Early resynchronization of non-pregnant beef cows based in corpus luteum blood flow evaluation 21 days after Timed-AI.

The study aimed to verify whether a hormone protocol started at Day 13 (D13) after Timed Artificial Insemination (TAI) influences the conception rate. Nelore cows (primiparous and multiparous) from two commercial beef farms (n = 1,431) were first TAI (D0). Timed AI was performed in lots (TAI Lots) ranging from 187 to 346 cows. On D13, regarding the TAI lot, cows were assigned for either receiving (Resynch group, n = 1,002) or not (Control group, a subset of approximately 30%, n = 429) another hormone protocol for resynchronization. The same hormone protocol was used for the first TAI and for the resynchronization, except for 1 mg instead of 2 mg of estradiol benzoate (EB) at the begging of the protocol. Eight days later (D21), the Resynch group was checked for corpus luteum blood flow by color Doppler ultrasonography, and in those detected as non-pregnant, the protocol was completed and a 2nd TAI was performed at D23. Pregnancy diagnosis was later (D30) performed by B-mode ultrasonography in the control group and confirmed in the presumptive pregnant cows from the 1st TAI of the Resynch group. The remaining cows were checked for pregnancy 30 days after the 2nd TAI (experimental Day 53). The statistical model to explain conception rate considered the effects of Group (Control or Resynch), Farm, Parity (primiparous or multiparous), Sire, Technician (who perform AI), TAI Lot and pertinent interactions (Group*Parity, Group*Farm and Group*TAI Lot). The statistical analyses of the model were performed using the Proc Glimmix (SAS virtual University Edition). The conception rate for the 1st TAI was similar (P > 0.4) between Control (50.3%, 216/429) and Resynch group (52.6%, 527/1002). The positive predictive diagnostic on D21 showed high relation with PD30 (90.7%, 527/581). In Resynch group, non-pregnant cows (n = 421, 1002 minus 581) were re-inseminated. The conception rate of the 2nd TAI (42.8%, 180/421) was affected (P < 0.002) by side effects of the Farm (48.5 vs. 33.1%) and Parity (51.2 vs. 40.3%, for multiparous vs. primiparous, p < 0.001). Nevertheless, after the 2 TAIs of the Resynch group, the cumulative conception rate was 70.5% (707/1002). In conclusion, the early resynchronization of cows with a low (1 mg) EB dose and progesterone device at D13 after TAI can be used as a strategy to reduce conception interval in beef cattle, and thus to increase the number of pregnant cows from artificial insemination after the breeding season.

Efficacy and limitations of different approaches to anticipate the diagnosis of pregnancy in cattle / Eficácia e limitações de diferentes abordagens para antecipar o diagnóstico de gestação em bovinos

The study evaluated sonographic and serologic exams performed for early (20 to 30d) diagnosis of pregnancy. One hundred-twenty (n= 120) bovine recipients were synchronized (estrous=D0) and timed embryo transferred (TET, D7) with fresh in vitro produced embryos. In the first trial (n= 46), diagnosis of pregnancy was performed on day 20 (D20) by detecting CL blood flow (BF) and by Pregnancy-Associated Glycoproteins (PAGs) serology. In the second trial (n= 30), pregnancy diagnosis was performed on D25 by ultrasound visualization of uterine contents and by PAGs serology. In the last trial, PAG's serology was performed on D30. Ultrasonographic detection of the uterine contents and embryo viability performed on D30 (DG30) was considered the gold standard. The PROC FREQ procedure was used to test the agreement between diagnostic methods. On D20, the Doppler ultrasonography of the CL had showed high sensitivity (100%), but only moderate specificity (53.3%). On the same day, serologic diagnostic had no agreement (k= -0.08, P< 0.46) with the gold standard, with very low sensitivity (6.3%). However, the sensitivity of the serologic exam increased dramatically (from 6.3 to 100%) from D20 to D25, and it contributed to detect false negatives from the ultrasound diagnosis, improving the overall accuracy from 90% to 96.7%.(AU)

O estudo foi planejado para correlacionar exames ultrassonográficos e sorológicos realizados para o diagnóstico precoce (20 a 30d) de gestação. Cento e vinte (n= 120) receptoras bovinas foram sincronizadas (estro=D0), e embriões frescos produzidos in vitro foram transferidos em tempo fixo (TETF, D7). No experimento 1 (n= 46), o diagnóstico de gestação foi realizado no D20, pela detecção do fluxo sanguíneo do CL e pela sorologia de glicoproteínas associadas à gestação (PAGs). No experimento 2 (n= 30), a detecção da gestação foi realizada por meio da visualização do conteúdo do útero e também pela sorologia para PAGs. No experimento 3, a sorologia para PAGs foi realizada no D30. Em todos os experimentos, a visualização ultrassonográfica da vesícula e da viabilidade embrionária, realizada no D30, foi considerada padrão-ouro. O procedimento PROC FREQ testou o nível de concordância dos métodos diagnósticos. No D20, o diagnóstico baseado na vascularização do CL mostrou alta sensibilidade (100%) e apenas moderada especificidade (53,3%). Nesse mesmo dia, o diagnóstico sorológico não apresentou concordância (k=-0,08, P<0,46) com o padrão-ouro, além de baixa sensibilidade (6,3%). No entanto, a sensibilidade do exame sorológico aumentou drasticamente (6,3 para 100%) do D20 para o D25, contribuindo para detectar falsos negativos diagnosticados pela ultrassonografia, melhorando a acurácia (90 para 96,7%).(AU)

