Thesis (M.S.)--University of Wyoming, 2008. / Title from PDF title page (viewed on August 4, 2009). Includes bibliographical references.

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The diagnostic performance of four serological assays for bovine brucellosis in African buffaloes, namely Rose-Bengal test (RBT), complement fixation test (CFT), indirect enzyme-linked immunosorbant assay (iELISA) and fluorescence polarisation assay (FPA) were evaluated and compared in a case-control study. The study followed the OIE assay validation pathway for validation of diagnostic tests applicable to wildlife species where there is a validated test available in a taxonomically closely related species. Two uninfected and four infected herds were recruited and an uninfected composite reference panel of 107 sera and infected composite reference panel of 93 were selected using composite reference standards. Diagnostic sensitivity (DSe) and diagnostic specificity (DSp) were calculated for individual tests and for different combinations of two tests in series and in parallel. Cut-off points were adjusted using receiver operating characteristics (ROC) analysis. Using these cut-off values, the index tests performed as follows: RBT DSe of 98.9% (95% CI 96.83% - 100%) and DSp of 98.1% (95% CI 95.6% - 100%), iELISA (cut-off >40.5%) DSe 98.9% (95% CI 94.2% - 100%) and DSp 100% (95% CI 96.6% - 100%), CFT (cut-off >0 iU/ml) DSe 74.2% (95% CI 64.1% - 82.7%) and DSp 100% (95% CI 96.6% - 100%) and FPA (cut-off >16 mP) DSe 97.9% (95% CI 94.2% - 99.7%) and DSp 100% (95% CI 96.6% - 100%). Based on performance index and area under the ROC curve, the iELISA performed best (198.9% and 1.0), followed closely by the FPA (197.9% and 0.989) and the RBT (197.0%). The CFT s lower performance (174.2%, and 0.871) was due to low DSe. Kappa values for test agreement between the index tests was above 0 for all combinations, and varied from unweighted Kappa of 0.685 (95% CI 0.608 0.762) between FPA and iELISA to 0.26 (0.136-0.383b) between CFT and RBT. Consideration of the indices for positive and negative test agreement between the index tests supported the differential specificity of tests for different immunoglobulin classes and higher in line with the findings in cattle. Positive predictive value in herd C and E were 100% for the iELISA, CFT and FPA, 97.3% in herd C and 98.4% in herd E for the RBT. Negative predictive values in herd C ranged from 89% for the CFT to 99.2% for the RBT and in herd E 73.1% for the CFT to 98.7% for the RBT. Overall repeatability was satisfactory, except for the FPA, which was considered the result of sample quality related to prolonged storage in a freezer. The index tests were all found fit for use to detect or confirm brucellosis in populations and individual animals. The values for DSe and DSp that were estimated will be of use in the interpretation of serological results and determination of diagnostic strategies in different circumstances. Different combinations of tests in series and parallel increased the DSp and DSe. Using the RBT in combination with the CFT/FPA/iELISA interpreted in series or in parallel in relation to the epidemiological setting and objective of testing is recommended. / Mini-dissertation (MSc)--University of Pretoria, 2015. / tm2016 / Veterinary Tropical Diseases / MSc / Unrestricted

Brucella spp. are zoonotic infectious agents, primarily of livestock, that cause the disease brucellosis. Bovine brucellosis, caused by Brucella abortus, is of greatest concern due to the disease’s significant economical and public health impact. Canada fully eradicated bovine brucellosis from domesticated cattle herds in 1985, however, continued surveillance through screening for B. abortus exposure is paramount to the maintenance of bovine brucellosis eradication nationwide. The Canadian Food Inspection Agency (CFIA) is responsible for the surveillance of bovine brucellosis outbreaks in Canada and the maintenance of eradication. Current B. abortus serodiagnostics and serological screening is mostly based on the detection of antibodies against Brucella lipopolysaccharide (LPS), a highly immunogenic component of the outer cellular membrane. Such tests face difficulties with false positive results due to cross reactivity with other Gram-negative bacteria that produce LPS. The purpose of the research presented here was to address this issue through identifying new B. abortus protein antigens for the improvement of serological test specificity. In this study, 101 candidates were identified through predictive bioinformatic analyses and selected for immunogenic evaluation. While none of the expressed candidates displayed positive serological activity with in-house brucellosis positive bovine serum panels, the workflow presented here can be used for continued research and the assessment of more proteins from B. abortus and other bacterial pathogens.

