Systemic inflammation in early lactation and its relation to the cows' oxidative and metabolic status, productive and reproductive performance as well as activity.

A dysregulated inflammatory response contributes to the occurrence of disorders in cows during the transition period from pregnancy to lactation. However, a detailed characterization of clinically healthy cows that exhibit enhanced inflammatory response during this critical period remains incomplete. In this experiment, a total of 99 individual transition dairy cows and 109 observations (18 cows monitored in 2 consecutive lactations), submitted to similar transition management were involved to evaluate the relationship between elevated inflammatory response and metabolic, oxidative status as well as transition outcomes. Blood was taken at -7, 3, 6, 9 and 21 d in milk (DIM) and concentrations of metabolic parameters (glucose, ß-hydroxybutyric acid (BHBA), nonesterified fatty acids (NEFA), insulin, insulin-like growth factor 1 (IGF-1) and fructosamine) were analyzed. Additionally, oxidative parameters (proportion of oxidized glutathione to total glutathione in red blood cells (GSSG (%)), the activity of glutathione peroxidase (GPx) and of superoxide dismutase (SOD), concentrations of malondialdehyde (MDA) and oxygen radical absorbance capacity (ORAC)) and acute phase proteins (APP) including haptoglobin (Hp), serum amyloid A (SAA) and albumin-to-globulin ratio (A:G) were determined in the blood of 21 DIM. The 3 APP parameters were used to group clinically healthy cows into 2 categories through k-medoids clustering, i.e., a group showing an acute phase response (APR, n = 39) and a group not showing such a response, i.e., non-APR (n = 50). Diseased cases (n = 20) were handled in a separate group. Lower SAA and Hp concentrations as well as higher A:G were observed in the non-APR group, although for Hp differences were observed from the APR group, not from the diseased group. Only one of the 5 oxidative parameters differed between the groups, with the non-APR group exhibiting lower GPx activity compared with the diseased group. The non-APR group showed the highest IGF-1 levels among the 3 groups, and lower NEFA concentrations compared with the diseased groups. The diseased group also showed reduced dry matter intake and milk yield compared with clinically healthy cows, regardless of their inflammatory status. Moreover, the APR group exhibited temporarily lower activity levels compared with the non-APR group. These findings highlight that cows with a lower inflammatory status after 21 DIM exhibited better metabolic health characteristics, productive performance as well as activity levels. Nevertheless, the detrimental effects of a higher inflammatory status in the absence of clinical symptoms are still relatively limited.

Effect of supplementing an α-amylase enzyme or a blend of essential oil components on the performance, nutrient digestibility and nitrogen balance of dairy cows.

Lowering the dietary protein content is a promising strategy to reduce N excretions in cattle but requires an improved N utilization by the animal. Feed enzymes (e.g., exogenous α-amylase) and plant extracts (e.g., essential oils (EO)) are 2 additives which may enhance rumen function and possibly also microbial protein yield. This may increase fat and protein corrected milk yield (MY) and milk nitrogen efficiency (MNE) and thus lower N losses from dairy cows. Both types of additives were studied in an experiment including 39 Holstein cows (average ± SD: 40.7 ± 7.95 kg/d MY, 89 ± 43 d in milk (DIM), 2.7 ± 1.5 lactations, 677 ± 68.6 kg of BW) consisting of a covariate (4 weeks) and treatment period (5 weeks). During the whole experiment cows were fed a typical Benelux diet (CTRL), supplemented with concentrates to meet individual requirements for energy and metabolizable protein, which were fulfilled for 100% and 101%, respectively. The total diet was low in crude protein (15.5%) and relatively high in starch (22.6% and 6.6% rumen bypass starch). Cows were balanced for parity, DIM, MY and roughage intake and randomly assigned to one of 3 groups, receiving the following treatments in the treatment period; (1) CTRL (n = 13); (2) CTRL + 14 g/cow/d Ronozyme RumiStar (α-amylase enzyme, DSM) (AMEZ, n = 13); (3) CTRL + 2.5 g/cow/d Crina Protect (blend of EO components, DSM) (ESOL, n = 13). Animal performance, ruminal pH and enteric gas emissions were monitored throughout the experiment. During the last week of the covariate and treatment period, nitrogen balances were conducted, total-tract nutrient digestibility was determined and urinary allantoin and uric acid were determined as indicators for microbial N production. The statistical model applied to these variables contained group and DIM during treatment period as fixed effects and the values from the covariate period as covariate. Post-hoc Dunnet corrected comparisons between each treatment group and the control group were explored. The α-amylase enzyme tended to increase apparent total-tract starch digestibility and increased milk lactose concentration. The EO blend tended to increase milk yield and increased milk N output, MNE and feed efficiency. Therefore, when feeding reduced dietary protein levels, EO have potential to improve the N-use efficiency in cattle, whereas the α-amylase enzyme might increase starch digestibility and milk lactose. However, additional research is necessary to substantiate our findings.

Combination of milk variables and on-farm data as an improved diagnostic tool for metabolic status evaluation in dairy cattle during the transition period.

