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Distinct blood and milk 18-carbon fatty acid proportions and buccal bacterial populations in dairy cows differing in reticulorumen pH response to dietary supplementation of rapidly fermentable carbohydrates
Laboratory for Animal Nutrition and Animal Product Quality (Lanupro), Department of Animal Sciences and Aquatic Ecology, Ghent University, 9000 Gent, Belgium
Laboratory for Animal Nutrition and Animal Product Quality (Lanupro), Department of Animal Sciences and Aquatic Ecology, Ghent University, 9000 Gent, Belgium
* Current address: Research Group Marine Biology, Department of Biology, Ghent University, 9000 Gent, Belgium.
B. Vlaeminck
Footnotes
* Current address: Research Group Marine Biology, Department of Biology, Ghent University, 9000 Gent, Belgium.
Affiliations
Laboratory for Animal Nutrition and Animal Product Quality (Lanupro), Department of Animal Sciences and Aquatic Ecology, Ghent University, 9000 Gent, Belgium
Laboratory for Animal Nutrition and Animal Product Quality (Lanupro), Department of Animal Sciences and Aquatic Ecology, Ghent University, 9000 Gent, Belgium
Laboratory for Animal Nutrition and Animal Product Quality (Lanupro), Department of Animal Sciences and Aquatic Ecology, Ghent University, 9000 Gent, Belgium
Nine Holstein dairy cows were fed diets with increasing proportions of rapidly fermentable carbohydrates (RFCH) to investigate the effect on reticular pH, milk fat content (MFC), 18-carbon fatty acid proportions in blood plasma and milk, and bacterial community in buccal swab samples. Inter-animal variation was expected in terms of reticular pH response upon higher RFCH proportions, which would be reflected in the occurrence or not of milk fat depression (MFD). Moreover, this variation in occurrence of MFD was hypothesized to be related to differences in blood and milk fatty acid proportions and in the bacterial community in buccal samples. Cows were fed a total mixed ration throughout the experiment, which consisted of 4 periods: adaptation (d 0–4) and low (d 5–18), increasing (d 19–24), and high RFCH (d 25–28). During the increasing RFCH period, the standard concentrate (211 g of starch/kg of dry matter) was gradually and partly replaced by a concentrate high in RFCH (486 g of starch/kg of dry matter). The reticular pH was measured using a bolus and the time below pH 6.00 was calculated on a daily basis. On d 13, 14, 25, 27, and 28, plasma and milk samples were collected and analyzed for 18-carbon fatty acid proportions, and buccal swabs were collected for bacterial community analysis based on 16S rRNA gene amplicon sequencing. Inter-animal variation was observed in terms of reticular pH, which allowed us to divide the cows into 2 groups: tolerant (time below pH 6.00 ≤ 0.1 h/d) and susceptible cows (time below pH 6.00 ≥ 1.26 h/d). The lower reticular pH of susceptible cows was accompanied by lower MFC. Both groups already differed in reticular pH and MFC during the low-RFCH period. Furthermore, higher RFCH amounts did not decrease the reticular pH in either of the 2 groups. Nevertheless, MFD was observed in both groups during the high-RFCH period compared with the low-RFCH period. Lower MFC in animals with lower reticular pH or during the high-RFCH period was associated with a shift in 18-carbon fatty acids toward trans-10 at the expense of trans-11 intermediates, which was observed in plasma as well as in milk samples. Moreover, lower MFC was accompanied by shifts in the relative abundance of specific bacteria in buccal samples. Genera Dialister, Sharpea, Carnobacterium, Acidaminococcus, and uncultured genera belonging to the Betaproteobacteria were more abundant in situations with greater trans-10 proportions.
Diet-induced milk fat depression (MFD) in dairy cattle is characterized by a reduction in milk fat content and yield, and alterations in milk fatty acid (FA) composition (
). The biohydrogenation theory established that MFD is caused by alterations in rumen biohydrogenation of dietary PUFA. When high-starch diets are fed or rumen pH is low, typical pathways of rumen biohydrogenation are altered to produce unique FA intermediates that inhibit milk fat synthesis (
). Under normal rumen conditions, 18:2n-6 is mainly isomerized to cis-9,trans-11 CLA, which is further hydrogenated to trans-11 18:1 and, ultimately, to 18:0 (
). However, under milk fat-depressing conditions, a shift in biohydrogenation pathways occurs toward the formation of trans-10 intermediates (i.e., trans-10,cis-12,cis-15 CLnA, trans-10,cis-15 18:2, trans-10,cis-12 CLA, and trans-10 18:1), referred to as the trans-11 to trans-10 shift (
Of extreme interest in this respect is the inter-animal variation in rumen pH response to dietary supplementation of rapidly fermentable carbohydrates (RFCH;
The relationship between rumen acidosis resistance and expression of genes involved in regulation of intracellular pH and butyrate metabolism of ruminal epithelial cells in steers.
Variability in susceptibility to acidosis among high producing mid-lactation dairy cows is associated with rumen pH, fermentation, feed intake, sorting activity, and milk fat percentage.
). Animals that experience less severe decreases in rumen pH upon a dietary RFCH challenge might also be less susceptible to alterations in ruminal biohydrogenation pathways and the associated MFD.
Rumen bacteria play an important role in biohydrogenation of PUFA (
). Indeed, in vitro studies with pure cultures of bacteria from the gastrointestinal tract of both ruminants and humans revealed that (Pseudo)butyrivibrio spp. are capable of trans-11 formation (e.g.,
) are capable of producing trans-10 isomers from 18:2n-6. However, the role of these microbes in ruminal biohydrogenation of 18-carbon FA is not well known. As such, the rumen microbial etiology of the trans-11 to trans-10 shift is not well understood yet.
The current study investigated the changes in 18-carbon FA proportions and alterations in bacterial populations of dairy cows upon a dietary RFCH challenge. We hypothesized that inter-animal variation would be observed in response to the diet in terms of reticular pH, and that this would be reflected in the occurrence or not of MFD. Moreover, this variation in occurrence of MFD was hypothesized to be related to differences in FA proportions and bacterial community. As such, we aimed to gain more insight into the microbiology of the trans-11 to trans-10 shift by investigating associations between specific 18-carbon FA proportions and bacterial populations.
