Browsing by Author "Fernandes, Ana Paula"

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    Antibody responses induced by Leish-Tec®, an A2-basedvaccine for visceral leishmaniasis, in a heterogeneous caninepopulation.
    (2014) Testasicca, Miriam Conceição de Souza; Santos, Mariana Silva dos; Machado, Leopoldo Marques; Serufo, Ângela Vieira; Doro, Daniel; Avelar, Daniel; Tibúrcio, Ana Maria Leonardi; Abrantes, Christiane de Freitas; Coelho, George Luiz Lins Machado; Grimaldi Junior, Gabriel; Gazzinelli, Ricardo Tostes; Fernandes, Ana Paula
    Zoonotic visceral leishmaniasis (VL) is a widespread disease, and dogs are the main reser-voirs for human parasite transmission. Hence, development of an effective vaccine thatprevents disease and reduces the transmission of VL is required. As euthanasia of seropos-itive dogs is recommended in Brazil for VL epidemiological control, to include anti-VLcanine vaccines as a mass control measure it is necessary to characterize the humoralresponses induced by vaccination and if they interfere with the reactivity of vaccinateddogs in serological diagnostic tests. Leish-Tec®is an amastigote-specific A2 recombinantprotein vaccine against canine visceral leishmaniasis (CVL) that is commercially availablein Brazil. Here, we tested the immunogenicity of Leish-Tec®in a heterogeneous dog popula-tion by measuring A2-specific antibody responses. Healthy dogs (n = 140) of various breedswere allocated to two groups: one group received Leish-Tec®(n = 70), and the other groupreceived a placebo (n = 70). Anti-A2 or anti-Leishmania promastigote antigen (LPA) antibodylevels were measured by ELISA in serum samples collected before and after vaccination.An immunochromatographic test (DPP) based on the recombinant K28 antigen was alsoused for serodiagnosis of CVL. Vaccinated animals, except one, remained seronegative foranti-LPA total IgG and anti-K28 antibodies. Conversely, seropositivity for anti-A2 total IgGantibodies was found in 98% of animals after vaccination. This value decreased to 81.13% at6 months before rising again (98%), after the vaccination boost. Anti-A2 IgG2 and IgG1 titers.
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    Antigenic extracts of Leishmania braziliensis and Leishmania amazonensis associated with saponin partially protects BALB/c mice against Leishmania chagasi infection by suppressing IL-10 and IL-4 production.
    (2010) Grenfell, Rafaella Fortini Queiroz; Silva, Eduardo de Almeida Marques da; Testasicca, Miriam Conceição de Souza; Coelho, Eduardo Antônio Ferraz; Fernandes, Ana Paula; Afonso, Luís Carlos Crocco; Rezende, Simone Aparecida
    This study evaluated two vaccine candidates for their effectiveness in protecting BALB/c mice against Leishma¬nia chagasi infection. These immunogenic preparations were composed of Leishmania amazonensis or Leishmania braziliensis antigenic extracts in association with saponin adjuvant. Mice were given three subcutaneous doses of one of these vaccine candidates weekly for three weeks and four weeks later challenged with promastigotes of L. chagasi by intravenous injection. We observed that both vaccine candidates induced a significant reduction in the parasite load of the liver, while the L. amazonensis antigenic extract also stimulated a reduction in spleen parasite load. This protection was associated with a suppression of both interleukin (IL)-10 and IL-4 cytokines by spleen cells in response to L. chagasi antigen. No change was detected in the production of IFN-γ. Our data show that these im-munogenic preparations reduce the type 2 immune response leading to the control of parasite replication.
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    Efficacy of nanoemulsion with Pterodon emarginatus Vogel oleoresin for topical treatment of cutaneous leishmaniasis.
    (2021) Kawakami, Monique Yoko Martins; Zamora, Lisset Ortiz; Araújo, Raquel Silva; Fernandes, Caio Pinho; Ricotta, Tiago Queiroga Nery; Oliveira, Leandro G. de; Queiroz Júnior, Celso Martins; Fernandes, Ana Paula; Conceição, Edemilson Cardoso da; Ferreira, Lucas Antônio Miranda; Barros, Andre Luis Branco de; Aguiar, Marta Marques Gontijo de; Oliveira, Anna Eliza Maciel de Faria Mota
    Cutaneous leishmaniasis (CL) is a neglected tropical skin disease caused by the protozoan genus Leishmania. The treatment is restricted to a handful number of drugs that exhibit toxic effects, limited efficacy, and drug resis- tance. Additionally, developing an effective topical treatment is still an enormous unmet medical challenge. Natural oils, e.g. the oleoresin from P. emarginatus fruits (SO), contain various bioactive molecules, especially terpenoid compounds such as diterpenes and sesquiterpenes. However, its use in topical formulations can be impaired due to the natural barrier of the skin for low water solubility compounds. Nanoemulsions (NE) are drug delivery systems able to increase penetration of lipophilic compounds throughout the skin, improving their topical effect. In this context, we propose the use of SO-containing NE (SO-NE) for CL treatment. The SO-NE was produced by a low energy method and presented suitable physicochemical characteristic: average diameter and polydispersity index lower than 180 nm and 0.2, respectively. Leishmania (Leishmania) amazonensis-infected BALB/c mice were given topical doses of SO or SO-NE. The topical use of a combination of SO-NE and intra- peritoneal meglumine antimoniate reduced lesion size by 41 % and tissue regeneration was proven by histo- pathological analyses. In addition, a reduction in the parasitic load and decreased in the level of IFN-γ in the lesion may be associated, as well as a lower level of the cytokine IL-10 may be associated with a less intense inflammatory process. The present study suggests that SO-NE in combination meglumine antimoniate represents a promising alternative for the topical treatment of CL caused by L. (L.) amazonensis.
