Molecular Identification of Parasites Causing Cutaneous Leishmaniasis in Panama

Abstract

Isolates from 475 cutaneous leishmaniasis (CL) patients from three endemic regions were studied by three typing techniques. The molecular analysis from lesion scrapings based on hsp70 PCR-restriction fragment length polymorphism (RFLP) showed that 78.1% (371/475) restriction patterns corresponded to Leishmania (Viannia) panamensis, 19% (90/475) to Leishmania (Viannia) guyanensis, and 3.0% (14/475) to Leishmania (Viannia) braziliensis. Promastigotes isolated by culture from lesions of 228 patients (48.0%, 228/475) were identified by multi-locus enzyme electrophoresis. Of them, 95.2% (217/228) were typified as L. (V.) panamensis, 1.3% (3/228) as L. (V.) guyanensis, 2.2% (5/228) as L. (V.) braziliensis, and 1.3% (3/228) as hybrids (L. [V.] braziliensis/L. [V.] panamensis). However, a partial sequencing analysis of the hsp70 gene from 77 selected samples showed 16.9% (13/77) typified as L. (V.) panamensis, 68.8% (53/77) as Leishmania (V.) sp., 1, 3.9% (3/77) as L. (V.) guyanensis, 1.3% (1/77) as L. (V.) braziliensis outlier, 2.6% (2/77) as Leishmania (Viannia) naiffi, 2.6% as (2/77) Leishmania (V.) sp., and 2 and 3.9% (3/77) hybrid isolates of L. (V.) braziliensis/L. (V.) guyanensis. These results confirm L. (V.) panamensis as the predominant species and cause of CL lesions in Panama and that L. (V.) guyanensis, L. (V.) braziliensis, and L. (V.) naiffi are circulating to a lower degree. Furthermore, the determination of parasite isolates belonging to atypical clusters and hybrid isolates suggests the circulation of genetic variants with important implications for the epidemiology and clinical follow-up of CL in Panama. No evidence of the existence of parasites of the Leishmania (Leishmania) mexicana complex in Panamanian territory was found in this study.

Figure 1.

Map of Panama showing location of the study sites and the cutaneous leishmaniasis transmission intensity based on the median number of confirmed cases (2015–2017). Map was created using ArcGIS Desktop 10.6.1 software (ArcGIS Desktop^© 10.6.1 software, 2018, Redlands, CA). Data were obtained from information reported by the Panamanian Ministry of Health-National Leishmaniasis Surveillance Services to the PAHO as published in PAHO, leishmaniasis, epidemiological report in the Americas, Washington, PAHO, 2019. Available at: www.paho.org/leishmaniasis. PAHO = Pan American Health Organization. This figure appears in color at www.ajtmh.org.

Figure 2.

Hsp70-based Bayesian phylogenetic tree of Leishmania species obtained constructed under the GTR (Nst = 6) model with a gamma rate of four. The Bayesian consensus tree was searched by the program Mr. Bayes after 8,000,000 cycles of the Markov Chain Monte Carlo algorithm with a subsampling frequency of 1,000. Clade credibility values are shown as values at each clade node. Reference sequence codes appear in black color. All sequence codes obtained in this study are highlighted in red color. Reference sequences are abbreviated as follows: bra: Leishmania braziliensis; braO: Leishmania braziliensis outlier; guy: Leishmania guyanensis; lai: Leishmania lainsoni; nai: Leishmania naiffi; pan: Leishmania panamensis; per: Leishmania peruviana. This figure appears in color at www.ajtmh.org.