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The objectives of the present study were (1) to evaluate the effect of prepartum cholecalciferol treatment on serum Ca concentration during the first 10 d after calving and (2) to evaluate the effect of treatment on subsequent health and performance. Multiparous Holstein cows (n = 377) from one dairy farm were fed a negative dietary cation-anion difference diet (−31 mEq/kg of DM) for the last 21 d of gestation. On d 275, the animals were randomly assigned to a control or a treatment group. Cows in the control group were left untreated, and cows in the treatment group received an injection of 12 × 106 IU of cholecalciferol intramuscularly on the day of enrollment. If treated cows did not deliver the calf within 6 d, they were reinjected with 10 × 106 IU of cholecalciferol. Blood samples were drawn on 1, 2, 3, 5, 7, and 10 days in milk (DIM) and analyzed for serum Ca, P, and Mg concentrations. In a subsample of cows (50 control cows, 35 cows treated once with cholecalciferol, and 15 cows treated twice) serum haptoglobin, nonesterified fatty acids, β-hydroxybutyrate, and 25-hydroxycholecalciferol concentrations were analyzed on 1, 5, and 10 DIM. Binary data [retained placenta (RP), metritis] were analyzed using logistic regression models. Repeated measures ANOVA with first-order autoregressive covariance was performed to evaluate the treatment effect on milk yield over the first 10 test days after parturition, 25-hydroxycholecalciferol, serum Ca, P, Mg, β-hydroxybutyrate, nonesterified fatty acids, and haptoglobin concentrations. Cox proportional hazards were used to model the time to event outcomes (time to pregnancy within 200 d, culling until 300 DIM). After enrollment of 31.4% of cows and a preliminary analysis, adverse reactions became apparent, and the study was stopped. Cows treated with cholecalciferol had a greater risk of incurring RP and metritis. The adjusted mean incidences were 2.0%, 7.7%, and 4.0% for RP, and 21.6%, 39.3%, and 33.3% for metritis for control cows, cows treated once, and cows treated twice with cholecalciferol, respectively. Compared with control cows, cows injected once with 12 × 106 IU of cholecalciferol produced less energy-corrected milk on the first (−3.76 kg) and second (−2.75 kg) test days, respectively. Cows injected twice with cholecalciferol (12 × 106 IU of cholecalciferol and 10 × 106 IU 1 wk later) had a reduced milk yield only at first test day (−3.80 kg). Treatment with cholecalciferol led to a significant increase in 25-hydroxycholecalciferol on d 1, 5, and 10 after calving. Serum Ca and P concentrations were significantly increased in cows treated with cholecalciferol, but serum Mg concentrations were significantly reduced. Haptoglobin concentrations were significantly increased on 5 DIM in cows injected once with 12 × 106 IU of cholecalciferol. Although we observed no effect of treatment on culling until 300 DIM, time to pregnancy was delayed by 34 d in cows injected once with 12 × 106 IU of cholecalciferol. In the present study, injection with 12 × 106 IU of cholecalciferol had detrimental effects on health and milk production despite the beneficial effects on Ca homeostasis.

The objective of this study was to investigate the effects of a milk byproduct–based starter pellet, high in lactose, on feed intakes, growth, and rumen development when calves were fed either a high or low level of milk. This was done by using a 2 × 2 factorial design, with 120 Holstein heifer calves randomly assigned to 1 of 2 milk treatments and 1 of 2 feed treatments upon enrollment at birth. Calves were either assigned to a low (LM) or high (HM) level of milk replacer by an automated milk-feeding system for individually-housed calves. The LM treatment calves were offered 6 L/d (150 g/L; 1.2 L, 5×/d), whereas HM treatment calves were offered 15 L/d (150 g/L; 3 L, 5×/d) of milk replacer. Calves were also assigned to a milk byproduct- (whey) based starter feed treatment (WF), or a grain-based starter ration (95% concentrate starter pellet, 5% chopped wheat straw) feed treatment (GF). All calves were offered their treatment feed starting on d 5, with water offered ad libitum from d 0. The WF treatment calves were offered 150 g/d of the milk byproduct–based pellet until they consumed the entirety for 2 out of 3 consecutive days. At this point, WF treatment calves received 150 g of the milk byproduct–based pellet top dressed onto the grain-based ration to allow for ad libitum feed consumption until weaned, when they only received the grain-based ration postweaning (d 64–77). Calves on the GF feed treatment received only the grain-based ration. All calves were gradually weaned from d 43 to 63. By design, calves on the HM treatment consumed greater levels of milk than LM treatment calves for the majority of the days in the preweaning phase (8.9 ± 0.11 vs. 5.5 ± 0.11 L/d; SE). Calves on the GF and WF treatments consumed similar amounts of milk in the preweaning phase (7.2 ± 0.11 vs. 7.2 ± 0.11 L/d); however, there was a milk and feed treatment interaction detected for milk consumption in the weaning phase, with HM-GF calves drinking more milk than the other treatments and HM-WF calves drinking more milk than both LM-GF and LM-WF calves. The WF treatment calves had greater solid feed intake during the preweaning phase than GF treatment calves, with no feed intake differences in the weaning or postweaning phases. The WF calves drank more water in the preweaning phase than GF calves. Average daily gain was similar throughout the study between the GF and WF treatment calves. Rumen physiological measures were all similar by the end of the trial (d 77) regardless of feed or milk treatment, suggesting similar rumen development postweaning. Overall, regardless of the level of milk fed, offering the milk byproduct–based starter pellet may increase preweaning feed intake, along with water consumption. However, both feed and milk treatments had similar performance and rumen development measures by the end of the trial.

International audience; Metabolome profiling in biological fluids is an interesting approach for exploring markers of methane emissions in ruminants. In this study, a multiplatform metabolomics approach was used for investigating changes in milk metabolic profiles related to methanogenesis in dairy cows. For this purpose, 25 primiparous Holstein cows at similar lactation stage were fed the same diet supplemented with (treated, n = 12) or without (control, n = 13) a specific antimethanogenic additive that reduced enteric methane production by 23% with no changes in intake, milk production, and health status. The study lasted 6 wk, with sampling and measures performed in wk 5 and 6. Milk samples were analyzed using 4 complementary analytical methods, including 2 untargeted (nuclear magnetic resonance and liquid chromatography coupled to a quadrupole-time-of-flight mass spectrometer) and 2 targeted (liquid chromatography-tandem mass spectrometry and gas chromatography coupled to a flame ionization detector) approaches. After filtration, variable selection and normalization data from each analytical platform were then analyzed using multivariate orthogonal partial least square discriminant analysis. All 4 analytical methods were able to differentiate cows from treated and control groups. Overall, 38 discriminant metabolites were identified, which affected 10 metabolic pathways including methane metabolism. Some of these metabolites such as dimethylsulfoxide, dimethylsulfone, and citramalic acid, detected by nuclear magnetic resonance or liquid chromatography-mass spectrometry methods, originated from the rumen microbiota or had a microbial-host animal co-metabolism that could be associated with methanogenesis. Also, discriminant milk fatty acids detected by targeted gas chromatography were mostly of ruminal microbial origin. Other metabolites and metabolic pathways significantly affected were associated with AA metabolism. These findings provide new insight on the potential role of milk metabolites as indicators of enteric methane modifications in dairy cows.

Subclinical mastitis (SCM) represents a significant burden and challenge to modern dairy management. Multidrug-resistant Escherichia coli (MDR E. coli) in milk poses a public health threat to humans especially via the consumption of unpasteurized dairy products. This study aimed to determine the occurrence of MDR E. coli in cows and buffalo in the households of the western part of the Chitwan district of Nepal. A total of 243 lactating cows and buffalo were included in this study. Milk samples (n = 972) were screened using the California Mastitis Test (CMT). The E. coli was isolated from milk samples that were positive for CMT using standard bacteriological protocols. A semi-structured questionnaire was administered to farmers to identify the risk factors associated with the occurrence of SCM in cows and buffalo. Of the 243 dairy animals screened, 42.8% (n = 104/243) showed positive CMT results. However, of the 972 quarters sampled, only 19.3% (n = 188/972) were positive for SCM. The prevalence of E. coli in these animals was found to be 16.5% in animals (n = 40/243). However, E. coli was isolated from only 5% (n = 49/972) of the quarters. Of the 49 E. coli isolated, the resistance to ceftriaxone (38.8%, n = 19/49) and ciprofloxacin (37.7%, n = 17/49) were the most prevalent. Animals with a history of mastitis were 3.57 times more likely to have SCM than other animals. Similarly, lactating animals with previous teat abrasions were 3.22 times more likely to develop SCM than animals without teat injuries. As expected, cleaning the barn once in 2 to 3 d was associated with an increased occurrence of SCM in lactating cows. This study reports the occurrence of MDR E. coli in SCM, which poses a public health threat. Creating awareness of milk pasteurization, and food safety practices are necessary among the farmers.

Low dietary energy and decreased intake of herbage have been attributed to the reduced performance of grazing dairy cattle. We hypothesized that grasses with inherently greater energy would interact in a complementary way with condensed tannins (CT) in birdsfoot trefoil to increase herbage intake by grazing dairy heifers. Eight pasture treatments comprising high-sugar perennial ryegrass (Lolium perenne L.), orchardgrass (Dactylis glomerata L.), meadow bromegrass (Bromus riparius Rehmann), and tall fescue [Schendonorus arundinaceus (Schreb.) Dumort] were established in Lewiston, Utah as monocultures and binary mixtures with birdsfoot trefoil (Lotus corniculatus L.; BFT). Pasture treatments were rotationally stocked by Jersey heifers for 105 d in 2017 and 2018, and herbage samples were collected pre- and postgrazing for each 7-d grazing period and analyzed for herbage mass, nutritive value, and apparent herbage intake. We observed differences among pasture treatments in herbage quantity and nutritive value, as well as differences in herbage intake by grazing Jersey heifers. On average, grass-BFT mixtures had greater herbage intake than grass monocultures, and every grass-BFT treatment individually had greater herbage intake than their respective grass monocultures. Using multivariate analyses, we determined that approximately 50% of the variation in herbage intake was due to nutritive and physical herbage characteristics, with the most explanatory being characteristics related to fiber and energy, followed by those related to the percent of BFT in the herbage. Grass monocultures exhibited a range of inherent dietary energy, but there was indication that an imbalance of energy to crude protein (e.g., protein deficient) reduced intake of grass monocultures. Moreover, there was some evidence of a complementary effect between increased dietary energy and CT; however, low CT levels made it impossible to determine the effect of CT on herbage intake per se. This study confirmed that chemical and physical characteristics inherent to different pasture species have a large effect on herbage intake by grazing cattle. Pastures planted to binary mixtures of nutritious grasses and birdsfoot trefoil increase herbage intake of temperate pastures by grazing Jersey heifers.

The objective of this study was to investigate the effects of delayed increase in milk allowance and type of gradual weaning program on the feed intake, health, and growth of male Holstein calves. Using a 2 × 2 factorial design, each male Holstein calf was randomly assigned to 1 of 2 milk level treatments and 1 of 2 weaning treatments upon arrival to a rearing facility. Calves were either assigned to a moderate (MM) or lower (LM) milk level treatment. All calves received the same milk program from arrival until d 12, increasing from 4 to 5 L/d (115 to 125 g/L) over the 12 d. On d 13, MM calves began receiving additional milk compared with LM calves until they were weaned, with MM offered milk increasing to a peak of 9 L/d (140 g/L). Calves on the MM treatment received a total of 35 kg of milk replacer over 45 d, compared with LM calves, who received a total of 25 kg of milk replacer over 45 d. Calves were also assigned to be weaned by either a 2-step gradual program or a 4-step gradual program. Calves assigned to the 2-step program had their milk reduced in 2 steps from d 30 to 45, and 4-step calves had their milk reduced in 4 steps from d 30 to 45. After d 46, calves received no milk but continued to be monitored until d 59. Calves had ad libitum access to water from arrival, and solid feed was offered from d 5 onward. No interactions of effects of milk level and weaning treatment were detected. By design, calves on the MM treatment consumed greater amounts of milk than LM calves in the preweaning phase from d 13 to 29 (6.1 vs. 4.8 L/d) and weaning phases from d 30 to 45 (4.7 vs. 3.2 L/d). The LM calves consumed greater amounts of solid feed than MM calves from d 22 to 29 of the preweaning phase, and from d 30 to 40 and d 43 to 45 of the weaning phase, whereas postweaning feed consumption was similar for both milk treatments. Calves in both milk treatments had similar blood β-hydroxybutyrate levels, indicating similar rumen development. Body weight and size measurements were similar between milk treatments across the study, although LM calves had a greater average daily gain than MM calves between d 38 and 46 (0.96 vs. 0.71 kg/d). Both gradual weaning treatments resulted in similar feed consumption, growth, and health parameters. Overall, the results indicate that delaying a moderate increase in milk until 12 d after arrival at a rearing facility provided no overall beneficial effect on growth or rumen development.

