Novel detection of Leishmania RNA virus-1 (LRV-1) in clinical isolates of Leishmania Viannia panamensis.

American tegumentary leishmaniasis comprises a discrete set of clinical presentations endemic to Latin America. Leishmania RNA virus-1 (LRV-1) is a double-stranded RNA virus identified in 20­25% of the Leishmania Viannia braziliensis and L. V. guyanensis, however not in L. V. panamensis. This is the first report of LRV-1 in L. V. panamensis and its associations with clinical phenotypes of ATL. Unique surplus discard clinical isolates of L. V. panamensis were identified from the Public Health Ontario Laboratory (PHOL) and the Leishmania Clinic of the Instituto de Medicina Tropical 'Alexander von Humboldt' between 2012 and 2019 and screened for LRV-1 by real-time polymerase chain reaction. Patient isolates were stratified according to clinical phenotype. Of 30 patients with L. V. panamensis, 14 (47%) and 16 (53%) patients had severe and non-severe ATL, respectively. Five (36%) of 14 severe cases and 2 (12%) of 16 non-severe cases were positive for LRV-1, respectively. No differences in sex were observed for clinical phenotype and LRV-1 status. Although an association between LRV-1 status and clinical phenotype was not demonstrated, this is the first description of the novel detection of LRV-1 in L. V. panamensis, a species that has been documented predominantly in Central America.

A fine mapping of single nucleotide variants and haplotype analysis of IL13 gene in patients with Leishmania guyanensis-cutaneous leishmaniasis and plasma cytokines IL-4, IL-5, and IL-13.

Introduction: Leishmaniasis continues to pose a substantial health burden in 97 countries worldwide. The progression and outcome of Leishmania infection are influenced by various factors, including the cytokine milieu, the skin microbiota at the infection site, the specific Leishmania species involved, the genetic background of the host, and the parasite load. In endemic regions to leishmaniasis, only a fraction of individuals infected actually develops the disease. Overexpression of IL-13 in naturally resistant C57BL/6 mice renders them susceptible to L. major infection. Haplotypes constructed from several single nucleotide variant (SNV) along a chromosome fragment may provide insight into any SNV near the fragment that may be genuinely associated with a phenotype in genetic association studies. Methods: We investigated nine SNVs (SNV1rs1881457A>C, SNV2rs1295687C>G, SNV3rs2069744C>T, SNV4rs2069747C>T, SNV5rs20541A>G, SNV6rs1295685A>G, SNV7rs848A>C, SNV8rs2069750G >C, and SNV9rs847T>C) spanning the entire IL13 gene in patients with L. guyanensis cutaneous leishmaniasis (Lg-CL). Results: Our analysis did not reveal any significant association between the SNVs and susceptibility/protection against Lg-CL development. However, haplotype analysis, excluding SNV4rs2069747 and SNV8rs2069750 due to low minor allele frequency, revealed that carriers of the haplotype CCCTAAC had a 93% reduced likelihood developing Lg-CL. Similarly, the haplotypes ACCCGCT (ORadj=0.02 [95% CI 0.00-0.07]; p-value, 6.0×10-19) and AGCTAAC (ORadj=0.00[95% CI 0.00-0.00]; p-value 2.7×10-12) appeared to provide protection against the development of Lg-CL. Conversely, carriers of haplotype ACCTGCC have 190% increased likelihood of developing Lg-CL (ORadj=2.9 [95%CI 1.68-5.2]; p-value, 2.5×10-6). Similarly, haplotype ACCCAAT (ORadj=2.7 [95%CI 1.5-4.7]; p-value, 3.2×10-5) and haplotype AGCCGCC are associated with susceptibility to the development of Lg-CL (ORadj=1.7[95%CI 1.04-2.8]; p-value, 0.01). In our investigation, we also found a correlation between the genotypes of rs2069744, rs20541, rs1295685, rs847, and rs848 and plasma IL-5 levels among Lg-Cl patients. Furthermore, rs20541 showed a correlation with plasma IL-13 levels among Lg-Cl patients, while rs2069744 and rs848 showed a correlation with plasma IL-4 levels among the same group. Conclusions: Overall, our study identifies three haplotypes of IL13 associated with resistance to disease development and three haplotypes linked to susceptibility. These findings suggest the possibility of a variant outside the gene region that may contribute, in conjunction with other genes, to differences in susceptibility and partially to the pathology.

