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Leishmania infantum chagasi is the primary etiological agent of visceral leishmaniasis in Latin America, a lethal disease that afflicts hundreds of thousands of people worldwide every year. Previous studies have shown that the parasite releases microvesicles known as exosomes, which prolong and exacerbate infection in the vertebrate vector. However, little is known of their role in the insect vector, the sand fly Lutzomyia longipalpis. Exosomes were isolated from cultured L. i. chagasi in logarithmic (procyclic) (LOG) and stationary phase (metacyclic-like) (STAT) growth stages, which are the parasite stages found in the vector, and submitted to proteomic analysis. Our studies showed that exosomes from LOG and STAT L. i. chagasi display discrete protein profiles. The presence of approximately 50 known virulence factors was detected, including molecules for immunomodulation and evasion (GP63, EF1α, Oligopeptidase), increased pathogenicity (Casein kinase, KMP-11, Cysteine Peptidase and BiP) and parasite protection (Peroxidoxin). Additionally, the majority of ontological terms were associated with both exosome phases, and no substantial ontological enrichment was observed associated with any of the two exosomal stages. We demonstrated that LOG exosomes show a marked increase in protein number and abundance, including many virulence factors, compared to STAT L. i. chagasi exosomes. SIGNIFICANCE: The knowledge of the role of Leishmania exosomes on leishmaniasis opened up a new world of potential and complexity regarding our understanding of the disease. In Brazil the majority of visceral leishmaniasis cases are caused by the parasite Leishmania infantum chagasi and transmitted by the vector Lutzomyia longipalpis. While Leishmania exosomes were found to play an active role in the mammalian host, little is understood about their effects on the sand fly, or how they might impact on the insect infection by the parasite. For this reason, we isolated exosomes from two developmental stages of L. i. chagasi that occur within the insect with a view to identifying and describing the alterations they undergo. We have identified many hundreds of proteins within both exosome phases and have developed a structure by which to examine potential candidates. Our findings regarding the composition of the exosome proteome raise many questions regarding their function and provide compelling evidence that exosomes play an active role in the parasite's development within the sand fly., (Copyright © 2020 The Authors. Published by Elsevier B.V. All rights reserved.)

Visceral leishmaniasis (VL) is a disease of great concern for public health because of its high incidence and lethality. Here, we performed a serologic study of domestic dogs in the municipality of Petrolina in northeastern Brazil to evaluate the possible risk factors associated with canine seropositivity for Leishmania infantum chagasi. Blood samples from 1245 dogs in urban and rural areas were collected and examined by indirect immunofluorescent antibody test (IFAT) and an indirect enzyme-linked immunosorbent assay (ELISA). The dogs were subjected to physical examination and classified according to their clinical manifestations. A questionnaire was administered to the owners to detect potential risk factors for infection with Leishmania spp. using logistic regression models. Of the 1245 dogs evaluated, 11.2% (140/1245) were seropositive in both tests (CI 95%: 9.5% to 13.1%). Approximately 60.7% of the reactive dogs were clinically suspect, with lymphadenomegaly, cutaneous ulcerations, onychogryphosis, pale mucous membranes and alopecia being the most obvious symptoms of infection. The seroprevalences in urban and rural areas were 5.4% (CI 95%: 4% to 7.1%) and 23.6% (CI 95%: 19.5% to 28.1%), respectively. The possible risk factors for the presence of anti L. infantum chagasi antibodies were the presence of a green area close to the home of the animal (OR=3.63; p<0.001), a mongrel breed (OR=2.11; p=0.025) and male gender (OR=1.51, p=0.034). The seroprevalence of L. infantum chagasi in the canine population is distributed in a heterogeneous manner, with a higher prevalence in rural areas., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Visceral leishmaniasis, a disease caused by Leishmania infantum chagasi, represents a major public health problem in many areas of the world. However, there is currently no vaccine for human use. The aim of this work was to purify the 503 antigen of Leishmania i. chagasi directly from unclarified Escherichia coli feedstock through expanded bed adsorption (EBA) chromatography. Batch experiments were performed to optimize the adsorption and elution conditions of the antigen onto a STREAMLINE Chelating resin using two central composite rotatable designs (CCRD). The results showed that the optimal binding conditions of the 503 antigen were pH 8.0 in the presence of 2.4 M NaCl. For the elution of the target protein, the optimized conditions included the presence of 600.0 mM imidazole. The adsorption isothermal data of the 503 antigen were fitted to the Langmuir adsorption isotherm. The EBA experiment successfully recovered 59.2% of the 503 antigen from the unclarified E. coli homogenate with a purification factor of 6.0., (Copyright © 2015 Elsevier B.V. All rights reserved.)