Effect of milk stasis on mammary gland involution and the microRNA profile.

The presence of an autocrine factor in milk that can trigger mammary gland involution was proposed more than 50 years ago. To provide evidences that one or more autocrine factor(s) exists, 10 multiparous cows in late lactation were quarter-milked for 7 d. Following this baseline period, the right front quarter of each cow was left unmilked while the other quarters were milked for 7 d. Before the last milking of that period, milk (mammary secretions) was collected aseptically from both front quarters. After that milking, 250 mL of the collected samples was infused in the cows' respective rear quarters. No quarters were milked for the following 7 d (milk stasis period), and then quarter milking was resumed in all quarters for the last 7 d of the experiment (remilking period). Quarter milk samples were collected during the baseline period, before the milk stasis period, and during the remilking period. These samples were used for measuring milk components and the concentration of involution markers (SCC, BSA and lactoferrin). Samples of mammary secretions were collected manually from the quarters during the milk stasis period for involution marker determination. RNA was extracted from samples collected from front quarters before the last milking before the milk stasis period for microRNA (miRNA) determination. As anticipated, the longer milk stasis period implemented for the right front quarter resulted in a more advanced involution than in the left front quarter, based on the concentration of involution markers in the mammary secretions, lower milk production recovery and changes in milk composition during the remilking period. All 3 involution marker concentrations in the mammary secretions increased in both rear quarters, but were greater in the right quarter secretions than in the left quarter secretions. Resuming milking reinitiated milk production in all quarters, but milk production recovery in the right rear quarters was less robust than that in the left rear quarters (54.3 ± 1.4% vs 61.6 ± 1.4%, respectively). Milk from the quarters infused with mammary secretions (right rear) had a lower lactose content, but a higher milk protein content and higher SCC than the quarters infused with milk. We detected a total of 359 miRNAs, 76 of which were differentially expressed in milk and mammary secretions. Expression of bta-miR-221 and bta-miR-223 were upregulated in mammary secretions 34- and 40-fold, respectively. The results of the present experiment support the contention that milk stasis leads to the accumulation of one or more factors that trigger involution. The results also indicate that milk stasis leads to changes in the miRNA profile of the milk, but whether such changes are a cause or a consequence of the involution process remains to be established.

Mechanism underlying the modulation of milk production by incomplete milking.

Mammary gland secretory activity is modulated by systemic and local factors; however, the relationship between these factors is unknown. The aim of this study was to determine how a local factor, such as incomplete milking, affects mammary epithelial cell activity, number, and responsiveness to blood prolactin (PRL). Eight cows in mid-lactation were differentially milked (i.e., their right quarters were milked incompletely at approximately 70%, and their left quarters were milked completely, twice daily for 4 wk). Throughout the experiment, milk yield was measured at the quarter level. Milk samples were collected from each quarter once a week to assess the milk components, and epithelial cell concentrations, as well as to isolate milk fat globule RNA. In the weeks before and after the experiment, mammary gland functional capacity was evaluated by measuring the volume of milk harvested after complete filling of the gland. At the end of the last experimental week, mammary gland biopsies were performed on each rear quarter. The milk production of quarters milked completely remained stable during the treatment period, whereas, as expected, the milk production of quarters milked incompletely was only 53% of completely milked quarters at the end of the period. Accordingly, the expression of genes related to milk synthesis (CSN2, LALBA, and ACACA) in milk fat was lower in the quarters that were milked incompletely. Incomplete milking decreased the milk lactose content, indicating a loss of integrity of tight junctions. The total yield of epithelial cells in milk was not affected, but their concentration in milk, the BAX:BCL2 gene expression ratio, and the loss of mammary functional capacity were greater in the quarters milked incompletely, suggesting an acceleration of involution in those quarters. The expression of the short isoform of the PRL receptor gene (PRLR) tended to be lower, and the expression of STAT5A and STAT5B tended to decline in the quarters milked incompletely. In mammary gland biopsy samples, the number of both short and long isoforms of the PRLR were not affected, nor were the amount and activation of STAT3 and STAT5. However, the ratio of PRLR short isoform to PRLR long isoform was lower in the quarters milked incompletely. The decrease in milk yield induced by incomplete milking is rapid and associated with a decrease in mammary epithelial cell activity and a decrease in the number of secretory epithelial cells. The results of this experiment provide only limited support for the hypothesis that modulation of the mammary gland's responsiveness to PRL is part of the mechanism by which local factors, such as incomplete milking, modulate milk synthesis.

Efficacy of bacteriocin-based formula for reducing staphylococci, streptococci, and total bacterial counts on teat skin of dairy cows.