Milk composition, particularly milk fatty acids, has been extensively studied as an indicator of the metabolic status of dairy cows during early lactation. In addition to milk biomarkers, on-farm sensor data also hold potential in providing insights into the metabolic health status of cows. While numerous studies have explored the collection of a wide range of sensor data from cows, the combination of milk biomarkers and on-farm sensor data remains relatively underexplored. Therefore, this study aims to identify associations between metabolic blood variables, milk variables, and various on-farm sensor data. Second, it seeks to examine the supplementary or substitutive potential of these data sources. Therefore, data from 85 lactations on metabolic status and on-farm data were collected during 3 wk before calving up to 5 wk after calving. Blood samples were taken on d 3, 6, 9, and 21 after calving for determination of ß-hydroxybutyrate (BHB), nonesterified fatty acids (NEFA), glucose, insulin-like growth factor-1 (IGF-1), insulin, and fructosamine. Milk samples were taken during the first 3 wk in lactation and analyzed by mid-infrared for fat, protein, lactose, urea, milk fatty acids, and BHB. Walking activity, feed intake, and body condition score (BCS) were monitored throughout the study. Linear mixed effect models were used to study the association between blood variables and (1) milk variables (i.e., milk models); (2) on-farm data (i.e., on-farm models) consisting of activity and dry matter intake analyzed during the dry period ([D]) and lactation ([L]) and BCS only analyzed during the dry period ([D]); and (3) the combination of both. In addition, to assess whether milk variables can clarify unexplained variation from the on-farm model and vice versa, Pearson marginal residuals from the milk and on-farm models were extracted and related to the on-farm and milk variables, respectively. The milk models had higher coefficient of determination (R2) than the on-farm models, except for IGF-1 and fructosamine. The highest marginal R2 values were found for BHB, glucose, and NEFA (0.508, 0.427, and 0.303 vs. 0.468, 0.358, and 0.225 for the milk models and on-farm models, respectively). Combining milk and on-farm data particularly increased R2 values of models assessing blood BHB, glucose, and NEFA concentrations with the fixed effects of the milk and on-farm variables mutually having marginal R2 values of 0.608, 0.566, and 0.327, respectively. Milk C18:1 was confirmed as an important milk variable in all models, but particularly for blood NEFA prediction. On-farm data were considerably more capable of describing the IGF-1 concentration than milk data (marginal R2 of 0.192 vs. 0.086), mainly due to dry matter intake before calving. The BCS [D] was the most important on-farm variable in relation to blood BHB and NEFA and could explain additional variation in blood BHB concentration compared with models solely based on milk variables. This study has shown that on-farm data combined with milk data can provide additional information concerning the metabolic health status of dairy cows. On-farm data are of interest to be further studied in predictive modeling, particularly because early warning predictions using milk data are highly challenging or even missing.

Milk yield residuals and their link with the metabolic status of dairy cows in the transition period.

The transition period is one of the most challenging periods in the lactation cycle of high-yielding dairy cows. It is commonly known to be associated with diminished animal welfare and economic performance of dairy farms. The development of data-driven health monitoring tools based on on-farm available milk yield development has shown potential in identifying health-perturbing events. As proof of principle, we explored the association of these milk yield residuals with the metabolic status of cows during the transition period. Over 2 yr, 117 transition periods from 99 multiparous Holstein-Friesian cows were monitored intensively. Pre- and postpartum dry matter intake was measured and blood samples were taken at regular intervals to determine ß-hydroxybutyrate, nonesterified fatty acids (NEFA), insulin, glucose, fructosamine, and IGF1 concentrations. The expected milk yield in the current transition period was predicted with 2 previously developed models (nextMILK and SLMYP) using low-frequency test-day (TD) data and high-frequency milk meter (MM) data from the animal's previous lactation, respectively. The expected milk yield was subtracted from the actual production to calculate the milk yield residuals in the transition period (MRT) for both TD and MM data, yielding MRTTD and MRTMM. When the MRT is negative, the realized milk yield is lower than the predicted milk yield, in contrast, when positive, the realized milk yield exceeded the predicted milk yield. First, blood plasma analytes, dry matter intake, and MRT were compared between clinically diseased and nonclinically diseased transitions. MRTTD and MRTMM, postpartum dry matter intake and IGF1 were significantly lower for clinically diseased versus nonclinically diseased transitions, whereas ß-hydroxybutyrate and NEFA concentrations were significantly higher. Next, linear models were used to link the MRTTD and MRTMM of the nonclinically diseased cows with the dry matter intake measurements and blood plasma analytes. After variable selection, a final model was constructed for MRTTD and MRTMM, resulting in an adjusted R2 of 0.47 and 0.73, respectively. While both final models were not identical the retained variables were similar and yielded comparable importance and direction. In summary, the most informative variables in these linear models were the dry matter intake postpartum and the lactation number. Moreover, in both models, lower and thus also more negative MRT were linked with lower dry matter intake and increasing lactation number. In the case of an increasing dry matter intake, MRTTD was positively associated with NEFA concentrations. Furthermore, IGF1, glucose, and insulin explained a significant part of the MRT. Results of the present study suggest that milk yield residuals at the start of a new lactation are indicative of the health and metabolic status of transitioning dairy cows in support of the development of a health monitoring tool. Future field studies including a higher number of cows from multiple herds are needed to validate these findings.

The effect of maternal supply of rumen-protected protein to Holstein Friesian cows during the dry period on the transfer of passive immunity and colostral microbial composition.

