An enhanced method for the identification of Leishmania spp. using real-time polymerase chain reaction and sequence analysis of the 7SL RNA gene region.
The accurate identification of Leishmania spp. is important for the treatment of infected patients. Molecular methods offer an alternative to time-consuming traditional laboratory techniques for species determination. We redesigned a 7SL RNA gene-based polymerase chain reaction and sequence assay for increased species identification. DNA extracted from 17 reference strains and 10 cultured clinical isolates was examined. Sequence comparison was used successfully to identify organisms to the complex level with intercomplex similarity ranging from 77.5% to 98.4%. Many species within each complex were discriminated accurately by this method including Leishmania major, Leishmania tropica, Leishmania aethiopica, Leishmania guyanensis, and the previously indistinguishable Leishmania braziliensis and Leishmania panamensis. The Leishmania donovani complex members remain indistinguishable by this method, as are the representatives of Leishmania amazonensis/Leishmania garnhami and Leishmania mexicana/Leishmania pifanoi.
Microbiology Service, Department of Laboratory Medicine, Clinical Center, National Institutes of Health, Bethesda, MD 20892, USA.
Lindsay G
LG
Stevenson
Daniel P
DP
Fedorko
Adrian M
AM
Zelazny
D016054:DNA, Protozoan / Q000737:chemistry / Q000235:genetics; D006801:Humans; D007891:Leishmania / Q000145:classification* / Q000235:genetics* / Q000302:isolation & purification; D007896:Leishmaniasis / Q000175:diagnosis / Q000469:parasitology*; D008969:Molecular Sequence Data; D010274:Parasitology / Q000379:methods*; D016133:Polymerase Chain Reaction / Q000379:methods*; D011110:Polymorphism, Genetic; D016053:RNA, Protozoan / Q000235:genetics*; D020733:RNA, Small Cytoplasmic / Q000235:genetics*; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA; D012689:Sequence Homology, Nucleic Acid; D018271:Signal Recognition Particle / Q000235:genetics*
C115990:7SL RNA; D016054:DNA, Protozoan; D016053:RNA, Protozoan; D020733:RNA, Small Cytoplasmic; D018271:Signal Recognition Particle