Presence of antibodies against Leishmania chagasi in haemodialysed patients

SOUZA,Roberto Mascarenhas et al.Presence of antibodies against Leishmania chagasi in haemodialysed patients.Transactions of the Royal Society of Tropical Medicine and Hygiene,v. 103, p.749-751, 2009.

Presence of antibodies against Leishmania chagasi in haemodialysed patients

SOUZA, R. M. et al. Presence of antibodies against Leishmania chagasi in haemodialysed patients. Transactions of the Royal Society of Tropical Medicine and Hygiene, v. 103, n.7, p. 749—751. ISSN 0035-9203.

Influência do vírus da hepatite G (GBV-C) na resposta imune frente à infecção por Leishmania chagasi

GB virus type C (GBV-C) appears to promote a Th1 response and is associated with prolonged survival in HIV-infected people. L. chagasi causes a spectrum of illness that varies from severe visceral leishmaniasis, a disease that in the majority of cases is fatal if not treated, to self resolution of infection and development of positive DTH response that is protective against symptomatic disease. To determine if GBV-C viremia might influence the outcome of Leishmania infection, we characterized GBV-C status in a cohort of subjects residing in a L. chagasi endemic area in Brazil. GBV-C viremia was more prevalent in blood donors from urban than in periurban regions of Natal, Brazil (16% and 7.5% respectively). Evidence of prior GBV-C (anti-E2 antibodies) was detected in 24% and 12%of these groups respectively. Anti-E2 increased with age (p= 0.0121). No difference in GBV-C viremia was found in the DTH+ and VL groups (p= 0.269); however, subjects with visceral leishmaniasis were more likely to have anti-E2 than DTH+ subjects (p=0.0012), and DTH induration was smaller in subjects with E2 antibodies (4.5 mm) compared those without (7.12 mm) (p= 0.002). Furthermore, the size of the Leishmania DTH response was greater in GBV-C viremica subjects (6.8 mm) compared to non-viremic subjects (3.3 mm; p= 0.0054). There findings suggest that GBV-C virus may promote a type 1 immune response that could influence the outcome of Leishmania infection

Estudo sobre a participação de rodedores na cadeia de transmissão de Leishmania infantum (Protozoa: Trypanosomatidae) no Rio Grande do Norte

American visceral leishmaniasis is a zoonosis caused by Leishmania infantum and transmitted by the bite of the sand flies Lutzomia longipalpis.The main domestic reservoir is the dog, while foxes and opposums are the known wild reservoirs. However, identification of natural infections with L. infantum in rodents appears for need of investigating the participation of these rodents how source of infection of the parasite. In the present work the Leishmania infantum infection was investigated in rodents captured in Rio Grande do Norte, aiming at to offer subsidies to the understanding of the epidemic chains of LVA in the State. Thirteen Galea spixii were distributed in four groups, being G1 the group control with four animals and the others, G2, G3 and G4, with three animals each. Those animals were intraperitoneally inoculated with 107 promastigotas of L. infantum and accompanied for, respectively, 30, 90 and 180 days. Weekly the animals were monitored as for the corporal weight and rectal temperature. At the end of each stipulated period the animals were killed. Blood were used for determination of the parameters biochemical and haematological, PCR, ELISA, microscopic examination and cultivation in NNN medium. Liver, spleen and lymph node were used in Giemsa-stained impression and cultivation in NNN medium. Liver and spleen fragments were still used in PCR and histopathological, respectively. At the same time 79 rodents of the species Rattus rattus, Bolomys lasiurus, Oligoryzomys nigripis, Oryzomys subflavus and Trichomys apereoides were captured in the Municipal districts of Brejinho, Campo Grande, Coronel Ezequiel, Passa e Fica and Vázea for identification of natural infection with L. infantum. Evidence of infection was checked by direct examination of Giemsa-stained impression of liver, spleen and blood and culture of these tissues in NNN medium. Antibodies were researched by ELISA. They were not found differences among the weigh corporal final, rectal temperature and biochemical and haematological parameters of the Galea spixii controls and infected. The rectal temperature of the animals varied from 36OC to 40OC. For the first time values of the haematocrit (33,6% to 42,8%), hemoglobin (10,2 to 14,5g/dl), erythrocyts number (4,67x106 to 6,90x106/mm3), total leukocytes (0,9x103 to 9,2x103/mm3), platelets (49x103 to 509x103/mm3) total proteins (1,56 to 6,06 g/dl), albumin (1,34 to 3,05 g/dl) and globulins (0,20 to 3,01 g/dl) of the Galea spixii were determined. The lymphocytes were the most abundant leucocytes. Infection for L. infantum was diagnosed in two animals euthanasied 180 days after the infection. In one of the animals was also identified antibodies anti-Leishmania. The parasite was not found in none of the five other species of rodents captured. Galea spixii are resistant to the infection for L. infantum and they are not good models for the study for visceral leishmaniose, although they can act as infection sources. More studies are necessary to determine the paper of the rodents in the epidemic chain of transmission of the visceral leishmaniose in the State of Rio Grande do Norte

