Thesis (Ph. D.)--Virginia Polytechnic Institute and State University, 1990. / Vita. Abstract. Includes bibliographical references (leaves 111-123). Also available via the Internet.

Thesis (M.S.)--University of Wisconsin--Madison, 1962. / Typescript. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaves 45-54).

Thesis (M.S.)--University of Wisconsin--Madison, 1970. / eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references.

No description available.

Thesis (Ph. D.)--University of Wisconsin--Madison, 1949. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaves [i]-viii).

Brucellosis is endemic in many livestock worldwide especially developing countries. The aims of this study were to estimate the seroprevalence of bovine brucellosis and risk factors associated with the seropositivity in rural and peri-urban buffaloes and cattle populations of Sindh. Firstly, a cross sectional study was conducted to estimate the seroprevalence of bovine brucellosis in cattle and buffaloes of Sindh province, Pakistan. Serum samples (2600) were tested using Rose Bengal Plate Test. The overall seroprevalence of brucellosis in Sindh province was 13.96% (95% C.I.; 11.55 - 16.37). Of the 917 herds tested, 232 or 25.30% herds (95%C.I.; 22.51-28.24) were positive for brucellosis. The adult animals were 2.05 (95% C.I.; 1.14-3.68, P= 0.02) times more likely to test positive for brucellosis. The animals in a peri-urban dairy production system were 2.07 times (95%C.I.; 1.09-3.90, P = 0.03) times more likely have brucellosis. The species or sex of animal did not appear to affect the risk of seropositivity in cattle or buffalo in this population. Secondly, a cross sectional survey was conducted to understand the structure and composition of farms, animal husbandry and management practices in peri-urban dairy colonies in Karachi and farmers’ awareness of zoonoses. The mean herd size was 93.58 animals and 88.01% of these animals were female buffaloes. Of 326 farms surveyed, only 37.42% were able to associate animals with transmission of diseases in human. The characteristics of peri-urban dairy farms in Karachi are discussed. Thirdly, the value of FTA® cards in detecting the Brucella DNA in milk samples was estimated by determining the detection limits of genus specific ERI PCR assay for FTA® cards and comparing the PCR results from whole sediments taken from culturing pooled milk samples with taking sediment on FTA® cards. The detection limits of this method ranged from 6.6 x 103 cfu/ml for B. abortus to 7.17 x 106 cfu/ml for B. suis. Assuming the results of ERI PCR for the whole sediment as gold standard (method 1), the method using sediment on FTA® cards as test samples (method 2) showed a diagnostic sensitivity of 81.44% (95% C.I.; 75.54-87.33) but a poor diagnostic specificity of 42.86% (95%C.I.; 16.95-68.78). The kappa value, κ, was 0.14 (p = 0.02) demonstrating a poor agreement between the two methods. Lastly, 181 bulk milk samples were used to estimate the herd level prevalence of bovine brucellosis in Landhi dairy colony, Karachi. The ERI PCR was used to test these samples. The herd prevalence was estimated as 92.26% (95% C.I.; 88.34-96.19). For each level (50 animals) increase in herd size, the risk of herd being brucellosis positive increases by 2.38 times. The herds that have a male animal for breeding are 0.09 times less likely to have brucellosis. The history of abortion, presence of small ruminants or the regions of animal purchase don’t appear to have any association with the risk of brucellosis at a herd level in this population at LDC, Karachi. A high seroprevalence of bovine brucellosis in livestock population in Sindh and a very high herd level prevalence in peri-urban dairy farms in particular poses a serious threat to the public health and livestock production in Sindh, Pakistan.

