Publications by authors named "Elisa Cupolillo"

77 Publications

In Vitro Susceptibility to Miltefosine of (syn.  ) Isolates from Different Geographical Areas in Brazil.

Microorganisms 2021 Jun 5;9(6). Epub 2021 Jun 5.

Departamento de Parasitologia, Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo 05508-000, Brazil.

Treatment of visceral leishmaniasis in Brazil still relies on meglumine antimoniate, with less than ideal efficacy and safety, making new therapeutic tools an urgent need. The oral drug miltefosine was assayed in a phase II clinical trial in Brazil with cure rates lower than previously demonstrated in India. The present study investigated the susceptibility to miltefosine in 73 Brazilian strains of from different geographic regions, using intracellular amastigote and promastigote assays. The EC for miltefosine of 13 of these strains evaluated in intracellular amastigotes varied between 1.41 and 4.57 μM. The EC of the 73 strains determined in promastigotes varied between 5.89 and 23.7 μM. No correlation between in vitro miltefosine susceptibility and the presence of the miltefosine sensitive locus was detected among the tested strains. The relatively low heterogeneity in miltefosine susceptibility observed for the 73 strains tested in this study suggests the absence of decreased susceptibility to miltefosine in Brazilian and does not exclude future clinical evaluation of miltefosine for VL treatment in Brazil.
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http://dx.doi.org/10.3390/microorganisms9061228DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8228039PMC
June 2021

The Maze Pathway of Coevolution: A Critical Review over the and Its Endosymbiotic History.

Genes (Basel) 2021 Apr 27;12(5). Epub 2021 Apr 27.

Research on Leishmaniasis Laboratory, Oswaldo Cruz Institute, FIOCRUZ, Rio de Janeiro 21040360, Brazil.

The description of the genus as the causative agent of leishmaniasis occurred in the modern age. However, evolutionary studies suggest that the origin of can be traced back to the Mesozoic era. Subsequently, during its evolutionary process, it achieved worldwide dispersion predating the breakup of the Gondwana supercontinent. It is assumed that this parasite evolved from monoxenic Trypanosomatidae. Phylogenetic studies locate dixenous in a well-supported clade, in the recently named subfamily Leishmaniinae, which also includes monoxenous trypanosomatids. Virus-like particles have been reported in many species of this family. To date, several species have been reported to be infected by RNA virus (LRV) and (LBV). Since the first descriptions of LRVs decades ago, differences in their genomic structures have been highlighted, leading to the designation of LRV1 in . () species and LRV2 in . () species. There are strong indications that viruses that infect spp. have the ability to enhance parasitic survival in humans as well as in experimental infections, through highly complex and specialized mechanisms. Phylogenetic analyses of these viruses have shown that their genomic differences correlate with the parasite species infected, suggesting a coevolutionary process. Herein, we will explore what has been described in the literature regarding the relationship between and endosymbiotic viruses and what is known about this association that could contribute to discussions about the worldwide dispersion of
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http://dx.doi.org/10.3390/genes12050657DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8146029PMC
April 2021

Colonization and genetic diversification processes of Leishmania infantum in the Americas.

Commun Biol 2021 01 29;4(1):139. Epub 2021 Jan 29.

Laboratório de Pesquisa em Leishmaniose, Instituto Oswaldo Cruz, FIOCRUZ, 21040-365, Rio de Janeiro, Brazil.

Leishmania infantum causes visceral leishmaniasis, a deadly vector-borne disease introduced to the Americas during the colonial era. This non-native trypanosomatid parasite has since established widespread transmission cycles using alternative vectors, and human infection has become a significant concern to public health, especially in Brazil. A multi-kilobase deletion was recently detected in Brazilian L. infantum genomes and is suggested to reduce susceptibility to the anti-leishmanial drug miltefosine. We show that deletion-carrying strains occur in at least 15 Brazilian states and describe diversity patterns suggesting that these derive from common ancestral mutants rather than from recurrent independent mutation events. We also show that the deleted locus and associated enzymatic activity is restored by hybridization with non-deletion type strains. Genetic exchange appears common in areas of secondary contact but also among closely related parasites. We examine demographic and ecological scenarios underlying this complex L. infantum population structure and discuss implications for disease control.
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http://dx.doi.org/10.1038/s42003-021-01658-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7846609PMC
January 2021

Comparison and clinical validation of qPCR assays targeting Leishmania 18S rDNA and HSP70 genes in patients with American Tegumentary Leishmaniasis.

PLoS Negl Trop Dis 2020 10 12;14(10):e0008750. Epub 2020 Oct 12.

Laboratório de Pesquisa em Leishmaniose, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz, Rio de Janeiro, Rio de Janeiro, Brazil.

Leishmaniasis is a worldwide neglected disease, encompassing asymptomatic infections and different clinical forms, such as American Tegumentary Leishmaniasis (ATL) which is part of the complex of diseases caused by protozoan parasites from Leishmania genus, transmitted by sand fly vectors. As a neglected disease, much effort is still needed in treatment and diagnosis. Currently, ATL diagnosis is mainly made by parasite detection by microscopy. The sensitivity of the method varies, and factors such as collection procedures interfere. Molecular approaches, specially based on Real Time PCR (qPCR) technique, has been widely used to detect Leishmania infection and to quantify parasite load, once it is a simple, rapid and sensitive methodology, capable to detect low parasite concentrations and less prone to variability. Although many studies have been already published addressing the use of this technique, an improvement on these methodologies, including an analytical validation, standardization and data association is demanded. Moreover, a proper validation by the assay by the use of clinical samples is still required. In this sense, the purpose of the present work is to compare the performance of qPCR using two commonly used targets (18S rDNA and HSP70) with an internal control (RNAse P) in multiplex reactions. Additionally, we validated reactions by assaying 88 samples from patients presenting different clinical forms of leishmaniasis (cutaneous, mucosal, recent and old lesions), representing the diversity found in Brazil's Amazon Region. Following the methodology proposed herein, the results indicate the use of both qPCR assays, 18S rDNA and HSP70, to achieve a very good net sensitivity (98.5%) and specificity (100%), performing simultaneous or sequential testing, respectively. With this approach, our main goal is to conclude the first step of a further multicenter study to propose the standardization of detection and quantification of Leishmania.
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http://dx.doi.org/10.1371/journal.pntd.0008750DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7581006PMC
October 2020

Ecological divergence and hybridization of Neotropical parasites.

Proc Natl Acad Sci U S A 2020 10 21;117(40):25159-25168. Epub 2020 Sep 21.

