Determination of the specificity and nature of Mycoplasma Mycoides Subsp. Mycoides adherence to heir Host Cells

Abstract

Mycoplasma mycoides subsp. mycoides is the etiological agent of contagious bovine pleuropneumonia (CBPP). The disease is a significant constraint to cattle production throughout most of Sub-Saharan Africa. The contribution of cytoadherence and molecular factors involved in pathogenesis of M mycoides subsp. mycoides is poorly understood. This study aimed at providing data about cytoadherence of M mycoides subsp. mycoides on primary cultured bovine bronchus epithelial (BBE) cells and compared adherence rates to different cell lines using flow cytometry. In this assay, M mycoides subsp. mycoides was grown in "pleuropneumonia - like organism" (PPLO) medium and incubated with BBE cells at 37°C. The BBE cells were then stained with polyclonal antibodies to mycoplasma shown to adhere specifically to the BBE cells. The binding was dependent on both the concentration of the mycoplasmas and time and up to 70-99% of epithelial cells were positive. The specificity of mycoplasma adherence was examined using three different cell lines; bovine skin fibroblast (BY 122 ISF pSV neo), bovine testicular endothelial cells (282 BTVE pSV neo and baby hamster kidney (BHK) cells. Adherence rate of the three cell lines were significantly lower than that of bovine bronchus epithelial cells. Inhibition of adherence was observed upon pre-incubation of mycoplasma with a monoclonal antibody against capsular polysaccharide (PK-2). Monoclonal antibody against a surface protein L-a-glycerophosphate oxidase (GlpO) failed to inhibit significantly the adherence to BBE cells. Proteolysis of the BBE cells surface using proteinase K partially inhibited binding of the M mycoides subsp. mycoides to the BBE cells. Treatment of the BBE cells using glycosidase F to cleave the carbohydrate on the surface did not show significant inhibition. The significant reduction of adherence rate by PK-2 directed against an epitope on capsular polysaccharide located on the surface of M mycoides subsp. mycoides, suggests involvement of mycoplasma carbohydrates in adherence to the primary culture of BBE cells. The results also indicate that at least some proteins on the BBE cell surface are involved inM mycoides subsp. mycoides adherence and may be a receptor for a mycoplasma ligand.