The use of teat dips is one of the most effective strategies to control mastitis by preventing new intramammary infections. Reducing bacterial load on teat skin helps control the spread of pathogens and spoilage and improves the quality of milk. The objective of this study was to evaluate the reduction of bacterial populations through the application of bacteriocin-based teat formulas. Teats of 12 Holstein cows received 2 different concentrations of bactofencin A, nisin, and reuterin alone or in combination, as well as iodine (positive control) and saline (negative control). Teat swabs were collected before and after application of teat formulas and analyzed for staphylococci, streptococci, and total bacteria counts. There were no differences for staphylococci, streptococci, and total bacterial counts for samples collected before application throughout the entire experiment. Reuterin-low and reuterin-high treatments reduced total bacterial count by 0.47 and 0.36 logs, respectively, whereas bactofencin A had no effect on any tested bacterial groups. Nisin-low treatment reduced staphylococci, streptococci, and total bacterial counts by 0.47, 0.30 and 0.50 logs, respectively. Nisin-high treatment resulted in 0.50, 0.50, and 0.47 log reduction for staphylococci, streptococci, and total bacterial counts. The bacteriocin consortium showed the highest reduction rates with 0.91, 0.54, and 0.90 log reductions obtained for staphylococci, streptococci, and total bacteria counts, respectively, for the low-concentration consortium. Similarly, the high-concentration consortium showed reduction rates with 0.95, 0.60, and 0.82 log reductions obtained for staphylococci, streptococci, and total bacteria counts, respectively. Thus, nisin and the bacteriocin consortium showed the most promise as a teat disinfectant by reducing staphylococci, streptococci, and total bacteria counts.

Effect of photoperiod before and during first gestation on milk production and prolactin concentration in dairy heifers.

Holstein heifers (n = 45) were subjected to treatments according to a 2 × 2 factorial design where the main effects were the photoperiod treatments during the second isometric (ISO, 52-61 wk of age) and the second allometric (ALLO, 62 wk of age to 8 wk before calving) periods of mammary gland development. During the ISO period, heifers were subjected to either a short-day photoperiod (SDP; 8 h light, 16 h dark; n = 22) or a long-day photoperiod (LDP; 16 h light, 8 h dark; n = 23). During the ALLO period, the photoperiodic treatments were either maintained (SDP:SDP, n = 11; LDP:LDP, n = 11) or switched (SDP:LDP, n = 11; LDP:SDP, n = 12). The treatments ended 8 wk before calving. All animals were then subjected to about 16 h of light per day. Serum prolactin (PRL) concentration during the ISO period was greater in heifers exposed to LDP than in those exposed to SDP. For the first 20 wk of the ALLO period, heifers exposed to LDP had greater serum concentration of PRL than those exposed to SDP. On the other hand, previous exposure to LDP during the ISO period reduced the concentration of PRL compared with those exposed to SDP during that period. During the second 20 wk of the ALLO period, PRL concentration remained greater in the serum of heifers then exposed to LDP than SDP, but serum PRL was greater in heifers exposed to LDP during the ISO period. During the last weeks before calving, when all animals were exposed to LDP, previous exposure to LDP during the ALLO period reduced serum PRL. Early-lactation milk (wk 1-5) and energy-corrected milk (wk 2-6) production were higher in the heifers exposed to SDP than in those exposed to LDP during ALLO. Photoperiod had no effect on milk production after that period. In conclusion, the results do not support to the hypothesis that photoperiod affects mammary gland development during the second allometric phase. However, they confirm that a short-day photoperiod in late gestation enhances milk production in the following lactation in primiparous heifers. Using serum PRL as an indicator of the photoperiodic response, we can conclude that responsiveness to the photoperiodic signal is still conditioned by a previous photoperiod several months after it ends.

Effect of heat stress in late gestation on subsequent lactation performance and mammary cell gene expression of Saanen goats.

Little is known about the effects of heat stress during the late gestation period on lactation in dairy goats. For this reason, 32 Saanen goats were randomly assigned to 1 of 2 groups, control (CT; n = 16) or heat stress (HS; n = 16), during late gestation. The HS goats were housed in a climatic chamber before parturition and subjected to heat stress for the last 45 d. After parturition, the HS goats were housed in the same conditions as the CT group. Mammary gland biopsies were performed on 7 goats per treatment at -30, -15, 15, and 30 d relative to parturition, so that the expression levels of several genes could be determined. The HS goats produced less milk than the CT goats did during the first half of lactation, but not during the rest of lactation. Before parturition, apoptosis-related transcripts (TP53 and BAX) were higher in the mammary glands of the HS goats than in those of the CT goats. The HS goats also had higher levels of HSPB1 gene expression during gestation and lactation. However, expression of the prolactin receptor gene was lower after parturition in the mammary glands of HS, suggesting downregulation of prolactin signaling. In summary, heat stress during final gestation reduces milk yield in the subsequent lactation. Although the upregulation of apoptosis signaling in the HS goats suggests that heat stress affects mammary cell number, the loss of the effect on milk production is more compatible with an effect on cell activity, which could be due to a downregulation of prolactin signaling.

