Controle de anaplasmose bovina através de imunização com Anaplasma centrale / Bovine anaplasmosis control through immunization with Anaplasma centrale

A anaplasmose bovina é uma das principais causas de perdas produtivas e mortes no Rio Grande do Sul em rebanhos bovinos. O Anaplasma marginale é o principal agente causador da enfermidade e provoca hipertermia, anemia, prostração, abortos e perdas produtivas nos bovinos acometidos. Tendo em vista o controle deste hemoparasita em uma propriedade leiteira localizada no município de Eldorado do Sul no Rio Grande do Sul, na qual a incidência da doença era alta, 471 animais foram imunizados com Anaplasma centrale na busca de desenvolvimento cruzado para Anaplasma marginale. No experimento foi verificado que a incidência que normalmente era acima de 30% na propriedade passou para níveis inferiores a 5%. No entanto, foram verificados abortos decorrentes da imunização, principalmente nos animais que possuíam menos de 90 dias de prenhes. Já o número de mortes globais na fazenda caiu consideravelmente tendo em vista que a principal causa de morte era a anaplasmose bovina. Dos animais inoculados com A. centrale em torno de 15% apresentaram sintomatologia clínica da enfermidade e precisaram ser tratados com oxitetraciclina no período entre 15 e 30 dias após a imunização. O custo com tratamento empregado na propriedade posterior à imunização caiu em torno de 85% o que provocou impacto significativo economicamente na propriedade.(AU)

Bovine anaplasmosis is a major cause of production losses and deaths in Rio Grande do Sul cattle herds. Anaplasma sp. is the main causative agent of cattle disease. It causes hyperthermia, anemia, prostration, abortions and reduces milk production in affected animals. In order to control this hemoparasite on a dairy farm located in the municipality of Eldorado do Sul in Rio Grande do Sul, where the disease incidence was high, 471 animals were immunized with Anaplasma centrale in the search for cross-protectiv immunity for Anaplasma marginale. The property anaplasmosis incidence, which usually was above 30%, became 5% after the immunization. However, abortions were observed resulting from innoculaition, especially in animals that had less than 90 days of pregnancy. The global number of deaths on the farm dropped considerably given that the main cause of death was the bovine anaplasmosis. 15% of animals inoculated with A. centrale showed clinical symptoms of the disease between 15 and 30 days after immunization and had to be treated with oxytetracycline. The amount of money spent with anaplasmosis treatment decay 85% after the immunization, which caused significant economic impact on the property.(AU)

Controle de anaplasmose bovina através de imunização com Anaplasma centrale / Bovine anaplasmosis control through immunization with Anaplasma centrale

A anaplasmose bovina é uma das principais causas de perdas produtivas e mortes no Rio Grande do Sul em rebanhos bovinos. O Anaplasma marginale é o principal agente causador da enfermidade e provoca hipertermia, anemia, prostração, abortos e perdas produtivas nos bovinos acometidos. Tendo em vista o controle deste hemoparasita em uma propriedade leiteira localizada no município de Eldorado do Sul no Rio Grande do Sul, na qual a incidência da doença era alta, 471 animais foram imunizados com Anaplasma centrale na busca de desenvolvimento cruzado para Anaplasma marginale. No experimento foi verificado que a incidência que normalmente era acima de 30% na propriedade passou para níveis inferiores a 5%. No entanto, foram verificados abortos decorrentes da imunização, principalmente nos animais que possuíam menos de 90 dias de prenhes. Já o número de mortes globais na fazenda caiu consideravelmente tendo em vista que a principal causa de morte era a anaplasmose bovina. Dos animais inoculados com A. centrale em torno de 15% apresentaram sintomatologia clínica da enfermidade e precisaram ser tratados com oxitetraciclina no período entre 15 e 30 dias após a imunização. O custo com tratamento empregado na propriedade posterior à imunização caiu em torno de 85% o que provocou impacto significativo economicamente na propriedade.(AU)

Bovine anaplasmosis is a major cause of production losses and deaths in Rio Grande do Sul cattle herds. Anaplasma sp. is the main causative agent of cattle disease. It causes hyperthermia, anemia, prostration, abortions and reduces milk production in affected animals. In order to control this hemoparasite on a dairy farm located in the municipality of Eldorado do Sul in Rio Grande do Sul, where the disease incidence was high, 471 animals were immunized with Anaplasma centrale in the search for cross-protectiv immunity for Anaplasma marginale. The property anaplasmosis incidence, which usually was above 30%, became 5% after the immunization. However, abortions were observed resulting from innoculaition, especially in animals that had less than 90 days of pregnancy. The global number of deaths on the farm dropped considerably given that the main cause of death was the bovine anaplasmosis. 15% of animals inoculated with A. centrale showed clinical symptoms of the disease between 15 and 30 days after immunization and had to be treated with oxytetracycline. The amount of money spent with anaplasmosis treatment decay 85% after the immunization, which caused significant economic impact on the property.(AU)