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    Epitope mapping and protective immunity elicited by adenovirus expressing the Leishmania amastigote specific A2 antigen : correlation with IFN- and cytolytic activity by CD8+ T cells.
    (2008) Resende, Daniela de Melo; Caetano, Bráulia Costa; Dutra, Míriam Santos; Abrantes, Christiane de Freitas; Verly, Rodrigo Moreira; Resende, Jarbas Magalhães; Veloso, Dorila Piló; Rezende, Simone Aparecida; Romero, Oscar Bruna; Fernandes, Ana Paula; Gazzinelli, Ricardo Tostes
    A2was identified as an amastigote virulence factor of Leishmania (Leishmania) donovani and as a candidate antigen for vaccine development against visceral leishmaniasis. Here, predicted hydrophilic, class I and II MHC-binding synthetic peptides were used to define epitopes recognized by A2-specific antibodies, CD8+ T and CD4+ T cells, respectively. Immunization of BALB/c mice with adenovirus expressing A2 (AdA2) resulted in lowantibody response, contrasting with high levels of IFN-_ producing CD4+ T and CD8+ T cells specific for A2. Further, A2-specific CD8+ T cells from immunized mice were capable of lysing sensitized target cells in vivo. Finally, we demonstrated an association of A2-specific T cell responses and reduced parasitism in both liver and spleen from mice immunized with AdA2 and challenged with L. (L.) chagasi.
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    Evaluation of immune responses and protection induced by A2 and nucleoside hydrolase (NH) DNA vaccines against Leishmania chagasi and Leishmania amazonensis experimental infections.
    (2007) Zanin, Francisca Helena Calheiros; Coelho, Eduardo Antônio Ferraz; Tavares, Carlos Alberto Pereira; Silva, Eduardo de Almeida Marques da; Costa, Miriam Maria Silva; Rezende, Simone Aparecida; Gazzinelli, Ricardo Tostes; Fernandes, Ana Paula
    Several antigens have been tested as vaccine candidates against Leishmania infections but controversial results have been reported when different antigens are co-administered in combined vaccination protocols. Immunization with A2 or nucleoside hydrolase (NH) antigens was previously shown to induce Th1 immune responses and protection in BALB/c mice against Leishmania donovani and L. amazonensis (A2) or L. donovani and L. mexicana (NH) infections. In this work, we investigated the protective efficacy of A2 and NH DNA vaccines, in BALB/c mice, against L. amazonensis or L. chagasi challenge infection. Immunization with either A2 (A2-pCDNA3) or NH (NH-VR1012) DNA induced an elevated IFN-g production before infection; however, only A2 DNA immunized mice were protected against both Leishmania species and displayed a sustained IFN-g production and very low IL-4 and IL-10 levels, after challenge. Mice immunized with NH/A2 DNA produced higher levels of IFN-g in response to both specific recombinant proteins (rNH or rA2), but displayed higher IL-4 and IL-10 levels and increased edema and parasite loads after L. amazonensis infection, as compared to A2 DNA immunized animals. These data extend the characterization of the immune responses induced by NH and A2 antigens as potential candidates to compose a defined vaccine and indicate that a highly polarized type 1 immune response is required for improvement of protective levels of combined vaccines against both L. amazonensis and L. chagasi infections.
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    Evaluation of three recombinant proteins for the development of ELISA and immunochromatographic tests for visceral leishmaniasis serodiagnosis.
