Optimizing ovulation to first GnRH improved outcomes to each hormonal injection of ovsynch in lactating dairy cows

Ovulatory response to the first GnRH of Ovsynch is the critical determinant for successful synchronization of ovulation in dairy cows. Our objective in this study was to develop a pre-Ovsynch treatment that increased the percentage of cows that ovulated in response to the first GnRH injection of Ovsynch. To accomplish our goal, we evaluated a hormonal strategy that consisted of PGF_2α and GnRH before the first GnRH of Ovsynch. Lactating dairy cows (n = 137) were assigned to receive either no treatment before Ovsynch (control) or 25 mg of PGF_2α (PreP) followed 2 d later by 100 μg of GnRH (PreG), administered 4 (G4G), 5 (G5G), or 6 (G6G) d before initiating the Ovsynch protocol. Transrectal ultrasonography was performed to assess follicular size and resulting ovulation, and blood samples were collected to measure circulating concentrations of progesterone and estradiol immediately before each hormonal injection. Cows were inseminated at a fixed time 16 h after final GnRH of Ovsynch. Pregnancy diagnosis was performed 35 d later by palpation per rectum of uterine contents. Proportion of cows that ovulated to first GnRH of Ovsynch was 56.0, 66.7, 84.6, and 53.8% for G4G, G5G, G6G, and controls, respectively, and was greater for G6G than for control cows. Luteolytic response to PGF_2α of Ovsynch was greater in all treated than control cows (92.0, 91.7, 96.2, and 69.2% for G4G, G5G, G6G, and control, respectively). Synchronization rate to Ovsynch was greater (92 vs. 69%, respectively) in G6G than in control cows. In addition, cows that ovulated in response to first GnRH of Ovsynch had greater response to PGF_2α of Ovsynch (92.7 vs. 77.1%, respectively) and greater synchronization rate to the overall protocol (87.9 vs. 62.9%, respectively) than those that did not ovulate. Concentrations of progesterone at PGF_2α of Ovsynch, and estradiol and follicle size at final GnRH of Ovsynch, were identified as significant predictors of probability of pregnancy 35 d after artificial insemination. In summary, a PGF_2α-and-GnRH based pre-Ovsynch strategy consisting of a 6-d interval between PreG and first GnRH of Ovsynch resulted in a greater ovulatory and luteolytic response to first GnRH and PGF_2α of Ovsynch, respectively, compared with control cows. This, in turn, optimized synchronization rate to Ovsynch.     

Short communication: Repeat breeder cows with fluid in the uterine lumen had poorer fertility

The objective of the present study was to test the association between the presence of fluid in uterine lumen and the fertility of repeat breeder cows (RBC). Grazing Holstein cows that were clinically normal and were not pregnant after at least 3 consecutive inseminations were defined as RBC and included in the study (n = 358). A cut-point ≥2 mm of uterine lumen visible by ultrasonography was used as indicative of the presence of fluid in the uterus. The effect of this fluid on the fertility of RBC was assessed by mixed logistic regression. Twenty percent of the RBC had ≥2 mm of fluid in their uterus, and this finding was associated with a marked reduction in the odds of conception (odds ratio = 0.46). Additional intrauterine sampling revealed that 24.0% of RBC showed signs of subclinical endometritis [≥5% neutrophils in endometrial cytology (SCE)]. However, we found no agreement between the finding of uterine fluid and the diagnosis of SCE (kappa coefficient = 0.03, 95% confidence interval = ?0.07 to 0.14). In conclusion, the presence of fluid (≥2 mm) in uterine lumen, detected by ultrasonography, was associated with lower fertility in RBC, but was not related to the diagnosis of SCE. Thus, ultrasonographic evaluation is an important tool to assist in assessment of RBC.     

Effects of dietary fats differing in n-6:n-3 ratio fed to high-yielding dairy cows on fatty acid composition of ovarian compartments, follicular status, and oocyte quality