The objective of this randomized clinical study was to compare the effect of 2 antimicrobial interventions, tildipirosin or florfenicol + flunixin meglumine, used for treatment of pneumonia and extralabel treatment for otitis on health parameters and upper respiratory tract (URT) microbiota of preweaned Holstein calves. Housed preweaned Holstein heifers diagnosed with either otitis or pneumonia were assigned into 1 of 2 treatment groups, receiving a single subcutaneous injection of either 4 mg/kg of tildipirosin (TLD; n = 444) or 40 mg/kg of florfenicol combined with 2.2 mg/kg of a nonsteroidal anti-inflammatory, flunixin meglumine (FLF; n = 442). Calves were enrolled and treated on the day of diagnosis of the first case of pneumonia or otitis. If a calf had a recurrent case, the opposite drug was administered, respecting an interval of 5 d between drug injections. Blood samples for leukocyte counts were collected at 0, 2, 4, and 6 d after treatment, and rectal temperature was measured daily during the 5 d after treatment. Ear scores were observed from calves with otitis. Additionally, swabs of the URT were collected from a subset of 20 calves in each treatment group at d 0, 3, 6, 9, and 11 following enrollment for analysis of URT microbiota through next-generation sequencing of the 16S rRNA gene and quantitative PCR. Swabs were also collected from a comparative group of 20 healthy calves that did not receive any drug. No differences were observed between groups for recurrence risk of either pneumonia (TLD = 32.4%; FLF = 29.7%) or otitis (TLD = 72.7%; FLF = 73.6%). Similarly, no differences were observed for the total number of treatments for pneumonia (TLD = 1.45; FLF = 1.42) or otitis (TLD = 2.96; FLF = 3.07). On the other hand, both drugs reduced rectal temperature, ear scores, and leukocyte counts, with FLF calves having a greater reduction in rectal temperature within 4 d after treatment. Both TLD and FLF reduced the total bacterial load when compared with healthy untreated calves, but no differences were observed between treatment groups. Furthermore, compared with the untreated group, treated calves had lower mean relative abundances (MRA) of the genera Mannheimia, Moraxella, and Pasteurella within 11, 9, and 3 d after treatment, respectively; however, no significant differences were observed between TLD and FLF. On the other hand, MRA of Mycoplasma was not decreased by both treatments compared to untreated animals, and a higher MRA was observed in the TLD group during 11 d after treatment in comparison to FLF and untreated calves. Based on this data, we concluded that both drugs used in the study were effective in reducing rectal temperature, ear scores, leukocyte counts, and MRA of the genera Mannheimia, Pasteurella, and Moraxella in the URT, and calves treated with FLF had a greater reduction in rectal temperature.

Ketosis is one of the most frequent metabolic diseases in high-yielding dairy cows and is characterized by high concentrations of ketone bodies in blood, urine, and milk, causing high economic losses. The search for polymorphic genes, whose alleles have different effects on resistance to developing the disease, is of extreme importance to help select less susceptible animals. The aims of this study were to identify genomic regions associated with clinical and subclinical ketosis (β-hydroxybutyrate concentration) in North American Holstein dairy cattle and to investigate these regions to identify candidate genes and metabolic pathways associated with these traits. To achieve this, a GWAS was performed for 4 traits: clinical ketosis lactation 1, clinical ketosis lactation 2 to 5, subclinical ketosis lactation 1, and subclinical ketosis lactation 2 to 5. The estimated breeding values from 77,277 cows and 7,704 bulls were deregressed and used as pseudophenotypes in the GWAS. The top-20 genomic regions explaining the largest proportion of the genetic variance were investigated for putative genes associated with the traits through functional analyses. Regions of interest were identified on chromosomes 2, 5, and 6 for clinical ketosis lactation 1; 3, 6, and 7 for clinical ketosis lactation 2 to 5; 1, 2, and 12 for subclinical ketosis lactation 1; and 20, 11, and 25 for subclinical ketosis lactation 2 to 5. The highlighted genes potentially related to clinical and subclinical ketosis included ACAT2 and IGF1. Enrichment analysis of the list of candidate genes for clinical and subclinical ketosis showed molecular functions and biological processes involved in fatty acid metabolism, lipid metabolism, and inflammatory response in dairy cattle. Several genomic regions and SNPs related to susceptibility to ketosis in dairy cattle that were previously described in other studies were confirmed. The novel genomic regions identified in this study aid to characterize the most important genes and pathways that explain the susceptibility to clinical and subclinical ketosis in dairy cattle.

The aim of this study was to analyze time-lagged heat stress (HS) effects during late gestation on genetic co(variance) components in dairy cattle across generations for production, female fertility, and health traits. The data set for production and female fertility traits considered 162,492 Holstein Friesian cows from calving years 2003 to 2012, kept in medium-sized family farms. The health data set included 69,986 cows from calving years 2008 to 2016, kept in participating large-scale co-operator herds. Production traits were milk yield (MKG), fat percentage (fat%), and somatic cell score (SCS) from the first official test-day in first lactation. Female fertility traits were the nonreturn rate after 56 d (NRR56) in heifers and the interval from calving to first insemination (ICFI) in first-parity cows. Health traits included clinical mastitis (MAST), digital dermatitis (DD), and endometritis (EM) in the early lactation period in first-parity cows. Meteorological data included temperature and humidity from public weather stations in closest herd distance. The HS indicator was the temperature-humidity index (THI) during dams' late gestation, also defined as in utero HS. For the genetic analyses of production, female fertility, and health traits in the offspring generation, a sire-maternal grandsire random regression model with Legendre polynomials of order 3 for the production and of order 2 for the fertility and health traits on prenatal THI, was applied. All statistical models additionally considered a random maternal effect. THI from late gestation (i.e., prenatal climate conditions), influenced genetic parameter estimates in the offspring generation. For MKG, heritabilities and additive genetic variances decreased in a wave-like pattern with increasing THI. Especially for THI >58, the decrease was very obvious with a minimal heritability of 0.08. For fat% and SCS, heritabilities increased slightly subjected to prenatal HS conditions at THI >67. The ICFI heritabilities differed marginally across THI [heritability (h2) = 0.02–0.04]. For NRR56, MAST, and DD, curves for heritabilities and genetic variances were U-shaped, with largest estimates at the extreme ends of the THI scale. For EM, heritability increased from THI 25 (h2 = 0.13) to THI 71 (h2 = 0.39). The trait-specific alterations of genetic parameters along the THI gradient indicate pronounced genetic differentiation due to intrauterine HS for NRR56, MAST, DD, and EM, but decreasing genetic variation for MKG and ICFI. Genetic correlations smaller than 0.80 for NRR56, MAST, DD, and EM between THI 65 with corresponding traits at remaining THI indicated genotype by environment interactions. The lowest genetic correlations were identified when considering the most distant THI. For MKG, fat%, SCS, and ICFI, genetic correlations throughout were larger than 0.80, disproving concerns for any genotype by environment interactions. Variations in genetic (co)variance components across prenatal THI may be due to epigenetic modifications in the offspring genome, triggered by in utero HS. Epigenetic modifications have a persistent effect on phenotypic responses, even for traits recorded late in life. However, it is imperative to infer the underlying epigenetic mechanisms in ongoing molecular experiments.

In rodents and humans, the gut bacteria-derived metabolite trimethylamine N-oxide (TMAO) has been implicated in the progression of cardiovascular disease, chronic kidney disease, fatty liver, and insulin resistance; however, the effects of TMAO on dairy cattle health and milk production have not been defined. We aimed to determine whether intravenous TMAO infusion modifies measures of liver health, glucose tolerance, and milk production in early-lactation cows. Eight early-lactation Holstein cows (30.4 ± 6.41 d in milk; 2.88 ± 0.83 lactations) were enrolled in a study with a replicated 4 × 4 Latin square design. Cows were intravenously infused TMAO at 0 (control), 20, 40, or 60 g/d for 6 d. Washout periods lasted 9 d. Intravenous glucose tolerance tests (GTT) occurred on d 5. Blood was collected daily. Milk was collected on d -1, 0, 5, and 6. Urine was collected on d -1 and 6. Circulating metabolites, milk components, and TMAO concentrations in milk, urine, and plasma were quantified. Data were analyzed using a mixed model that included the fixed effects of treatment. Concentrations of TMAO in plasma, milk, and urine increased linearly with increasing dose. Dry matter intake and milk production were not modified by treatment. Daily plasma triacylglycerol, fatty acid (FA), and glucose concentrations were not modified. Serum albumin, total protein, globulin, total bilirubin, direct bilirubin, aspartate aminotransferase, γ-glutamyl transferase, and glutamate dehydrogenase concentrations were also not modified by treatment. Serum GTT glucose, FA, and insulin concentrations were not modified by treatment. Plasma total, reduced, and oxidized glutathione concentrations were also not modified by treatment. We conclude that a 6-d intravenous infusion of TMAO does not influence measures of liver health, glucose tolerance, or milk production in early-lactation dairy cows.

The aim of this study was to investigate the effect of controlled daily alternations in dietary starch level on changes in rumen environment, blood, urine, and milk metabolites of dairy cows. Six multiparous midlactation Holstein cows were used in a replicated 3 × 3 Latin square design with 14-d periods and 3 alternatinglevels of dietary starch as treatments. Each 14-d period consisted of a 7-d baseline period and 7-d alternating period where diets alternated day to day. During the baseline period, all cows were fed a control diet containing 21% starch (dry matter basis). During the alternating period, the control diet was replaced with 1 of the 3 experimental diets on d 8, 10, 12, and 14. The 3 experimental diets contained 28% (low), 35% (medium), and 42% (high) starch (dry matter basis). At d7 (baseline), 8 (ALT1), and 14 (ALT4) of each period,rumen fluid, blood, urine, and quarter milk (i.e., back right quarter) samples were collected at −0.5, 1, 2.5, 4, 5.5, and 7 h relative to morning feeding (0800 h). No differences were observed in dry matter intake, milk yield, and milk chemical composition. Rumen medial pH was lower in the high alternation level compared with the low or medium alternation levels at ALT1 but did not differ among starch alternation levels at ALT4. Similarly, the difference between rumen pH in medial and ventral contents was reduced at ALT1 with high alternation level but was not affected at ALT4. Total volatile fatty acid (VFA) concentrations were higher in the rumen medial fluid of the high alternation level at 7 h relative to morning feeding compared with those from the low and medium alternation levels. Similarly, total VFA concentrations constantly increased and were thehighest in the ventral rumen fluid at 7 h relative to morning feeding, although no differences were detected among starch alternation levels. In both rumen medialand ventral fluids, the high alternation level showed higher propionate and lower acetate proportions compared with low and medium alternation levels. No differences in blood pH were detected among starch alternation levels. However, glucose concentrations tended to be higher in cows from the high alternation level. l-Lactate concentrations in blood were higher in ALT1 than in ALT4 but were not affected by the starch alternation level. In urine, no differences in pH or l-lactate concentrations were detected among alternation levels (i.e., low, medium, and high). Similarly, no differences in milk pH were detected among alternation levels. According to these results, it seems that the daily dietarystarch alternation from 21% up to 42% (dry matter basis) is able to affect the ruminal fluid, especially during the first alternation. However, these changes in rumen fluid did not cause any effect on the variables measured in blood, urine, or milk. This study indicates that cows can cope with day-to-day alternations in type of rumen fermentable organic matter; however, longer-term effects on performance and health should be addressed in future studies.