The objective of this study was to analyze if maternal supply of rumen-protected protein during the dry period can affect the IgG concentration and microbial composition of colostrum and the IgG absorption and fecal microbial composition in the calf. Seventy-four multiparous Holstein Friesian (HF) dairy cows were stratified per parity and randomly assigned to one of 2 different dry period diets, a diet with a low crude protein (CP) level (LP) and a diet with a high CP level (HP) by addition of rumen-undegraded protein (RUP; formaldehyde-treated soybean meal, Mervobest, Nuscience, Drongen, Belgium). Colostrum was collected within 1 h after calving and IgG concentration was quantified by radial immunodiffusion analysis. Forty-nine calves (23 female and 26 male) were enrolled in the trial with a 2 × 2 factorial design, with prenatal and postnatal treatment as the 2 independent variables. This led to 4 experimental groups: LPLP, LPHP, HPLP, and HPHP, in which the first 2 letters refer to the prenatal treatment (diet of the dam) and the last 2 refer to the postnatal treatment (diet of the colostrum-producing cow). Calves received 3× 2 L of colostrum within 2, 6, and 24 h after birth. Meconium and feces were collected solely from female calves (n = 18) by digital palpation of the rectum, immediately after birth and before colostrum administration and at d 3 of age. Microbial DNA was extracted from meconium (n = 9), feces (n = 15), and colostrum (n = 49). Amplicon sequencing of the bacterial V3-V4 region of the 16S rRNA gene was performed for characterization of the bacterial communities. Colostrum IgG concentration was higher in cows that were supplemented with RUP, especially in cows entering their second lactation (LSM ± SEM 61.3 ± 2.3 vs. 55.2 ± 2.8 g of IgG/L). Calves born out of LP cows that received colostrum from HP cows (LPHP) had a lower serum IgG level compared with HPHP and LPLP calves (LSM ± SEM 14.2 ± 1.3 vs. 18.8 ± 1.2 and 20.9 ± 1.3 g of IgG/L in HPHP and LPLP, respectively). The most abundant phyla in colostrum were Proteobacteria (48.2%), Firmicutes (24.8%), Bacteroidetes (9.5%), and Actinobacteria (5.0%). The most abundant phyla in calf meconium and feces were Firmicutes (42.5 and 47.5%), Proteobacteria (21.7% and 33.7%), Bacteroidetes (16.8% and 15.7%), and Actinobacteria (2.9% and 3.1%). There was no difference in the overall microbial communities between colostrum from HP and LP cows. However, 2 genera (both members of the family Lachnospiraceae) were more abundant in colostrum from HP cows compared with LP cows. The microbial composition of meconium, feces and colostrum differed from each other. Fecal samples were more similar to each other and are characterized by a lower intersample diversity compared with colostrum and meconium samples. To conclude, increasing the CP level by addition of RUP in the dry period diet affected the colostrum IgG concentration and the transfer of passive immunity, but did not change the overall microbial composition of colostrum nor of meconium and feces in the calf.

Transition cow clusters with distinctive antioxidant ability and their relation to performance and metabolic status in early lactation.

Metabolic and oxidative stress have been characterized as risk factors during the transition period from pregnancy to lactation. Although mutual relations between both types of stress have been suggested, they rarely have been studied concomitantly. For this, a total of 99 individual transition dairy cows (117 cases, 18 cows sampled during 2 consecutive lactations) were included in this experiment. Blood samples were taken at -7, 3, 6, 9, and 21 d relative to calving and concentrations of metabolic parameters (glucose, ß-hydroxybutyric acid (BHBA), nonesterified fatty acids, insulin, insulin-like growth factor 1, and fructosamine) were determined. In the blood samples of d 21, biochemical profiles related to liver function and parameters related to oxidative status were determined. First, cases were allocated to 2 different BHBA groups (ketotic vs. nonketotic, N:n = 20:33) consisting of animals with an average postpartum BHBA concentration and at least 2 out of 4 postpartum sampling points exceeding 1.2 mmol/L or remaining below 0.8 mmol/L, respectively. Second, oxidative parameters [proportion of oxidized glutathione to total glutathione in red blood cells (%)], activity of glutathione peroxidase, and of superoxide dismutase, concentrations of malondialdehyde and oxygen radical absorbance capacity were used to perform a fuzzy C-means clustering. From this, 2 groups were obtained [i.e., lower antioxidant ability (LAA80%, n = 31) and higher antioxidant ability (HAA80%, n = 19)], with 80% referring to the cutoff value for cluster membership. Increased concentrations of malondialdehyde, decreased superoxide dismutase activity, and impaired oxygen radical absorbance capacity were observed in the ketotic group compared with the nonketotic group, and inversely, the LAA80% group showed increased concentrations of BHBA. In addition, the concentration of aspartate transaminase was higher in the LAA80% group compared with the HAA80% group. Both the ketotic and LAA80% groups showed lower dry matter intake. However, a lower milk yield was observed in the LAA80% group but not in the ketotic group. Only 1 out of 19 (5.3%) and 3 out of 31 (9.7%) cases from the HAA80% and LAA80% clusters belong to the ketotic and nonketotic group, respectively. These findings suggested that dairy cows vary in oxidative status at the beginning of the lactation, and fuzzy C-means clustering allows to classify observations with distinctive oxidative status. Dairy cows with higher antioxidant capacity in early lactation rarely develop ketosis.

Prediction of metabolic status of dairy cows in early lactation using milk fatty acids and test-day variables.