This investigation was performed in the Altiplano Altoandino sub-humid region in the Ulla Ulla district, Pelechuco township, Franz Tomayo province, department of La Paz, Bolivia. The objective of this study was to quantify and compare the presence of Brucellosis in male and female alpacas (Lama pacos) at ages 2, 3, 4, 5, 6, and 7 years. Blood samples were taken from the jugular vein of 500 alpacas from the Hichucollo, Huacuchani, Ucha Ucha, and Ulla Ulla communities. The serum from each sample was separated and a serological diagnosis was performed by rapid agglutination with Rose Bengal chemical stain. In the Ulla Ulla district, 11.0% of alpacas were suspected of being infected. Of that 11.0%, 9.6% represented female alpacas while the remaining 1.4% represented male alpacas. By community, the overall results were 0.6%, 0.2%, 7.4%, and 2.8% for Hichucollo, Huacuchani, Ucha Ucha, and Ulla Ulla respectively. With regards to gender in each community, 1.4%, 1.6%, 13.3%, and 11.1% of female alpacas and 2.7%, 0.0%, 1.2%, and 1.9 % of male alpacas were suspected of being infected in Hichucollo, Huacuchani, Ucha Ucha, and Ulla Ulla, respectively. According to age, 0.8%, 2.6%, 3.8%, 2.0%, 11.8%, and 0.0% were suspected of Brucellosis in alpacas 2, 3, 4, 5, 6, and 7 years old, respectively in the Ulla Ulla district. The analysis of variance among the results does not present significant differences (p ≥ 0.05), failing to reject the given hypothesis. The final prevalence point found in the Ulla Ulla district was 11.0%. It was concluded that the prevalence of suspected cases of Brucellosis in alpacas is a consequence of the antigenic characteristics of each biotype. Furthermore, the results affirm that the poor management of livestock and livestock health lead to significant problems with alpaca health. It is therefore necessary to implement control strategies as well as disease monitoring in communities by following a detailed program of diagnostic tests for Brucellosis. Also, non-infected flocks must be protected against infection.

Vaccine Brucella abortus strain RB51, unlike the wild strain 2308 and another vaccine strain (strain 19) does not induce anti-O-chain antibodies. An efficacious vaccine strain that fails to produce an O-chain and thus a lack of an anti-O-chain humoral response greatly simplifies identification of vaccinated versus field strain infected animals. The three primary objectives of this research were the following: 1) to develop a serological assay to detect anti-RB51 antibodies in vaccinated elk (Cervus elaphus nelsoni), 2) to identify potential antigenic alterations in RB51 after vaccination of elk and BALB/c mice, and 3) to confirm the general stability of RB51. Elk were divided into four groups based upon gender and the route of inoculation (subcutaneous or ballistic) of RB51 bacteria. This study developed a highly reliable ELISA (using a monoclonal anti-bovine IgG 1 antibody and acetone killed whole RB51 bacteria) which can identify RB51-vaccinated elk. Also, isolates recovered from RB51-vaccinated elk were inoculated into female BALB/c mice whose spleens were then cultured. All elk and mice isolates were bacteriologically, biochemically, and serologically evaluated. This study showed that RB51 is a highly stable strain, which does not revert to smooth morphology or initiate synthesis of LPS-O-chain, maintains it biochemical characteristics, does not undergo detectable antigenic variations, and remains attenuated even after successive passages in elk and mice. Overall, this research indicates that RB51 is a vaccine candidate for the prevention of brucellosis in elk. Further studies are needed to determine the protective capabilities of RB51 in elk. / Master of Science

