Effects of additional gonadotropin-releasing hormone and prostaglandin F2α treatment to an estradiol/progesterone-based embryo transfer protocol for recipient lactating dairy cows.

This study was designed to evaluate whether the utilization of a second PGF2α treatment at the end of an estradiol/progesterone (E2/P4)-based protocol with or without GnRH at the beginning of the protocol would improve pregnancy rates of lactating Holstein cows assigned to timed embryo transfer. A total of 501 lactating Holstein cows in 5 farms were enrolled in the experiment. Within farm, cows were blocked by parity and, within block, were assigned randomly to (1) insertion of an intravaginal P4 device (controlled internal drug-releasing device; CIDR) and estradiol benzoate on d -11, PGF2α on d -4, CIDR withdrawal and an injection of estradiol cypionate on d -2, and timed embryo transfer on d 7 (1-PGF; n = 164); (2) the same treatments as 1-PGF, but with PGF2α administered on d -4 and -2 (2-PGF; n = 171); and (3) 2-PGF with the addition of a GnRH treatment on d -11 (GnRH+2-PGF; n = 166). Ovaries were scanned by transrectal ultrasonography on d -11, -4, and 7, and blood samples were collected on d -11, -4, 0, and 7 for P4 determination. Treatment comparisons were performed using contrasts. The proportion of cows with a new corpus luteum on d -4 was greater in GnRH+2-PGF cows. Cows in 1-PGF had a greater P4 concentration on d 0 but lesser P4 on d 7 compared with cows in the other groups. Cows assigned to receive 2-PGF (2-PGF and GnRH+2-PGF) had greater estrus expression, and a greater proportion of cows ovulated to estradiol cypionate. No further contrast effects were observed for follicle diameter, double ovulation rate, pregnancy per embryo transfer (P/ET) on d 32 and 60, or pregnancy loss. As P4 concentration on d -4 increased, P/ET on d 60 tended to increase. Cows with P4 ≥3.66 ng/mL on d -4 had greater P/ET on d 32 and 60 than those with P4 below that threshold. Regardless of treatment, cows with P4 concentration ≥3.66 ng/mL also had a greater pregnancy per synchronized protocol (P/SP) on d 60. Also, a P4 concentration on d -4 (low or high) × follicle diameter (continuous) interaction tendency was observed when evaluating P/ET. Although P/ET did not differ among cows with different follicles sizes with reduced P4 concentration on d -4 (<3.66 ng/mL), it increased in cows with larger follicles exposed to increased P4 concentration (≥3.66 ng/mL). When P4 on d 0 was evaluated, P/ET on d 32 and 60 was greater for cows with low (≤0.09 ng/mL) versus high (>0.21 ng/mL) P4; as P4 concentration on d 0 increased, P/ET linearly decreased. In summary, cows with increased P4 concentrations during growth of the ovulatory follicular wave had improved P/ET. Administering a second PGF2α dose reduced P4 concentration on d 0 and increased ovulatory response to the protocol, but no benefits were observed on P/ET or P/SP.

Proposal of an iELISA for Mycoplasma bovis diagnosis in dairy cattle and associated risk factors / [Proposta de um iELISA para diagnóstico de Mycoplasma bovis em bovinos leiteiros e fatores de risco associados]

Mycoplasma bovis is a highly contagious agent associated with several pathologies in cattle. The detection of reactive antibodies to M. bovis by Indirect Enzyme-Linked Immunosorbent Assay (iELISA) identifies if there was an exposure to the microorganism. The current study aimed to optimize an iELISA from M. bovis total cell antigen, applying it to bovine serum samples, and to evaluate risk factors. Serum samples were obtained from 400 cows from 17 herds from Southeast Brazil. In the optimization of iELISA, the following was established: 2 µg/mL of antigen, sera dilution 1:300, and conjugate dilution 1:15000. The frequency was 62.3% (249/400) of reactive animals and 100% (17/17) of reactive herds. Risk factors were: herds with more than 100 animals (OR= 3.1; CI= 95%); Holstein breed (OR= 72.5; CI= 95%); cows (OR= 29.7; CI= 95%); intensive breeding system (OR= 3.3; CI= 95%); associated small ruminant production (OR= 4.4; CI= 95%); milk production above 500L (OR= 2.9; CI= 95%); no quarantine (OR= 1.5; CI= 95%); mechanical milking (OR= 5.5; CI= 95%) and cases of mastitis (OR= 5.5; CI= 95%). The proposed iELISA was able to detect antibodies reactive to M. bovis in bovine serum. Knowledge of these risk factors can assist in the implementation of prophylactic measures.(AU)