The adoption of automated milk feeders and group housing of preweaning dairy calves has become more common in Canada; however, disease detection in group-housed calves remains a challenge. The aim of this cross-sectional study was to assess whether feeding behavior data collected from a single point in time could be used to aid in the detection of neonatal calf diarrhea (NCD), bovine respiratory disease (BRD), and general disease, in preweaning group-housed calves being fed via an automated milk feeder. The data used was collected in an earlier study. A total of 8 dairy farms recruited from an online survey of calf-management practices were enrolled into the study. There was a total of 523 observations with 130 events of NCD, 115 events of BRD, and 210 events of general disease. Each farm was visited once in each of the fall, winter, spring, and summer, when the calves' health was scored, and the data were collected from the automated milk feeders. Mixed linear regression models were used to identify associations between feeding behavior data (milk consumption, time spent at the feeder, drinking speed, and the number of rewarded and unrewarded visits) and the presence of NCD, BRD, or general disease (having one or more of NCD, BRD, or umbilical infection), on the day of health scoring. Generalized linear mixed models were used to analyze the percentage of milk the calf consumed from their daily milk allotment. Calves with BRD consumed 63% less of their daily allotment of milk, had 2 fewer unrewarded visits to the automated milk feeder, and drank milk 152 mL/min slower compared with calves without BRD. Calves with NCD consumed 57% less of their daily milk allotment, consumed 758 mL less per day, and drank 92 mL/min slower than calves compared with calves without NCD. Calves with general disease drank 50% less of their daily milk allowance, consumed 496 mL less per day, drank 80 mL/min slower, and had 2 fewer unrewarded visits to the automated milk feeder, when compared with calves without disease. No significant associations were found between the presence of NCD, BRD, or general disease and time spent at the feeder or number of rewarded visits. Sensitivity and specificity values for disease identification were low when evaluating the feeding behaviors individually, so parallel testing was completed. To do so, if any significant feeding behavior was below the optimal cut point for disease detection as determined using a ROC curve, the calf was considered positive for disease and the sensitivity and specificity were recalculated. Parallel testing resulted in a sensitivity of 0.82, 0.78, and 0.84, and a specificity of 0.26, 0.23, and 0.21, for BRD, NCD, and general disease, respectively. This suggests that automated milk feeders may serve as a useful preliminary tool in the detection of diseased calves. For example, producers could use feeding behavior data to identify calves requiring further inspection; however, they should not use feeding behavior data as a sole disease detection method.

Intensified milk replacer (MR) feeding in calves has nutritional long-term effects and is suggested to increase milk production later in life. However, the underlying mechanisms are not completely understood. The aim of our study was to investigate whether MR feeding intensity has long-term effects on energy metabolism and energy use efficiency of dairy calves. Newborn female Holstein calves (n = 28) were randomly assigned to 2 liquid feeding groups offered daily either 10% of body weight (BW) colostrum followed by 10% of BW MR (10%-MR) or 12% of BW colostrum followed by 20% of BW MR (20%-MR). Calves were housed individually. Weaning was completed by the end of wk 12. Hay and calf starter were fed from d 1 until the end of wk 14 and 16, respectively. A total mixed ration was fed from wk 11 onward, and the metabolizable energy intake (MEI) was determined daily. Energy metabolism of calves was measured in respiratory chambers before weaning in wk 6 and 9, and after weaning in wk 14 and 22. The MEI/BW0.75 was higher before weaning but lower during and shortly after weaning in 20%-MR calves. During the preweaning period, the 20%-MR animals had higher average daily gain, BW, back fat thickness and muscle diameter, but lower plasma β-hydroxybutyrate concentrations. The group difference in average daily gain ceased in wk 9, differences in back fat thickness and muscle diameter ceased after weaning, whereas difference in BW0.75 persisted until wk 23. The energy conversion ratio (BW gain/MEI) was not different before weaning, but was lower during and after weaning in 20%-MR calves. The higher MEI and BW0.75 in 20%-MR calves resulted in higher heat production (HP), as well as in higher carbohydrate oxidation (COX) and fat oxidation during the preweaning period. Gas exchange variables normalized to BW0.75 or MEI differed between groups only during preweaning. The energy balance was lower in 10%-MR calves in wk 6 and 9. The HP/BW0.75 and COX/BW0.75 were higher, whereas HP/MEI was lower in 20%-MR calves in wk 6. When normalized to BW0.75 and MEI, HP in wk 6 and 9, and COX in wk 9 was lower in 20%-MR calves. In conclusion, 20%-MR calves showed greater efficiency estimates preweaning, but this effect did not occur after weaning, suggesting that energy use efficiency does not persist until later stages in life.

The objective of our study was to determine the effects of altering the ratio of stearic (C18:0; SA) and oleic (cis-9 C18:1; OA) acids in supplemental fatty acid (FA) blends on FA digestibility and milk yield of dairy cows. Eight multiparous Holstein cows (mean ± SD; 157 ± 11.8 d in milk) were randomly assigned to treatment sequence in a replicated 4 × 4 Latin square design with 14-d periods. Digestibility and production data were collected during the last 4 d of each period. The treatments were an unsupplemented control diet (CON), and 3 diets incorporating FA supplement blends at 1.4% of diet dry matter (DM) containing (as a % of total FA) 50% SA and 10% OA, 40% SA and 20% OA, or 30% SA and 30% OA. The FA blends were balanced to contain 33% palmitic, 5% linoleic, and

ComX can improve bacterial competence by modulating global gene expression. Although competence induction may also be a protective mechanism under stress, this has not been investigated in detail. Here, we demonstrated that ComX improved the acid tolerance and nisin yield of Lactococcus lactis, which is an important gram-positive bacterium increasingly used in modern biotechnological applications. We found that overexpression of comX could improve the survival rate up to 36.5% at pH 4.0, compared with only 5.4% and 1.1% with the wild-type and comX knockout strains, respectively. Moreover, quantitative real-time PCR results indicated that comX overexpression stimulated the expression of late competence genes synergistically with exposure to acid stress. Finally, electrophoretic mobility shift assay demonstrated the binding of purified ComX to the cin-box in the promoters of these genes. Taken together, our results reveal a regulation mechanism by which ComX and acid stress can synergistically modulate the expression of late competence genes to enhance cells' acid tolerance and nisin yield.

The growing amount of genomic information in dairy cattle has increased computational and modeling challenges in the single-step evaluations. The computational challenges are due to the dense inverses of genomic (G) and pedigree (A22) relationship matrices of genotyped animals in the single-step mixed model equations. An equivalent mixed model equation is given by single-step genomic BLUP that are based on the T matrix (ssGTBLUP), where these inverses are avoided by expressing G−1 through a product of 2 rectangular matrices, and (A22)−1 through sparse matrix blocks of the inverse of full relationship matrix A−1. A proper way to account genetic groups through unknown parent groups (UPG) after the Quaas-Pollak transformation (QP) is one key factor in a single-step model. When the UPG effects are incompletely accounted, the iterative solving method may have convergence problems. In this study, we investigated computational and predictive performance of ssGTBLUP with residual polygenic (RPG) effect and UPG. The QP transformation used A−1 and, in the complete form, T and (A22)−1 matrices as well. The models were tested with official Nordic Holstein milk production test-day data and model. The results show that UPG can be easily implemented in ssGTBLUP having RPG. The complete QP transformation was computationally feasible when preconditioned conjugate gradient iteration and iteration on data without explicitly setting up G or A22 matrices were used. Furthermore, for good convergence of the preconditioned conjugate gradient method, a complete QP transformation was necessary.

Understanding the regulation of cellular AA uptake as protein supply changes is critical for predicting milk component yields because intracellular supplies partly regulate protein synthesis. Our objective was to evaluate cellular uptake and kinetic behavior of individual AA when cells are presented with varying extracellular AA supplies. Bovine primary mammary epithelial cells were grown to confluency and transferred to medium with an AA profile and concentration similar to that of plasma from dairy cows for 24 h. Treatments were 4 AA concentrations, 0.36, 2.30, 4.28, and 6.24 mM, which represented 16, 100, 186, and 271% of typical plasma AA concentrations, respectively, in lactating dairy cows. Twenty-four plates of cells (89.4 × 19.2 mm) were assigned to each treatment. Cells were first subjected to treatment medium enriched with 15N-labeled AA for 24 h and then incubated with treatment medium enriched with 13C-labeled AA for 0, 15, 60, 300, 900, 1,800, and 3,600 s. Intracellular free AA, intracellular protein-bound AA, and extracellular medium free AA were analyzed for concentrations and isotopic enrichment using gas chromatography-mass spectrometry. A dynamic, 12-pool model was fitted to the data for 14 AA to derive unidirectional uptake and efflux, protein turnover, transamination, oxidation, and synthesis. The derived concentration for half the maximal uptake (km) indicated no saturation of AA uptake at typical in vivo concentrations for 11 of the 14 AA. Arginine, Pro, and Val appeared to exhibit saturation kinetics. Net uptake of all essential AA except Phe was positive across treatments. Most nonessential AA exhibited negative net uptake values. Efflux of AA was quite high, with several AA exhibiting greater than 100% efflux of the respective influx. Intracellular pool turnover was rapid for most AA (e.g., 2 min for Arg), demonstrating plasticity in matching needs for protein translation to supplies. Intracellular AA concentrations increased linearly in response to treatment for most AA, whereas 9 AA exhibited quadratic responses. Amino acid uptake is responsive to varying extracellular supplies to maintain homeostasis. No saturation of uptake was evident for most AA, indicating that transporter capacity is likely not a limitation for most AA except possibly Arg, Val, and Pro in mammary epithelial cells.

Maintenance energy is the energy required to conserve the state of an animal when no work is completed. Dietary energy must be supplied to meet maintenance requirements before milk can be produced. The objectives of the current experiment were to quantify the maintenance energy requirement of Jersey cows when lactating or dry. Energetic measures were collected on 8 Jersey cows and evaluated across 3 physiological phases and nutritional planes: lactation, dry cows fed at maintenance, and fasted dry cows. Through total collection of feces and urine as well as using headbox-style indirect calorimeters, energy balance and heat production data were measured across all phases. Lactation data were collected across four 28-d periods. Data for cows fed at maintenance were collected after 14 d and fasting heat production was measured during the last 24 h of a 96-h fast. Net energy for maintenance (NEM) requirements, and the efficiency of converting metabolizable energy (ME) into net energy were compared between lactating and dry (maintenance or fasting phase) cows. Heat production of dry cows fed at maintenance, which represents ME for maintenance, was 0.146 ± 0.0087 Mcal per unit of metabolic body weight (BW0.75, MBW). Fasting heat production, which represents NEM, was 0.102 ± 0.0071 Mcal/MBW. Energy balance was calculated as tissue energy plus milk energy. When estimated via regressing energy balance on ME intake, NEM was not different between dry and lactating cows (0.120 ± 0.32 vs. 0.103 ± 0.0052 Mcal/MBW). However, the slope of the regression of energy balance on ME intake was greater for dry compared with lactating cows (0.714 ± 0.046 vs. 0.685 ± 0.010) when evaluated with a fixed intercept. This suggests that dry cows were more efficient at converting ME into net energy and that the efficiency of utilizing ME for maintenance may be greater than for lactation. Our measurements of NEM and the slope of ME on energy balance were greater than the value used by the National Research Council (2001), which are 0.080 Mcal/MBW for NEM and approximately 0.64 for the slope. Results of this study suggest that NEM and the efficiency of converting ME into NEM of modern lactating Jersey cows are similar to recent measurements on modern Holstein cows and greater than previous measurements.