Controle de anaplasmose bovina através de imunização com Anaplasma centrale

ABSTRACT: Bovine anaplasmosis is a major cause of production losses and deaths in Rio Grande do Sul cattle herds. Anaplasma sp. is the main causative agent of cattle disease. It causes hyperthermia, anemia, prostration, abortions and reduces milk production in affected animals. In order to control this hemoparasite on a dairy farm located in the municipality of Eldorado do Sul in Rio Grande do Sul, where the disease incidence was high, 471 animals were immunized with Anaplasma centrale in the search for cross-protectiv immunity for Anaplasma marginale. The property anaplasmosis incidence, which usually was above 30%, became 5% after the immunization. However, abortions were observed resulting from innoculaition, especially in animals that had less than 90 days of pregnancy. The global number of deaths on the farm dropped considerably given that the main cause of death was the bovine anaplasmosis. 15% of animals inoculated with A. centrale showed clinical symptoms of the disease between 15 and 30 days after immunization and had to be treated with oxytetracycline. The amount of money spent with anaplasmosis treatment decay 85% after the immunization, which caused significant economic impact on the property.

RESUMO: A anaplasmose bovina é uma das principais causas de perdas produtivas e mortes no Rio Grande do Sul em rebanhos bovinos. O Anaplasma marginale é o principal agente causador da enfermidade e provoca hipertermia, anemia, prostração, abortos e perdas produtivas nos bovinos acometidos. Tendo em vista o controle deste hemoparasita em uma propriedade leiteira localizada no município de Eldorado do Sul no Rio Grande do Sul, na qual a incidência da doença era alta, 471 animais foram imunizados com Anaplasma centrale na busca de desenvolvimento cruzado para Anaplasma marginale. No experimento foi verificado que a incidência que normalmente era acima de 30% na propriedade passou para níveis inferiores a 5%. No entanto, foram verificados abortos decorrentes da imunização, principalmente nos animais que possuíam menos de 90 dias de prenhes. Já o número de mortes globais na fazenda caiu consideravelmente tendo em vista que a principal causa de morte era a anaplasmose bovina. Dos animais inoculados com A. centrale em torno de 15% apresentaram sintomatologia clínica da enfermidade e precisaram ser tratados com oxitetraciclina no período entre 15 e 30 dias após a imunização. O custo com tratamento empregado na propriedade posterior à imunização caiu em torno de 85% o que provocou impacto significativo economicamente na propriedade.

Effects of oocyte source, cell origin, and embryo reconstruction procedures on in vitro and in vivo embryo survival after goat cloning

The birth of cloned goats has been well documented, but the overall goat cloning efficiency by somatic cell nuclear transfer procedures is still low, which may be further intensified in extreme environments. The aim of this study was to produce cloned goats under the conditions of the Brazilian Semi Arid region, in a transgenic program for the expression of human lysozyme in the milk to target childhood diarrhea and malnutrition, comparing the effects of oocyte source, cell type, and embryo reconstruction procedures on in vitro and in vivo embryo survival after cloning by micromanipulation or by handmade cloning. The use of in vitro-matured oocytes resulted in more viable embryos after cloning than in vivo-matured cytoplasts, but no differences in pregnancy rates on day 23 were seen between oocyte sources (77.5 vs. 77.8%, respectively). The presence or absence of the zona pellucida for embryo reconstruction (78.8 vs. 76.0%, respectively) did not affect pregnancy outcome after transfer. However, pregnancy rate on day 23 was higher for embryos chemically activated by a conventional than a modified protocol (88.1 vs. 50.0%), and for embryos reconstructed with mesenchymal stem cells and fetal fibroblasts (100.0 and 93.3%) than with adult fibroblasts (64.7%). Although most pregnancies were lost, the birth of a cloned female was obtained from embryos reconstructed by micromanipulation using non-transgenic control cells and in vitro-matured oocytes with intact zona pellucida, after conventional activation and transfer at the 1-cell stage.

Effects of oocyte source, cell origin, and embryo reconstruction procedures on in vitro and in vivo embryo survival after goat cloning