Resposta imune à saliva de flebotomíneos (diptera: psychodidae) em indivíduos residentes em áreas urbana e periurbana no nordeste do brasil

Visceral leishmaniasis (VL) has a wide geographical distribution in tropical and subtropical areas of the planet, which is a protozoan parasite of the genus Leishmania. This pathogen is transmitted to the host through the sandflies bite, with its saliva, the immune response that leads to both. In the state of Rio Grande do Norte, 85% of the sand flies captured is Lutzomyia longipalpis, but the second most abundant, Lutzomyia evandroi, it deserves emphasis because its wide distribution and eclectic behavior. The exposure of people living in endemic areas for the insect vector VL greatly increases the chances of infection. This study aimed to evaluate aspects of the epidemiological profile of VL in endemic areas of human and nonendemic in the metropolitan area of Natal, as well as verify the abundance and seasonal fluctuations of sandflies species in two counties endemic for VL. Were collected in the municipalities of Nísia Floresta, Parnamirim, São Gonçalo do Amarante and Macaíba, of which groups of females were separated for further dissection of the salivary glands and identification of species. The blood samples used were from individuals of two Natal s districts where it has never been reported cases of VL and neighborhoods of Parnamirim applicants who present cases of VL. In the municipality of Nísia Floresta, the most abundant species was L. evandroi with 38.39%, followed by L. longipalpis with 36.22%, L. walkeri 19.67% L. lenti 3.81%, L. wellcomei 1.39% and L. whitmani 0.52%. Already in Parnamirim the proportions were L. walkeri with 73.15%, L. evandroi with 10.55%, L. wellcomei 7.63%, L. longipalpis 6.37%, L. whitmani 1.46%, L. sordellii 0.52%, L. intermedia 0.21 and L. shanonni 0.1%. In both municipalities was observed higher abundance of species distributed in the initial months of the year, as February and March. The study showed that no difference in exposure to the vector of VL among individuals from endemic and non endemic area for this disease. But there are differences in exposure between individuals of L. longipalpis and L. evandroi, confirming the great powers of the first vector. It was also characterized as predominant phenotype in the population of endemic areas who had negative serologic responses to antigens of Leishmania and result in negative Montenegro skin test (DTH), indicating that much of the population hasn t been bitten by infected insects

Estado nutricional e efeito da vitamina A na resposta imune frente à infecção por Leishmania Infantum

