Il periparto è la parte più critica della vita della bovina da latte. I cambiamenti importanti ed improvvisi nel metabolismo e nelle condizioni di vita possono favorire l’insorgenza di problemi di salute e di uno stato infiammatorio. Una risposta eccessiva dell’organismo all’infiammazione può sottrarre risorse fondamentali, causando un peggioramento delle condizioni di salute e un calo della produzione di latte. Nella prima parte della tesi si propone un nuovo indice basato su alcuni parametri plasmatici utile a descrivere meglio la risposta all’infiammazione nel post-parto. Una identificazione rapida degli animali caratterizzati da una risposta eccessiva all’infiammazione, specialmente quelli senza sintomi esterni, permette un intervento efficace per evitare ulteriori conseguenze negative. La seconda e terza parte della tesi descrivono due prove sperimentali finalizzate a ridurre la risposta all’infiammazione impiegando acidi grassi ω-3 e/o acido acetilsalicilico. La somministrazione di acidi grassi ω-3 a cavallo del parto ha ridotto la risposta all’infiammazione e migliorato il bilancio energetico, mentre la loro somministrazione esclusivamente prima del parto ha evidenziato alcuni leggeri miglioramenti nella risposta all’infiammazione e nel bilancio energetico. La somministrazione prima del parto di acido acetilsalicilico ha causato l’insorgenza di alcuni problemi e pertanto dovrebbe essere somministrata solo dopo il parto. / The peripartum period is the most critical stage of the dairy cows. The sudden and major changes in metabolism and life conditions may favor the onset of health problems together with an inflammatory status. An excessive response by the organism to inflammation may subtract primary resources, resulting in a worsening of health status and in a lower milk yield. In the first part of the thesis a new index based on some plasma parameters is proposed to better describe the response to inflammation in the post-calving of dairy cows. An early identification of the animals characterized by a severe response to inflammation, especially without any external symptom, allows an effective intervention in order to avoid further negative consequences. The second and the third part of the thesis describe two experiments aiming to reduce the response to inflammation using ω-3 fatty acids and/or acetylsalicylic acid. The administration of ω-3 fatty acids around calving reduced the inflammatory response and improved the energy balance, while their pre-calving only administration highlighted some slight improvements in inflammatory response and energy balance. The pre-calving administration of acetylsalicylic acid caused the onset of some problems and should be used only after calving.

Incidences of uterine infections in dairy cattle are high between parturition and Day 21 postpartum. Dairy cows with uterine infections are at risk for prolonged periods of days open and multiple services before becoming pregnant. Neutrophils are the first wave of immune system defense against uterine contamination. Neutrophil function seems to be mediated by reproductive hormones and good uterine health is related to properly functioning neutrophils. To elucidate the interaction between reproductive hormones, neutrophils and uterine health in dairy cows the objectives of this research were to evaluate: 1) changes in circulating white blood cell populations during the estrous cycle, 2) the effects of prostaglandin F₂[subscript α](PGF₂[subscript α]) on circulating white blood cell populations and 3) the effects of a two-injection PGF₂[subscript α] regimen on uterine neutrophil and bacterial populations and numbers of days open and services per conception. In the first experiment, the effect of stage of the estrous cycle on plasma neutrophil numbers was evaluated. Neutrophils were observed throughout the entire estrous cycle and numbers were greatest (P<0.05) on Day 14 (mid-cycle), when the corpus luteum was the dominant ovarian structure and plasma progesterone was at its acme. In the second experiment, plasma neutrophil numbers were examined in cows after injections of saline or the PGF₂[subscript α] pharmaceutical product, Lutalyse. Compared to saline, numbers of neutrophils were greater (P<0.05) 4 and 8 hr after Lutalyse injection. In the third experiment, neutrophil numbers were examined after injections of saline, Lutalyse or the PGF₂[subscript α] analog, Estrumate. Compared to saline, numbers of neutrophils did not differ (P>0.10) from cows injected with Lutalyse or Estrumate. In the fourth experiment, uterine bacterial populations and numbers of neutrophils were quantified in cows treated with Lutalyse or saline on Days 0 and 14 or 14 and 28 postpartum. Compared to saline, Lutalyse treatment decreased (P<0.05) total bacteria present in the uterus and increased (P<0.05) the number of uterine neutrophils. In experiment five, numbers of days open and services per conception were evaluated in cows treated with Lutalyse or saline on Days 0 and 14 or 14 and 28 postpartum. Compared to saline, Lutalyse decreased days open (154.7 ± 14.1 vs. 120.1 ± 7.9 days, respectively; P<0.05) and services per conception (3.0 ± 0.4 vs. 2.3 ± 0.2 services, respectively; P=0.09). These results suggest PGF₂[subscript α] treatment can increase neutrophil and depress bacterial cell populations in favor of the dairy cow's uterine health and may explain why fertility is improved when PGF₂α is administered early in the postpartum period. / Graduation date: 2012