Early lactation metabolic imbalance is an important physiological change affecting the health, production, and reproduction of dairy cows. The aims of this study were (1) to evaluate the potential of test-day (TD) variables with or without milk fatty acids (FA) content to classify metabolically imbalanced cows and (2) to evaluate the robustness of the metabolic classification with external data. A data set was compiled from 3 experiments containing plasma ß-hydroxybutyrate, nonesterified FA, glucose, insulin-like growth factor-I, FA proportions in milk fat, and TD variables collected from 244 lactations in wk 2 after calving. Based on the plasma metabolites, 3 metabolic clusters were identified using fuzzy c-means clustering and the probabilistic membership value of each cow to the 3 clusters was determined. Comparing the mean concentration of the plasma metabolites, the clusters were differentiated into metabolically imbalanced, moderately impacted, and balanced. Following this, the 2 metabolic status groups identified were imbalanced cows (n = 42), which were separated from what we refer to as "others" (n = 202) based on the membership value of each cow for the imbalanced cluster using a threshold of 0.5. The following 2 FA data sets were composed: (1) FA (groups) having high prediction accuracy by Fourier-transform infrared spectroscopy and, thus, have practical significance, and (2) FA (groups) formerly identified as associated with metabolic changes in early lactation. Metabolic status prediction models were built using FA alone or combined with TD variables as predictors of metabolic groups. Comparison was made among models and external evaluations were performed using an independent data set of 115 lactations. The area under the receiver operating characteristics curve of the models was between 75 and 91%, indicating their moderate to high accuracy as a diagnostic test for metabolic imbalance. The addition of FA groups to the TD models enhanced the accuracy of the models. Models with FA and TD variables showed high sensitivities (80-88%). Specificities of these models (73-79%) were also moderate and acceptable. The accuracy of the FA models on the external data set was high (area under the receiver operating characteristics curve between 76 and 84). The persistently good performance of models with Fourier-transform infrared spectroscopy-quantifiable FA on the external data set showed their robustness and potential for routine screening of metabolically imbalanced cows in early lactation.

Improved cattle behaviour monitoring by combining Ultra-Wideband location and accelerometer data.

Cattle behaviour is fundamentally linked to the cows' health, (re)production, and welfare. The aim of this study was to present an efficient method to incorporate Ultra-Wideband (UWB) indoor location and accelerometer data for improved cattle behaviour monitoring systems. In total, 30 dairy cows were fitted with UWB Pozyx wearable tracking tags (Pozyx, Ghent, Belgium) on the upper (dorsal) side of the cow's neck. In addition to the location data, the Pozyx tag reports accelerometer data as well. The combination of both sensor data was performed in two steps. In the first step, the actual time spent in the different barn areas was calculated using location data. In the second step, accelerometer data were used to classify cow behaviour using the location information of step 1 (e.g., a cow located in the cubicles cannot be classified as feeding, or drinking). A total of 156 hours of video recordings were used for the validation. For each hour of data, the total time each cow spent in each area and performing which behaviours (feeding, drinking, ruminating, resting, and eating concentrates) were computed using the sensors and compared against annotated video recordings. Bland-Altman plots for the correlation and difference between the sensors and the video recording were then computed for the performance analysis. The overall performance of locating the animals into the correct functional areas was very high. The R2 was 0.99 (P < 0.001), and the root-mean-square error (RMSE) was 1.4 min (7.5% of the total time). The best performance was obtained for the feeding and lying areas (R2 = 0.99, P < 0.001). Performance was lower in the drinking area (R2 = 0.90, P < 0.01) and the concentrate feeder (R2 = 0.85, P < 0.05). For the combined location + accelerometer data, high overall performance (all behaviours) was obtained with an R2 of 0.99 (P < 0.001) and a RMSE of 1.6 min (12% of the total time). The combination of location and accelerometer data improved the RMSE of the feeding time and ruminating time compared to the accelerometer data alone (2.6-1.4 min). Moreover, the combination of location and accelerometer enabled accurate classification of additional behaviours that are difficult to detect using the accelerometer alone, such as eating concentrates and drinking (R2 = 0.85 and 0.90, respectively). This study demonstrates the potential of combining accelerometer and UWB location data for the design of a robust monitoring system for dairy cattle.

Effect of supplementing rumen-protected methionine, lysine, and histidine to low-protein diets on the performance and nitrogen balance of dairy cows.

Lowering the dietary protein content can reduce N excretions and NH3 emissions from manure and increase milk N efficiency of dairy cows. However, milk yield (MY) and composition can be compromised due to AA deficiency. Methionine and Lys are known as first limiting EAA for dairy cows, and recently His is also mentioned as limiting, especially in grass-based or low-protein diets. To examine this, a trial was conducted with a 3-wk pre-experimental adaptation period (diet 16.5% crude protein), followed by a depletion period of 4 wk, in which 39 cows (average ± standard deviation: 116 ± 29.3 d in milk, 1.8 ± 1.2 lactations, 638 ± 73.2 kg of body weight, and 32.7 ± 5.75 kg MY/d) received a low-protein diet (CTRL) (14.5% crude protein). Then, taking into account parity, His plasma concentration, and MY, cows were randomly assigned to 1 of 3 treatment groups during the rumen-protected (RP) AA period of 7 wk; (1) CTRL; (2) CTRL + RP-Met + RP-Lys (MetLys); (3) CTRL + RP-Met + RP-Lys + RP-His (MetLysHis). Products were dosed, assuming requirements for digestible (d) Met, dLys, and dHis being, respectively, 2.4%, 7.0%, and 2.4% of intestinal digestible protein. In the cross-back period of 5 wk, all cows received the CTRL diet. During the last week of each period, a N balance was conducted by collecting total urine and spot samples of feces. Total feces production was calculated using the inert marker TiO2. Statistical analysis was performed with a linear mixed model with cow as random effect and data of the last week of the pre-experimental period used as covariate for the animal performance variables. No effect of supplementing RP-Met and RP-Lys nor RP-Met, RP-Lys, and RP-His on feed intake, milk performance, or milk N efficiency was observed. However, the plasma AA profile indicated additional supply of dMet, dLys, and dHis. Nevertheless, evaluation of the AA uptake relative to the cow's requirements showed that most EAA (exclusive Arg and Thr) were limiting over the whole experiment. Only dHis was sufficiently supplemented during the RP-AA period due to an overestimation of the diet's dMet and dLys supply in the beginning of the trial. The numerically increased milk urea N and urinary N excretion when RP-Met, RP-Lys, and RP-His were added to the low-protein diet suggest an increased catabolism of the excess His.

