Systemic changes caused by artificial insemination in beef cows (Bos indicus) and their impact on animal welfare / Alterações sistêmicas causadas pela inseminação artificial em vacas de corte (Bos indicus) e seu impacto no bem-estar animal

This study aimed to verify if the process of artificial insemination (AI) characterized here as animal immobilization, the passage of the semen applicator through the cervix, and deposition of the semen in the uterus, affected cows' welfare. For this, 18 beef calved cows were selected and divided into two groups: inseminated cows (AIG, n = 9), and not inseminated cows, the control group (CG, n = 9). Body condition score, uterus, and ovary evaluation were performed. Later, both groups were submitted into an estrus synchronization protocol and only the AIG group was inseminated. Blood components of urea, creatinine, AST, GGT, CK, glucose, triglycerides, cholesterol, HDL, LDL, VLDL, NEFA, BHB, cortisol, estradiol, progesterone, albumin, and total protein were measured 30 h before AI, and 4, 24, 48 and 168 h after AI. Statistical differences were considered when P <0.05. No differences between AIG and CG were observed. On the other hand, when the moment of insemination was evaluated, differences were observed for urea, creatinine, AST, GGT, CK, glucose, triglycerides, NEFA, BHB, albumin, and total protein. There was an oscillation of metabolic profiles depending on the time and procedures to which animals were exposed, even though it could be inferred that the AI process was incapable of altering those metabolic components on animals that were inseminated. Still, we can affirm that artificial insemination cannot be categorized as a negative reproduction tool on animal welfare. However, the containment and management procedures for AI may alter the metabolic profile of cows, especially the increase of CK.(AU)

O objetivo deste estudo foi verificar se o processo de inseminação artificial (IA) caracterizado como imobilização do animal, passagem do aplicador de sêmen pelo colo do útero e deposição do sêmen no útero, afetou o bem-estar de bovinos. Para tanto, foram selecionadas 18 vacas de corte paridas, divididas em dois grupos: grupo de animais inseminados (AIG, n = 9) e grupo de animais não inseminados, grupo controle (GC, n = 9). Foram avaliados o escore de condição corporal, útero e ovário. Posteriormente, ambos os grupos foram submetidos a um protocolo de sincronização de cio e apenas o grupo AIG foi inseminado. Componentes metabólicos como ureia, creatinina, AST, GGT, CK, glicose, triglicerídeos, colesterol, HDL, LDL, VLDL, NEFA, BHB, cortisol, estradiol, progesterona, albumina e proteína total foram mensurados 30 horas antes da IA e 4, 24, 48 e 168 horas após a IA. Diferenças estatísticas foram consideradas quando P <0,05. Não foram observadas diferenças entre os dois grupos, por outro lado, quando o momento da inseminação foi avaliado, diferenças foram observadas para ureia, creatinina, AST, GGT, CK, glicose, triglicerídeos, NEFA, BHB, albumina e proteína total. Houve uma variação dos perfis metabólicos em função do tempo e dos procedimentos que os animais foram submetidos, embora pode-se inferir que o processo de IA não foi capaz de alterar esses componentes metabólicos os animais inseminados. Ainda assim, observou-se que o processo de IA não foi categorizado como uma ferramenta negativa de reprodução com relação ao bem-estar animal. Porém, ainda assim, os procedimentos de contenção e manejo da IA podem alterar o perfil metabólico das vacas, principalmente o aumento da CK.(AU)

Assessment of the usage of biodegradable polymeric matrix in vaginal devices to sustain progesterone release in cows.