Effects of milking frequency and domperidone injections on milk production and prolactin signaling in the mammary gland of dairy cows.

Eleven mid-lactation Holstein cows were milked twice daily during the first 2 experimental weeks. During wk 3 to 10, the cows were differentially milked: right quarters were milked thrice daily (3×) and left quarters were milked once daily (1×). During wk 11 to 14, all quarters were milked twice daily. After 4 wk of differential milking, the cows received daily i.m. injections of the dopamine antagonist domperidone (DOMP; 300 mg; n = 6) or of dimethyl sulfoxide as the control (CTL; n = 5) for 8 wk (wk 7-14). During the differential milking period (wk 3-6), milk production was greater for quarters milked 3× than for those milked 1× but did not differ between DOMP and CTL cows. During the differential milking + injection period (wk 7-10), milk production continued to differ according to milking frequency. However, DOMP injection did not have an effect or interact with milking frequency on milk production. During the injection period (wk 11-14), milk production remained greater in the quarters previously milked 3× and milk production increased in DOMP injected cows but not in CTL cows. Injections of DOMP increased prolactin concentration, which was greater in the serum of DOMP cows than in that of CTL cows during the differential milking + injection and the injection periods. The expression of genes that are directly related to milk synthesis (CSN2, LALBA, and ACACA) was greater in the 3× quarters than in the 1× quarters. In addition, DOMP increased CSN2 expression during the injection period. The expression of both isoforms of the PRLR gene was greater in the 3× quarters during the differential milking + injection and the injection periods. At the protein level, injections of DOMP tended to increase the number of long PRLR isoform during the differential milking + injection period. The number of short PRLR isoform was greater in the 1× quarters than in the 3× quarters during the differential milking, the differential milking + injection, and the injection periods. The total amount of STAT3 protein was greater in the 1× quarters during the differential milking and the differential milking + injection periods. The amount of phosphorylated STAT3 protein was greater in the 1× quarters during the differential milking period. The total amount of phosphorylated STAT5 protein was greater in the 3× quarters during the differential milking and the differential milking + injection periods. The results of this experiment support the hypothesis that the responsiveness of the mammary gland to PRL is modulated by milking frequency, although the underlying mechanism remains to be determined.

Effects of feed restriction and supplementary folic acid and vitamin B12 on immune cell functions and blood cell populations in dairy cows.

Cows undergoing a negative energy balance (NEB) often experience a state of immunosuppression and are at greater risk of infectious diseases. The present study aimed to evaluate the impact of a folic acid and vitamin B12 supplement and feed restriction on several immune parameters. Sixteen cows at 45 ± 3 days in milk were assigned to 8 blocks of 2 cows each according to each cow's milk production in the previous week, and within each block, the cows randomly received weekly intramuscular injections of either saline or 320 mg of folic acid and 10 mg of vitamin B12 for 5 weeks. During week 5, the cows were fed 75% of their ad libitum intake for 4 days. Blood samples were taken before the beginning of the experiment, just before feed restriction and after 3 days of feed restriction, in order to evaluate blood cell populations, the phagocytosis capacity and oxidative burst of polymorphonuclear leukocytes (PMNs), the proliferation of peripheral blood mononuclear cells (PBMCs) and concentrations of non-esterified fatty acids (NEFAs) and ß-hydroxybutyrate. The vitamin supplement did not affect any of the tested variables except milk fat and lactose content. Feed restriction reduced milk production and increased the concentration of NEFAs. Feed restriction did not affect blood cell populations but did reduce the percentage of PMN positive for oxidative burst after stimulation with phorbol 12-myristate 13-acetate. The proliferation of PBMCs was reduced when the cell culture medium was supplemented with sera collected during the feed restriction. In conclusion, feed restriction affected the functions of PMN and PBMC and this effect was not prevented by the folic acid and vitamin B12 supplement. These results support the hypothesis that the greater risk of infectious diseases in cows experiencing a NEB is related to impaired immune cell functions by high circulating concentration of NEFAs.

Review: Inhibition of prolactin as a management tool in dairy husbandry.

