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Some factors influencing the in vitro infectivity and replication of Encephalitozoon cuniculi
Abstract:
Rabbit Encephalitozoon cuniculi were propagated in vitro using rabbit choroid plexus (RCP) cells. The organisms reached maximum titer and numbers by 15 days. The source and in vitro passage level of RCP cells moderately influenced the sensitivity of the cells to infection. Cells less than 1 week old were significantly less sensitive than older cells. A moderate increase in infectivity for RCP cells was demonstrated with increasing organism passage level in vitro. Rabbit E. cuniculi were not affected by penicillin-streptomycin or gentamicin in the culture medium. The organism survived more than 9 days in buffer at 37 C and least 24 days at 4 and 20 C. Storage at -70 C or in liquid nitrogen was successful for at least 6 months. Encephalitozoon cuniculi survived 60 but not 120 min at 56 C. They were killed after 10 min of autoclaving and by 2% (v/v) Lysol, 10% (v/v) formalin and 70% (v/v) ethyl alcohol. The organisms survived at least 24 h at pH 9 or pH 4 and were not affected by sonication, freezing and thawing, or distilled water but lost significant infectivity after 24 h in CsCl or 40% (w/v) sucrose.
Insights
Encephalitozoon cuniculi propagation in vitro using rabbit choroid plexus cells is feasible. This study details optimal conditions and factors affecting parasite survival and infectivity for research applications.
Area of Science:
- Microbiology
- Cell Biology
- Parasitology
Background:
- Encephalitozoon cuniculi is an important microsporidian parasite.
- Understanding its in vitro propagation is crucial for research.
- Rabbit choroid plexus cells offer a viable model system.
Purpose of the Study:
- To establish and optimize in vitro propagation of Rabbit Encephalitozoon cuniculi.
- To determine factors influencing cell sensitivity and parasite infectivity.
- To assess the viability and storage conditions for the parasite.
Main Methods:
- In vitro propagation of E. cuniculi using rabbit choroid plexus (RCP) cells.
- Evaluation of cell age and passage level on infection sensitivity.
- Assessment of parasite survival under various temperature, pH, and chemical conditions.
- Testing of cryopreservation and disinfectant efficacy.
Main Results:
- Maximum E. cuniculi titer achieved by 15 days post-infection.
- Younger RCP cells (<1 week) showed reduced sensitivity to infection.
- Increased parasite passage level enhanced RCP cell infectivity.
- E. cuniculi survived >9 days at 37°C, >24 days at 4°C/20°C, and 6 months with cryopreservation.
- Parasite was inactivated by heat (56°C for 120 min), autoclaving, Lysol, formalin, and ethanol.
- E. cuniculi survived extreme pH (4 and 9) and sonication but lost infectivity in CsCl and sucrose.
Conclusions:
- Rabbit choroid plexus cells support robust in vitro propagation of E. cuniculi.
- Cell age and parasite passage level are key factors for successful infection.
- Specific storage and inactivation methods are identified for E. cuniculi handling.
- These findings provide a foundation for further research on E. cuniculi pathogenesis and treatment.