Department of Biomedical Sciences, Institute of Tropical Medicine, 2000 Antwerp, Belgium;

The tropical Andes are an important natural laboratory to understand speciation in many taxa. Here we examined the evolutionary history of parasites of the species complex based on whole-genome sequencing of 67 isolates from 47 localities in Peru. We first show the origin of Andean as a clade of near-clonal lineages that diverged from admixed Amazonian ancestors, accompanied by a significant reduction in genome diversity and large structural variations implicated in host-parasite interactions. Within the Andean species, patterns of population structure were strongly associated with biogeographical origin. Molecular clock and ecological niche modeling suggested that the history of diversification of the Andean lineages is limited to the Late Pleistocene and intimately associated with habitat contractions driven by climate change. These results suggest that changes in forestation over the past 150,000 y have influenced speciation and diversity of these Neotropical parasites. Second, genome-scale analyses provided evidence of meiotic-like recombination between Andean and Amazonian species, resulting in full-genome hybrids. The mitochondrial genome of these hybrids consisted of homogeneous uniparental maxicircles, but minicircles originated from both parental species. We further show that mitochondrial minicircles-but not maxicircles-show a similar evolutionary pattern to the nuclear genome, suggesting that compatibility between nuclear-encoded mitochondrial genes and minicircle-encoded guide RNA genes is essential to maintain efficient respiration. By comparing full nuclear and mitochondrial genome ancestries, our data expand our appreciation on the genetic consequences of diversification and hybridization in parasitic protozoa.
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http://dx.doi.org/10.1073/pnas.1920136117DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7547230PMC
October 2020

In-depth quantitative proteomics uncovers specie-specific metabolic programs in Leishmania (Viannia) species.

PLoS Negl Trop Dis 2020 08 17;14(8):e0008509. Epub 2020 Aug 17.

Laboratório de Pesquisa em Leishmanioses, Instituto Oswaldo Cruz, Fiocruz, Rio de Janeiro, RJ, Brazil.

Leishmania species are responsible for a broad spectrum of diseases, denominated Leishmaniasis, affecting over 12 million people worldwide. During the last decade, there have been impressive efforts for sequencing the genome of most of the pathogenic Leishmania spp. as well as hundreds of strains, but large-scale proteomics analyses did not follow these achievements and the Leishmania proteome remained mostly uncharacterized. Here, we report a comprehensive comparative study of the proteomes of strains representing L. braziliensis, L. panamensis and L. guyanensis species. Proteins extracted by SDS-mediated lysis were processed following the multi-enzyme digestion-filter aided sample preparation (FASP) procedure and analysed by high accuracy mass spectrometry. "Total Protein Approach" and "Proteomic Ruler" were applied for absolute quantification of proteins. Principal component analysis demonstrated very high reproducibility among biological replicates and a very clear differentiation of the three species. Our dataset comprises near 7000 proteins, representing the most complete Leishmania proteome yet known, and provides a comprehensive quantitative picture of the proteomes of the three species in terms of protein concentration and copy numbers. Analysis of the abundance of proteins from the major energy metabolic processes allow us to highlight remarkably differences among the species and suggest that these parasites depend on distinct energy substrates to obtain ATP. Whereas L. braziliensis relies the more on glycolysis, L. panamensis and L. guyanensis seem to depend mainly on mitochondrial respiration. These results were confirmed by biochemical assays showing opposite profiles for glucose uptake and O2 consumption in these species. In addition, we provide quantitative data about different membrane proteins, transporters, and lipids, all of which contribute for significant species-specific differences and provide rich substrate for explore new molecules for diagnosing purposes. Data are available via ProteomeXchange with identifier PXD017696.
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http://dx.doi.org/10.1371/journal.pntd.0008509DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7451982PMC
August 2020

Occurrence of multiple genotype infection caused by Leishmania infantum in naturally infected dogs.

PLoS Negl Trop Dis 2020 07 27;14(7):e0007986. Epub 2020 Jul 27.

Laboratório de Pesquisa em Leishmaniose, Instituto Oswaldo Cruz, Fiocruz, Rio de Janeiro, RJ, Brasil.

Genetic polymorphisms in natural Leishmania populations have been reported in endemic areas. Microsatellite typing is a useful tool to elucidate the genetic variability of parasite strains, due to its capability for high-resolution mapping of genomic targets. The present study employed multilocus microsatellite typing (MLMT) to explore the genotypic composition of Leishmania infantum in naturally infected dogs by genotyping parasites infecting different tissues with or without in vitro expansion. Eighty-six samples were collected from 46 animals in an endemic region of visceral leishmaniasis (VL). MLMT was performed for 38 spleen samples and 48 L. infantum cultures isolated from different tissues. Of the 86 samples, 23 were effectively genotyped by MLMT, identifying nine multilocus genotypes (MLG; referred to as MLG A-I). MLGs A, B and C were detected in more than one type of tissue and in more than one sample. Conversely, MLG D-I were uniquely detected in one sample each. The results showed that multiple genotype infections occur within a single host and tissue. Paired sample analysis revealed the presence of different MLMT alleles in 14 dogs, while the same MLG allele was present in 15 animals. STRUCTURE analysis demonstrated the presence of two populations; 13 samples displayed a similar admixture of both ancestral populations, and these were not assigned to any population. Only samples for which Q ≥ 0.70 after CLUMPP alignment were considered to be part of Population 1 (POP1) or Population 2 (POP2). POP2 comprised the majority of samples (n = 54) compared to POP1 (n = 19). This study presents evidence of multiple genotype infections (caused by L. infantum) in dogs in an area with high VL transmission. Further investigations must be undertaken to determine the effects of multiple infection on the host immune response and disease dynamics and treatment.
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http://dx.doi.org/10.1371/journal.pntd.0007986DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7410330PMC
July 2020

Trans-Atlantic Spill Over: Deconstructing the Ecological Adaptation of in the Americas.

Genes (Basel) 2019 12 19;11(1). Epub 2019 Dec 19.

Laboratory of Research on Leishmaniasis, Oswaldo Cruz Institute, FIOCRUZ, 21040-360 Rio de Janeiro, Brazil.

