Transformation of Anaplasma Phagocytophilum

Overview

Journal BMC Biotechnol

Publisher Biomed Central

Specialty Biotechnology

Date 2006 Nov 2

PMID 17076894

Citations 67

Authors

Roderick F Felsheim

Michael J Herron

Curtis M Nelson

Nicole Y Burkhardt

Anthony F Barbet

Timothy J Kurtti

Ulrike G Munderloh

Affiliations

Soon will be listed here.

Abstract

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.

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