Accumulating evidence supports that the hormone prolactin (PRL) is galactopoietic in dairy ruminants. Accordingly, the inhibition of PRL secretion by the dopamine agonists quinagolide and cabergoline causes a sharp decline in milk production and could be useful in several critical periods. First, PRL inhibition may reduce the incidence during the periparturient period of metabolic disorders caused by the abrupt increase in energy demand for milk production. Metabolic disturbances can be lessened by reducing milk output by milking once a day or incompletely in the first few days of lactation. The injection of cows with quinagolide for the first 4 days of lactation reduced milk production during the first week of lactation without any residual effects. Blood glucose and calcium concentrations were higher and ß-hydroxybutyric acid concentration was lower in the quinagolide-treated cows. Second, PRL inhibition may help sick or injured lactating cows, considering that they can fall into severe negative energy balance when they are unable to consume enough feed to support their milk production. This leads to a weakened immune system and increased susceptibility to diseases. When cows were subjected to feed restriction and were treated with quinagolide, the decrease in milk production was accelerated without any residual effects. The quinagolide-treated cows had higher glucose and lower ß-hydroxybutyric acid and non-esterified fatty acid concentrations than the control cows did. Third, PRL inhibition may facilitate drying-off in high-yielding cows, because they are often dried off while still producing significant quantities of milk, which delays mammary involution and increases risk of mastitis. Therefore, strategies that reduce milk production before drying-off and accelerate mammary gland involution could be an important management tool. In this context, inhibition of PRL was utilised to accelerate mammary gland dry-off. Quinagolide decreased milk production within the first day of treatment, and both quinagolide and cabergoline induced more rapid changes in several markers of mammary gland involution after drying-off. In addition, quinagolide improved the animals' resistance to intramammary infection. These results suggest that the inhibition of PRL could be a strategy for facilitating drying-off, reducing metabolic stress during the postpartum period, and alleviating acute nutritional stress during illness without compromising the overall productivity of dairy ruminants.

Review: the cellular mechanisms underlying mammary tissue plasticity during lactation in ruminants.

The mammary tissue is characterized by its capacity to adapt in response to a wide variety of changing conditions. This adaptation capacity is referred to as the plasticity of mammary tissue. In dairy ruminants, lactation is challenged by modifications that can either be induced on purpose, such as by modifying management practices, or occur involuntarily, when adverse environmental constraints arise. These modifications can elicit both immediate changes in milk yield and composition and carryover effects that persist after the end of the challenge. This review focuses on the current knowledge concerning the cellular mechanisms underlying mammary tissue plasticity. The main mechanisms contributing to this phenomenon are changes in the activity and number of mammary epithelial cells (MECs). Changes in the number of these cells result from variations in the rates of cell proliferation and death as well as changes in the rate MEC exfoliation. The number of MECs also depends on the number of resident adult mammary stem cells and their progenitors, which can regenerate the pools of the various mammary cells. Several challenges, including changes in milking frequency, changes in level of feed supply and hormonal manipulations, have been shown to modulate milk yield together with changes in mammary cell activity, turnover and exfoliation. Epigenetic changes may be an additional mechanism of adaptation. Indeed, changes in DNA methylation and reductions in milk yield have been observed during once-daily milking and during mastitis in dairy cows and may affect cell activity persistently. In contrast to what has been assumed for a long time, no carryover effect on milk yield were observed after feed supply challenges in dairy cows and modification of milking frequency in dairy goats, even though the number of mammary cells was affected. In addition, mammary tissue plasticity has been shown to be influenced by the stage of lactation, health status and genetic factors. In conclusion, the cellular mechanisms underlying mammary tissue plasticity are diverse, and the mammary tissue either does or does not show elastic properties (with no permanent deformation), in response to environmental changes.

Invited review: Accelerating mammary gland involution after drying-off in dairy cattle.

Bovine mammary gland involution, as a part of the reproductive cycle in dairy cows, is a very important remodeling transformation of the mammary gland for the subsequent lactation. There is considerable incentive to accelerate mammary gland involution to improve udder health, shorten the dry period, and simplify the management process by reducing dietary changes. The complex process of mammary involution is characterized by morphological changes in the epithelial cells and mammary tissue, changes in the composition of mammary secretions, and changes in the integrity of tight junctions. Involution is facilitated by elements of the immune system and several types of proteases and is coordinated by various types of hormones. This review first describes the involution process and then argues for the need to accelerate it. Last, this review focuses on various intervention methods for accelerating involution. Our aim is to provide a comprehensive overview of bovine mammary gland involution as well as potential techniques and new opinions for dry cow management.

Glucose and insulin responses to an intravenous glucose tolerance test administered to feed-restricted dairy cows receiving folic acid and vitamin B12 supplements.

