Exploiting genetic polymorphisms in metabolic enzymes for rapid screening of Leishmania infantum genotypes

Leishmania infantum is the aetiological agent of visceral leishmaniasis (VL) and cutaneous leishmaniasis (CL). Numerous strains and/or zymodemes have been identified and characterized by multilocus enzyme electrophoresis (MLEE). MLEE is considered the reference method for L. infantum parasite typing and it is based upon enzyme electrophoretic mobility analysis from promastigote cultures. However, the MLEE technique is cumbersome, time-consuming and does not detect silent genetic mutations or nucleotide changes that give rise to amino acid changes that do not alter electrophoretic mobility. As a result of these difficulties, many DNA-based typing methods have been developed over the past few years. However, relative to the enzymes utilized in MLEE analysis, we observed a shortage of DNA sequences available in the GenBank database or an absolute lack of sequences belonging to specific zymodemes. The aims of the present study were to (i) implement the number of sequences coding for metabolic enzymes used in MLEE; (ii) identify polymorphisms that characterize L. infantum zymodemes most prevalent in the Mediterranean basin; and (iii) exploit these polymorphisms to develop a rapid screening test that would give results comparable with existing MLEE typing.