Antioxidant action in diets with ground soybeans on ruminal microbial production, digestion, and fermentation in buffaloes

The objective of this study was to evaluate the effect of adding ground soybeans and antioxidants to diet of buffaloes on intake, ruminal microbial production, total and ruminal digestibility of nutrients, and ruminal fermentation parameters. Four crossbred buffaloes with a mean weight of 506±29 kg were distributed in a 4×4 Latin square. Four diets were tested: control; diet with ground soybeans; diet with ground soybeans and supplementation with yerba mate; and diet with ground soybeans and supplementation with yerba mate and vitamin E. The addition of ground soybeans had negative effects on intake of dry matter, organic matter, crude protein, neutral detergent fiber, and acid detergent fiber, as well as ruminal digestibility of neutral detergent fiber and total carbohydrates but did not influence total digestibility of nutrients. Yet, the ground soybeans diet increased the concentration of butyrate and microbial production in the rumen. The diet with ground soybeans supplemented with yerba mate decreased the concentration of acetate and increased the concentration of propionate and increased the efficiency of synthesis and the production of microbial protein in the rumen. There was a positive additive effect of vitamin E in the presence of yerba mate, enhancing the synthesis and production efficiency of microbial protein. Thus, the addition of ground soybeans and antioxidants to diet of buffaloes improves the efficiency of microbial protein synthesis and increases the production of butyrate, without, in general, altering the total digestibility of nutrient

Effects of calcium-magnesium carbonate and calciummagnesium hydroxide as supplemental sources of magnesium on ruminal microbiome

Our objective was to evaluate the inclusion of calcium-magnesium carbonate [CaMg(CO3 ) 2 ] and calcium-magnesium hydroxide [CaMg(OH)4 ] in corn silage-based diets and their impact on ruminal microbiome. Our previous work showed a lower pH and molar proportion of butyrate from diets supplemented with [CaMg(CO3 ) 2 ] compared to [CaMg(OH)4 ]; therefore, we hypothesized that ruminal microbiome would be affected by Mg source. Four continuous culture fermenters were arranged in a 4 × 4 Latin square with the following treatments defined by the supplemental source of Mg: 1) Control (100% MgO, plus sodium sesquicarbonate as a buffer); 2) CO3 [100% CaMg(CO3 ) 2 ]; 3) OH [100% CaMg(OH)4 ]; and 4) CO3 /OH [50% Mg from CaMg(CO3 ) 2 , 50% Mg from CaMg(OH)4 ]. Diet nutrient concentration was held constant across treatments (16% CP, 30% NDF, 1.66 MCal NEl/kg, 0.67% Ca, and 0.25% Mg). We conducted four fermentation periods of 10 d, with the last 3 d for collection of samples of solid and liquid digesta effluents for DNA extraction. Overall, 16 solid and 16 liquid samples were analyzed by amplification of the V4 variable region of bacterial 16S rRNA. Data were analyzed with R and SAS to determine treatment effects on taxa relative abundance of liquid and solid fractions. Correlation of butyrate molar proportion with taxa relative abundance was also analyzed. Treatments did not affect alpha and beta diversities or relative abundance of phylum, class and order in either liquid or solid fractions. At the family level, relative abundance of Lachnospiraceae in solid fraction was lower for CO3 and CO3 /OH compared to OH and Control (P < 0.01). For genera, abundance of Butyrivibrio (P = 0.01) and Lachnospiraceae ND3007 (P < 0.01) (both from Lachnospiraceae family) was lower and unclassified Ruminococcaceae (P = 0.03) was greater in CO3 than Control and OH in solid fraction; while abundance of Pseudobutyrivibrio (P = 0.10) and Lachnospiraceae FD2005 (P = 0.09) (both from Lachnospiraceae family) and Ruminobacter (P = 0.09) tended to decrease in CO3 compared to Control in liquid fraction. Butyrate molar proportion was negatively correlated to Ruminococcaceae (r = –0.55) in solid fraction and positively correlated to Pseudobutyrivibrio (r = 0.61) and Lachnospiraceae FD2005 (r = 0.61) in liquid. Our results indicate that source of Mg has an impact on bacterial taxa associated with ruminal butyrate synthesis, which is important for epithelial health and fatty acid synthesis.UCR::Vicerrectoría de Docencia::Ciencias Agroalimentarias::Facultad de Ciencias Agroalimentarias::Escuela de ZootecniaUCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Agroalimentarias::Centro de Investigación en Nutrición Animal (CINA

