Abstract
Seven isolates from patients with American cutaneous leishmaniasis in the Amazon region of Brazil were phenotypically suggestive of Leishmania (Viannia) guyanensis/L. (V.) shawi hybrids. In this work, two molecular targets were employed to check the hybrid identity of the putative hybrids. Heat shock protein 70 (hsp70) gene sequences were analyzed by three different polymerase chain reaction (PCR) approaches, and two different patterns of inherited hsp70 alleles were found. Three isolates presented heterozygous L. (V.) guyanensis/L. (V.) shawi patterns, and four presented homozygous hsp70 patterns involving only L. (V.) shawi alleles. The amplicon sequences confirmed the RFLP patterns. The high-resolution melting method detected variant heterozygous and homozygous profiles. Single-nucleotide polymorphism genotyping/cleaved amplified polymorphic site analysis suggested a higher contribution from L. (V.) guyanensis in hsp70 heterozygous hybrids. Additionally, PCR-RFLP analysis targeting the enzyme mannose phosphate isomerase (mpi) gene indicated heterozygous and homozygous cleavage patterns for L. (V.) shawi and L. (V.) guyanensis, corroborating the hsp70 findings. In this communication, we present molecular findings based on partial informative regions of the coding sequences of hsp70 and mpi as markers confirming that some of the parasite strains from the Brazilian Amazon region are indeed hybrids between L. (V.) guyanensis and L. (V.) shawi.
Highlights
The genus Leishmania presents highly diverse nonsexual mechanisms for the generation of diversity
These strains included 21 reference strains of the main species present in the lower Amazon region, including four different reference strains of L. (V.) guyanensis, two of L. (V.) shawi shawi, two of L. (V.) shawi santarensis and the seven putative L. guyanensis /L. shawi hybrids that were isolated from the skin lesions of patients with localized cutaneous leishmaniasis
The hsp70 polymerase chain reaction (PCR) products obtained from the seven isolates generated two RFLP profiles, one that was characteristic of heterozygous alleles, while the other was characteristic of homozygous profile of the L. (V). shawi alleles (Figure 1A)
Summary
The genus Leishmania presents highly diverse nonsexual mechanisms for the generation of diversity These mechanisms include the occurrence of tandem repeat genes, gene amplification, gene duplication, mini-chromosome generation and mosaic aneuploidy (Delgado et al, 1997; Victoir and Dujardin, 2002; Dujardin et al, 2007; Sterkers et al, 2011). This high genome plasticity is achieved through both sexual-like and nonsexual characteristics, providing great complexity that is reflected in the wide geographical distribution of Leishmania spp., the diversity of its hosts and the complexity of illnesses associated with the parasites. Recent studies have shown that intraspecies hybrids are prone to hybridization events, while interspecies hybrids seem to be sterile (Inbar, et al, 2019)
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