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Infection and Immunity, October 2005, p. 6736-6741, Vol. 73, No. 10
0019-9567/05/$08.00+0     doi:10.1128/IAI.73.10.6736-6741.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Isocitrate Lyase Activity Is Required for Virulence of the Intracellular Pathogen Rhodococcus equi

Daniel M. Wall, Pamela S. Duffy,1,{dagger} Chris DuPont,2 John F. Prescott,2 and Wim G. Meijer1*

Department of Industrial Microbiology, Conway Institute of Biomolecular and Biomedical Research, University College Dublin, Dublin 4, Ireland,1 Department of Pathobiology, University of Guelph, Guelph, Ontario N1G 2W1, Canada2

Received 30 March 2005/ Returned for modification 16 May 2005/ Accepted 13 July 2005

Rhodococcus equi is an important pathogen of foals, causing severe pyogranulomatous pneumonia. Virulent R. equi strains grow within macrophages, a process which remains poorly characterized. A potential source of carbon for intramacrophage R. equi is membrane lipid-derived fatty acids, which following ß oxidation are assimilated via the glyoxylate bypass. To assess the importance of isocitrate lyase, the first enzyme of the glyoxylate bypass, in virulence of a foal isolate of R. equi, a mutant was constructed by a strategy of single homologous recombination using a suicide plasmid containing an internal fragment of the R. equi aceA gene encoding isocitrate lyase. Complementation of the resulting mutant with aceA showed that the mutant was specific for this gene. Assessment of virulence in a mouse macrophage cell line showed that the mutant was killed, in contrast to the parent strain. Studies in the liver of intravenously infected mice showed enhanced clearance of the mutant. When four 3-week-old foals were infected intrabronchially, the aceA mutant was completely attenuated, in contrast to the parent strain. In conclusion, the aceA gene was shown to be essential for virulence of R. equi, suggesting that membrane lipids may be an important source of carbon for phagocytosed R. equi.

* Corresponding author. Mailing address: Department of Industrial Microbiology, University College Dublin, Dublin 4, Ireland. Phone: 353-1716-1364. Fax: 353-1716-1183. E-mail: wim.meijer{at}ucd.ie.

Editor: J. B. Bliska

{dagger} Present address: Department of Pediatric Gastroenterology and Nutrition, Massachusetts General Hospital, 114 16th St. (114-3503), Charlestown, MA 02129.

Infection and Immunity, October 2005, p. 6736-6741, Vol. 73, No. 10
0019-9567/05/$08.00+0     doi:10.1128/IAI.73.10.6736-6741.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.



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