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Anaplasma marginale in tick cell culture
R J Hidalgo1, E W Jones, J E Brown
1College of Veterinary Medicine, Mississippi State University 39762.
Abstract:
Anaplasma marginale was propagated in a tick cell line derived from Dermacentor variabilis embryos. The rickettsial organism was identified and monitored in culture by transmission electron microscopy and the indirect immunofluorescence technique, using specific monoclonal antibodies. Inoculation of the embryonic tick cell line with midguts of infected adult ticks (culture 1), nymphal ticks (culture 2) and adult ticks that were infected as nymphs and dissected as adults (culture 3) resulted in 3 continuous cultures of A marginale. Culture 1 had been maintained through 22 passages over a 11-month period; cultures 2 and 3 had been maintained for 18 passages over a 9-month period. Growth of A marginale in the cell line began in the area of the nuclear membrane at approximately 4 days after inoculation or transfer. Thereafter, the organisms were observed in inclusions scattered throughout the cytoplasm of the host cells. Maximal growth of the organism occurred at 7 to 14 days, after which numbers of inclusions rapidly decreased to minimal or undetectable levels. The organism began new cycles of growth with each 1:5 to 1:10 split and transfer of the host cells. Electron microscopy of recently infected cells revealed a morphology of the organism that closely resembled that observed in marginal bodies of infected erythrocytes. After several passages, A marginale organisms had a varied morphology and resembled the organism described in midgut cells of naturally infected ticks.(ABSTRACT TRUNCATED AT 250 WORDS)
Insights
Anaplasma marginale was successfully cultured in a tick cell line from Dermacentor variabilis embryos. This research establishes a continuous Anaplasma marginale culture for further study.
Area of Science:
- Veterinary Entomology
- Microbiology
- Cell Biology
Background:
- Anaplasma marginale is an economically significant tick-borne pathogen.
- Establishing continuous cell cultures is crucial for studying tick-borne rickettsial organisms.
Purpose of the Study:
- To propagate Anaplasma marginale in a tick cell line.
- To characterize the growth and morphology of Anaplasma marginale in vitro.
Main Methods:
- Propagation of Anaplasma marginale in a Dermacentor variabilis embryonic cell line.
- Monitoring using transmission electron microscopy and indirect immunofluorescence with monoclonal antibodies.
- Continuous culture maintenance through multiple passages over extended periods.
Main Results:
- Three continuous Anaplasma marginale cultures were established from infected tick tissues.
- Organisms initiated growth near the nuclear membrane and formed cytoplasmic inclusions.
- Maximal growth occurred between 7-14 days, with new growth cycles initiated upon cell transfer.
- Morphology varied from erythrocyte-associated forms to midgut-like forms after several passages.
Conclusions:
- A tick cell line derived from Dermacentor variabilis embryos supports continuous Anaplasma marginale propagation.
- The established cell culture system provides a valuable tool for studying Anaplasma marginale.
- In vitro culture allows for observation of Anaplasma marginale's life cycle and morphological changes.