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My dissertation hypothesis is that bovine trophoblast cells express cell-surface and secreted non-classical major histocompatibility complex class I (MHC-Ib) proteins which inhibit NK cells and other leukocytes by binding to inhibitory receptors (e.g., LILRB1, LILRB2, KIR2DL4, and/or CD94/NKG2A). Extremely polymorphic and ubiquitously expressed classical MHC class I (MHC-Ia) proteins, which present foreign antigenic peptides to CD8+ T lymphocytes, are involved in acceptance or rejection of tissue grafts. Non-classical MHC class I (MHC-Ib) glycoproteins, such as Human Leukocyte Antigen-G (HLA-G) and murine Qa-2, are important modulators of the maternal immune system during pregnancy. MHC-Ib proteins are: (a) oligomorphic or monomorphic, (b) expressed in specific tissues under specific condtions, and (c) produced as surface and/or soluble isoforms due to alternative splicing. Third trimester-bovine trophoblast cells express both MHC-Ia and MHC-Ib proteins. The MHC-Ib proteins expressed by trophoblast cells during the third trimester of pregnancy are encoded by four bovine leukocyte antigen (BoLA) loci: BoLA-NC1, BoLA-NC2, BoLA-NC3, and BoLA-NC4. Two MHC-Ia (N*01701 and N*01802) and three MHC-Ib (NC1*00501, NC3*00101 and NC4*00201) proteins showed cell-surface expression in transfection studies performed in murine P815 and human K562 cells. Two additional isoforms, NC1*00401 and NC2*00102, were not detected on the surface of these cells. Nevertheless, both class Ia proteins, N*01701 and N*01802, and five class Ib proteins, NC1*00401, NC1*00501, NC2*00102, NC3*00101, and NC4*00201, were detected in crude cell lysates on Western blots. Precipitation of proteins from culture supernatants showed that cell-surface MHC-Ia (N*01701 and N*01802) and MHC-Ib proteins (NC1*00501, NC3*00101, and NC4*00201) are shed from the surface of these cells into the media. The mechanism of shedding of these proteins is, however, not known. Monoclonal antibodies W6/32, IL-A88, H1A, H6A, H11A, H58A, and PT-85A recognized surface MHC-I isoforms with varying affinity. We were able to develop a sandwich enzyme-linked immunosorbent assay (ELISA) using either H1A or IL-A88 antibody as the capture antibody and the W6/32 antibody for detection. We produced monoclonal antibodies against cattle NC1*00501 and NC3*00101 proteins. One monoclonal antibody generated against BoLA-NC3*00101 was highly specific. Unfortunately, due to failure to clone the NC3*00101- hybridoma, we no longer have an infinite source of this monoclonal antibody for NC3*00101. We eluted peptides from NC3*00101-transfected MHC-null K562 cells and identified peptides using liquid chromatography-mass spectrum (LC-MS) analysis. Analysis of peptide binding data using the SAS Proc mixed statistical program, suggested that the peptide EVTNQLVVL is a potential peptide ligand, which can be used to make tetramers for enumeration of antigen-specific leukocytes.

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Bovine colostrum collected within 12h of parturition was de-fatted, de-caseinated, and ultrafiltered (UF) using a 5 kDa cut-off membrane; the resulting UF permeate was freeze dried to create a powder with possible use as a functional food ingredient. Samples representative of five lots of this powdered “colostrum low molecular weight fraction” (CLMWF) were analyzed for chemical composition and nutrient profile. The average contents of fat, moisture, and ash were 0.6%, 1.7%, and 8.3% w/w, respectively. Carbohydrate analysis showed an average of 58.2% w/w lactose monohydrate with no monosaccharides, other disaccharides, trioses, or tetroses detected. The total nitrogen content averaged 1.13% w/w, with 74% of this in the non-protein nitrogen fraction, producing a true protein content of 1.9% w/w. A significant mass fraction of the material (~29% w/w) remains to be characterized. The CLMWF powders were found to contain significant quantities of the minerals calcium (average 870 mg/100g), magnesium, (311 mg/100g), phosphorus (1473 mg/100g), potassium (1705 mg/100g) and sodium (690 mg/100g), the nutrients taurine (average 26.5 mg/100g), L-carnitine (40.5 mg/100g), thiamine (648 mcg/100g) and riboflavin (6991 mcg/100g), and the nucleos(t)ides uridine (55.2 mg/100g) and 5’UMP (18.8 mg/100g), cytidine (3.33 mg/100g) and 5’CMP (4.83 mg/100g) and guanosine (3.45 mg/100g) and 5’GMP (3.57 mg/100g).

