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Antimicrobial Agents and Chemotherapy, September 2003, p. 2725-2731, Vol. 47, No. 9
0066-4804/03/$08.00+0     DOI: 10.1128/AAC.47.9.2725-2731.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Recapitulation in Saccharomyces cerevisiae of Cytochrome b Mutations Conferring Resistance to Atovaquone in Pneumocystis jiroveci

Philip Hill,1 Jacques Kessl,2 Nicholas Fisher,1 Steven Meshnick,3 Bernard L. Trumpower,2* and Brigitte Meunier1*

Wolfson Institute for Biomedical Research, University College London, London, United Kingdom,1 Department of Biochemistry, Dartmouth Medical School, Hanover, New Hampshire,2 Department of Microbiology and Immunology, University of North Carolina, Chapel Hill, North Carolina3

Received 31 March 2003/ Returned for modification 12 May 2003/ Accepted 19 June 2003

Pneumocystis jiroveci (human-derived P. carinii) is an opportunistic pathogenic fungus which causes pneumonia and is life-threatening in immunocompromised individuals. Spontaneously acquired resistance to atovaquone, a hydroxynaphthoquinone that is used to treat P. jiroveci infections, was linked to mutations in the mitochondrially encoded cytochrome b gene. Because P. jiroveci cannot be easily cultivated, we have developed Saccharomyces cerevisiae as an alternative system to study atovaquone resistance mutations. In this work, we introduced seven mutations linked with atovaquone resistance in P. jiroveci into the S. cerevisiae cytochrome b gene. The effects of the mutations on the respiratory function and on the sensitivity to the inhibitor were then characterized. Six of the reported mutations lowered the sensitivity of the S. cerevisiae bc1 complex to atovaquone, while one mutation had no effect on the drug resistance. These results were confirmed by monitoring the in vivo resistance of S. cerevisiae mutants which carried both the cytochrome b mutations and a deletion of the ABC transporter genes, allowing the drug to bypass the weakened efflux pump system. S. cerevisiae thus provides an easy-to-use system to characterize in vivo and in vitro cytochrome b mutations reported in pathogens and to assess their role in drug resistance.

* Corresponding authors. Mailing address for B. Meunier: Wolfson Institute for Biomedical Research, University College London, London, United Kingdom. Phone: 44 20 7679 6860. Fax: 44 20 7679 6860. E-mail: b.meunier{at}ucl.ac.uk. Mailing address for B. L. Trumpower: Department of Biochemistry, Dartmouth Medical School, Hanover, N.H. E-mail: Trumpower{at}Dartmouth.edu.

Antimicrobial Agents and Chemotherapy, September 2003, p. 2725-2731, Vol. 47, No. 9
0066-4804/03/$08.00+0     DOI: 10.1128/AAC.47.9.2725-2731.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



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