    (2019) Santos, Anna Raquel Ribeiro dos; Serufo, Ângela Vieira; Figueiredo, Maria Marta; Godoi, Lara Carvalho; Vitório, Jéssica Gardone; Marcelino, Andreza Pain; Avelar, Daniel Moreira de; Rodrigues, Fernandes Tenório Gomes; Coelho, George Luiz Lins Machado; Medeiros, Fernanda Alvarenga Cardoso; Jeronimo, Selma Maria Bezerra; Oliveira, Edward José de; Nascimento, Frederico Crepaldi; Teixeira, Santuza Maria Ribeiro; Gazzinelli, Ricardo Tostes; Nagem, Ronaldo Alves Pinto; Fernandes, Ana Paula
    BACKGROUND Visceral Leishmaniasis (VL) is an infectious disease that is a significant cause of death among infants aged under 1 year and the elderly in Brazil. Serodiagnosis is a mainstay of VL elimination programs; however, it has significant limitations due to low accuracy. OBJECTIVE This study aimed to evaluate three recombinant Leishmania infantum proteins (rFc, rC9, and rA2) selected from previous proteomics and genomics analyses to develop enzyme-linked immunosorbent assay (ELISA) and immunochromatographic tests (ICT) for the serodiagnosis of human VL (HVL) and canine VL (CVL). METHODS A total of 186 human (70 L. infantum-infected symptomatic, 20 other disease-infected, and 96 healthy) and 185 canine (82 L. infantum-infected symptomatic, 27 L. infantum-infected asymptomatic, and 76 healthy) sera samples were used for antibody detection. FINDINGS Of the three proteins, rA2 (91.5% sensitivity and 87% specificity) and rC9 (95.7% sensitivity and 87.5% specificity) displayed the best performance in ELISA-HVL and ELISA-CVL, respectively. ICT-rA2 also displayed the best performance for HVL diagnosis (92.3% sensitivity and 88.0% specificity) and had high concordance with immunofluorescence antibody tests (IFAT), ELISA-rK39, IT-LEISH®, and ELISAEXT. ICT-rFc, ICT-rC9, and ICT-rA2 had sensitivities of 88.6%, 86.5%, and 87.0%, respectively, with specificity values of 84.0%, 92.0%, and 100%, respectively for CVL diagnosis. MAIN CONCLUSIONS The three antigens selected by us are promising candidates for VL diagnosis regardless of the test format, although the antigen combinations and test parameters may warrant further optimisation.
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    Field randomized trial to evaluate the efficacy of the Leish - Tec vaccine against canine visceral leishmaniasis in anendemic area of Brazil.
    (2016) Silva, Shara Regina; Feres, Ana Maria Leonardi Tibúrcio; Silva, João Carlos França da; Dias, Edelberto Santos; Michalsky, Érika Monteiro; Andrade, Hélida Monteiro de; Coelho, Eduardo Antônio Ferraz; Ribeiro, Gustavo Meirelles; Fernandes, Ana Paula; Coelho, George Luiz Lins Machado
    Background: A canine vaccine remains a promising approach for effective control of visceral leishmaniasis(VL), given its complex epidemiology in areas where zoonotic VL is prevalent. Leish-Tec®is a recombi-nant vaccine, based on the Leishmania A2 antigen, against canine VL (CVL). It is, since 2014, the singlecommercial vaccine licensed in Brazil. Here, Leish-Tec®efficacy was estimated through a randomizedfield trial (RFT), in a highly VL endemic area.Methods: The RFT was conducted from 2008 to 2010 in an endemic area of southeastern Brazil, presentinga CVL seroprevalence of 41.9%. Eight hundred forty-seven seronegative dogs were randomly selected toreceive Leish-Tec®(n = 429) or placebo (n = 418). Animals were followed up by clinical, serological, andparasitological exams for 18 months. The CVL incidence in both groups was compared through proportionanalysis.Results: A significant reduction in the number of cases of CVL was observed in the vaccine group, ascompared with the placebo group, whether efficacy was estimated according to parasitological results(71.4%; 95% CI: 34.9–87.3%; p = 0.001; risk ratio = 0.287), by adding results of xenodiagnosis and parasito-logical exams (58.1%; 95% CI: 26.0–76.3%; p = 0.002; risk ratio = 0.419). Among the animals that convertedto a positive anti-A2 serology, efficacy reached 80.8% (95% CI: 37.6–94.1%, p = 0.001; risk ratio = 0.192).Xenodiagnosis has detected a reduction of 46.6% (p = 0.05) in transmission to sand flies from vaccinatedanimals presenting anti-A2 positive serology.Conclusion: The Leish-Tec®vaccine proved significantly effective for prophylaxis of CVL, after naturalchallenge assured by transmission of Leishmania parasites, in a highly endemic area. Noteworthy, thisreport has unveiled the complexity of performing a RFT for anti-CVL vaccines in Brazil, which may behelpful for designing of future studies.
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    Intramuscular immunization with p36(LACK) DNA vaccine induces IFN-gama production but does not protect BALB/c mice against Leishmania chagasi intravenous challenge.