The objectives were to determine the incorporation of dietary encapsulated fats differing in n-6:n-3 ratio into milk fat, plasma, and various ovarian compartments and to examine the effects on ovarian follicular status, preovulatory follicle characteristics, and oocyte quality. Twenty-four multiparous Israeli Holstein cows, averaging 114 d in milk, were assigned to 1 of 3 treatment groups: 1) control (n = 7), in which cows were fed a lactating cow diet; 2) E-FLAX (n = 8), in which cows were fed a lactating cow diet that consisted of 1 kg/d of encapsulated fat (3.8% of dry matter) containing 40.8% flaxseed oil, providing 242.2 g of C18:3n-3 (low n-6:n-3 ratio); or 3) E-SUN (n = 9), in which cows were fed a lactating cow diet that consisted of 1 kg/d of encapsulated fat (3.8% of dry matter) containing 40.8% sunflower oil, providing 260.0 g of C18:2n-6 (high n-6:n-3 ratio). Ovaries were monitored by ultrasonography for follicular status, and after synchronization, follicles >7 mm were aspirated and evaluated. Ovum pickup was performed (19 sessions for the control and E-FLAX groups and 11 for the E-SUN group), and in vitro maturation and oocyte fertilization were conducted. The E-FLAX treatment increased the proportions of C18:3n-3 (5.8 fold), C20:5n-3, and C22:5n-3 (approximately 4-fold) in milk fat as compared with the other 2 treatments. The proportion of C18:3n-3 fatty acid in plasma increased dramatically with the E-FLAX treatment, from 1.43 and 1.49% in the control and E-SUN groups, respectively, to 7.98% in the E-FLAX group. Consequently, the n-6:n-3 ratio in plasma was reduced from approximately 42 in the control and E-SUN groups to 6.74 in the E-FLAX group. Proportions of C18:3n-3 in follicular fluid and granulosa cells were approximately 5-fold higher in the E-FLAX group than in the other 2 groups. The percentage of C18:2n-6 in cumulus-oocyte complexes of cows in the E-SUN group was 54% higher than that in the E-FLAX group and was 2.4-fold higher than that in the control group; the proportion of C18:3n-3 in the E-FLAX group was 4.73% and was not detected in the other groups. The average numbers of 2- to 5-mm follicles on d 5 and 9 of the cycle were higher in the E-FLAX group than in the E-SUN group, whereas the average numbers of follicles ≥10 mm on d 5, 9, and 13 were higher in the E-SUN group than in the other 2 groups. The estrous cycles of the cows were synchronized and PGF_2α was injected on d 16 to 17 of the cycle. The interval from PGF_2α injection to behavioral estrus was longer in the E-FLAX group than in the E-SUN group, and the beginning of the luteal phase of the subsequent cycle was delayed. Concentrations of estradiol in follicular fluid of the preovulatory follicles were higher in the E-SUN group than in the E-FLAX group. The number of follicles aspirated by ovum pickup was higher in the E-FLAX group than in the control group, and the cleavage rate in the E-FLAX group was higher than in the control group, but not the E-SUN group. In conclusion, dietary n-3 fatty acids influenced the follicular status and increased the cleavage rate of oocytes as compared with those of control cows. These findings could be related to modifications of the fatty acid composition in plasma and ovarian compartments in response to dietary supplementation.     

Effect of reducing the period of follicle dominance in a timed artificial insemination protocol on reproduction of dairy cows

Objectives were to determine the effect of reducing the period of follicle dominance in a timed artificial insemination (AI) protocol on pregnancy per AI (P/AI) in Holstein cows. In experiment 1, 165 cows received 2 injections of PGF_2α at 36 and 50 d in milk (DIM). At 61 DIM, cows were assigned randomly to Cosynch 72 h (CoS72: d 61 GnRH, d 68 PGF_2α, d 71 GnRH) or to a 5-d Cosynch 72 h with 1 (5dCoS1: d 61 GnRH, d 66 PGF_2α, d 69 GnRH) or 2 injections of PGF_2α (5dCoS2: d 61 GnRH, d 66 and 67 PGF_2α, d 69 GnRH). Blood was sampled at the first GnRH, first PGF_2α, and at the second GnRH of the protocols and assayed for progesterone. Ovulatory responses to GnRH were evaluated by ultrasonography. Cows were considered synchronized if they had concentrations of progesterone ≥1 ng/mL and <1 ng/mL on the days of the PGF_2α, and the second GnRH of the protocols, respectively, and if they ovulated within 48 h of the second GnRH injection. In experiment 2, 933 cows were assigned randomly to CoS72 or 5dCoS2. Blood was assayed for progesterone and ovaries were scanned as in experiment 1. Plasma on the days of the first PGF_2α and final GnRH of the timed AI protocols was assayed for estradiol in 75 cows. Pregnancy was diagnosed on d 38 and 66 after AI. In experiment 1, the proportions of cows with corpora lutea (CL) regression on the day of AI differed and were 79.0, 59.1, and 95.7% for CoS72, 5dCoS1, and 5dCoS2, respectively. Cows that ovulated to the first GnRH of the Cosynch tended to have lesser CL regression than cows that did not ovulate (73.0 vs. 86.4%). Protocol synchronization differed between treatments and they were greater for CoS72 (69.4%) and 5dCoS2 (78.4%) than for 5dCoS1 (42.3%). In experiment 2, CL regression was lesser (91.5 vs. 96.3%) but detection of estrus at timed AI (30.9 vs. 23.6%) was greater for CoS72 than 5dCoS2, and cows in estrus had increased P/AI (46.2 vs. 31.9%). Cows in CoS72 ovulated a larger follicle and had greater concentrations of estradiol on the day of AI than cows in 5dCoS2, but protocol synchronization tended to increase in cows receiving the 5dCoS2. When all 933 cows were evaluated, P/AI was greater for 5dCoS2 than for CoS72 (37.9 vs. 30.9%). Similarly, when only cows with progesterone <1 ng/mL on the day of AI were evaluated, P/AI was greater for 5dCoS2 than for CoS72 (39.3 vs. 33.9%). Treatment with PGF_2α on d 5 and 6 after GnRH resulted in increased luteolysis and allowed for reducing the interval from GnRH to timed AI, which increased P/AI. Reducing time of follicle dominance in timed AI protocols improves fertility of lactating dairy cows.     

