Volume 72, Issue 4
  • ISSN: 0002-9637
  • E-ISSN: 1476-1645


We evaluated the -specific primers B1-B2 to detect in normal skin and peripheral blood monocytes of patients with active cutaneous leishmaniasis. Southern blotting and sequencing of polymerase chain reaction (PCR) products confirmed the specificity of kinetoplast DNA (kDNA) amplification from tissue fluid from healthy skin, whereas the PCR with monocytes also amplified a human sequence of a size similar (718 basepairs) to the expected kDNA product (750 basepairs), resulting in false-positive results. Although B1 was not homologous to any human DNA sequence, B2 showed homology to a human chromosome 2 intergenic region (AC010878) at positions 35,881-36,599, which are spaced 718 nucleotides apart. Amplification of the human art3fact from monocyte DNA was confirmed using the primer B2 alone. Examination of other primers reported for the PCR of kDNA from various species of showed that six of seven were homologous to human DNA sequences. These findings underscore the importance of exploiting sequencing, bioinformatics, and DNA probes to refine molecular amplification techniques and to validate the performance of primers when used for new applications.


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  1. Desjeux P, 2001. Worldwide increasing risk factors for leishmaniasis. Med Microbiol Immunol (Berl) 190 : 77–79. [Google Scholar]
  2. Herwaldt BL, 1999. Leishmaniasis. Lancet 354 : 1191–1199. [Google Scholar]
  3. Brujin M, Barker D, 1992. Diagnosis of new world leishmaniasis: specific detection of species of the Leishmania braziliensis complex by amplification of kinetoplast DNA. Acta Trop 52 : 45–48. [Google Scholar]
  4. Castilho T, Shaw MJ, Floeter-Winter LM, 2003. New PCR assay using glucose-6-phosphate dehydrogenase for identification of Leishmania species. J Clin Microbiol 41 : 540–546. [Google Scholar]
  5. Marturf J, Nasereddin A, Niederwieser I, Jaffe CL, Beck HP, Felger I, 2003. Identification and differentiation of Leishmania species in clinical samples by PCR amplification of the miniexon sequence and subsequent restriction fragment length polymorphism analysis. J Clin Microbiol 41 : 3147–3153. [Google Scholar]
  6. Weigle K, Labrada LA, Lozano C, Santrich C, Barker D, 2002. PCR-based diagnosis of acute and chronic cutaneous leishmaniasis caused by Leishmania (Viannia). J Clin Microbiol 40 : 601–606. [Google Scholar]
  7. Belli A, Rodriguez B, Aviles H, Harris E, 1998. Simplified polymerase chain reaction detection of new world Leishmania in clinical specimens of cutaneous leishmaniasis. Am J Trop Med Hyg 58 : 102–109. [Google Scholar]
  8. Rodgers MR, Pepper SJ, Wirth DF, 1990. Amplification of kine-toplastic DNA as a tool in the detection and diagnosis of Leishmania. Exp Parasitol 71 : 267–275. [Google Scholar]
  9. Uliana SRB, Nelson K, Beverley SM, Camargo EP, Floeter-Winter LM, 1994. Discrimination amongst Leishmania by polymerase chain reaction and hybridization with small subunit ribosomal DNA derived oligonucleotides. J Eukaryot Microbiol 41 : 324–330. [Google Scholar]
  10. Mendonca MG, de Brito ME, Rodrigues EH, Bandeira V, Jardim ML, Abath FG, 2004. Persistence of Leishmania parasites in scars after clinical cure of American cutaneous leishmaniasis: is there sterile cure? J Infect Dis 189 : 1018–1023. [Google Scholar]
  11. Palma G, Gutierrez Y, 1991. Laboratory diagnosis of Leishmania. Clin Lab Med 11 : 909–922. [Google Scholar]
  12. Ausubel F, Brent MR, Kingston RE, Moor DD, Seidman JG, Smith JA, Struhl K, 1987. Current Protocols in Molecular Biology. New York: John Wiley & Sons Inc., 6.2.1–6.2.3.
  13. Altschul SF, Gish W, Miller W, Myers EW, Lipman DJ, 1990. Basic local alignment search tool. J Mol Biol 215 : 403–410. [Google Scholar]
  14. Higgins D, Thompson J, Gibson T, Thompson JD, Higgins DG, Gibson TJ, 1994. CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res 22 : 4673–4680. [Google Scholar]
  15. Gomes-Rodrigues EH, Felinto de Brito ME, Mendonça MG, Wekhäuser RP, Coutinho EM, Souza WV, Militão de Albuquerque Mde F, Jardim ML, Abath FG, 2002. Evaluation of PCR for diagnosis of American cutaneous leishmaniasis in an area of endemicity in northeastern Brazil. J Clin Microbiol 40 : 3572–3576. [Google Scholar]
  16. Guevara P, Rojas E, Gonzalez N, Scorza JV, Añez N, Valera M, Ramirez JL, 1994. Presence of Leishmania braziliensis in blood samples from cures patients at different stages of immunotherapy. Clin Diag Lab Immunol 1 : 385–389. [Google Scholar]
  17. Salotra P, Sreenivas G, Pogue G, Lee N, Nakhasi H, Ramesh V, Negi NS, 2001. Development of a species -specific PCR assay for detection of Leishmania donovani in clinical samples from patients with kalaazar and post-kalaazar dermal leishmaniasis. J Clin Microbiol 39 : 849–854. [Google Scholar]
  18. Galindo I, Rangel-Aldao R, Ramirez JL, 1993. A combined polymerase chain reaction-colour development hybridization assay in a microtiter format for the detection of Clostridium sp. Appl Microbiol Biotechnol 39 : 553–557. [Google Scholar]

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  • Received : 18 Jun 2004
  • Accepted : 24 Oct 2004

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