Development of fixed-time artificial insemination protocols for locally adapted Curraleiro Pé-Duro cows / Desenvolvimento de protocolos de inseminação artificial em tempo fixo em vacas da raça localmente adaptada Curraleiro Pé-Duro

Two experiments were conducted aiming to evaluate the effects of two ovulatory inducers (Exp.1) and equine chorionic gonadotropin (eCG; Exp.2) on follicular and luteal dynamics in a fixed-time AI (FTAI) protocol in locally adapted Curraleiro Pé-Duro cows. In Exp. 1 multiparous cows (n=12) received an intravaginal device containing 1g of progesterone (P4) for 8 days and 2mg of estradiol benzoate (EB) intramuscularly (IM) at device insertion (Day 0). At device removal (Day 8) 0.150mg of Sodium D-Cloprostenol was administered IM and the cows were randomly assigned to receive 1mg of EB (EB8) or 1mg of estradiol cypionate (EC8) IM, or to not receive any ovulatory inducer (Control). All the animals participated in all treatments (crossover). The interval from P4 removal to ovulation was shorter and less variable in the EB8 treatment group (P≤0.05). In Exp. 2 (crossover), multiparous cows (n=12) received the same hormonal treatment as the EB8 group in Exp.1. At device removal (Day 8) cows were randomly assigned to receive 300UI of eCG IM or to not receive eCG (Control). No difference was ascertained on follicular and luteal parameters in Exp. 2 (P>0.05). We concluded that EB can be used as the ovulatory inducer (Exp. 1) in a FTAI protocol in Curraleiro Pé-Duro cows. However, eCG (Exp. 2) was not able to stimulate follicular and luteal development. This result is probably due to the adaptive capacity of Curraleiro Pé-Duro cows that maintained a satisfactory body condition score even in dry and hot environments.(AU)

Foram realizados dois experimentos com o objetivo de avaliar o efeito de dois indutores da ovulação e da gonadotrofina coriônica equina (eCG) na dinâmica folicular e luteal, em um protocolo de inseminação artificial em tempo fixo (IATF) em vacas localmente adaptadas da raça Curraleiro Pé-Duro. No experimento 1, vacas pluríparas receberam um dispositivo intravaginal contendo 1g de progesterona (P4) durante oito dias e 2mg de benzoato de estradiol (BE) intramuscular (IM) no momento da inserção do dispositivo (dia zero). Na retirada do dispositivo (dia oito), as vacas receberam 0,150mg de D-cloprostenol sódico IM e foram separadas aleatoriamente para receber 1mg de BE IM (BE8) ou 1mg de cipionato de estradiol IM (CE8), ou nenhum indutor da ovulação (controle). Todos os animais participaram de todos os tratamentos (crossover). O intervalo entre a retirada da P4 e a ovulação foi menor e menos variável no tratamento BE8 (P≤0,05). O momento da ovulação foi mais precoce e mais concentrado nos animais do grupo BE 8. No experimento 2 (crossover), vacas pluríparas receberam o mesmo tratamento hormonal do grupo BE8 do experimento1. Na retirada do dispositivo (dia 8), as vacas foram separadas aleatoriamente para receberem 300UI de eCG IM, enquanto o controle não. Não houve diferença nos parâmetros foliculares e luteais avaliados no experimento 2 (P>0,05). Em conclusão, o BE pode ser utilizado como indutor da ovulação (experimento 1) em protocolos de IATF em vacas Curraleiras Pé-Duro. Entretanto, o eCG (experimento 2) não foi capaz de estimular o desenvolvimento folicular e luteal. Esse resultado é devido provavelmente à capacidade adaptativa das vacas Curraleiras Pé-Duro em manter uma condição corporal satisfatória mesmo em condições de clima seco e quente.(AU)

Estudo etnofarmacológico e etnobotânico de Himatanthus drasticus (Mart. ) Plumel (janaguba) / Ethnopharmacological and ethnobotanical study of Himatanthus drasticus (Mart. ) Plumel (janaguba).