The birth of cloned goats has been well documented, but the overall goat cloning efficiency by somatic cell nuclear transfer procedures is still low, which may be further intensified in extreme environments. The aim of this study was to produce cloned goats under the conditions of the Brazilian Semi Arid region, in a transgenic program for the expression of human lysozyme in the milk to target childhood diarrhea and malnutrition, comparing the effects of oocyte source, cell type, and embryo reconstruction procedures on in vitro and in vivo embryo survival after cloning by micromanipulation or by handmade cloning. The use of in vitro-matured oocytes resulted in more viable embryos after cloning than in vivo-matured cytoplasts, but no differences in pregnancy rates on day 23 were seen between oocyte sources (77.5 vs. 77.8%, respectively). The presence or absence of the zona pellucida for embryo reconstruction (78.8 vs. 76.0%, respectively) did not affect pregnancy outcome after transfer. However, pregnancy rate on day 23 was higher for embryos chemically activated by a conventional than a modified protocol (88.1 vs. 50.0%), and for embryos reconstructed with mesenchymal stem cells and fetal fibroblasts (100.0 and 93.3%) than with adult fibroblasts (64.7%). Although most pregnancies were lost, the birth of a cloned female was obtained from embryos reconstructed by micromanipulation using non-transgenic control cells and in vitro-matured oocytes with intact zona pellucida, after conventional activation and transfer at the 1-cell stage.(AU)

Effectiveness of microwell-based in vitro culture systems for zona-free cloned bovine embryos

Background: Modified embryo in vitro culture (IVC) systems have been devised for the culture of individual embryos, suchas in microdroplets, capillary tubes, or co-cultures systems. However, the development of the Well-of-the-Well (WOW)system takes advantage of both the individual and the grouped culture systems, as embryos are cultured into microwellsdisposed into a larger well. The aim of this study was to evaluate the in vitro and in vivo developmental potential of handmade cloning (HMC)-derived cloned bovine embryos after the IVC in three microwell (WOW) systems.Material, Methods & Results: Adipose tissue-derived mesenchymal stem cells (MSCs) isolated after subcutaneous biopsy from a Nellore adult female were used as nucleus donor cells for cloning. Collected adipose tissue was incubated in0.075% collagenase for 60 min at 39oC. After digestion, cell suspension was centrifuged at 80 g for 10 min, supernatantwas discarded, and 2 mL of Red Blood Cells Lysis Buffer (RBC) was added to the pellet, remaining for 2 min. The RBCbuffer was diluted with 20 mL PBS, with cell suspension spun at 80 g for 5 min. Supernatant was discarded and the pelletre-suspended in DMEM culture medium + 10% fetal calf serum (FCS). Following cell counting, 5 x 105 cells were seededin 25-cm² bottles containing 6 mL DMEM + 10% FCS, and cultured at 38.3°C, 5% CO2 in air and saturated humidity.After IVM, oocytes were denuded and incubated in demecolcine, followed by zona pellucida removal, oocyte bisection,embryo reconstruction, electrofusion and chemical activation. Cloned embryos were allocated to one of three IVC groups:cWOW: conventional microwells (250 µm, round); mWOW: modified microwells (130 µm, conical); and WOW-PDMS:microwells in polydimethylsiloxane chips (170 µm, cylindrical, with interconnecting microchannels); IVF embryos wereused as controls. Cleavage (D2), blastocyst (D7) and pregnancy (D30) rates were analyzed by the χ2 test, for P < 0.05...(AU)

Effectiveness of microwell-based in vitro culture systems for zona-free cloned bovine embryos

Background: Modified embryo in vitro culture (IVC) systems have been devised for the culture of individual embryos, suchas in microdroplets, capillary tubes, or co-cultures systems. However, the development of the Well-of-the-Well (WOW)system takes advantage of both the individual and the grouped culture systems, as embryos are cultured into microwellsdisposed into a larger well. The aim of this study was to evaluate the in vitro and in vivo developmental potential of handmade cloning (HMC)-derived cloned bovine embryos after the IVC in three microwell (WOW) systems.Material, Methods & Results: Adipose tissue-derived mesenchymal stem cells (MSCs) isolated after subcutaneous biopsy from a Nellore adult female were used as nucleus donor cells for cloning. Collected adipose tissue was incubated in0.075% collagenase for 60 min at 39oC. After digestion, cell suspension was centrifuged at 80 g for 10 min, supernatantwas discarded, and 2 mL of Red Blood Cells Lysis Buffer (RBC) was added to the pellet, remaining for 2 min. The RBCbuffer was diluted with 20 mL PBS, with cell suspension spun at 80 g for 5 min. Supernatant was discarded and the pelletre-suspended in DMEM culture medium + 10% fetal calf serum (FCS). Following cell counting, 5 x 105 cells were seededin 25-cm² bottles containing 6 mL DMEM + 10% FCS, and cultured at 38.3°C, 5% CO2 in air and saturated humidity.After IVM, oocytes were denuded and incubated in demecolcine, followed by zona pellucida removal, oocyte bisection,embryo reconstruction, electrofusion and chemical activation. Cloned embryos were allocated to one of three IVC groups:cWOW: conventional microwells (250 µm, round); mWOW: modified microwells (130 µm, conical); and WOW-PDMS:microwells in polydimethylsiloxane chips (170 µm, cylindrical, with interconnecting microchannels); IVF embryos wereused as controls. Cleavage (D2), blastocyst (D7) and pregnancy (D30) rates were analyzed by the χ2 test, for P < 0.05...

