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Updated: Aug 15, 2026
Aseptic Laboratory Techniques: Plating Methods
Published on: May 11, 2012
Effects of various suspending media on plaque formation by rickettsiae in tissue culture
Abstract:
Effects of some media used for suspending rickettsiae during purification, for metabolic studies, and in titrations of infectious rickettsiae were examined with respect to the plaque-forming ability of Rickettsia rickettsi and R. typhi in primary chicken embryo tissue cultures and the infectivity of R. typhi in mice. Brain heart infusion broth (BHI) was found superior to all other media tested in preventing both a significant decrease in plaque-forming units (PFU) and a delay in plaque formation. Skim milk, egg yolk, and some metabolic media were effective in maintaining PFU at 0 C, but did not prevent a significant delay in plaque formation. However, infectivity of R. typhi for tissue culture and mice was markedly decreased when suspended in metabolic media at 26 C. Addition of BHI to the routine tissue culture overlay reversed the deleterious effects of sucrose-phosphate solutions. The effects of Mg(2+), Mn(2+), K(+), Na(+), sucrose, and glutamate were also examined. No significant differences were observed between R. rickettsi and R. typhi in their responses to different media. The results of this study suggest the necessity for a reappraisal of previous studies of metabolism and infectivity of rickettsiae in these media.
Insights
Brain heart infusion broth (BHI) best preserves Rickettsia plaque-forming ability and infectivity. Other media, like skim milk or metabolic media, can delay plaque formation or decrease infectivity, necessitating a review of past rickettsial studies.
Area of Science:
- Microbiology
- Infectious Diseases
- Cell Culture
Background:
- Rickettsiae are obligate intracellular bacteria causing significant human diseases.
- Previous studies on rickettsial metabolism and infectivity may be influenced by the suspending media used.
- Standardization of media is crucial for accurate assessment of rickettsial viability and infectivity.
Purpose of the Study:
- To evaluate the impact of various suspending media on the plaque-forming ability of Rickettsia rickettsi and Rickettsia typhi.
- To assess the effect of different media on the infectivity of Rickettsia typhi in mice and tissue cultures.
- To determine the optimal medium for maintaining rickettsial viability and infectivity for experimental purposes.
Main Methods:
- Rickettsiae were suspended in different media, including brain heart infusion broth (BHI), skim milk, egg yolk, and metabolic media.
- Plaque-forming ability was assessed in primary chicken embryo tissue cultures.
- Infectivity of R. typhi was evaluated in mice and tissue cultures.
- The effects of temperature and specific ions (Mg2+, Mn2+, K+, Na+) and nutrients (sucrose, glutamate) were examined.
Main Results:
- Brain heart infusion broth (BHI) significantly outperformed other tested media in maintaining plaque-forming units (PFU) and preventing delayed plaque formation.
- Skim milk and egg yolk maintained PFU at 0°C but did not prevent delayed plaque formation.
- Metabolic media at 26°C markedly decreased R. typhi infectivity for tissue culture and mice.
- Addition of BHI to sucrose-phosphate solutions reversed deleterious effects.
Conclusions:
- Brain heart infusion broth (BHI) is superior for suspending rickettsiae in plaque assays and infectivity studies.
- The choice of suspending medium significantly impacts rickettsial viability and experimental outcomes.
- Previous studies utilizing suboptimal media for rickettsial metabolism and infectivity may require re-evaluation.