RESUMO Himatanthus drasticus (Mart.) Plumel, janaguba, produz um látex em seu caule que é usado para o tratamento de alguns tipos de câncer, vermes intestinais, febre, menstruações irregulares, infertilidade feminina e úlceras gástricas. Estudos fitoquímicos revelaram a presença de acetato de lupeol e proteínas; e estudos farmacológicos, os efeitos antiúlcera, antitumoral, imunomodulador, analgésico e anti-inflamatório. O presente trabalho objetivou realizar levantamento etnobotânico e etnofarmacológico sobre H. drasticus como contribuição ao conhecimento científico da referida espécie. Foi elaborado um questionário, que foi aplicado em entrevista a 21 raizeiros nos mercados das cidades de Crato, Fortaleza e Pacajus do estado do Ceará (CE). Observou-se que 38,1% dos raizeiros têm mais de 10 anos de serviço e 47,6%, estão na faixa etária de 40 a 59 anos, o que representa um fator positivo quanto à experiência com plantas medicinais. Aproximadamente 85,7% dos raizeiros relataram o uso do látex da H. drasticus para tratamento de úlcera; 66,7% para inflamação; 66,7% para câncer. Verificou-se que as principais indicações terapêuticas relatadas pelos raizeiros sobre esta espécie estão de acordo com as informações científicas. Os levantamentos etnobotânicos e etnofarmacológicos exercem um papel primordial no resgate do conhecimento tradicional, nos meios rurais e urbanos, tanto por seu valor histórico-cultural, como também pela necessidade de confirmação das indicações de uso.

ABSTRACT The Himatanthus drasticus (Mart.) Plumel, janaguba, produces a latex whose stem is used to treat some types of cancer, intestinal worms, fever, irregular menses, female infertility and gastric ulcers. Phytochemical studies revealed the presence of lupeol acetate and proteins in the latex; and pharmacological studies indicated antiulcer, anti-tumor, immunomodulating, analgesic and anti-inflammatory effects. This study aimed to conduct surveys on ethnopharmacological and ethnobotanical species H. drasticus as a contribution to the scientific knowledge about the species. A questionnaire and interviews were made with 21 healers in the markets of the cities of Crato, Fortaleza and Pacajús, in state of Ceara (CE). It was observed that 38.1% of the healers have more than 10 years of service, and 47.6% are between 40-59 years old, which is a positive factor about the experience with medicinal plants. Approximately 85.7% of the healers reported to apply latex H. drasticus for the treatment of ulcers, 66.7% for inflammation, and 66.7% for cancer. It was found that the main therapeutic indications mentioned by the healers about this species are consistent with the scientific information available. The ethnobotanical and ethnopharmacological surveys play an essential role on keeping the traditional knowledge in rural and urban areas, both for its historical and cultural value, but also for the need of confirmation of the intended use.

Aspectos clínicos e laboratoriais da mastite induzida por inoculação intramamária de Staphylococcus aureus em vacas e búfalas / Clinical and laboratory aspects of mastitis induced by intramammary inoculation of Staphylococcus aureus in cows and buffaloes

Vários aspectos relacionados à mastite bubalina precisam ser esclarecidos. Muitas condutas são adaptadas a essa espécie pelo conhecimento adquirido com a espécie bovina. Com este experimento, objetivou-se evidenciar particularidades da mastite em búfalas e vacas que tiveram uma glândula inoculada com 1,0 x 103 UFC de S. aureus. Os animais foram monitorados por cultura bacteriana do leite, California Mastitis Test (CMT), e escores para avaliação da severidade da mastite. Foram utilizados os seguintes critérios: temperatura retal, apetite, produção de leite (resposta sistêmica à inflamação), contagem de células somáticas (CCS), aparência/consistência da glândula e aparência da secreção láctea (resposta localizada à inflamação). Todos os animais desenvolveram mastite clínica superaguda. A bactéria foi recuperada de todas as glândulas desafiadas, sem diferença significativa no percentual de isolamento entre as espécies até o 11º dia pós-inoculação, porém com diferença (P<0,001) no 30º dia pós-inoculação. A CCS pelo método eletrônico e o CMT detectaram nas búfalas resposta mais intensa logo após a inoculação e diminuição mais rápida da contagem ao final do experimento, quando se comparou essa espécie com as vacas. Ambos os testes demonstraram correlação com a cultura bacteriana. Na avaliação da severidade da mastite, as médias do escore total, do escore da resposta local e do escore da resposta sistêmica foram mais elevadas na espécie bovina (P<0,05). Ao longo do período de observação, verificou-se uma capacidade superior das búfalas em recuperar o status sanitário adequado, chegando, ao final do experimento, com os parâmetros avaliados mais próximos dos fisiológicos...

Several aspects related to bubaline mastits need to be clarified. Many conducts are adapted to this species by the knowledge adquired with the bovine species. This experiment aimed to evidence particularities of mastits in buffaloes and cows submitted to one-gland inoculation with 1.0 x 103 CFU of S. aureus. The animals were monitored through bacterial culturing milk, California Mastitis Test (CMT) and scores to assess the severity of mastitis. The following criteria were used: rectal temperature, appetite, milk production (systemic response to inflammation), somatic cell count (SCC), appearance/consistency of the gland and appearance of the milk secretion (localized response to inflammation). All animals developed hyperacute clinical mastitis. The bacteria was recovered from all challenged glands, with no significant difference in the percentage of isolation between the species until day 11 after inoculation, but with a difference (p<0.001) on day 30 post inoculation. The SCC with the electronic method and CMT, detected in buffaloes a more intense response right after the inoculation and a faster decrease of counting by the end of the experiment, when compared to cows. In both tests correlation with the bacterial culture was detected. In assessing the severity of mastitis, the averages of total score, of the score of local response and the systemic response score were higher in the bovine species (P<0.05). Throughout the observation period, there was a higher capacity of buffaloes in reaching the appropriate sanitary status, coming to the end of the experiment with the parameters closer to the physiological...

