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Updated: Oct 6, 2025
Antigen-Capture Enzyme-Linked Immunosorbent Assay for Specific Detection of Mycoplasma pneumoniae
Published on: February 24, 2023
Mycoplasma synoviae dihydrolipoamide dehydrogenase is an immunogenic fibronectin/plasminogen binding protein and a
Jingjing Qi1, Yu Wang2, Haoran Li3
1Shanghai Veterinary Research Institute, the Chinese Academy of Agricultural Sciences (CAAS), 518 Ziyue Road, Shanghai, 200241, PR China.
Abstract:
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.
Insights
Mycoplasma synoviae dihydrolipoamide dehydrogenase (PdhD) is a surface protein involved in host cell adhesion and binds fibronectin and plasminogen. Antibodies against PdhD inhibit M. synoviae attachment and exhibit mycoplasmacidal activity.
Area of Science:
- Avian Pathogen Research
- Molecular Biology of Microbial Pathogenesis
- Enzyme Function in Bacterial Adhesion
Background:
- Mycoplasma synoviae (M. synoviae) causes significant economic losses in the poultry industry due to arthritis and airsacculitis.
- Cytoadherence is a critical step in M. synoviae infection, yet the mechanisms are not fully understood.
- Dihydrolipoamide dehydrogenase (PdhD), essential for energy metabolism, has an uncharacterized role in M. synoviae cytoadherence.
Purpose of the Study:
- To investigate the role of M. synoviae dihydrolipoamide dehydrogenase (MSPdhD) in bacterial cytoadherence and pathogenesis.
- To characterize MSPdhD as a potential target for therapeutic intervention against M. synoviae infections.
Main Methods:
- Recombinant MSPdhD (rMSPdhD) expression and antibody production in rabbits.
- Immunoblot, mycoplasmacidal assays, suspension immunofluorescence, and subcellular localization analyses.
- Enzymatic activity assays, indirect immunofluorescence, and binding assays (Western blot, ELISA) with host cell proteins fibronectin and plasminogen.
Main Results:
- MSPdhD is an immunogenic, surface-localized protein with enzymatic activity.
- rMSPdhD adheres to chicken embryo fibroblast cells (DF-1), and M. synoviae attachment is inhibited by anti-rMSPdhD serum.
- MSPdhD binds to fibronectin and plasminogen in a dose-dependent manner.
- Rabbit anti-rMSPdhD serum demonstrated high complement-dependent mycoplasmacidal activity (88.5%).
Conclusions:
- MSPdhD is a multifunctional protein involved in M. synoviae cytoadherence, host protein binding, and potentially pathogenesis.
- Anti-MSPdhD antibodies show promise in inhibiting M. synoviae adhesion and possess significant bactericidal activity.
- MSPdhD represents a potential novel target for controlling M. synoviae infections in poultry.
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