Diagnostic milk biomarkers for predicting the metabolic health status of dairy cattle during early lactation.

Data on metabolic profiles of blood sampled at d 3, 6, 9, and 21 in lactation from 117 lactations (99 cows) were used for unsupervised k-means clustering. Blood metabolic parameters included ß-hydroxybutyrate (BHB), nonesterified fatty acids, glucose, insulin-like growth factor-1 (IGF-1) and insulin. Clustering relied on the average and range of the 5 blood parameters of all 4 sampling days. The clusters were labeled as imbalanced (n = 42) and balanced (n = 72) metabolic status based on the values of the blood parameters. Various random forest models were built to predict the metabolic cluster of cows during early lactation from the milk composition. All the models were evaluated using a leave-group-out cross-validation, meaning data from a single cow were always present in either train or test data to avoid any data leakage. Features were either milk fatty acids (MFA) determined by gas chromatography (MFA [GC]) or features that could be determined during a routine dairy herd improvement (DHI) analysis, such as concentration of fat, protein, lactose, fat/protein ratio, urea, and somatic cell count (determined and reported routinely in DHI registrations), either or not in combination with MFA and BHB determined by mid-infrared (MIR), denoted as MFA [MIR] and BHB [MIR], respectively, which are routinely analyzed but not routinely reported in DHI registrations yet. Models solely based on fat, protein, lactose, fat/protein ratio, urea and somatic cell count (i.e., DHI model) were characterized by the lowest predictive performance [area under the receiver operating characteristic curve (AUCROC) = 0.69]. The combination of the features of the DHI model with BHB [MIR] and MFA [MIR] powerfully increased the predictive performance (AUCROC = 0.81). The model based on the detailed MFA profile determined by GC analysis did not outperform (AUCROC = 0.81) the model using the DHI-features in combination with BHB [MIR] and MFA [MIR]. Predictions solely based on samples at d 3 were characterized by lower performance (AUCROC DHI + BHB [MIR] + MFA [MIR] model at d 3: 0.75; AUCROC MFA [GC] model at d 3: 0.73). High predictive performance was found using samples from d 9 and 21. To conclude, overall, the DHI + BHB [MIR] + MFA [MIR] model allowed to predict metabolic status during early lactation. Accordingly, these parameters show potential for routine prediction of metabolic status.

Different reticuloruminal pH metrics of high-yielding dairy cattle during the transition period in relation to metabolic health, activity, and feed intake.

The measurement of pH in the reticulorumen in combination with a time-pH threshold has been widely applied in research to diagnose subacute ruminal acidosis. However, other pH metrics also have biological value. In this study, 44 animals were monitored during the transition period using reticuloruminal pH boluses. Traditional and more complex pH characteristics were calculated to characterize the reticuloruminal pH profile: time pH <6, slope of a logistic cumulative pH curve (ß0), and deviations [squared error (SqEr)] from pH predictions based on a harmonic static model. In this study, we aimed to examine the associations between those pH metrics and metabolic health parameters, feed intake, and activity. Finally, to describe the reticuloruminal pH dynamically, we also constructed a dynamic linear model. The results of this model were studied in relation to feed intake. All pH parameters were mutually correlated (particularly ß0 and SqEr; mean Pearson correlation of -0.52). pH patterns, rather than time pH <6, were associated with metabolic health and feed intake: high variation in daily pH (ß0 parameter) was reflected in higher blood concentrations of nonesterified fatty acids. Moreover, pH deviations of the harmonic model were negatively associated with feed intake and rumination behavior. This research confirms the biological importance of pH metrics focusing on pH variation and pH deviations and provides deeper insight into its associations with metabolic health status, feed intake, and activity during early lactation.

Subacute ruminal acidosis phenotypes in periparturient dairy cows differ in ruminal and salivary bacteria and in the in vitro fermentative activity of their ruminal microbiota.

