The objectives of this study were: 1) to compare the effects of live yeast (LY), yeast fermentation product (YFP), a mix of Lactobacillus acidophilus and Propionibacterium freudenreichii (MLP), and Lactobacillus plantarum included as additives in dairy cows’ diets on in vitro ruminal fermentation and gas production (GP); and 2) to evaluate the effects of L. plantarum as direct-fed microbials (DFM) in dairy cows’ diets on in vitro ruminal fermentation, GP, nutrient digestibility, and N metabolism. Three experiments were carried out: Exp. 1 had the objective to compare all additives regarding ruminal fermentation parameters: an Ankom GP system was used in a completely randomized design, consisting of four 48 h incubations, and eight replications per treatment. There were eight treatments: a basal diet without additive (CTRL) or with one of the following additives: LY, YFP, MLP, or L. plantarum at four levels (% of diet Dry Matter (DM)): 0.05% (L1), 0.10% (L2), 0.15% (L3), and 0.20% (L4). In Exp. 2, a batch culture was used to evaluate ruminal fermentation, and CO2 and CH4 production using the same treatments and a similar experimental design, except for having 16 replications per treatment. Based on Exp. 1 and 2 results, Exp. 3 aimed at evaluating the effects of the L. plantarum on ruminal true nutrient digestibility and N utilization in order to evaluate the use of L. plantarum as DFM. The treatments CTRL, MLP, L1, and L2 were used in a replicated 4 × 4 Latin square design using a dual-flow continuous culture system. Data were analyzed using linear and nonlinear regression; treatment means were compared through contrasts, and L treatments in Exp. 1 and 2 were tested for linear, quadratic, and cubic effects. In Exp. 1, all treatments containing additives tended to reduce OM digestibility as well as reduced total volatile fatty acids (VFA) concentration and total GP. The YFP had greater OM digestibility than LY, and MLP treatment had greater total VFA concentration compared to L. plantarum treatments. In Exp. 2, additives reduced CO2 production, and there were no major differences in CH4. In Exp. 3, all additives reduced NH3-N concentration. In conclusion, pH and lactate concentration were not affected in all three experiments regardless of additive tested, suggesting that these additives may not improve ruminal fermentation by pH modulation; and L. plantarum may improve ruminal N metabolism when used as DFM in high-producing dairy cows’ diets, mainly by reducing NH3-N concentration.

Undesirable interactions between trace mineral elements and ruminal contents may occur during digestion when mineral salts are supplemented. Antimicrobial effects of copper sulfate (CuSO 4 ) may affect ruminal digestibility of nutrients when fed as a source of copper (Cu), while sodium selenite (Na 2 SeO 3 ) may be reduced in the rumen to less available forms of selenium (Se). Our objective was to evaluate if protection of CuSO 4 and Na 2 SeO 3 by lipid-microencapsulation would induce changes on ruminal microbial fermentation. We used 8 fermentors in a dual-flow continuous-culture system in a 4 × 4 duplicated Latin square with a 2 × 2 factorial arrangement of treatments. Factors were CuSO 4 protection (unprotected and protected by lipid-microencapsulation) and Na 2 SeO 3 protection (unprotected and protected by lipid-microencapsulation). Treatments consisted of supplementation with 15 mg/kg of Cu and 0.3 mg/kg of Se from either unprotected or protected (lipid-microencapsulated) sources, as follows: (1) Control (unprotected CuSO 4 + unprotected Na 2 SeO 3 ); (2) Cu-P (protected CuSO 4 + unprotected Na 2 SeO 3 ); (3) Se-P (unprotected CuSO 4 + protected Na 2 SeO 3 ); (4) (Cu+Se)-P (protected CuSO 4 + protected Na 2 SeO 3 ). All diets had the same nutrient composition and fermentors were fed 106 g of dry matter/d. Each experimental period was 10 d (7 d of adaptation and 3 d for sample collections). Daily pooled samples of effluents were analyzed for pH, NH 3 -N, nutrient digestibility, and flows (g/d) of total N, NH 3 -N, nonammonia N (NAN), bacterial N, dietary N, and bacterial efficiency. Kinetics of volatile fatty acids was analyzed in samples collected daily at 0, 1, 2, 4, 6, and 8 h after feeding. Main effects of Cu protection, Se protection, and their interaction were tested for all response variables. Kinetics data were analyzed as repeated measures. Protection of Cu decreased acetate molar proportion, increased butyrate proportion, and tended to decrease acetate: propionate ratio in samples of kinetics, but did not modify nutrient digestibility. Protection of Se tended to decrease NH 3 -N concentration, NH 3 -N flow, and CP digestibility; and to increase flows of nonammonia N and dietary N. Our results indicate that protection of CuSO 4 may increase butyrate concentration at expenses of acetate, while protection of Na 2 SeO 3 tended to reduce ruminal degradation of N. Further research is needed to determine the effects of lipid-microencapsulation on intestinal absorption, tissue distribution of Cu and Se, and animal performance.