Pathogen fitness landscapes change when transmission cycles establish in non-native environments or spill over into new vectors and hosts. The introduction of in the Americas into the Neotropics during European colonization represents a unique case study to investigate the mechanisms of ecological adaptation of this important parasite. Defining the evolutionary trajectories that drive fitness in this new environment are of great public health importance as they will allow unique insight into pathways of host/pathogen co-evolution and their consequences for region-specific changes in disease manifestation. This review summarizes current knowledge on genetic and phenotypic diversity in the Americas and its possible role in the unique epidemiology of visceral leishmaniasis (VL) in the New World. We highlight the importance of appreciating adaptive molecular mechanisms in to understand the parasites' successful establishment on the continent.
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http://dx.doi.org/10.3390/genes11010004DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7017240PMC
December 2019

Premature deaths by visceral leishmaniasis in Brazil investigated through a cohort study: A challenging opportunity?

PLoS Negl Trop Dis 2019 12 19;13(12):e0007841. Epub 2019 Dec 19.

Pan-American Foot-and-Mouth Disease Center-PANAFTOSA/PAHO, Duque de Caxias, Rio de Janeiro, Brazil.

Background: Visceral Leishmaniasis (VL) is the most severe form of leishmaniasis because it can lead to death. In the Americas, 96% of cases are in Brazil, and despite efforts, the fatality rate has increased in the past years. We analyzed deaths associated to VL in Brazil and investigated the factors that could influence on the timeliness of fatal outcome with emphasis on time (tStoD).

Methodology: The registered deaths by VL were sourced from the Brazilian National Notification System from 2007-2014. Through a retrospective cohort study, univariate and multivariable Cox proportional hazards model analysis were performed and investigated the factors that could influence the time (tStoD). These factors were analyzed through survival models.

Results: Out of the 1,589 reported deaths, the median for onset of the symptoms and the case notification date (tStoN) is 25 days (10-61), and for date of case notification and death (tNotD) is 9 days (4-17). The time (tStoN) to event investigation for HIV non-infected individuals was 1.4 (1.16-1.68) greater than the HIV positive group. At the same time peri-urban and urban area were 0.83 (0.47-1.44) and 1.33 (1.16-1.52), respectively. The explorations revealed apparent differences between the time to event investigation (both for tStoN and tNotD) and the age at the onset of the symptoms. According to the tStoN the rate of notification is 1.73 times greater in patients under 5 years old at the onset of the clinical symptoms compared to older patients.

Conclusion: VL patients under 5 years old were diagnosed earlier and had shorter survival. It could mean that in younger population, although properly diagnosed, the fatality pattern might be related to the severity of the disease. Main host characteristics were evaluated, and age and co-infections seem to have an impact in the disease progression.
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http://dx.doi.org/10.1371/journal.pntd.0007841DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6922316PMC
December 2019

Pro-Cellular Exhaustion Markers are Associated with Splenic Microarchitecture Disorganization and Parasite Load in Dogs with Visceral Leishmaniasis.

Sci Rep 2019 09 10;9(1):12962. Epub 2019 Sep 10.

Laboratório de Pesquisa em Leishmaniose, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz, Rio de Janeiro, Brazil.

In canine visceral leishmaniasis (CVL), splenic white pulp (SWP) disorganization has been associated with disease progression, reduced cytokine and chemokine expression and failure to control the parasite load. This profile is compatible with the cellular exhaustion previously shown in human visceral leishmaniasis. The present study aimed to evaluate the in situ expression of cellular exhaustion markers and their relation to clinical signs, SWP disorganization and parasite load. Forty dogs naturally infected by Leishmania infantum were grouped according to levels of SWP organization and parasite load. SWP disorganization was associated with reductions in the periarteriolar lymphatic sheath and lymphoid follicles/mm and worsening of the disease. Apoptotic cells expressing CTLA-4 increased in dogs with disorganized SWP and a high parasite load. In the same group, PD-L1 and LAG-3 gene expression were reduced. A higher number of CD21TIM-3 B cells was detected in disorganized spleens than in organized spleens. Apoptosis is involved in periarteriolar lymphatic sheath reduction and lymphoid follicle atrophy and is associated with CTLA-4 cell reductions in the splenic tissue of dogs with visceral leishmaniasis (VL). Failure to control the parasite load was observed, suggesting that cell exhaustion followed by T and B cell apoptosis plays a role in the immunosuppression observed in CVL.
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http://dx.doi.org/10.1038/s41598-019-49344-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6736856PMC
September 2019

First report of Leishmania (Viannia) lindenbergi causing tegumentary leishmaniasis in the Brazilian western Amazon region.

Parasite 2019 23;26:30. Epub 2019 May 23.

Fundação Oswaldo Cruz, Instituto Oswaldo Cruz, IOC, Rio de Janeiro, Brazil.

Tegumentary Leishmaniasis (TL) in the Brazilian Amazon region is associated with several Leishmania species. In this report, we describe two cases of TL related to Leishmania lindenbergi occurring in different locations of Rondônia state. After clinical diagnosis, lesion samples were collected for parasitological diagnoses via direct microscopic visualization, parasite isolation, and PCR. PCR reactions were positive in both clinical samples. Parasite isolation was possible for both patients, and isolates were submitted to species identification by isoenzyme electrophoresis and DNA sequencing. This report is the first to describe human infections caused by L. lindenbergi since the initial description and record of human infection by this species in 2002.
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http://dx.doi.org/10.1051/parasite/2019030DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6532396PMC
August 2019

A novel multilocus sequence typing scheme identifying genetic diversity amongst Leishmania donovani isolates from a genetically homogeneous population in the Indian subcontinent.

Int J Parasitol 2019 06 18;49(7):555-567. Epub 2019 May 18.

Parasitology Department, Centre for Infectious Diseases and Microbiology Laboratory Services (CIDMLS), Institute of Clinical Pathology and Medical Research (ICPMR), Westmead Hospital, Westmead, Sydney, NSW, Australia; Molecular Mycology Research Laboratory, Centre for Infectious Diseases and Microbiology, Faculty of Medicine and Health, Sydney Medical School, Westmead Clinical School, Marie Bashir Institute for Emerging Infectious Diseases and Biosecurity, Westmead Hospital (Research and Education Network), The University of Sydney, Sydney, NSW, Australia. Electronic address:

In the Indian subcontinent, infection with Leishmania donovani can cause fatal visceral leishmaniasis. Genetic variation in L. donovani is believed to occur rapidly from environmental changes and through selective drug pressures, thereby allowing continued disease occurrence in this region. All previous molecular markers that are commonly in use multilocus microsatellite typing and multilocus sequence typing, were monomorphic in L. donovani originating from the Indian subcontinent (with only a few exceptions) and hence are not suitable for this region. An multilocus sequence typing scheme consisting of a new set of seven housekeeping genes was developed in this study, based on recent findings from whole genome sequencing data. This new scheme was used to assess the genetic diversity amongst 22 autochthonous L. donovani isolates from Bangladesh. Nineteen additional isolates of the L. donovani complex (including sequences of L. donovani reference strain BPK282A1) from other countries were included for comparison. By using restriction fragment length polymorphism of the internal transcribed spacer 1 region (ITS1-RFLP) and ITS1 sequencing, all Bangladeshi isolates were confirmed to be L. donovani. Population genetic analyses of 41 isolates using the seven new MLST loci clearly separated L. donovani from Leishmania infantum. With this multilocus sequence typing scheme, seven genotypes were identified amongst Bangladeshi L. donovani isolates, and these isolates were found to be phylogenetically different compared with those from India, Nepal, Iraq and Africa. This novel multilocus sequence typing approach can detect intra- and inter-species variations within the L. donovani complex, but most importantly these molecular markers can be applied to resolve the phylogenetically very homogeneous L. donovani strains from the Indian subcontinent. Four of these markers were found suitable to differentiate strains originating from Bangladesh, with marker A2P being the most discriminative one.
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http://dx.doi.org/10.1016/j.ijpara.2019.02.010DOI Listing
June 2019

Draft Whole-Genome Sequence of Leishmania (Viannia) braziliensis Presenting Leishmania RNA Virus 1, from Western Amazon, Brazil.

Microbiol Resour Announc 2018 Sep 6;7(9). Epub 2018 Sep 6.

Laboratório de Epidemiologia Genética, Fundação Oswaldo Cruz, Fiocruz Rondônia, Porto Velho, Brazil.

Leishmania (Viannia) braziliensis is the main etiological agent of tegumentary leishmaniasis in the neotropics. Here, we report a draft genome sequence (31.2 Mb) of an L. braziliensis strain from the western Amazon region of Brazil. This genome sequence will complement those available for other Leishmania species and contribute to further studies focusing on this parasite and the neglected diseases associated with it.
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http://dx.doi.org/10.1128/MRA.00924-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6256520PMC
September 2018

Genome Dynamics during Environmental Adaptation Reveal Strain-Specific Differences in Gene Copy Number Variation, Karyotype Instability, and Telomeric Amplification.

mBio 2018 11 6;9(6). Epub 2018 Nov 6.

Unité de Parasitologiemoléculaire et Signalisation, Institut Pasteur, Paris, France

Protozoan parasites of the genus adapt to environmental change through chromosome and gene copy number variations. Only little is known about external or intrinsic factors that govern genomic adaptation. Here, by conducting longitudinal genome analyses of 10 new clinical isolates, we uncovered important differences in gene copy number among genetically highly related strains and revealed gain and loss of gene copies as potential drivers of long-term environmental adaptation in the field. In contrast, chromosome rather than gene amplification was associated with short-term environmental adaptation to culture. Karyotypic solutions were highly reproducible but unique for a given strain, suggesting that chromosome amplification is under positive selection and dependent on species- and strain-specific intrinsic factors. We revealed a progressive increase in read depth towards the chromosome ends for various isolates, which may represent a nonclassical mechanism of telomere maintenance that can preserve integrity of chromosome ends during selection for fast growth. Together our data draw a complex picture of genomic adaptation in the field and in culture, which is driven by a combination of intrinsic genetic factors that generate strain-specific phenotypic variations, which are under environmental selection and allow for fitness gain. Protozoan parasites of the genus cause severe human and veterinary diseases worldwide, termed leishmaniases. A hallmark of biology is its capacity to adapt to a variety of unpredictable fluctuations inside its human host, notably pharmacological interventions, thus, causing drug resistance. Here we investigated mechanisms of environmental adaptation using a comparative genomics approach by sequencing 10 new clinical isolates of the , , and complexes that were sampled across eight distinct geographical regions. Our data provide new evidence that parasites adapt to environmental change in the field and in culture through a combination of chromosome and gene amplification that likely causes phenotypic variation and drives parasite fitness gains in response to environmental constraints. This novel form of gene expression regulation through genomic change compensates for the absence of classical transcriptional control in these early-branching eukaryotes and opens new venues for biomarker discovery.
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http://dx.doi.org/10.1128/mBio.01399-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6222132PMC
November 2018

New insights into the genetic diversity of Leishmania RNA Virus 1 and its species-specific relationship with Leishmania parasites.

PLoS One 2018 18;13(6):e0198727. Epub 2018 Jun 18.

Laboratório de Pesquisa em Leishmaniose, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz, Rio de Janeiro, Rio de Janeiro, Brazil.

Cutaneous leishmaniasis is a neglected parasitic disease that manifests in infected individuals under different phenotypes, with a range of factors contributing to its broad clinical spectrum. One factor, Leishmania RNA Virus 1 (LRV1), has been described as an endosymbiont present in different species of Leishmania. LRV1 significantly worsens the lesion, exacerbating the immune response in both experimentally infected animals and infected individuals. Little is known about the composition and genetic diversity of these viruses. Here, we investigated the relationship between the genetic composition of LRV1 detected in strains of Leishmania (Viannia) braziliensis and L. (V.) guyanensis and the interaction between the endosymbiont and the parasitic species, analyzing an approximately 850 base pair region of the viral genome. We also included one LRV1 sequence detected in L. (V.) shawi, representing the first report of LRV1 in a species other than L. braziliensis and L. guyanensis. The results illustrate the genetic diversity of the LRV1 strains analyzed here, with smaller divergences detected among viral sequences from the same parasite species. Phylogenetic analyses showed that the LRV1 sequences are grouped according to the parasite species and possibly according to the population of the parasite in which the virus was detected, corroborating the hypothesis of joint evolution of the viruses with the speciation of Leishmania parasites.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0198727PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6005476PMC
January 2019

Morphophysiological changes in the splenic extracellular matrix of Leishmania infantum-naturally infected dogs is associated with alterations in lymphoid niches and the CD4+ T cell frequency in spleens.

PLoS Negl Trop Dis 2018 04 20;12(4):e0006445. Epub 2018 Apr 20.

Laboratório de Pesquisas em Leishmaniose, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz, Rio de Janeiro, Rio de Janeiro, Brazil.