Effects of high dietary crude protein on the characteristics of preovulatory follicles in dairy heifers

The objectives were to examine the effect of high dietary crude protein on characteristics of preovulatory follicles in dairy heifers. Eight Israeli-Holstein heifers, 4 fitted with rumen fistula and 4 intact, were assigned to 1 of 3 treatments in a replicated (n = 2) 4 × 3 incomplete Latin square design with 39-d periods. Treatments were: low (6.0%; LP), moderate (13.0%; MP), and high (20.0%; HP) crude-protein diets, containing 1.27 Mcal NE_L/kg dry matter. Diets were based on approximately 66% wheat straw and various proportions of ground corn grain and soybean meal. The estrous cycles of the heifers were synchronized, and 14 d after behavioral estrus, heifers received PGF_2α injections. After a further 40 h, at d 39 of each period, follicular fluid (FF) was aspirated from follicles of diameter >7 mm. The intake of the LP diet was 9% lower than that of MP and HP diets. Rumen ammonia and plasma urea nitrogen concentrations were highest in the HP and lowest in the LP, with intermediate levels in MP diets. No differences were found between treatments in plasma and FF concentrations of glucose and nonesterified fatty acids. High-protein diets increased urea concentrations very similarly in preovulatory FF and in plasma. No differences were observed between diets, in preovulatory follicle diameters and concentrations of androstenedione. However, higher estradiol and progesterone concentrations in FF were observed under the HP than under the MP diet, with no difference between diets in estrogen to progesterone ratio. It can be concluded that high concentrations of urea in plasma, caused by high dietary crude protein, penetrated into preovulatory follicles, but did not impair preovulatory characteristics. This lack of detrimental effects might be attributed to the use in this study of nonlactating heifers, which have fewer nutritional and physiological constraints and eliminate negative effects of potential interactions with high urea on dairy cows’ reproductive systems.     

Effects of whole cottonseed diet and recombinant bovine somatotropin on ovarian follicles in lactating dairy cows

The effects of whole cottonseed (WCS) in the diet and the administration of bovine somatotropin (bST) on ovarian follicular dynamics and plasma progesterone (P4) concentrations were examined in cows during a period of synchronized follicular growth. Lactating Holstein cows (n = 28) were randomly assigned to treatments in a 2 x 2 factorial arrangement. Diets consisted of WCS (15% of dry matter) or no WCS, and bST at a dose of 0 or 208 mg/14 d. Dietary treatments began within 24 h of calving and bST treatments began within 7 d postpartum. Cows received GnRH at 65 +/- 3 d postpartum (d 0), PGF2alpha, (d 7), a second dose of GnRH (d 9), and were inseminated 16 h later (d 10). Ovarian changes were monitored daily by ultrasonography from d 0 to 9. On d 9,93% of cows had a preovulatory follicle and 86% ovulated. For Class 2 (6 to 9 mm) follicles, a diet x bST interaction was detected, with bST stimulating Class 2 follicles in cows fed WCS, but not in cows on the control diet. Neither diet nor bST affected numbers of Class 1 (2 to 5 mm) or Class 3 (> or = 10 mm) follicles or sizes of the subordinate and dominant follicles. During the luteal phase of the cycle, lactating cows fed WCS tended to have elevated concentrations of plasma P4, whereas bST was without effect. Plasma concentrations of high-density lipoprotein cholesterol were increased in cows fed WCS. Number and diameter of corpora lutea did not differ among treatments.     

Effects of dietary unsaturated fatty acids on oocyte quality and follicular development in lactating dairy cows in summer

Dietary sources of fatty acids were evaluated for their influence on oocyte quality and follicular development using 54 lactating cows in summer. Fat supplements were 1) sunflower oil (80% cis 18:1), 2) Ca salt of transoctadecenoic acids (57% trans 18:1), 3) Ca salt of vegetable oils (30% 18:2), and 4) linseed oil (56% 18:3 and 16% 18:2). Fats were fed at 1.35% of dietary dry matter beginning at 5 wk prior to expected calving date and at 1.5% (oils) and 1.75% (Ca salts) of dietary dry matter for 15 wk after parturition. Four days following a programmed induced ovulation, 5 transvaginal oocyte aspirations were performed 3 or 4 d apart. Three days after the last aspiration, PGF_2α was injected, followed 3 d later by a GnRH injection and a timed artificial insemination (d 0) 16 to 20 h later. For the first 4 aspirations, oocytes grading 1 or 2 were used for in vitro embryo production. Total cell number and the proportion of terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL)-positive blastomeres were analyzed at d 8. At the fifth aspiration, the occurrence of metaphase II, group II caspase activity, and TUNEL labeling were determined after oocyte maturation. A total of 1,011 oocytes were collected. The proportion of oocytes with high caspase activity was greater for grade 3 compared with grades 1 and 2 (37.5 vs. 1.54 and 1.61%). Feeding polyunsaturated fatty acids, as compared with monosaturated fatty acids, failed to affect oocyte quality, as demonstrated by subsequent embryo development. Cows fed 18:2- or 18:3-enriched diets had a larger preovulatory follicle at insemination and subsequent volume of the corpus luteum compared with those fed cis 18:1 or trans 18:1 diets (16.8, 16.2 vs. 15.0, 14.9 ± 0.7 mm; 7,323, 8,208 vs. 6,033, 5,495 ± 644 mm³, respectively). The previously documented benefits of polyunsaturated fatty acids on reproductive performance appear to reflect actions at alternative biological windows in lactating dairy cows.     

