Viable Mycoplasma mycoides ssp. mycoides small colony-mediated depression of the bovine cell responsiveness to the mitogen concanavalin A.

Mycoplasma mycoides ssp. mycoides biotype Small Colony (MmmSC) is the causative agent of contagious bovine pleuropneumonia (CBPP), which is still a major tropical cattle disease. Development of an efficient vaccine requires an understanding of the immunopathology of CBPP as MmmSC presents a strong ability to escape the host immune response. The objective of this study was to determine whether the presence of MmmSC can modulate the immune response induced by the mitogen Concanavalin A (ConA) on bovine immune cells [peripheral blood mononuclear cells (PBMC) and lymph node (LN) cells]. Comparative analysis of the immunomodulating properties of viable versus heat-killed MmmSC on ConA-stimulated immune cells revealed that while heat-killed MmmSC had no effect, viable MmmSC strongly depressed, in a concentration-dependent manner, the ConA mitogenic activity (blastogenesis and interferon-gamma production). Both B-cell and T-cell activation were affected with the highest impact on the CD4 T cells. The phenotypic analysis showed that the ConA-induced proliferation of CD25(+) cells was strongly reduced when co-exposed to viable MmmSC, confirming that events associated with ConA-induced cell activation were suppressed by the pathogen. This study thus demonstrated that viable MmmSC is able to inhibit the polyclonal mitogenic activity of the ConA on bovine PBMC and LN cells. This finding strongly suggests that the persistence of viable MmmSC may also thus inhibit the bovine immune response directed towards inactivated MmmSC, whether dead or in the form of antigens, also present during infection. This study confirmed that MmmSC has evolved an efficient mechanism to prevent its elimination from the host.

Mycoplasma mycoides ssp. mycoides biotype small colony-secreted components induce apoptotic cell death in bovine leucocytes.

Contagious bovine pleuropneumonia, caused by Mycoplasma mycoides ssp. mycoides biotype small colony (MmmSC), is one of the most serious cattle diseases in Africa. Several observations suggested that MmmSC had evolved an efficient way to escape the bovine immune responses by triggering host-cell cytotoxicity. This study was implemented to determine whether the cytotoxic effect was due to apoptotic cell death. To that end, bovine blood cells were cultured for up to 3 days in the presence of viable or heat-killed MmmSC compared to unstimulated cultures. The findings provided evidence for a viable MmmSC-induced, time-dependent apoptosis in bovine blood leucocytes, whereas heat-killed MmmSC had no effect. Morphological and physiological changes (evidenced by TUNEL and annexin V staining) typical of apoptosis were observed in response to viable MmmSC. All the lymphocyte subsets as well as the monocyte/granulocyte subset exhibited extensive apoptosis after exposure to viable MmmSC. Our results demonstrated a potential role for MmmSC-secreted components as pathogenic factors able to induce programmed cell death in bovine blood leucocytes.

Gamma interferon-producing CD4 T-cells correlate with resistance to Mycoplasma mycoides subsp. mycoides S.C. infection in cattle.

Contagious bovine pleuropneumonia (CBPP), caused by Mycoplasma mycoides subsp. mycoides SC (MmmSC), is one of the most significant cattle disease in Africa. The control measures, which led to eradication from numerous countries are not feasible in Africa where the only prophylaxis relies on vaccination. However, the attenuated vaccines, used up to now in Africa, are of low efficiency. The development of an improved vaccine is, therefore, a necessity. The purpose of this study was to compare some immunological parameters in MmmSC-infected cattle (endobronchial versus natural in-contact infection) and assess the response in correlation with the clinical outcome (death versus recovery). Characterization of the immune parameters elicited in recovered animals, known to be refractory to new infection, will be an important step towards development of new vaccines against CBPP. A significant outcome of this study was the demonstration that all MmmSC-infected cattle developed a MmmSC-specific cell-mediated immune response. A kinetic analysis of the MmmSC responsiveness showed that the main difference between endobronchially- and in-contact infected animals was the delay before the onset of the MmmSC-specific immune response. The first MmmSC-responding PBMC sample was selected from each animal for cell phenotyping. The phenotypic analysis of this early MmmSC-induced response revealed the predominant contribution of the CD4 T-cells in all animals whereas IFNgamma was only constantly produced in recovered animals. Evolution of this early MmmSC-specific immune response was then followed by a kinetic analysis of the MmmSC-induced CD4 T-cell response and IFNgamma released. The results demonstrated that in recovered animals, the MmmSC-specific CD4 Th1-like T-cell response was maintained until slaughtering whereas in animals with acute disease, progression of CBPP was associated with a decreased ability of the PBMC to produce IFNgamma. The results led to the identification of immune parameters, which correlate with protection against CBPP and to a relevant strategy for the development of improved vaccines against this disease.