The present experiment was conducted to determine whether, during periods of negative energy balance, the increase in glucose availability, despite similar DMI and greater milk production, induced by a combined supplement of folic acid and vitamin B12 was related to effects of insulin on metabolism. Sixteen multiparous Holstein cows averaging 45 days in milk (standard deviation: 3) were assigned to 8 blocks of 2 animals each according to their milk production (45 kg/d; standard deviation: 6) during the week preceding the beginning of the experiment. Within each block, they received weekly intramuscular injections of either saline (CON) or folic acid and vitamin B12 (VIT) during 5 consecutive weeks. During the last week, the cows were fed 75% of their ad libitum intake during 4 d. Blood samples were taken the morning before starting the feed restriction and on the third day of feed restriction. On the fourth day of feed restriction, the daily meal was not served and an intravenous glucose tolerance test was performed. During the 4 wk preceding the feed restriction, milk production and DMI were not affected by treatments. During the feed restriction, the vitamin supplement tended to decrease milk fat concentration and increase milk concentration of lactose. Plasma concentrations of homocysteine, Ile, Leu, Val, and branched-chain AA increased in VIT cows during the restriction but not in CON cows. During the glucose tolerance test, insulin peak height was lower and insulin incremental positive area under the curve tended to be lower for VIT than for CON [83 (95% confidence interval, CI: 64-108) vs. 123 (95% CI: 84-180) µg·180 min/L, respectively]. Free fatty acid nadir was reached earlier for VIT than for CON [34 (95% CI: 26-43) vs. 46 (95% CI: 31-57) min, respectively]. Glucose area under the curve, clearance rate and peak height, insulin time to reach the peak and clearance rate, and free fatty acid nadir did not differ between VIT and CON. The reduction in insulin release during a glucose tolerance test without changes in glucose clearance rate or area under the curve suggests that the vitamin supplement improved insulin sensitivity in feed-restricted lactating dairy cows.

A randomized controlled trial on the effect of incomplete milking during the first 5 days in milk on reproductive performance of dairy cows.

The main objective of the current study was to measure the effect of incomplete milking on luteal activity and on pregnancy hazard. We also aimed to study the effect of early-lactation hyperketonemia (i.e., ß-hydroxybutyrate blood concentration ≥1.4 mmol/L during the first 3 wk in milk) on those reproductive outcomes. Multiparous Holstein cows (n = 853) from 13 commercial herds were enrolled in a randomized controlled trial. Cows were assigned to a control or a treatment group, incompletely milked (10-14 L of milk collected/d) from 1 to 5 DIM. Blood samples were collected once a week during weeks in milk 1 to 3 for ß-hydroxybutyrate blood concentration, and a threshold of 1.4 mmol/L was used to define hyperketonemia. During weeks in milk 5 and 7, cows were sampled for progesterone blood concentration, and a threshold of 1 ng/mL was used to define luteal activity. Reproductive information and culling dates were obtained through herd records. Logistic regression models and survival analyses were used to assess the effect of treatment on luteal activity and on pregnancy hazard, respectively. Analogous models were used to investigate the effect of early-lactation hyperketonemia on reproductive outcomes. The odds of luteal activity for incompletely milked cows were 1.1 (95% confidence interval: 0.72-1.7) times those of conventionally milked cows. The effect of treatment on pregnancy hazard varied as a function of time, parity, and start of the breeding period. In second-parity cows that started the breeding period <55 d in milk, the pregnancy hazard (95% confidence interval) in incompletely milked cows was 576.3 (240.0-1,383.7), 36.9 (18.9-72.1), 6.8 (3.3-13.8), 2.5 (1.0-5.9), and 0.13 (0.07-0.26) times that of conventionally milked cows at 1 to 21, 22 to 43, 44 to 65, 66 to 87, and >87 d after the voluntary waiting period, respectively. The treatment did not have an effect on pregnancy hazard in cows in third parity or greater or in those starting the breeding period ≥55 d in milk. Early-lactation hyperketonemia was not associated with any of the reproductive outcomes. In conclusion, the incomplete milking protocol had no effect on luteal activity and had a positive effect on pregnancy hazard in second-parity cows in herds with a short voluntary waiting period (<55 d). We did not observe an effect of early-lactation hyperketonemia on luteal activity or on pregnancy hazard.

Effect of incomplete milking during the first 5 days in milk on udder and reproductive tract health: Results from a randomized controlled trial.