Parity-related changes in bovine follicle and oocyte populations,oocyte quality,and hormones to 90 days postpartum

Ultrasound-guided transvaginal follicular aspiration was used to obtain oocytes from Holstein cows to study follicular development and oocyte morphology during early lactation. A single aspiration was conducted on each of the 120 cows during wk 1 to 12 postpartum. On the day of aspiration, the number of follicles and their sizes were recorded. The collected oocytes were morphologically classified into four grades. Blood and follicular fluid samples were collected for hormone and metabolite assays. Greater serum nonesterified fatty acids concentrations for first- and second-parity cattle on d 7 implied a more negative energy balance status than for third-parity cattle. The numbers of follicles and oocytes retrieved were affected by linear days postpartum x parity interactions, with second-parity cattle having increases compared with decreases for third-parity cattle. Oocyte quality score was affected by the quadratic days postpartum x parity interaction. First- and second-parity cattle had greater oocyte quality scores from d 20 to 70 postpartum compared with third-parity cattle, but third-parity cattle had greater quality oocytes near the end of the 12-wk period. In contrast to cattle subjected to multiple aspirations, first- and second-parity cattle had higher estradiol and insulin-like growth factor I concentrations in follicular fluid associated with greater numbers of oocytes retrieved than third-parity cattle. This study demonstrated that conditions related to early lactation have a negative effect on oocyte quality and endocrine measures of dairy cattle and that animals of various parities may be differentially affected.     

Follicular wave of the ovulatory follicle and not cyclic status influences fertility of dairy cows

Two experiments evaluated the influence of follicular wave at artificial insemination (AI) on fertility of dairy cows. In experiment 1, data from 5,607 lactating cows enrolled in estrous and ovulation synchronization programs for AI were evaluated. Cows’ blood was analyzed for progesterone 7 to 14 d apart, with the second sample collected on the day of the first GnRH (GnRH1) of the synchronization protocol. Cows were classified as cyclic if progesterone was ≥1 ng/mL in at least 1 of the 2 samples and as anovular if both samples were <1 ng/mL. Cyclic cows were categorized as low (CLOW; < 1 ng/mL) or high (CHIGH; ≥ 1 ng/mL) progesterone on the day of GnRH1, which would result in ovulation of the dominant follicle of the first (FW) and second (SW) follicular waves, respectively, at AI. Pregnancy per AI (P/AI) was determined 30 and 53 d after AI. In experiment 2, 220 cyclic Holstein cows received 2 injections of PGF_2α administered 14 d apart. The Ovsynch protocol (d 0 GnRH, d 7 PGF_2α, d 9 GnRH, d 9.5 timed AI) was initiated either 3 or 10 d after the second PGF_2α of the presynchronization to result in insemination to the FW or SW dominant follicles. Blood was analyzed for progesterone and ovaries were scanned to determine ovulatory responses and follicle diameter. Pregnancy was determined on d 32 and 67 after timed AI. In experiment 1, P/AI on d 30 was greater for CHIGH cows than for CLOW and anovular cows (43.0, 31.3, and 29.7%, respectively), but because of pregnancy loss, P/AI on d 53 was lowest for anovular cows. Proportions of cows with short reinsemination intervals differed among groups and were 7.1, 15.7, and 11.9% for CHIGH, CLOW, and anovular cows, respectively. Pregnancy loss was greater for anovular cows than for CLOW cows (15.0 vs. 10.0%) and was intermediate for CHIGH cows (13.5%). In experiment 2, 9.8 and 97.2% of the FW and SW cows, respectively, had progesterone ≥1 ng/mL at GnRH1. Concentrations of progesterone at the GnRH1 and PGF_2α injections of the Ovsynch protocol were greater for SW cows than FW cows. Pregnancy per AI was greater for SW cows than for FW cows (41.7 vs. 30.4%) despite less ovulation to GnRH1 in SW cows than in FW cows (78.7 vs. 88.4%). Collectively, these data indicate that follicular wave of the ovulatory follicle and not cyclic status caused the greatest reduction in P/AI in dairy cows. Whether the culprit is the follicle itself or the hormonal milieu characteristic of the first follicular wave and the early stage of the estrous cycle remains to be elucidated. Synchronization programs that induced ovulation of the FW follicle at AI reduced P/AI in lactating dairy cows, and ovulation of the FW follicle, or development of the ovulatory follicle under low progesterone concentrations, or both, might be mechanisms for reduced fertility in anovular cows.     