MATERIALS AND METHODS
Animals, Experimental Design, Diets, and Management
All experimental procedures involving animals were approved by the Central Committee on Animal Experiments (AVD24246002017848, appendix 2) and the Institute for Animal Welfare of the Schothorst Feed Research B.V. (RM17–15-LRA-57). Ten Holstein-Friesian dairy cows (1 primiparous, 9 multiparous; 638 ± 74 kg of BW; 157 ± 43 DIM; 27.43 ± 7.77 kg/d of milk yield) were selected from a herd of 125 cows based on their milk FA profile in early lactation (as described by
Susceptibility of dairy cows to subacute ruminal acidosis is reflected in milk fatty acid proportions, with C18:1 trans-10 as primary and C15:0 and C18:1 trans-11 as secondary indicators.
Susceptibility of dairy cows to subacute ruminal acidosis is reflected in milk fatty acid proportions, with C18:1 trans-10 as primary and C15:0 and C18:1 trans-11 as secondary indicators.
), it was hypothesized to observe distinct rumen pH profiles within those animals. In early lactation, half of the selected animals (n = 5) showed a high proportion in milk fat of trans-10 18:1 at 3 wk in milk (>0.31 g/100 g of FA), a high level of 15:0 (on average, ≥1.18 g/100 g of FA over the 4 wk in milk), and a sharp decrease of trans-11 18:1 (Δ ≥0.25 g/100 g of FA during the 4 wk in milk). Those animals were hypothesized to be more susceptible to increased amounts of RFCH. Their counterparts (n = 5) had a low milk fat proportion of trans-10 18:1 at 3 wk in milk (<0.23 g/100 g of FA), an average 15:0 proportion of 0.99 g/100 g of FA or lower, and a rather stable trans-11 18:1 proportion in milk fat during early lactation. In the current experiment, the selected animals were housed in a freestall barn and were subjected to increasing amounts of RFCH in the diet (explained below). Unfortunately, 1 cow had to be withdrawn from the experiment due to an accident unrelated to the experiment.
The experiment lasted for 28 d and was divided into 4 periods based on the relative amount of RFCH in the diet: (1) adaptation period (d 0–4); (2) low-RFCH period (d 5–18); (3) increasing RFCH period (d 19–24); and (4) high-RFCH period (d 25–28). Cows were fed a TMR (basal diet/concentrate, on average, 670/330, wt/wt DM basis) ad libitum with a limited amount of residual feed to prevent cows from feed selection. Refusals of TMR were weighed daily before the morning feeding and used to adjust the amounts of feed offered. The basal diet (Table 1) consisted of (g/kg of DM) grass silage (514), maize silage (367), soybean meal (56.5), rapeseed meal (56.5), and a mineral premix (6.0). Two different concentrates were used in our study: a standard concentrate (concentrate A) and a concentrate high in RFCH (particularly supplied through wheat; concentrate B; Table 1). Both concentrates were formulated to be isoenergetic and isoproteic. During the low-RFCH period, cows only received concentrate A, which was gradually and partly replaced by concentrate B during the increasing RFCH period. The ratio of concentrate A to concentrate B remained constant in the high-RFCH period. The experimental protocol is presented in Table 2. Cows were continuously monitored based on their milk production, reticular pH, and occurrence of health problems. Some cows showed very low reticular pH values and were therefore fed less concentrate than originally planned. On the other hand, other cows did not respond on increasing RFCH in the diet, according to the reticular pH monitoring, and extra concentrate B was provided to these animals. Hence, due to large variation between cows in response to increasing amounts of concentrate B, differences in the ratio of concentrate A and concentrate B within the same period occurred (Supplemental Table S1; https://doi.org/10.3168/jds.2018-15823).
Table 1Ingredients and chemical composition of the basal diet and concentrates A and B
Two different concentrates were used in our study: a standard concentrate (concentrate A) and a concentrate high in rapidly fermentable carbohydrates (particularly supplied through wheat; concentrate B)
Contains: 140 g of Ca, 0 g of P, 14 g of Mg, 7.5 g of Na, 0.3 g of K, 11.5 g of Cl, 0.3 g of S, 2,000 mg of Cu, 4,025 mg of Zn, 3,050 mg of Mn, 40 mg of Se, 75 mg of Co, 124 mg of I, 600,000 IU of vitamin A, 120,000 IU of vitamin D3, and 5,000 IU of vitamin E.
NEL calculated based on the Dutch net energy evaluation system (VEM; Van Es, 1975).
MJ/kg of DM
6.7
6.7
Fatty acid composition, g/kg
16:0
3.01
2.84
1.20
18:0
0.40
0.66
0.28
cis-9 18:1
2.71
4.82
0.99
18:2n-6
6.16
9.72
3.67
18:3n-3
6.27
0.97
0.35
Total fatty acids
19.0
19.0
6.35
1 Two different concentrates were used in our study: a standard concentrate (concentrate A) and a concentrate high in rapidly fermentable carbohydrates (particularly supplied through wheat; concentrate B)
2 Contains: 140 g of Ca, 0 g of P, 14 g of Mg, 7.5 g of Na, 0.3 g of K, 11.5 g of Cl, 0.3 g of S, 2,000 mg of Cu, 4,025 mg of Zn, 3,050 mg of Mn, 40 mg of Se, 75 mg of Co, 124 mg of I, 600,000 IU of vitamin A, 120,000 IU of vitamin D3, and 5,000 IU of vitamin E.
3 NEL calculated based on the Dutch net energy evaluation system (VEM;
Two different concentrates were used in our study: a standard concentrate (concentrate A) and a concentrate high in RFCH (particularly supplied through wheat; concentrate B).
Mean values are reported for the low- and high-RFCH period. The value of the last day is reported for the increasing RFCH period.
Day
Period
Basal diet
Concentrate A
Concentrate B
5–18
Low RFCH
676
324
0
19–24
Increasing RFCH
678
77
245
25–28
High RFCH
659
75
266
1 Two different concentrates were used in our study: a standard concentrate (concentrate A) and a concentrate high in RFCH (particularly supplied through wheat; concentrate B).
2 RFCH = rapidly fermentable carbohydrates.
3 Mean values are reported for the low- and high-RFCH period. The value of the last day is reported for the increasing RFCH period.
Diets were offered as 2 equal meals at 0730 and 1500 h. Animals had access to a constant supply of fresh water and were milked twice a day at 0530 and 1600 h. Individual milk yield was recorded at each milking.