P-gp modulating drugs greatly potentiate the in vitro effect of ivermectin against resistant larvae of Haemonchus placei.

Since its production in the 1980s, ivermectin (IVM) has been used indiscriminately and the selection pressure to which bovine gastrointestinal nematodes have been exposed has been intense, resulting in considerable economic losses due to parasitic resistance. One possibility for the control of resistant parasites is the use of P-glycoprotein (P-gp) modulators, because one of the main biochemical changes in ivermectin-resistant parasites is the increased activity of membrane proteins responsible for the efflux of drugs and xenobiotics. This study aimed to evaluate the in vitro effect of eight P-gp modulating drugs to potentiate IVM efficacy against an IVM-resistant field isolate of Haemonchus placei (Nematoda: Trichostrongylidae). The association of IVM with cyclosporin-A, ceftriaxone, dexamethasone, diminazene aceturate, quercetin, trifluoperazine, verapamil, or vinblastine resulted in increased IVM (10(-4)M) efficacy of 5.1%, 49.06%, 76.42%, 3.31%, 28.85%, 13.74%, 45.64% and 43.61%, respectively, and reduced the IVM half maximal effective concentration (EC50) from 4.381 × 10(-6)M to 9.877 × 10(-8), 2.739 × 10(-7), 1.240 × 10(-6), 1.651 × 10(-6), 2.710 × 10(-7), 1.159 × 10(-7), 1.026 × 10(-6) and 7.136 × 10(-7)M, respectively. Only diminazene aceturate did not significantly reduce the number of migrating larvae when associated with IVM (P > 0.05). The effect of P-gp modulating drugs depended on IVM concentration, with greater potentiating effect at lower IVM concentrations. The in vitro application of trifluoperazine, dexamethasone, quercetin, verapamil, cyclosporin A, vinblastine, and ceftriaxone potentiated IVM efficacy against an IVM-resistant field isolate of H. placei, resulting in higher efficacy and lower IVM EC50.

A new polymorphism in the Growth and Differentiation Factor 9 (GDF9) gene is associated with increased ovulation rate and prolificacy in homozygous sheep.

Brazilian Santa Inês (SI) sheep are very well-adapted to the tropical conditions of Brazil and are an important source of animal protein. A high rate of twin births was reported in some SI flocks. Growth and Differentiation Factor 9 (GDF9) and Bone Morphogenetic Protein 15 (BMP15) are the first two genes expressed by the oocyte to be associated with an increased ovulation rate in sheep. All GDF9 and BMP15 variants characterized, until now, present the same phenotype: the heterozygote ewes have an increased ovulation rate and the mutated homozygotes are sterile. In this study, we have found a new allele of GDF9, named FecG(E) (Embrapa), which leads to a substitution of a phenylalanine with a cysteine in a conservative position of the mature peptide. Homozygote ewes presenting the FecG(E) allele have shown an increase in their ovulation rate (82%) and prolificacy (58%). This new phenotype can be very useful in better understanding the genetic control of follicular development; the mechanisms involved in the control of ovulation rate in mammals; and for the improvement of sheep production.

Protein kinase C (PKC) role in bovine oocyte maturation and early embryo development.

The aims of the present study were to determine the role of protein kinase C (PKC) on meiotic resumption and its effects on pronuclear formation and cleavage in the bovine. Oocytes were matured in the presence of 0, 1, 10 and 100 nM of phorbol 12-myristate 13-acetate (PMA), to evaluate the percentage of germinal vesicle breakdown. To study pronuclear formation and cleavage, oocytes were randomly distributed in four groups and matured in modified TCM-199 with LH and FSH (negative control); 10% of estrous cow serum (positive control); 100 nM of PMA (treatment); 100 nM of 4alpha-PDD (phorbol ester control). Oocytes were also matured in positive control medium, fertilized and transferred to KSOM with increasing concentrations of a PKC inhibitor. The protein profile and the presence of PKC at the end of maturation period were determined by SDS-PAGE followed by Silver Stain and Western blot, respectively. PMA stimulated meiotic resumption in a concentration-dependent manner. PKC stimulation during oocyte maturation caused an increase in pronuclear formation and did not cause parthenogenetic activation. Inhibitor of PKC (MyrPKC) inhibited cleavage in a dose-dependent and irreversible manner. A protein band around 74 kDa was not detected in PMA-treated oocytes and PKC was not detected by Western blot at the end of the maturation period. In conclusion, meiotic resumption was accelerated and the rate of oocytes with two pronuclei was increased when PKC was activated during oocyte maturation. Moreover, cleavage was inhibited in the presence of PMA.

Sexing of Dorper sheep fetuses derived from natural mating and embryo transfer by ultrasonography.