The spleen is one of the main affected organs in canine visceral leishmaniasis (CVL). Disorganization of the splenic white pulp (SWP) has been associated with immunosuppression and disease progression. This study aims to assess structural and cellular changes in the splenic extracellular matrix of dogs with CVL, correlating these changes with the parasite load and clinical signs. Splenic fragments were collected from 41 naturally infected animals for parasite load quantification by quantitative PCR, histopathological analysis and immunohistochemistry for CD3+, CD4+, and CD8+ T cells; CD21+ B cells; Ki-67+, IFN-γ+, and IL-10+ cells; and the MMP-9 and ADAM-10 enzymes. Laminin, collagen and fibronectin deposition were also evaluated. The animals were grouped according to the level of SWP organization. SWP disorganization was accompanied by a reduction in the quantity of lymphoid follicles/mm2 (p > 0.0001). Animals with moderate to intense SWP disorganization showed more clinical signs (p = 0.021), higher laminin (p = 0.045) and collagen deposition (p = 0.036), higher MMP-9 expression (p = 0.035) and lower numbers of CD4+ T cells (p = 0.027) in the spleen than the animals with organized SWP. These data suggest that splenic structure and function are drastically altered and compromised during CVL.
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http://dx.doi.org/10.1371/journal.pntd.0006445DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5931683PMC
April 2018

The applicability of real-time PCR in the diagnostic of cutaneous leishmaniasis and parasite quantification for clinical management: Current status and perspectives.

Acta Trop 2018 Aug 29;184:29-37. Epub 2017 Sep 29.

Laboratório de Pesquisas em Leishmanioses, Instituto Oswaldo Cruz, Fiocruz, Rio de Janeiro, Brazil. Electronic address:

Cutaneous leishmaniasis (CL) is spread worldwide and is the most common manifestation of leishmaniasis. Diagnosis is performed by combining clinical and epidemiological features, and through the detection of Leishmania parasites (or DNA) in tissue specimens or trough parasite isolation in culture medium. Diagnosis of CL is challenging, reflecting the pleomorphic clinical manifestations of this disease. Skin lesions vary in severity, clinical appearance, and duration, and in some cases, they can be indistinguishable from lesions related to other diseases. Over the past few decades, PCR-based methods, including real-time PCR assays, have been developed for Leishmania detection, quantification and species identification, improving the molecular diagnosis of CL. This review provides an overview of many real-time PCR methods reported for the diagnostic evaluation of CL and some recommendations for the application of these methods for quantification purposes for clinical management and epidemiological studies. Furthermore, the use of real-time PCR for Leishmania species identification is also presented. The advantages of real-time PCR protocols are numerous, including increased sensitivity and specificity and simpler standardization of diagnostic procedures. However, despite the numerous assays described, there is still no consensus regarding the methods employed. Furthermore, the analytical and clinical validation of CL molecular diagnosis has not followed international guidelines so far. A consensus methodology comprising a DNA extraction protocol with an exogenous quality control and an internal reference to normalize parasite load is still needed. In addition, the analytical and clinical performance of any consensus methodology must be accurately assessed. This review shows that a standardization initiative is essential to guide researchers and clinical laboratories towards the achievement of a robust and reproducible methodology, which will permit further evaluation of parasite load as a surrogate marker of prognosis and monitoring of aetiological treatment, particularly in multi-centric observational studies and clinical trials.
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http://dx.doi.org/10.1016/j.actatropica.2017.09.020DOI Listing
August 2018

Influences of climate change on the potential distribution of Lutzomyia longipalpis sensu lato (Psychodidae: Phlebotominae).

Int J Parasitol 2017 09 29;47(10-11):667-674. Epub 2017 Jun 29.

Parasitology Department, Universidade de São Paulo, São Paulo, Brazil.

This study explores the present day distribution of Lutzomyia longipalpis in relation to climate, and transfers the knowledge gained to likely future climatic conditions to predict changes in the species' potential distribution. We used ecological niche models calibrated based on occurrences of the species complex from across its known geographic range. Anticipated distributional changes varied by region, from stability to expansion or decline. Overall, models indicated no significant north-south expansion beyond present boundaries. However, some areas suitable both at present and in the future (e.g., Pacific coast of Ecuador and Peru) may offer opportunities for distributional expansion. Our models anticipated potential range expansion in southern Brazil and Argentina, but were variably successful in anticipating specific cases. The most significant climate-related change anticipated in the species' range was with regard to range continuity in the Amazon Basin, which is likely to increase in coming decades. Rather than making detailed forecasts of actual locations where Lu. longipalpis will appear in coming years, our models make interesting and potentially important predictions of broader-scale distributional tendencies that can inform heath policy and mitigation efforts.
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http://dx.doi.org/10.1016/j.ijpara.2017.04.007DOI Listing
September 2017

Hepatozoon canis and Leishmania spp. coinfection in dogs diagnosed with visceral leishmaniasis.

Rev Bras Parasitol Vet 2016 Oct-Dec;25(4):450-458. Epub 2016 Dec 1.

Laboratório de Pesquisa em Leishmaniose, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz - FIOCRUZ, Rio de Janeiro, RJ, Brasil.

This study describes the occurrence of dogs naturally co-infected with Hepatozoon canis and two Leishmania species: L. infantum or L. braziliensis. Four dogs serologically diagnosed with Visceral Leishmaniasis were euthanized. Liver and spleen samples were collected for histopathological analysis and DNA isolation. H. canis meronts were observed in tissues from all four dogs. H. canis infection was confirmed by PCR followed by sequencing of a fragment of 18S rRNA gene. Leishmania detection and typing was confirmed by ITS1' PCR-RFLP and parasite burden was calculated using ssrRNA quantitative qPCR. A DPP - Dual Path platform test was performed. One out (Dog #2) of four animals was asymptomatic. Dogs #1 and #4 were infected by L. infantum and were DPP test positive. Dogs #2 and #3 were infected by L. braziliensis and were DPP test negative. Furthermore, visceral dissemination was observed in Dogs #2 and #3, since L. braziliensis was detected in liver and spleen samples. The visceral dissemination of L. braziliensis associated with systemic signs suggested that this co-infection could influence the parasite burden and disease progression.
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http://dx.doi.org/10.1590/S1984-29612016065DOI Listing
May 2018

Leishmania (Viannia) naiffi: rare enough to be neglected?

Mem Inst Oswaldo Cruz 2015 Sep;110(6):797-800

Laboratório Interdisciplinar de Pesquisas Médicas, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz, Rio de Janeiro, RJ, Brasil.