Metabolism of silage alcohols in lactating dairy cows

Dairy cows fed silage are subjected to various alcohols and low molecular weight esters. Four lactating Holstein cows fitted with ruminal cannulas and permanent indwelling catheters in the hepatic portal vein, hepatic vein, mesenteric vein, and mesenteric artery were used to study the absorption of alcohols into portal blood and the metabolism of feed alcohols in the rumen and splanchnic tissues. The cows were allocated to 4 experimental treatments in a Latin square design. All treatments were formulated as total mixed rations with the same overall nutrient composition, differing by the source of corn silage. Treatments were a control silage and 3 qualities of problematic corn silage (silage with Fusarium toxin, Penicillium-infected silage, and silage with a high propanol content). Feeding was followed by a decreasing ruminal pH, as well as decreasing molar proportions of ruminal acetate and isobutyrate. The ruminal concentrations of total VFA, ethanol, propanol, 2-butanol, ethyl acetate, propyl acetate, glucose, and l-lactate, and molar proportions of propionate, butyrate, isovalerate, valerate, and caproate increased after feeding. Treatments affected ruminal concentrations of propanol, propyl acetate, and butyrate and a strong correlation was observed between ruminal propyl acetate and the molar proportion of butyrate (r = −0.79). Arterial concentrations of ethanol, propanol, propanal, acetone (sum of acetone and acetoacetate), 3-hydroxybutyrate, l-lactate, glutamate, and glutamine increased, and the arterial concentration of glucose decreased after feeding, but no effects of treatment were observed for arterial variables. The postprandial increase in arterial ethanol was maintained for 5 h. The net portal release of ethanol tended to decrease with the treatment with the lowest ethanol content, and the net splanchnic release of ethanol increased after feeding, but overall, the net splanchnic flux of ethanol was not different from zero, in agreement with the liver being the major organ for alcohol metabolism. The net portal flux and net hepatic flux of propanol were affected by treatment. All dietary ethanol and propanol were accounted for by absorption of the respective alcohol into the portal blood. The hepatic extraction ratios of ethanol and propanol were, on average, 63 to 66%, and no indications of saturation of hepatic alcohol metabolism were observed at any time. We concluded that typical amounts of alcohols in corn silage do not interfere with splanchnic metabolism of any of the measured variables and do not saturate hepatic pathways for alcohol metabolism. However, even low concentrations of alcohols in feed might affect ruminal metabolism and are followed by hours of elevated peripheral blood concentrations of alcohols.     

Cow characteristics associated with the variation in number of contacts between dairy cows

In modern freestall barns where large groups of cows are housed together, the behavior displayed by herd mates can influence the welfare and production of other individuals. Therefore, understanding social interactions in groups of dairy cows is important to enhance herd management and optimize the outcomes of both animal health and welfare in the future. Many factors can affect the number of social contacts in a group. This study aimed to identify which characteristics of a cow are associated with the number of contacts it has with other group members in 2 different functional areas (feeding and resting area) to increase our understanding of the social behavior of dairy cows. Inside 2 herds housed in freestall barns with around 200 lactating cows each, cow positions were recorded with an ultra-wideband real-time location system collecting all cows' positions every second over 2 wk. Using the positioning data of the cows, we quantified the number of contacts between them, assuming that cows spending time in proximity to one another (within a distance of 2.5 m for at least 10 min per day) were interacting socially. We documented in which barn areas these interactions occurred and used linear mixed models to investigate if lactation stage, parity, breed, pregnancy status, estrus, udder health, and claw health affect the number of contacts. We found variation in the number of contacts a cow had between individuals in both functional areas. Cows in later lactation had more contacts in the feeding area than cows in early lactation. Furthermore, in one herd, higher parity cows had fewer contacts in the feeding area than first parity cows, and in the other herd, cows in third parity or higher had more contacts in the resting area. This study indicates that cow characteristics such as parity and days in milk are associated with the number of contacts a cow has daily to its herd mates and provides useful information for further research on social interactions of dairy cows.     

Atypical progesterone profiles and fertility in Swedish dairy cows

The incidence of normal and atypical progesterone profiles in Swedish dairy cows was studied. Data were collected from an experimental herd over 15 yr, and included 1,049 postpartum periods from 183 Swedish Holstein and 326 Swedish Red and White dairy cows. Milk progesterone samples were taken twice weekly until initiation of cyclical ovarian activity and less frequently thereafter. Progesterone profiles were 1) normal profile: first rise in milk progesterone above the threshold value before d 56 postpartum, followed by regular cyclical ovarian activity (70.4%); 2) delayed onset of cyclical ovarian activity: low milk progesterone the first 56 d postpartum (15.6%); 3) cessation of cyclical ovarian activity: ovarian activity resumed within 56 d postpartum, but ceased for a period of 14 d or more (6.6%); and 4) prolonged luteal phase: ovarian activity resumed within 56 d postpartum, but milk progesterone remained elevated in the nonpregnant cow for a period of 20 d or more (7.3%). Swedish Holsteins had 1.5 times higher risk of atypical profile than Swedish Red and Whites. Risk of atypical profiles was 0.5 and 0.7 times lower for older cows compared with first-parity cows; 2.3 times higher for cows in tie-stalls compared with those in loose housing; 2.6 times higher for cows calving during winter compared with summer; 0.5 times lower for cows in earlier (1994-1999) calving-year groups compared with the most recent (2000-2002); 2.5 times higher for cows with planned extended calving interval compared with conventional calving interval; and 2.2 times higher for an atypical profile in previous lactation compared with a normal profile. Cows with atypical profiles had a 15-d increase in interval from calving to first artificial insemination and an 18-d increase in interval from calving to conception. Progesterone samples taken within the first 60 d postpartum were used to calculate the percentage of samples above the threshold value of luteal activity. This measure had a significantly different mean in profiles and can be used to separate delayed onset of cyclical ovarian activity profiles and prolonged luteal phase profiles from normal. Thereby, it may be a more effective tool than measurements based only on the onset of ovarian cyclical activity in genetic evaluation of early postpartum fertility in dairy cows.     