Both ruminal microbial structure and functionality might play a role in inter-individual variation in susceptibility for subacute rumen acidosis (SARA) observed in dairy cows. The aims of this study were to determine whether differences between cows with distinct SARA susceptibility were reflected in distinct (1) ruminal microbial communities, (2) salivary bacterial communities, and (3) fermentative capacity of ruminal microbiota assessed in vitro. To test this hypothesis, rumen samples were collected via an esophageal tube on 21 d postpartum from 38 multiparous Holstein cows, which were classified into 4 groups differing in median and mean time of reticular pH below 6 as well as area under the curve of pH below 6.0. During the 21 d postpartum, all cows within a group fulfilled following criteria: susceptible (S, n = 10; mean or median ≥180 min/d), moderately susceptible (MS, n = 7; 60 min/d < mean time of pH below 6 < 180 min/d, and median time of pH below 6 <180 min/d), moderately unsusceptible (MU, n = 11; 10 min/d < mean < 60 min/d, and median time of pH below 6 ≤30 min/d), or unsusceptible (U, n = 10; median = 0 min/d, and mean <10 min/d). Groups did not differ in total daily dry matter intake nor in total, roughage, or concentrate intake during daily 6-h time intervals. Rumen bacterial α-diversity did not differ among groups, but ß-diversity varied and bacterial 16S rRNA gene copy numbers were lower in S compared with U cows. The relative abundance of genera Streptococcus, Sharpea, Prevotellaceae_YAB2003, Succinivibrionaceae_UCG-001, Ruminococcus, and Ruminococcaceae_UCG-001 were higher in S compared with U cows. In contrast, Lachnospiraceae_ND3007 and Oscillospiraceae_V9D2013 were more abundant in U cows. Although pH-associated, inter-animal differences were also observed in the salivary bacteria, common differences in ruminal and salivary bacterial genera were limited. The functionality of the rumen microbiota was evaluated in vitro through exposure of the microbial inoculum of S and U cows to an anaerobic buffer at pH 5.8 and 6.8, in the presence of sterile supernatant of their own and of dry cows' rumen fluid (2 × 2 design). Generally, the S inoculum produced more volatile fatty acids, except at low pH with dry cows' supernatant, where volatile fatty acid production was completely impaired and lactate accumulation was highest. Compared with the microbes of U cows, microbes of S cows showed less fermentative activity in situations with 2 stress factors (low pH and an unfamiliar environment, i.e., rumen fluid supernatant of dry cows).

Invited review: MicroRNAs in bovine colostrum-Focus on their origin and potential health benefits for the calf.

Colostrum is the first milk produced by a cow after she gives birth. Compared with mature milk, it has a high concentration of immunoglobulin G. Calves are born without circulating antibodies, thus ingestion of colostrum is necessary to protect the calf against pathogens in the first challenging weeks of life. In addition to the life-saving supply of antibodies, colostrum contains minerals, vitamins, growth factors, and immune cells. Recently, microRNAs (miRNAs) were added to that list. MicroRNAs are short, non-coding RNA molecules that can regulate gene expression at the post-transcriptional level. They are thought to act as key regulators of diverse biological and developmental processes. Colostrum contains higher amounts of miRNAs than mature milk; immune- and development-related miRNAs are prominent. Their expression pattern in milk is likely to be influenced by maternal nutrition and environment. The fat content of the maternal diet appears to have a major effect on expression of miRNAs in milk and in the neonate. The immunological state of the mammary gland seems to affect miRNA expression as well. In cows diagnosed with subclinical mastitis, alterations in the expression of miRNAs in milk have been observed. It is believed that miRNAs in colostrum and milk are signaling molecules passed from mother to newborn. They are packaged in extracellular vesicles, which makes them resistant to the harsh conditions in the gastrointestinal tract. Therefore, they can reach the small intestine, where they are absorbed and transferred into the bloodstream. MicroRNAs are important for the development of the intestines. For example, miRNAs stimulate cell viability, proliferation, and stem cell activity of the intestinal epithelium. Furthermore, miRNAs seem to act as key players in the development of the complete immune system. They can, among other things, regulate B- and T-cell differentiation and affect interleukin production of macrophages. The abundance of miRNAs in colostrum and milk and the possibility for their absorption in the intestines of the neonate supports the hypothesis that these tiny molecules are important for the development of the newborn. The probable relation of diet to the expression of miRNAs by the mother creates a possible avenue to optimize expression of miRNAs and improve neonatal maturation.

Reducing enteric methane emissions from dairy cattle: Two ways to supplement 3-nitrooxypropanol.

The aim of this work was to determine the effect of 3-nitrooxypropanol (3-NOP) on the enteric methane (CH4) emissions and performance of lactating dairy cows when mixed in with roughage or incorporated into a concentrate pellet. After 2 pretreatment weeks without 3-NOP supplementation, 30 Holstein Friesian cows were divided into 3 homogeneous treatment groups: no additive, 3-NOP mixed in with the basal diet (roughage; NOPbas), and 3-NOP incorporated into a concentrate pellet (NOPconc). The pretreatment period was followed by a 10-wk treatment period in which the NOPbas and NOPconc cows were fed 1.6 g of 3-NOP/cow per day. After the treatment period, a 2-wk washout period followed without 3-NOP supplementation. The CH4 emissions were measured using a GreenFeed unit (C-Lock Inc., Rapid City, SD) installed in a freestall with cubicles during the entire experimental period. On average for the total treatment period and compared with the no-additive group, CH4 production (g/d) was 28 and 23% lower for NOPbas and NOPconc, respectively. Methane yield (g/kg of dry matter intake) and methane intensity (g/kg of milk) were 23 and 24% lower for NOPbas, respectively, and 21 and 22% lower for NOPconc, respectively. No differences were found between NOPbas and NOPconc. Moreover, supplying 3-NOP did not affect total dry matter intake, milk production, or milk composition. The results of this experiment show that 3-NOP can reduce enteric CH4 emissions of dairy cattle when incorporated into a concentrate pellet and that this reduction is not different from the effect of mixing in 3-NOP with the basal diet (roughage). This broadens the possibilities for using 3-NOP in the dairy sector worldwide, as it is not always feasible to provide an additive mixed in with the basal diet.

