Electron tomography of negatively stained complex viruses: application in their diagnosis

BACKGROUND: Electron tomographic analysis can be combined with the simple and rapid negative staining technique used in electron microscopy based virus diagnosis. METHODS: Standard negative staining of representative examples of parapoxviruses and paramyxoviruses was combined with electron tomographic analysis. RESULTS: Digital sectioning of reconstructions of these viruses at a selected height demonstrated the viral ultrastructure in detail, including the characteristic diagnostic features like the surface threads on C-particles of a parapoxvirus and individual glycoproteins and the internal nucleoprotein strand of Newcastle disease virus. For both viruses, deformation and flattening were observed. CONCLUSION: The combination of negative staining of complex viruses with electron tomographic analysis, allows visualizing and measuring artifacts typical for negative staining. This approach allows sharp visualisation of structures in a subnanometer-thick plane, avoiding blurring due to superposition which is inherent to TEM. In selected examples, such analyses can improve diagnosis of viral agents.