Nutritional status is an important determinant to the response against Leishmania infection, although few studies have characterized the molecular basis for the association found between malnutrition and the disease. Vitamin A supplementation has long been used in developing countries to prevent mortality by diarrheal and respiratory diseases, but there are no studies on the role of vitamin A in Leishmania infection, although we and others have found vitamin A deficiency in visceral Leishmaniasis (VL). Regulatory T cells are induced in vitro by vitamin A metabolites and are considered important cells implicated T CD4+ cell suppression in human VL. This work aimed to examine the correlation of nutritional status and the effect of vitamin A in the response against Leishmania infantum infection. A total of 179 children were studied: 31 had active VL, 33 VL history, 44 were DTH+ and 71 were DTH- and had negative antibody to Leishmania (DTH-/Ac-). Peripheral blood monuclear cells were isolated in a subgroup of 10 active VL and 16 DTH-/Ac- children and cultivated for 20h under 5 different conditions: 1) Medium, 2) Soluble promastigote L. infantum antigens (SLA), 3) All-trans retinoic acid (ATRA), 4) SLA + ATRA and 5) Concanavalin A. T CD4+CD25highFoxp3+, T CD4+CD25-Foxp3- and CD14+ monocytes were stained and studied by flow cytometry for IL-10, TGF-β and IL-17 production. Nutritional status was compromised in VL children, which presented lower BMI/Age and retinol concentrations when compared to healthy controls. We found a negative correlation between nutritional status (measured by BMI/Age and serum retinol) and anti-Leishmania antibodies and acute phase proteins. There was no correlation between nutritional status and parasite load. ATRA presented a dual effect in Treg cells and monocytes: In healthy children (DTH-/Ac-), it induced a regulatory response, increasing IL-10 and TGF-β production; in VL children it modulated the immune response, preventing increased IL-10 production after SLA stimulation. Furthermore, we found a positive correlation between BMI/Age and IL-17 production and negative correlation between serum retinol and IL-10 and TGF-β production in T CD4+CD25highFoxp3+ cells after SLA stimulus. Our results show a potential dual role of vitamin A in the immune system: improvement of regulatory profile during homeostasis and down modulation of IL-10 in Treg cells and monocytes during symptomatic VL. Therefore, the use of vitamin A concomitant to VL therapy might improve recovery from disease status in Leishmania infantum infection

Anticorpos anti-trypanosoma cruzi como preditivos de infecção por leishmania infantum

Leishmania infantum and Trypanosoma cruzi are trypanosomatids of medical importance and are, respectively, the etiologic agents of visceral leishmaniasis (VL) and Chagas disease (CD) in Brazil. People infected with L. infantum or T. cruzi may develop asymptomatically, enabling the transmission of pathogens through blood transfusion and / or organs. The assessment of the infection by T. cruzi is included among the tests performed for screening blood donors in Brazil, however, there is no availability of tests for Leishmania. Serological tests for T. cruzi are very sensitive, but not specific, and may have cross-reactions with other microorganisms. Thus, the aim of this study was to determine the prevalence of Leishmania infection in blood donors and assess whether the serological test for T. cruzi detect L. infantum. Among the 300 blood samples from donors, discarded in 2011, 61 were T. cruzi positive, 203 were from donors with other infections and 36 were from handbags with low blood volume, but without infection. We also assessed 144 samples from donors without infections and able to donate blood, totaling 444 subjects. DNA was extracted from blood samples of all to perform quantitative PCR (qPCR) to detect Leishmania DNA. The buffy coat obtained from all samples was grown in Schneider medium supplemented and NNN. All samples were evaluated for the presence of anti-Leishmania antibody. The serological results indicate a percentage of 22% of Leishmania infection in blood samples obtained from discarded bags. A total of 60% of samples positive in ELISA for T. cruzi were negative by IFI, used as confirmatory test, ie 60% false positive for Chagas. Among these samples false positive for Chagas, 72% were positive by ELISA for Leishmania characterizing the occurrence of cross reaction between serologic assays. Of the 300 cultures performed, 18 grew parasites that were typed by qPCR and specific isoenzymes, found the species Leishmania infantum crops. Among the 18 cultures, 4 were purged from scholarships for low volume and all negative serology blood bank, thus demonstrating that there is a real risk of Leishmania transmission via transfusion. It is concluded that in an area endemic for leishmaniasis in Brazil, serological diagnosis performed to detect infection by T. cruzi among blood donors can identify infection by L. infantum and although cause false positive for Chagas, this cross-reactivity reduces the risk of Leishmania infection via blood transfusion, since tests are not applied specific detection of the parasite. In this way, there remains the need to discuss the implementation of a specific serological screening test for Leishmania in endemic countries such as Brazil

Flebotomíneos (Diptera: Psychodidae) em um sistema agroflorestal da Região Metropolitana de Natal, Rio Grande do Norte, Brasil