Supplementation of DHA-Gold pre and/or postnatally to goat kids modifies in vitro methane production and rumen morphology until 6 mo old.

This study aimed to investigate the effect of pre and/or postnatal supplementation of a dry whole cell algae (DHA-Gold) to goat kids, on in vitro methane (CH4) production, animal growth, and rumen morphology at the age of 6 mo. Furthermore, the in vitro retreatment effect of DHA-Gold was evaluated. Twenty pregnant Saanen goats giving birth to 2 male kids were used. Half of these does were supplemented (D+) with 18.2 g/d of DHA-Gold in the last 3 wk of pregnancy, whereas the other half was not (D-). After kidding, one goat kid per doe in both groups was supplemented daily with 0.28 g/kg of body weight of DHA-Gold (k+) until 12 wk, whereas the other goat kids were untreated (k-). This resulted in 4 experimental groups D+k+, D+k-, D-k+, and D-k-. In vitro incubations were performed at the ages of 4 wk, 11 wk, and 6 mo. At the age of 6 mo, goat kids were euthanized and additional incubations were performed supplementing 4 doses of DHA-Gold (0, 0.4, 0.8, and 1.6 mg/mL). Additionally, rumen tissue of the atrium ruminis, ventral rumen, and dorsal blind sac were collected to assess rumen morphology. Rumen inocula of 4-wk-old goat kids supplemented D+ showed lower (P < 0.05) in vitro CH4 production, however, this was mainly due to a reduction in the overall fermentation, while CH4 expressed relatively to total volatile fatty acids (VFA) was higher when goat kids were treated D+ or k+. The detrimental D+ effect on VFA production diminished at 11 wk old but remained a tendency (0.05 < P < 0.1). As for 4 wk D+ as well as k+ supplementation of DHA-Gold stimulated rather than inhibited in vitro CH4 production expressed relative to total VFA. Supplementation of DHA-Gold either D+ or k+ decreased density, width, and surface area of the ruminal papillae. However, no effect on animal growth was observed. Moreover, detrimental effects of D+ or k+ treatment on VFA production or stimulation of relative CH4 production were no longer observed at 6 mo old. Nevertheless, direct exposure of DHA-Gold to 6-mo-old inoculum linearly (P < 0.05) decreased CH4 and VFA production, which tended (P = 0.06) to be greater when using D-rumen inoculum. Accordingly, neither D+ nor k+ DHA-Gold supplementation showed potential for reduction of rumen methanogenesis. Furthermore, this early life intervention could represent some risk for impaired rumen papillae development, which, however, did not impair animal performance.

Polyunsaturated fatty acids are less effective to reduce methanogenesis in rumen inoculum from calves exposed to a similar treatment early in life.

The aim of this study was to evaluate the dose response on in vitro methane (CH) production of PUFA to which the inoculum donor animals had been exposed early in life. Sixteen Holstein calves (160 ± 3 and 365 ± 2 kg BW) at 6 and 12 mo of age were used as inoculum donors. Half of the calves were given increasing amounts of extruded linseed from birth (22 g/d) until 4 mo of age (578 g/d) first mixed with milk and then included in their concentrate. Linseed oil (LSO) was supplemented in vitro at 5 different doses (0, 0.6, 1.2, 2.4, and 4.8 mg/mL). Supplementation of LSO in the rumen inocula at both ages linearly decreased ( < 0.05) the in vitro CH production. Total in vitro VFA production was not affected by LSO supplementation. Inhibition of CH was smaller when using the rumen inoculum from calves that had received a similar treatment early in life ( < 0.05). Differences in response to in vitro supplementation of a type of fatty acids similar to those applied during early life suggest some "changes" in the functioning of the rumen microbial community.

Season of birth is associated with first-lactation milk yield in Holstein Friesian cattle.

The aim of the present research was to assess factors associated with first-lactation milk yield in dairy heifers, including maternal and environmental factors, factors related to the development of the heifer and factors related to its offspring such as gender of the calf. In addition, the potential underlying mechanism, in particular metabolic adaptations, was further explored. Data on body growth, reproduction and milk yield of 74 Holstein Friesian heifers on three herds in Flanders (Belgium) were collected. At birth, body measurements of the heifers were recorded and blood samples were taken (in order) to determine basal glucose and insulin concentrations. Body measurements were assessed every 3 months until first calving, and gender and weight of their first calf were recorded. Information on fertility and milk yield of the heifer and its dam were collected from the herd databases. Daily temperature and photoperiod were recorded from the database of the Belgian Royal Meteorological Institute. Linear mixed models were run with herd as a random factor, to account for differences in herd management. Heifers grew 867±80.7 g/day during their first year of life and were inseminated at 14.8±1.34 months. First calving took place at 24.5±1.93 months, at a weight of 642±61.5 kg and heifers produced 8506±1064 kg energy corrected milk during their first 305-day lactation. Regression models revealed that none of the maternal factors such as milk yield and parity, nor the growth of the heifer during the 1st year of life were associated with milk yield during first lactation. Age, and to a lesser extent BW at first parturition were positively associated with first-lactation milk yield. In addition, the season of birth, but not calving, had a significant influence on milk yield, with winter-born heifers producing less than heifers born in any other season. The lower yielding winter-born heifers had higher insulin concentrations at birth, whereas glucose concentrations were similar, the latter being suggestive for lower insulin sensitivity of the peripheral tissues. Furthermore, environmental temperature at the end of gestation was negatively correlated with neonatal insulin concentrations. In conclusion, results of the present study suggest heifers born during the hotter months are born with a higher peripheral insulin sensitivity, finally leading to a higher first-lactation milk yield.

