests and a field trial of the BF in a sweet pepper (Capsicum annuum L.) crop at two different locations. AD at 50% dilution, supplemented with 2.3% sugar beet molasses, was the optimum growth medium for producing the BF, with a bacterial concentration of 10⁹ cfu mL⁻¹. In the field trial, the treatments inoculated with BF and fertilized with decreased mineral N (80%) produced significantly better yields per ha than the controls with decreased N (80%) and full N (100%) without BF. This indicates improved efficiency of N use by the crop, as a consequence of the use of BF.     

Collection,Analysis, and Utilization of Biogas from Anaerobic Treatment of Crab Processing Waters

Abstract

Two laboratory-scale, upflow anaerobic reactors were constructed to treat blue crab (Callinectes sapidus) cooking wastewater for determination of the volume and composition of the resulting bio-gas. Both systems (A and B) consisted of two 4-liter (L) upflow anaerobic reactors and one 8-L aerobic reactor in series. The first anaerobic reactor of System A contained 240,12.7 mm foam cubes, while the first anaerobic reactor of System B contained 180 foam pieces (25.4 ×25.4 ×12.7 mm). Biogas production rates for System A ranged from 6.6 L gas/L feed to 10.0 L gas/L feed. Mean values for System B ranged from 7.1 L gas/L feed to 11.9 L gas/L feed. Methane and carbon dioxide comprised 68% and 28%, respectively, of the gas from both systems. Hydrogen sulfide averaged 1.5% and 1.4% for systems A and B, respectively. A pilot-scale biogas collection and utilization system was also constructed and successfully demonstrated. Details of the pilot-scale system design are provided in the paper to help processors better understand how the technology might be applied.     

泔脚厌氧发酵中试系统设计及运行研究

在小型试验的基础上,进行了总容积1300 L的预酸化两相连续发酵的中试设备构建设计;运行试验结果表明,在中温发酵条件下,泔脚废物的预酸化两相处理,在负荷1.66 kg.m-3d-1和35℃时,每吨泔脚能产生102 m3沼气。与城市有机废物大型发酵工艺的技术与运营指标对比定性分析表明,本工艺条件下的泔脚厌氧发酵处置具有广阔的社会产业化应用前景。     

动物营养研究中有关消化与代谢概念的辨正

消化试验与代谢试验是动物营养研究中最常应用的方法,两种方法的原理和试验步骤都源自营养物质(与能量)在动物消化道与体组织内的转化过程。但近年国内一些研究报道(特别是禽方面)中,忽略了被吸收的有机养分分别汇集入碳代谢池与氮代谢池的事实,误认为能测出消化率的养分也能被测定代谢(利用)率。文章就此进行了讨论,以便有助于辨别有关研究中的正误,提高其科学性。     

藏猪对饲粮纤维的消化及其与肠道微生物的相关性研究

旨在研究藏猪对饲粮纤维素和半纤维素的消化及其与粪便细菌的相关性,探索藏猪具备较强纤维降解能力的潜在因素。采用消化试验测定150 d放牧藏猪、舍饲藏猪和商品猪（杜×长×大猪,DLY猪）对饲粮纤维的表观消化率。采集粪便样品利用单分子实时测序技术,测定粪便细菌16S rRNA基因全长序列,分析粪便细菌群落的结构和多样性,采用Spearman相关分析获取饲粮纤维表观消化率与粪便中细菌群落的相关性。结果表明,放牧藏猪对饲粮纤维素和半纤维素的表观消化率显著高于舍饲藏猪与DLY猪（P<0.05）。在放牧藏猪、舍饲藏猪与DLY猪粪便细菌中共鉴定出15个门、26个纲、48个目、87个科、190个属、419个种。放牧藏猪粪便细菌中有1个门纤维杆菌门（Fibrobacteres）、3个属拟普雷沃菌属（Alloprevotella）、纤维杆菌属（Fibrobacter）与琥珀酸弧菌属（Succinivibrio）、3个种（Alloprevotella rava、Fibrobacter intestinalis、Succinivibrio dextrinosolvens）相对丰度显著高于舍饲藏猪与DLY猪（P<0.05）,并与饲粮纤维表观消化率呈显著正相关（P<0.05）。综上所述,放牧藏猪具备较强的纤维消化能力,这种能力与粪便中的纤维降解菌密切相关。     

奶牛乳腺上皮细胞的原代培养及其生物学特性分析

旨在从奶牛乳腺组织中分离原代乳腺上皮细胞（bovine mammary epithelial cells,BMECs）并传代培养后探究其生物学特性。本研究从屠宰场采集健康泌乳奶牛乳腺并采用改进的酶消化法从乳腺中分离得到原代奶牛乳腺上皮细胞,通过形态学观察、免疫荧光以及染色体核型分析的方法对其进行鉴定。同时,研究第3、第6和第9代乳腺上皮细胞的生长曲线、群体倍增时间和冻存复苏活力,检测不同代次细胞分泌乳蛋白、乳脂、乳糖的功能及泌乳相关基因的表达。结果表明,所分离的奶牛乳腺上皮细胞纯度较好,细胞生长呈现S型,3个代次细胞的群体倍增时间依次为34.87、41.45和65.04 h,冻存复苏活力为88%~93%;在细胞分泌功能方面,诱导培养2 d后均能检测到酪蛋白、甘油三酯和乳糖,且各代次间无显著差异;此外,3个代次的细胞诱导后均能表达乳成分合成相关基因。本研究成功培养了原代奶牛乳腺上皮细胞,并证明直到第9代细胞仍然具有正常的生物学功能,为体外探究乳腺细胞增殖与分化机制提供了良好的试验材料和技术支撑。