Screening Yeasts from Ruminal Fluid of Dairy Heifer Fed a Different Ratio Roughage to Concentrate Diets

The objective of these studies were to identify ruminal yeast in varying ratios of roughage to concentrate in TMR diets in order to explore yeast diversity by using molecular technique with similarity of rDNA sequence. The experiment was assigned to four 98.6% of cross bred Holstein Friesian heifers with 2 levels and two replicates of roughage to concentrate ratios as： 10：90 （T1） and 50：50 （T2）. The experimental period was 14 days. Rumen fluid sample was collected by stomach tube for total DNA extraction by using silica gel method, and analysis of quantity and quality of DNA by Nanovue and agarose gel electrophoresis. The divergent DI/D2 domain of 26S rDNA was amplified by primers NL-1（5＇-GCA TAT CAA TAA GCG GAG GAA AAG-Y） and NL4 （5＇-GGT CCG TGT TTCAAG ACG G-3＇） by polymerase chain reaction （PCR）. The nucleotide sequences of D l/D2 domain of 26S rDNA were determined using PCR products. Generated sequences were aligned with related species by using the CLUSTAL W. The result showed that an average dry matter intake of TI was 7.00 kg/d and T2 was 6.99 kg/d. DNA concentrate from TIRI, TIR2, T2RI and T2R2 were 106, 131.5, 84 and 182.5 ng//aL, respectively. The purity of DNA was 1.57, 1.76, 1.78 and 1.86, respectively. The divergent D1/D2 domain of 26S rDNA of treatment could be amplified for T1R1 and T2R1 but could not for T1R2 and T2R2. The sequences of D1/D2 domain of 26S rDNA were compared with nucleotide database by BLAST programs （http：//www.ncbi.nlrn.nih.gov/BLAST）, the T2RI yeast-strain was closest to Yarrowia lipolytica. However, yeast strain in T1R1 could not be specifically identified because D 1/132 domain of 26S rDNA seem to represent variable region with number of nucleotide sequence showing 2-3 substitution from known species. The phylogenetic tree based on the sequences of the DI/D2 domain of 26S rDNA showed that TIR! was related to Pichia and Candida （96%） and T2R1 was related to Yarrowia lypolytica （100%）. This study indicated that ruminal yeast strains could be found varying in different ratio of roughage to concentrate.

Methane Emissions from Grazing Holstein-Friesian Heifers at Different Ages Estimated Using the Sulfur Hexafluoride Tracer Technique

Although the effect of animal and diet factors on enteric methane （CH4） emissions from confined cattle has been extensively examined, less data is available regarding CH4 emissions from grazing young cattle. A study was undertaken to evaluate the effect of the physiological state of Holstein-Friesian heifers on their enteric CH4 emissions while grazing a perennial ryegrass sward. Two experiments were conducted： Experiment 1 ran from May 2011 for 11 weeks and Experiment 2 ran from August 2011 for 10 weeks. In each experiment, Holstein-Friesian heifers were divided into three treatment groups （12 animals/group） consisting of calves, yearling heifers, and in-calf heifers （average ages： 8.5, 14.5, and 20.5 months, respectively）. Methane emissions were estimated for each animal in the final week of each experiment using the sulfur hexafluoride tracer technique. Dry matter （DM） intake was estimated using the calculated metabolizable energy （ME） requirement divided by the ME concentration in the grazed grass. As expected, live weight increased with increasing animal age （P 〈 0.001）; however, there was no difference in live weight gain among the three groups in Experiment 1, although in Experiment 2, this variable decreased with increasing animal age （P 〈 0.001 ）. In Experiment 1, yearling heifers had the highest CH4 emissions （g·d^-1） and in-calf heifers produced more than calves （P 〈 0.001 ）. When expressed as CH4 emissions per unit of live weight, DM intake, and gross energy （GE） intake, yearling heifers had higher emission rates than calves and in-calf heifers （P 〈 0.001）. However, the effects on CH4 emissions were different in Experiment 2, in which CH, emissions （g·d^-1） increased linearly with increasing animal age （P 〈 0.001）, although the difference between yearling and in-calf heifers was not significant. The CH4/live weight ratio was lower in in-calf heifers than in the other two groups （P 〈 0.001 ）, while CH4 energy output as a proportion of GE intake was lower in calves than in yearling and in-calf heifers （P 〈 0.05）. All data were then pooled and used to develop prediction equations for CH4 emissions. All relationships are significant （P 〈 0.001）, with R2 values ranging from 0.630 to 0.682. These models indicate that CH4 emissions could be increased by 0.252 g.d-1 with an increase of I kg live weight or by 14.9 g·d^-1 with an increase of 1 kg·d^-1 of DM intake; or, the CH4 energy output could be increased by 0.046 MJ·d^-1 with an increase of 1 MJ·d^-1 of GE intake. These results provide an alternative approach for estimating CH4 emissions from grazine dairy heifers when actual CH, emission data are not available.

Preference, Feeding Behavior, and in Vitro Fermentation Characteristics of Pelleted Feeds Containing Alkaline Aqueous Lignin By-Product from Paper Processing in Holstein Heifers

The objectives of this study were to evaluate the effect of using multiple dietary inclusion rates of alkaline aqueous lignin by-product(AALB),in replacing a commercial binder,on the preference and feeding behavior of dairy heifers,and in vitro digestibility.In the experiment I,five pelleted diets including negative control(no-binder),commercial lignosulfonate binder,and 1.6%,3.2%,and 4.8%of AALB were tested.In the experiment II,five pelleted diets including negative control,positive control(2%molasses),and 1%,2.4%,and 3.8%of AALB(all combined with 2%of molasses)were tested.Feeding behavior and preference were determined for 60 min per animal per evaluation.In vitro digestibility was determined for each pelleted diet for 48 h over five incubation runs.Results from Experiment I showed animals receiving 1.6%of AALB diet showed the greatest preference(dry matter(DM),intake)and a greater number of approaches with eating than other diets.The preference was lower when animals received control or 4.8%AALB diets relative to other diets.Results from Experiment II revealed animals receiving 3.8%AALB combined with molasses showed the greatest DM intake preference over 60 min,and DM intake per approach.Diets that included 2.4%and 3.8%of AALB combined with molasses presented the greatest in vitro DM digestibility.Overall,heifers showed a greater intake when receiving pelleted diets with 1.6%AALB.Furthermore,preference,intake over 60 min,and in vitro DM digestibility,were greater in the pelleted diets that contained higher rates of AALB when combined with molasses.