In vitro development and cell allocation after aggregation of syngeneic wild type and fluorescence-expressing bovine cloned embryos

Background: The in vitro production (IVP) of embryos by in vitro fertilization or cloning procedures has been known to cause epigenetic changes in the conceptus that in turn are associated with abnormalities in pre-and postnatal development. Handmade cloning (HMC) procedures and the culture of zona-free embryos in individual microwells provide excellent tools for studies in developmental biology, since embryo development and cell allocation patterns can be evaluated under a wide range of embryo reconstruction arrangements and in in vitro embryo culture conditions. As disturbances in embryonic cell allocation after in vitro embryo manipulations and unusual in vivo conditions during the first third of pregnancy appear to be associated with large offspring, embryo aggregation procedures may allow a compensation for epigenetic defects between aggregated embryos or even may influence more favorable cell allocation in embryonic lineages, favoring subsequent development. Thus, the aim of this study was to evaluate in vitro embryo developmental potential and the pattern of cell allocation in blastocysts developed after the aggregation of handmade cloned embryos produced using syngeneic wild type and/or transgenic somatic cells. Materials, Methods & Results: In vitro-matured bovine cumulus-oocyte complexes (COC) were manually bisected after cumulus and zona pellucida removal; then, two enucleated hemi-oocytes were paired and fused with either a wild type (WT) or a GFP-expressing (GFP) fetal skin cell at the 11th and 19th passages, respectively. Following chemical activation, reconstructed cloned embryos and zona-free parthenote embryos were in vitro-cultured in microwells, for 7 days, either individually (1 x 100%) or after the aggregation of two structures (2 x 100%) per microwell, as follows: (G1) one WT cloned embryo; (G2) two aggregated WT embryos; (G3) one GFP cloned embryo; (G4) two aggregated GFP embryos; (G5) aggregation of a WT embryo and a GFP embryo; (G6) one parthenote embryo; or (G7) two aggregated parthenote embryos. Fusion (clones), cleavage (Day 2), and blastocyst (Day 7) rates, and embryonic cell allocation were compared by the x² or Fisher tests. Total cell number (TCN) in blastocysts was analyzed by the Student's test (P < 0.05). Fusion and cleavage rates, and cell allocation were similar between groups. On a per WOW basis, development to the blastocyst stage was similar between groups, except for lower rates of development seen in G3. However, when based on number of embryos per group (one or two), blastocyst development was higher in G1 than all other groups, which were similar between one another. Cloned GFP embryos had lower in vitro development to the blastocyst stage than WT embryos, which had more TCN than parthenote or aggregated chimeric WT/GFP embryos. Aggregated GFP embryos had fewer cells than the other embryo groups. Discussion: The in vitro development of GFP cloned embryos was lower than WT embryos, with no effects on cell allocation in resulting blastocysts. Differences in blastocyst rate between groups were likely due to lower GFP-expressing cell viability, as GFP donor cells were at high population cell doublings when used for cloning. On a per embryo basis, embryo aggregation on Day 1 resulted in blastocyst development similar to non-aggregated embryos on Day 7, with no differences in cell proportion between groups. The use of GFP-expressing cells was proven a promising strategy for the study of cell allocation during embryo development, which may assist in the elucidation of mechanisms of abnormalities after in vitro embryo manipulations, leading to the development of improved protocols for the in vitro production (IVP) of bovine embryos.

In vitro development and cell allocation after aggregation of syngeneic wild type and fluorescence-expressing bovine cloned embryos / In vitro development and cell allocation after aggregation of syngeneic wild type and fluorescence-expressing bovine cloned embryos

Materials, Methods & Results: In vitro-matured bovine cumulus-oocyte complexes (COC) were manually bisected after cumulus and zona pellucida removal; then, two enucleated hemi-oocytes were paired and fused with either a wild type (WT) or a GFP-expressing (GFP) fetal skin cell at the 11th and 19th passages, respectively. Following chemical activation, reconstructed cloned embryos and zona-free parthenote embryos were in vitro-cultured in microwells, for 7 days, either individually (1 x 100%) or after the aggregation of two structures (2 x 100%) per microwell, as follows: (G1) one WT cloned embryo; (G2) two aggregated WT embryos; (G3) one GFP cloned embryo; (G4) two aggregated GFP embryos; (G5) aggregation of a WT embryo and a GFP embryo; (G6) one parthenote embryo; or (G7) two aggregated parthenote embryos. Fusion (clones), cleavage (Day 2), and blastocyst (Day 7) rates, and embryonic cell allocation were compared by the 2 or Fisher tests. Total cell number (TCN) in blastocysts was analyzed by the Student´s test (P 0.05). Fusion and cleavage rates, and cell allocation were similar between groups. On a per WOW basis, development to the blastocyst stage was similar between groups, except for lower rates of development seen in G3. However, when based on number of embryos per group (one or two), blastocyst development was higher in G1 than all other groups, which were simi