Disease management along the boundaries of wildlife reserves is a growing conservation problem worldwide, as infected wildlife can migrate outside protected areas and pose a threat to livestock and human health. The bison Bison bison population in Yellowstone National Park has long been infected with Brucella abortus, the bacterium causing bovine brucellosis. Concern over migratory bison transmitting B. abortus to cattle herds on lands adjacent to Yellowstone has led to proposals for bison vaccination. Model simulations suggest that vaccination is unlikely to eradicate B. abortus from Yellowstone bison but could be an effective tool for reducing the level of infection and eliminating unpopular management practices such as lethal culling. The culling of Yellowstone bison to reduce the risk of brucellosis transmission to cattle is negatively affecting long-term bison conservation because of difficulties in diagnosing actively infected animals. Age-specific serology and B. abortus culture assays from slaughtered bison were used to develop a diagnostic tool to estimate whether particular animals are infective. Findings suggest that active B. abortus infection is age-dependent, which allows true infection probabilities to be estimated based on age and quantitative diagnostic tests. Active brucellosis infection was associated with below-average nutritional condition, with the intensity of B. abortus infection being influenced by seasonal reductions in dietary protein and energy. The reproductive strategy of Yellowstone bison is linked with the seasonal availability of food, which increases bison fitness but may have consequences for B. abortus infection. Seasonal food restriction may also influence the ability of vaccinated bison to recall protective immune responses when later exposed to B. abortus. The rate of fat metabolism was an important factor influencing cell-mediated responses. Thus, individual variation and the seasonal availability of food may reduce vaccine efficacy when vaccination is applied at the population level. Consequently, effective management practices will require a diverse range of integrated methods, which include maintaining separation of livestock and wildlife, managing habitat to reduce brucellosis transmission, and reducing disease prevalence in wildlife. The long-term success of these management practices will depend on sound science and support of the stakeholders involved.

The global One Health movement has become firmly entrenched in both political and scientific discourse pertaining to emerging infectious diseases in the past decade. Since the discovery of the H5N1 strain of Highly Pathogenic Avian Influenza in Hong Kong in 1997, the promotion of more holistic programmes for the control of emerging infectious disease has garnered “unprecedented support” in terms of donor funding and political mobilisation (Scoones 2010). Advocates of One Health argue that intersectoral approaches promoting better communication between the veterinary, medical and environmental disciplines at all levels of governance make not only sound economic sense, they are fundamental to the “new approach” required to address the growing disease threats of the 21st century. However, despite international endorsement of the One Health rhetoric, there is growing pressure to now “turn the rhetoric into reality” (Okello et al 2011). Using a multiple, embedded case study methodology, this thesis seeks to examine questions surrounding the practical implementation of One Health interventions, particularly in developing countries which experience limited resources and competing health priorities. Through examining the livestock and public health policy processes at both local and national levels in Uganda and Nigeria, I attempt to identify whether policy spaces exist for the formal inclusion of One Health approaches in future policy decisions. Furthermore, by scrutinising the current internationally dominant One Health narratives in light of global health governance perspectives and the emerging One Health Global Network, I question whether One Health can be better “packaged” to include endemic diseases and a more focussed sustainable livelihoods approach; arguably inciting greater motivation for developing countries to truly participate. Data from my three empirical chapters are presented in the context of three overriding “One Health propositions” for consideration; by questioning “whose world, whose health”, I aim to delve further into the issues of not whether, but how this “new health paradigm” can be operationalised, and how to address the potential gaps which may ultimately prevent One Health from becoming a truly global phenomenon.