The objective of this experiment was to determine the effect of increasing digestible His (dHis) doses on milk production, milk composition, and plasma AA concentrations in lactating dairy cows fed diets that meet or exceed their energy and metabolizable protein (MP) requirements. In a companion paper (Räisänen et al., 2021) results are presented on the effect of increasing dHis dose with an MP-deficient basal diet. In this experiment, 16 Holstein cows (72 ± 15 d in milk) were used in a replicated 4 × 4 Latin square design experiment with four 28-d periods. Treatments were as follows: (1) control, total mixed ration (TMR) with 1.8% dHis of MP (TMR1; dHis1.8); (2) a different TMR with 2.2% dHis (TMR2; dHis2.2); (3) TMR2 supplemented with rumen-protected His (RP-His) to supply 2.6% dHis (dHis2.6); and (4) TMR2 supplemented with RP-His to supply 3.0% dHis of MP (dHis3.0). Estimated dHis intakes calculated at the end of the experiment were 46, 58, 69, and 79 g/d for dHis1.8, dHis2.2, dHis2.6, and dHis3.0, respectively. Contrasts were used to compare TMR1 with TMR2 and to test the linear and quadratic effects of RP-His inclusion rate on TMR2. We detected no effects of TMR or dHis dose on dry matter intake or milk yield, whereas energy-corrected milk (ECM) yield was quadratically increased, being greatest for cows on treatment dHis2.6. Milk true protein and lactose concentrations and milk true protein yield were not affected by TMR or dHis dose. Milk fat concentration and yield increased quadratically, and lactose yield tended to increase quadratically with increasing dHis dose. Calculated apparent efficiency of His utilization decreased quadratically with increasing dHis supply. Further, plasma concentration of His was greater for cows on TMR2 compared with TMR1. When an MP-adequate diet was fed to dairy cows, milk true protein concentration and yield were not affected by dHis supply, but milk fat and ECM yields of dairy cows were optimized at dHis supply of 69 g/d or 2.65% of MP.

Four sialylated lactuloses [N-acetylneuraminic acid-α2,3-lactulose (Neu5Acα2,3lactulose), N-acetylneuraminic acid-α2,6-lactulose (Neu5Acα2,6lactulose), deaminoneuraminc acid-α2,3-lactulose (Kdnα2,3lactulose), and deaminoneuraminc acid-α-2,6-lactulose (Kdnα2,6lactulose)] were reported to modulate the immunity of mice. The influences of cytokine expression, cell immunity, humoral immunity, and nonspecific immunity were investigated in our study using several techniques. Analysis via ELISA showed that cytokine expression was induced by sialylated lactulose treatment consistently in the serum and spleen. Among the 4 tested sialylated lactuloses, Neu5Acα2,6lactulose performed the best, simultaneously and appropriately promoting the expression of proinflammatory and anti-inflammatory factors in the serum and spleen. Kdnα2,3lactulose showed the best antioxidant activity according to detection of the activity of superoxide dismutase, myeloperoxidase, peroxidase, and alkaline phosphatase. Flow cytometry revealed that only Kdnα2,3lactulose significantly boosted the CD3+ T lymphocyte ratio similarly to that of lactulose. Analysis of the hemolysin content to characterize humoral immunity revealed that Kdnα2,3lactulose notably increased hemolysin content compared with that in the control group. To evaluate the nonspecific immune effects of the 4 sialylated lactuloses, a fluorescence microsphere phagocytosis assay was used to analyze the phagocytosis of macrophages. Kdnα2,3lactulose still performed the best in enhancing the phagocytosis of macrophages, showing markedly increased phagocytic percentage and phagocytic index values compared with those in the control and lactulose groups. Comparing the differences of these 4 sialylated lactuloses in affecting immunity in mice revealed that Kdnα2,3lactulose had the best overall performance in influencing cytokine expression, cell immunity, humoral immunity, and nonspecific immunity. This study provides critical support for use of sialylated lactuloses as potential immunomodulators in foods.

Suppression of appetite, or hypophagia, is among the most recognizable effects of disease in livestock, with the potential to impair growth, reproduction, and lactation. The continued evolution of the field of immunology has led to a greater understanding of the immune and endocrine signaling networks underlying this conserved response to disease. Inflammatory mediators, especially including the cytokines tumor necrosis factor-α and interleukin-1β, are likely pivotal to disease-induced hypophagia, based on findings in both rodents and cattle. However, the specific mechanisms linking a cytokine surge to decreased feeding behavior are more difficult to pin down and likely include direct effects on appetite centers in the brain, alteration of gastric motility, and modulation of other endocrine factors that influence appetite and satiety. These insights into the mechanisms for disease-induced hypophagia have great relevance for management of neonatal calves, mature cows transitioning to lactation, and cows experiencing mastitis; however, it is not necessarily the case that increasing feed intake by any means possible will improve health outcomes for diseased cattle. We explore conflicting effects of hypophagia on immune responses, which may be impaired by the lack of specific substrates, versus apparent benefits for controlling the growth of some pathogens. Anti-inflammatory strategies have shown promise for promoting recovery of feed intake following some conditions but not others. Finally, we explore the potential for early disease detection through automated monitoring of feeding behavior and consider which strategies may be implemented to respond to early hypophagia.

Our primary objective was to perform a meta-analysis and meta-regression to evaluate the effects of diets supplemented with calcium salts of palm fatty acids (CSPF) compared with nonfat supplemented control diets (CON) on nutrient digestibility and production responses of lactating dairy cows. Our secondary objective was to perform a meta-analysis to evaluate whether experimental design affects production responses to supplemental CSPF. The data set was formed from 33 peer-reviewed publications with CSPF supplemented at ≤3% diet dry matter. We analyzed the interaction between experimental design (continuous vs. change-over) and treatments (CON vs. CSPF) to evaluate whether experimental design affects responses to CSPF (Meta.1). Regardless of experimental design, we evaluated the effects of CSPF compared with CON on nutrient digestibility and production responses of lactating dairy cows by meta-analysis (Meta.2) and meta-regression (Meta.3) approaches. In Meta.1, there was no interaction between treatments and experimental design for any variable. In Meta.2, compared with CON, CSPF reduced dry matter intake [DMI, 0.56 ± 0.21 kg/d (±SE)] and milk protein content (0.05 ± 0.02 g/100 g), increased neutral detergent fiber (NDF) digestibility (1.60 ± 0.57 percentage units), the yields of milk (1.53 ± 0.56 kg/d), milk fat (0.04 ± 0.02 kg/d), and 3.5% fat corrected milk (FCM, 1.28 ± 0.60 kg/d), and improved feed efficiency [energy corrected milk (ECM)/DMI, 0.08 kg/kg ± 0.03]. There was no effect of treatment for milk protein yield, milk fat content, body weight, body weight change, or body condition score. Compared with CON, CSPF reduced the yield of de novo milk fatty acids (FA) and increased the yields of mixed and preformed milk FA. In Meta.3, we observed that each 1-percentage-unit increase of CSPF in diet dry matter reduced DMI, increased NDF digestibility, tended to increase FA digestibility, increased the yields of milk, milk fat, and 3.5% FCM, reduced the content of milk protein, reduced the yield of de novo milk FA, and increased the yields of mixed and preformed milk FA. In conclusion, our results indicate no reason for the restrictive use of change-over designs in CSPF supplementation studies or meta-analysis. Feeding CSPF increased NDF digestibility, tended to increase FA digestibility, and increased the yields of milk, milk fat, and 3.5% FCM. Additionally, CSPF increased milk fat yield by increasing the yields of mixed and preformed milk FA.

Selective use of antibiotic dry cow treatment can be implemented at the cow or quarter level, with the latter having the potential to further reduce antibiotic use. Our objective was to compare these 2 approaches in 6 herds in the United Kingdom in which environmental mastitis predominated. Eight hundred seven cows were enrolled and categorized as having a high cell count (n = 401) or low cell count (n = 406) in the last 3 mo of lactation and clinical mastitis history. All quarters of all enrolled cows received an internal teat sealant. Within each category, cows were randomly allocated to 1 of 3 groups; in one group antibiotic treatment was allocated at cow level (i.e., all 4 quarters received antibiotic), whereas in the 2 remaining groups antibiotic treatment was allocated at quarter level, based on California Mastitis Test (CMT) findings. Two different thresholds, score 1 and 2, were used to determine likely infection status. Quarter milk samples were collected at dry off and postcalving for bacteriological culture and somatic cell count (SCC). Cows were monitored for clinical mastitis from dry off until 100 d in milk. Cow level SCC and milk yield data were collated from farm records. Within each category, the 2 quarter level treatment groups were compared with cow level treatment at dry off. Leaving quarters untreated with intramammary antibiotic in cows in the high cell count group, with a CMT2 or1, reduced antibiotic use by 55% and 31%, respectively, and resulted in no difference in the odds of being infected with any pathogen postcalving, but was associated with a higher SCC at the first test day. Intramammary antibiotic treatment of quarters with a CMT ≥1 in cows in the low cell count category at dry off was not associated with any reduction in the odds of being infected with a major pathogen postcalving but was associated with a decrease in the odds of being infected with a minor mastitis pathogen postcalving. The use of antibiotics in quarters of cows categorized as low cell count at dry off, increased the proportion of quarters treated with antibiotic from 0% at cow level to 31% (CMT ≥ 1) and 12% (CMT ≥ 2) at quarter level, only resulting in a reduction in SCC of around 20,000 cells/mL at the first test day, if all quarters with CMT score ≥1 were treated with antibiotic. No differences in clinical mastitis incidence and milk yield in the first 100 d in milk were detected between any of the treatment groups. These study findings support selective quarter level dry off treatment only in cows with cow level SCC200,000 cells/mL at dry off.

We evaluated the effects of fatty acid (FA) supplement blends containing 60% palmitic acid (C16:0) and either 30% stearic acid (C18:0) or 30% oleic acid (cis-9 C18:1) on nutrient digestibility and milk production of low- and high-producing dairy cows. Twenty-four multiparous Holstein cows [118 ± 44 d in milk (DIM)] were divided into 2 blocks by milk production and then randomly assigned to treatment sequence in four 3 × 3 Latin squares within production level, balanced for carryover effects in three consecutive 21-d periods. Cows were blocked by milk yield and assigned to 1 of 2 groups (n = 12 per group): (a) low group (42.5 ± 3.54 kg/d; 147 ± 42 DIM) and (b) high group (55.8 ± 3.04 kg/d; 101 ± 34 DIM). Commercially available fat supplements were combined to provide treatments that consisted of the following: (1) control (CON; diet with no supplemental FA), (2) FA supplement blend containing 60% C16:0 and 30% C18:0 (PA+SA), and (3) FA supplement blend containing 60% C16:0 and 30% cis-9 C18:1 (PA+OA) The FA blends were fed at 1.5% of dry matter (DM) and replaced soyhulls from CON. Preplanned contrasts were (1) overall effect of FA treatments [CON vs. the average of the FA treatments (FAT); 1/2 (PA+SA + PA+OA)], and (2) effect of FA supplement (PA+SA vs. PA+OA). Regardless of production level, overall FAT reduced DMI compared with CON. Also, regardless of level of milk production, PA+OA increased total-tract FA digestibility compared with PA+SA. Treatment by production level interactions were observed for neutral detergent fiber (NDF) digestibility, total FA intake, and the yields of 3.5% fat-corrected milk (FCM), energy-corrected milk (ECM), and milk fat. In low-producing cows, FAT increased DM and NDF digestibility compared with CON. In high-producing cows PA+SA increased DM and NDF digestibility compared with PA+OA. In low-producing cows, PA+SA increased 3.5% FCM, ECM, and milk fat yield compared with PA+OA. However, in high-producing cows PA+OA tended to increase 3.5% FCM compared with PA+SA. In conclusion, low-producing cows responded better to a FA blend containing 60% C16:0 and 30% C18:0, whereas high-producing dairy cows responded more favorably to a FA blend containing 60% C16:0 and 30% cis-9 C18:1. However, further research is required to validate our observations that higher-yielding cows have improved production responses when supplemented with cis-9 C18:1 compared with C18:0.