Background: The in vitro production (IVP) of embryos by in vitro fertilization or cloning procedures has been known to cause epigenetic changes in the conceptus that in turn are associated with abnormalities in pre- and postnatal development. Handmade cloning (HMC) procedures and the culture of zona-free embryos in individual microwells provide excellent tools for studies in developmental biology, since embryo development and cell allocation patterns can be evaluated under a wide range of embryo reconstruction arrangements and in in vitro embryo culture conditions. As disturbances in embryonic cell allocation after in vitro embryo manipulations and unusual in vivo conditions during the fi rst third of pregnancy appear to be associated with large offspring, embryo aggregation procedures may allow a compensation for epigenetic defects between aggregated embryos or even may infl uence more favorable cell allocation in embryonic lineages, favoring subsequent development. Thus, the aim of this study was to evaluate in vitro embryo developmental potential and the pattern of cell allocation in blastocysts developed after the aggregation of handmade cloned embryos produced using syngeneic wild type and/or transgenic somatic cells.Materials, Methods & Results: In vitro-matured bovine cumulus-oocyte complexes (COC) were manually bisected after cumulus and zona pellucida removal; then

Clonagem animal: a sobrevivência dos mais aptos / Animal cloning: survival of the fittest

A clonagem animal por transferência nuclear de célula somática (TNCS) apresenta inúmeras aplicações científicas e comerciais, incluindo a produção de animais transgênicos, a preservação de animais de genética desejável, rara ou em extinção, ou mesmo a aplicação para o estudo de aspectos básicos em biologia molecular, celular e do desenvolvimento. Não obstante, a clonagem por TNCS ainda é ineficiente, com menos de 5% dos embriões clones produzidos resultando em animais nascidos vivos. O sucesso na clonagem exige o exímio domínio técnico e científico de várias disciplinas e áreas de conhecimento, havendo pelo menos cinco etapas críticas no processo associadas a falhas de desenvolvimento, desde a produção in vitro dos embriões até o nascimento de um animal viável. A identificação de fatores associados às falhas em cada etapa, em especial aqueles relacionados ao oócito receptor (citoplasto), à célula doadora (carioplasto) e aos procedimentos técnicos per se de produção de embriões clones, além da observação cuidadosa dos sinais de anormalidades subsequentes à transferência dos embriões para fêmeas receptoras, é essencial para a optimização de todos os procedimentos para a obtenção, em seu final, de um animal clonado viável e que sobreviva até a vida adulta. Esta revisão visa descrever alguns eventos técnicos e biológicos associados ao sucesso e/ou insucesso da clonagem animal.

Animal cloning by somatic cell nuclear transfer (SCNT) has numerous scientific and commercial applications, including the production of transgenic animals, preservation of animals from desirable or rare gene pools, and animals in risk of extinction, or even for the study of basic aspects in molecular, cell and developmental biology. Nevertheless, cloning by SCNT is still inefficient, with less than 5% of cloned embryos resulting in liveborn animals. The cloning success depends on a proficient technical and scientific know-how of a number of disciplines and areas of knowledge, with at least five critical steps in the process associated with developmental failures, from the in vitro production of cloned embryos through the birth of a viable animal. The identification of factors associated with failures in each step, in special to those related to the recipient oocyte (cytoplast), to the nucleus donor cell (karyoplast), and to the technical procedures for the production of cloned embryos per se, along with the careful observation of signs of abnormalities following the transfer of embryos to recipient females, is essential for the optimization of procedures that, ultimately, may result in a cloned animal that survives to adulthood. This review aims to discuss some technical and biological events associated with success and/or failure in animal cloning.

Clonagem animal: a sobrevivência dos mais aptos / Animal cloning: survival of the fittest

A clonagem animal por transferência nuclear de célula somática (TNCS) apresenta inúmeras aplicações científicas e comerciais, incluindo a produção de animais transgênicos, a preservação de animais de genética desejável, rara ou em extinção, ou mesmo a aplicação para o estudo de aspectos básicos em biologia molecular, celular e do desenvolvimento. Não obstante, a clonagem por TNCS ainda é ineficiente, com menos de 5% dos embriões clones produzidos resultando em animais nascidos vivos. O sucesso na clonagem exige o exímio domínio técnico e científico de várias disciplinas e áreas de conhecimento, havendo pelo menos cinco etapas críticas no processo associadas a falhas de desenvolvimento, desde a produção in vitro dos embriões até o nascimento de um animal viável. A identificação de fatores associados às falhas em cada etapa, em especial aqueles relacionados ao oócito receptor (citoplasto), à célula doadora (carioplasto) e aos procedimentos técnicos per se de produção de embriões clones, além da observação cuidadosa dos sinais de anormalidades subsequentes à transferência dos embriões para fêmeas receptoras, é essencial para a optimização de todos os procedimentos para a obtenção, em seu final, de um animal clonado viável e que sobreviva até a vida adulta. Esta revisão visa descrever alguns eventos técnicos e biológicos associados ao sucesso e/ou insucesso da clonagem animal.(AU)