Reduction of Mycobacterium avium ssp. paratuberculosis in colostrum: Development and validation of 2 methods, one based on curdling and one based on centrifugation.

The aim of this study was to develop and validate 2 protocols (for use on-farm and at a central location) for the reduction of Mycobacterium avium ssp. paratuberculosis (MAP) in colostrum while preserving beneficial immunoglobulins (IgG). The on-farm protocol was based on curdling of the colostrum, where the IgG remain in the whey and the MAP bacteria are trapped in the curd. First, the colostrum was diluted with water (2 volumes colostrum to 1 volume water) and 2% rennet was added. After incubation (1 h at 32°C), the curd was cut and incubated again, after which whey and curd were separated using a cheesecloth. The curd was removed and milk powder was added to the whey. Approximately 1 log reduction in MAP counts was achieved. A reduction in total proteins and IgG was observed due to initial dilution of the colostrum. After curd formation, more than 95% of the immunoglobulins remained in the whey fraction. The semi-industrial protocol was based on centrifugation, which causes MAP to precipitate, while the IgG remain in the supernatant. This protocol was first developed in the laboratory. The colostrum was diluted with skimmed colostrum (2 volumes colostrum to 1 volume skimmed colostrum), then skimmed and centrifuged (at 15,600 × g for 30 min at room temperature). We observed on average 1.5 log reduction in the MAP counts and a limited reduction in proteins and IgG in the supernatant. To obtain a semi-industrial protocol, dairy pilot appliances were evaluated and the following changes were applied to the protocol: after 2:1 dilution as above, the colostrum was skimmed and subsequently clarified, after which the cream was heat treated and added to the supernatant. To investigate the effect of the colostrum treatment on the nutritional value and palatability of the colostrum and the IgG transfer, an animal experiment was conducted with 24 calves. Six received the dam's colostrum, 6 were given untreated purchased colostrum (control), and 2 groups of 6 calves received colostrum treated according to both of the above-mentioned methods. No significant differences were found between the test groups and the dam's colostrum group in terms of animal health, IgG uptake in the blood serum, milk, or forage uptake. Two protocols to reduce MAP in colostrum (for use on-farm or at a central location) were developed. Both methods preserve the vital IgG.

Environmental and maternal factors associated with gross placental morphology in dairy cattle.

This article reports on a study of gross placental morphology of 282 expelled placentas from 89 primi- and 193 multiparous Holstein dams immediately after calving and examines associations with environmental factors such as typical herd features and season of calving, and maternal factors such as age at calving, level of milk yield at conception and cumulative amount of milk produced during gestation. The highest correlation between calf measurements and placental characteristics was found between the weight of the calf and the total cotyledonary surface (r = .643; p < .001), confirming the high importance of the cotyledonary surface available for nutrient transfer to the developing foetus. Younger age in adolescent and smaller heart girth in multiparous dams were associated with a higher cotyledon number, suggesting placental compensation in dams with lower capacities in terms of dry matter intake. No significant association between milk yield during gestation in multiparous animals and gross placental morphometrics could be detected, indicating that factors such as the amount of milk produced during gestation affect placental development less than foetal weight close to term. Therefore, placental growth may be sustained at the expense of other tissues in an attempt to maintain pregnancy and minimize the adverse consequences for the foetus. This study offers evidence concerning factors affecting the placental surface size for nutrient transfer from dam to calf in dairy cattle based on gross morphometrics, but needs confirmation from studies in which this surface size is more profoundly assessed by measuring the branches present in the cotyledonary villi.

The effect of maize silage type on the performances and methane emission of dairy cattle.

To examine whether type of maize silage is important for milk production performances, maize silage LG30224 (LG) was compared with Falkone (FA), the latter having a 4.0% points lower rumen NDF digestibility and 19 g/kg dry matter (DM) more starch. To bridge the lower energy content of FA, a third treatment was involved by adding maize meal (MM) in a ratio of 92/8 on DM (FA+MM). Maize and grass silage were fed ad libitum in a ratio of 65/35 on DM basis. Concentrates were supplemented individually to meet energy and protein requirements. The experiment was set up as a Latin square with three groups of nine Holstein cows during three periods of 3 weeks. In the last 2 weeks of each period, DM intake (DMI) and milk performances were measured. Each group included one cannulated cow to study effects on rumen fermentation. During the last 4 days of each period, two cows from each group were placed in gas exchange chambers to measure nutrient digestibility and methane production. Total DMI was higher (p < 0.05) for FA+MM (20.8 kg/day) than for FA (20.3 kg/day), while DMI for LG was intermediate (20.6 kg/day). Treatment did not affect milk production nor composition, whereas fat-protein-corrected milk was higher for LG (30.5 kg/day) and FA+MM (30.3 kg/day) than for FA (29.9 kg/day). The ration did not affect pH nor volatile fatty acid composition in the rumen. Further, total tract digestibility of OM, crude protein, NDF and starch did not differ among treatments. The ration with LG gave higher methane production per day and per kg NDF intake than both rations with FA, but the difference was not significant when expressed per kg DMI or FPCM. Thus, maize silage type is of little importance for milk production if energy and physical structure requirements are met.