Silage treated with lactic acid bacteria inoculants has been reported to increase ruminal microbial biomass when tested in vitro. Therefore, we tested if alfalfa silage inoculated with Lactobacillus plantarum MTD-1 would improve ruminal N metabolism and increase milk production in high-producing dairy cows. Twenty-eight early lactation Holstein cows (8 ruminally cannulated) were blocked by DIM and milk production; animals were used in a double crossover design consisting of four 28-d periods. Animals in each block were randomly assigned to 2 treatments: a diet containing uninoculated alfalfa silage (control) and a diet containing alfalfa silage inoculated with L. plantarum MTD-1 (LP). Diets were formulated to contain 50% of alfalfa silage, 16% crude protein, and 25% neutral detergent fiber (dry matter basis). Milk production and dry matter intake were recorded in the last 14 d of each period. Milk samples were collected twice at both daily milkings on d 20, 21, 27, and 28 of each period. On d 22, omasal samples were collected from the cannulated animals over a period of 3 d to quantify ruminal digestibility and nutrient flows. Data were analyzed using mixed models of SAS 9.4 (SAS Institute). Compared to the control, cows receiving the LP treatment had greater milk production (40.4 vs. 39.6 kg/d) and lower milk urea nitrogen concentration (11.6 vs. 12.7 mg/dL), despite minor changes in energy-corrected milk. Milk lactose concentration was greater in the milk produced by cows fed the LP treatment, which reflected a tendency for increased milk lactose yield. Although milk true protein concentration was lower for cows in the LP treatment, milk true protein yield was the same on both control and LP treatments. Improvements in milk production of animals under the LP treatment were associated with greater organic matter truly digested in the rumen, especially ruminal neutral detergent fiber digestion. Minor changes were observed in total omasal microbial nonammonia N flow in cows receiving the LP treatment. Therefore, alfalfa silage treated with L. plantarum MTD-1 may improve ruminal fermentation and milk production; however, because of a lack of response in ruminal N metabolism, these changes did not result in greater energy-corrected milk in high-producing dairy cows.

Nonreplacement dairy calves, or bobby calves, are fasted and transported to abattoirs from as young as 5 d of age in Australia. The aims of this cross-sectional observational study were (1) to assess the welfare status, as measured by blood parameters, of bobby calves in the commercial supply chain after transport and lairage, and (2) to assess whether distance and duration of transport are risk factors for poor bobby calf welfare, as measured by blood parameters. We hypothesized that bobby calves transported greater distances would be more likely to show evidence of compromised welfare, as measured by blood indicators of hydration, energy status, and muscle fatigue or damage. We also hypothesized that there would be a large amount of variability in indicators of energy status between calves from different farms. We analyzed blood samples collected at slaughter over a spring and an autumn calving period from 4,484 Australian bobby calves aged approximately 5 to 14 d old from 3 different states, after transport, fasting, and lairage. Packed cell volume (PCV), plasma glucose, and serum urea, total protein, β-hydroxybutyrate (BHB), and creatine kinase (CK) were measured. Radio frequency identification ear tag data were used to estimate the distance that the calves were transported and to identify the farm of origin. Data were analyzed using linear mixed models, except for BHB, which was analyzed using a Goodman-Kruskal gamma test due to left censoring of the data. Twelve percent of calves showed evidence of anemia (PCV less than 0.23 L/L), and 11% had urea concentrations consistent with dehydration (urea more than 7.7 mmol/L). Thirty-six percent of calves had CK activity above normal resting values, and 1% of calves had CK >2,000 U/L, indicating muscle fatigue or damage. Distance transported had significant effects on all blood variables except urea and BHB. With increasing distance transported, calves were more likely to show evidence of a negative energy balance (low plasma glucose) or dehydration (high PCV or total protein). The estimated effect of distance overall was small, but for calves transported more than 500 km, plasma glucose concentration declined more per kilometer. The calves' farm of origin accounted for a reasonable amount of the random variation between calves for plasma glucose (20%). Our results suggest that longer transport distances may increase the risk of poor calf welfare (dehydration, negative energy balance) after transport, and on-farm calf management (e.g., nutrition, timing of feeding before transport) may affect transported calves' energy status; improving this area could result in better energy availability during fasting.

California is the leading state for the production of almonds, with more than 400,000 bearing hectares of orchards that produced approximately 1 billion kilograms of shelled nuts in 2017. Almond hulls (AH) are a regional by-product feedstuff fed predominantly to dairy cattle in California. A 2012 study surveyed 40 dairy farms in California and found that 39 out of 104 total mixed rations contained AH, with a mean daily feeding rate of 1.45 kg/cow. In 2017, approximately 2 billion kilograms of AH was produced. At a feeding rate of 1.45 kg/cow daily, even if all 1.7 million lactating cows in California are consuming AH, there will be a surplus of AH on the market as the approximately 130,000 nonbearing hectares come into nut production. Therefore, the potential of feeding varying amounts of AH to lactating dairy cows was investigated using 12 Holstein cows with 4 primiparous and 8 multiparous cows. The dietary treatments were 4 total mixed rations containing 0, 7, 13, or 20% AH. The AH used contained 12.8% crude fiber (as-is basis), which was below the 15% legal limit set by state feed regulations. Diets were formulated so that as the inclusion rate of AH increased, the amount of steam-flaked corn and soyhull pellets decreased and soybean meal inclusion increased. Experimental design was a replicated 4 × 4 Latin square. Diet had a cubic effect on actual milk yield, energy-corrected milk yield, and dry matter intake, with the 7% AH diet having the highest values and the 13% AH diet having the lowest. The percent and yield of total solids and the yields of lactose and fat did not differ with diet, but percent and yield of protein declined linearly with increased AH inclusion, and fat percent increased linearly. Apparent total-tract digestibilities of dry matter and organic matter were higher with the inclusion of AH in the diet. Total percentage of the day spent ruminating increased linearly with higher amounts of AH. Overall, this work demonstrated that AH can be fed at varying amounts, up to 20% of the diet, to lactating dairy cows to support high levels of milk production and that increasing amounts of AH (up to 20%) in the diet could lead to improved digestibility and milk fat percentage but decreased milk protein production.

Disruption of endoplasmic reticulum (ER) homeostasis, a condition termed "ER stress," contributes to the development of liver injury in nonruminants. Because liver injury is a prominent pathological feature associated with overproduction of ketone bodies in dairy cows with ketosis, understanding the ER stress state and its functional consequences on liver injury is of particular interest. Here, 30 multiparous cows (within 3 wk postpartum) classified based on blood β-hydroxybutyrate (BHB) as healthy (n = 15, BHB0.6 mM) or clinically ketotic (n = 15, BHB3.0 mM) were used. Compared with healthy cows, ketotic cows had greater levels of serum fatty acids and activities of serum aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, γ-glutamyl transferase, and glutamate dehydrogenase but lower serum glucose. Furthermore, dairy cows with ketosis had greater protein abundance of ER stress markers in liver tissue, including protein kinase RNA-like ER kinase (PERK), inositol-requiring protein-1α (IRE1α), and cleaved activating transcription factor-6 (ATF6). Cows with ketosis also had higher mRNA levels of hepatic 78-kDa glucose-regulated protein (GRP78) and spliced X-box binding protein 1 (sXBP1). These data confirmed an enhanced ER stress state in clinically ketotic cows. To explore whether enhanced hepatic ER stress was induced by elevated ketone bodies and the possible contribution of ER stress to liver injury, in vitro experiments were then performed using isolated primary calf hepatocytes treated with incremental concentrations of BHB (0, 0.6, 1.2, 3.0, and 4.8 mM) for 12 h with or without overexpression of GRP78 (the master regulator of unfolded protein response). Phosphorylation levels of PERK and IRE1α proteins, level of cleaved ATF6 protein, and mRNA abundance of GRP78 and sXBP1 in hepatocytes increased after treatment with high (3.0 and 4.8 mM) BHB, indicating a mechanistic link between excessive BHB and enhanced hepatic ER stress. Furthermore, treatment with 3.0 and 4.8 mM BHB markedly elevated activities of aspartate aminotransferase and alanine aminotransferase in cell supernatant, indicating exacerbated hepatocyte damage after ER stress was enhanced. Overexpression of GRP78 attenuated both BHB-induced ER stress and the ensuing cellular damage, suggesting that hepatocyte damage caused by excessive BHB can be mediated via enhanced ER stress. Overall, the present study revealed that ER stress may exacerbate liver injury development in clinically ketotic cows, underscoring the biological relevance of this pathway in the context of liver injury.

This study approaches the interrelation patterns between composition of milk and whey, curd yield, chromaticity, syneresis, and technological quality of Manchega sheep milk using multivariate factor analysis. In addition, the effect of the main husbandry components (flock, prolificacy, season of the year, stage of lactation, and parity) on the common latent factors that define the pattern of variation of Manchega milk was assessed. For this purpose, 1,200 individual Manchega ewe milk samples from 4 different flocks registered under the Protected Designation of Origin Queso Manchego were analyzed (50 ewes/flock). Samples were collected in 2 different seasons of the year (spring and autumn) and at 3 time points per season: early, mid-, and late lactation. The obtained results suggested that curd yield mainly depends on milk composition, and the retention of water in the curd is related to coagulation traits. Thus, composition and moisture content could be useful indicators to assess the efficiency and quality of milk intended for cheesemaking, regardless of the analysis of coagulation properties. Finally, in terms of husbandry, a direct effect of flock and stage of lactation was observed on all analyzed factors, with a lower influence of season and parity.

The aim of this study was to explore the effect of colostrum feeding time on the ileal microbiome of neonatal calves. In this study, 22 male Holstein calves were randomly assigned to different colostrum feeding time treatments: after birth (at 45 min, n = 7); at 6 h after birth (n = 8); and at 12 h after birth (TRT12h; n = 7). At 51 h after birth, calves were killed and ileum digesta was collected for microbiome analysis using shotgun metagenomic sequencing. Bacteria, archaea, eukaryotes, and viruses were identified from the ileum microbiome. For the bacteriome, Firmicutes and Proteobacteria were the predominant phyla, and Escherichia, Streptococcus, Lactobacillus were the 3 most abundant genera. For the archaeal community, Euryarchaeota and Crenarchaeota were the 2 major phyla, and Methanosarcina, Methanobrevibacter, and Methanocorpusculum were the 3 most abundant genera. In total, 116 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways were identified from the ileal microbiome, with "biosynthesis of vancomycin group antibiotics," "biosynthesis of ansamycins," "valine, leucine, and isoleucine biosynthesis," "ribosome," and "d-alanine metabolism" as the top 5 functions. When the ileal microbiomes were compared among the 3 treatments, the relative abundance of Enterococcus was higher in TRT12h calves, suggesting that calves may have a higher abundance of opportunistic pathogens when the feeding of colostrum is delayed for 12 h. Moreover, among all KEGG pathways, the enriched "taurine and hypotaurine metabolism" (KO00430) pathway was identified in the ileal microbiome of TRT12h calves; however, future studies are needed to understand the effect on the host. Additionally, 2 distinct ileal microbial profiles were identified across all samples, indicating that that host factors may play a significant role in driving varied microbiome changes in response to colostrum feeding time. Whether such microbiome shifts affect long-term gut function and calf performance warrants future studies.