Animal cloning by somatic cell nuclear transfer (SCNT) has numerous scientific and commercial applications, including the production of transgenic animals, preservation of animals from desirable or rare gene pools, and animals in risk of extinction, or even for the study of basic aspects in molecular, cell and developmental biology. Nevertheless, cloning by SCNT is still inefficient, with less than 5% of cloned embryos resulting in liveborn animals. The cloning success depends on a proficient technical and scientific know-how of a number of disciplines and areas of knowledge, with at least five critical steps in the process associated with developmental failures, from the in vitro production of cloned embryos through the birth of a viable animal. The identification of factors associated with failures in each step, in special to those related to the recipient oocyte (cytoplast), to the nucleus donor cell (karyoplast), and to the technical procedures for the production of cloned embryos per se, along with the careful observation of signs of abnormalities following the transfer of embryos to recipient females, is essential for the optimization of procedures that, ultimately, may result in a cloned animal that survives to adulthood. This review aims to discuss some technical and biological events associated with success and/or failure in animal cloning.(AU)

Intoxicação por Baccharis megapotamica var. weiriiem ovinos / Spontaneous Baccharis megapotamica var. weirii poisoning in sheep

As intoxicações por Baccharis coridifolia afetam especialmente animais de fazenda famintos ou curiosos e que não haviam tido contato prévio com a planta. B. coridifolia ocorre usualmente em terrenos secos de coxilhas no Rio Grande do Sul e estados ou países vizinhos. Uma forma indistinguível e esporádica da doença tem sido associada com a ingestão de Baccharis megapotamica que ocorre em áreas úmidas. Relata-se a intoxicação natural de quatro cordeiros após ingestão de Baccharis megapotamica var. weirii. A doença foi observada em uma propriedade localizada em Barra do Ribeiro, Rio Grande do Sul. O rebanho era composto por 220 ovinos, os quais eram mantidos em área de pastagem nativa sem qualquer suplementação. Uma rápida doença clínica caracterizada por anorexia, cólica, diarréia e desidratação causou a morte de três cordeiros em um período de 8 a 24 horas, o outro foi encontrado morto. A necropsia revelou alterações significativas no rúmen, no qual havia edema de serosa e hemorragias equimóticas na submucosa. Microscopicamente, o rúmen apresentou edema de submucosa, além de edema, tumefação, vacuolização e necrose de mucosa. O diagnóstico foi fundamentado nos achados clínicos, patológicos e epidemiológicos.(AU)

Spontaneous poisoning of livestock by Baccharis coridifolia affects mostly hungry or curious animals that have not entered in contact with the plant previously. The plant occurs usually in dried hilly soils of Rio Grande do Sul and neighboring states or countries. An indistinguishable and sporadic form of the disease has been associated with the ingestion of Baccharis megapotamica, a species occurring in moist areas. This communication reports the spontaneous poisoning of four lambs after ingestion of B. megapotamica var. weirii. Clinical signs were observed in three lambs, the other was found dead. A rapid clinical disease characterized by anorexia, colic, dehydration, and diarrhea resulted in death after 8-24 hours. Necropsy revealed significant changes in the rumen, in which serosal edema, and submucosal echimotic hemorrhages were observed. Microscopically, the rumen showed submucosal edema and mucosal tumefaction, vacuolization, and necrosis. Diagnosis was based on clinical, pathological and epidemiological findings.(AU)

Uso de oócitos bovinos como citoplasma receptor na produção de embriões por transferência nuclear de célula somática interespécie (NTSCi)

Interspecies embryo clones have been produced by research groups with relative success in some species. Bovine oocytes matured in vitro and enucleated by micromanipulation were used in three experiments as recipeient cytoplasm in nuclear transfer of ovine, caprine and porcine fibroblasts. The fibroblasts were cultivated until the third passage before being frozen and used. The electrofusion was induced by an application of a 20V pulse during 45 ms. The activation was done with 5 mM ionomycin and subsequently 2 mM 6DMAP. NTSC bovine embryos, NTSCi caprine and ovine embryos were cultivated in SOF medium and NTSCi porcine embryos were cultivated in NCSU23 medium. The fusion rates of the reconstructed complexes with bovine cells did not differ from those observed with ovine cells (88.2%), caprine cells (74.1%) and porcine cells (79.4%). The cleavage rates in ovine (60.3%), caprine (68.4%) and porcine (57.1%) NTSCi groups did not differ from the control group NTSC bovine. The blastocyst rate observed in the group of NTSCi ovine embryos (10.3%) was similar to the group of NTSC bovine embryos (12.7%). In NTSCi caprine embryos, 5.3% of the embryos developed up to the blastocyst stage, while in the NTSCi porcine group there was no development up to the blastocyst stage. In conclusion, the bovine cytoplasm was able to support the embryo development in NTSCi up to the blastocyst stage usi