In order to improve fetal sexing in the Dorper sheep breed, the objective of the present study was to determine, by repeated ultrasonographic examinations, the migration period of the genital tubercle (GT) in sheep fetuses derived from natural mating or embryo transfer and to compare the accuracy of a single examination with repeated examinations at short intervals. For this purpose, transrectal ultrasound was performed, using a double-frequency linear transducer (6.0 and 8.0 MHz) for monitoring 51 sheep fetuses distributed in three experimental groups (EI, EII and EIII). The fetuses in EI (n = 23) and EII (n = 18) derived, respectively, from natural mating and embryo transfer were monitored at 48-h intervals from the 30th to 60th day of gestation and sexed based on the final location of the GT. The fetuses in EIII (n = 10), which originated from embryo transfer, were examined only once on the 65th day of gestation and sexed taking into consideration the final position of the GT and/or by identification of anatomical structures of external genitalia. The accuracy of fetal sexing was 91.3% (21 fetuses sexed/23 quantified) in EI, 88.9% (16 sexed/18 quantified) in EII and 100% (10 sexed/10 quantified) in EIII, without significant difference (P > 0.05) between experiments. Migration of the GT occurred earlier (P < 0.05) in fetuses produced by natural mating (43.0 +/- 2.8 days) than in those derived from embryo transfer (46.1 +/- 4.7 days). The results show that fetal sexing can be done from the 50th day onward in fetuses produced by natural mating and from the 60th day onward in fetuses derived from frozen embryos. It can also be concluded that repeated ultrasonographic exams in short time intervals do not maximise the accuracy of fetal sexing. In addition, real-time ultrasonography is a reliable tool for fetal sex determination in sheep after Day 50 of gestation, taking into account both the location of the GT and the identification of external genital structures.

Chronic administration of recombinant ovine leptin in growing beef heifers: effects on secretion of LH, metabolic hormones, and timing of puberty.

Serum concentrations of leptin increase linearly from approximately 16 wk before until the week of pubertal ovulation in beef heifers. To test the hypothesis that exogenous leptin can hasten the onset of puberty in heifers, we examined the effects of chronic administration of recombinant ovine leptin (oleptin) on timing of puberty, pulsatile and GnRH-mediated release of LH, and plasma concentrations of GH, IGF-I, and insulin. Fourteen fall-born, prepubertal heifers (Brahman x Hereford, 12 to 13 mo; 304.7+/-4.12 kg) were used. Heifers were stratified by age and BW and assigned randomly to one of two groups (seven animals per group): 1) Control; heifers received s.c. injections of saline twice daily (0700 and 1900) for 40 d; and 2) Leptin; heifers received s.c. injections of oleptin (19.2 microg/kg) twice daily at 0700 and 1900 for 40 d. Blood samples were collected at 10-min intervals for 5 h on. d 0, 5, 10, 20, 30, and 40, and twice daily, just before each treatment injection, throughout the study. On d 41, heifers received i.v. injections of GnRH at 0 (0.0011 microg/kg) and 90 min (0.22 microg/kg), with additional sampling for 5.5 h to examine releasable pools of LH. Diets promoted a gain of 0.32+/-0.09 kg/d, which did not differ between groups. Plasma concentrations of leptin increased markedly in leptin-treated heifers and were greater (P < 0.001) than controls throughout (27.8+/-0.8 vs. 4.9+/-0.12 ng/mL). None of the heifers reached puberty during the experiment, but did so within 45 d of its termination. Mean concentrations of plasma LH, GH, IGF-I, and insulin were not affected by treatment, nor was there an overall effect on the frequency of LH pulses. However, a treatment x day interaction (P = 0.02) revealed that the frequency of LH pulses (pulses/ 5 h) was greater (P = 0.03) in controls (3.6+/-0.36) than in leptin-treated heifers (1.7+/- 0.28) on d 10. Characteristics of GnRH-induced release of LH were not affected by treatment. In summary, chronically administered leptin failed to induce puberty or alter endocrine characteristics in beef heifers nearing the time of expected puberty.

Leptin prevents fasting-mediated reductions in pulsatile secretion of luteinizing hormone and enhances its gonadotropin-releasing hormone-mediated release in heifers.

We tested the hypothesis that leptin could prevent fasting-mediated reductions in pulsatile secretion and modify GnRH-mediated release of LH in heifers approaching puberty. Thirteen crossbred, prepubertal heifers (13.5-16 mo; 280-350 kg) exhibiting frequencies of pulses of LH between 0.67 and 1 pulse/h, were assigned randomly to two groups: 1). control (n = 6), fasted for 72 h with s.c. injections of saline at 12-h intervals, and 2). leptin (n = 7), fasted for 72 h with s.c. injections of oleptin (19.2 microg/kg) at 12-h intervals. Blood samples were collected intensively for 6 h on Days 0 and 3. This was followed on Day 3 with sequential administration of physiological (0.0011 microg/kg, i.v.) and pharmacological (0.22 microg/kg, i.v.) doses of GnRH and additional blood sampling. Leptin treatment increased (P = 0.0003) plasma concentrations of leptin 5-6-fold compared to controls. Fasting caused a marked decline (P = 0.01) between Days 0 and 3 in the frequency of LH pulses in controls; however, this effect was prevented in the leptin group, with pulse frequency increasing (P < 0.008) from Day 0 to 3. Leptin treatment increased GnRH-induced release of LH at both low (P = 0.04) and high (P = 0.02) doses. Plasma insulin and insulin-like growth factor-1 were reduced by fasting and unaffected by leptin. Leptin increased mean concentrations of growth hormone. Results indicate, for the first time, that exogenous leptin can prevent fasting-mediated reductions in the frequency of LH pulses and modify GnRH-mediated release of LH in intact, prepubertal heifers.