Mycoplasma bovis é um agente altamente contagioso relacionado a várias patologias em bovinos. A detecção de anticorpos reativos a M. bovis por Ensaio de Imunoadsorção Enzimática Indireto (iELISA) identifica se houve exposição ao microrganismo. O presente estudo teve como objetivo otimizar um iELISA de antígeno celular total de M. bovis, aplicando-o a amostras de soro bovino, bem como avaliar fatores de risco. Amostras de soro foram obtidas de 400 vacas de 17 rebanhos da Região Sudeste do Brasil. Na otimização do iELISA foram obtidos: 2µg/mL de antígeno, diluição dos soros 1:300 e do conjugado 1:15000. A frequência de animais reativos foi de 62,3% (249/400) e de 100% (17/17) para os rebanhos. Os fatores de risco foram: rebanhos com mais de 100 animais (OR= 3,1; IC= 95%); raça Holandesa (OR= 72,5;IC= 95%); vacas (OR= 29,7;IC= 95%); sistema intensivo (OR= 3,3; C= 95%); produção de pequenos ruminantes (OR= 4,4;IC=95%); produção de leite acima de 500L (OR= 2,9;IC= 95%); sem quarentena (OR= 1,5;IC= 95%); ordenha mecânica (OR= 5,5;IC= 95%) e casos de mastite (OR= 5,5;IC= 95%). O iELISA proposto foi capaz de detectar anticorpos reativos a M. bovis no soro bovino. O conhecimento desses fatores de risco pode auxiliar na implementação de medidas profiláticas.(AU)

Somatic cell count and type of intramammary infection impacts fertility from in vitro produced embryo transfer.

The objective of this study was to assess the impact of mastitis-causing bacteria and somatic cell count (SCC) on pregnancy per embryo transfer (P/ET) in Holstein-Gir crossbred (Girolando) lactating dairy cows. Cows (n = 1397) were subjected to a timed-embryo transfer protocol. Milk samples were collected two days before embryo transfer for SCC and bacteriological culture analyses. Pregnancy diagnosis was performed on days 31 and 66 after timed-embryo transfer. The animals were grouped according to the National Mastitis Council recommendations: Gram-positive environmental (EV+), Gram-negative environmental (EV-), Gram-positive contagious (C+), coagulase-negative staphylococci (CNS) and control (no bacterial growth). Additional analysis was made by categorizing bacteria based on degree of pathogenicity (Major or Minor). Bacterial growth reduced P/ET (P < .01) at both 31 and 66 days of gestation. The P/ET was lower (P < .05) at 31 days in EV- (30.1%) and EV+ (29.9%) groups and tended (P = .09) to be lower in the C+ group (36.6%) than the control group (44.0%). The P/ET from the Major group at 31 days of gestation was lower (P = .03) compared with the Minor and control groups (32.1 vs 41.1 vs 43.2%, respectively). Cows with SCC > 400,000 cells/mL had lower P/ET (P < .01) than animals with SCC < 200,000 cells/mL at both 31 (30.4% vs 40.8%) and 66 days (24.7% vs 32.2%) of gestation. Pregnancy loss was not different between bacterial isolates and SCC categories. Elevated SCC significantly reduced P/ET, whereas environmental agents and those with Major pathogenicity yielded the greatest reduction in P/ET.

Effect of adding a gonadotropin-releasing-hormone treatment at the beginning and a second prostaglandin F2α treatment at the end of an estradiol-based protocol for timed artificial insemination in lactating dairy cows during cool or hot seasons of the year.