Variants of CARD8 in Leishmania guyanensis-cutaneous leishmaniasis and influence of the variants genotypes on circulating plasma cytokines IL-1ß, TNFα and IL-8.

Nucleotide-binding oligomerization domain, leucine-rich repeat-containing protein family (NLR) are intracellular pathogen recognition receptors mediating innate immunity, releasing proinflammatory cytokines IL-1ß and IL-18, and promoting pyroptotic cell death, upon sensing pathogenic or endogenous danger signals. In animal models, NLRP3 inflammasome has a dual role, pathogenic or protective in Leishmania-infection, depending on the Leishmania species and mice strain. Caspase recruitment containing domain 8 (CARD8) is a negative regulator of NLRP3 inflammasome and also an inhibitor of transcription factor NFĸB, a major transcription factor of proinflammatory cytokines. We investigated whether single nucleotide variants in CARD8 may partially explain why only a proportion of individuals coming from the same area of endemicity of leishmaniasis develop cutaneous leishmaniasis caused by Leishmania guyanensis. We genotyped four single nucleotide variants of the CARD8 gene by direct nucleotide sequencing in 1741 individuals from an endemic area of leishmaniasis, constituting 850 patients with CL and 891 healthy controls. The frequencies of the genotypes of the variants rs2288877 T>C, rs73944113 C>T, and rs2043211 A>T are similar among the patients with CL and HC, while the variant rs2288876 A>G) reveals an excess of the genotype AA among the patients with CL (44%) compared to 37% in the HC group. Allele A of the variant rs2288876 A>G) is associated with susceptibility to CL (OR = 1.2 [95%CI 1.03-1.4]; P = 0.01). Haplotype analysis showed that individuals harboring the haplotype CCAA have 280% odds of developing CL caused by L. guyanensis (OR = 3.8 [95% CI 2.0-7.7]; p = 0.00004). The variants rs2288877 T>C and rs2288876 A>G correlate with the plasma level of IL-8. Spearman correlation showed a significant positive correlation between the rs2288876 A>G allele A and the level of IL-8 (ρ = 0.22; p = 0.0002). CARD8 may partially contribute to the development of CL caused by L. guyanensis.

Detection of Leishmania RNA Virus 1 in Leishmania (Viannia) panamensis Isolates, Panama.

We detected Leishmania RNA virus 1 (LRV1) in 11 isolates of Leishmania (Viannia) panamensis collected during 2014-2019 from patients from different geographic areas in Panama. The distribution suggested a spread of LRV1 in L. (V.) panamensis parasites. We found no association between LRV1 and an increase in clinical pathology.

Diversity and geographical distribution of Leishmania species and the emergence of Leishmania (Leishmania) infantum and L. (Viannia) panamensis in Central-Western Venezuela.

Transmission of cutaneous leishmaniasis in Venezuela reveals diverse and changing epidemiological landscapes, as well as a spectrum of clinical phenotypes presumed to be linked to a variety of Leishmania species. Central-western Venezuela constitutes one of the highest endemic epicenters in the country, and updated molecular epidemiological information is still lacking. Therefore, in this study we aimed to characterize the landscape of circulating Leishmania species across central-western Venezuela through the last two decades, performed comparisons of haplotype and nucleotide diversity, and built a geospatial map of parasite species distribution. A total of 120 clinical samples were collected from patients across the cutaneous disease spectrum, retrieving parasitic DNA, and further characterizing by PCR and sequencing of the HSP70 gene fragment. This data was later collated with further genetic, geospatial and epidemiological analyses. A peculiar pattern of species occurrence including Leishmania (Leishmania) amazonensis (77.63% N=59), Leishmania (Leishmania) infantum (14.47% N=11), Leishmania (Viannia) panamensis (5.26% N=4) and Leishmania (Viannia) braziliensis (2.63% N=2) was revealed, also highlighting a very low genetic diversity amongst all analyzed sequences. Geographical distribution showed that most cases are widely distributed across the greater urban-sub urban area of the Irribaren municipality. L.(L.) amazonensis appears to be widely dispersed throughout Lara state. Statistical analyses failed to reveal significance for any comparisons, leading to conclude a lack of association between the infective Leishmania species and clinical phenotypes. To the best of our knowledge, this is an unprecedented study which addresses comprehensively the geographical distribution of Leishmania species in central-western Venezuela throughout the last two decades, and the first to incriminate L. (L.) infantum as an etiologic agent of cutaneous leishmaniasis in this region. Our findings support that Leishmania endemism in central-western Venezuela is caused mainly by L.(L.) amazonensis. Future studies are needed to unveil additional details on the ecological intricacies and transmission aspects of leishmaniasis (i.e. sampling phlebotomines and mammals) and to adopt adequate public health prevention and control strategies and mitigate disease impact in this endemic region.