Measurement of Reticular pH and Grouping of Animals
During the whole experiment, the reticular pH was measured every 10 min using a SmaXtec Premium bolus (SmaXtec Animal Care GmbH, Graz, Austria). Boli were introduced before the experiment using an oral balling gun. Various reticular pH parameters were calculated on a daily basis during the low- and high-RFCH period (i.e., mean pH, pH nadir, time below pH 5.60, time below pH 5.80, and time below pH 6.00; Table 3). Based on the time below pH 6.00, the animals were divided into 2 groups; that is, tolerant (n = 4) and susceptible (n = 5) cows. During both the low- and high-RFCH periods, the reticular pH of tolerant cows hardly decreased below 6.00 (time below pH 6.00 ≤0.10 h/d), whereas the reticular pH of susceptible cows was lower than 6.00 at least during 1 hour per day, both in the high- as well as in the low-RFCH period (Table 3).
Table 3Lactation and mean reticular pH parameters of the individual cows that were subjected to increased amounts of rapidly fermentable carbohydrates (RFCH) in the diet during different periods (low and high RFCH)
As our cows were not ruminally cannulated and oral intubation or rumenocentesis could not be applied for ethical reasons, we collected alternative samples (i.e., milk, saliva, blood, and buccal swabs). As the diet of the cows changed daily during the increasing RFCH period, samples were only collected during the low- and high-RFCH periods, in which the ratio of basal diet to concentrate was more stable.
From d 11 to 18 (low-RFCH period) and 25 to 28 (high-RFCH period), milk samples were collected daily at each milking (morning and afternoon) and stored with preservative (Bronopol; Qlip, Zutphen, the Netherlands) at −20°C until analysis of long-chain FA (LCFA).
On d 13 and 14 (low-RFCH period) and 25, 27, and 28 (high-RFCH period), samples of saliva, blood plasma, and buccal swabs were collected 3 h after feeding (based on
). Saliva was collected using a sponge (Koala Universal Sponge; Essef, Sint-Eloois-Winkel, Belgium), which was cut into smaller pieces (approximately 10 × 5 × 2 cm). The sponge was inserted into the mouth of the animal for approximately 1 min using a forceps (∼25 cm long). The collected saliva was then pressed out into plastic tubes, which were placed on ice and then transferred to −20°C until further analysis. After thawing, subsamples (1 mL) were collected in glass tubes, stored at −20°C, and freeze-dried before LCFA analysis. As ruminants regularly regurgitate large amounts of rumen material to their oral cavities, those saliva samples were used as a noninvasive alternative to assess the 18-carbon FA composition of the rumen.
Blood was collected in lithium heparin–containing tubes. After collection, blood was centrifuged at 3,000 × g for 10 min at room temperature. The supernatant was separated and transferred into plastic tubes and stored at −20°C until analysis of LCFA.
Buccal swabs were collected using a sterile cotton roll (10 × 38 mm; Alan dental rolls, Alan & Co., Verviers, Belgium) according to
. The sterile cotton roll was held with a sterile forceps (∼25 cm long) and inserted into the mouth of the animal and swabbed several times across the inner side of the left cheek. The cotton roll was then placed in a sterile glass tube, which was placed on ice and then transferred to −20°C until further analysis. These buccal swabs were used as a noninvasive proxy to assess the composition of the rumen bacterial community (
Fatty acid methyl esters were prepared from freeze-dried saliva samples and plasma samples using a direct transesterification procedure as described by
. Briefly, toluene (2 mL) containing the internal standard (21:0; Sigma Aldrich, Diegem, Belgium) and methanolic NaOH (2 mL; 0.5 M) were added and the mixture was incubated at 70°C for 60 min. This was followed by 30 min of incubation at 50°C after addition of methanolic HCl (3 mL), prepared by dissolving acetyl chloride in methanol (5/1, vol/vol). The FAME were extracted with hexane.
In addition, total plasma lipids were extracted using the BUME method (adapted from
) before separation into 4 lipid classes (i.e., free FA, phospholipids, cholesterol esters, and triacylglycerols) using solid phase extraction columns according to
. The internal standard (TAG-13:0; Sigma Aldrich) was added before milk fat extraction to quantify the total fat content based on the sum of individual FA.
Composition Analysis
Analysis of FAME was carried out using a GC (HP7890A; Agilent Technologies, Diegem, Belgium) equipped with a fused silica capillary column (SP-2560; 75 m × 0.18 mm i.d. × 0.14 µm film thickness; Supelco Analytical, Bellefonte, PA) and a flame ionization detector. A combination of 2 oven temperature programs was used in our study to achieve separation of most 18:1 isomers (
Combining results of two GC separations partly achieves determination of all cis and trans 16:1, 18:1, 18:2 and 18:3 except CLA isomers of milk fat as demonstrated using Ag-ion SPE fractionation.
). For the first GC run, the temperature program was initially 70°C for 2 min, increasing by 15°C/min to 150°C, followed by a second increase at 1°C/min up to 165°C and holding for 12 min, followed by a third increase at 2°C/min to 170°C, held at 170°C for 5 min, increased at 5°C/min to 215°C, and held at 215°C for 20 min. For the second GC run, the temperature program was, at the time of sample injection, column temperature of 70°C, increased at 50°C/min to 175°C and maintained for 13 min, followed by a final increase at 5°C/min to 215°C, and maintained for 20 min. For both GC runs, inlet and detector temperatures were 250°C and 255°C, respectively. The injection volume and split ratio depended on sample type and GC run. Hydrogen was used as the carrier gas at a flow rate of 1 mL/min. Identities of peaks were determined using mixtures of methyl ester standards (GLC463, Nu-Chek Prep, Elysian, MN; cis-9,trans-11 CLA and trans-10,cis-12 CLA, Larodan 279, Fine Chemicals AB, Malmö, Sweden). Quantification of FA was based on the area of the internal standard and on the conversion of peak areas to the weight of FA by a theoretical response factor for each FA (
Evaluation of sequential methods for the determination of butterfat fatty acid composition with emphasis on trans-18:1 acids. Application to the study of seasonal variations in French butters.
Before genomic DNA extraction, the cotton roll was removed from the tube with a sterile forceps, and approximately one-third of the swab was cut off using a sterile pair of scissors and immediately submerged in 2 mL of PBS buffer (
) for approximately 18 h. After centrifugation at 4°C for 10 min at 14,000 × g, the supernatant was transferred into a sterile 7-mL screw-cap tube containing zirconia beads, after which genomic DNA extraction was performed using the repeated bead beating plus column purification (RBB+C) method (adapted from
). The yield and quality of extracted DNA was determined using a NanoDrop spectrophotometer (VWR International BVBA, Leuven, Belgium).
