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Virulence of Mycobacterium tuberculosis for guinea pigs: a quantitative modification of the assay developed by
V Balasubramanian1, W Guo-Zhi, E Wiegeshaus
1Department of Medical Microbiology and Immunology, University of Wisconsin, Madison.
Abstract:
One of the more accepted methods of assay of virulence of tubercle bacilli is one developed by Mitchison in which guinea pigs were infected by the intramuscular route with 1.0 mg of tubercle bacilli freshly harvested from Lowenstein Jensen medium and in which virulence was based on a subjective score of the extent of gross disease in the animal 6 weeks after infection. Due to the practical difficulties involved in such an assay when routinely performed, the following modifications were made: frozen stocks of the culture were prepared prior to the date of infection, the inoculum was quantitated by means of colony forming units, and virulence was based on the number of tubercle bacilli recovered from the spleen of infected animals 6 weeks after infection. A significant correlation was obtained between the findings from the Mitchison assay of virulence and the modification. The latter assay is recommended for its quantitative and objective features.
Insights
This study refines a method for assessing the virulence of tubercle bacilli. The modified assay uses quantitative colony forming units and spleen bacterial counts for objective and reliable virulence measurement in guinea pigs.
Area of Science:
- Microbiology
- Immunology
- Infectious Diseases
Background:
- The Mitchison assay is a standard method for evaluating tubercle bacilli virulence.
- This assay relies on subjective scoring of gross disease in guinea pigs six weeks post-infection.
- Practical challenges exist in the routine application of the original Mitchison assay.
Purpose of the Study:
- To develop a more practical, quantitative, and objective assay for tubercle bacilli virulence.
- To validate a modified virulence assay against the established Mitchison method.
Main Methods:
- Prepared frozen stocks of tubercle bacilli cultures prior to infection.
- Quantitated the inoculum using colony forming units (CFUs).
- Assessed virulence by enumerating tubercle bacilli in the spleens of infected guinea pigs after six weeks.
Main Results:
- A significant correlation was found between the original Mitchison assay and the modified assay.
- The modified assay demonstrated quantitative and objective measurement of virulence.
- The modified assay overcomes practical difficulties of the original method.
Conclusions:
- The modified assay provides a reliable, objective, and quantitative method for assessing tubercle bacilli virulence.
- This refined method is recommended for routine virulence assays.
- The quantitative approach enhances reproducibility and comparability of virulence data.
