Transformation of Anaplasma phagocytophilum

Roderick F. Felsheim, Michael J. Herron, Curtis M. Nelson, Nicole Y. Burkhardt, Anthony F. Barbet, Timothy J. Kurtti, Ulrike G. Munderloh

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Background: Tick-borne pathogens cause emerging zoonoses, and include fastidious organisms such as Anaplasma phagocytophilum. Because of their obligate intracellular nature, methods for mutagenesis and transformation have not been available. Results: To facilitate genetic manipulation, we transformed A. phagocytophilum (Ap) to express a green fluorescent protein (GFP) with the Himar1 transposase system and selection with the clinically irrelevant antibiotic spectinomycin. Conclusion: These transformed bacteria (GFP/Ap) grow at normal rates and are brightly fluorescent in human, monkey, and tick cell culture. Molecular characterization of the GFP/Ap genomic DNA confirmed transposition and the flanking genomic insertion locations were sequenced. Three mice inoculated with GFP/Ap by intraperitoneal injection became infected as demonstrated by the appearance of morulae in a peripheral blood neutrophil and re-isolation of the bacteria in culture.

Original languageEnglish (US)
Article number42
JournalBMC Biotechnology
StatePublished - Oct 31 2006

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    Felsheim, R. F., Herron, M. J., Nelson, C. M., Burkhardt, N. Y., Barbet, A. F., Kurtti, T. J., & Munderloh, U. G. (2006). Transformation of Anaplasma phagocytophilum. BMC Biotechnology, 6, [42].