Effects of lactic acid-producing bacteria as direct-fed microbials on the ruminal microbiome

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 × 109 cfu/g, and Propionibacterium freudenreichii, 2 × 109 cfu/g [MLP = 0.01% of diet dry matter (DM)]; and 2 different levels of L. plantarum, 1.35 × 109 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 Butyrivibrio_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 NH3-N daily outflow from the fermentors. 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 NH3-N concentration without altering pH and lactic acid concentration.UCR::Vicerrectoría de Docencia::Ciencias Agroalimentarias::Facultad de Ciencias Agroalimentarias::Escuela de Zootecni

Production, physiological response, and calcium and magnesium balance of lactating Holstein cows fed different sources of supplemental magnesium with or without ruminal buffer

The objective of this study was to evaluate the effects of dietary replacement of magnesium oxide (MgO) with calcium-magnesium hydroxide [CaMg(OH)2] and its interaction with ruminal buffer (sodium sesquicarbonate) supplementation on production, Ca and Mg balance, and overall physiological response of mid-lactation Holstein dairy cows. Sixty cows averaging 40.5 ± 7.0 kg of milk/d were used. Treatments were assigned following a 2 × 2 factorial arrangement: (1) MgO, (2) MgO + buffer, (3) CaMg(OH)2, or (4) CaMg(OH)2 + buffer. Diets were formulated to have 16.5% of crude protein, 1.82 Mcal/kg of net energy for lactation, 0.67% Ca, 0.39% P, and 0.25% Mg, all on a dry matter (DM) basis. Treatments were individually top dressed. Milk production, composition, and DM intake were evaluated. A subsample of 20 cows were randomly selected for the evaluation of Ca and Mg balance, blood gases, and electrolytes. Ruminal fluid was also collected for evaluation of pH and Ca and Mg solubility. Effects of Mg source, buffer, and the interaction Mg source × buffer were analyzed through orthogonal contrasts. An interaction of Mg source × buffer was found for DM intake and feed efficiency, in which cows fed CaMg(OH)2 had a similar feed efficiency regardless of ruminal buffer inclusion; however, when cows were fed MgO, the inclusion of buffer reduced feed efficiency. No effects on body weight and milk yield were observed. Buffer addition tended to increase the concentrations of fat, protein, and solids-not-fat, without affecting the yields of these milk components. Magnesium source and buffer did not affect ruminal fluid, blood, urine, or fecal pH; however, buffer supplementation increased urinary pH. Treatment with CaMg(OH)2 increased blood concentration of HCO3 −, total CO2, and base excess compared with cows fed MgO. No differences were observed in the ruminal solubility of Ca and Mg or on milk or urinary Ca and Mg excretion. Greater plasma Mg concentration was observed for animals fed MgO compared with cows fed CaMg(OH)2; however, both sources were above the threshold recommended in the literature for dairy cows. Also, a reduction in fecal Mg excretion was observed in animals fed CaMg(OH)2. In summary, we provide evidence that CaMg(OH)2 could replace MgO without affecting performance, overall physiological response, or Ca and Mg balance of mid-lactating dairy Holstein cows.UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Agroalimentarias::Centro de Investigación en Nutrición Animal (CINA)UCR::Vicerrectoría de Docencia::Ciencias Agroalimentarias::Facultad de Ciencias Agroalimentarias::Escuela de Zootecni

Effects of partially replacing dietary corn with molasses, condensed whey permeate, or treated condensed whey permeate on ruminal microbial fermentation

