Transformation of Anaplasma phagocytophilum

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. To facilitate genetic manipulation, we transformed A. phagocytophilum...

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Bibliographic Details
Published in:BMC biotechnology 2006-10, Vol.6 (1), p.42-42
Main Authors: Felsheim, Roderick F, Herron, Michael J, Nelson, Curtis M, Burkhardt, Nicole Y, Barbet, Anthony F, Kurtti, Timothy J, Munderloh, Ulrike G
Format: Article
Language:English
Subjects:
Anaplasma
Anaplasma phagocytophilum
Anaplasma phagocytophilum - genetics
Anaplasma phagocytophilum - metabolism
Animals
Green Fluorescent Proteins - genetics
Green Fluorescent Proteins - metabolism
Haplorhini
Humans
Ixodidae
Methodology
Ticks
Transformation, Bacterial - physiology
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Summary: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. 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. 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.
ISSN:1472-6750
1472-6750
DOI:10.1186/1472-6750-6-42