Otimização do sistema de produção de clones por transferência nuclear de célula somática (NTSC)

A técnica de transferência nuclear é uma ferramenta que possibilita a produção de embriões clones que podem ser utilizados tanto na clonagem reprodutiva como no modelo para o estudo de diversos mecanismos fisiológicos durante o desenvolvimento embrionário. Neste sentido, embriões clones bovinos foram produzidos por transferência nuclear, com o objetivo de estabelecer a técnica de clonagem, bem como otimizála para as condições do Laboratório de Embriologia e Biotécnicas de Reprodução da UFRGS. Durante este estudo, 1.123 estruturas foram reconstruídas em diferentes condições, sendo que 95 blastocistos foram produzidos em 56 replicações. O primeiro blastocisto foi produzido na 5ª rotina. Após a transferência de parte destes embriões, 13 prenhezes foram estabelecidas, entretanto, a maioria delas foi interrompida no terço inicial da gestação. Uma prenhez gemelar alcançou 260 dias, momento em que os fetos foram abortados. Outra gestação foi a termo, com o nascimento de um clone vivo, porém, o animal veio a óbito 42 horas após o nascimento.

Bovine in vitro embryo production protocol: does it really influence embryo cryotolerance?

The protocol of  in vitro production (IVP) of bovine embryos is one of the critical factors determining embryo viability after cryopreservation. In this study were used two differents protocols to produce IVP bovine embryos, with variations in protein source, oocyte/zygote density per media volume, with the aim to determine the in vitro and in vivo embryo survival after vitrification using hand-pulled glass micropipettes. Expanded blastocysts (D7) were morphologically selected by size (?180 ?m) and osmotic behavior before they were randomly allocated to sub-groups by protocol: non-vitrified embryos (control; C) and vitrified embryos (V). For the evaluation of the in vitro survival, control embryos and a group of warmed vitrified embryos were in vitro-cultured (IVC) for 72 h. Re-expansion rates of warmed embryos at 24 h of IVC were 94.8% and 93.2% for Protocols 1 and 2, respectively. Hatching rates at 72 h of IVC of embryos from Protocol 1 (C=80% and V=75.8%) tended to be higher (P=0.0561, Chi2 test) than those from Protocol 2 (C=67.2% and V=59.3%). For the evaluation of in vivo survival, 21 vitrified embryos per protocol were singly non-surgically transferred to synchronized recipients (n=42) after the in-straw cryoprotectant dilution, resulting in 4 (19%) pregnancies per group on Day 60 of gestation. In conclusion, despite a lower variation on in vitro embryo development betwe

Bovine in vitro embryo production protocol: does it really influence embryo cryotolerance?

The protocol of  in vitro production (IVP) of bovine embryos is one of the critical factors determining embryo viability after cryopreservation. In this study were used two differents protocols to produce IVP bovine embryos, with variations in protein source, oocyte/zygote density per media volume, with the aim to determine the in vitro and in vivo embryo survival after vitrification using hand-pulled glass micropipettes. Expanded blastocysts (D7) were morphologically selected by size (?180 ?m) and osmotic behavior before they were randomly allocated to sub-groups by protocol: non-vitrified embryos (control; C) and vitrified embryos (V). For the evaluation of the in vitro survival, control embryos and a group of warmed vitrified embryos were in vitro-cultured (IVC) for 72 h. Re-expansion rates of warmed embryos at 24 h of IVC were 94.8% and 93.2% for Protocols 1 and 2, respectively. Hatching rates at 72 h of IVC of embryos from Protocol 1 (C=80% and V=75.8%) tended to be higher (P=0.0561, Chi2 test) than those from Protocol 2 (C=67.2% and V=59.3%). For the evaluation of in vivo survival, 21 vitrified embryos per protocol were singly non-surgically transferred to synchronized recipients (n=42) after the in-straw cryoprotectant dilution, resulting in 4 (19%) pregnancies per group on Day 60 of gestation. In conclusion, despite a lower variation on in vitro embryo development betwe

Bovine in vitro embryo production protocol: does it really influence embryo cryotolerance?

