In Colombia, nine species of parasites of the genus Leishmania circulate in more than 20 sand fly species, putting at risk of contracting the disease approximately 60% of the population. The Federico Lleras Acosta Dermatological Center, a reference center in Colombia, has been treating patients with cutaneous and mucosal leishmaniasis for more than 15 years, identifying the infecting Leishmania species from different clinical samples, and recording systematically all the epidemiological and geographic information related to each diagnosed patient. With this valuable information, the objective of this work was to perform a long term and large-scale study, aiming to identify the Leishmania species circulating in Colombia from clinical samples from 1999 to 2016, and to assess their current and potential spatial distribution. In all, four Leishmania species were identified in 688 samples from 183 municipalities distributed in 28 of the 32 departments of the country, and 387 records were georeferenced, from 20 departments. The most widespread species was L . (V . ) braziliensis , showing new collection records, and the species related to areas with highest leishmaniasis transmission was L . (V . ) panamensis . Ecological niche models were built for the three species that had more than 20 georeferenced records, showing a potential distribution for L . (V . ) braziliensis on 42% of the national territory mainly in the interandean valleys, and the Orinoquia and Amazon regions. Leishmania (V . ) guyanensis potential distribution covers 36% of Colombia continental territory with a spatial distribution similar to that of L . (V . ) braziliensis . There was a marked tendency of L . (V . ) panamensis to be distributed in the northwest of the country occupying 35% of the national area and mainly in areas of transformed ecosystems. Species were identified in patients from areas where the occurrence of cases was unprecedented, which suggests that the distribution of Leishmania may be greater than currently known. To improve the predictive capacity of the models, we suggest incorporating, in future studies, Leishmania samples from vectors and reservoirs that have a greater dependence on environmental variables. Our results are an important tool for health systems because they allow potential areas of transmission and information gaps to be identified.

Different Leishmania species cause distinct clinical manifestations of the infectious disease leishmaniasis. It is fundamentally important to understand the mechanisms governing the interaction between Leishmania and its host cell. Little is known about this interaction between Leishmania (Viannia) braziliensis and human macrophages. In this study, we aimed to identify differential gene expression between non-infected and L. (V) braziliensis-infected U937-derived macrophages. We deployed a whole human transcriptome microarray analysis using 72 hours post-infection samples and compared those samples with their non-infected counterparts. We found that 218 genes were differentially expressed between infected and non-infected macrophages. A total of 71.6% of these genes were down-regulated in the infected macrophages. Functional enrichment analyses identified the steroid and sterol/cholesterol biosynthetic processes between regulatory networks down-regulated in infected macrophages. RT-qPCR further confirmed this down-regulation in genes belonging to these pathways. These findings contrast with those from studies involving other Leishmania species at earlier infection stages, where gene up-regulation for this metabolic pathway has been reported. Sterol biosynthesis could be an important biological process associated with the expression profile of macrophages infected by L. (V.) braziliensis. Differential transcriptional results suggest a negative regulation of the genetic regulatory network involved in cholesterol biosynthesis.

The discrimination of Leishmania species from patient samples has epidemiological and clinical relevance. In this study, different gene target PCR-restriction fragment length polymorphism (RFLP) protocols were evaluated for their robustness as Leishmania species discriminators in 61 patients with cutaneous leishmaniasis. We modified the hsp70-PCR-RFLP protocol and found it to be the most reliable protocol for species identification.H uman infections by Leishmania spp. produce a pleiomorphic syndrome in which symptomatology depends on the parasite species and the immunological stage of the host. The symptoms range from completely asymptomatic to cutaneous, mucocutaneous, and visceral (1). Several authors have reported differences in the treatment outcomes linked to the parasite species (2-5). Furthermore, mucocutaneous leishmaniasis (MCL) is a belated complication associated with specific parasite species (3, 6) most commonly occurring in infections caused by the Leishmania (Viannia) subgenus. American cutaneous leishmaniasis (ACL) cases are usually the result of infections produced by this subgenus, and species identification is useful for treatment and prognosis. Molecular techniques may become a routine way to confirm suspected cases of ACL (7-10); the present study describes the best PCR-restriction fragment length polymorphism (RFLP) gene target for determining the species of Leishmania present in clinical samples from ACL lesions in a set of Colombian patients.The study was approved by the boards of ethical conduct of the Hospital Militar Central-Bogota-Colombia (HOMIC) and Centro Dermatologico Federico Lleras Acosta Bogota-Colombia (CDFLL) in accordance with national (resolution 008430 of the Colombian Health Ministry) and international (Declaration of Helsinki and amendments, World Medical Association, South Korea, 2008) guidelines. DNA was extracted from skin biopsy specimens from the internal border of the lesions from 42 adult patients with a clinical diagnosis of ACL. The diagnosis was confirmed microscopically in 35 patients and by PCR detection of the parasite in 7 patients. All patients voluntarily participated in the study and signed an informed consent.The CDFLL biobank provided 19 Giemsa-stained slide smears from cutaneous lesions. In 17 of them, the presence of Leishmania sp. amastigotes was microscopically confirmed, and in 2 smears, the detection of the parasite was established by PCR. DNA was recovered from the Giemsa-stained smears.All PCRs performed included DNA from 2 negative-control patients from CDFLL (with confirmed diagnoses of sporotrichosis and ecthyma gangrenosum) and from three healthy volunteers. The entire group of patients had been infected within the Colombian borders.We selected genes and sequences previously reported to be useful markers for species identification by PCR-RFLP of Leishmania species for further evaluation. We analyzed zinc-metalloprotease (gp63) (11), spliced leader (SL) (12), cysteine protease B1 (cpb) (13), and heat shock protein 70 (hsp70) (14, 15) for the...

We consider PCR-miniexon as a diagnostic method of first choice for mucocutaneous leishmaniasis due to its excellent diagnostic performance and its ability to discriminate between Leishmania and Viannia subgenera as well as between species belonging to Leishmania subgenus.