Our hypothesis was that fertility could be increased in a timed artificial insemination (TAI) protocol based on estradiol (E2) and progesterone (P4) by combining GnRH with E2-benzoate at the start of the protocol to increase circulating P4 during preovulatory follicle development and by using 2 prostaglandin F2α (PGF) treatments at the end to decrease P4 near TAI. Lactating Holstein cows (n=1,808) were randomly assigned during the cool or hot season of the year to receive TAI (d 0) following 1 of 3 treatments: (1) control: controlled internal drug-release insert + 2mg of E2-benzoate on d -11, PGF on d -4, controlled internal drug-release insert withdrawal + 1.0mg of E2-cypionate on d -2, and TAI on d 0; (2) 2PGF: identical to control protocol with addition of a second PGF treatment on d -2; (3) GnRH: identical to 2PGF protocol with addition of a 100-µg GnRH treatment on d -11. Pregnancy diagnoses were performed on d 32 and 60 after TAI. Season had major effects on many reproductive measures, with cool season greater than hot season in percentage of cows with corpus luteum (CL) at PGF (62.9 vs. 56.2%), ovulatory follicle diameter (15.7 vs. 14.8mm), expression of estrus (86.7 vs. 79.9%), ovulation following the protocol (89.7 vs. 84.3%), and pregnancies per artificial insemination (P/AI; 45.4 vs. 21.4%). The GnRH protocol increased percentage of cows with CL (control=56.9%; 2PGF=55.8%; GnRH=70.5%) and P4 at PGF (control=3.28±0.22; 2PGF=3.35±0.22; GnRH=3.70±0.21ng/mL), compared with control and 2PGF protocols. The GnRH protocol increased P/AI at the pregnancy diagnosis at 32d [37.3% (219/595)] and 60d [31% (179/595)] after TAI, compared with control [30.0% (177/604); 25.1% (145/604)], with intermediate results with 2PGF protocol [33.2% (196/609); 28.0% (164/609)]. The positive effects of GnRH treatment on P/AI were only detected during the cool season (GnRH=50.9%; 2PGF=44.2%; control=41.0%) and not during the hot season. In addition, the effect of GnRH was only observed in cows with low P4 (<3ng/mL) at the start of the protocol and not in cows that began the protocol with high P4. Furthermore, presence of CL at PGF interacted with follicle diameter such that cows with a CL at PGF had greater P/AI if they ovulated larger rather than smaller follicles near TAI. Thus, fertility to TAI can be improved by inducing ovulation at the beginning of an E2/P4-based protocol using GnRH treatment, particularly during the cool season of the year and in cows with low P4 at the start of the protocol.

The slope of the VO2 slow component is associated with exercise intolerance during severe-intensity exercise.

The aim of this study was to analyze the relationship between the slope of the VO2 slow component (VO2sc) and exercise tolerance (tlim) during constant-work-rate (CWR) exercise performed within the severe intensity domain. Fifteen active subjects (VO2max = 41.2 ± 5.1 ml.kg-1.min-1) performed the following tests: 1) an incremental test to determine the VO2max and the work rate associated with the VO2max (IVO2max) and; 2) two CWR transitions at 95% of the IVO2max to determine the slope of the VO2 slow component and the tlim. Three tlims were obtained: tlim1 = CWR1; tlim2 = CWR2; and tlim1+2 = (CWR1 + CWR2) / 2. There was no significant difference between the VO2max (3271.7 ± 410.7 mL·min-1) and VO2peak obtained during the CWR tests (CWR1 = 3356.3 ± 448.8 mL·min-1, CWR2 = 3362.2 ± 393.4 mL·min-1, p > 0.05). Significant correlations (p < 0.05) were found among the VO2sc kinetics and tlim1 (r = -0.53), tlim2 (r = -0.49) and tlim1+2 (r = -0.55). Thus, exercise tolerance during CWR performed within the severe intensity domain is partially explained by the slope of the VO2 slow component.

Consumo de oxigênio durante ciclismo na máxima fase estável de lactato sanguíneo até a exaustão: modelo contínuo vs. intermitente / Consumo de oxígeno durante ciclismo en la máxima fase estable de lactato sanguíneo hasta el agotamiento: modelo continuo vs. intermitente / Oxygen uptake during cycling at maximal lactate steady state to exhaustion: continuous vs. intermittent model

Objetivo. Analisar o consumo de oxigênio (VO2) acumulado e o tempo em que o VO2 é mantido (TMcg) em elevados valores durante o exercício realizado na máxima fase estável de lactato (MLSS) determinada de forma contínua (MLSScon) e intermitente (MLSSint) até à exaustão. Método. Catorze ciclistas treinados realizaram um teste incremental máximo no cicloergômetro; 2‐5 testes contínuos e 2‐4 testes intermitentes, para determinação da MLSScon e MLSSint, respectivamente; por fim mais 2 testes até a exaustão (TTE) (contínuo e intermitente), conduzidos nas respectivas cargas de MLSS. Resultados. O TTE contínuo foi significativamente menor do que o TTE intermitente (54,7 ± 10,9 vs. 67,8 ± 14,3, respectivamente; p = 0,001). O VO2 da carga (VO2cg) foi significantemente maior na condição intermitente (p < 0,05), em ambas as situações (MLSS e TTE). Por outro lado, o TMcg e o VO2 acumulado na carga (VO2ACcg) foram maiores na condição contínua (p < 0,0001), em ambas as situações. O total de VO2 acumulado (VO2ACt) foi similar entre as condições contínuas e intermitentes (104,7 ± 8,7 L vs. 103,9 ± 41,1 L para a MLSS e 193,8 ± 41,1 L vs. 226,4 ± 53,0 L para o TTE, respectivamente). Conclusão. Os exercícios contínuo e intermitente realizados com a mesma duração ou até a exaustão, e em condições metabólicas similares, apresentam valores de VO2ACt similares, embora o TMcg e o VO2ACcg sejam inferiores no exercício intermitente (AU)