The protocol of  in vitro production (IVP) of bovine embryos is one of the critical factors determining embryo viability after cryopreservation. In this study were used two differents protocols to produce IVP bovine embryos, with variations in protein source, oocyte/zygote density per media volume, with the aim to determine the in vitro and in vivo embryo survival after vitrification using hand-pulled glass micropipettes. Expanded blastocysts (D7) were morphologically selected by size (?180 ?m) and osmotic behavior before they were randomly allocated to sub-groups by protocol: non-vitrified embryos (control; C) and vitrified embryos (V). For the evaluation of the in vitro survival, control embryos and a group of warmed vitrified embryos were in vitro-cultured (IVC) for 72 h. Re-expansion rates of warmed embryos at 24 h of IVC were 94.8% and 93.2% for Protocols 1 and 2, respectively. Hatching rates at 72 h of IVC of embryos from Protocol 1 (C=80% and V=75.8%) tended to be higher (P=0.0561, Chi2 test) than those from Protocol 2 (C=67.2% and V=59.3%). For the evaluation of in vivo survival, 21 vitrified embryos per protocol were singly non-surgically transferred to synchronized recipients (n=42) after the in-straw cryoprotectant dilution, resulting in 4 (19%) pregnancies per group on Day 60 of gestation. In conclusion, despite a lower variation on in vitro embryo development betwe

Otimização do sistema de produção de clones por transferência nuclear de célula somática (NTSC)

A técnica de transferência nuclear é uma ferramenta que possibilita a produção de embriões clones que podem ser utilizados tanto na clonagem reprodutiva como no modelo para o estudo de diversos mecanismos fisiológicos durante o desenvolvimento embrionário. Neste sentido, embriões clones bovinos foram produzidos por transferência nuclear, com o objetivo de estabelecer a técnica de clonagem, bem como otimizála para as condições do Laboratório de Embriologia e Biotécnicas de Reprodução da UFRGS. Durante este estudo, 1.123 estruturas foram reconstruídas em diferentes condições, sendo que 95 blastocistos foram produzidos em 56 replicações. O primeiro blastocisto foi produzido na 5ª rotina. Após a transferência de parte destes embriões, 13 prenhezes foram estabelecidas, entretanto, a maioria delas foi interrompida no terço inicial da gestação. Uma prenhez gemelar alcançou 260 dias, momento em que os fetos foram abortados. Outra gestação foi a termo, com o nascimento de um clone vivo, porém, o animal veio a óbito 42 horas após o nascimento.

Uso de oócitos bovinos como citoplasma receptor na produção de embriões por transferência nuclear de célula somática interespécie (NTSCi)

Interspecies embryo clones have been produced by research groups with relative success in some species. Bovine oocytes matured in vitro and enucleated by micromanipulation were used in three experiments as recipeient cytoplasm in nuclear transfer of ovine, caprine and porcine fibroblasts. The fibroblasts were cultivated until the third passage before being frozen and used. The electrofusion was induced by an application of a 20V pulse during 45 ms. The activation was done with 5 mM ionomycin and subsequently 2 mM 6DMAP. NTSC bovine embryos, NTSCi caprine and ovine embryos were cultivated in SOF medium and NTSCi porcine embryos were cultivated in NCSU23 medium. The fusion rates of the reconstructed complexes with bovine cells did not differ from those observed with ovine cells (88.2%), caprine cells (74.1%) and porcine cells (79.4%). The cleavage rates in ovine (60.3%), caprine (68.4%) and porcine (57.1%) NTSCi groups did not differ from the control group NTSC bovine. The blastocyst rate observed in the group of NTSCi ovine embryos (10.3%) was similar to the group of NTSC bovine embryos (12.7%). In NTSCi caprine embryos, 5.3% of the embryos developed up to the blastocyst stage, while in the NTSCi porcine group there was no development up to the blastocyst stage. In conclusion, the bovine cytoplasm was able to support the embryo development in NTSCi up to the blastocyst stage usi

Otimização do sistema de produção de clones por transferência nuclear de célula somática (NTSC)

A técnica de transferência nuclear é uma ferramenta que possibilita a produção de embriões clones que podem ser utilizados tanto na clonagem reprodutiva como no modelo para o estudo de diversos mecanismos fisiológicos durante o desenvolvimento embrionário. Neste sentido, embriões clones bovinos foram produzidos por transferência nuclear, com o objetivo de estabelecer a técnica de clonagem, bem como otimizála para as condições do Laboratório de Embriologia e Biotécnicas de Reprodução da UFRGS. Durante este estudo, 1.123 estruturas foram reconstruídas em diferentes condições, sendo que 95 blastocistos foram produzidos em 56 replicações. O primeiro blastocisto foi produzido na 5ª rotina. Após a transferência de parte destes embriões, 13 prenhezes foram estabelecidas, entretanto, a maioria delas foi interrompida no terço inicial da gestação. Uma prenhez gemelar alcançou 260 dias, momento em que os fetos foram abortados. Outra gestação foi a termo, com o nascimento de um clone vivo, porém, o animal veio a óbito 42 horas após o nascimento.