Doctor of Philosophy / Department of Diagnostic Medicine and Pathobiology / Brad J. White and Robert L. Larson / Bovine respiratory disease (BRD) is the most costly disease of cattle in US feedyards and diagnosis based on clinical signs of illness is challenging. Over the course of five independent studies we evaluated the precision of multiple observers assigning clinical illness scores (CIS) to calves with induced Mycoplasma bovis pneumonia. We also evaluated the accuracy of CIS in relation to lung lesions at necropsy. Agreement among observers over all five studies was slight ({kappa]= 0.16; 95% confidence interval, 0.10 to 0.24) and ranged from 0.10 to 0.21 for individual trials. The accuracy of CIS varied based on the pulmonary consolidation score chosen to represent a truly ill animal. Inflammation associated with BRD can lead to significant pulmonary damage and reduced lung function. Treatment for BRD frequently involves antimicrobial administration and occasionally non-steroidal anti-inflammatory drugs. We evaluated how calves experimentally challenged with Mannheimia haemolytica respond to treatment with flunixin meglumine, alone or in combination with the antimicrobial florfenicol. Individual calf response to bacterial pneumonia was highly variable in this study. None of the changes in serum biomarkers, CBC or chemistry parameters provided reliable indicators of the pulmonary inflammation associated with the mild severity of bronchopneumonia in our study. Metaphylaxis is frequently administered to manage the risk of BRD within cohorts of cattle. We evaluated the impact of metaphylactic antimicrobial administration 10 days prior to experimental Mannheimia haemolytica inoculation to mitigate pulmonary lesions. We found that calves receiving tildipirosin had less lung damage and fewer clinical signs of illness compared to calves treated with tulathromycin or saline. Finally, the ability to predict those animals that would not finish the production cycle normally would provide benefits in effectively managing cattle. We evaluated the ability of classification algorithms to accurately predict an individual calf’s outcome based on data available at first identification of and treatment for BRD. We found accuracy of classifiers was dependent on the data recorded by the feedyard and there are sub-groups of calves within feedyard populations where classifiers were highly accurate. These data suggest the importance of pairing the proper classifier with the data available.

Nigeria recently became the leading economy in Sub-Saharan Africa with a total GDP of 522.64 billion of US dollars (Tradingeconomics.com). As GDP increases, population rises and food demand intensifies. Within this context it is of critical importance to achieve food security. However, Nigeria heavily relies in exportations in order to meet the growing food demand, especially of meat products, a situation which is not desirable. The livestock industry, although one of the largest in Sub-Saharan Africa, still constrained by several endemic livestock diseases which result in annual economic loses for value of 140 million of US dollars (Fadiga et al., 2013). Within this group, bovine and porcine trypasosomiasis alone has been estimated to be responsible for 50 million of US dollars in economic loses in Nigeria (Fadiga et al., 2013). However, the real epidemiological situation, and hence the possibility of developing a rational control programme, remains largely unknown across the country due to the absence of large epidemiological studies. Majority of the trypanosomiasis research studies in Nigeria employ the Haematocrit technique or the Buffy coat technique and Giemsa stain as a diagnostic method. These techniques possess a high specificity but a much lower sensitivity than the molecular method employed in this research study. In fact, better epidemiological studies employing molecular techniques have been conducted in recent times such (Majekodumni et al., 2013a; Takeet et al., 2013) and results displayed much higher trypanosomiasis prevalence than previously detected by microscopy. In many sub-Saharan countries the majority of national livestock herds are owned by mobile communities; however, the trypanosome status of cattle owned by mobile pastoralist communities have been less thoroughly studied when compared to those of sedentary livestock keepers. In this doctoral work, the epidemiology of trypanosomiasis was studied, in transhumant herds located in two different Nigerian enclaves: the Kachia grazing reserve and the Jos Plateau, both located in North-central Nigeria. Within Kachia, the ecology appears to determine the presence of infection with a spatially differentiated distribution of the detected trypanosome species being observed across the reserve that appears not to be related to the migration of livestock. While upon the Jos Plateau, the current reduction in trypanosome prevalence suggests an abrupt change in the trypanosome infection rates in this part of the country. The hypothesis established in this doctoral work is that these epidemiologically different scenarios are the result of land pressures that have ultimately resulted in the habitat destruction of the vector. Longitudinal data was also collected in order to assess the effectivity of different formulations of synthetic pyrethroids for the combined control of trypanosomiasis and tick-borne diseases. Insecticide treated cattle represents at the moment the best long-term and cost-effective method for the control of the vector responsible for the transmission of trypanosomiasis, the tsetse fly. Since no data exist about the efficacy of the insecticide or the compliance of the pastoralist population with its application under migratory conditions, its performance was assessed in this doctoral work. In addition, animal health outcomes were monitored to stablish the possible relationship between clinical symptoms and disease outcome and socio-economic data relevant for the dynamics of disease such as migration trends, husbandry practices, awareness and administered treatment has been also analysed. The compiled information of this data will establish the risk associated with contracting the disease and provide further indications for the control of African bovine trypanosomiasis in the specific context of transhumant pastoral systems of sub-humid sub-Saharan African.