Illumina Library Generation and Data Mining
Bacterial 16S rRNA gene amplicon sequencing (V3-V4 region) using Illumina MiSeq (Illumina Inc., San Diego, CA) technology (2 × 300 bp) was performed by Macrogen Sequencing Service (Macrogen Korea, Seoul, Korea). Preparation of the amplicon barcoded library (primers: 344F and 806R;
The amplicon sequencing data set was demultiplexed and barcodes were clipped off by the sequencing service provider. Reads were analyzed using the Quantitative Insights Into Microbial Ecology (QIIME) bioinformatics pipeline, version 1.9.1 (
). This resulted in an average of 76,142 ± 11,216 sequences per sample (mean ± SD). The subsequent analysis, picking operational taxonomic units (OTU), assigning taxonomy, inferring phylogeny, and creating OTU tables, were also performed by QIIME software (
. Afterward, all potential rumen bacteria were retained based on cow rumen samples from our laboratory (J. Jeyanathan, Ghent University, Ghent, Belgium, personal communication) and based on
Global Rumen Census Collaborators Rumen microbial community composition varies with diet and host, but a core microbiome is found across a wide geographical range.
. The OTU with less than 0.005% of the total number of sequences were removed. To ensure the comparability between samples, rarefied OTU sets (2,227 sequences per sample) were used for further analysis.
Two measures of α diversity were calculated in QIIME (
): (1) Shannon entropy, an indicator of species richness and evenness in community structure, and (2) the number of OTU observed. Next to this, β diversity indices between samples were determined in QIIME (
). Sequence data have been deposited in the National Center for Biotechnology Information (NCBI; https://www.ncbi.nlm.nih.gov/) database under accession number SRP136158.
Statistical Analysis
DMI, Reticular pH, Milk Fat Content, Milk Production, and 18-Carbon FA
Data were analyzed using the MIXED procedure of SAS (version Enterprise Guide 7.1; SAS Institute Inc., Cary, NC) by the following model
Yijkl = µ + Si + Pej + Dk + Tl + Si × Pej + εijkl,
where Yijkl = response variable, µ = average, Si is the fixed effect of animal group (i = tolerant or susceptible), Pej is the fixed effect of period (j = low or high RFCH), Dk is the random effect of sampling day, Tl is the random effect of time of sampling (l = morning or afternoon, only applicable for milk samples), Si × Pej is the interaction between group and period, and εijkl is the residual error term. Least squares means are reported with treatment effects declared significant at P < 0.05. A Tukey-Kramer multiple comparison test was used to evaluate significant differences.
Bacterial Community
Both α diversity indices were analyzed in SAS (version Enterprise Guide 7.1; SAS Institute Inc.) by the model described above. In addition, the nonparametric permutational MANOVA-based statistical test ANOSIM was used in QIIME (
) to determine differences in overall bacterial community between groups or periods based on β diversity indices. To analyze for differences in taxa relative abundance (at species level), a first screening was performed by the Kruskal-Wallis test in QIIME (
). Bacterial taxa that showed differences between groups or periods or both based on this analysis were considered for further analysis in SAS (version Enterprise Guide 7.1; SAS Institute Inc.) by the model described above. When assumptions of normality or constant variance were not fulfilled, a square root or a sin-square root transformation was performed before analysis. In addition, Spearman rank correlation was used to check the correlation between different milk and plasma parameters and the abundance of different bacterial taxa (at species level) using QIIME (
The mean reticular pH parameters are presented in Table 4. Grouping of the animals was based on the time below pH 6.00. Moreover, the discrepancy in reticular pH between groups was observed in all pH parameters (group: P < 0.05): mean pH, pH nadir, and time below pH 5.60, 5.80, and 6.00. Unexpectedly, gradual shifting from the low- to the high-RFCH period did not decrease the mean or minimum reticular pH or increase the time below a certain pH threshold (period: P > 0.05) in both groups of animals. In tolerant cows, the mean reticular pH was even higher in the high-RFCH period as compared with the low-RFCH period (group × period: P = 0.031).
Table 4Reticular pH variables from cows which were tolerant or susceptible
The DMI, milk fat content, and milk yield in the low- and high-RFCH period are presented in Table 5. Susceptible cows showed a lower DMI (group: P = 0.004), milk fat content (group: P = 0.004), and milk production (group: P = 0.010) as compared with the tolerant ones. Irrespective of group of cows, the milk fat content decreased in the high-RFCH period (period: P < 0.001) without changing milk production (period: P = 0.684). In that period, the milk fat content of tolerant cows decreased to the level of the susceptible cows in the low-RFCH period, whereas the milk fat content of susceptible cows decreased further.
Table 5Dry matter intake, milk fat content, and milk production from cows that were tolerant or susceptible
As mainly 18-carbon FA are known to induce MFD, we traced shifts within this group of FA in particular. Hence, we expressed the proportions of these FA relative to the sum of all 18-carbon FA to avoid confounding factors when expressed relative to total FA (e.g., alterations in odd- and branched-chain FA, which might also be influenced by varying dietary RFCH).
Blood Plasma
Proportions of different 18-carbon FA and total 18-carbon FA in blood plasma are presented in Table 6. Total 18-carbon FA (group: P < 0.001) and the proportion of 18:2n-6 (group: P = 0.072) and 18:3n-3 (group: P < 0.001) were higher or tended to be higher in tolerant cows as compared with susceptible cows. Plasma proportions of total trans-11 (t11) and total trans-10 (t10) intermediates were higher (group: P = 0.032 and P < 0.001, respectively) in susceptible cows as compared with tolerant cows. As the difference between groups was greater for t10 intermediates than for t11 intermediates, the ratio of t10 to t11 intermediates was also higher in plasma of susceptible cows (group: P = 0.002). The higher proportion of t11 intermediates was mainly caused by a higher proportion of cis-9,trans-11 CLA (group: P = 0.002), as no differences were observed between groups in other t11 intermediates. The higher proportion of t10 intermediates in susceptible cows was caused by higher proportions of trans-10,cis-12,cis-15 CLnA (group: P < 0.001) and trans-10 18:1 (group: P = 0.006). Plasma proportions of trans-10,cis-12 CLA were extremely low in all samples (<0.10 g/100 g of C18, data not shown). In addition, the proportions of total 18:1 FA and 18:0 were also higher in plasma of susceptible cows (group: P = 0.029 and P < 0.001, respectively).
Table 6Proportions of different 18-carbon fatty acids (g/100 g of C18) in blood plasma collected from cows that were tolerant or susceptible
c = cis; t = trans; CLnA = conjugated linolenic acid; t10 and t11 = sum of all 18-carbon biohydrogenation intermediates containing a double bond in the trans configuration either at the 10th or the 11th carbon atom (from the carboxyl end); t10:t11 = ratio of trans-10 intermediates to trans-11 intermediates.