Raising the suspicion of a non-autochthonous infection: identification of Leishmania guyanensis from Costa Rica exhibits a Leishmaniavirus related to Brazilian north-east and French Guiana viral genotypes.

BACKGROUND: Costa Rica has a history of neglecting prevention, control and research of leishmaniasis, including limited understanding on Leishmania species causing human disease across the country and a complete lack of knowledge on the Leishmania RNA virus, described as a factor linked to the worsening and metastasis of leishmanial lesions. OBJECTIVES: The aim of this work was to describe a case of cutaneous leishmaniasis by Leishmania (Viannia) guyanensis, bearing infection with Leishmaniavirus 1 (LRV1) in Costa Rica, raising the suspicion of imported parasites in the region. METHODS: The Leishmania strain was previously identified by routine hsp70 polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) in Costa Rica and subsequently characterised by isoenzyme electrophoresis and Sanger sequencing in Brazil. Screening for LRV1 was conducted with a dual RT-PCR approach and sequencing of the fragment obtained. FINDINGS: Since 2016 Costa Rica performs Leishmania isolation and typing as part of its epidemiological surveillance activities. Amongst 113 strains typed until 2019, only one was characterised as a L. (V.) guyanensis, corresponding to the first confirmed report of this species in the country. Interestingly, the same strain tested positive for LRV1. Sequencing of the viral orf1 and 2, clustered this sample with other LRV1 genotypes of South American origin, from the Northeast of Brazil and French Guiana. MAIN CONCLUSION: The unique characteristics of this finding raised the suspicion that it was not an autochthonous strain. Notwithstanding its presumed origin, this report points to the occurrence of said endosymbiont in Central American Leishmania strains. The possibility of its local dispersion represents one more challenge faced by regional health authorities in preventing and controlling leishmaniasis.

Sequencing of hsp70 for discernment of species from the Leishmania (Viannia) guyanensis complex from endemic areas in Colombia.

BACKGROUND: Colombia is ranked very high among countries with the highest numbers of endemic Leishmania species (n = 9) causing human disease. Although much effort has been devoted to generating simple and specific tools for Leishmania species identification, challenges remain in the discrimination of species belonging to the Leishmania (Viannia) guyanensis complex: L. (V.) guyanensis and L. (V.) panamensis. METHODS: A set of seven reference strains of species belonging to the L. (Leishmania) and L. (Viannia) subgenera, clinical strains from human cases of cutaneous leishmaniasis (CL; n = 26) and samples collected from sylvatic mammals and sand flies (n = 7) from endemic areas in Colombia were analyzed in this study. The heat-shock protein 70 gene (hsp70) was amplified by PCR from DNA extracted from logarithmic-phase promastigotes or tissue samples, and the PCR products were sequenced. Sequence alignment was performed against a set of previously published and curated sequences, and phylogenetic analysis based on the maximum-likelihood and Bayesian inference approaches was conducted. Haplotype diversity among strains and species of the L. (V.) guyanensis complex was explored using a median-joining network. RESULTS: Sequencing of the hsp70 gene for L. (Viannia) spp. typing was comparable to species identification using isoenzyme electrophoresis or monoclonal antibodies. Complete species matching was found, except for one sylvatic sample with an identity yet unsolved. Among the L. (V.) panamensis clinical strains, two distinctive phylogenetic clusters were found to correlate with two different zymodemes: L. (V.) panamensis Z2.2 and Z2.3. Analysis of samples from sylvatic environments identified novel records of naturally infected wild mammal and sand fly species. CONCLUSIONS: Our results support the adequacy of hsp70 gene sequencing as a single-locus approach for discrimination of L. (Viannia) spp., as well as for exploring the genetic diversity within the L. (V.) guyanensis complex.