Phenotypic and molecular characterization of bovine Campylobacter fetus strains isolated in Brazil.

The objective of the present study was to characterize the phenotypic and molecular aspects of Campylobacter fetus strains isolated from bovine herds with reproductive problems. Thirty-one Brazilian field isolates, together with one reference strain of each subspecies of C. fetus, were analyzed. The strains were submitted to phenotypic identification followed by subspecies characterization using the polymerase chain reaction (PCR) and numeric evaluation of restriction fragment length polymorphism (RFLP) separated by pulsed-field gel electrophoresis (PFGE). Phenotypically, 4 isolates (12.1%) were classified as C. fetus subsp. fetus, and 29 isolates (87.9%) were classified as C. fetus subsp. venerealis. However, according to molecular analysis, only 1 isolate (3.0%) was classified as C. fetus subsp. fetus (the reference strain), whereas 32 isolates (97.0%) were considered C. fetus subsp. venerealis. SalI digestion of C. fetus genomic DNA, obtained from the 33 strains, yielded 7-10 DNA fragments ranging in size from 40 to 373kb, with 12 distinct patterns. Furthermore, the numeric analysis by neighbor-joining of the DNA from the 33 strains resulted in a dendrogram in which 2 distinct groups were identified. It was concluded that phenotypic characterization of C. fetus subspecies might lead to erroneous classification of field isolates. Although RFLP-PFGE is a powerful and reliable technique to characterize C. fetus, it has the inconvenience of being time consuming and laborious. Whereas PCR, besides providing rapid results, was found to be reliable and convenient for the characterization of field isolates of C. fetus.

Follicular development and steroid concentrations in cows with different levels of fertility raised under nutritional stress.

The aim of the present study was to characterize ovarian follicular development and steroid concentrations during postpartum and the estrous cycle of Brangus Ibagé cows (3/8 Nelore + 5/8 Aberdeen Angus) with different levels of fertility. Cows were classified as having high or low fertility according to the calving interval (CI). The average CI of the herd from which cows used in this study were selected was 404.6+/-5.44 and 711.2+/-20.89 days for the high and low fertility groups, respectively. Four cows of high fertility and five cows of low fertility had calves removed between 70 and 100 days after parturition. Ovarian activity was monitored daily by ultrasound for 16 days after calf removal. Days to emergency of the first follicular wave after calf removal, number of follicles with diameter >9 mm, growth rate of largest follicle, maximum diameter of largest follicle, length (days) and number of follicular waves were recorded. During this period, blood was collected daily for measurements of serum progesterone (P(4)) and estradiol (E(2)) concentrations. In another experiment, ovarian activity and P(4) and E(2) concentrations were examined during estrous cycle in five cows of high fertility and four cows of low fertility. Ovarian activity and steroid concentrations were assessed from the day prior to estrus to the 15th day of the estrous cycle (estrus = day 0). In postpartum cows of high fertility, the total number of follicles >5mm and the maximum diameter of the largest follicle were higher than in cows of low fertility (P < 0.05). Concentrations of P(4) and E(2) did not differ between groups in the postpartum cows. However, E(2) increased 5 days after calf removal (around 90 days of postpartum) in the high fertility group, followed by an increase in P(4) with average values indicating ovulation around 100 days postpartum. In cycling cows, the profile of follicular development was similar between cows of high and low fertility. There was no difference between groups for number of follicles >5mm, but the day effect was significant (P < 0.01). Plasma concentrations of P(4) and E(2) were similar in both groups. These data suggest that cows, from a population raised in the same environment have different fertility as a consequence of individual physiological characteristics.

Efeito da somatotrofina bovina (bST-r), do implante de progestágeno e do desmame por 72 horas na induçäo do estro e na taxa de prenhez em vacas / Effect of bovine somatotropin (bST-r), progestagen intravaginal device and temporary calf removal on estrous induction and on pregnancy rate in cows