The aim of this study was to investigate the effect of an incomplete milking on risk of mastitis and reproductive tract disease. Multiparous dairy cows (n = 878) from 13 commercial herds were enrolled in a randomized controlled trial. Cows were randomly assigned to either a control (milked conventionally) or a treatment group, which consisted of an incomplete milking (10-14 L of milk collected/d) from 1 to 5 d in milk (DIM). Quarter milk samples were collected at approximately 11 and 18 DIM to measure somatic cell count (SCC). Quarters were considered negative for intramammary infection if SCC was <100,000 cells/mL and positive if SCC was ≥200,000 cells/mL. To calculate intramammary infection incidence, negative quarters of the initial samples collected were tested again 1 wk later. This was done to deter incidence of positive quarters. To calculate elimination rate, positive quarters were tested again 1 wk later to detect mastitis elimination. Farmers recorded clinical mastitis events. Cows were also examined at approximately 35 DIM with a Metricheck device (Simcro, Hamilton, New Zealand) for detection of purulent vaginal discharge (PVD) and with an endometrial cytobrush for presence of leukocytes [endometrial cytology for smear (ENDO) and for leukocyte esterase test (LE)]. A threshold ≥3 was used to define a positive PVD or LE test, whereas a polymorphonuclear cell count ≥6% was used to define a positive ENDO. Five generalized mixed models with cow or herd as random intercepts were used to determine the effects of incomplete milking on odds of new intramammary infection, odds of intramammary infection elimination, and odds of a positive PVD, LE, or ENDO status. To investigate time until first clinical mastitis event, a Cox model with a herd frailty term was used. The odds of new intramammary infection and intramammary infection elimination for incompletely milked cows were 0.90 [95% confidence interval (CI): 0.49, 1.7] and 2.9 (95% CI: 1.4, 6.0) times those of conventionally milked cows, respectively. The hazard of clinical mastitis in incompletely milked cows was 0.96 (95% CI: 0.59, 1.6) times that of conventionally milked cows. The odds of PVD, LE, and ENDO for incompletely milked cows were 1.4 (95% CI: 0.89, 2.1), 1.3 (95% CI: 0.88, 1.8), and 1.2 (95% CI: 0.81, 1.7) times those of conventionally milked cows. These results suggest that incomplete milking during the first 5 DIM increases the odds of a decrease in SCC from 11 to 18 DIM but does not affect odds of increase in SCC in the same period. The incomplete milking had no effect on clinical mastitis incidence in the first 90 DIM or on reproductive tract health at 35 DIM.

A randomized controlled trial on the effect of incomplete milking during early lactation on ketonemia and body condition loss in Holstein dairy cows.

Limiting milk production for a short period of time in early lactation could be a relevant strategy to prevent hyperketonemia (HYK). From December 2013 to March 2015, 838 multiparous Holstein cows from 13 herds were enrolled in a randomized controlled trial evaluating the effect of incomplete milking in early lactation on ketonemia and its effect on body condition score (BCS) loss. Cows were randomly assigned 4 wk before expected calving date to 1 of 2 treatment groups, (1) a conventional milking protocol (CON) for which cows were completely milked or (2) an incomplete milking protocol (INC) for which a maximum of 10 to 14 kg of milk/d were withdrawn during the first 5 d in milk (DIM). ß-Hydroxybutyrate (BHB) concentrations were measured from blood samples collected on each cow 3 times at weekly intervals. Hyperketonemia was defined as BHB ≥1.4 mmol/L. Body condition score variation in the postcalving period was calculated by subtracting BCS assessed at wk 7 from BCS assessed at first week after calving. Effect of treatment on ketonemia and prevalence of HYK were evaluated for 4 specific time periods: 1 to 3, 4 to 7, 8 to 17, and 18 to 26 DIM. Effect of treatment on ketonemia was investigated using linear mixed models with natural logarithm of BHB measurements as outcome and treatment groups as fixed effect. Generalized linear mixed models with HYK as outcome, using logit link, and treatment groups as fixed effect were used to investigate effect of treatment on odds of HYK. A logistic regression model with BCS loss (<0.75 or ≥0.75) as outcome and treatment groups and herd as fixed effects was used to study effect of INC on odds of having BCS loss ≥0.75. A total of 813 lactations had complete data and were used for statistical analysis of ketonemia and HYK. A total of 709 lactations had complete data and were used for analysis of BCS loss. Geometric means of blood BHB concentrations during the 1 to 3, 4 to 7, 8 to 17, and 18 to 26 DIM periods were, respectively, 0.72 (95% confidence interval = 0.66, 0.80), 0.66 (0.60, 0.73), 0.90 (0.80, 1.01), and 0.93 (0.83, 1.05) mmol/L for INC, and 0.65 (0.59, 0.72), 0.79 (0.72, 0.87), 0.94 (0.84, 1.06), and 0.92 (0.82, 1.04) mmol/L for CON. Cows in INC group had lower ketonemia during the 4 to 7 DIM period. Predicted prevalence of HYK during the 1 to 3, 4 to 7, 8 to 17, and 18 to 26 DIM periods were, respectively, 2.8 (3.2, 15.1), 4.6 (2.0, 10.0), 13.4 (8.4, 20.0), and 23.0% (17.4, 29.7) for INC and 2.6 (2.5, 13.8), 10.7 (5.6, 19.3), 19.4 (13.0, 27.9), and 21.3% (16.0, 27.8) for CON. The INC treatment reduced the prevalence of HYK during the 4 to 7 and 8 to 17 DIM periods. No association was observed between INC and BCS loss in the postcalving period. Overall, the incomplete milking protocol was effective for reducing ketonemia and prevalence of HYK during the early postpartum period.