Genomic diversity and genetic variation of Leishmania panamensis within its endemic range.

Species belonging to the Leishmania (Viannia) subgenus are important causative agents of cutaneous and mucocutaneous leishmaniasis in Central and South America. These parasites possess several distinctive biological features that are influenced by their genetics, population structure, and genome instability. To date, several studies have revealed varying degrees of genetic diversity within Leishmania species. Particularly, in species of the L. (Viannia) subgenus, a generalized high intraspecific genetic diversity has been reported, although, conflicting conclusions have been drawn using different molecular techniques. Despite being the most common Leishmania species circulating in Panama and Colombia, few studies have analyzed clinical samples of Leishmania panamensis using whole-genome sequencing, and their restricted number of samples has limited the information they can provide to understand the population structure of L. panamensis. Here, we used next generation sequencing (NGS) to explore the genetic diversity of L. panamensis within its endemic range, analyzing data from 43 isolates of Colombian and Panamanian origin. Our results show the occurrence of three well-defined geographically correlated groups, and suggests the possible occurrence of additional phylogeographic groups. Furthermore, these results support the existence of a mixed mode of reproduction in L. panamensis, with varying frequencies of events of genetic recombination occurring primarily within subpopulations of closely related strains. This study offers important insights into the population genetics and reproduction mode of L. panamensis, paving the way to better understand their population structure and the emergence and maintenance of key eco-epidemiological traits.

IL-23R Variant rs11805303 Is Associated With Susceptibility to the Development of Cutaneous Leishmaniasis in Leishmania guyanensis-Infected Individuals.

BACKGROUND: Emerging evidence suggests that the interleukin (IL) 17/ IL-23 axis may play a role in the pathogenesis of leishmaniasis. Our aim was to investigate whether the IL-23R variant rs11805303 is a risk factor for the development of cutaneous leishmaniasis (CL) in Leishmania guyanensis-infected individuals. METHODS: We genotyped by polymerase chain reaction-restriction fragment length polymorphism the rs11805303 C/T in 828 patients with CL and 806 healthy individuals. Plasma tumor necrosis factor-α, IL-6, interferon-γ, IL-1ß, and IL-17 were measured with the Bioplex assay. RESULTS: The distribution of the genotypes differed between patients with CL and healthy controls with a common odds ratio of 1.78 (P = 2.2 × 10-11) for the disease-associated T allele. Leishmania guyanensis-infected individuals homozygous for the T allele show a 200% increased risk of progressing to disease development, with a 95% confidence interval ranging from 81% to 400% (P = 9.9 × 10-6) in comparison to individuals homozygous for the C allele. Males homozygous for the T allele have higher plasma levels of IL-17 compared with heterozygous or homozygous CC individuals. CONCLUSIONS: The present association of the IL-23R variant rs11805303 with the development of CL suggests that the IL-17/IL-23 axis may play an important role in the pathogenesis of CL.

Filling the gaps in Leishmania naiffi and Leishmania guyanensis genome plasticity.

Insufficient and irregular data reports on Leishmaniasis, issuing from the developing world, have left much to be desired in terms of understanding the molecular signatures producing distinct infectious phenotypes of the disease. Herein, we report on the complete genome sequencing of Leishmania naiffi and Leishmania guyanensis, sampled from patients in regions of Colombia and Venezuela. In this study, the isolates of cutaneous lesions from both species presented limited structural variation at the chromosomal level, low gene copy number variation, and high genetic heterogeneity. We compared these sequences to the reference genomes hitherto related from Brazil and French Guyana. Although of the same species, we note a consequential genomic disparity between the Venezuelan and French Guyanese isolates of L. guyanensis. Although less significant on the global schema of cutaneous and mucosal disease, such genomic studies of L. naiffi and L. guyanensis substantiate the gaps in understanding of the molecular architecture and multivariate clinical pictures of Leishmaniasis, on an international scale.

Comparison of Whole Genome Sequencing versus Standard Molecular Diagnostics for Species Identification in the Leishmania Viannia Subgenus.