Este trabalho teve por objetivo avaliar a eficiência de tratamentos hormonais sobre a fertilidade de vacas de corte no pós-parto com diferentes condiçöes corporais, durante a estaçäo de monta de outono. Setenta e três vacas pluríparas cruzadas (Hereford x Nelore) criadas extensivamente, com condiçäo corporal entre 2 e 4, foram pesadas e distribuídas em três grupos experimentais. O grupo GSED, constituído por 25 vacas, recebeu pessário vaginal (dia 0) contendo 250mg de acetato de medroxiprogesterona e 500mg de somatotropina bovina recombinante (bST-r). Na retirada dos pessários (dia 7), as vacas receberam 0,5mg de cipionato de estradiol e procedeu-se o desmame temporário dos bezerros por 72 horas. No grupo SED, 25 vacas receberam tratamento semelhante ao grupo GSED, porém näo receberam bST-r. No grupo-controle, as 23 vacas somente foram separadas dos seus bezerros por 72h. Quando da retirada dos pessários as vacas foram colocadas com touros por 30 dias. Os animais foram pesados e avaliados quanto à condiçäo corporal no início do experimento e naretirada dos touros (dia 37). Foi constatada perda média de peso de 0,648 Kg/dia e os percentuais de estro foram de 26,1 por cento, 33,3 por cento e 56,5 por cento, respectivamente, para os grupos controle, SED e GSED. O diagnóstico de gestaçäo, realizado pela palpaçäo retal 60 dias após a retirada dos touros, indicou percentuais de prenhez de 13,0 por cento, 8,3 por cento e 21,7 por cento, respectivamente, para os grupos controle, SED e GSED (P=0,16), demonstrando que os programas hormonais adotados näo foram eficientes no incremento das taxas de prenhez de vacas que perdiam peso entre 50 e 70 dias após o parto

Fator de crescimento derivado das plaquetas, retinol e insulina na regulaçäo da maturaçäo nuclear de oócitos bovinos e suas conseqüências no desenvolvimento embrionário / Platelet-derived growth factor, retinol and insulin in the regulation of bovine oocyte nuclear maturation and their consequent effect in the embryonic development

O objetivo desse trabalho foi verificar as açöes do fator de crescimento derivado das plaquetas (PDGF;P), da insulina (I), do retinol (R) e de suas associaçöes(PI, PIR, IR e PR) na maturaçäo nuclear (MN)de oócitos bovinos e suas consequências no desenvolvimento embrionário (DE). O meio básico para maturaçäo dos oócitos nos diferentes tratamentos foi o TCM-199 modificado acrescido de PVA (controle). No DE, foram utilizados os grupos R, PIR, IR, um controle negativo (PVA) e um controle positivo, contendo soro fetal bovino e gonadotrofinas (SFBHOR). Os fatores P, I, R e suas associaçöes näo aceleraram a MN em 7h mas sim após 18h (P<0,001), com exceçäo dos tratamentos R e PR, nos quais as percentagens de metáfase II foram, respectivamente, de 4,7(porcento) e 8,3(porcento), similares à obtida no grupo-controle (0,0(porcento)). Considerando um nível de significância de P<0,0001 em comparaçäo ao grupo-controle, os maiores índices de matáfase II foram obtidos na presença das associaçöes IR (19,0(porcento)) e PIR (21,3(porcento)). No DE, R (18,3(porcento)), PIR (13,9(porcento)) e IR (10,6(porcento)) incrementaram os índices de clivagem, comparados ao PVA (0,0(porcento);P<0,001), porém näo atingiram os índices do grupo SFBHOR (53,8(porcento);P<0,001). Conclui-se que insulina e PDGF aceleram a MN e suas açöes säo potencializadas pelo retinol. Os índices de clivagem de oócitos maturados na presença de R, IR e PIR säo superiores aos do PVA, mas significativamente inferiores aos maturados em SFBHOR

Experimental infection of pregnant ewes with bovine viral diarrhea virus type-2 (BVDV-2): effects on the pregnancy and fetus.

The reproduction effects of bovine viral diarrhea virus type-2 (BVDV-2) infection were investigated in ewes inoculated with a non-cytopathic BVDV-2 isolate at three stages of gestation. Virus inoculation was followed by a transient viremia, accompanied by a transient and mild hyperthermia and nasal discharge in a few animals. Some ewes were sacrificed at different time-points after virus inoculation to study the kinetics of fetal infection. Infectivity and viral antigens were detected in placentomes from day 7 to 36 post-inoculation (pi) and in fetal fluids and tissues between days 10 and 28 pi. Cardiac petechial hemorrhages and hemoperitoneum accompanied by a severe fibrinous ulcerative placentitis were observed in fetuses examined at days 21, 28 and 36 pi. Inoculation of ewes at days 55-60 of gestation resulted in a prolonged virus replication in placentomes and fetal tissues; ewes that were allowed to proceed with pregnancy had 77% of abortions or fetal and perinatal deaths. Seven stillbirths, unviable and viable lambs born to these ewes were virus-positive at birth. Infectious virus was repeatedly isolated from leukocytes of two lambs up to 2 and 6 months of age, indicating they were persistently infected. Ewes inoculated at days 65-70 of gestation had 66.6% of fetal and perinatal losses. Three viable lambs born to these ewes were healthy, BVDV antibody-positive and virus-negative. A transient viral replication in placentomes and in a few fetal tissues, followed by the rise of fetal neutralizing antibodies and virus clearance was the result of inoculating ewes at days 120-125 of gestation. Lambs born to these ewes were healthy, antibody-positive and virus-negative. These results demonstrate that the biology of BVDV-2 infection in pregnant sheep is essentially similar to that of BVDV-1 in pregnant cattle and sheep. These features make this species an attractive animal model for studying the pathogenesis of congenital BVDV-2 infection.

Implantable cardioverter-defibrillator: beyond efficacy.