Camelina is an oil seed crop that belongs to the Brassica family (Cruciferae). Camelina meal is a by-product from the biofuel industry that contains on average 38% crude protein and between 10 to 20% of residual fat, which limits the inclusion levels of camelina meal in dairy cow diets as the main protein supplement. Thus, we conducted a solvent extraction on ground camelina seed on a laboratory scale. The objectives of this study were (1) to assess the effects of replacing canola meal (CM) with solvent-extracted camelina meal (SCAM) in lactating dairy cow diets; and (2) to determine the effects of SCAM on microbial fermentation and AA flow in a dual-flow continuous culture system. Diets were randomly assigned to 6 fermentors in a replicated 3 × 3 Latin square with three 10-d experimental periods consisting of 7 d for diet adaptation and 3 d for sample collection. Treatments were 0, 50, and 100% SCAM inclusion, replacing CM as the protein supplement. Diets contained 55:45 forage:concentrate, and fermentors were fed 72 g of dry matter/d equally divided in 2 feeding times. On d 8, 9, and 10 of each period, samples were collected for analyses of pH, volatile fatty acids (VFA), N metabolism, NH-N digestibility, and AA flow. Statistical analysis was performed using the MIXED procedure of SAS (SAS Institute Inc., Cary, NC), and linear and quadratic effects of SCAM inclusion were assessed. Total VFA concentration and pH were not affected by diets. Molar proportion of acetate decreased, whereas molar proportion of propionate increased with SCAM inclusion. Total branched-chain VFA concentration was the least in fermentors fed diet 0, and greatest in fermentors fed diet 50. Digestibility of NDF decreased in fermentors fed SCAM diets, and dry matter, organic matter, and crude protein true digestibility were similar across diets. Concentration of NH-N linearly decreased, and non-NH-N linearly increased with SCAM inclusion. Bacterial efficiency (calculated as g of bacterial N flow/kg of organic matter truly digested) tended to be greater in fermentors fed diet 100. Outflow of Arg linearly increased with SCAM inclusion, whereas overall AA flow was not affected by diet. In conclusion, replacing CM with SCAM increased propionate molar proportion and non-NH-N flow, and decreased NH-N flow and concentration, which may improve animal energy status and N utilization. Inclusion of SCAM did not change most AA flow, indicating that it can be a potential replacement for CM.

