Abstract
The objective of this study will be to evaluate the inclusion of the Yerba-mate extract and their effects on fermentation, degradability of the diet and microbial populations in the dual-flow continuous culture system. The experiment will be conducted in the Faciola's Lab at the Department of Animal Science, University of Florida with 8 fermenters, each fermenter unit will be randomly assign to receive each diet once over the 4 periods in a replicate 4x4 Latin square design. Each 10-day period will be consist 7-day diet adaptation period followed by a 3-day sampling period. Experimental treatments given on a dry matter basis were will be use a control diet, without yerba-mate extract and 3 diets with increasing levels (1.5; 3 and 4.5% of yerba-mate extract). Fermentation conditions will kept constant with temperature set at 39°C, individual pH meters will be used to monitor the pH of each fermenter. Artificial saliva will be continuously infused into fermenters at rate of 2 mL/min. Each fermenter will manually fed a total of 72 g DM/d equally divided in two meals per day. Rumen fluid will be collected approximately 2 h after morning feeding from 2 ruminally cannulated dairy cows. 1,250 mL of rumen fluid will be then poured into each of the fermentation jars until it cleared the overflow spout. During the last 3 days of each period, liquid and solid effluents from each fermenter will combined and homogenized. Then, will be sample stored for the N-NH3 and volatile fatty acids analyses. At the end of each period, digesta effluent from the three sampling days will be composited by fermenter and freeze, for measurement of dry matter, ash, crude protein, ether extract, neutral detergent fiber, and acid detergent fiber, and will be calculated organic matter and component degradability. At the same time to the sample collection for evaluation of the fermentation (day 8, 9, and 10), at 2, 6, and 10 h after morning feeding. At the same time points, solid particles will collected from solid effluent containers of each fermenter. Liquid and solid samples will stored at -80°C for further DNA extraction. For the DNA extraction will used the Phenol-chloroform method. The 16S and 18S rRNA gene will amplified for the bacterial, archaea, and protozoa DNA. All statistical procedures will be carried out using SAS 9.2 for windows, with ± = 0.05.
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