The prognosis and treatment of New World tegumentary leishmaniasis is dependent on the infecting species, yet such species identification in the Leishmania Viannia subgenus poses a diagnostic challenge. Currently, speciation relies on standard molecular techniques such as restriction fragment length polymorphism (RFLP) analysis, and Sanger sequencing (SS). Whole-genome sequencing (WGS) is a robust and increasingly cost-efficient tool that may improve Leishmania species identification. We evaluated WGS versus standard RFLP-SS for species identification in three reference and five clinical strains of Leishmania Viannia spp. Internal transcribed spacer1 (its1), cysteine proteinase b (cpb), and heat shock protein 70 (hsp70) polymerase chain reaction-restriction fragment length polymorphism (RFLP) was performed, followed by SS of the its2, cpb, hsp70, and mannose phosphate isomerase (mpi) loci. After de novo assembly, sequences were mapped, and homology compared with both reference strains and reference genomes on National Center for Biotechnology Information. All American Type Culture Collection strains were confirmed to be single-species of L. V. braziliensis, L. V. guyanensis, or L. V. panamensis by WGS. Conversely, RFLP-SS was able to definitively identify one of three isolates to the species level. Clinical samples were identified as either single-species (N = 3), mixed (N = 1), or hybrid (N = 1) infections by WGS, while standard molecular diagnosis required multi-target composite analysis for identification due to loci-dependent results by RFLP-SS. We have corroborated the utility of WGS as a diagnostic tool to speciate members of the L. Viannia subgenus and to discriminate between mixed and hybrid infections. WGS is a potentially useful complement to multistaged RFLP-SS for species identification in Leishmania infections.

The mutation G133D on Leishmania guyanensis AQP1 is highly destabilizing as revealed by molecular modeling and hypo-osmotic shock assay.

The Leishmania aquaglyceroporin 1 (AQP1) plays an important role in osmoregulation and antimony (Sb) uptake, being determinant for resistance to antimony. We have previously demonstrated that G133D mutation on L. guyanensis AQP1 (LgAQP1) leads to reduced Sb uptake. Here, we investigated the effects of G133D mutation on LgAQP1 structure, associated with Sb uptake and alterations in osmoregulation capacity. High confidence molecular models of wild-type LgAQP1 as well as the LgAQP1::G133D mutant were constructed and optimized via comparative homology modeling. Computational methods from the mCSM platform were used to evaluate the effects on protein stability and on its ability to bind to glycerol. Functional validation of the disruptive effect of the mutation on LgAQP1 was done by challenging the parasites with hypo-osmotic chock. Glycine 133 is on transmembrane helix 3, buried in the membrane in both open and closed conformation. G133D mutation was predicted to be highly destabilizing, as it alters the helical bundling arrangement in order to accommodate the aspartic acid side chain. The shift in helices also resulted in fewer favorable contacts with glycerol in the channel, which would explain the reduced affinity for similar small molecules as SbO3. Under hypo-osmotic condition, L. guyanensis AQP1G133D presented a 3-fold increase in cellular volume and pronounced delay to recover osmosis homeostasis when compared to the wild-type, a profile that was enhanced in LgAQP1-/- mutants. In conclusion, G133D is a highly disruptive mutation that will destabilize the monomer, compromise tetramer formation and alter pore conformation, leading to reduced Sb uptake and deficient osmoregulation.

Diagnosis and identification of Leishmania species in patients with cutaneous leishmaniasis in the state of Roraima, Brazil's Amazon Region.

BACKGROUND: Cutaneous leishmaniasis (CL) is an endemic disease in Brazil that is highly prevalent in the northern region of the country. Although there is a continuous and growing number of cases registered in the state of Roraima, there is limited information regarding the species of Leishmania that affect the human population. In this study, we aimed to characterize which Leishmania species cause human disease in those presenting with cutaneous leishmaniasis in endemic areas of the State of Roraima. METHODS: We conducted a prospective surveillance study between 2016 to 2018 in health centers located in the State of Roraima, Brazil. Participants with clinical suspicion of CL were enrolled and provided lesion samples for parasitological detection by microscopy. A subset of the samples was tested by polymerase chain reaction and sequencing of the internal transcribed spacer 1 (ITS-1 PCR) for molecular species identification. RESULTS: A total of 262 participants were enrolled in this study. Of those, 129 (49.27%) were positive by parasitological examination. Most positive subjects (81.58%) were male, and most cases presented a single lesion (80.26%). ITS-1 PCR and sequencing on a subset of 76 samples allowed us to detect nine different species of Leishmania: L. (V.) braziliensis, L (V.) panamensis, L. (V.) guyanensis, L. (V.) naiffi, L. (V.) shawi, L.(V.) utingensis, L. (V.) lindenbergi, L. (L.) amazonensis and L. (L.) mexicana. CONCLUSIONS: Our study provides the first assessment of circulating species of Leishmania in the State of Roraima, Brazil, and shows the high diversity in this region. This study opens the path for further research on the transmission of leishmaniasis in the northernmost Brazilian State including vector and reservoir surveillance as well as for intensification of investigation and control activities against CL in the region.