The objective of this systems-scale study was to investigate grazing season timeframes on pasture and milk production and on milk processability of dairy systems with compact spring-calving dairy cows grazing white clover (Trifolium repens L.) based grassland. Fifty-four primiparous and multiparous Holstein-Friesian dairy cows were used in a one-factor study with 3 systems (n = 18) and repeated over 2 yr (2008/09 and 2009/10). The 3 systems were: early spring calving with annual fertilizer N input of 100 kg·ha-1 applied in spring (ES100N; 2.1 cows·ha-1; grazing February to November), early spring calving without fertilizer N (ES0N; 1.6 cows·ha-1; grazing February to November) and late spring calving without fertilizer N (LS0N; 1.53 cows·ha-1; grazing April to January). Annual pasture production was affected by an interaction between grazing system and year: Mean annual pasture yields for 2008 and 2009 were ES100N; 10.35 and 9.88, ES0N; 8.88 and 8.63, LS0N; 9.18 and 10.31 t of dry matter (DM)·ha-1 (SEM 0.39). LS0N had higher pasture DM yield in 2009 due to higher clover DM production and biological N fixation compared with the other systems. Clover stolon and root mass in the following February was correlated with stolon and root mass in the previous November with 64% of stolon mass present on LS0N in February (R2 = 0.84). There were no detectable differences in per-lactation milk yield (6,335 kg·cow-1), fat, protein and lactose yields (271, 226, 297 kg·cow-1, respectively), cow liveweight (585 kg) or body condition score (3.02). Although winter grazing favored subsequent clover DM production, biological N fixation and pasture DM production, delaying calving date in spring and extending lactation into the following winter led to inefficient use of this pasture by the grazing herd and lowered the quality of late-lactation milk for processing purposes. Hence, a mean calving date in mid- to late-February is recommended for zero-fertilizer N input clover-based grassland.

Escherichia coli O157:H7 (E. coli O157:H7), one of the most widespread foodborne pathogens, can cause a series of diseases and even lead to death. In this study, a highly sensitive method was developed by combining aptamer-exonuclease III (Exo III)-assisted amplification with lateral flow assay (LFA) based on gold nanoparticles (AuNP). The compound of single-stranded (ss) DNA-anti-E. coli O157:H7 aptamer (ssDNA-aptamer) was formed by hybridization between designed target ssDNA and aptamer. When E. coli O157:H7 was present, target bacteria were bound with the aptamer, and the free target ssDNA was hybridized with the probes of the designed hairpin (HP) structure. Exo III digests the 3' double-stranded blunt end of the complex and releases the enzyme product. Because the remaining sequence of the HP of the designed enzyme product was the same as the target ssDNA sequence, the target ssDNA could be amplified. Finally, the enhanced target ssDNA was combined with AuNP-LFA to achieve visual detection of E. coli O157:H7. The quantitative ability of this platform for E. coli O157:H7 was 7.6 × 101 cfu/mL in pure culture, and the detection limit in milk was 8.35 × 102 cfu/mL. This LFA was highly specific to E. coli O157:H7, and the time for detection of E. coli O157:H7 in milk was 4 h. Hence, this system has important application prospects in the detection of pathogenic bacteria in dairy products.

Establishment of a vascular system within the corpus luteum (CL) is critical for progesterone (P4) secretion. Measurement of luteal blood flow (LBF) may be a feasible way to determine luteolysis induced with cloprostenol sodium (CLO). Our overall objective was to establish timelines to assess luteolysis via Doppler ultrasonography. Estrous cycles were synchronized in 11- to 12-mo-old Holstein heifers (n = 37). Heifers were injected CLO at a random stage of the estrous cycle 12 d from treatment. Gonadotropin-releasing hormone (GnRH) was administered 2 (d −10) and 8 d (d −4) after the initial CLO. This satisfied the study objective of inducing simultaneous presence of CL at d 4 of development (D4 CL) and CL at d 10 of development (D10 CL) on the day of treatment with different CLO doses (hereafter referred to as d 0). Heifers were randomly assigned to 1 of 5 treatments on d 0: negative control (NC) consisting of no treatment with CLO (n = 8); a quarter dose of CLO (0.125 mg; n = 8); half dose of CLO (0.25 mg; n = 8); full dose of CLO (0.5 mg; n = 8); or positive control (PC) consisting of 4 doses of 0.5 mg of CLO at 24-h intervals starting at d 0 (n = 5). Data collection was performed at d 0 (before and 1 h after treatment) and 2, 4, 6, and 8 d following treatment, to determine luteal volume (LV), LBF, and circulating concentrations of P4. Both NC and PC were efficient in mimicking physiological scenarios that occur during normal luteal development and luteolysis. Heifers that received PC had complete LBF disappearance of both D4 and D10 CL between d 2 and 4 after the first of 4 CLO treatments given 24 h apart (average 4.0 ± 0.0 and 3.2 ± 0.7 d, respectively). Complete LBF disappearance was used as a luteolysis marker. Treatment with different doses of CLO did not impair luteal development of the D4 CL. However, concurrent complete LBF disappearance for D10 CL in heifers treated with half (5/8 heifers) and full doses of CLO (8/8 heifers) resulted in less LBF in the half dose, and less LV and LBF in the full-dose treatment, in D4 CL at d 8 post-treatment, compared with NC. Treatment with various doses of CLO induced an acute increase in LBF 1 h after treatment, regardless of dose in D10 but not in D4 CL. We found a lack of dose response in LV reduction of D4 and D10 CL. Interestingly, LV of the D10 CL decreased in untreated NC between d 0 and 8 after treatment (d 10–18 of luteal development). Assessment with color Doppler ultrasound was sensitive enough to identify dose-response patterns in Holstein heifers (absence, partial, or complete luteolysis) following various doses of CLO. Variability in time to complete LBF disappearance of mature D10 CL following a full dose of CLO limits the use of Doppler ultrasonography to detect luteolysis at a single time point following treatment.

Our objective was to measure whey protein removal percentage from separated sweet whey using spiral-wound (SW) polymeric microfiltration (MF) membranes using a 3-stage, 3× process at 50°C and to compare the performance of polymeric membranes with ceramic membranes. Pasteurized, separated Cheddar cheese whey (1,080 kg) was microfiltered using a polymeric 0.3-μm polyvinylidene (PVDF) fluoride SW membrane and a 3×, 3-stage MF process. Cheese making and whey processing were replicated 3 times. There was no detectable level of lactoferrin and no intact α- or β-casein detected in the MF permeate from the 0.3-μm SW PVDF membranes used in this study. We found BSA and IgG in both the retentate and permeate. The β-lactoglobulin (β-LG) and α-lactalbumin (α-LA) partitioned between retentate and permeate, but β-LG passage through the membrane was retarded more than α-LA because the ratio of β-LG to α-LA was higher in the MF retentate than either in the sweet whey feed or the MF permeate. About 69% of the crude protein present in the pasteurized separated sweet whey was removed using a 3×, 3-stage, 0.3-μm SW PVDF MF process at 50°C compared with 0.1-μm ceramic graded permeability MF that removed about 85% of crude protein from sweet whey. The polymeric SW membranes used in this study achieve approximately 20% lower yield of whey protein isolate (WPI) and a 50% higher yield of whey protein phospholipid concentrate (WPPC) under the same MF processing conditions as ceramic MF membranes used in the comparison study. Total gross revenue from the sale of WPI plus WPPC produced with polymeric versus ceramic membranes is influenced by both the absolute market price for each product and the ratio of market price of these 2 products. The combination of the market price of WPPC versus WPI and the influence of difference in yield of WPPC and WPI produced with polymeric versus ceramic membranes yielded a price ratio of WPPC versus WPI of 0.556 as the cross over point that determined which membrane type achieves higher total gross revenue return from production of these 2 products from separated sweet whey. A complete economic engineering study comparison of the WPI and WPPC manufacturing costs for polymeric versus ceramic MF membranes is needed to determine the effect of membrane material selection on long-term processing costs, which will affect net revenue and profit when the same quantity of sweet whey is processed under various market price conditions.

Yak milk is an essential and predominant food resource for Tibetan people for subsistence purposes and to combat altitude-induced challenges. Due to its unique qualities, yak milk has recently been gaining broader attention from consumers across China as well in other parts of the world. One of the key characteristics of yak milk is the protein content, which is about 40 to 60% higher than that of native bovine milk. In this work, a sensitive and reproducible high-throughput analytical method was developed employing both ultra high-performance liquid chromatography Orbitrap (Thermo Fisher Scientific) high-resolution accurate mass spectroscopy (UHPLC-HRAM-MS) and UHPLC coupled with triple quadrupole tandem MS (UHPLC-QqQ-MS) to simultaneously analyze 8 milk proteins. A total of 15 Maiwa yak milk samples and 15 bovine milk samples were qualitatively and quantitatively analyzed using targeted proteomics and compared for α-lactalbumin, β-lactoglobulin, αS1-casein, αS2-casein, β-casein, κ-casein, lactoferrin, and osteopontin. Peptides of β-lactoglobulin were used to specifically distinguish yak and bovine milk. The results showed that this novel detection method could quantitatively detect these major and minor milk proteins with >0.99 linear correlation coefficient and a recovery rate between 90 and 120%, with relative standard deviations typically less than 10%. The data revealed that yak milk not only had higher overall milk protein content than bovine milk but higher lactoferrin and osteopontin contents as well. The lactoferrin content of yak milk was about 30% higher than that of bovine milk, and the osteopontin content of yak milk was nearly twice that of bovine milk. The application of this method demonstrates that UHPLC-HRAM-MS and UHPLC-QqQ-MS are suitable for high-throughput qualitative and quantitative analysis of major and minor proteins of yak and bovine milk.

Effective testing tools for Escherichia coli O157:H7 can prevent outbreaks of foodborne illness. In this paper, a smartphone-based colorimetric aptasensor was developed using functionalized gold nanoparticles (GNP) and multi-walled carbon nanotubes (MWCNT) for monitoring E. coli O157:H7 in milk. The maximum absorption peak of GNP bonded with aptamer (Apt) generated evident transformation from 518 to 524 nm. The excess GNP-Apt was removed by functionalized MWCNT magnetized with carbonyl iron powder (CIP) and hybridized with a DNA probe, whereas the GNP-Apt immobilized on E. coli O157:H7 remained in the system. In the presence of a high-salt solution, the GNP-Apt that captured E. coli O157:H7 remained red, but the free GNP-Apt aggregated and appeared blue. The chromogenic results were analyzed by a smartphone-based colorimetric device that was fabricated using acrylic plates, a light-emitting diode, and a mobile power pack. To our knowledge, this was the first attempt to use a smartphone-based colorimetric aptasensor employing the capture of GNP-Apt coupled with separation of MWCNT@CIP probe to detect E. coli O157:H7. The aptasensor exhibited good reproducibility and no cross-reaction for other bacteria. A concentration of 8.43 × 103 cfu/mL of E. coli O157:H7 could be tested in pure culture, and 5.24 × 102 cfu/mL of E. coli O157:H7 could be detected in artificially contaminated milk after 1 h of incubation. Therefore, the smartphone-based colorimetric aptasensor was an efficient tool for the detection of E. coli O157:H7 in milk.