Risk factors associated with postpartum subclinical hypocalcemia in dairy cows

Our study objectives were to evaluate the association of prepartum plasma Mg concentrations with subclinical hypocalcemia (SCH) classification at parturition and to evaluate the association of other cow-level risk factors with SCH classification at calving or at 2 d in milk (DIM). A total of 301 animals from 2 dairy herds located in New York were enrolled in a cohort study. Blood samples were collected at approximately 1 wk before the expected calving date, within 4 h of calving, and at 2 DIM. Prepartum samples had plasma macromineral concentrations (Ca, K, Mg, P), albumin, and β-hydroxybutyrate analyzed. Samples collected at calving were analyzed for Ca only, and samples from 2 DIM had macromineral and albumin concentrations determined. Postpartum SCH was defined as Ca concentrations ≤2.1 mmol/L. The prevalence of SCH at calving was 2, 40, and 66% for first, second, and third or greater parities, respectively. Only 4% of cows could be classified with prepartum subclinical hypomagnesemia (Mg concentrations <0.8 mmol/L), which did not provide enough power to appropriately determine the association of plasma Mg with postpartum Ca concentrations and its effect on SCH classification. Multiparous cows with Ca concentrations ≤2.4 mmol/L in the prepartum period and third or greater parity cows had a higher risk of being categorized as SCH at calving [relative risk (RR) = 1.4 and 1.7, respectively]. The risk of SCH at 2 DIM was associated with the interaction of Ca status at calving and lameness score. Nonlame cows with Ca concentrations ≤2.1 mmol/L (RR = 3.2) and normocalcemic lame cows at parturition (RR = 3.4) were more likely to be SCH at 2 DIM compared with nonlame normocalcemic cows. In conclusion, we identified a prepartum Ca cut-point for identification of cows that are more likely to be classified as SCH at calving. Different risk factors were associated with SCH depending on the timing of diagnosis relative to parturition.     

Production effects and bioavailability of N-acetyl-l-methionine in lactating dairy cows

Two experiments were conducted to investigate the production effects of N-acetyl-l-methionine (NALM; experiment 1) and to estimate its bioavailability (BA) and rumen escape (RE; experiment 2), respectively, in lactating dairy cows. In experiment 1, 18 multiparous Holstein cows were used in a replicated, 3 × 3 Latin square design experiment with three 28-d periods. Treatments were (1) basal diet estimated to supply 45 g/d digestible Met (dMet) or 1.47% of metabolizable protein (MP; control), (2) basal diet top-dressed with 32 g/d of NALM to achieve dMet supply of 2.2% of MP, and (3) basal diet top-dressed with 56 g/d of NALM to achieve dMet supply of 2.6% of MP. The NALM treatments supplied estimated 17 and 29 g/d dMet from NALM, respectively, based on manufacturer's specifications. In experiment 2, 4 rumen-cannulated lactating Holstein cows were used in a 4 × 4 Latin square design experiment with four 12-d periods. A 12-d period for baseline data collection and 4 d for determination of RE of NALM preceded the Latin square experiment. For determination of RE, 30 g of NALM were dosed into the rumen simultaneously with Cr-EDTA (used as a rumen fluid kinetics marker) and samples of ruminal contents were collected at 0 (before dosing), 1, 2, 4, 6, 8, 10, 14, 18, and 24 h after dosing. Rumen escape of NALM was calculated using the estimated passage rate based on the measured Cr rate of disappearance. Bioavailability of abomasally dosed NALM was determined using the area under the curve of plasma Met concentration technique. Two doses of l-Met (providing 7.5 and 15 g of dMet) and 2 doses of NALM (11.2 and 14.4 g of dMet) were separately pulse-dosed into the abomasum of the cows and blood was collected from the jugular vein for Met concentration analysis at 0 (before dosing), 1, 2, 4, 6, 8, 10, 12, 14, 18, and 24 h after dosing. Supplementation of NALM did not affect DMI, milk yield, feed efficiency, or milk protein and lactose concentrations and yields in experiment 1. Milk fat concentration and energy-corrected milk yield decreased linearly with NALM dose. Plasma Met concentration was not affected by NALM dose. The estimated relative BA of abomasally dosed NALM (experiment 2) was 50% when dosed at 14.4 g/cow (11.2 g/d dMet from NALM) and 24% when dosed at 28.8 g/cow (14.4 g/d dMet from NALM). The estimated RE of NALM was 19% based on the measured k_p of Cr at 11%/h. The total availability of ingested NALM was estimated at 9.5% for the lower NALM dose when taking into account RE (19%) and bioavailability in the small intestine (50%). Overall, NALM supplementation to mid-lactation dairy cows fed a MP-adequate basal diet below NRC (2001) recommendations (45 g/d or 1.47% Met of MP) decreased milk fat and energy-corrected milk yields but did not affect milk or milk true protein yields. Further evaluation of BA of NALM at different doses is warranted. In addition, intestinal conversion of NALM to Met needs additional investigation to establish a possible saturation of the enzyme aminoacylase I at higher NALM doses.     