A brucelose bovina é uma zoonose que apresenta importância econômica por ocasionar perdas reprodutivas nas espécies animais acometidas. Em 2001, no Brasil, foi criado o Programa Nacional de Controle e Erradicação da Brucelose e da Tuberculose Animal. Uma das principais medidas instituídas pelo programa é a vacinação de fêmeas entre três e oito meses de idade pela cepa B19. Posteriormente, em 2007, ficaram definidas as normas para a utilização da cepa RB51 em fêmeas maiores que oito meses de idade, e em propriedades que apresentem foco da doença. Sabe-se que esta medida, se utilizada, abreviaria o período de tempo necessário para que a redução da prevalência da brucelose bovina fosse atingida. A fim de estimar a magnitude desta redução, bem como aferir o acumulado de animais a serem protegidos para que a diminuição na prevalência ocorra, foi realizado um modelo matemático associando a proteção de 80% das bezerras por B19, à proteção de 10, 40 e 80% das demais fêmeas por RB51. Para as simulações, foram consideradas as Unidades Federativas que apresentam prevalência da brucelose bovina acima de 2%. Comparando os cenários analisados, pôde-se concluir que a redução do período de tempo necessário para que se atingisse 2% de prevalência pouco diferiu quanto à cobertura vacinal de 40 ou 80% das fêmeas vacináveis. Isto faz com que a primeira alternativa possa, economicamente, ser tida como mais interessante, uma vez que abrangeria um acumulado menor de animais protegidos. Considerando o Estado de Goiás, que apresenta prevalência de 3,01% e um efetivo de 8.336.632 fêmeas bovinas, a proteção das fêmeas jovens por B19 juntamente à proteção de 40% das demais por RB51, faria com que a redução da prevalência alcançasse 2% em quatro anos, com um acumulado de 1.302.130 animais protegidos. Em relação ao Estado de Mato Grosso, que apresenta a maior prevalência, 10,25%, e 8.377.433 fêmeas com mais de 24 meses de idade, foi observado que a frequência da brucelose chegaria a 2% em quinze anos caso a proporção de fêmeas protegidas por RB51 fosse 40%, resultando em um acumulado de 1.656.098 animais. Em comparação à utilização da B19 como única medida preventiva, utilizando a RB51 em 40% das fêmeas vacináveis, se reduziria o período de tempo para se atingir 2% de prevalência de sete para quatro anos no Estado de Goiás, e de 22 para 15 anos no Mato Grosso. Além disso, a determinação dos acumulados a serem vacinados permite que se avalie economicamente a viabilidade ao uso da RB51 / Bovine brucellosis is a zoonosis that play important economic role for causing reproductive losses on animal species that are affected. In 2001, in Brazil, Brazilian National Program for the Control and Eradication of Bovine Brucellosis and Tuberculosis was created. One of the main action proposed by the program is vaccination of female between three to eight months of age by B19 strain. Later, in 2007, were defined the roles for RB51 strain use in females older than eight months, and in herds that are in outbreak situation. It is known that this measure, if used, would short the time needed for bovine brucellosis reduction. In order to estimate the magnitude of this reduction, as well as to measure the accumulated of protected animals for brucellosis prevalence decrease, a mathematical model was done associating protection by B19 in 80% of heifers, to protection by RB51 in 10, 40 and 80% of other females. For simulations, states that presented brucellosis prevalence superior than 2% were considered. Comparing these scenarios was possible to conclude that the time needed to reduce prevalence to 2% were similar between 40 and 80%. This turns the former alternative economically more interesting, once it will cover lower number of protected animals. Considering Goiás State, that presents prevalence of 3.01% and a amount of 8336,632 female bovine, protection of 80% of heifers by B19, along protection of 40% of other females by RB51, would make prevalence reach 2% in four years, with a total of 1302,130 protected animals. Relative to Mato Grosso State, that has the major prevalence, 10.25%, and 8377,433 female older than 24 months of age, were observed that brucellosis frequency would reduce to 2% in 15 years in case of 40% of female protected by RB51, resulting in an amount of 1656,098 animals. In comparison with the use of B19 as single preventive measure, time to reduce prevalence to 2% will reduce from seven to four years in Goiás, and from 22 to 15 years in Mato Grosso. Moreover, determination of accumulated of protected animals allows that RB51 use would be economically evaluated

Com o objetivo de se aperfeiçoar a PCR para detecção de Brucella spp. em leite bubalino, foram estudados diferentes protocolos de extração de DNA em leite bubalino experimentalmente contaminado com Brucella abortus amostra 1119-3. Esses protocolos basearam-se na utilização de isotiocianato de guanidina, fervura e proteinase K, com algumas variações. O critério de avaliação utilizado foi a sensibilidade analítica. O protocolo utilizando fervura com filtração inicial da amostra em colchão de sacarose a 40% apresentou sensibilidade analítica de 103 UFC/ml. Os protocolos utilizando proteinase K para extração de DNA apresentaram sensibilidade analítica de 104 UFC/ml. A maior sensibilidade analítica (10 UFC/ml) e a melhor visualização do produto amplificado foram obtidos com a utilização de isotiocianato de guanidina com precipitação imediata em álcool sem adição de Tween 20 na lavagem inicial da amostra. / To improve the PCR performance for detection of Brucella spp. in buffalo milk, different DNA extraction protocols were carried out in buffalo milk spiked with Brucella abortus strain 1119-3. These protocols were based on utilization of guanidine isothiocyanate, boil and proteinase K, with some variations. The analytical sensitivity was the evaluation criteria. Boiling with initial filtration of the sample through a solution of sacarosis 40% presented analytical sensitivity of 103 CFU/ml. The protocols based on proteinase K presented analytical sensitivity of 104 CFU/ml. The highest analytical sensitivity (10 CFU/ml) and best visualization of the amplified product were verified for the protocol using guanidine isothyocianate with immediate precipitation in alcohol without addition of Tween 20 in the initial sample wash.