DIAGNOSIS OF KALA-AZAR BY NESTED PCR BASED ON AMPLIFICATION OF THE LEISHMANIA MINI-EXON GENE

To diagnose visceral leishmaniasis (kala-azar), we have developed a ne sted PCR method based on amplification of the mini-exon gene, which is unique and tandomly repeated in the Leishmania genome. Nested PCR was sufficiently sensitive for the detection of DNA in an amount equivale nt to a single Leishmania parasite or less. We examined the usefulness of this PCR method using bone marrow aspirates and buffy coat cells c ollected from kala-azar patients who had or had not received chemother apy in northwest China. We obtained PCR positivity for all of the para sitologically positive bone marrow samples from the patients. Some amb iguities with the primary PCR results were eliminated by the subsequen t nested PCR. The buffy coat samples from 7 of 12 patients with spleno megaly were positive by the nested PCR, although only 2 of them were p ositive for parasites by culture. However, buffy coat samples from nin e children, whose splenomegaly has been reduced and clinically cured b y antimony treatment, were all negative. Thus; this nested PCR method represents a new tool for the diagnosis of kala-azar,vith patient bloo d samples instead of bone marrow or spleen aspirates obtained by more invasive procedures.