Short communication: Serum osteoprotegerin concentrations in periparturient dairy cows

Dairy cows experience hypocalcemia at the onset of lactation; however, the contribution of bone metabolism to circulating Ca concentrations during the periparturient period is not well understood. Osteoprotegerin (OPG) protects against loss of skeletal mass by regulating osteoclastogenesis. We previously reported that the bone resorption increased around parturition in dairy cows from the perspective of bone markers in blood. The aim of this study was to explore osteoclastic bone resorption in periparturient dairy cows by measuring changes in serum OPG concentrations around calving. The OPG concentrations were evaluated in primiparous (n = 9) and multiparous (n = 9) cows at 21 d precalving and over 5 d after calving. Based on mixed model analyses, the multiparous cows had a higher serum OPG concentration than primiparous cows at precalving and exhibited a significant decline in serum OPG and Ca concentrations after calving. In contrast to multiparous cows, primiparous cows did not exhibit these changes. Therefore, we hypothesized that multiparous cows preserve their bone mineral content during late pregnancy and activate the osteoclastic bone resorption after calving. Based on our serum OPG data, we identified the different periparturient osteoclastic bone resorption patterns in healthy primiparous and multiparous dairy cows.     

Effect of direct-fed microbials on performance, diet digestibility, and rumen characteristics of Holstein dairy cows

The objective of this study was to determine the effect of feeding direct-fed microbial (DFM) products containing Lactobacillus acidophilus and Propionibacteria freudenreichii on the performance, nutrient digestibility, and rumen fermentation of Holstein dairy cows in midlactation. Experiments were conducted from February to May 2003. Cows were fed 1 of 3 dietary treatments: 1) 1 × 10⁹ colony-forming units (cfu)/d of live L. acidophilus strain LA747 and 2 × 10⁹ cfu/d of live P. freudenreichii strain PF24 (DFM1); 2) 1 × 10⁹ cfu/d of live L. acidophilus strain LA747, 2 × 10⁹ cfu/d of live P. freudenreichii strain PF24, and 5 × 10⁸ cfu/d of L. acidophilus strain LA45 (DFM2); or 3) lactose (control). Treatments were administered by mixing 45 g of finely ground corn with 5 g of DFM products or lactose and top dressing on the total mixed rations once daily. All cows received the same total mixed ration: 12.7% alfalfa hay, 46.2% corn silage, and 41.1% concentrate on a dry matter (DM) basis. In study 1 (lactation study), 39 multiparous and 18 primiparous Holstein cows were blocked by parity and randomly assigned to treatments for 84 d. Starting on d 35, fecal grab samples were collected from each cow at 5- to 8-h intervals over 48 h for digestibility measurements. A rumen fermentation study (study 2) was conducted concurrently with the lactation study. Three rumenfistulated, multiparous Holstein cows were randomly assigned to dietary treatments DFM1, DFM2, and control in a 3 × 3 Latin square design with 28-d periods. In study 1, there was no difference in average DM intake (23.9, 23.6, and 24.2 kg/d) or 4% fat-corrected milk (36.8, 35.3, and 36.2 kg/d) for treatments DFM1, DFM2, and control. Percentage or yield of milk components also did not differ among treatments. Feed efficiency averaged 1.52 kg of 4% fat-corrected milk/kg of DM intake and did not differ among treatments. There were no differences in apparent DM, crude protein, neutral detergent fiber, or starch digestibility among treatments. In study 2, there was no difference in rumen pH and concentrations of ammonia or total volatile fatty acids measured at 0, 1, 3, and 6 h after feeding. Under the conditions of these studies, supplementing midlactation cows with DFM products containing L. acidophilus and P. freudenreichii did not affect cow performance, diet digestibility, or rumen fermentation.     

NutriDense corn grain and corn silage for dairy cows

Twenty multiparous Holstein cows, 4 of them surgically fitted with ruminal cannulas, were used in a replicated 4 × 4 Latin square to compare the effects of whole-plant silage and grain produced from NutriDense (ND), leafy NutriDense (LND), or a conventional yellow dent (YD) hybrid on ruminal fermentation, total tract nutrient digestibility, and performance of lactating dairy cows. On a DM basis, diets contained 30.6% corn silage and 27.7% corn grain provided from the 3 hybrids according to the following combinations: 1) YD grain and YD silage, 2) YD grain and LND silage, 3) ND grain and YD silage, and 4) ND grain and LND silage. The average concentrations of crude protein, neutral and acid detergent fiber, and ether extract of LND silage and ND grain were higher, but the contents of nonfibrous carbohydrates and starch were lower than those of their YD counterparts. Although DM intake was similar among treatments, feeding ND grain, LND silage, or both reduced the intakes of nonfibrous carbohydrates and starch but increased the intake of ether extract. Apparent digestibility of starch in the total tract was highest for the diet that contained LND silage and YD grain, whereas the amount and percentage of ether extract that were apparently digested in the total tract was increased and tended to be increased, respectively, by the addition of ND grain, LND silage, or both to the diets. Ruminal fermentation parameters were unaffected by treatments except for the concentration of ammonia nitrogen in the ruminal fluid, which tended to be increased by the feeding of ND grain, LND silage, or both. Production of milk, crude and true protein, fat, lactose, and total solids did not differ among diets. Concentration of milk urea nitrogen increased when the ND grain, LND silage, or both were fed to the cows. Results indicate that ND grain and LND silage were similar to the conventional grain and silage for the feeding of lactating dairy cows.     

