Mycoplasma synoviae dihydrolipoamide dehydrogenase is an immunogenic fibronectin/plasminogen binding protein and a putative adhesin
Mycoplasma synoviae ( M. synoviae ) is an important avian pathogen that causes arthritis and airsacculitis in young chickens and turkeys. Infection by M. synoviae results in considerable economic losses to the poultry industry worldwide. Cytoadherence is a crucial stage during mycoplasma infection. Dihydrolipoamide dehydrogenase (PdhD) is a flavin-dependent enzyme that is critical for energy metabolism and redox balance. To date, its role in cytoadherence is poorly understood. In this study, recombinant PdhD from M. synoviae (rMSPdhD) was expressed in the supernatant component of E. coli BL21 and rabbit anti-rMSPdhD serum was prepared. rMSPdhD was shown to be an immunogenic protein by immunoblot assays, while the mycoplasmacidal assay revealed that the rabbit anti-rMSPdhD serum had a high complement-dependent mycoplasmacidal rate (88.5 %). Using a suspension immunofluorescence assay and subcellular localization analysis, MSPdhD was shown to be a surface-localized protein distributed in both the cytoplasm and cell membrane of M. synoviae . The enzymatic activity of rMSPdhD was determined by measuring its ability to reduce lipoamide to dihydrolipoamide and convert NADH to NAD + . Using an indirect immunofluorescence assay, rMSPdhD was shown to adhere to DF-1 chicken embryo fibroblast cells. Furthermore, the attachment of M. synoviae to DF-1 cells was significantly inhibited by rabbit anti-rMSPdhD serum. Western blot and ELISA binding assays confirmed that rMSPdhD also bound to fibronectin (Fn) and plasminogen (Plg) in a dose-dependent manner. In conclusion, our data show that MSPdhD is not only a biological enzyme, but also an immunogenic surface-exposed protein that can bind to Fn and Plg as well as adhere to host cells. In addition, we show that rabbit anti-rMSPdhD serum can inhibit the adhesion of M. synoviae to DF-1 cells and has a significant complement-dependent bactericidal activity. Our findings suggest that MSPdhD may be involved in the pathogenesis of M. synoviae .
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