Evaluation of cytochrome b sequence to identify Leishmania species and variants: the case of Panama

BACKGROUND: The genetic heterogeneity of Leishmania parasites is a major factor responsible for the wide variety of Leishmania-associated manifestations. Consequently, understanding the genetic make-up of Leishmania species using suitable molecular markers is an important component of realising local and regional scale disease risk. The cytochrome b (cytb) is frequently used to type New World Leishmania species. However, its potential to discriminate Leishmania species and variants requires further evaluation. OBJECTIVES: To explore the capacity of cytb gene to identify New World Leishmania species and variants and to develop an approach able to type local Leishmania species and variants. METHODS: We retrieved 360 partial and complete Leishmania cytb gene sequences publicly available in GenBank database to study all single nucleotide polymorphisms (SNPs) across the cytb gene that differentiate New World Leishmania species. This information was used to develop an approach based upon the polymorphisms found in a DNA segment of 948bp. We also compared the typing results found with this technique with the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) profiling obtained using HSP70 gene as target. One hundred Panamanian isolates were used to both typed Leishmania species and assess local genetic variability. FINDINGS: We found complete agreement between our cytb approach and the PCR-RFLP profiling method based on HSP70 for Leishmania species identification. Ninety-two isolates were identified as L. panamensis, although other Viannia species were found circulating at a lower frequency. Three L. panamensis haplotypes were identified in Panamanian provinces. We also provide an initial report of L. guyanensis haplotypes circulating in Panama. MAIN CONCLUSIONS: Cytb gene sequence encompasses key main SNPs that aid to identify Leishmania species. The cytb approach developed with this information was able to identify and assess genetic variability of local Leishmania species found in this study.