The results from 1 cow at 1 sampling time were omitted before statistical analysis (outlier).
RFCH period
SEM
P-value
Tolerant (n = 4)
Susceptible (n = 5)
Low
High
Low
High
Group (S)
Period (Pe)
S × Pe
Total C18 fatty acids, mg/mL
2.05
1.90
1.45
1.30
0.080
<0.001
0.067
0.991
18:0
18.26
17.81
19.36
19.10
0.312
<0.001
0.272
0.758
t6 18:1 + t7 18:1 + t8 18:1
0.17
0.19
0.18
0.20
0.014
0.382
0.273
0.931
t9 18:1
0.11
0.09
0.11
0.10
0.010
0.167
0.268
0.562
t10 18:1
0.09
0.10
0.13
0.16
0.016
0.006
0.252
0.475
t11 18:1
0.53
0.40
0.50
0.45
0.027
0.767
0.002
0.155
t12 18:1
0.53
0.55
0.62
0.61
0.024
0.004
0.808
0.667
c9 18:1 + t13 18:1 + t14 18:1
10.02
8.80
10.26
9.90
0.426
0.128
0.072
0.325
t15 18:1 + c11 18:1
0.76
0.78
0.85
0.92
0.034
0.002
0.175
0.510
c12 18:1
0.55
0.53
0.56
0.54
0.021
0.573
0.263
0.876
c13 18:1
0.10
0.10
0.13
0.12
0.007
<0.001
0.888
0.596
c14 18:1 + t16 18:1
0.15
0.15
0.19
0.19
0.007
<0.001
0.907
0.393
c15 18:1
0.08
0.11
0.13
0.13
0.007
<0.001
0.179
0.072
Sum 18:1
13.09
11.80
13.68
13.32
0.459
0.029
0.082
0.315
t11,c15 18:2
0.20
0.18
0.21
0.18
0.012
0.561
0.189
0.581
c9,t11 CLA
0.28
0.30
0.34
0.40
0.022
0.002
0.124
0.351
c9,t11,c15 CLnA
0.06
0.05
0.06
0.06
0.008
0.377
0.827
0.799
t10,c12,c15 CLnA
0.21
0.22
0.26
0.31
0.022
<0.001
0.186
0.386
18:2n-6
57.61
59.62
56.55
57.98
0.723
0.072
0.023
0.697
18:3n-3
10.12
9.83
9.36
8.43
0.286
<0.001
0.042
0.272
t11
1.07
0.94
1.11
1.09
0.044
0.032
0.095
0.202
t10
0.30
0.34
0.39
0.47
0.027
<0.001
0.035
0.486
t10:t11
0.28
0.36
0.36
0.43
0.022
0.002
0.002
0.747
1 Cows were divided into 2 groups based on the time below reticular pH 6.00. Tolerant cows (n = 4): ≤0.10 h/d; susceptible cows (n = 5): ≥1.26 h/d.
2 c = cis; t = trans; CLnA = conjugated linolenic acid; t10 and t11 = sum of all 18-carbon biohydrogenation intermediates containing a double bond in the trans configuration either at the 10th or the 11th carbon atom (from the carboxyl end); t10:t11 = ratio of trans-10 intermediates to trans-11 intermediates.
3 The results from 1 cow at 1 sampling time were omitted before statistical analysis (outlier).
Irrespective of group of cows, plasma proportions of trans-11 18:1 (period: P = 0.002) and 18:3n-3 (period: P = 0.042) were lower in the high-RFCH period, whereas proportions of 18:2n-6 (period: P = 0.023), total t10 intermediates (period: P = 0.035) and the ratio of t10 to t11 intermediates (period: P = 0.002) were higher during the high-RFCH period.
The effects of animal group and RFCH period on proportions of different 18-carbon FA in 4 blood plasma fractions (free FA, phospholipids, cholesterol esters, and triacylglycerols) are available as supplementary material (Supplemental Table S2; https://doi.org/10.3168/jds.2018-15823).
Milk
Proportions of different 18-carbon FA and total 18-carbon FA in milk fat are shown in Table 7. Total 18-carbon FA were higher in milk fat of susceptible cows as compared with tolerant cows (group: P < 0.001) and decreased in the high-RFCH period in both groups (period: P < 0.001). Both 18:2n-6 and 18:3n-3 proportions were lower in milk of susceptible cows (group: P < 0.05) and increased in the high-RFCH period (period: P < 0.001). The proportion of total t11 intermediates in milk was higher in tolerant cows at the beginning of the experiment, but decreased to the level observed in the susceptible cows during the high-RFCH period (group × period: P = 0.003). This interaction effect in total t11 intermediates was caused by both cis-9,trans-11 CLA (group × period: P = 0.003) and trans-11 18:1 (group × period: P = 0.009). The milk proportion of trans-11,cis-15 18:2 was also higher in tolerant cows (group: P = 0.014), but was not lower in the high-RFCH period. The total proportion of t10 intermediates and the ratio of t10 to t11 intermediates were higher in milk from susceptible cows (group: P < 0.001) and increased in the high-RFCH period in both groups (period: P < 0.001). This was mainly caused by higher proportions of trans-10 18:1 (group: P < 0.001; period: P < 0.001), as the milk proportion of trans-10,cis-12 CLA was lower in susceptible cows (group: P < 0.001). However, the latter milk FA only represented very minor proportions in milk fat (on average, 0.05 to 0.06 g/100 g of C18; minimum = 0.00 g/100 g of C18; maximum = 0.09 g/100 g of C18). Total 18:1 FA increased in the high-RFCH period only in susceptible cows (group × period: P = 0.037), whereas 18:0 decreased during the high-RFCH period in both groups of cows (period: P = 0.003), although this decrease tended (group × period: P = 0.059) to be larger for susceptible cows.
Table 7Proportions of different 18-carbon fatty acids (FA; g/100 g of C18) in milk collected from cows that were tolerant or susceptible
c = cis; t = trans; CLnA = conjugated linolenic acid; t10 and t11 = sum of all 18-carbon biohydrogenation intermediates containing a double bond in the trans configuration either at the 10th or the 11th carbon atom (from the carboxyl end); t10:t11 = ratio of trans-10 intermediates to trans-11 intermediates.
Means within a row with different superscripts differ (P < 0.05).
0.104
0.491
<0.001
0.003
t10
0.94
1.05
1.11
1.44
0.058
<0.001
<0.001
0.071
t10:t11
0.21
0.29
0.26
0.36
0.014
<0.001
<0.001
0.503
a–c Means within a row with different superscripts differ (P < 0.05).