Predicting dry matter intake (DMI) and feed efficiency by leveraging the use of data streams available on farm could aid efforts to improve the feed efficiency of dairy cattle. Residual feed intake (RFI) is the difference between predicted and observed feed intake after accounting for body size, body weight change, and milk production, making it a valuable metric for feed efficiency research. Our objective was to develop and evaluate DMI and RFI prediction models using multiple linear regression (MLR), partial least squares regression, artificial neural networks, and stacked ensembles using different combinations of cow descriptive, performance, sensor-derived behavioral (SMARTBOW; Zoetis), and blood metabolite data. Data were collected from mid-lactation Holstein cows (n = 124; 102 multiparous, 22 primiparous) split equally between 2 replicates of 45-d duration with ad libitum access to feed. Within each predictive approach, 4 data streams were added in sequence: dataset M (week of lactation, parity, milk yield, and milk components), dataset MB (dataset M plus body condition score and metabolic body weight), dataset MBS (dataset MB plus sensor-derived behavioral variables), and dataset MBSP (dataset MBS plus physiological blood metabolites). The combination of 4 datasets and 4 analytical approaches resulted in 16 analyses of DMI and RFI, using variables averaged within cow across the study period. Additional models using weekly averaged data within cow and study were built using all predictive approaches for datasets M, MB, and MBS. Model performance was assessed using the coefficient of determination, concordance correlation coefficient, and root mean square error of prediction. Predictive models of DMI performed similarly across all approaches, and models using dataset MBS had the greatest model performance. The best approach–dataset combination was MLR–dataset MBS, although several models performed similarly. Weekly DMI models had the greatest performance with MLR and partial least squares regression approaches. Dataset MBS models had incrementally better performance than datasets MB and M. Within each approach–dataset combination, models with DMI averaged over the study period had slightly greater model performance than DMI averaged weekly. Predictive performance of all RFI models was poor, but slight improvements when using MLR applied to dataset MBS suggest that rumination and activity behaviors may explain some of the variation in RFI. Overall, similar performance of MLR, compared with machine learning techniques, indicates MLR may be sufficient to predict DMI. The improvement in model performance with each additional data stream supports the idea of integrating data streams to improve model predictions and farm management decisions.

Selecting for lower methane (CH4) emitting animals is one of the best approaches to reduce CH4 given that genetic progress is permanent and cumulative over generations. As genetic selection requires a large number of animals with records and few countries actively record CH4, combining data from different countries could help to expedite accurate genetic parameters for CH4 traits and build a future genomic reference population. Additionally, if we want to include CH4 in the breeding goal, it is important to know the genetic correlations of CH4 traits with other economically important traits. Therefore, the aim of this study was first to estimate genetic parameters of 7 suggested methane traits, as well as genetic correlations between methane traits and production, maintenance, and efficiency traits using a multicountry database. The second aim was to estimate genetic correlations within parities and stages of lactation for CH4. The third aim was to evaluate the expected response of economically important traits by including CH4 traits in the breeding goal. A total of 15,320 methane production (MeP, g/d) records from 2,990 cows belonging to 4 countries (Canada, Australia, Switzerland, and Denmark) were analyzed. Records on dry matter intake (DMI), body weight (BW), body condition score, and milk yield (MY) were also available. Additional traits such as methane yield (MeY; g/kg DMI), methane intensity (MeI; g/kg energy-corrected milk), a genetic standardized methane production, and 3 definitions of residual methane production (g/d), residual feed intake, metabolic BW (MBW), BW change, and energy-corrected milk were calculated. The estimated heritability of MeP was 0.21, whereas heritability estimates for MeY and MeI were 0.30 and 0.38, and for the residual methane traits heritability ranged from 0.13 to 0.16. Genetic correlations between different methane traits were moderate to high (0.41 to 0.97). Genetic correlations between MeP and economically important traits ranged from 0.29 (MY) to 0.65 (BW and MBW), being 0.41 for DMI. Selection index calculations showed that residual methane had the most potential for inclusion in the breeding goal when compared with MeP, MeY, and MeI, as residual methane allows for selection of low methane emitting animals without compromising other economically important traits. Inclusion of residual feed intake in the breeding goal could further reduce methane, as the correlation with residual methane is moderate and elicits a favorable correlated response. Adding a negative economic value for methane could facilitate a substantial reduction in methane emissions while maintaining an increase in milk production.

The objective of the study was to develop a static empirical model for the estimation of net energy content of compound feeds in a dynamic feeding system using net energy for lactation at 20 kg of dry matter intake/d (NEL20) values calculated by the Nordic Feed Evaluation System (NorFor) model. In the NorFor system, NEL20 is a standardized value used to describe net energy content of feeds. The static model would allow prediction of the net energy value of compound feeds without access to the input data needed for the dynamic models. Our hypothesis was that NEL20 values of compound feeds can be predicted using organic matter digestibility (in vitro) and chemical components of the compound feeds as input variables. For this, 75 compound feeds and their 108 associated ingredients were collected across Scandinavia for model development. The proposed best model for prediction of compound feed NEL20 included crude fat, neutral detergent fiber, digestible organic matter measured in vitro, and crude protein (urea corrected) as independent variables. Lack of additivity of chemical components between values analyzed directly in the compound feed and values calculated by the weighted sum of ingredients was detected as the main source of error in the model, emphasizing the importance of accurate chemical analysis and sampling practices. Results from practical use of the model show that it may be a valuable tool that could be used by several actors in the feeding sector using the NorFor system. Feed manufacturers could use it to monitor the net energy content in their final product, and farmers could use it to check the net energy content of the purchased compound feed. However, validation of this model against an independent set of samples is lacking in this study and its prediction performance should be further evaluated. The model will need recalibration if the feed parameters used in the dynamic model for the estimation of reference values change, as this would not be reflected in the predicted values of the created model.

The objective of this study was to examine the effects of including a molasses-based liquid feed on sorting of a mixed diet of starter and hay, and to compare the nutrient composition of the ration consumed by calves offered hay and starter according to different feed presentations. Holstein bull and heifer calves were exposed for the first 8 wk of life to 1 of 3 feed presentation treatments: (1) starter and hay provided in separate buckets (n = 15), (2) a mixed diet of 80% starter and 20% hay (n = 16), or (3) a mixed diet containing the same ratio of starter and hay, with inclusion of 10% (as-is basis) liquid molasses (n = 14). Calves received 8 L/d of pasteurized waste milk and were weaned over a 10-d period beginning at 42 d of age. Intake was recorded daily and calves were weighed weekly. Fresh and refused feed were sampled on 3 consecutive days during wk 4 and 6 of the preweaning period and wk 8 immediately postweaning. Sorting was assessed through nutrient analysis of the feed samples, with additional sorting indices reflective of sorting for starter and hay components calculated based on weighted averages of individual nutrient intakes as a percent of predicated intakes. Dry matter intake and average daily gain did not differ among treatments. During the preweaning period, addition of molasses to the mixed diet did not affect the extent of feed sorting, with calves provided both mixed diets sorting, on average, in favor of starter and against hay. Postweaning, addition of molasses tended to reduce the extent of feed sorting, although calves on both treatments persisted in sorting for starter and against hay. Across treatments, there was an association between the pre- and postweaning extent of feed sorting, suggesting a degree of individual consistency in sorting behavior. Calves provided starter and hay separately consumed a diet consisting of approximately 18 to 19% hay preweaning and 14% hay postweaning, and generally had greater fiber intake and lower intake of nonfiber carbohydrates compared with calves provided a mixed diet. These results indicate that addition of molasses slightly reduced sorting during the postweaning period only. On average, calves provided both mixed diets sorted their feed in favor of starter and against hay, which mirrored dietary selection exhibited by calves offered hay and starter separately.

The association between dry period length (DPL) and time to culling and pregnancy in the subsequent lactation may be important for the economically optimal length of the dry period. Therefore, this study aimed to (1) quantify the association between DPL and hazard of culling and pregnancy in the subsequent lactation; (2) develop continuous functions of DPL for the hazard ratios of culling and pregnancy; and (3) investigate the effect of a cause-specific hazards model and a subdistribution model to analyze competing events. The data used in this observational cohort study were from dairy herd improvement milk test lactation records from 40 states in the United States. After edits, there remained 1,108,515 records from 6,730 herds with the last days dry in 2014 or 2015. The records from 2 adjacent lactations (current, subsequent) were concatenated with the DPL of interest, 21 to 100 d, in between both lactations. We defined 8 DPL categories of 10 d each. Kaplan-Meier survival curves were used to show associations between DPL and time to culling or pregnancy for 3 lactation groups: lactation 1 and 2, lactation 2 and 3, and lactation 3 and greater. To control for confounding factors in Cox proportional models, we included 6 current lactation covariates and 3 time-dependent variables in the survival models. Hazard ratios of culling were estimated for 4 days in milk (DIM) categories from 1 to 450 DIM. Hazard ratios of pregnancy were estimated for 3 DIM categories from 61 to 300 DIM. Competing risk analysis of 8 disposal codes (i.e., farmer reported reasons) for culling and the culling event for pregnancy were conducted by a cause-specific hazards model and a subdistribution model. Hazard ratios were also estimated as quadratic polynomials of DPL. Compared with the reference DPL category of 51 to 60 d, hazard ratios of culling and pregnancy of the other 7 DPL categories ranged between 0.70 and 1.49, and 0.93 and 1.15, respectively. Short DPL were associated with lower risk of culling in the early lactation but not over the entire lactation. Short DPL were associated with greater hazard of pregnancy. Trends in hazard ratios over the ranges of the 8 DPL categories were not always consistent. Competing risk analysis with both models provided little differences in hazard ratios of culling and pregnancy. In conclusion, variations in DPL were associated with meaningful differences in the hazard ratios for culling and pregnancy and minor differences in the relative frequency of disposal codes. Subdistribution hazards models produced hazard ratios similar to cause-specific hazard models. The quadratic polynomials may be useful for decision support on customization of DPL for individual cows.

The aims of this study were to evaluate histopathologic changes during the transition period, describe the histopathological features of the metabotypes identified in Part I ( Scharen et al., 2021b ), and investigate effects of a metaphylactic treatment with butaphosphan and cyanocobalamin (BCC) on the liver parenchyma. Eighty German Holstein cows (mean 305-d production: 10,957 kg, range: 6,480–15,193 kg; mean lactation number: 3.9, range: 2–9) from a commercial dairy farm in Saxony, Germany, were enrolled in a randomized, prospective, triple-blinded study. Two groups received a treatment with BCC (5 or 10 mL/100 kg of body weight 10% butaphosphan and 0.005% cyanocobalamin, Catosal, Bayer Animal Health, n = 20 each) and one group a placebo treatment (NaCl 0.9%, n = 40). Liver biopsy specimens were collected 14 d antepartum (AP) and 7, 28, and 42 d postpartum (PP), routinely processed for histologic examination, and stained with hematoxylin and eosin, Sudan III, periodic acid-Schiff, and picrosirius red stains. The sections were assessed for fat and glycogen content and degenerative, inflammatory, fibrotic, and proliferative changes. The statistical analysis included the effects of the sampling day, the lactation number, the treatment, and the metabotype (A = medium, B = minor, C = large alterations in the liver metabolome profile between AP and PP status). There was mild to moderate fat infiltration in the liver of 37% of cows in the last 2 wk AP, and moderate to severe fat infiltration in 66% of cows in the first days PP. The degree of fat infiltration increased from 2 wk AP until the end of the first week PP, and then decreased until the end of the study period, at which time about 25% of cows had moderate to severe fatty infiltration. Lipidosis was positively correlated with the severity of liver cell degeneration, and negatively correlated with the degree of glycogen deposits. Complete glycogen depletion of hepatocytes was not observed in cows, even in the presence of severe hepatic lipidosis. Moderate to severe lymphocytic hepatitis was seen in 39% of cows throughout the study period, and cows with lactation numbers 5 or greater had perisinusoidal fibrosis more often than younger cows. Severe fibrosis and cirrhosis of the liver did not occur. Metabotype B animals exhibited a higher chance of fatty infiltration, lower glycogen storage, and perisinusoidal fibrosis and for this metabotype positive correlations were calculated between increased fat deposition in the liver and marked glycogen depletion, and increased degenerative, inflammatory, fibrotic, and proliferative changes of hepatic tissue. For the treatment with BCC, no significant effect was observed. In summary, during the transition period, the liver of dairy cows is characterized by fat accumulation and glycogen depletion and histologic signs of hepatitis and hepatocyte degeneration. These histomorphologic changes were accentuated in animals exhibiting little alterations in their liver metabolome profile across the transition period (metabotype B) and support the assumption of a decreased grass silage quality as a causative factor.