A randomized controlled trial on the effect of incomplete milking during the first 5 days in milk on culling hazard and on milk production and composition of dairy cows.

An incomplete milking in early lactation could help limit negative energy balance in dairy cattle, but its potential effects on culling hazard and on milk production and composition throughout the entire lactation are unknown. The objective of this study was to evaluate the effect of an incomplete milking during the first 5 d in milk on culling hazard, milk weight, milk fat and protein concentrations, and energy-corrected milk (ECM) yield during the whole lactation. A randomized controlled trial was conducted in 13 dairy farms near St-Hyacinthe, Quebec, Canada. Approximately 1 mo before expected calving, Holstein multiparous cows calving between December 2013 and March 2015 (n = 846 cow lactations) were randomly assigned to a control or a treatment group. Cows in the control group were milked conventionally, whereas cows in the treatment group were submitted to an incomplete milking protocol (maximum of 10, 12, and 14 L/d of milk was collected on days in milk 1-3, 4, and 5, respectively). All farms were registered on Dairy Herd Improvement Association, which was used to obtain records on culling, monthly milk yield, and milk fat and protein concentrations. In addition, daily milk yield records were available for 6 farms. A Cox proportional hazards model with a herd frailty term was fitted to the data to compare culling hazard among treatment groups. Regarding milk production and composition, 4 linear mixed models with herd as a fixed effect, cow as a random effect, and using an autoregressive covariance structure were used to study the effect of the incomplete milking on (1) milk weight, (2) milk fat concentration, (3) milk protein concentration, and (4) ECM yield. Culling hazard did not differ among treatment groups (hazard ratio = 1.0; 95% CI = 0.82, 1.3). We observed no differences in milk weight, milk fat, or protein concentration among treatment groups between weeks in milk (WIM) 2 and 44 (the studied period). We noted a difference in ECM between treatment groups for WIM 38, with incompletely milked cows producing less milk than conventionally milked cows (-2.7 kg/d; 95% CI = -0.02, -5.2 kg/d), but no differences were found for any of the other WIM. These results suggest that this strategy for controlling the negative energy balance has negligible effect on cow productivity.

Effect of the concentration of circulating prolactin on dairy cows' responsiveness to domperidone injection.

The objective of this study was to determine whether the responsiveness of the mammary gland to prolactin (PRL) is affected by the concentration of the hormone. After 1 pre-experimental week (d -7 to -1), 18 Holstein cows in mid to late lactation were injected intramuscularly twice daily with either 0.5 mg of quinagolide (QN) or 2 mL of water (control) for 2 wk (d 1 to 14; treatment period). After the treatment period, all cows received daily subcutaneous injections of 300 mg of domperidone (DOMP) for 3 wk (d 15 to 35; DOMP period). The cows were monitored for an additional 2 wk as a posttreatment period (d 36 to 49). Blood and milk samples were collected 3 times per week. Additionally, blood samples were collected during the a.m. milking on d -4, 14, and 35. Milk production was not affected by QN during the treatment period but was increased during the DOMP and posttreatment periods in the QN cows. With respect to milk composition, the treatments affected only the protein content, which was greater in the QN cows during the treatment period. Blood PRL concentration declined during QN injections and was lower in the QN cows than in the control cows between d 5 and 14. The basal concentration of PRL was increased by DOMP injections during the DOMP and posttreatment periods but was not affected by previous QN injections. Prolactin concentration in milk was not affected by the QN treatments but was increased by DOMP injections during the DOMP and posttreatment periods. Milking-induced PRL release was decreased by QN on d 14. On d 35, milking did not induce a significant release of PRL above the baseline for both treatments. In conclusion, the results of this experiment support the contention that the mammary gland's responsiveness to PRL is modulated by the previous level of the hormone.

Effect of 17ß-estradiol on milk production, hormone secretion, and mammary gland gene expression in dairy cows.

