title: Diagnosis of visceral leishmaniasis by polymerase chain reaction of DNA extracted from Giemsa's solution-stained slides.
abstract: Visceral leishmaniasis (VL) is caused by the protozoan parasite Leishmania donovani and is a potentially fatal disease in endemic areas of the world. Nepal is an endemic area in which VL causes major public health problems in the lowland areas of the southeast regions. The aim of the present study was to evaluate the sensitivity of polymerase chain reaction (PCR) amplification for the detection of Leishmania DNA from Giemsa's solution-stained bone marrow slides. Bone marrow samples were aspirated from a total of 115 VL suspected patients and used to prepare smears on glass slides and for the initiation of in vitro culture. Bone marrow slides were used for microscopic observation, DNA extraction, and subsequent PCR amplification. PCR analysis showed that all the positive samples were of Leishmania parasites. The PCR assay also showed a higher sensitivity (69%) than microscopic examination (57%) and culture (21%). In addition, PCR was able to detect VL in 12% of samples which were negative by microscopy. PCR of DNA extracted from Giemsa's solution-stained bone marrow slides is a suitable tool for confirming diagnosis in patients with VL and may also be useful in the diagnosis of difficult cases. Bone marrow smears are easily stored and can be easily sent to research centers where PCR is available. This makes PCR a good option for diagnosis in the field.
authors.affiliation: Department of Immunogenetics, Institute of Tropical Medicine (NEKKEN) and the Global COE Program, Nagasaki University, Sakamoto, Nagasaki, Japan. pandey_kishor@hotmail.com
authors.forename: Kishor
authors.initials: K
authors.lastname: Pandey
authors.forename: Basu Dev
authors.initials: BD
authors.lastname: Pandey
authors.forename: Arun Kumar
authors.initials: AK
authors.lastname: Mallik
authors.forename: Osamu
authors.initials: O
authors.lastname: Kaneko
authors.forename: Haruki
authors.initials: H
authors.lastname: Uemura
authors.forename: Hiroji
authors.initials: H
authors.lastname: Kanbara
authors.forename: Tetsuo
authors.initials: T
authors.lastname: Yanagi
authors.forename: Kenji
authors.initials: K
authors.lastname: Hirayama
mesh_terms: D000818:Animals; D001399:Azure Stains; D001853:Bone Marrow / Q000469:parasitology; D004396:Coloring Agents; D016054:DNA, Protozoan / Q000032:analysis / Q000302:isolation & purification*; D006801:Humans; D007893:Leishmania donovani / Q000235:genetics / Q000302:isolation & purification*; D007898:Leishmaniasis, Visceral / Q000175:diagnosis*; D008853:Microscopy / Q000379:methods; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity
chemical_list: D001399:Azure Stains; D004396:Coloring Agents; D016054:DNA, Protozoan