Objetivo. Analizar el consumo de oxígeno (VO2) acumulado y el tiempo en que el VO2 es mantenido (TMcg) en valores elevados, durante el ejercicio realizado en la máxima fase estable de lactato (MLSS), determinada de forma continua (MLSScon) e intermitente (MLSSint) hasta el agotamiento. Método. Catorce ciclistas entrenados realizaron un test incremental máximo en el cicloergómetro; 2‐5 test continuos y 2‐4 test intermitentes, para determinación de la MLSScon y MLSSint, respectivamente; por último realizaron 2 test hasta el agotamiento (TTE) (continuo e intermitente), a las respectivas cargas de MLSS. Resultados. El TTE continuo fue significativamente menor que el TTE intermitente (54,7 ± 10,9 vs. 67,8 ± 14,3, respectivamente; p = 0,001). El VO2 de la carga (VO2cg) fue significativamente mayor en la condición intermitente (p < 0,05), en ambas situaciones (MLSS y TTE). Por otro lado, el TMcg y el VO2 acumulado en la carga (VO2ACcg) fueron mayores en la condición continua (p < 0,0001), en ambas las situaciones. El total de VO2 acumulado (VO2ACt) fue similar entre las condiciones continuas e intermitentes (104,7 ± 8,7 L vs. 103,9 ± 41,1 L para a MLSS y 193,8 ± 44,1 L vs. 226,4 ± 53,0 L para el TTE, respectivamente). Conclusión. Los ejercicios continuo e intermitente realizados con la misma duración o hasta el agotamiento, y en condiciones metabólicas similares presentan valores de VO2ACt similares, aunque el TMcg y el VO2ACcg sean inferiores en el ejercicio intermitente (AU)

Objective. The aim of this study was to analyse the accumulated oxygen uptake (VO2), as well the maintained time (TMcg) at high levels during a maximal lactate steady state (MLSS) exercise, determined in continuous (MLSScon) and intermittent (MLSSint) modes, until exhaustion. Method. Fourteen trained cyclists performed an incremental maximal testing; 2‐5 continuous and 2‐4 intermittent constant workload trials, in order to identify the MLSScon and MLSSint, respectively; two tests until exhaustion (TTE) (continuous and intermittent) were conducted using their respective MLSS workloads. Results. The continuous TTE was significantly lower than TTE at intermittent protocol (54.7 ± 10.9 vs. 67.8 ± 14.3 min; p = 0.001). The VO2 at target workload (VO2cg) was higher at intermittent exercise (p < 0.05), at both situations (MLSS and TTE). On the other hand, TMcg and the VO2 accumulated in MLSS workloads (VO2ACcg) were higher during continuous mode (p < 0.0001), at both situations. The total VO2 accumulated (VO2ACt) was similar between intermittent and continuous cycling (MLSS: 104.7 ± 8.7 L vs. 103.9 ± 41.1 L and TTE: 193.8 ± 41.1 L vs. 226.4 ± 53.0 L, respectively). Conclusion. When MLSS continuous and intermittent were performed with same duration or until exhaustion, the VO2ACt was similar, although the TMcg and VO2ACcg were smaller during intermittent exercise (AU)

Effect of feeding according to energy balance on performance, nutrient excretion, and feeding behavior of early lactation dairy cows.