1 Cows were divided into 2 groups based on the time below reticular pH 6.00. Tolerant cows (n = 4): ≤0.10 h/d; susceptible cows (n = 5): ≥1.26 h/d.
2 c = cis; t = trans; CLnA = conjugated linolenic acid; t10 and t11 = sum of all 18-carbon biohydrogenation intermediates containing a double bond in the trans configuration either at the 10th or the 11th carbon atom (from the carboxyl end); t10:t11 = ratio of trans-10 intermediates to trans-11 intermediates.
Proportions of different 18-carbon FA and total 18-carbon FA in saliva are presented in Supplemental Table S3 (https://doi.org/10.3168/jds.2018-15823). Total 18-carbon FA and proportions of different isomers (i.e., total t11 intermediates, cis-9,trans-11 CLA, trans-11 18:1, and trans-10,cis-12 CLA) decreased or tended to decrease in the high-RFCH period, irrespective of group (period: P < 0.10). The saliva proportion of total 18:1 FA tended to be higher in susceptible cows as compared with tolerant cows (group: P = 0.068).
Bacterial Community of Susceptible and Tolerant Cows Fed RFCH
No differences (P > 0.05) were observed in species richness (Figure 1A) or Shannon diversity (Figure 1B) between tolerant and susceptible cows or between the low- and high-RFCH period. In addition, the statistical test ANOSIM revealed no differences in overall bacterial community composition between susceptible and tolerant cows (group: P = 0.255) or between periods (period: P = 0.392).
Figure 1Alpha diversity of bacterial communities by the number of observed species (A) and by Shannon diversity (B) in buccal swab samples collected from cows that were tolerant or susceptible to increased amounts of rapidly fermentable carbohydrates (RFCH) in the diet during different periods (low RFCH = black; high RFCH = gray). Cows were divided into 2 groups based on the time below reticular pH 6.00. Tolerant cows (n = 4): ≤10 h/d; susceptible cows (n = 5): ≥1.26 h/d. Data are presented as mean ± SD. No significant differences were observed (group: P > 0.05; period: P > 0.05; group × period: P > 0.05).
Nevertheless, differences were observed in relative abundance of specific bacterial taxa between groups or periods or both (Table 8). The relative abundance of the genus Butyrivibrio decreased during the high-RFCH period, irrespective of group (period: P = 0.035), whereas the genus Pseudobutyrivibrio did not differ between groups or periods (P > 0.05). Relative abundances of the genera Dialister and Megasphaera were both higher in buccal swab samples from susceptible cows as compared with tolerant cows (group: P = 0.043), whereas Propionibacterium acnes was more abundant in tolerant cows (group: P = 0.007). Several other taxa, mainly within the phylum Firmicutes, were also more abundant in buccal swabs from susceptible cows compared with tolerant cows (group: P < 0.05; e.g., Staphylococcus epidermidis, uncultured Enterococcaceae, uncultured Lactobacillales, and uncultured Streptococcaceae). In contrast, uncultured RFP12 within the phylum Verrucomicrobia was more abundant in tolerant cows (group: P = 0.007). Decreases in relative abundance upon increased fermentability of the diet (period: P < 0.05) were observed for taxa within the phyla Bacteroidetes (e.g., CF231, uncultured Bacteroidales), Firmicutes (e.g., Ruminococcus, uncultured Erysipelotrichaceae), and Proteobacteria (e.g., Ruminobacter). The genus Carnobacterium was more abundant in buccal fluid from susceptible cows (group: P = 0.034) and increased in the high-RFCH period (period: P = 0.023), whereas opposite effects were observed for uncultured RF32 (group: P = 0.037; period: P = 0.023). The genus Anaeroplasma was more abundant in buccal fluid from tolerant cows in the low-RFCH period, but decreased to the level of susceptible cows in the high-RFCH period (group × period: P = 0.036).
Table 8Average relative abundance (%) of different bacterial taxa in buccal swab samples collected from cows that were tolerant or susceptible
Associations Between 18-Carbon FA and Bacterial Taxa
Several genera within the phyla Bacteroidetes (i.e., CF231, uncultured p-2534–18B5, uncultured Porphyromonadaceae) and Firmicutes (i.e., Oscillospira, uncultured Erysipelotrichaceae) correlated positively with milk fat content and proportions of total t11 intermediates in both milk and plasma (Table 9). Butyrivibrio and Pseudobutyrivibrio only showed positive correlations with milk fat content and milk t11 proportion, respectively. In contrast, Acidaminococcus, Dialister, Sharpea p-3329–23G2, and uncultured Betaproteobacteria showed positive correlations with total t10 intermediates in milk, plasma, or both, whereas uncultured Fibrobacter species additionally correlated negatively with milk fat content. Notably, Megasphaera did not correlate with any of the discussed milk or plasma FA proportions.
Table 9Spearman rank correlations between relative abundance of bacterial taxa in buccal swab samples and milk fat content and total proportions of trans-11 and trans-10 intermediates in milk and plasma
When high-starch diets are fed to dairy cows, it is known that typical pathways of rumen biohydrogenation are altered to produce unique FA intermediates (e.g., trans-10,cis-12 CLA), of which some might inhibit milk fat synthesis when reaching the mammary gland (
The relationship between rumen acidosis resistance and expression of genes involved in regulation of intracellular pH and butyrate metabolism of ruminal epithelial cells in steers.
Variability in susceptibility to acidosis among high producing mid-lactation dairy cows is associated with rumen pH, fermentation, feed intake, sorting activity, and milk fat percentage.
Susceptibility of dairy cows to subacute ruminal acidosis is reflected in milk fatty acid proportions, with C18:1 trans-10 as primary and C15:0 and C18:1 trans-11 as secondary indicators.