The usage of timed artificial insemination (TAI) at a low cost leading to better reproductive rates has been the aim of several research groups in the field. Usually during TAI protocols, sustained progesterone (P4 ) release devices are employed. Most devices are constituted of a nylon skeleton covered with a silicon layer with P4 . A device based on biopolymers was developed in order to reduce costs and decrease its environmental impact. In this study, we compared the kinetics of sustained progesterone release among devices manufactured with a polymeric blend made of polyhydroxybutyrate-valerate (PHBV) and poly-ε-caprolactone (PCL) (DISP) which were compared with DIB® (Internal Bovine Device) used as the control. In the in vitro and in vivo progesterone release tests, two types of biopolymer-based devices with a superficial area of 147 cm2 were used: DISP8 (46% PHBV, 46% PCL and 8% P4 ; n = 4), DISP10 (45% PHBV, 45% PCL, 10% P4 ; n = 4) and DIB® (1 g P4 , 120 cm2 area; n = 3). The in vitro tests were carried out according to USP XXIII specifications and were performed in a dissolutor sink using an alcohol/water mixture (60/40 v/v) as a release media and samples were collected at 2 min, 2, 4, 8, 12, 24, 48, 60, 72, 84 and 96 h. P4 concentrations were measured through spectrophotometry in a 244 nm long wave. Three to 3 comparisons of angular coefficients of the straight lines obtained by regression analysis of accumulated P4 concentrations as a function of square root of time were carried out. Furthermore, the diffusion coefficient values of P4 were also determined for DISP8 and DISP10. The results showed that the concentrations of P4 were higher in the DISP10 (774.63 ± 45.26 µg/cm2 /t1/2 ) compared to DISP8 (566.17 ± 3.68 µg/cm2 /t1/2 ) (P < 0.05). However, both DISP10 and DISP8 P4 concentrations did not differ from DIB® (677.39 ± 16.13 µg/cm2 /t1/2 ). For the analysis of released quantities per day of the in vitro test, four periods were considered: 0-24, 24-48, 48-72 and 72-96 h. In the first 24 h, DISP8 released significantly less P4 than DISP10 or DIB®, which did not differ among them. Between 24 and 48 h, DISP10 released significantly more P4 than DIB®. DISP8 released an intermediate P4 amount and did not differ significantly from DIB® or DISP10. Between 48 and 72 h, P4 quantity released by DISP10 was significant higher (P < 0.01) than that of DIB® and DISP8, which did not differ among themselves. Between 72 and 96 h, DISP10 released significantly more P4 than DIB®, and DISP8 released an intermediate amount which did not differ from DIB® or DISP10 (P < 0.01). There was interaction between treatment and time (P = 0.0024). The diffusion coefficient values were: 1.36 × 10-8 (cm2 /s) for DISP10 and 1.12 × 10-8 (cm2 /s) for DISP8. For the in vivo test, ovariectomized crossbred cows received DIB® (n = 4) or DISP8 (n = 8) in an alternate design with a non-balanced sequence (cross-over) added of measures repeated in time referring to 16 days of blood samples collection. Samples were analyzed through radioimmunoassay in solid phase using the commercial kit of DPC (Diagnostics Products Corporation). Plasma concentrations of P4 peaked at 4 h after the placement of the device, this being the only time in which plasma P4 concentrations differed between DIB® (11.45 ± 1.96) compared with DISP8 (9.23 ± 1.15 ng/mL) (P = 0.027). On day 8, plasma P4 concentrations were similar for DIB® (2.44 ± 0.09) and DISP8 (1.89 ± 0.13 ng/mL) (P = 0.58) showing that both devices were able to keep P4 concentrations above 1 ng/mL in the plasma of the cow during the 16 day in vivo test. In conclusion, devices manufactured with the blend of PHBV/PCL biopolymers can sustain the release of P4 in a similar manner as silicon.

Use of FSH in two different regimens for ovarian superstimulation prior to ovum pick up and in vitro embryo production in Holstein cows.

We aimed with the present study to evaluate the effects of FSH treatment (200 mg) split in four or six administrations on ovarian follicle stimulation and in vitro oocyte competence for embryo production in dairy cows with synchronized follicular wave emergence. On random days of the estrous cycle (Day 0), non-lactating Holstein cows received a progesterone (P4)-releasing intravaginal device and 2 mg estradiol benzoate IM. On Day 3, they received 0.530 mg sodium cloprostenol (PGF2α) IM. Control cows (n = 35) received no further treatments, whereas FSH-treated cows received 200 mg FSH split in four (FSH4 group; n = 33) or six (FSH6 group; n = 33) administration regimens. Starting on Day 4, cows in FSH4 group received 200 mg FSH split in four equivalent doses of 50 mg 12 h apart. Cows in FSH6 group received the same total FSH dose split in six equivalent doses of 33.3 mg 12 h apart, but treatments started on Day 3. On Day 7 AM (36 h of "coasting" period for FSH-treated groups), the P4 devices were removed and cows were subjected to ovum pick up (OPU). Viable oocytes were in vitro fertilized using sexed-sorted semen. Although FSH treatment did not (P > 0.1) increase the total number of follicles (Control, 53.2 ± 4.5 vs. FSH-treated, 51.4 ± 3.1), the two hormonal stimulation regimens, FSH4 and FSH6, increased the number of medium follicles (6-10 mm; 5.2 ± 0.5 vs. 18.1 ± 1.4; P < 0.0001) and reduced the number of small follicles (2-5.9 mm; 46.3 ± 5.1 vs. 31.0 ± 2.4 P < 0.0001). Also, FSH treatment or regimen did not increase (P > 0.1) the number of viable oocytes (Control, 12.6 ± 1.26 vs. FSH-treated, 12.70 ± 1.03), recovery rate (Control, 36.5% vs. FSH-treated, 36%) and the number of in vitro produced blastocyst (Control, 4.1 ± 0.52 vs. FSH-treated 4.3 ± 0.5). We concluded that FSH stimulation protocol proposed herein is effective to stimulate the growth of small antral follicle population prior to OPU, but it was ineffective to improve in vitro oocyte competence for embryo production in non-lactating Holstein cows with synchronized follicular wave emergence.

