Genomic Analysis of Colombian Leishmania panamensis strains with different level of virulence

Abstract

The establishment of Leishmania infection in mammalian hosts and the subsequent manifestation of clinical symptoms require internalization into macrophages, immune evasion and parasite survival and replication. Although many of the genes involved in these processes have been described, the genetic and genomic variability associated to differences in virulence is largely unknown. Here we present the genomic variation of four Leishmania (Viannia) panamensis strains exhibiting different levels of virulence in BALB/c mice and its application to predict novel genes related to virulence. De novo DNA sequencing and assembly of the most virulent strain allowed comparative genomics analysis with sequenced L. (Viannia) panamensis and L. (Viannia) braziliensis strains, and showed important variations at intra and interspecific levels. Moreover, the mutation detection and a CNV search revealed both base and structural genomic variation within the species. Interestingly, we found differences in the copy number and protein diversity of some genes previously related to virulence. Several machine-learning approaches were applied to combine previous knowledge with features derived from genomic variation and predict a curated set of 66 novel genes related to virulence. These genes can be prioritized for validation experiments and could potentially become promising drug and immune targets for the development of novel prophylactic and therapeutic interventions.

Intraspecific Genomic Divergence and Minor Structural Variations in Leishmania (Viannia) panamensis

“…Another interesting finding was associated with the 225 SNPs annotated, shared between the three clades analyzed ( Figure 5A,B. Table S4), most of them previously described in the literature and involved in the different biological processes of the parasite [27,68].…”

“…Despite these findings, we analyzed the genes with CNVs and observed that 44 of them were shared among all the genomes. Interestingly, some of them have been previously identified by other authors and include genes involved in parasite survival and virulence [27,28], which indicate their importance during the adaptation to human hosts. Additionally, the ontology enrichment analysis revealed that most genes with CNVs were involved in cellular ion zinc homeostasis in many clinical isolates; some of these genes were previously identified in L. infantum [63], as reported by a previous study, wherein the authors have described the need of these genes in the parasite to maintain the intracellular concentrations of zinc and avoid the toxic effects of this metal.…”

“…The paired-end Illumina reads of 19 Colombian clinical isolates and of the three Panamanian genomes (data downloaded from DDBJ/ENA/GenBank (http://www.ebi.ac.uk/ena) and TriTrypDB (http: //tritrypdb.org) databases) were mapped to the reference UA946 L. panamensis genome [27] (this genome was used as reference due to its high quality of assembly and annotation as reported elsewhere [27]) and assembled with the SMALT program (version 0.7.4) (www.sanger.ac.uk/resources/software/smalt/). The mapping involved the following parameters: exhaustive search option (−x and −y of 0.8), a reference hash index of 13 bases, and a sliding step of 3.…”

“…These analyses were carried out in SplitsTree5 [36], using the neighbor-net method. We included as reference sequence the UA946 (LpanUA) sequence from Urrea et al [27]. Additionally, we used as outgroup the L. guyanensis genome assembly from DDBJ/ENA/GenBank database (http://www.ebi.ac.uk/ena) under Run accession SRR8179913 (Lguy_SRR8179913).…”

“…However, despite the number of studies reported to date, only few studies using the above-mentioned approaches have focused on evaluating such genomic variability in the clinical isolates of Leishmania species from the subgenus Viannia, such as the published study by de Figueiredo de Sá et al who highlighted the high genetic diversity of L. braziliensis within restricted or adjacent areas [26], the study published by Urrea et al who evaluated the genomic variations of four L. panamensis strains exhibiting different levels of virulence [27], and the recently published study by Restrepo et al who described the genetic variations occurred in two L. panamensis clinical samples [28].…”