In the Brazilian Amazon, American tegumentary leishmaniasis (ATL) is endemic and presents a wide spectrum of clinical manifestations due, in part, to the circulation of at least seven Leishmania species. Few reports of Leishmania (Viannia) naiffi infection suggest that its occurrence is uncommon and the reported cases present a benign clinical course and a good response to treatment. This study aimed to strengthen the clinical and epidemiological importance of L. (V.) naiffi in the Amazon Region (Manaus, state of Amazonas) and to report therapeutic failure in patients infected with this species. Thirty Leishmania spp samples isolated from cutaneous lesions were characterised by multilocus enzyme electrophoresis. As expected, the most common species was Leishmania (V.) guyanensis (20 cases). However, a relevant number of L. (V.) naiffi patients (8 cases) was observed, thus demonstrating that this species is not uncommon in the region. No patient infected with L. (V.) naiffi evolved to spontaneous cure until the start of treatment, which indicated that this species may not have a self-limiting nature. In addition, two of the patients experienced a poor response to antimonial or pentamidine therapy. Thus, either ATL cases due to L. (V.) naiffi cannot be as uncommon as previously thought or this species is currently expanding in this region.
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http://dx.doi.org/10.1590/0074-02760150128DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4667584PMC
September 2015

Successful isolation of Leishmania infantum from Rhipicephalus sanguineus sensu lato (Acari: Ixodidae) collected from naturally infected dogs.

BMC Vet Res 2015 Oct 9;11:258. Epub 2015 Oct 9.

Programa de Pós-Graduação em Medicina Tropical, Universidade de Brasília, Brasília, DF, Brazil.

Background: The main transmission route of Leishmania infantum is through the bites of sand flies. However, alternative mechanisms are being investigated, such as through the bites of ticks, which could have epidemiological relevance. The objective of this work was to verify the presence of Leishmania spp. in Rhipicephalus sanguineus sensu lato collected from naturally infected dogs in the Federal District of Brazil.

Methods: Ticks were dissected to remove their intestines and salivary glands for DNA extraction and the subsequent amplification of the conserved region of 120 bp of kDNA and 234 bp of the hsp70 gene of Leishmania spp. The amplified kDNA products were digested with endonucleases HaeIII and BstUI and were submitted to DNA sequencing. Isolated Leishmania parasites from these ticks were analyzed by multilocus enzyme electrophoresis, and the DNA obtained from this culture was subjected to microsatellite analyses.

Results: Overall, 130 specimens of R. sanguineus were collected from 27 dogs. Leishmania spp. were successfully isolated in culture from five pools of salivary glands and the intestines of ticks collected from four dogs. The amplified kDNA products from the dog blood samples and from the tick cultures, when digested by HaeIII and BstUI, revealed the presence of L. braziliensis and L. infantum. One strain was cultivated and characterized as L. infantum by enzyme electrophoresis. The amplified kDNA products from the blood of one dog showed a sequence homology with L. braziliensis; however, the amplified kDNA from the ticks collected from this dog showed a sequence homology to L. infantum.

Conclusion: The results confirm that the specimens of R. sanguineus that feed on dogs naturally infected by L. infantum contain the parasite DNA in their intestines and salivary glands, and viable L. infantum can be successfully isolated from these ectoparasites.
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http://dx.doi.org/10.1186/s12917-015-0576-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4600268PMC
October 2015

Further Evidence of an Association between the Presence of Leishmania RNA Virus 1 and the Mucosal Manifestations in Tegumentary Leishmaniasis Patients.

PLoS Negl Trop Dis 2015 15;9(9):e0004079. Epub 2015 Sep 15.

Fundação Oswaldo Cruz, Unidade Rondonia, Porto Velho, Rondonia, Brazil.

Tegumentary Leishmaniasis (TL) is endemic in Latin America, and Brazil contributes approximately 20 thousand cases per year. The pathogenesis of TL, however, is still not fully understood. Clinical manifestations vary from cutaneous leishmaniasis (CL) to more severe outcomes, such as disseminated leishmaniasis (DL), mucosal leishmaniasis (ML) and diffuse cutaneous leishmaniasis (DCL). Many factors have been associated with the severity of the disease and the development of lesions. Recent studies have reported that the presence of Leishmania RNA virus 1 infecting Leishmania (Leishmania RNA virus 1, LRV1) is an important factor associated with the severity of ML in experimental animal models. In the present study, 156 patients who attended Rondonia's Hospital of Tropical Medicine with both leishmaniasis clinical diagnoses (109 CL; 38 ML; 5 CL+ML; 3 DL and 1 DCL) and molecular diagnoses were investigated. The clinical diagnosis were confirmed by PCR by targeting hsp70 and kDNA DNA sequences and the species causing the infection were determined by HSP70 PCR-RFPL. The presence of LVR1 was tested by RT-PCR. Five Leishmania species were detected: 121 (77.6%) samples were positive for Leishmania (Viannia) braziliensis, 18 (11.5%) were positive for Leishmania (V.) guyanensis, 3 (1.8%) for Leishmania (V.) lainsoni, 2 (1.3%) for Leishmania (Leishmania) amazonensis and 2 (1.3%) for Leishmania (V.) shawi. Six (3.9%) samples were positive for Leishmania sp. but the species could not be determined, and 4 (2.6%) samples were suggestive of mixed infection by L. (V.) braziliensis and L. (V.) guyanensis. The virus was detected in L. braziliensis (N = 54), L. guyanensis (N = 5), L. amazonensis (N = 2), L. lainsoni (N = 1) and inconclusive samples (N = 6). Patients presenting with CL+ML, DL and DCL were excluded from further analysis. Association between the presence of the virus and the disease outcome were tested among the remaining 147 patients (CL = 109 and ML = 38). Of them, 71.1% (n = 27) mucosal lesions were positive for LRV1, and 28.9% (n = 11) were negative. In cutaneous lesions, 36.7% (n = 40) were positive and 63.3% (n = 69) were negative for LRV1. The ratio P(ML|LRV1+)/P(ML|LRV1-) was 2.93 (CI95% 1.57...5.46; p<0.001), thus corroborating the hypothesis of the association between LRV1 and the occurrence of mucosal leishmaniasis, as previously described in animal models; it also indicates that LRV1 is not the only factor contributing to the disease outcome.
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http://dx.doi.org/10.1371/journal.pntd.0004079DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4570810PMC
March 2016

Parasite load induces progressive spleen architecture breakage and impairs cytokine mRNA expression in Leishmania infantum-naturally infected dogs.

PLoS One 2015 13;10(4):e0123009. Epub 2015 Apr 13.