Uterine lavage is efficient to recover endometrial cytology sample and does not interfere with fertility rate after artificial insemination in cows.

Productivity rates directly depend on the fertility of a herd, which in turn can be influenced by many factors. Semen deposited in the female reproductive tract is foreign to the body and, in response to this invasion, produces an inflammatory reaction, which is characterized by rapid infusion of polymorphonuclear (PMN) cells. Techniques to obtain an endometrial sample are usually invasive and can mask the true inflammatory response. Ultrasound is a noninvasive technique and can contribute to the diagnosis of postartificial insemination (AI) inflammatory response in cattle. The present study was divided into two experiments. The aim of experiment 1 was to compare two methods of endometrial cytology collection, uterine cytobrush (UC) and uterine lavage (UL), and their effects on uterine hemodynamics that provide information about blood flow. The two methods were evaluated by Doppler ultrasound using the spectral and color modes. For that purpose, 19 Nellore cows were synchronized for timed AI and subjected to UC (n = 9) or UL (n = 10). The techniques were performed 4 hours after AI. The results showed that both techniques allow collection of a good quality sample and with enough PMN cells to perform counting. More PMN cells were obtained by UL than UC. There was no difference in uterine blood flow between the UC and UL groups in any of the periods evaluated (34 hours before and 4, 24, and 48 hours after collection of uterine sample). On the basis of results of experiment 1, the effect of UL on fertility was studied in experiment 2. A total of 128 Nellore cows were synchronized for TAI; 35 cows were subjected to endometrial cytology by UL 4 hours after AI, and 93 were not submitted to any procedure (control). Pregnancy diagnosis was performed by transrectal ultrasound 30 days after AI. Pregnancy rates did not differ between UL (54.29%) and control (56.99%) groups. The results of this study showed that UL allows the collection of more representative cells of the surface of the uterus than UC technique and causes no damage to the reproductive tract. Moreover, UL did not affect pregnancy rate when performed 4 hours after AI.

Follicle and corpus luteum size and vascularity as predictors of fertility at the time of artificial insemination and embryo transfer in beef cattle / Tamanho e vascularização do folículo e corpo lúteo como preditores de fertilidade após inseminação artificial e transferência de embriões em gado de corte

Two ultrasound based fertility prediction methods were tested prior to embryo transfer (ET) and artificial insemination (AI) in cattle. Female bovines were submitted to estrous synchronization prior to ET and AI. Animals were scanned immediately before ET and AI procedure to target follicle and corpus luteum (CL) size and vascularity. In addition, inseminated animals were also scanned eleven days after insemination to target CL size and vascularity. All data was compared with fertility by using gestational diagnosis 35 days after ovulation. Prior to ET, CL vascularity showed a positive correlation with fertility, and no pregnancy occurred in animals with less than 40% of CL vascularity. Prior to AI and also eleven days after AI, no relationship with fertility was seen in all parameters analyzed (follicle and CL size and vascularity), and contrary, cows with CL vascularity greater than 70% exhibit lower fertility. In inseminated animals, follicle size and vascularity was positive related with CL size and vascularity, as shown by the presence of greater CL size and vascularity originated from follicle with also greater size and vascularity. This is the first time that ultrasound based fertility prediction methods were tested prior to ET and AI and showed an application in ET, but not in AI programs. Further studies are needed including hormone profile evaluation to improve conclusion.

Duas técnicas de predição de fertilidade, baseadas em ultrassonografia, foram testadas no momento da transferência de embriões (TE) e inseminação artificial (IA) em bovinos. Fêmeas bovinas foram submetidas a protocolos de sincronização de estro para TE e IA. Os animais foram escaneados por ultrassonografia imediatamente antes do procedimento de TE e IA para identificar o tamanho e vascularização do folículo e corpo lúteo (CL). Além disso, os animais inseminados foram escaneados onze dias após a inseminação para identificar o tamanho e vascularização do CL. Todos os dados foram comparados com a fertilidade utilizando-se do diagnóstico gestacional 35 dias após a ovulação. No momento da TE, a vascularização do CL apresentou-se positivamente relacionada com a fertilidade, sendo que animais com menos de 40% de vascularização do CL não ficaram gestantes. No momento da IA, assim como onze dias após a IA, nenhuma relação foi encontrada entre fertilidade e os parâmetros analizados (tamanho e vascularização do folículo e CL), enquanto que contrariamente, houve uma queda na fertilidade em vacas com vascularização do CL acima de 70%. Nos animais inseminados, o tamanho e vascularização do folículo foi positivamente relacionado ao tamanho e vascularização do CL, demonstrado pela presença de um CL maior e mais vascularizado proveniente de um folículo maior e mais vascularizado. O presente estudo é o primeiro a mostrar métodos de predição de fertilidade baseados em ultrassonografia no momento da IA e TE, demonstrando uma aplicabilidade prática no momento da TE. Novos estudos são necessários para suportar os resultados mostrados, incluindo análises hormonais.