), we hypothesized to observe distinct reticular pH profiles within the limited number of cows that had been selected for the trial (n = 10, of which 1 died). As expected, inter-animal differences were observed, which allowed us to divide cows into 2 groups based on their reticular pH (i.e., tolerant and susceptible cows). Interestingly, the discrepancy in reticular pH between susceptible and tolerant cows was already observed during the low-RFCH period, whereas higher amounts of RFCH did not decrease the reticular pH in any of the 2 groups of cows. Tolerant cows even showed higher reticular pH values during the high-RFCH period. Similar effects were observed in the study of
, when considering individual cow data: cows experiencing SARA at the end of the SARA induction experiment already showed longer periods of rumen pH below 5.60 during the preinduction period. On the other hand, the SARA-induction protocol failed to induce SARA (defined as pH <5.60 for more than 283 min/d) in cows that had less than 50 min/d below 5.60 during the preinduction period (
), we hypothesized that the inter-animal variation in reticular pH would be reflected in the occurrence or not of MFD. Indeed, in line with a lower reticular pH, the susceptible cows also showed a lower milk fat content in comparison with the tolerant cows (35.44 vs. 38.37 mg/g, −7.64%). This lower milk fat content was not a dilution effect, as total milk yield was also lower in susceptible cows (29.95 vs. 34.12 kg/d, −12.22%), which might be related to the lower DMI of these cows (21.27 vs. 23.19 kg/d, −8.28%). This lower DMI of susceptible cows might be the result of reduced fiber digestibility occurring with lower rumen pH values (
), which increases the ruminal retention time and, thus, limits DMI. However, fiber digestibility was not measured in our study. In contrast to the lack of response of increasing amounts of dietary RFCH on reticular pH, MFD (i.e., decreased milk fat content without changes in milk yield) was observed when shifting from the low- to the high-RFCH diet in both groups of cows (35.07 vs. 38.75 mg/g, −9.50%). This 9.50% reduction in milk fat was lower than milk fat reductions observed in other reports [e.g., −34%, from 40.3 to 28.3 mg/g in
Induction of and recovery from milk fat depression occurs progressively in dairy cows switched between diets that differ in fiber and oil concentration.
]. However, in the current study, the relative amount of RFCH was gradually increased over a period of 7 d, whereas cows were abruptly switched to a diet high in starch in the experiments of
Induction of and recovery from milk fat depression occurs progressively in dairy cows switched between diets that differ in fiber and oil concentration.
. In addition, these cows were also supplemented with plant oils (i.e., soybean oil or sunflower oil), which was not the case in the current experiment. Hence, the response of the milk fat content to the increased amounts of dietary RFCH without a concomitant decrease in reticular pH suggests that a low rumen pH is not the main determinant of MFD.
Previous studies suggested that diet-induced MFD is caused by alterations in rumen biohydrogenation pathways of dietary PUFA, resulting in the formation of specific FA isomers, of which some might inhibit milk fat synthesis (
); therefore, we focused on proportions of 18-carbon FA in the current study. Two distinct pathways of biohydrogenation have been described for 18:2n-6 and 18:3n-3 (
). Under normal rumen conditions, 18:2n-6 is mainly isomerized to cis-9,trans-11 CLA, which is further hydrogenated to trans-11 18:1 and, ultimately, to 18:0 (
). Under altered ruminal conditions (e.g., supply of marine products, a low rumen pH, high amounts of starch in the diet), a shift occurs toward the formation of t10 intermediates (i.e., trans-10,cis-12,cis-15 CLnA, trans-10,cis-15 18:2, trans-10,cis-12 CLA, and trans-10 18:1) at the expense of t11 intermediates (
). As the milk fat content was generally lower in susceptible cows compared with tolerant cows and decreased in both groups of cows during the high-RFCH period, differences in rumen proportions of t11 and t10 intermediates were expected (1) between susceptible and tolerant cows, irrespective of period, and (2) between the low- and high-RFCH period, irrespective of animal group.
As rumen samples were not available in the current trial, alternative samples were collected to assess possible shifts in the rumen FA metabolism. Fatty acid profiles of blood and milk samples might be acceptable proxies to assess a shift in the rumen biohydrogenation pathways, despite postabsorptive conversions that can take place. Indeed, in the mammary gland, trans-11 18:1 might be converted to cis-9,trans-11 CLA through the action of Δ9-desaturase (
Eicosapentaenoic acid and 3,10 dithia stearic acid inhibit the desaturation of trans-vaccenic acid into cis-9,trans-11-conjugated linoleic acid through different pathways in Caco-2 and T84 cells.
), which affects the FA distribution of blood plasma. To bypass these issues, we focused on the sum of t11 and t10 intermediates in the current discussion, rather than specific t11 and t10 isomers. As expected, we found lower milk fat concentrations associated with a t11 to t10 shift both in milk as well as in blood plasma, which might be related to alterations in rumen biohydrogenation pathways. Total t10 intermediates and the ratio of t10 to t11 intermediates were higher in plasma and milk (1) from susceptible cows, irrespective of RFCH period, and (2) during the high-RFCH period, irrespective of animal group. On the other hand, total t11 intermediates were lower in milk (1) from susceptible cows and (2) during the high-RFCH period.
As ruminants regularly regurgitate large amounts of rumen material to their oral cavities, oral samples might also be an alternative to rumen sampling for the analysis of the rumen FA composition. To our knowledge, this is the first time that saliva samples were collected for this purpose. Unfortunately, proportions of particular trans FA (e.g., total t10 and t11 intermediates) in saliva did not reflect the differences that were expected based on milk and plasma FA proportions. Furthermore, the variation in saliva samples was higher as compared with plasma and milk samples. This suggests that saliva, collected using the current sampling procedure, could not represent an alternative to milk or blood and, ultimately, to rumen samples. To achieve representative saliva samples, rumen fluid should be present in the sample, which can be identified by its green-brown color. With the current sampling procedure, most cows were nervous at the time of sampling and were, as a consequence, not ruminating, which might be the main reason of limited coherence between FA profiles of saliva versus blood and milk FA profiles.
As bacteria play an important role in rumen biohydrogenation of PUFA (
), differences in bacterial composition might be related to the shift in 18-carbon FA metabolism from its main biohydrogenation pathway to the alternative pathway. In the current study, buccal swab samples were used as a noninvasive method to determine the rumen bacterial community (
, we observed no differences in species richness and diversity of bacterial communities and no difference in overall bacterial community composition when gradually switching from the low- to high-RFCH period. However, at the species level, some differences in relative abundance were observed between susceptible and tolerant cows or upon increased amounts of dietary RFCH. This discussion will focus on changes in bacterial taxa that could potentially be involved in ruminal biohydrogenation of 18-carbon FA. Decreased abundance of the genus Butyrivibrio upon increasing the fermentability of the diet aligns with former observations of
, in which decreases of Butyrivibrio fibrisolvens populations were observed when animals were adapted to a high-concentrate diet. This might indicate a potential role of Butyrivibrio spp. in ruminal t11 formation, as suggested by other reports (
Dietary fish oil supplements modify ruminal biohydrogenation, alter the flow of fatty acids at the omasum, and induce changes in the ruminal Butyrivibrio population in lactating cows.
