Mouse Model for Cytomegalovirus Infection

Cytomegaloviruses (CMVs) are prototypes of the P subgroup of herpesviruses. They possess large DNA genomes of ~230 kb with more than 200 open reading frames (Mocarski, 1996). Thus, they range among the mammalian viruses with the largest coding capacity. Although a high degree of species specificity is characteristic for cytomegaloviruses, viruses with a similar genomic structure and pathobiology are found in virtually all mammalian species analyzed. In vivo, CMVs infect a broad range of cells and tissues including fibroblasts, smooth muscle cells, endothelial and epithelial cells, stromal cells, and macrophages. As for herpesviruses in general, these viruses are not eliminated from an organism following primary infection. Instead, the virus persists in its host in a state of latency, where the full genome but no infectious progeny are detectable. Reactivation from latency occurs frequently and results in virus shedding and recurrent disease (Britt and Alford, 1996). Manifestation of the primary infection, as well as frequency and severity of recurrent disease, are controlled by the immune system. Thus, CMVs have proved to be valuable tools for studying virus-host interactions and multiple aspects of immunological control of infection.

The strict species specificity of CMVs precludes experimental infection of animals with human cytomegalovirus (hCMV). Therefore, rodent cytomegaloviruses (e.g., mouse, rat, and guinea pig) have been used as model systems to understand hCMV disease and to answer open questions that cannot easily be addressed in clinical research. Among these, murine cytomegalovirus (mCMV) is by far the best studied. This unit describes procedures for infecting newborn and adult mice with mCMV (Basic Protocol). Methods are included for propagating mCMV in cell cultures (Support Protocol 1) and for preparing a more virulent form of mCMV from salivary glands of infected mice (Support Protocol 2). Support Protocol 3 describes a plaque-forming cell (PFC) assay for measuring mCMV titers of infected tissues or virus stocks. Finally, Support Protocol 4 describes a method for preparing the murine embryonic fibroblasts used for propagating mCMV and for the PFC assay. The protocols should serve as a starting point for immunologists interested in the immune control of CMV infection, for geneticists analyzing immunodeficient mouse mutants, for researchers developing antiviral drug and immunotherapies, and finally for cell biologists who want to study virus-cell interactions in vitro. References to more specialized virological and immunological procedures are given (see Commentary; Reddehase and Lucin, 1993).

NOTE: mCMV is not a hazardous agent as it does not infect humans. Growth and purification of mCMV in vitro can be performed in any tissue culture laboratory with basic equipment including a laminar flow hood, a microscope, and a cell culture incubator. This is especially true if permissive fibroblast cell lines are used instead of primary murine embryonic fibroblasts (MEFs). Productive infection of fibroblasts results in cell rounding, plaque formation, and finally cell death, which can be followed by microscopic inspection. Analysis of mCMV infection and dissemination in vivo, by contrast, requires at least good basic experience in animal handling.

NOTE: All tissue culture procedures and preparation of virus and cells must be performed under sterile conditions in a laminar flow hood. Unless specified otherwise, cells and virus are grown at 37°C in a humidified atmosphere with 5% CO2.

Animal Models for Infectious Diseases

Contributed by Wolfram Brune, Hartmut Hengel, and Ulrich H. Koszinowski

Current Protocols in Immunology (1999) 19.7.1-19.7.13 Copyright © 1999 by John Wiley & Sons, Inc.

Diabetes Sustenance

Diabetes Sustenance

Get All The Support And Guidance You Need To Be A Success At Dealing With Diabetes The Healthy Way. This Book Is One Of The Most Valuable Resources In The World When It Comes To Learning How Nutritional Supplements Can Control Sugar Levels.

Get My Free Ebook


Post a comment