Estradiol inhibits milk production in dairy cows. The present study evaluated the effect of 17ß-estradiol (E2) injections on prolactin (PRL) secretion and the mammary gland response to this hormone. Eight mid-lactation cows were used in a crossover design. During each experimental period, the cows were injected daily with either E2 (2.5 mg) or soy oil (2.5 mL; control) for 7 d. For each period, blood and milk samples were collected from d -4 to 14 (relative to the first injection) to measure PRL, insulin-like growth factor-1, and cortisol concentrations. In addition, blood samples were collected during morning milking on d -4, 2, and 7 to determine the milking-induced PRL release. Mammary gland biopsies were collected on the last day of injections. Milk fat samples were collected from d 1 to 7 and on d 14. The mRNA levels of genes encoding proteins related to mammary activity (α-lactalbumin, ß-casein, and acetyl-coenzyme A carboxylase), apoptosis (Bax, Bcl2, and caspase-3), PRL receptors (PRLR; long and short forms), signal transducer and activator of transcription 5 (STAT5A and STAT5B), and suppressors of cytokine signaling (SOCS2 and SOCS3) were evaluated by real-time reverse transcription PCR using RNA extracted from milk fat and mammary biopsies. Milk production was decreased moderately (about 9%) by E2 injections during the treatment period. Estradiol injections increased basal PRL levels in serum and milk but did not affect milking-induced PRL release. Estradiol injections increased the plasma concentration of insulin-like growth factor-1 but did not affect cortisol concentration during the treatment period. In mammary tissue, the expression of Bcl2 was downregulated, whereas that of STAT5A and B and the Bax:Bcl2 mRNA ratio was higher during E2 injections. The total STAT5 protein content in mammary tissue was elevated by E2 injections. We found no significant difference observed for the other genes in mammary tissue or milk fat. The present data do not support the contention that E2 injections inhibit milk production by interfering with PRL signaling, but enhanced basal PRL concentration and STAT5 gene expression in mammary tissue.

Innovative dairy cow management to improve resistance to metabolic and infectious diseases during the transition period.

The incidence of metabolic and infectious diseases varies greatly during the lactation cycle. Most new cases of clinical mastitis appear at the beginning of lactation, and the incidence increases with the level of milk production. In addition to mastitis, many other infectious diseases become clinically apparent during the first 2weeks of lactation. During this time, cows are in a negative energy balance and must mobilize body reserves to balance the deficit between food energy intake and energy required for milk production. The relationships between energy deficit and metabolic diseases, such as ketosis and hepatic lipidosis, are well known. Furthermore, cows in energy deficit have a weakened immune system and are therefore more susceptible to infections. There is now good evidence that the increase in circulating non-esterified fatty acids impairs immune cell functions. Therefore, management approaches that reduce the negative energy balance and the increase in non-esterified fatty acids at the beginning of lactation are likely to improve resistance to infection. Improving the nutrient supply through periparturient nutritional management has been the subject of considerable research. However, another way to reduce the imbalance between nutrient supply and demand is to temporarily decrease the latter. In this review, we examine how management strategies such as conjugated linoleic acid feeding, prepartum milking, or limiting postpartum milk production could be used to reduce metabolic perturbations and immunosuppression during the transition period. At this stage, it appears that reducing the amount of milk harvested postpartum by means of partial milking in the first days after calving is the most promising approach to reduce metabolic stress and immunosuppression without compromising the productivity of high-yielding dairy cows.

Effects of photoperiod modulation and melatonin feeding around drying-off on bovine mammary gland involution.

The risk for a dairy cow to acquire new intramammary infections is high during the transition from lactation to the dry period, because of udder engorgement and altered immune functions. Once the gland is fully involuted, it becomes much more resistant to intramammary infections. Therefore, strategies to depress milk yield before drying-off and accelerate the involution process after drying-off could be beneficial for udder health. The objective of this study was to assess the effect of photoperiod manipulation and melatonin feeding from 14 d before to 14 d after drying-off on the speed of the involution process. Thirty Holstein cows in late lactation were randomly allocated to one of the following treatments: (1) a long-day photoperiod (16 h of light: 8 h of darkness), (2) a short-day photoperiod (8 h of light: 16 h of darkness), and (3) a long-day photoperiod supplemented by melatonin feeding (4 mg/kg of body weight). Milk and blood samples were collected on d -26, -19, -12, -5, -1, 1, 3, 5, 7, 10, and 14 relative to the last milking to determine concentrations of mammary gland involution markers and serum prolactin. Additional blood samples were taken around milking on d -15, before the start of the treatments, and on d -1, before drying-off, to evaluate the treatment effects on milking-induced prolactin release. The short-day photoperiod slightly decreased milk production and basal prolactin secretion during the dry period. The milking-induced prolactin surge was smaller on d -1 than on d -15 regardless of the treatments. Lactoferrin concentration, somatic cell count, and BSA concentration as well as matrix metalloproteinase-2 and -9 activities increased in mammary secretions during the first 2 wk of the dry period, whereas milk citrate concentration and the citrate:lactoferrin molar ratio decreased. The rates of change of these parameters were not significantly affected by the treatments. The long-day photoperiod supplemented by melatonin feeding did not affect milk production, prolactin secretion, or mammary gland involution. Under the conditions in this study, photoperiod modulation and melatonin feeding did not appear to affect the rate of mammary gland involution.