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Optimization of Microculture and Evaluation of Miniculture for the Isolation of Leishmania Parasites from Cutaneous Lesions in Peru

Traditional culture of Leishmania parasites is labor-intensive and shows poor sensitivity. We evaluated microculture and novel miniculture methods for diagnosis of cutaneous leishmaniasis (CL). Consecutive patients who came to the Leishmaniasis Clinic, Hospital Nacional Cayetano Heredia, Lima, Peru, were enrolled. Lesion aspirates were cultured in traditional tubes containing Novy-MacNeal-Nicolle medium and in miniculture tubes (Eppendorf, Hamburg, Germany) and capillary tubes (microculture) containing RPMI 1640 medium containing 20% fetal bovine serum. The reference standard was positive results in two of four tests (smear, culture, polymerase chain reaction, or leishmanin skin test). Outcome measures were sensitivity and time to positivity. Fifty-five patients with 74 lesions were enrolled. Of 59 lesions that fulfilled reference criteria for CL, 50 were positive by microculture (sensitivity = 84.7%; P = 0.001), 45 by miniculture (sensitivity = 76.3%; P = 0.042), and 35 by traditional culture (sensitivity = 59.3%). Median time to positivity was three days by microculture and miniculture and five days by traditional culture (P < 0.001). Microculture and miniculture are sensitive and efficient means of diagnosing CL.

Received July 14, 2008. Accepted for publication August 27, 2008.

Acknowledgments: We thank Dr. Eduardo Gotuzzo and Ana Luz Quispe (Instituto de Medicina Tropical Alexander von Humboldt) for logistical support throughout this study.

^* Address correspondence to Andrea K. Boggild, Tropical Diseases Unit, Toronto General Hospital, 200 Elizabeth Street, North Wing, 13th Floor, Room 1350, Toronto, Ontario M5G 2C4, Canada. E-mail: andrea.boggild{at}utoronto.ca

Authors’ addresses: Andrea K. Boggild, Tropical Diseases Unit, Toronto General Hospital, 200 Elizabeth Street, North Wing, 13th Floor, Room 1350, Toronto, Ontario M5G 2C4, Canada, E-mail: andrea.boggild{at}utoronto.ca. Donald E. Low, Department of Microbiology, Mount Sinai Hospital, 600 University Avenue, Room 1485, Toronto, Ontario M5G 1X5, Canada, E-mail: Dlow{at}mtsinai.on.ca. Cesar Miranda-Verastegui, Diego Espinosa, Dalila Martinez-Medina, and Alejandro Llanos-Cuentas, Instituto de Medicina Tropical Alexander von Humboldt, Universidad Peruana Cayetano Heredia, Av. Honorio Delgado #430, URB Ingenieria, Lima 31, Peru, E-mails: cesar.miranda{at}mail.mcgill.ca, diegoespinosa{at}gmail.com, dalmaext{at}hotmail.com, and allanos{at}upch.edu.pe. Jorge Arevalo, Departmento de Bioquimica, Biologia Molecular y Farmacologia, Facultad de Ciencias, Universidad Peruana Cayetano Heredia, Av. Honorio Delgado #430, URB Ingenieria, Lima 31, Peru, E-mail: biomoljazz{at}gmail.com.