Leishmania donovani: Intraspecific polymorphisms of Sudanese isolates revealed by PCR-based analyses and DNA sequencing

Four polymerase chain reaction (PCR)-based approaches were used to analyze diversity within 23 Sudanese isolates of Leishmania donovani. Methods compa red were fingerprinting with single nonspecific primers, restriction analys is of the amplified ribosomal internal transcribed spacer (ITS) locus, sing le-stranded conformation polymorphism (SSCP), and sequencing of the ITS reg ion. When PCR fingerprinting and restriction analysis of ITS were applied, highly similar fragment patterns were observed for all strains of L. donova ni studied. The ITS1 locus gave five different SSCP profiles among the 23 S udanese isolates, whereas the ITS2 locus was highly conserved with the exce ption of 1 isolate. Strains of L. donovani derived from other geographical areas were found to have different ITS2 patterns. SSCP analysis correlated well with results of DNA sequencing and confirmed that SSCP was able to det ect genetic diversity at the level of a single nucleotide. SSCP had advanta ges over the other methods employed for investigation of sequence variation within the species L. donovani. There was no correlation between the form of clinical manifestation of the disease and the PCR fingerprinting, ITS-RF LP, or ITS-SSCP characteristics. (C) 2001 Academic Press.