Sandflies stand out as important vectors of leishmaniasis. The females need to ingest blood meals, enabling them to transmit protozoa of the genus Leishmania, which may give rise to visceral leishmaniasis (VL) or American tegumentary leishmaniasis (ATL), in addition to transmitting other parasites. Leishmaniasis are important infirmities, distributed worldwide, whose infection results from the interaction of reservoir animals, the vector insect, parasitic protozoa and the healthy host. In the state of Rio Grande do Norte (RN), Brazil, these insects are important transmitters of VL, which usually presents in the most serious form. It occurs mainly in metropolitan areas, with the dog as its main reservoir and Lutzomyia longipalpis as the vector. ATL is most present in the highland areas of the state. In addition to hematophagia, engaged in by the females, both sexes need to ingest carbohydrates, which are essential to the sand flies energy requirements and may interfere in the development of Leishmania. The aim of this study was to determine the occurrence and abundance of sand flies in different environments on the farm belonging to the Empresa de Pesquisas Agropecuárias do RN (Institute of Agricultural Research of RN), in the municipality of Parnamirim, in order to relate this occurrence with climatological and biological references and eating habits. Three consecutive monthly collections were carried out with CDC traps in a fragment of the Atlantic Forest, in a residence, on a goat breeding farm and on cashew, dwarf and giant coconut, mango, banana, eucalyptus, acacia and bean plantations. A total of 1241 sandflies from eight species (Lutzomyia evandroi, Lutzomyia longipalpis, Lutzomyia shannoni, Lutzomyia sordellii Lutzomyia walkeri, Lutzomyia wellcomei, Lutzomyia whitmani, and Lutzomyia intermedia) were collected, most in the forest environment. L. longipalpis, the main VL transmitter, was confirmed as a species adapted to anthropic environments, whereas others such as L. wellcomei, the vector of ATL, occurred predominantly in forests. Carbohydrate characterization of the sand flies and plants of the region demonstrated that a number of exotic plants such as hay and eucalyptus may play some role in the adaptation of these species to modified environments. Breeding in laboratory showed a mean biological cycle of 53.5 days from egg to adulthood for L. shannoni and the possibility of diapause behavior in L. wellcomei. This study serves as a source of information that may contribute to the epidemiological vigilance of tegumentary and visceral leishmaniasis in the state, given that it analyzes the bioecology of transmitting species, as well as their potential to adapt to new environments

Comportamento adaptativo de Lutzomyia longipalpis (Diptera: Psychodidae) às áreas urbanas da cidade de Teresina-PI.

The sand fly Lutzomyia longipalpis (Diptera: Psychodidae) is currently appointed as the main vector of visceral leishmaniasis in the Americas. The growth of cities in areas originally endemics to American Visceral Leishmaniasis (AVL) resulted in the spread of the disease at the same time that observed the adaptation of this species to the urban environment.Changes in behavior of L.longipalpis that enabled the adapt to increasing losings of biodiversity, as well as the frequent exposure of the vector to insecticides evident in urban areas, could justify the increasing population of the species and consequently the spread of disease for these environments .Thus, we selected sixty houses spread among three areas with increasing stages of occupation of an area endemic for AVL in Teresina-PI. We evaluated the correlation between the density of L.longipalpis captured and different aspects, such as population density of animals, vegetation cover and socio-economic aspects in each house. In addition to the correlations, the feeding preference of the vector between the predominant plant species in the neighborhoods, as well as the presence of metabolic mechanisms of resistance among the captured insects were tested. The results showed that over the growing occupations, represented by three areas, L.longipalpis demonstrate its adaptive nature through an apparent opportunistic behavior in relation to sources of carbohydrates and blood. On the evolutionary point of view, this behavior may have favored its vector competence in urban areas among the limited presence of food sources, as well as in various environments encountered.

Antígenos da forma amastigota de Leishmania chagasi identificados por dupla varredura da biblioteca de cDNA

Control of human visceral leishmaniasis in endemic regions is hampered in part by the lack of knowledge with respect of the role reservoirs and vector. In addition, there is not yet an understanding of how non-symptomatic subclinical infection might influence the maintenance of infection in a particular locality. Of worrisome is the limited accessibility to medical care in places with emerging drug resistance. There is still no available protective vaccine either for humans or other reservoirs. Leishmania species are protozoa that express multiple antigens which are recognized by the vertebrate immune system. Since there is not one immunodominant epitope recognized by most hosts, strategies must be developed to optimize selection of antigens for prevention and immunodiagnosis. For this reason, we generated a cDNA library from the intracellular amastigote form of Leishmania chagasi, the causative agent of South American visceral leishmaniasis. We employed a two-step expression screen of the library to systematically identify T and T-dependent B cell antigens. The first step was aimed at identifying the largest possible number of clones producing an epitope-containing polypeptide with a pool of sera from Brazilians with documented visceral leishmaniasis. After removal of clones encoding heat shock proteins, positive clones underwent a second step screen for their ability to cause proliferation and IFN-γ responses of T cells from immune mice. Six unique clones were selected from the second screen for further analysis. The clones encoded part of the coding sequence of glutamine synthetase, transitional endoplasmic reticulum ATPase, elongation factor 1γ, kinesin K-39, repetitive protein A2, and a hypothetical conserved protein. Humans naturally infected with L. chagasi mounted both cellular and antibody responses to these protein Preparations containing multiple antigens may be optimal for immunodiagnosis and protective vaccines against Leishmania

