Cellular and Cytokine Responses in the Granulomas of Asymptomatic Cattle Naturally Infected with Mycobacterium bovis in Ethiopia

Abstract

Cells (CD3+ T cell and CD68+ macrophages), cytokines (IFN-γ+ and TNF-α+) and effector molecule (iNOS+) responses were evaluated in the lymph nodes and tissue of cattle naturally infected with Mycobacterium bovis. Detailed post mortem and immunohistochemical examinations of lesions were performed on 16 cows positive for single intradermal cervical comparative tuberculin (SICCT) test which were identified from dairy farms located around the Addis Ababa City. The severity of the gross lesion was significantly higher (p=0.003) in M. bovis culture positive (n=12) cows than in culture negative (n=4). Immunohistochemical techniques showed that in culture positive cows, the mean immunolabeling fraction of CD3+ T cells decreased as the stage of granuloma increased from stage I to stage IV (p<0.001). In contrast, the immunolabelling fraction of CD68+ macrophages, IFN-γ+, TNF-α+ and iNOS+ increased from stage I to stage IV (p< 0.001). In culture negative cows, early stages showed a significantly higher fraction of CD68+ macrophages (p=0.03) and iNOS+ (p=0.007) when compared to culture positive cows. Similarly, at advanced granuloma stages, culture negative cows demonstrated significantly higher mean proportions of CD3+ T cells (p< 0.001) compared to culture positive cows. Thus, this study demonstrates that following natural infection of cows with M. bovis, as the stage of granuloma increases from stage I to stage IV, the immunolabelling fraction of CD3+ cells decreases while the immunolabeling fraction of CD68+ macrophages, IFN-γ+, TNF-α+ and iNOS+ increases.

Heterogeneity of Pulmonary Granulomas in Cattle Experimentally Infected With Mycobacterium bovis

“…The NHP model has also demonstrated a higher bacterial burden in lesions arising at earlier time points and a significant reduction in bacterial burden after the onset of adaptive immunity, consistent with the findings in this study (12). Previous studies of tuberculoid granulomas of cattle have suggested that bacterial burdens are increased in late stage IV granulomas, while others have found no correlation between bacterial numbers and granuloma stage (26,41). Those studies, however, estimated bacterial burden by enumerating AFB observed in microscopic tissue sections.…”

“…Prior studies in cattle have described the morphological heterogeneity of tuberculoid granulomas in both experimental and natural infections (26,37,41,42). The present study examined bacterial burdens of individual pulmonary tuberculoid granulomas at 30, 90, 180 and 270 days after experimental aerosol infection and revealed heterogeneity in bacterial burdens at all time points.…”

“…Previous cattle studies using IHC to measure IFN-γ protein within granulomas showed no difference in expression based on granuloma stage; however, there was increased IFN-γ expression in calves receiving a higher dose (1,000 CFU vs. 1 CFU) (42). A separate study using IHC showed an increase in IFN-γ protein immunolabeling between stages I and IV (41). Yet, another study, also using IHC, showed IFN-γ in granulomas of all stages, with a qualitative, but not significant decrease in labeling in late stage IV granulomas compared to earlier stages (47).…”

“…Other studies using granulomas from cattle collected at a single time-point after experimental infection, or from naturally infected cattle of unknown disease duration have shown decreasing trends, increasing trends, or no difference, respectively, between TNF-α expression in granulomas of stage I to IV. (24,41,47,49).…”

Epidemiology of Bovine Tuberculosis and Its Zoonotic Implication in Addis Ababa Milkshed, Central Ethiopia

“…The severity of the lesion was higher in the lymph nodes of the thoracic region. This observation is related to the route of infection, which is predominantly a respiratory route especially in dairy cattle kept in intensive dairy farms ( 28 ). Thus, the thoracic lymph nodes are affected predominantly as they are draining the lungs.…”