OBJECTIVES: The introduction of the implantable cardioverter-defibrillator was a very important advance in the treatment of malignant ventricular arrhythmias. However, its use is associated with some possible adverse events, which should be taken into consideration when a patient is proposed for implantation. These complications may occur early after implantation and be associated with the procedure itself, or they may be late complications, usually associated with the device or the arrhythmia. It was our objective to assess the incidence of these complications in our population of patients. POPULATION AND METHODS: We describe the complications found in 98 patients (55.9 +/- 13.9 years, 89% male) with an implantable cardioverter-defibrillator and compare our results with the incidence of complications described by other authors. RESULTS: We found complications associated with the presence of the implantable cardioverter-defibrillator in 32% of patients. The most frequent complication was inappropriate shocks in 13%. The presence of infection was detected in 4%, lead insulation faults in 5%, need for lead extraction in 2%, repositioning in 1% and re-establishment of the connection with the generator in 2%. In 5% of patients, there was inefficacy of the device, 3% due to non-detected ventricular tachycardias (slow tachycardias) and 2% due to electrical storms. There was syncope in 2% of patients. The total mortality in a 2.9 +/- 1.9 year follow-up was 13% (sudden death in 3%). CONCLUSIONS: Major complications associated with implantable cardioverter-defibrillators were in our experience relatively rare, our results agreeing with those of other centers.

Cava endoluminal balloon occlusion as an adjunct technique to perform less invasive cardiopulmonary bypass procedures.

OBJECTIVES: A cardiopulmonary bypass technique was developed with the aim of achieving a less invasive approach to right-sided lesions of the heart or to avoid extensive dissection in reoperations. METHODS: Cavae endoluminal balloon occlusion (CEBO) was achieved with balloon catheters, introduced through cannulas inserted in the right jugular and one of the femoral veins. This procedure can be used in association with a standard femoral cardiopulmonary bypass or with the port-access technique. RESULTS: Twenty-two patients were operated, through a right thoracotomy, for atrial septal defect (19) and tricuspid valve regurgitation associated with mitral valve pathology (3). There were two intra-operative complications, partially related with the development of this technique and lack of appropriate designed material. There was no mortality and a shorter and easier postoperative care was achieved in almost all patients. CONCLUSIONS: The CEBO technique is an useful and safe adjunct to perform miniinvasive or complex reoperations on the right side of the heart.

Effect of fetal age and method of recovery on isolation of preantral follicles from bovine ovaries.

The objective of this study was to compare enzymatic and mechanical methods, at distinct fetal ages, on isolation of different developmental stages of preantral follicles from bovine ovaries. Fetal ovaries were obtained from pregnant cattle at 150 to 270 d of gestation, and 135,521 preantral follicles at different stages of development were studied. The dissociation of ovaries with a mechanical procedure resulted in an average of 938.16 prenatral follicles. In contrast, 3,715.56 follicles were obtained when enzymatic digestion was used (P = 0.0001). Histological evaluation confirmed follicular stages and demonstrated that both mechanical and mechanical-enzymatic procedure did not affect the cellular integrity of the follicles. Granulosa cell-oocyte complexes surrounded by a basal membrane, were considered preantral follicles in this study. The ratio of different stages of isolated preantral follicles was significantly (P = 0.0001) correlated to fetal age. The earliest fetal age at which tertiary follicles were identified was at 210 d of gestation. The results confirm previous observation that follicular development and atresia are initiated during fetal development. These data provide information on methodologies to isolate intact bovine preantral follicles for investigating the control and regulation of follicular development and the growth of preantral follicles in vitro.

Mechanisms underlying degeneration of cryopreserved vascular homografts.

OBJECTIVE: To analyze the mechanism(s) underlying homograft degeneration, we designed an experimental model in which the behavior of cryopreserved autografts and homografts, as well as fresh autografts, implanted in the same animal was compared. METHODS: A cryopreserved homograft was implanted in the aorta of 14 sheep. The excised aortic autologous segment was then subjected to cryopreservation, and 1 to 8 weeks later it was implanted 1 to 2 cm below the cryopreserved homograft. The intermediate segment of the native aorta, the fresh autograft, was dissected at this point. Animals were put to death at different times and the implanted segments were harvested together with a portion of native aorta. Histologic and immunohistochemical analyses, as well as cell viability assessments, were then performed on the explanted segments. Similar studies were also conducted on fragments of cryopreserved autografts and homografts before implantation. RESULTS: With the exception of a partial loss of the endothelium, cryopreserved specimens retained cell viability and morphologic integrity before implantation. Explanted cryopreserved homografts showed profound changes affecting all strata, as well as a decline in cell viability. Lymphocyte infiltrates were found up to 12 months after implantation. Endothelium was always absent in cryopreserved homografts. However, a reendothelialization of the cryopreserved autografts was observed. After an initial period of neuronal degeneration, reenervation of the cryopreserved autograft segment occurred 6 to 12 months after the operation. Findings regarding the fresh autografts were similar to those of the cryopreserved autografts. CONCLUSION: Our results suggest that the immunologic reaction rather than the cryopreservation process is responsible for the degenerative process occurring in cryopreserved homografts.