First Report of Canine Infection by Leishmania (Viannia) guyanensis in the Brazilian Amazon.

The American cutaneous (CL) and visceral leishmaniasis (VL) are zooanthroponoses transmitted by sand flies. Brazil records thousands of human leishmaniasis cases annually. Dogs are reservoirs of Leishmania infantum, which causes VL, but their role in the transmission cycle of CL is debatable. Wild mammals are considered reservoirs of the aetiological agents of CL (Leishmania spp.). OBJECTIVE: To describe the aetiology of leishmaniasis in dogs in an endemic area for CL and VL in the Amazon, Brazil. METHODS: Clinical evaluation and blood collection of 40 dogs from the villages Ubim (20) and Socorro (20), city of Tomé-Açu, state of Pará, were carried out. The DNA extracted from the blood was used for PCR with Leishmania-specific primers targeting the hsp70-234 gene sequence. Products were sequenced (ABI3500XL), and the sequences were aligned, edited (BioEdit), and analyzed (Blastn). RESULTS: Of the 34 amplified samples, 21 were sequenced, namely Leishmania infantum (12), L. guyanensis (5), L. braziliensis (3), and Leishmania sp. (01). CONCLUSION: Given the diversity of circulating pathogens, elucidation of the role of the dog in the Leishmania spp. cycle in Amazonian villages is imperative to the surveillance of CL in the region. We present the first report in Brazil, confirmed by sequencing, of canine infection by L. guyanensis, a species highly resistant to treatment in humans, with the drug of first choice (Glucantime®).

Identification of Leishmania species in patients derived to the National Institute of Health, Peru. / Identificación de especies de Leishmania en pacientes derivados al Instituto Nacional de Salud del Perú.

In Peru, leishmaniasis is a metaxenic disease that represents a serious public health problem, due to its wide distribution and the number of people in danger of contracting the disease, being the vulnerable population mainly those with low economic resources. The study was conducted from patients who were derived to Peru's National Institute of Health between 2006 and 2011 so that the specialized diagnosis could be carried out. The identification of the species of infectious Leishmania was developed through the analysis of the High-Resolution Melting Analysis obtained from the genomic DNA of promastigotes and amastigotes, which allows to identify the species of Leishmania (Viannia) braziliensis, Leishmania (V.) guyanensis, Leishmania (V.) peruviana as more prevalent, in addition to Leishmania (V.) lainsoni and Leishmania (L.) amazonensis.

En el Perú, la leishmaniasis es una enfermedad metaxénica que representa un serio problema de salud pública, debido a su amplia distribución y al número de personas en riesgo de contraer la enfermedad, siendo la población vulnerable principalmente las personas de bajos recursos económicos. El estudio se realizó a partir de pacientes que fueron derivados al Instituto Nacional de Salud entre el 2006 y el 2011 para que se les realizara el diagnóstico especializado. La identificación de la especie de Leishmania infectante se desarrolló mediante el análisis de las curvas de disociación (HRMA) obtenidas a partir del ADN genómico de promastigotes y amastigotes, lo que permitió identificar las especies de Leishmania (Viannia) braziliensis, Leishmania (V.) guyanensis, Leishmania (V.) peruviana como las más prevalentes, además de Leishmania (V.) lainsoni y Leishmania (L.) amazonensis.

Case Report: Successful Treatment with Miltefosine of Severe New World Mucosal Leishmaniasis Caused by Leishmania guyanensis.