Corn is a feedstuff commonly fed to dairy cows as a source of energy. The objective of this study was to evaluate whether partially replacing dietary corn with molasses or condensed whey permeate, in lactating dairy cow diets in a dual-flow continuous culture system, can maintain nutrient digestibility by ruminal microorganisms. Furthermore, this study evaluated whether treating condensed whey permeate before feeding could aid the fermentation of the condensed whey permeate in the rumen. Eight fermentors were used in a 4 × 4 replicated Latin square with 4 periods of 10 d each. The control diet (CON) was formulated with corn grain, and the other diets were formulated by replacing corn grain with either sugarcane molasses (MOL), condensed whey permeate (CWP), or treated condensed whey permeate (TCWP). Diets were formulated by replacing 4% of the diet dry matter (DM) in the form of starch from corn with sugars from the byproducts. Sugars were defined as water-soluble carbohydrates (WSC) in the rations. The fermentors were fed 52 g of DM twice daily of diets containing 17% crude protein, 28% neutral detergent fiber, and 45% nonfiber carbohydrates. Liquid treatments were pipetted into each fermentor. After 7 d of adaptation, samples were collected for analyses of volatile fatty acids (VFA), lactate, and ammonia, and fermentors’ pH were measured at time points after the morning feeding for 3 d. Pooled samples from effluent containers were collected for similar analyses, nutrient flow, and N metabolism. Data were statistically analyzed using Proc MIXED of SAS version 9.4 (SAS Institute Inc.); fixed effects included treatment and time, and random effects included fermentor, period, and square. The interaction of treatment and time was included for the kinetics samples. The TCWP and MOL treatments maintained greater fermentor pH compared with CWP. Total VFA concentration was increased in CWP compared with MOL. The acetate:propionate ratio was increased in TCWP compared with CON, due to tendencies of increased acetate molar proportion and decreased propionate molar proportion in TCWP. Lactate concentration was increased in MOL. Digestibility of WSC was increased in the diets that replaced corn with byproducts. The partial replacement of 4% of DM from corn starch with the sugars in byproducts had minimal effects on ruminal microbial fermentation and increased pH. Treated CWP had similar effects to molasses.UCR::Vicerrectoría de Docencia::Ciencias Agroalimentarias::Facultad de Ciencias Agroalimentarias::Escuela de Zootecni

Effects of replacing magnesium oxide with calcium-magnesium carbonate with or without sodium bicarbonate on ruminal fermentation and nutrient flow in vitro

The objective of this study was to evaluate the effects of replacing magnesium oxide (MgO) with calcium-magnesium carbonate [CaMg(CO3)2] on ruminal fermentation with or without the addition of sodium bicarbonate (NaHCO3). Eight fermentors of a dualflow continuous-culture system were distributed in a replicated (2) 4 × 4 Latin square design in a 2 × 2 factorial arrangement of treatments (magnesium sources × NaHCO3). The treatments tested were 0.21% MgO [MgO; dry matter (DM) basis; 144.8 mEq of dietary cation-anion difference (DCAD)]; 0.21% MgO + 0.50% NaHCO3 (MgO+NaHCO3; DM basis; 205.6 mEq of DCAD); 1.00% CaMg(CO3)2 [CaMg(CO3)2; DM basis; 144.8 mEq of DCAD]; and 1.00% CaMg(CO3)2 + 0.50% NaHCO3 [CaMg(CO3)2+NaHCO3; DM basis; 205.6 mEq of DCAD]. Diets were formulated to have a total of 0.28% of Mg (DM basis). The experiment consisted of 40 d, which was divided into 4 periods of 10 d each, where 7 d were used for adaptation and 3 d for sampling to determine pH, volatile fatty acids (VFA), ammonia (NH3-N), lactate, mineral solubility, N metabolism, and nutrient digestibility. The effects of Mg source [MgO vs. CaMg(CO3)2], NaHCO3 (with vs. without), and the interaction were tested with the MIXED procedure of SAS version 9.4 (SAS Institute). There was no Mg source × NaHCO3 interaction in the pH variables and mineral solubility, and Mg sources evaluated did not affect the variables related to ruminal pH and solubility of Mg. On the other hand, the inclusion of NaHCO3 increased the pH daily average, independent of Mg source, which led to a reduced time that pH was below 5.8 and decreased area under the curve. Total VFA and lactate concentration were similar among treatments regardless of NaHCO3 and Mg source; however, the molar proportion of isobutyrate and NH3-N concentration were lower in diets with CaMg(CO3)2 compared with MgO. Moreover, NaHCO3 inclusion increased NH3-N, total daily NH3-N flow, isobutyrate concentration, and acid detergent fiber digestibility. Our results showed that CaMg(CO3)2 leads to a lower NH3-N concentration and isobutyrate proportion. Therefore, because most of the tested variables were not significantly different between MgO and CaMg(CO3)2 when combined or not with NaHCO3, CaMg(CO3)2 can be a viable alternative source to replace MgO in dairy cow diets without affecting mineral solubility, ruminal pH, nutrient digestibility, total VFA, and the main ruminal VFA. Although Mg sources are known to have an alkalizing effect, NaHCO3 inclusion in diets with Mg supplementation allowed an increase in ruminal pH, as well as an increase in isobutyrate and NH3-N flowUCR::Vicerrectoría de Docencia::Ciencias Agroalimentarias::Facultad de Ciencias Agroalimentarias::Escuela de Zootecni