Concentration of progesterone during the development of the ovulatory follicle: II. Ovarian and uterine responses

Two experiments evaluated the influence of altering the concentrations of progesterone during the development of the ovulatory follicle on the composition of the follicular fluid, circulating LH and PGF_2α metabolite (PGFM), and expression of endometrial progesterone receptor and estrogen receptor-α. In both experiments, the estrous cycles were presynchronized (GnRH and progesterone insert followed by insert removal and PGF_2α 7 d later, and GnRH after 48 h) and cows were then enrolled in 1 of 2 treatments 7 d later (study d −16): high progesterone (HP) or low progesterone (LP). In experiment 1 (n = 19), cows had their estrous cycle synchronized starting on study d −9 (GnRH and progesterone insert on d −9, and insert removal and PGF_2α on d −2). In experiment 2 (n = 25), cows were submitted to the same synchronization protocol as in experiment 1, but had ovulation induced with GnRH on study d 0. In experiment 1, plasma was sampled on d −4 and analyzed for concentrations of LH; the dominant follicle was aspirated on d 0 and the fluid analyzed for concentrations of progesterone, estradiol, and free and total IGF-1. In experiment 2, follicular development and concentrations of progesterone and estradiol in plasma were evaluated until study d 16. Uterine biopsies were collected on d 12 and 16 for progesterone receptor and estrogen receptor-α protein abundance. An estradiol/oxytocin challenge for PGFM measurements in plasma was performed on d 16. In experiments 1 and 2, LP cows had lower plasma concentrations of progesterone and greater concentrations of estradiol, and had larger ovulatory follicle diameter (20.4 vs. 17.2 mm) at the end of the synchronization protocol than HP cows. Concentration of LH tended to be greater for LP than HP cows (0.98 vs. 0.84 ng/mL). The dominant follicle of LP cows had greater concentration of estradiol (387.5 vs. 330.9 ng/mL) and a lower concentration of total IGF-1 (40.9 vs. 51.7 ng/mL) than that of HP cows. In experiment 2, estradiol and progesterone concentrations did not differ between treatments from d 0 to 16; however, the proportion of cows with a short luteal phase tended to increase in LP than HP (25 vs. 0%). Concentrations of PGFM were greater for LP than HP. Uterine biopsies had a greater abundance of progesterone receptor, and tended to have less estrogen receptor-α abundance on d 12 compared with d 16. An interaction between treatment and day of collection was detected for estrogen receptor-α because of an earlier increase in protein abundance on d 12. Reduced concentrations of progesterone during the development of the ovulatory follicle altered follicular dynamics and follicular fluid composition, increased basal LH concentrations, and prematurely increased estrogen receptor-α abundance and exacerbated PGF_2α release in the subsequent estrous cycle.     

Persistency of methane mitigation by dietary nitrate supplementation in dairy cows

Feeding nitrate to dairy cows may lower ruminal methane production by competing for reducing equivalents with methanogenesis. Twenty lactating Holstein-Friesian dairy cows (33.2 ± 6.0 kg of milk/d; 104 ± 58 d in milk at the start of the experiment) were fed a total mixed ration (corn silage-based; forage to concentrate ratio 66:34), containing either a dietary urea or a dietary nitrate source [21 g of nitrate/kg of dry matter (DM)] during 4 successive 24-d periods, to assess the methane-mitigating potential of dietary nitrate and its persistency. The study was conducted as paired comparisons in a randomized design with repeated measurements. Cows were blocked by parity, lactation stage, and milk production at the start of the experiment. A 4-wk adaptation period allowed the rumen microbes to adapt to dietary urea and nitrate. Diets were isoenergetic and isonitrogenous. Methane production, energy balance, and diet digestibility were measured in open-circuit indirect calorimetry chambers. Cows were limit-fed during measurements. Nitrate persistently decreased methane production by 16%, whether expressed in grams per day, grams per kilogram of dry matter intake (DMI), or as percentage of gross energy intake, which was sustained for the full experimental period (mean 368 vs. 310 ± 12.5 g/d; 19.4 vs. 16.2 ± 0.47 g/kg of DMI; 5.9 vs.4.9 ± 0.15% of gross energy intake for urea vs. nitrate, respectively). This decrease was smaller than the stoichiometrical methane mitigation potential of nitrate (full potential = 28% methane reduction). The decreased energy loss from methane resulted in an improved conversion of dietary energy intake into metabolizable energy (57.3 vs. 58.6 ± 0.70%, urea vs. nitrate, respectively). Despite this, milk energy output or energy retention was not affected by dietary nitrate. Nitrate did not affect milk yield or apparent digestibility of crude fat, neutral detergent fiber, and starch. Milk protein content (3.21 vs. 3.05 ± 0.058%, urea vs. nitrate respectively) but not protein yield was lower for dietary nitrate. Hydrogen production between morning and afternoon milking was measured during the last experimental period. Cows fed nitrate emitted more hydrogen. Cows fed nitrate displayed higher blood methemoglobin levels (0.5 vs. 4.0 ± 1.07% of hemoglobin, urea vs. nitrate respectively) and lower hemoglobin levels (7.1 vs. 6.3 ± 0.11 mmol/L, urea vs. nitrate respectively). Dietary nitrate persistently decreased methane production from lactating dairy cows fed restricted amounts of feed, but the reduction in energy losses did not improve milk production or energy balance.