An 88-year-old man with mutilating mucosal leishmaniasis (ML) involving septal perforation, with granulomas in the pharynx and larynx, was treated with oral miltefosine, 50 mg three times/day for 28 days. Miltefosine, an antineoplastic agent, is considered an alternative option for the treatment of ML, showing efficacies of 75-92% in Bolivia, Brazil, and Argentina. The patient denied having previous cutaneous (CL) leishmaniasis, and no CL lesions were recognized by physical examination. Parasites obtained from mucosal lesions were identified by cytochrome b gene sequencing as Leishmania guyanensis. Clinical cure was observed 2 months posttreatment, and no evidence of reactivation was observed in the 3-year follow-up. Adverse effects such as nausea, loss of appetite, and epigastric pain were experienced during treatment with miltefosine. There is a need for improved access to miltefosine in leishmaniasis-endemic areas of Latin America and a greater awareness of ML and its treatment among physicians working in endemic countries.

Identificación de especies de Leishmania en pacientes derivados al Instituto Nacional de Salud del Perú / Identification of Leishmania species in patients derived to the National Institute of Health, Peru

RESUMEN En el Perú, la leishmaniasis es una enfermedad metaxénica que representa un serio problema de salud pública, debido a su amplia distribución y al número de personas en riesgo de contraer la enfermedad, siendo la población vulnerable principalmente las personas de bajos recursos económicos. El estudio se realizó a partir de pacientes que fueron derivados al Instituto Nacional de Salud entre el 2006 y el 2011 para que se les realizara el diagnóstico especializado. La identificación de la especie de Leishmania infectante se desarrolló mediante el análisis de las curvas de disociación (HRMA) obtenidas a partir del ADN genómico de promastigotes y amastigotes, lo que permitió identificar las especies de Leishmania (Viannia) braziliensis, Leishmania (V.) guyanensis, Leishmania (V.) peruviana como las más prevalentes, además de Leishmania (V.) lainsoni y Leishmania (L.) amazonensis.

ABSTRACT In Peru, leishmaniasis is a metaxenic disease that represents a serious public health problem, due to its wide distribution and the number of people in danger of contracting the disease, being the vulnerable population mainly those with low economic resources. The study was conducted from patients who were derived to Peru's National Institute of Health between 2006 and 2011 so that the specialized diagnosis could be carried out. The identification of the species of infectious Leishmania was developed through the analysis of the High-Resolution Melting Analysis obtained from the genomic DNA of promastigotes and amastigotes, which allows to identify the species of Leishmania (Viannia) braziliensis, Leishmania (V.) guyanensis, Leishmania (V.) peruviana as more prevalent, in addition to Leishmania (V.) lainsoni and Leishmania (L.) amazonensis.

A Single Haplotype of IFNG Correlating With Low Circulating Levels of Interferon-γ Is Associated With Susceptibility to Cutaneous Leishmaniasis Caused by Leishmania guyanensis.

BACKGROUND: Interferon-γ (IFN-γ) plays an important role in the control of Leishmania infection. Blockade of IFN-γ signaling in mice increases lesion size and parasite load. In endemic areas of Leishmaniasis, only a fraction of the population develop the disease. This suggest that host genetics may play a role in this response. We investigated whether single nucleotide polymorphisms (SNPs) in IFNG may be associated with elevated or decrease risk in the development of cutaneous leishmaniasis (CL). METHODS: We assessed 9 SNP and cytosine-adenine (CA) repeats in IFNG by nucleotide sequencing in 647 patients with CL caused by Leishmania guyanensis and 629 controls. Circulating plasma IFN-γ levels were also assayed in 400 patients with CL and 400 controls. RESULTS: The rs2069705TT genotype is associated with elevated risk of developing CL compared with the rs2069705CC genotype (OR, 1.7; 95% CI, 1.3-2.4; P = .0008). There is a 70% chance that this genotype raises the risk of developing CL. In a dominant model, carriers of the rs2069705T allele compared with the rs2069705CC genotype showed a 50% (range, 20-100%) increased risk of developing CL (OR, 1.5; 95% CI, 1.2-2.0; P = .0004). Haplotype analysis showed 1 haplotype (H1) associated with low levels of IFN-γ presented an increased risk of 60% of developing CL (OR, 1.6; 95% CI, 1.3-1.9; P = 5 × 10-5) compared with non-H1. CONCLUSIONS: IFNG variant rs2069705 seems to be a genetic modifier of clinical outcome of Leishmania infection; individuals with the H1 haplotype, associated with low levels of IFN-γ, have a 60% risk of developing CL.