The Effects of Incremental Doses of Aflatoxin B1 on In Vitro Ruminal Nutrient Digestibility and Fermentation Profile of a Lactating Dairy Cow Diet in a Dual-Flow Continuous Culture System

Aflatoxin B1 (AFB1 ) is a mycotoxin known to impair human and animal health. It is also believed to have a deleterious effect on ruminal nutrient digestibility under in vitro batch culture systems. The objective of this study was to evaluate the effects of increasing the dose of AFB1 on ruminal dry matter and nutrient digestibility, fermentation profile, and N flows using a dual-flow continuous culture system fed a diet formulated for lactating dairy cows. Eight fermenter vessels were used in a replicated 4 × 4 Latin square design with 10 d periods (7 d adaptation and 3 d sample collection). Treatments were randomly applied to fermenters on diet DM basis: (1) 0 µg of AFB1/kg of DM (Control); (2) 50 µg of AFB1/kg of DM (AF50); (3) 100 µg of AFB1/kg of DM (AF100); and (4) 150 µg of AFB1/kg of DM (AF150). Treatments did not affect nutrient digestibility, fermentation, and N flows. Aflatoxin B1 concentration in ruminal fluid increased with dose but decreased to undetectable levels after 4 h post-dosing. In conclusion, adding incremental doses of AFB1 did not affect ruminal fermentation, digestibility of nutrients, and N flows in a dual-flow continuous culture system fed diets formulated for lactating dairy cows.UCR::Vicerrectoría de Docencia::Ciencias Agroalimentarias::Facultad de Ciencias Agroalimentarias::Escuela de Zootecni

Effects of exogenous amylolytic or fibrolytic enzymes inclusion on in vitro fermentation of lactating dairy cow diets in a dual-flow continuous-culture system

The objective of this study was to determine the effects of including exogenous amylolytic or fibrolytic enzymes in a diet for high-producing dairy cows on in vitro ruminal fermentation. Eight dual-flow continuousculture fermentors were used in a replicated 4 × 4 Latin square. The treatments were control (CON), a xylanase and glucanase mixture (T1), an α-amylase mixture (T2), or a xylanase, glucanase, and α-amylase mixture (T3). Treatments were included at a rate of 0.008% of diet dry matter (DM) for T1 and T2 and at 0.02% for T3. All treatments replaced the equivalent amount of soybean meal in the diet compared with CON. All diets were balanced to have the same nutrient composition [30.2% neutral detergent fiber (NDF), 16.1% crude protein (CP), and 30% starch; DM basis], and fermentors were fed 106 g/d divided into 2 feedings. At each feeding, T2 was pipetted into the respective fermentor and an equivalent amount of deionized water was added to each fermentor to eliminate potential variation. Experimental periods were 10 d (7 d for adaptation and 3 d for sample collection). Composite samples of daily effluent were collected and analyzed for volatile fatty acids (VFA), NH3-N, and lactate concentrations, degradability of DM, organic matter, NDF, CP, and starch, and flow and metabolism of N. Samples of fermentor contents were collected from each fermentor at 0, 1, 2, 4, 6, and 8 h after feeding to determine kinetics of pH, NH3-N, lactate, and VFA concentrations over time. All data were analyzed using PROC GLIMMIX of SAS (SAS Institute Inc.), and the repeated variable of time was included for kinetics measurements. Treatment did not affect mean pH, degradability, N flow and metabolism, or the concentrations of VFA, NH3-N, or lactate in the effluent samples. Treatment did not affect pH, acetate:propionate ratio, or the concentrations of lactate, NH3-N, total VFA, acetate, propionate, butyrate, isobutyrate, valerate, or caproate. However, the concentration of total VFA tended to change at each time point depending upon the treatment, and T2 tended to have a greater proportion of 2-methylbutyrate and isovalerate than CON, T1, or T3. As 2-methylbutyrate and isovalerate are branchedchain VFA that are synthesized from branched-chain amino acids, T2 may have an increased fermentation of branched-chain amino acids or decreased uptake by fibrolytic microorganisms. Although we did not observe changes in N metabolism due to the enzymes, there could be changes in microbial populations that utilize branched-chain VFA. Overall, the tested enzymes did not improve in vitro ruminal fermentation in the diet of high-producing dairy cows.UCR::Vicerrectoría de Docencia::Ciencias Agroalimentarias::Facultad de Ciencias Agroalimentarias::Escuela de Zootecni