    (2005) Silva, Eduardo de Almeida Marques da; Coelho, Eduardo Antônio Ferraz; Gomes, Daniel Cláudio de Oliveira; Vilela, Márcia de Carvalho; Masioli, Cássio Zumerle; Tavares, Carlos Alberto Pereira; Fernandes, Ana Paula; Afonso, Luís Carlos Crocco; Rezende, Simone Aparecida
    Acute visceral leishmaniasis is a progressive disease caused by Leishmania chagasi in South America. The acquisition of immunity following infection suggests that vaccination is a feasible approach to protect against this disease. Since Leishmania homologue of receptors for activated C kinase (LACK) antigen is of particular interest as a vaccine candidate because of the prominent role it plays in the pathogenesis of experimental Leishmania major infection, we evaluated the potential of a p36(LACK) DNA vaccine in protecting BALB/c mice challenged with L. chagasi. In this study, mice received intramuscular (i.m.) or subcutaneous (s.c.) doses of LACK DNA vaccine. We evaluated the production of vaccine-induced cytokines and whether this immunization was able to reduce parasite load in liver and spleen. We detected a significant production of interferon gamma by splenocytes from i.m. vaccinated mice in response to L. chagasi antigen and to rLACK protein. However, we did not observe a reduction in parasite load neither in liver nor in the spleen of vaccinated animals. The lack of protection observed may be explained by a significant production of IL-10 induced by the vaccine.
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    Making an anti-amastigote vaccine for visceral leishmaniasis : rational, update and perspectives.
    (2012) Fernandes, Ana Paula; Coelho, Eduardo Antônio Ferraz; Coelho, George Luiz Lins Machado; Grimaldi Junior, Gabriel; Gazzinelli, Ricardo Tostes
    Visceral leishmaniasis is a major health problem in Latina America, as well as the Mediterranean region of Europe and Asia. We aimed to develop a vaccine against visceral leishmaniasis targeting the intracellular amastigotes, which is the parasite stage that persists throughout infections with Leishmania parasites. With this in mind, we identified an amastigote specific antigen (A2) that contains an immunogenic epitope for CD4+ T helper (Th) cells and multiple repetitive units encoding CD8+ cytotoxic T lymphocyte (CTL) epitopes. Vaccine formulations containing the recombinant A2 associated with saponin, alum and IL-12 or expressed by attenuated adenovirus were shown to be protective in mice, dogs and nonhuman-primates. We are currently identifying novel amastigote specific immunogenic proteins that could be aggregated to A2 to further improve the level of vaccineinduced cell-mediated immunity and protection against visceral leishmaniasis.
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    Sensitive and specific serodiagnosis of Leishmania infantum infection in dogs using peptides selected from hypothetical proteins identified by an immunoproteomic approach.
    (2013) Chávez Fumagalli, Miguel Angel; Martins, Vívian Tamietti; Testasicca, Miriam Conceição de Souza; Lage, Daniela Pagliara; Costa, Lourena Emanuele; Lage, Paula Souza; Duarte, Mariana Costa; Ker, Henrique Gama; Ribeiro, Tatiana Gomes; Carvalho, Fernando A. A.; Régis, Wiliam César Bento; Reis, Alexandre Barbosa; Tavares, Carlos Alberto Pereira; Soto, Manuel; Fernandes, Ana Paula; Coelho, Eduardo Antônio Ferraz
    In Brazil, the percentage of infected dogs living in areas where canine visceral leishmaniasis (CVL) is endemic ranges from 10 to 62%; however, the prevalence of infection in dogs is probably higher than figures reported from serological studies. In addition, problems with the occurrence of false-positive or false-negative results in the serodiagnosis of CVL have been reported. The present work analyzed the potential of synthetic peptides mapped from hypothetical proteins for improvement of the serodiagnosis of Leishmania infantum infection in dogs. From 26 identified leishmanial proteins, eight were selected, considering that no homologies between these proteins and others from trypanosomatide sequence databases were encountered. The sequences of these proteins were mapped to identify linear B-cell epitopes, and 17 peptides were synthesized and tested in enzyme-linked immunosorbent assays (ELISAs) for the serodiagnosis of L. infantum infection in dogs. Of these, three exhibited sensitivity and specificity values higher than 75% and 90%, respectively, to differentiate L. infantum-infected animals from Trypanosoma cruziinfected animals and healthy animals. Soluble Leishmania antigen (SLA) showed poor sensitivity (4%) and specificity (36%) to differentiate L. infantum-infected dogs from healthy and T. cruzi-infected dogs. Lastly, the three selected peptides were combined in different mixtures and higher sensitivity and specificity values were obtained, even when sera from T. cruzi-infected dogs were used. The study’s findings suggest that these three peptides can constitute a potential tool for more sensitive and specific serodiagnosis of L. infantum infection in dogs.