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Antimicrobial Agents and Chemotherapy, August 2005, p. 3274-3280, Vol. 49, No. 8
0066-4804/05/$08.00+0     doi:10.1128/AAC.49.8.3274-3280.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Production and Characterization of Stable Amphotericin-Resistant Amastigotes and Promastigotes of Leishmania mexicana

Hamdan I. Al-Mohammed, Michael L. Chance, and Paul A. Bates^*

Liverpool School of Tropical Medicine, Pembroke Place, Liverpool L3 5QA, United Kingdom

Received 15 March 2005/ Returned for modification 12 April 2005/ Accepted 11 May 2005

The sensitivities of Leishmania mexicana amastigote and promastigote forms to amphotericin B were investigated in vitro and found to be strongly influenced by the culture media used. When differences in culture media were minimized, there was no significant difference in the 50% inhibitory concentration values between the two life cycle stages. Stable amphotericin B-resistant amastigote and promastigote lines were produced by the application of increasing drug pressure to long-term cultures. Lines capable of growth in concentrations of amphotericin B lethal to normal parasites were produced. Compared to normal parasites, these amphotericin-resistant lines showed marked differences in membrane sterol compositions, with very high levels of 4,14,dimethyl-cholesta-8,24-dienol and other methyl sterols. They also showed a consistent morphological feature, the presence of multilamellar membrane-like material in the flagellar pocket, revealed by transmission electron microscopy. Amphotericin-resistant parasites were capable of infecting BALB/c mice, but the resulting lesion growth was slower than that after infection with normal parasites. However, unlike normal parasites, the amphotericin-resistant parasites were unaffected by experimental chemotherapy with amphotericin B. These results show that amphotericin B resistance could arise as a result of increased clinical use of amphotericin B therapy.

Present address: Department of Biology, College of Science, King Faisal University, P.O. Box 10590, Riyadh 11443, Saudi Arabia.