Viral detection by electron microscopy: past, present and future - PubMed (original) (raw)

Viral detection by electron microscopy: past, present and future

Philippe Roingeard. Biol Cell. 2008 Aug.

Abstract

Viruses are very small and most of them can be seen only by TEM (transmission electron microscopy). TEM has therefore made a major contribution to virology, including the discovery of many viruses, the diagnosis of various viral infections and fundamental investigations of virus-host cell interactions. However, TEM has gradually been replaced by more sensitive methods, such as the PCR. In research, new imaging techniques for fluorescence light microscopy have supplanted TEM, making it possible to study live cells and dynamic interactions between viruses and the cellular machinery. Nevertheless, TEM remains essential for certain aspects of virology. It is very useful for the initial identification of unknown viral agents in particular outbreaks, and is recommended by regulatory agencies for investigation of the viral safety of biological products and/or the cells used to produce them. In research, only TEM has a resolution sufficiently high for discrimination between aggregated viral proteins and structured viral particles. Recent examples of different viral assembly models illustrate the value of TEM for improving our understanding of virus-cell interactions.

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Figures

Figure 1

Figure 1

Diagnosis of virus infections by examination of ultrathin sections of human tissues or cells (A) Parapoxvirus (Orf virus) infection on a human skin biopsy specimen. Multiple oval viral particles (arrow), comprising a dense core surrounded by an envelope (inset, high magnification), are observed in an infected cell. The Orf virus is a parapoxivirus that causes a common skin disease of sheep and goats, and is occasionally transmitted to human. (B) Polyomavirus (BK virus) infection in cells pelleted from a urine sample taken from an organ‐transplant patient. The presence of a large number of viral particles leads to their arrangement into a crystal‐like structure (inset, high magnification).

Figure 2

Figure 2

Direct negative staining of virus in fluid recovered from human skin vesicles (A and B) or from stool samples (C and D) (A and B) Rapid morphological diagnosis and differential diagnosis from a herpesvirus (Varicella, A) and a parapoxvirus (Orf virus, B). The penetration of the negative stain into the herpesvirus particle may reveal the presence of the viral capsid within the envelope. (C and D) Negative staining of viruses involved in gastroenteritis reveals the surface detail of the subunit arrangement of the adenovirus core particle (C), showing its icosahedral form clearly, whereas the rotavirus displays its typical ‘wheel‐like’ appearance (D).

Figure 3

Figure 3

Detection of retroviruses in rodent hybridoma cells used for the production of biological products Ultrathin sections of cells of different origins may show intracisternal A‐type retroviral particles (A) or C‐type retroviral particles budding at the cell surface (B). The C‐type particles released by the cells can be detected by negative staining in the cell supernatant (B, inset).

Figure 4

Figure 4

Ultrastructural changes associated with viral replication or viral factories (A) The Semliki forest virus, an alphavirus, induces the formation of a cytopathic vacuole (CPV), surrounded by the ER. Numerous viral replication complexes (arrow) are anchored in the internal membrane of the CPV. (B) The non‐structural proteins of HCV, a flavivirus, induce the formation of a membranous web in the perinuclear area.

Figure 5

Figure 5

Budding of HIV The viral particle at the top shows virus formation with distortion of a cellular membrane away from the cytoplasm. The budding particle and the particle at the bottom are immature viral particles, whereas the two particles in the centre are mature and have a truncated cone‐shaped core. Thus maturation of the core by the viral protease occurs shortly after the release of the particle from the host cell membrane.

Figure 6

Figure 6

Budding of HCV Ultrastructural analysis of cells producing the HCV core protein shows that this protein self‐assembles into HCV‐like particles (arrows) at convoluted and electron‐dense ER membranes surrounding the lipid droplets (LD) present in the perinuclear area.

Figure 7

Figure 7

Hepatitis B subviral envelope particle morphogenesis and intracellular trafficking Ultrastructural analysis of cells producing HBV major envelope protein shows that this protein self‐assembles in the ER into filaments packed into crystal‐like structures (A, see also a high magnification of these packed filaments in the inset). These filaments are transported to the ERGIC, where they are unpacked and relaxed (B).

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