Previous analysis of the bovine SLC11A1 complementary DNA (cDNA) failed to identify any nucleotide variations other than a microsatellite length variation within the 3' untranslated region functionally associated with bovine brucellosis. In this study I set out to identify mutations in the genomic complement of the gene that may be associated with resistance or susceptibility to bovine brucellosis, and to determine if the microsatellite length polymorphism in the 3'UTR of bovine SLC11A1 modulates gene expression and subsequent disease resistance in a phase dependent manner. The results of this study demonstrate that there are seventy-five total single nucleotide polymorphic (SNP) sites (excluding indels) located within the bovine genomic SLC11A1 sequence of a Brucella abortus resistant bull and a susceptible cow. Twenty of these SNPs segregated between resistant and susceptible populations, with 3 non-synonymous SNPs significantly associating with resistance or susceptibility to B. abortus infection. An A695G within exon 2 resulted in a histidine (resistant allele) to arginine (susceptible allele) amino acid substitution and was in significant linkage disequilibrium with the previously described 3' untranslated region (UTR) microsatellite length variation associated with brucellosis resistance. A transcriptional element search in the 3' UTR revealed a ETS-domain PU.1 site, an IFN-γ activation site (GAS), an Interferon Consensus Sequence Binding Protein site (ICSBP) and several Initiation Response sites (Inr), suggesting a possible function for this region in regulation of the expression of SLC11A1. A mobility shift assay confirmed sequence-specific DNA-protein interaction within this region. A luciferase reporter assay indicated that the 3'UTR of SLC11A1 could act as a downstream enhancer for expression. Macrophage killing assays with RAW264.7 cells expressing bovine SLC11A1 demonstrated that the microsatellite repeat is functionally associated with the macrophage killing efficiency, but not in a phase-dependent manner, suggesting that these length polymorphisms do not affect the angular orientation between cooperatively binding transcription factors, and leaves the possibility that the 3'UTR microsatellites regulate SLC11A1 transcription through some alternate mechanism, possibly mRNA stability.

No description available.

Bacillus anthracis is a facultative intracellular bacterial pathogen that can cause cutaneous, gastrointestinal or respiratory disease in many vertebrates, including humans. Commercially available anthrax vaccines for immunization of humans are of limited duration and do not protect against the respiratory form of the disease. Brucella abortus is a facultative intracellular bacterium that causes chronic infection in animals and humans. As with other intracellular pathogens, cell mediated immune responses (CMI) are crucial in affording protection against brucellosis. B. abortus strain RB51 has been shown to be useful in eliciting protective cell mediated immunity and humoral responses against Brucella in cattle and other animal species. Since the protective antigen (PA) of B. anthracis is known to induce protective antibodies, it was decided that the objective of this research was to test whether the gene encoding PA could be expressed in Brucella producing a bivalent vaccine to protect against both brucellosis and anthrax. The pag gene was transcriptionally fused to promoters of genes encoding superoxide dismutase or heat shock protein groE, subcloned into a broad host range plasmid (pBBR1MCS) and shown to express in E. coli by immunoblotting using antiserum specific for PA. The immunoblot results revealed that E. coli produced a PA protein of the expected size. In addition, the culture medium was shown to contain the same PA protein using immunoblotting. These results show that E. coli is capable of expressing B. anthracis PA in both the cellular and extracellular forms. The pBB/PA plasmid was used to transform B. abortus RB51 and CmR clones screened for the expression of PA by immunoblotting. Twenty clones of strain RB51/pBBSOD were show to express a 30kDa PA protein. Three clones of strain RB51/pBBGroE-PA were shown to express a 63-83kDa protein as detected by antiserum specific for PA. Using the A/J mouse, an immunocompromised vertebrate model, immunization and challenge studies were performed. Preliminary results demonstrate that the bivalent vaccine is capable of producing protection against a live challenge with B. abortus and some protection against live non-disease producing spores of B. anthracis. / Master of Science

B. abortus is an intracellular facultative bacterial pathogen which causes abortion in cattle and undulant fever in humans. Cattle vaccines such as B. abortus strains 19 and RB51 are live vaccine strains which protect approximately 75% of the vaccinated animals. No effective vaccines are available for the prevention of brucellosis in humans. We are developing vaccinia virus recombinants expressing various B. abortus proteins to prevent brucellosis in susceptible mammalian species. In this work the B. abortus groEL gene encoding the antigenic heat shock protein GroEL was subcloned into vaccinia virus via homologous recombination. Expression of the GroEL protein in vaccinia infected cells in-vivo was confirmed by immunoblotting. Groups of 5 female BALB/C mice were injected with the vaccinia recombinant or appropriate positive and negative control vaccines. Mice were bled and their humoral immune responses assessed. In addition, mice were challenged with virulent B. abortus strain 2308 and protection measured by the rate of splenic clearance of live Brucella. In spite of demonstrating specific GroEL antibodies in recombinant vaccinia injected mice, no significant level of protection was demonstrable. Preliminary lymphocyte transformation assays were carried out to establish if a cell mediated immune response to GroEL was induced in the vaccinated animals. / Master of Science