Browsing by Author "McFadyen, Ross"
Now showing 1 - 2 of 2
Results Per Page
Sort Options
- ItemImmunological tools for the characterization of the humoral immune response to Mycobacterium bovis infection in African buffaloes (Syncerus caffer)(Stellenbosch : Stellenbosch University, 2016-03) McFadyen, Ross; Miller, Michele; Parsons, Sven; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Biomedical Sciences: Molecular Biology and Human GeneticsENGLISH ABSTRACT : Bovine tuberculosis (bTB), caused by Mycobacterium bovis, is a chronic infectious disease known to occur in free-ranging mammals, captive wildlife, and livestock. M. bovis forms part of the Mycobacterium tuberculosis complex (MTC), a genetically related group of bacteria that cause tuberculosis in humans or other species. bTB eradication efforts are complicated by the occurrence of wildlife reservoirs of the disease such as the African buffalo (Syncerus caffer). Buffalo in the Kruger National Park and Hluhluwe-iMfolozi Game Reserve are affected by endemic bTB and there is the potential for M. bovis to spill-over into other domestic or wild animals, including zoonotic transmission to humans. Current standard diagnostic tests lack the required sensitivity to successfully detect all infected animals and are known to be ineffective in animals who have become anergic in the late stages of disease. Antibodies have been shown to correlate with disease pathology due to a high antigen burden and are able to be detected in anergic animals. The aim of this study was to characterize the humoral (antibody mediated) immune response in bTB-positive buffalo using different molecular techniques. Towards this aim, an in-house PPD ELISA was developed to detect antibodies against M. bovis and was compared to the Chembio® DPP VetTB, a commercial antibody assay. Both tests were able to detect M. bovis-specific antibodies in bTB-positive buffalo and there was good correlation between the two. We aimed to further assess the humoral response by investigating the diagnostic performance of monocyte chemotactic protein 1 (MCP-1), a cytokine involved in the activation of B-cell proliferation and class switching. There was a significant difference in antigen-specific MCP-1 release between bTB-positive and bTB-negative buffalos, suggesting there is promise for the use of MCP-1 as a biomarker, although sensitivity was low. Furthermore we evaluated the plasma stability of the MCP-1 protein during storage on protein saver cards (PSC) as well as after heat-treatment at 65°C. After 11 days at room temperature on the PSC, MCP-1 was still detected at similar levels. MCP-1 concentrations were higher after heat-treatment and correlated with the dilution factor. Lastly, we aimed to determine the stability of 8 reference genes to be used in a whole-blood qPCR gene expression assay in buffaloes. Selected gene transcripts of the African buffalo were sequenced using primers from the domestic cow and it was shown that PPIA is the most stable reference genes whereas GAPDH is the most unsuitable for use under our experimental conditions. In conclusion, the use of serological tests for the detection of M. bovis infected buffalo are disadvantaged by their low sensitivity, although using them in combination with conventional tests may provide useful information on the immune status of the animal. MCP-1 shows promise as a biomarker of disease in buffalo and its stability means that transporting of plasma samples can be made safe and efficient by heat-treating samples and storing them on PSC. Further research is needed to fully optimize a diagnostic GEA for bTB in buffaloes, however, we have shown that PPIA is a suitable reference gene for whole-blood stimulation in African buffaloes.
- ItemThe kinetics of the humoral and interferon-gamma immune responses to experimental Mycobacterium bovis infection in the white rhinoceros (Ceratotherium simum)(Frontiers, 2017-12) Parsons, Sven D. C.; Morar-Leather, Darshana; Buss, Peter E.; Hofmeyr, Jennifer; McFadyen, Ross; Rutten, Victor P. M. G.; Van Helden, Paul D.; Miller, Michele Ann; Michel, Anita LuiseENGLISH ABSTRACT: Mycobacterium bovis is the cause of tuberculosis (TB) in a wide range of species, including white rhinoceroses (Ceratotherium simum). Control of the disease relies on the indirect detection of infection by measuring pathogen-specific responses of the host. These are poorly described in the white rhinoceros and this study aimed to characterize the kinetics of immune responses to M. bovis infection in this species. Three white rhinoceroses were infected with M. bovis and their immune sensitization to this pathogen was measured monthly for 20 months. Cell-mediated immunity was characterized in whole blood samples as the differential release of interferon-gamma in response to bovine purified protein derivative (PPDb) and avian PPD (PPDa) as well as the release of this cytokine in response to the M. bovis proteins 6 kDa early secretory antigenic target (ESAT-6)/10 kDa culture filtrate protein (CFP-10). Humoral immunity was quantified as the occurrence or the magnitude of antibody responses to the proteins ESAT-6/CFP-10, MPB83, MPB83/MPB70, and PPDb. The magnitude and duration of immune reactivity varied between individuals; however, peak responses to these antigens were detected in all animals circa 5–9 months postinfection. Hereafter, they gradually declined to low or undetectable levels. This pattern was associated with limited TB-like pathology at postmortem examination and appeared to reflect the control of M. bovis infection following the development of the adaptive immune response. Measurement of these markers could prove useful for assessing the disease status or treatment of naturally infected animals. Moreover, immune responses identified in this study might be used to detect infection; however, further studies are required to confirm their diagnostic utility.