Laboratório de Pesquisas em Leishmaniose, Instituto Oswaldo Cruz, Fiocruz, Rio de Janeiro, Brasil.

Canine Visceral Leishmaniasis (CVL) shares many aspects with the human disease and dogs are considered the main urban reservoir of L. infantum in zoonotic VL. Infected dogs develop progressive disease with a large clinical spectrum. A complex balance between the parasite and the genetic/immunological background of the host are decisive for infection evolution and clinical outcome. This study comprised 92 Leishmania infected mongrel dogs of various ages from Mato Grosso, Brazil. Spleen samples were collected for determining parasite load, humoral response, cytokine mRNA expression and histopathology alterations. By real-time PCR for the ssrRNA Leishmania gene, two groups were defined; a low (lowP, n = 46) and a high parasite load groups (highP, n = 42). When comparing these groups, results show variable individual humoral immune response with higher specific IgG production in infected animals but with a notable difference in CVL rapid test optical densities (DPP) between highP and lowP groups. Splenic architecture disruption was characterized by disorganization of white pulp, more evident in animals with high parasitism. All cytokine transcripts in spleen were less expressed in highP than lowP groups with a large heterogeneous variation in response. Individual correlation analysis between cytokine expression and parasite load revealed a negative correlation for both pro-inflammatory cytokines: IFNγ, IL-12, IL-6; and anti-inflammatory cytokines: IL-10 and TGFβ. TNF showed the best negative correlation (r2 = 0.231; p<0.001). Herein we describe impairment on mRNA cytokine expression in leishmania infected dogs with high parasite load associated with a structural modification in the splenic lymphoid micro-architecture. We also discuss the possible mechanism responsible for the uncontrolled parasite growth and clinical outcome.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0123009PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4395300PMC
January 2016

Distinct Leishmania species infecting wild caviomorph rodents (Rodentia: Hystricognathi) from Brazil.

PLoS Negl Trop Dis 2014 Dec 11;8(12):e3389. Epub 2014 Dec 11.

Laboratory of Trypanosomatid Biology, Oswaldo Cruz Institute, Rio de Janeiro, Rio de Janeiro, Brazil.

Background: Caviomorph rodents, some of the oldest Leishmania spp. hosts, are widely dispersed in Brazil. Despite both experimental and field studies having suggested that these rodents are potential reservoirs of Leishmania parasites, not more than 88 specimens were analyzed in the few studies of natural infection. Our hypothesis was that caviomorph rodents are inserted in the transmission cycles of Leishmania in different regions, more so than is currently recognized.

Methodology: We investigated the Leishmania infection in spleen fragments of 373 caviomorph rodents from 20 different species collected in five Brazilian biomes in a period of 13 years. PCR reactions targeting kDNA of Leishmania sp. were used to diagnose infection, while Leishmania species identification was performed by DNA sequencing of the amplified products obtained in the HSP70 (234) targeting. Serology by IFAT was performed on the available serum of these rodents.

Principal Findings: In 13 caviomorph rodents, DNA sequencing analyses allowed the identification of 4 species of the subgenus L. (Viannia): L. shawi, L. guyanensis, L. naiffi, and L. braziliensis; and 1 species of the subgenus L. (Leishmania): L. infantum. These include the description of parasite species in areas not previously included in their known distribution: L. shawi in Thrichomys inermis from Northeastern Brazil and L. naiffi in T. fosteri from Western Brazil. From the four other positive rodents, two were positive for HSP70 (234) targeting but did not generate sequences that enabled the species identification, and another two were positive only in kDNA targeting.

Conclusions/significance: The infection rate demonstrated by the serology (51.3%) points out that the natural Leishmania infection in caviomorph rodents is much higher than that observed in the molecular diagnosis (4.6%), highlighting that, in terms of the host species responsible for maintaining Leishmania species in the wild, our current knowledge represents only the "tip of the iceberg."
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http://dx.doi.org/10.1371/journal.pntd.0003389DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4263410PMC
December 2014

Screening and characterization of RAPD markers in viscerotropic Leishmania parasites.

PLoS One 2014 14;9(10):e109773. Epub 2014 Oct 14.

Laboratoire d'Epidémiologie Moléculaire et de Pathologie Expérimentale (LR11IPT04)/Laboratoire d'Epidémiologie et d'Ecologie Parasitaire (LR00SP04) -Institut Pasteur de Tunis, Université Tunis el Manar, Tunis, Tunisia.

Visceral leishmaniasis (VL) is mainly due to the Leishmania donovani complex. VL is endemic in many countries worldwide including East Africa and the Mediterranean region where the epidemiology is complex. Taxonomy of these pathogens is under controversy but there is a correlation between their genetic diversity and geographical origin. With steady increase in genome knowledge, RAPD is still a useful approach to identify and characterize novel DNA markers. Our aim was to identify and characterize polymorphic DNA markers in VL Leishmania parasites in diverse geographic regions using RAPD in order to constitute a pool of PCR targets having the potential to differentiate among the VL parasites. 100 different oligonucleotide decamers having arbitrary DNA sequences were screened for reproducible amplification and a selection of 28 was used to amplify DNA from 12 L. donovani, L. archibaldi and L. infantum strains having diverse origins. A total of 155 bands were amplified of which 60.65% appeared polymorphic. 7 out of 28 primers provided monomorphic patterns. Phenetic analysis allowed clustering the parasites according to their geographical origin. Differentially amplified bands were selected, among them 22 RAPD products were successfully cloned and sequenced. Bioinformatic analysis allowed mapping of the markers and sequences and priming sites analysis. This study was complemented with Southern-blot to confirm assignment of markers to the kDNA. The bioinformatic analysis identified 16 nuclear and 3 minicircle markers. Analysis of these markers highlighted polymorphisms at RAPD priming sites with mainly 5' end transversions, and presence of inter- and intra- taxonomic complex sequence and microsatellites variations; a bias in transitions over transversions and indels between the different sequences compared is observed, which is however less marked between L. infantum and L. donovani. The study delivers a pool of well-documented polymorphic DNA markers, to develop molecular diagnostics assays to characterize and differentiate VL causing agents.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0109773PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4196940PMC
June 2015

Polymorphisms and ambiguous sites present in DNA sequences of Leishmania clones: looking closer.

Infect Genet Evol 2014 Jul 21;25:110-6. Epub 2014 Apr 21.