). However, as no difference was observed between susceptible and tolerant cows, no correlation between Butyrivibrio spp. and t11 intermediates was observed in our study, concurring with other reports (
As yet uncultured bacteria phylogenetically classified as Prevotella, Lachnospiraceae incertae sedis and unclassified Bacteroidales, Clostridiales and Ruminococcaceae may play a predominant role in ruminal biohydrogenation.
), correlated positively with t11 intermediates in the current trial. An alternative explanation is that Butyrivibrio spp. are the main bacteria involved in ruminal t11 production, and low rumen pH, as encountered by susceptible cows, could decrease their activity (i.e., t11 production;
), which is not reflected in relative abundance data.
The ruminal bacteria responsible for the formation of t10 intermediates are not well known. Increased concentrations in blood and milk of t10 intermediates in the susceptible cows in comparison with tolerant ones coincided with a higher relative abundance of Megasphaera spp. Others also observed such concomitant increases of these FA isomers and the relative abundance of Megasphaera spp. in rumen content (
Nutritional models of experimentally-induced subacute ruminal acidosis (SARA) differ in their impact on rumen and hindgut bacterial communities in dairy cows.
, in which higher amounts of Megasphaera (elsdenii) were observed in the rumen of cows upon a high-grain diet, the abundance of this genus did not increase when shifting from the low- to high-RFCH period in the current study. Furthermore, Megasphaera did not linearly correlate with t10 intermediates in milk or blood in the current study. Together with contrasting results about the capability of Megasphaera spp. to produce trans-10,cis-12 CLA in vitro (
Dietary fish oil supplements modify ruminal biohydrogenation, alter the flow of fatty acids at the omasum, and induce changes in the ruminal Butyrivibrio population in lactating cows.
). Surprisingly, in the current study, P. acnes abundance was comparable with the abundance of Butyrivibrio spp., and its abundance was higher in tolerant cows compared with susceptible cows. This might indicate that P. acnes is also present in the oral cavity of cows, as observed in humans (
). As such, buccal swab samples might not be a good alternative to assess the rumen abundance of this particular species. In a former experiment in our laboratory (
), Dialister spp. and Sharpea spp. were more abundant in the rumen of goats in situations with greater t10 accumulation, which was confirmed in the current study; nevertheless, contrasting results were observed for Lactobacillus spp. and Bifidobacterium spp. (
). According to the current study, Carnobacterium spp., Acidaminococcus spp., and uncultured genera belonging to the Betaproteobacteria were more abundant in situations with greater t10 accumulation, potentially indicating a role of these bacteria in ruminal t10 formation. However, in vitro experiments with pure cultures are required to confirm the capacity of these bacteria to produce t10 intermediates.
CONCLUSIONS
Inter-animal variation was observed in our study in terms of reticular pH, which allowed us to divide cows into 2 groups (i.e., tolerant cows showing higher pH values and susceptible cows with lower pH values). In line with this, susceptible cows also showed lower milk fat concentrations in comparison with tolerant cows. This discrepancy in reticular pH between cows was observed throughout the experiment, irrespective of the dietary RFCH level. Even more, gradually switching from the low- to high-RFCH period did not decrease the reticular pH in either of the 2 groups. Nevertheless, MFD was observed in both groups of cows upon increasing RFCH amounts in the diet, which indicates that a low rumen pH is not the main determinant of MFD. Lower milk fat concentrations were associated with a shift in biohydrogenation pathways toward the formation of t10 intermediates at the expense of t11 intermediates. Moreover, lower milk fat concentrations were also accompanied by significant shifts in the relative abundance of specific bacterial taxa, which were correlated with either t10 or t11 biohydrogenation intermediates. Dialister spp., Sharpea spp., Carnobacterium spp., Acidaminococcus spp., and uncultured genera belonging to the Betaproteobacteria were more abundant in situations with greater t10 accumulation, suggesting their potential role in altered biohydrogenation pathways.
ACKNOWLEDGMENTS
The doctoral research of Lore Dewanckele was financed by the Special Research Fund of Ghent University (Bijzonder Onderzoeksfonds, BOF, Belgium; BOF15/DOC/246 to LD); the PhD research of Longhui Jing was cofounded by the Chinese Scholarship Council (CSC, Beijing, China) and by the Special Research Fund of Ghent University (BOF, Gent, Belgium). The authors gratefully acknowledge Janine Koppes and the staff from Schothorst Feed Research B.V. (Lelystad, the Netherlands) for their help during the experiment. We also thank Charlotte Melis (Laboratory for Animal Nutrition and Animal Product Quality, Department of Animal Sciences and Aquatic Ecology, Ghent University, Gent, Belgium) for performing the fatty acid extractions.
As yet uncultured bacteria phylogenetically classified as Prevotella, Lachnospiraceae incertae sedis and unclassified Bacteroidales, Clostridiales and Ruminococcaceae may play a predominant role in ruminal biohydrogenation.
Susceptibility of dairy cows to subacute ruminal acidosis is reflected in milk fatty acid proportions, with C18:1 trans-10 as primary and C15:0 and C18:1 trans-11 as secondary indicators.
Combining results of two GC separations partly achieves determination of all cis and trans 16:1, 18:1, 18:2 and 18:3 except CLA isomers of milk fat as demonstrated using Ag-ion SPE fractionation.
Variability in susceptibility to acidosis among high producing mid-lactation dairy cows is associated with rumen pH, fermentation, feed intake, sorting activity, and milk fat percentage.
Nutritional models of experimentally-induced subacute ruminal acidosis (SARA) differ in their impact on rumen and hindgut bacterial communities in dairy cows.
Eicosapentaenoic acid and 3,10 dithia stearic acid inhibit the desaturation of trans-vaccenic acid into cis-9,trans-11-conjugated linoleic acid through different pathways in Caco-2 and T84 cells.
Induction of and recovery from milk fat depression occurs progressively in dairy cows switched between diets that differ in fiber and oil concentration.
The relationship between rumen acidosis resistance and expression of genes involved in regulation of intracellular pH and butyrate metabolism of ruminal epithelial cells in steers.
Dietary fish oil supplements modify ruminal biohydrogenation, alter the flow of fatty acids at the omasum, and induce changes in the ruminal Butyrivibrio population in lactating cows.
Evaluation of sequential methods for the determination of butterfat fatty acid composition with emphasis on trans-18:1 acids. Application to the study of seasonal variations in French butters.
Trans-11 to trans-10 shift of ruminal biohydrogenation of fatty acids is linked to changes in rumen microbiota.
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Clermont-Ferrand, France2016: 150 (Rowett Institute of Nutrition and Health, Aberdeen, Scotland, UK)
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