Fodder beet (FB) is a source of readily fermentable carbohydrate that can mitigate early spring herbage deficits and correct the negative energy balance experienced during early lactation in pastoral dairy systems of New Zealand. However, the low-fiber and high-soluble carbohydrate content of both FB bulb and spring herbage are factors that promote subacute ruminal acidosis, impairing rumen function and limiting the marginal milk production response to supplement. In a crossover experiment, 8 Holstein Friesian × Jersey early-lactation dairy cows were used to test the effect of supplementing 16 kg of dry matter (DM) of a grazed perennial ryegrass herbage with 6 kg of DM/d of FB bulb (FBH) versus herbage only (HO) on changes in rumen function and grazing behavior. Following 20 d of adaptation to diets, DM disappearance (%) of FB bulb (FBH cows only) and herbage were measured in sacco, separately. Cows were fasted overnight, and the ruminal contents were bailed the following morning (~0930 h) again to determine the pool size of volatile fatty acids, ammonia, and particle size of digesta, as well as to estimate the rate of ruminal outflow and degradation of neutral detergent fiber. The FBH diet did not alter DM intake, milk yield, or milk solid (fat + protein) production compared with HO. Supplementation of herbage with FB reduced ruminal pH compared with HO between ~0800 h and 1300 h each day. During each period, 1 cow experienced severe subacute ruminal acidosis (pH 180 min/d) during final adaptation to the target FB allocation. The FBH diet reduced the ruminal pool of acetate and ammonia, but increased the ruminal pool of butyrate and lactate compared with HO. When fed FB, rumination and grazing time increased and grazing intensity declined compared with cows fed HO. Despite increased rumination, the comminution of large particles declined 28% between the first and second rumen bailing when cows were fed FB, and in sacco DM disappearance of perennial ryegrass declined 18% compared with cows fed HO. These results indicate that grazing dairy cows supplemented with FB (40% of daily intake) increase rumination and mastication intensity to counteract reduced ruminal degradation of ryegrass herbage due to low ruminal fluid pH.

Probiotics can ferment nondigestible carbohydrates and produce short-chain fatty acids (SCFA; acetate, propionate, and butyrate) in the human colon. In this study, the levels of SCFA were determined in the following yogurts fermented with different combinations of probiotics: (1) cocultures of Streptococcus thermophilus and Lactobacillus bulgaricus (control, C); (2) S. thermophilus, L. bulgaricus, and Bifidobacterium bifidum (C-Bb); (3) S. thermophilus, L. bulgaricus, and Lactobacillus acidophilus (C-La); and (4) S. thermophilus, L. bulgaricus, and Lactobacillus gasseri (C-Lg). Results showed that the acetate levels were significantly higher in C-Bb, C-La, and C-Lg yogurts than in C yogurt. Fermentation and physicochemical characteristics of all yogurts were identical. Treatment of mucus-secreting colon epithelial cells (HT29-MTX) with C-Bb, C-La, and C-Lg yogurt supernatants resulted in an increase in the expression of MUC2 and CDX2 and the production of mucin proteins. The adhesion of probiotics onto HT29-MTX cells increased following treatment with C-Bb, C-La, and C-Lg yogurt supernatants. Our data suggest that a yogurt diet rich in acetate improves the protective function of the intestinal epithelium.

Novel antihypercholesterolemic bioactive peptides (BAP) from peptic camel whey protein hydrolysates (CWPH) were generated at different time, temperature, and enzyme concentration (%). Hydrolysates showed higher pancreatic lipase- (PL; except 3 CWPH) and cholesterol esterase (CE)-inhibiting potential, as depicted by lower half-maximal inhibitory concentration values (IC50 values) compared with nonhydrolyzed camel whey proteins (CWP). Peptide sequencing and in silico data depicted that most BAP from CWPH could bind active site of PL, whereas as only 3 peptides could bind the active site of CE. Based on higher number of reactive residues in the BAP and greater number of substrate binding sites, FCCLGPVPP was identified as a potential CE-inhibitory peptide, and PAGNFLPPVAAAPVM, MLPLMLPFTMGY, and LRFPL were identified as PL inhibitors. Molecular docking of selected peptides showed hydrophilic and hydrophobic interactions between peptides and target enzymes. Thus, peptides derived from CWPH warrant further investigation as potential candidates for adjunct therapy for hypercholesterolemia.

To study the key aroma components and flavor profile differences of Cheddar cheese with different maturity and from different countries, the flavor components of 25 imported commercial Cheddar cheese samples in the China market were determined by gas chromatography-mass spectrometry. The quality and quantity of 40 flavor compounds were analyzed by gas chromatography-olfactometry among 71 aroma compounds determined by gas chromatography-mass spectrometry. Combined with odor activity value calculation, principal component analysis (PCA) was conducted to analyze the relationship among 26 flavor compounds with odor activity values >1 and the maturity of Cheddar cheese. The PCA results showed significant differences between the group of mild Cheddar cheese and the groups of medium Cheddar cheese and mature Cheddar cheese, and no significant differences were observed between medium Cheddar cheese and mature Cheddar cheese. According to the results of PCA and consumers' preference test, representative Cheddar cheese samples with different ripening times were selected for the flavor profile analysis. Partial least squares regression analysis was conducted to obtain the relationship between sensory properties and flavor compounds of different Cheddar cheeses. Based on partial least squares regression analysis, 1-octen-3-one, hexanal, acetic acid, 3-methylindole, and acetoin were positively correlated with milky, sour, and yogurt of mild Cheddar cheese. Dimethyl trisulfide, phenylacetaldehyde, ethyl caproate, octanoic acid, and furaneol and other compounds were positively correlated with fruity, caramel, rancid, and nutty notes of the medium and mature Cheddar cheeses.

The detection of coliforms in young cheese is a potential indication of undesirable microbial growth within the processing environment. The aim of this study was to investigate sources and conditions that lead to the intermittent detection of coliforms (1-3 log cfu/g) in young Cheddar cheese at a single commercial facility. Analysis of historical production data, in combination with iterative investigative sampling events, was performed to determine coliform levels in milk, whey, curd, and surfaces at the beginning, middle, and end of the production day. After sanitation, conveyor belt pieces from the draining and matting conveyor (DMC) were collected and evaluated for bacterial survivors using culture-based methods and scanning electron microscopy. Production data analysis indicated that cheese produced later in the production day (≥16 h) was significantly more likely to test positive for coliforms than cheese made earlier in the production day (

Regular veterinary visits to improve herd health and production management (HHPM) are important management components on many dairy cattle farms. These visits provide opportunities for constructive conversations between veterinarians and farmers and for shifting management from a reactionary approach to proactively optimizing health and welfare. However, little is known about the structure of HHPM farm visits and to what extent veterinarians provide assistance beyond purely technical services. Therefore, our aims in this cross-sectional study were to describe HHPM farm visit structure, determine which dairy-specific topics were discussed, and assess whether the focus of the visits aligned with farmers' priorities. Veterinary practitioners (n = 14) were recruited to record audio and video of regularly scheduled HHPM farm visits (n = 70) using an action camera attached to their chest or head. A questionnaire was distributed to farmers containing closed- and open-ended questions to assess their goals and perceptions related to farm management and HHPM farm visits. Descriptive statistics and negative binomial and Poisson regression models were used to study dairy-specific topics initiated by the farmer or veterinarian during various activities. A mean of 51% of the visit duration was dedicated to transrectal pregnancy and fertility diagnostics, and a considerable amount of time (30%) was spent on visit preparation, transitions between tasks, and leaving. A total of 488 discussions were initiated by either the veterinarian (55%) or the farmer (45%). Mean length of discussions was 2 min, and only 17% of the HHPM visit duration was spent discussing dairy-specific topics. Veterinarians initiated 62% of their discussions about herd issues, whereas farmer-initiated discussions revolved around herd health in 39% of the discussions. Discussion topics most frequently raised by participants included fertility, udder health, calf health and management, and transition diseases. Consistently, farmers' answers to a rank question regarding their main HHPM farm visit goals indicated that their priorities were to have transrectal pregnancy and fertility diagnostics performed and to improve herd fertility and general herd health. Answers to an open-ended question revealed that additional aims of many farmers were to receive information, have questions answered, and identify and discuss problems. A farmer's belief that HHPM farm visits were "absolutely" tailored toward his or her goals was positively associated with number of discussions during the visit and their conviction that they "always" voiced their wishes and needs to the veterinarian. Opportunities to broaden the focus of HHPM farm visits and improve communication between farmers and veterinarians should be identified and veterinarians should be trained accordingly, which would increase veterinarians' ability to add value during HHPM farm visits.

There has been a growing interest in cofermentation of starter cultures with probiotics in milk. In this study, we analyzed the effects of adding the single probiotics Lactobacillus casei Zhang (Zhang) or Bifidobacterium animalis ssp. lactis Probio-M8 (M8) or a combination of Zhang and M8 to starter cultures on volatile and nonvolatile metabolomic profiles after 14 d of storage at 4°C and compared using a liquid chromatography-tandem mass spectrometry (LC-MS) and GC-MS-based metabolomics approach. Principal component analysis, heatmap plots, and Spearman correlation results showed that Zhang alone had a greater effect on volatile and nonvolatile metabolomic profiles than M8 alone. The combination of Zhang and M8 had additive effects on the production of metabolites. For volatile metabolites, the levels of acetaldehyde, diacetyl, acetoin, and acetic acid were higher for the combination of Zhang and M8 compared with either single probiotic culture. Significantly increased nonvolatile components induced by adding Zhang were identified were enriched in the galactose, amino- and nucleotide sugar, fructose and mannose, purine, phenylalanine metabolism, and arginine biosynthesis pathways. The metabolism and biosynthesis of starch, sucrose, tyrosine, galactose metabolism, and aminoacyl-tRNA biosynthesis were significantly upregulated by adding the combination of Zhang and M8. This work provides a detailed insight into different effects of Zhang and M8 used alone or in combination on the volatile and nonvolatile metabolomic profiles of yogurts.

The present research aimed at developing practical and feasible models to optimize feeding adequacy to maintain desired rumen pH conditions and prevent subacute ruminal acidosis (SARA) in dairy cows. We conducted 2 meta-analyses, one using data from recent published literatures (study 1) to investigate the prediction of SARA based on nutrient components and dietary physical and chemical characteristics, and another using internal data of our 5 different published experiments (study 2) to obtain adjustments based on cow status. The results of study 1 revealed that physically effective neutral detergent fiber inclusive of particles8 mm (peNDF8) and dietary starch [% of dry matter (DM)] were sufficient for predicting daily mean ruminal pH {y = 5.960 - (0.00781 × starch) + (0.03743 × peNDF8) - [0.00061 × (peNDF8 × peNDF8)]}. The model for time of pH suppression (5.8 for ruminal pH or6.0 for reticular pH, min/d) can be predicted with additionally including DMI (kg/d): 124.7 + (1.7007 × DMI) + (20.9270 × starch) + (0.2959 × peNDF8) - [0.0437 × (DMI × starch × peNDF8)]. As a rule of thumb, when taken separately, we propose 15 to 18% peNDF8 as a safe range for diet formulation to prevent SARA, when starch or NFC levels are within 20 to 25% and 35 to 40% ranges, respectively. At dietary starch content below 20% of DM, grain type was insignificant in affecting ruminal pH. However, increasing dietary starch contents by using corn as the sole grain source could lead to more severe drops of pH compared with using grain mix based on barley and wheat, as underlined by an interaction between starch content and grain type. Data from study 2 emphasized an increased risk of SARA for cows in the first and second lactation with lower mean pH (0.2 units) and double amounts of time at pH5.8 compared with the cows with ≥3 parities. Given that a lower ruminal pH is expected in these high-risk cows, it is advisable to keep the lower end of recommended starch (20%) and higher peNDF8 (18%) contents in the diet of these cows. Overall, the present study underlines the possibility of predicting SARA based on dietary factors including peNDF8 and starch contents, as well as DMI of the cows, which can be practically implemented for optimal diet formulation for dairy cows. With more data available, future studies should attempt to improve the predictions by including additional key dietary and cow factors in the models.