Downstream purification and analysis of a model biopharmaceutical protein (recombinant human myelin basic protein) is described. The recombinant protein was expressed in the milk of transgenic cows and was found exclusively associated with the casein micellar phase. Binding of milk calcium to the active sites of a cation exchanger resin was used beneficially in this study in order to gently disrupt the casein micelles and liberate the recombinant protein. This approach was found superior to the conventional micelle disruption procedures with respect to product recovery, resin fouling due to milk components and column hydrodynamic properties. Further purification was carried out using Ni2+ affinity chromatography and resulted in purity more than 90% and a total recovery of 78%. A capillary electrophoresis total protein assay employing large volume sample stacking and a microsphere-based, sandwich-type immunoassay were developed and validated. Both methods were successfully integrated with the downstream purification protocol in order to evaluate various quality attributes of the recombinant protein. A onestep capillary isoelectric focusing protocol was developed in order to monitor the recombinant protein in milk samples. The results showed extra protein bands in the transgenic milk that had isoelectric points significantly lower than the theoretically calculated one which indicated that the protein had been modified during expression. The association between the recombinant protein and bovine milk caseins was explored at the molecular level using the surface plasmon resonance technique. Results showed a calciummediated interaction between the recombinant protein and the phosphorylated caseins. This selective interaction was not noted between the human myelin basic protein and milk caseins which indicated mammary gland-related posttranslational modifications, most likely phosphorylation. The co-expression of the recombinant protein and caseins in the mammary gland, along with the ability of the recombinant protein to form calcium bridges with caseins explained its association with the casein micellar phase in the transgenic milk. Despite this and owing to the low expression levels of the recombinant protein in milk, light scattering investigations using diffusing wave spectroscopy showed no significant differences between the transgenic and the non-transgenic milk samples with respect to the average micelle size and the micelle surface charges.

A positive phenotypic correlation between milk protein percentage and reproductive performance in dairy cattle, especially during early lactation has been recently reported. The objective of this study was to quantify the relationship between milk protein percentage and different measures of fertility in Irish, seasonal calving, dairy cattle using data from experiments comparing strains of Holstein-Friesian cows under different feeding systems. The relationships between body condition score, milk production and fertility were also investigated. The data used in this study consisted of 584 lactation records over a 5-yr period. Principal component analysis and logistic regression was used to study the relationship between milk protein percentage and fertility performance of the cow. Greater milk protein percentage during the first 60 days post-calving was associated with better reproductive performance. The probability of a cow being submitted in the first 21 days of the breeding season increased with increased milk protein percentage during early lactation. Similarly, the probability of a cow becoming pregnant to its first service or to the whole breeding season also increased. Cows were classified as either high or low milk protein percentage based on their protein percentage over the whole lactation. Cows in the high milk protein group had a 7% greater conception rate compared to cows in the low protein percentage group. In conclusion, cows with higher protein percentage, especially during early lactation are submitted earlier in the breeding season, and have a higher conception rate. Physiologically, the shortage of glucose caused by negative energy balance restricts the synthesis of milk protein in the udder. On the other side, negative energy balance also causes the reduction of IGF-I, LH and oestradiol, which consequently delay the ovarian follicular development and finally reduces fertility. Therefore, there is a biological explanation for the association between milk protein percentage and fertility performance.

Diss. (sammanfattning) Umeå : Sveriges lantbruksuniv., 2007. / Härtill 3 uppsatser.

Diss. (sammanfattning) Uppsala : Sveriges lantbruksuniversitet, 2008. / Härtill 4 uppsatser.

Diss. (sammanfattning) Uppsala : Sveriges lantbruksuniv. / Härtill 5 uppsatser.

Diss. (sammanfattning). Uppsala : Sveriges lantbruksuniv. / Härtill 4 uppsatser.