Laboratório de Pesquisa em Leishmaniose, Instituto Oswaldo Cruz (IOC), Fundação Oswaldo Cruz (FIOCRUZ), Rio de Janeiro, Brazil.

In genetic studies of Leishmania parasites, co-dominant markers are chosen for their ability to detect heterozygous polymorphisms, to infer the occurrence of inbreeding and to resolve genetic variability. The majority of DNA sequence based reports perform conventional dye terminator cycle sequencing where perfectly ambiguous sites or double peaks in the chromatogram are interpreted as heterozygous strains. However, molecular peculiarities of the parasite such as aneuploidy, mixed populations and homologous recombination advise that data from regular DNA sequence analysis should be carefully evaluated. We report here a closer look at ambiguous sites observed in 6pgd DNA sequences obtained for a multilocus sequence analysis project on Leishmania (Viannia) strains. After comparing 286 DNA sequences from biological and molecular clones of six L. (Viannia) strains we could distinguish events that contribute to genetic variation in Leishmania (recombination, mutation, chromosomal mosaics). Also, the results suggest how diversity might not be completely revealed through regular DNA sequence analysis and demonstrate the importance for molecular epidemiology research to be aware of such possibilities while choosing samples for studies.
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http://dx.doi.org/10.1016/j.meegid.2014.04.011DOI Listing
July 2014

Multilocus sequence analysis for Leishmania braziliensis outbreak investigation.

PLoS Negl Trop Dis 2014 Feb 13;8(2):e2695. Epub 2014 Feb 13.

Laboratório de Pesquisa em Leishmaniose, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz, Rio de Janeiro, Brazil.

With the emergence of leishmaniasis in new regions around the world, molecular epidemiological methods with adequate discriminatory power, reproducibility, high throughput and inter-laboratory comparability are needed for outbreak investigation of this complex parasitic disease. As multilocus sequence analysis (MLSA) has been projected as the future gold standard technique for Leishmania species characterization, we propose a MLSA panel of six housekeeping gene loci (6pgd, mpi, icd, hsp70, mdhmt, mdhnc) for investigating intraspecific genetic variation of L. (Viannia) braziliensis strains and compare the resulting genetic clusters with several epidemiological factors relevant to outbreak investigation. The recent outbreak of cutaneous leishmaniasis caused by L. (V.) braziliensis in the southern Brazilian state of Santa Catarina is used to demonstrate the applicability of this technique. Sequenced fragments from six genetic markers from 86 L. (V.) braziliensis strains from twelve Brazilian states, including 33 strains from Santa Catarina, were used to determine clonal complexes, genetic structure, and phylogenic networks. Associations between genetic clusters and networks with epidemiological characteristics of patients were investigated. MLSA revealed epidemiological patterns among L. (V.) braziliensis strains, even identifying strains from imported cases among the Santa Catarina strains that presented extensive homogeneity. Evidence presented here has demonstrated MLSA possesses adequate discriminatory power for outbreak investigation, as well as other potential uses in the molecular epidemiology of leishmaniasis.
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http://dx.doi.org/10.1371/journal.pntd.0002695DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3923721PMC
February 2014

Population structure and evidence for both clonality and recombination among Brazilian strains of the subgenus Leishmania (Viannia).

PLoS Negl Trop Dis 2013 31;7(10):e2490. Epub 2013 Oct 31.

Institut für Mikrobiologie und Hygiene, Charité Universitätsmedizin Berlin, Berlin, Germany.

Background/objectives: Parasites of the subgenus Leishmania (Viannia) cause varying clinical symptoms ranging from cutaneous leishmaniases (CL) with single or few lesions, disseminated CL (DL) with multiple lesions to disfiguring forms of mucocutaneous leishmaniasis (MCL). In this population genetics study, 37 strains of L. (V.) guyanensis, 63 of L. (V.) braziliensis, four of L. (V.) shawi, six of L. (V.) lainsoni, seven of L. (V.) naiffi, one each of L. (V.) utingensis and L. (V.) lindenbergi, and one L. (V.) lainsoni/L. naiffi hybrid from different endemic foci in Brazil were examined for variation at 15 hyper-variable microsatellite markers.

Methodology/principal Findings: The multilocus microsatellite profiles obtained for the 120 strains were analysed using both model- and distance-based methods. Significant genetic diversity was observed for all L. (Viannia) strains studied. The two cluster analysis approaches identified two principal genetic groups or populations, one consisting of strains of L. (V.) guyanensis from the Amazon region and the other of strains of L. (V.) braziliensis isolated along the Atlantic coast of Brazil. A third group comprised a heterogeneous assembly of species, including other strains of L. braziliensis isolated from the north of Brazil, which were extremely polymorphic. The latter strains seemed to be more closely related to those of L. (V.) shawi, L. (V.) naiffi, and L. (V.) lainsoni, also isolated in northern Brazilian foci. The MLMT approach identified an epidemic clone consisting of 13 strains of L. braziliensis from Minas Gerais, but evidence for recombination was obtained for the populations of L. (V.) braziliensis from the Atlantic coast and for L. (V.) guyanensis.

Conclusions/significance: Different levels of recombination versus clonality seem to occur within the subgenus L. (Viannia). Though clearly departing from panmixia, sporadic, but long-term sustained recombination might explain the tremendous genetic diversity and limited population structure found for such L. (Viannia) strains.
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http://dx.doi.org/10.1371/journal.pntd.0002490DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3814519PMC
June 2014

Severity of tegumentary leishmaniasis is not exclusively associated with Leishmania RNA virus 1 infection in Brazil.

Mem Inst Oswaldo Cruz 2013 Aug;108(5):665-7

Laboratório Interdisciplinar de Pesquisas Médicas, Instituto Oswaldo Cruz, Rio de Janeiro, RJ, Brasil.

Leishmania RNA virus (LRV) has been shown to be a symbiotic component of Leishmania parasites in South America. Nested retro-transcription polymerase chain reaction was employed to investigate LRV1 presence in leishmaniasis lesions from Brazil. In endemic areas of Rio de Janeiro (RJ), no LRV1 infection was observed even with mucosal involvement. LRV1 was only detected in Leishmania (V.) guyanensis cutaneous lesions from the northern region, which were obtained from patients presenting with disease reactivation after clinical cure of their primary lesions. Our results indicated that the severity of leishmaniasis in some areas of RJ, where Leishmania (V.) brazi-liensis is the primary etiological agent, was not associated with Leishmania LRV1 infection.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3970596PMC
http://dx.doi.org/10.1590/0074-0276108052013021DOI Listing
August 2013