The objectives of this study were to evaluate 2 feeding strategies for early lactation cows on performance and efficiency of nutrient utilization. Fifty-eight Holsteins cows were blocked by parity and production during the pretreatment period and then randomly assigned at 21 d postpartum to a control diet [n=29; 16.2% crude protein, 1.64 Mcal of net energy for lactation (NE(L)), 22% starch, and 19% forage neutral detergent fiber (NDF)] or a diet with caloric density manipulated weekly (precision diet; n=29; 16.2% crude protein; 1.59 to 1.68 NE(L); 18 to 26% starch; and 16 to 22% forage NDF) to promote a calculated positive energy balance of 5 Mcal/day. Diets were fed as total mixed rations and precision cows had their diets adjusted individually once a week, by feeding additional grain supplementation from 0 to 25% of daily dry matter (DM) offered, according to the energy balance of the preceding week. Energy balance was calculated daily and then averaged weekly. The study lasted from wk 3 to 19 postpartum, and nutrient digestibility, rumen fluid composition, urinary output, estimates of microbial protein synthesis, and feeding behavior were evaluated between wk 9 and 13 postpartum. Compared with controls, precision cows had similar DM intake (24.3 kg/d), but NE(L) intake tended to be greater primarily between wk 4 and 8 postpartum. Yields of milk (45.2 vs. 41.9 kg/d), milk components, 3.5% fat-corrected milk (44.0 vs. 40.8 kg/d), and energy-corrected milk (43.4 vs. 40.2) were all greater for precision than control cows, resulting in greater energy-corrected milk production per kilogram of diet DM consumed (1.79 vs. 1.72). Precision cows produced more milk calories per kilogram of metabolic weight (0.227 vs. 0.213 Mcal of NE(L)/kg), although the amount of consumed calories partitioned into milk (82.3%) and measures of energy status did not differ between treatments throughout the study. Glucose concentrations were greater throughout the day in precision cows compared with controls at 6 wk, but not 13 wk postpartum. Apparent digestibility of nutrients, composition of rumen fluid, mean and low rumen pH, and estimated rumen microbial N synthesis remained mostly unaltered by treatments. Although precision cows produced more milk true protein, measures of efficiency of dietary N use were not influenced by treatment. On wk 13 postpartum, precision cows consumed a diet with longer NDF particles, which resulted in a tendency for greater intake of NDF >8mm because of less sorting against the long particles than control cows. Meal pattern differed with treatment, and precision cows consumed feed more sparsely throughout the day, spent more time ruminating lying, and had similar meal duration (mean of 36.3 min/meal) compared with control cows, but smaller meal size (3.33 vs. 3.64 kg/meal). Results from the current study indicate that allocating dietary resources according to the individual needs of cows based on energy balance improves lactation performance compared with feeding a single total mixed ration, despite similar average nutrient intake between treatments. Improvements in performance are likely related to allocation of calories based on the needs of the cow and on shifts of feeding behavior that might favor intake of smaller meals.

Targeted progesterone supplementation improves fertility in lactating dairy cows without a corpus luteum at the initiation of the timed artificial insemination protocol.

The objectives of this study were to determine the effect of supplemental progesterone on fertility in lactating dairy cows lacking a corpus luteum (CL) at the initiation of the timed artificial insemination (AI) program. Holstein cows were subjected to the 5-d timed AI program (d -8 GnRH, d -3 and -2 PGF2α, d 0 GnRH and AI). Cows had their ovaries scanned by ultrasonography on d -8 and those bearing a CL were considered to be in diestrus (DI; n=946). Cows that lacked a CL on d -8 were assigned to remain as untreated control (CON; n=234) or receive 2 controlled internal drug release (CIDR) inserts containing progesterone (2CIDR; n=218) from d -8 to -3, as a single insert has been proven insufficient to modulate fertility in cows without CL. Blood was analyzed for progesterone and estradiol concentrations. Pregnancy was diagnosed on d 34 and 62 after AI. Progesterone concentrations during the timed AI program were lowest for CON, intermediate for 2CIDR, and highest for DI. Supplementation increased progesterone concentrations between d -7 and -3 compared with CON (2.65 vs. 0.51 ng/mL). Ovulation to the first GnRH was not affected by treatment. However, a greater proportion of CON and 2CIDR cows had a new CL on d -3 compared with DI cows (66.7 vs. 61.9 vs. 52.0%). In cows with a new CL, the diameter of the ovulatory follicle was larger for CON than 2CIDR, and intermediate for DI (18.7 vs. 16.5 vs. 17.7 mm). Concentrations of estradiol on d -3 did not differ among treatments; however, DI cows had greater estradiol concentrations at AI compared with CON or 2CIDR cows. Pregnancy per AI was less for CON compared with 2CIDR or DI on d 32 (30.8 vs. 46.8 vs. 49.9%) and 64 (28.6 vs. 43.7 vs. 47.3%), indicating that supplementation with progesterone reestablished fertility in cows lacking a CL on d -8. A greater proportion of nonpregnant CON cows had a short reinsemination interval compared with 2CIDR or DI (11.1 vs. 3.5 vs. 5.7%). Treatment did not affect pregnancy loss between d 34 and 62 of gestation. A single ultrasound exam was effective in identifying a low-fertility cohort of cows based on the absence of CL at the first GnRH injection of the timed AI protocol. Progesterone supplementation with 2 CIDR inserts increased progesterone in plasma to 2.65 ng/mL and restored fertility in lactating dairy cows lacking a CL at the initiation of the timed AI program similar to that of cows in diestrus.