Cultivation of Viruses
المؤلف:
Stefan Riedel, Jeffery A. Hobden, Steve Miller, Stephen A. Morse, Timothy A. Mietzner, Barbara Detrick, Thomas G. Mitchell, Judy A. Sakanari, Peter Hotez, Rojelio Mejia
المصدر:
Jawetz, Melnick, & Adelberg’s Medical Microbiology
الجزء والصفحة:
28e , p423-424
2025-10-21
83
Many viruses can be grown in cell cultures or in fertile eggs under strictly controlled conditions. Virus growth in animals is still used for the primary isolation of certain viruses and for studies of the pathogenesis of viral diseases and of viral oncogenesis. Diagnostic laboratories may attempt to recover viruses from clinical samples to establish disease causes. Research laboratories cultivate viruses as the basis for detailed analyses of viral replication and protein function.
Cells grown in vitro are central to the cultivation and characterization of viruses. There are three basic types of cell cultures. Primary cultures are made by dispersing cells (usually with trypsin) from freshly removed host tissues. In general, they are unable to grow for more than a few passages in culture. Diploid cell lines are secondary cultures that have undergone a change that allows their limited culture (up to 50 passages) but that retain their normal chromo some pattern. Continuous cell lines are cultures capable of more prolonged—perhaps indefinite—growth that have been derived from diploid cell lines or from malignant tissues. These have altered and irregular numbers of chromosomes. The type of cell culture used for viral cultivation depends on the sensitivity of the cells to a particular virus.
A. Detection of Virus-Infected Cells
Multiplication of a virus can be monitored in a variety of ways:
1. Development of cytopathic effects (ie, morphologic changes in the cells). Types of virus-induced cytopathic effects include cell lysis or necrosis, inclusion body formation, giant cell formation, and cytoplasmic vacuolization (Figure 1A, B, and C).
2. Appearance of a virus-encoded protein, such as the hem agglutinin of influenza virus. Specific antisera can be used to detect the synthesis of viral proteins in infected cells.
3. Detection of virus-specific nucleic acid. Molecular-based assays such as polymerase chain reaction provide rapid, sensitive, and specific methods of detection.
4. Adsorption of erythrocytes to infected cells, called hemadsorption, caused by the presence of virus-encoded hemagglutinin (parainfluenza, influenza) in cellular membranes. This reaction becomes positive before cytopathic changes are visible and in some cases occurs in the absence of cytopathic effects (Figure 1D).
5. Viral growth in an embryonated chick egg may result in death of the embryo (eg, encephalitis viruses), production of pocks or plaques on the chorioallantoic membrane (eg, herpes, smallpox, and vaccinia), or development of hemagglutinins in the embryonic fluids or tissues (eg, influenza).
Fig1. Cytopathic effects produced in monolayers of cultured cells by different viruses. The cultures are shown as they would normally be viewed in the laboratory, unfixed and unstained (60×). A: Enterovirus—rapid rounding of cells progressing to complete cell destruction. B: Herpesvirus—focal areas of swollen, rounded cells. C: Paramyxovirus—focal areas of fused cells (syncytia). D: Hemadsorption. Erythrocytes adhere to those cells in the monolayer that are infected by a virus that causes a hemagglutinin to be incorporated into the plasma membrane. Many enveloped viruses that mature by budding from cytoplasmic membranes produce hemadsorption. (Courtesy of I Jack.)
B. Inclusion Body Formation
In the course of viral multiplication within cells, virus specific structures called inclusion bodies may be produced. They become far larger than the individual virus particle and often have an affinity for acid dyes (eg, eosin). They may be situated in the nucleus (herpesvirus), in the cytoplasm (poxvirus, rabies virus), or in both (measles virus). In many viral infections, the inclusion bodies are the site of development of the virions (the viral factories). Variations in the appearance of inclusion material depend on the tissue fixative and stain used.
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