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / The objective of this research was to evaluate three grazing systems with Coastcross-1 (CC) + 100 kg N/ha/year + common vetch; CC + 100 kg N/ha/year + arrowleaf clover; and CC + 200 kg N/ha/year. Lactating Holstein cows were used in the evaluation. The grazing method was the rotative stocking, with one to two days of occupation. Thirteen grazing cycles were performed during the experimental period (354 days). The actual mean value of forage on offer was 5.3% of body weight. The forage mass, botanical composition, leaf/stem ratio; daily accumulation rate, production and forage intake; grazing efficiency, stocking rate and nutritive value were evaluated. Experimental design was completely randomized with three treatments (grazing systems), three replicates (paddocks) in completely split-plot time (grazing/seasons). The average of forage production and stocking rate were 21.0, 20.3 and 24.3 t/ha/year; 7.0, 6.5 e 7.7 animal units/ha/day, respectively. Crude protein, neutral detergent fiber and in situ digestibility of dry matter average values were 18.1, 16.7 and 17.6%; 57.8, 58.9 and 58.7%; 79.6, 78.9 and 80.6%, for the respective systems. High productivity were found on Coastcross-1 mixed with common vetch + 100 kg N/ha/year and Coastcross-1 alone + 200 kg N/ha/year. Better results to nutritive value were found on winter, especially on Coastcross-1 mixed with common vetch. / Objetivou-se com esta pesquisa avaliar três sistemas forrageiros constituídos por: Coastcross-1(CC) + ervilhaca comum + 100 kg de N/ha/ano; CC + trevo vesiculoso + 100 kg de N/ha/ano e CC + 200 kg de N/ha/ano. Para avaliação foram utilizadas vacas em lactação da raça Holandesa. O método de pastejo utilizado foi o de lotação rotacionada, com um a dois dias de ocupação. Durante o período experimental (345 dias) foram realizados treze pastejos. O valor real médio de oferta de forragem foi de 5,3% do peso corporal. Foram avaliadas a massa de forragem, composição botânica, relação folha/colmo; taxa de acúmulo diário, produção e consumo de foragem; eficiência de pastejo, taxa de lotação e valor nutritivo da forragem. O delineamento experimental utilizado foi o inteiramente casualizado com três tratamentos (sistemas forrageiros), três repetições (piquetes) e parcelas subdivididas no tempo (pastejos/estações). Os valores médios de produção de forragem e taxa de lotação foram de 21,0; 20,3 e 24,3 t/ha/ano; 7,0; 6,5 e 7,7 unidades animais/ha/dia, respectivamente. Para proteína bruta, fibra em detergente neutro e digestibilidade in situ da matéria seca os valores médios obtidos foram de 18,1; 16,7 e 17,6%; 57,8; 58,9 e 58,7%; 79,6; 78,9 e 80,6%, para os respectivos sistemas. Em termos de produtividade melhores resultados foram obtidos no consórcio de ervilhaca comum com CC recebendo 100 kg de N/ha/ano e na pastagem de CC recebendo 200 kg de N/ha/ano. Melhores resultados de valor nutritivo foram obtidos no inverno, em especial para o consórcio de Coastcross-1 com ervilhaca.

The objective of this research was to evaluate the forage yield and nutritive value of elephantgrass on conventional and organic systems. In the organic system, species with complementary growth periods were combined; elephantgrass was planted in lines 3m apart from each other; between the rows of elephantgrass, during the winter period, ryegrass was sown and in the summer period the development of species of spontaneous growth was allowed. In the conventional production, two pastures were studied, one with the same strategy of organic production, but with chemical fertilization and another with elephantgrass cultivated alone. 100 kg of N ha-1 were applied with mineral and organic fertilization (bovine manure and pig slurry) in conventional and organic systems, respectively. Holstein cows receiving 0.9% of body weight complementary concentrate feed were used in the evaluation. The experimental design was completely randomized, with three treatments (forage systems), three repetitions (paddocks) and with repeated measures in time (seasons). Forage mass parameters of pre and post grazing, botanical composition, forage production and nutritive value were evaluated. Forage samples were collected by hand-plucking technique for the determination of organic matter, crude protein and in situ digestibility of organic matter. During the experimental period (337 days) seven grazing cycles were performed. Forage production and stocking rate were 12548; 10270; 19168 kg ha-1 and 3.3; 2.1; 4.5 AU ha-1 dia-1, respectively, for the forage systems. Pastures mixed, on conventional and organic production, present best forage distribution throughout the seasons. Higher value of forage production was observed in elephantgrass cultivated alone, on conventional system. Better value of nutritional value parameters were observed in organic pasture. / O objetivo desta pesquisa foi avaliar a produção de forragem e o valor nutritivo do capim-elefante nos sistemas convencional e orgânico. No sistema orgânico, combinaram-se espécies com períodos de crescimento complementares; o capim-elefante foi plantado em linhas com 3m de distância uma da outra; entre as fileiras de capim-elefante, durante o período de inverno, foi semeado azevém e no período de verão permitiu-se o desenvolvimento de espécies de crescimento espontâneo. Na produção convencional, foram estudadas duas pastagens, uma com a mesma estratégia de produção orgânica, porém com adubação química e outra de capim-elefante sob cultivo singular. Foram aplicados 100 kg de N ha-1 com adubação mineral e orgânica (esterco de bovinos e dejetos de suínos) nos sistemas convencional e orgânico, respectivamente. Como animais experimentais foram utilizados vacas da raça Holandesa, recebendo alimentação complementar na forma de concentrado à razão de 0,9% do peso corporal. O delineamento experimental foi o inteiramente casualizado, com três tratamentos (sistemas forrageiros), três repetições (piquetes) e medidas repetidas no tempo (estações). Foram avaliados os parâmetros de massa de forragem pré e pós-pastejo, composição botânica, produção de forragem e valor nutritivo. Para a determinação de matéria orgânica, proteína bruta e digestibilidade in situ da matéria orgânica, foram coletadas manualmente amostras de forragem mediante técnica de simulação de pastejo. Durante o período experimental (337 dias) foram realizados sete ciclos de pastejo. A produção de forragem e a taxa de lotação foram de 12548; 10270; 19168 kg ha-1 e 3,3; 2,1; 4,5 UA ha-1 dia-1, para os sistemas forrageiros. Pastos com misturas forrageiras, sob produção convencional e orgânica, apresentam a melhor distribuição de forragem ao longo das estações. O maior valor de produção de forragem foi observado em pastagens cultivadas singularmente, no sistema convencional. Melhores parâmetros de valor nutricional foram observados em pastagens orgânicas.