Fertility and uterine hemodynamic in cows after artificial insemination with semen assessed by fluorescent probes.

Fluorescent probes (propidium iodide, Hoechst 33342, fluorescein isothiocyanate-conjugated Pisum sativum agglutinin, and 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolcarbocyanine iodide) were used to simultaneously evaluate the integrity of plasma and acrosomal membranes as well as mitochondrial membrane function in cryopreserved bovine semen and to verify its influence on fertility and postinsemination uterine vascularization. One hundred eighty-two Nellore cows were distributed for artificial insemination (AI) using semen batches separated according to the cell percentage presenting intact plasma membrane, intact acrosome, and high mitochondrial function (IPIAH): group G (44.5% IPIAH, n = 68), group M (23.0% IPIAH, n = 56), and group R (8.5% IPIAH, n = 58). The uterine hemodynamic was evaluated by Doppler sonogram in three periods: 30 hours before AI, 4 and 24 hours after AI were considered the resistance index and the uterine vascularization score. The pregnancy rate of group G (64.7%) was greater (P > 0.05) compared with group R (36.2%), but both did not differ from group M (50.0%). There was no effect (P > 0.05) of semen quality on uterine vascularization. Greater vascularization was noticed 4 hours after AI than 30 hours before and 24 hours after AI. Semen evaluation using fluorescent probes contributes to predicting fertilizing potential of semen. The use of semen with less percentage of IPIAH sperm does not alter uterine hemodynamic in cows.

Parâmetros sanguíneos de vacas leiteiras suplementadas com diferentes fontes de gordura / Blood parameters of dairy cows supplemented with different fat sources

O objetivo deste trabalho foi avaliar a utilização de diferentes fontes de gordura nas rações de vacas em lactação e seus efeitos sobre o consumo de matéria seca e as concentrações dos parâmetros sangüíneos. Foram utilizadas 12 vacas da raça Holandês, agrupadas em três quadrados latinos balanceados 4x4 e alimentadas com as seguintes rações: 1) controle; 2) óleo de soja refinado; 3) grão de soja in natura; e 4) sais de cálcio de ácidos graxos (Megalac-E). As concentrações de colesterol total, colesterol, lipoproteína de baixa densidade (LDL) e colesterol de alta densidade (HDL) foram maiores (P<0,05) para as vacas alimentadas com as rações contendo fontes de gordura em relação à ração controle. As concentrações de uréia e nitrogênio ureico no soro foram semelhantes entre as rações utilizadas, com exceção da ração contendo sais de cálcio de ácidos graxos, que apresentou concentrações menores (P<0,05). O consumo de matéria seca foi menor (P<0,05) nas vacas que receberam sais de cálcio nas rações. Os animais que receberam a ração contendo grão de soja como fonte de gordura apresentaram menor (P<0,05) produção de leite quando comparada às demais rações utilizadas. A utilização de fontes de gordura em rações de vacas em lactação alterou os parâmetros sanguíneos, dependendo da fonte de gordura utilizada.

This study was carried out to evaluate the use of different fat sources in dairy cows rations and its effects on dry matter intake, milk yield and composition, and blood parameter. Twelve Holstein cows were allocated in three balanced latin square 4x4, and fed with the following rations: 1) Control, with 2.5 percent; 2) Refined soybean oil; 3) Whole soybean; and 4), Calcium salts of fatty acids (Megalac-E). The concentrations of total cholesterol and lipoprotein low density (LDL) were higher (P<0.05) for cows that received rations with fat sources, which also causes increased (P<0.05) e concentration of high density lipoprotein (HDL). The concentrations of urea and blood urea nitrogen were similar among the rations, except for the diet containing calcium salts of fatty acids, which showed lower (P<0.05). The dry matter intake was lower (P<0.05) in cows that received calcium salts in the rations. The animals that received a diet containing whole soybeans as a source of fat had lower (P<0.05) yield milk when compared to other rations used. The use of fat sources in dairy cows rations influenced the blood parameters, especially for parameters related to lipidogram.