The objectives of this study were to evaluate the effects of lipopolysaccharide (LPS) dosing on bacterial fermentation and bacterial community composition (BCC), to set up a subacute ruminal acidosis (SARA) nutritional model in vitro, and to determine the best sampling time for LPS dosing in a dual-flow continuous culture system. Diets were randomly assigned to 6 fermentors in a replicated 3 × 3 Latin square with three 11-d experimental periods that consisted of 7 d for diet adaptation and 4 d for sample collection. Treatments were control diet (CON), wheat and barley diet (WBD) to induce SARA, and control diet + LPS (LPSD). Fermenters were fed 72 g of dry matter/d. The forage: concentrate ratio of CON was 65:35. The WBD diet was achieved by replacing 40% of dry matter of the CON diet with 50% ground wheat and 50% ground barley. The LPS concentration in LPSD was 200,000 endotoxin units, which was similar to that observed in cows with SARA. The SARA inducing and LPS dosing started at d 8. The BCC was determined by sequencing the V4 region of the 16S rRNA gene using the Illumina MiSeq platform (Illumina Inc., San Diego, CA). The LPSD and CON maintained pH above 6 for the entire experimental period, and the WBD kept pH between 5.2 and 5.6 for 4 h/d, successfully inducing SARA. Digestibility of neutral detergent fiber and crude protein in LPSD were not different from WBD but tended to be lower than CON. Lipopolysaccharide dosing had no effect on pool of VFA concentrations and profiles but decreased bacterial N; the pattern changes of VFA and LPS in LPSD started to increase and be similar to WBD 6 h after LPS dosing. Pool of LPS concentration was around 11-fold higher in WBD and 4-fold higher in LPSD than CON. In the solid fraction, the BCC of LPSD was different from WBD and tended to be different from CON. In the liquid fraction, the BCC was different among treatments. The LPS dosing increased the relative abundance of Succinimonas, Anaeroplasma, Succinivibrio, Succiniclasticum, and Ruminobacter, which are main gram-negative bacteria related to starch digestion. Our results suggest that LPS dosing does not affect pH alone. However, LPS could drive the development of SARA by affecting bacteria and bacterial fermentation. For future studies, samples are suggested to be taken 6 h after LPS dosing in a dualflow continuous culture system.

The objective of this study was to evaluate ruminal microbiome changes associated with feeding Lactobacillus plantarum GB-LP1 as direct-fed microbials (DFM) in high-producing dairy cow diets. A dual-flow continuous culture system was used in a replicated 4 × 4 Latin square design. A basal diet was formulated to meet the requirements of a cow producing 45 kg of milk per day (16% crude protein and 28% starch). There were 4 experimental treatments: the basal diet without any DFM (CTRL); a mixture of Lactobacillus acidophilus, 1 × 10 9 cfu/g, and Propionibacterium freudenreichii, 2 × 10 9 cfu/g [MLP = 0.01% of diet dry matter (DM)]; and 2 different levels of L. plantarum, 1.35 × 10 9 cfu/g (L1 = 0.05% and L2 = 0.10% of diet DM). Bacterial samples were collected from the fluid and particulate effluents before feeding and at 2, 4, 6, and 8 h after feeding; a composite of all time points was made for each fermentor within their respective fractionations. Bacterial community composition was analyzed through sequencing the V4 region of the 16S rRNA gene using the Illumina MiSeq platform. Sequenced data were analyzed on DADA2, and statistical analyses were performed in R (RStudio 3.0.1, https: / / www .r -project .org/ ) and SAS 9.4 (SAS Institute Inc.); orthogonal contrasts were used to compare treatments. Different than in other fermentation scenarios (e.g., silage or beef cattle high-grain diets), treatments did not affect pH or lactic acid concentration. Effects were mainly from overall DFM inclusion, and they were mostly observed in the fluid phase. The relative abundance of the phylum Firmicutes, family Lachnospiraceae, and 6 genera decreased with DFM inclusion, with emphasis on Bu-tyrivibrio_2, Saccharofermentans, and Ruminococcus_1 that are fibrolytic and may display peptidase activity during fermentation. Lachnospiraceae_AC2044_group and Lachnospiraceae_XPB1014_group also decreased in the fluid phase, and their relative abundances were positively correlated with NH 3 -N daily outflow from the fermenters. Specific effects of MLP and L. plantarum were mostly in specific bacteria associated with proteolytic and fibrolytic functions in the rumen. These findings help to explain why, in the previous results from this study, DFM inclusion decreased NH 3 -N concentration without altering pH and lactic acid concentration.