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Immunoelectron microscopic localization of variable proteins on the surface of Mycoplasma bovis
A Behrens1, M Heller, R Rosenbusch
1Institut für Mikrobiologie und Tierseuchen, Tierärztliche Hochschule Hannover, Germany.
Abstract:
The ultrastructural distribution and immunological accessibility of the variable surface proteins VspA, VspB, VspC and VspD were determined by immunoelectron microscopy on the surface of negatively stained cells of Mycoplasma bovis PG45 and 18 subclones, expressing either one or two of the Vsps. The variable proteins VspA, VspB, VspC and VspD, recognized by two monoclonal antibodies (mAb 1E5 and mAb 87-2) and visualized by goat-anti-murine-lgM labelled with gold particles, showed identical distribution patterns on the surfaces of the cells of all M. bovis clones investigated. Gold particles were distributed over the whole cell surface, arranged in clusters. The cell form seemed not to have an influence on the decoration pattern. Gold particles were also observed in irregular distributions around the cells. All clones showed unlabelled cells as well as strongly and weakly labelled cells. There were in general, however, no significant differences in the percentages of unlabelled, weakly labelled and strongly labelled cells, either between clones expressing different Vsps or between individual clones. No correlations were found between the numbers of labelled cells in immunoelectron microscopy and the numbers of labelled colonies in immunobinding assay (IBA) originating from the same broth cultures. The percentage of positive colonies in IBA was generally much higher than the percentage of positive cells in immunoelectron microscopy. The results show that the cells of the M. bovis clones are not identical, but differ in their surface antigens, and reveal the high variable potential of this species.
Insights
Mycoplasma bovis variable surface proteins (Vsps) are distributed in clusters across the cell surface. Despite this, individual M. bovis cells exhibit distinct surface antigen profiles, highlighting the species' high variability.
Area of Science:
- Microbiology
- Immunology
- Cell Biology
Background:
- Mycoplasma bovis is a significant veterinary pathogen.
- Variable surface proteins (Vsps) are crucial for M. bovis pathogenesis and immune evasion.
- Understanding Vsp distribution is key to developing effective control strategies.
Purpose of the Study:
- To investigate the ultrastructural distribution and immunological accessibility of VspA, VspB, VspC, and VspD on M. bovis cells.
- To compare Vsp expression and accessibility across different M. bovis subclones.
- To correlate immunoelectron microscopy findings with immunobinding assay results.
Main Methods:
- Immunoelectron microscopy using gold-labeled antibodies to visualize Vsps on negatively stained M. bovis cells.
- Analysis of 18 M. bovis subclones expressing one or two Vsps.
- Immunobinding assay (IBA) to quantify Vsp-positive colonies.
Main Results:
- Vsps (VspA-D) exhibited clustered distribution patterns across the entire cell surface, irrespective of cell morphology or Vsp type.
- Significant heterogeneity in Vsp labeling intensity (unlabelled, weakly, strongly labelled) was observed among cells within all clones.
- No correlation was found between cell surface labeling in immunoelectron microscopy and colony positivity in IBA, with IBA showing a much higher percentage of positive results.
Conclusions:
- M. bovis cells within clones are antigenically distinct, demonstrating high surface antigen variability.
- The distribution pattern of Vsps is consistent, but individual cell accessibility varies.
- Discrepancies between immunoelectron microscopy and IBA suggest different sensitivities or detection mechanisms for surface antigens.