Identification and differentiation of Iranian Leishmania species by PCR amplification of kDNA

We describe the specific identification of Leishmania species in Iran using PCR DNA amplification of kDNA. For this purpose, we designed a pair of pri mers - upstream 5 ' TCGCAGAACGCCCCTACC 3 ' and downstream 5 ' -AGGGGTTGGT-G TAAAATAGGC 3 ' - specific for conserved sequences of kDNA of Leishmania. Us ing this primer, we identified 3 different amplified fragments from the kDN A of the WHO reference Leishmania species. Two bands at 620 and 850 bp were identified for L. major (MRHO/IR/64/Nadim-1 strain) and only 1 band at 620 bp was identified for L. major (P strain). Therefore, we could differentia te 2 Leishmania species. Also, 1 band at 830 bp was identified for L. tropi ca (MHOM/Sudan/58/OD strain). We determined the sequence analysis of 2 DNA bands (620 and 850 bp) obtained from kDNA of L. major ((MRHO/IR/64/Nadim-1) . A total of 157 bp from the 5 ' site and 234 bp from the 3 ' site were seq uenced and showed about 28% homology between 620 and 850 bp fragments. This technique could amplify as little as I fg of DNA and was used to different iate kDNA samples isolated from Iranian patients with. cutaneous leishmania sis. These data indicate that the primer used for PCR amplification of kDNA is specific and can be used for diagnostic and epidemiological purposes.