Associação entre fatores nutricionais e a resposta frente à infecção por Leishmania chagasi

Leishmania chagasi infection presents a wide spectrum of clinical outcomes, ranging from asymptomatic self resolving infection to disease, visceral leishmaniasis (VL). The exact mechanisms that lead the evolution of infection to disease are not understood. It is believed that malnutrition is a risk factor associated with VL development, although there are few human studies in the area. We aimed to assess the nutritional factors associated with the response to L. chagasi infection in Rio Grande do Norte. The study was conducted from December 2006 to January 2008. 149 children were assessed: 20 active VL cases, 33 children with VL history, 40 DTH+ asymptomatic children and 56 DTH-. Nutritional status was assessed using z scores for Weight/Age, Weight/Height, Height/Age, Body Mass Index (BMI), and mid-upper arm circumference/height (MUAC/height). Vitamin A status was determined by serum retinol concentrations and the modified-relative-dose-esponse test (MRDR). Breastfeeding time and birth weight were also evaluated. VL children presented compromised nutritional status when compared to the other groups using BMI and MUAC/age, with means -1,53 ± 1,10 and -1,48 ± 1,28 z scores, respectively (ANOVA, p < 0,05). VL children also showed lower vitamin A levels: 43% presented serum retinol < 20 µg/dL and 15% MRDR > 0,060. Birth weight was inverserly associated with the risk to belong the VL group (β = -0,00; OR = 0,84; 95% CI 0,73 - 0,99; p = 0,047), whereas more breastfeeding time was directly associated with the risk to belong to the DTH+ group (β = 0,02; OD = 1,16; 95% CI 1,01 - 1,33; p = 0,036). The nutritional variables evaluated were associated with the response to the L. chagasi infection, with malnutrition and compromised vitamin A status as markers of children who present with VL. Higher birth weight was associated with protection to disease, and higher breastfeeding time was associated with increased likelihood of an asymptomatic infection. The results show that modifiable nutritional aspects in the study population are associated with the response to the L. chagasi infection

Determinantes envolvidos na resposta imune celular humana à infecção por Leishmania infantum chagasi

Visceral leishmaniasis (VL) is a disease caused by protozoa of the Leishmania donovani complex, whose infection has clinical spectrum ranging from asymptomatic infection to active disease characterized by fever, cachexia, hepatosplenomegaly, and immunosuppression. The healing or protective immunity require an antigen-specific type 1. The Montenegro skin test (DTH) has been interpreted as a marker of protective immunity. However, there is no known correlation between the DTH response to type 1 and DTH and immunity of type 1 are maintained in the long term. Thus, a longitudinal study of 8 years, nested in a cohort family held in Brazil, documented the status of DTH and cytokine production by peripheral blood mononuclear cells in response to antigen-specific stimulation. This study was the interdisciplinary approach of physicians, biochemists, nutritionists, veterinary medicine, biology and statistics. The results show that 46.2% of subjects were analyzed DTH positive at baseline. The prevalence of positive and DTH induration size increased with age (p = 0.0021). 15.7% of individuals positive DTH "retro-converted" the negative and 50.4% (64) of individuals negative DTH became positive. The size of DTH induration was correlated significantly with the antigen-induced production of IFN-γ (r = 0.6186, p = 0.0001). IL-6 was secreted at higher levels in peripheral blood mononuclear cells of individuals who "retro-converted" DTH positive to negative than individuals who remained stable DTH status (p = 0.005). Thus, IFN-γ produced by peripheral blood mononuclear cells, may be a surrogate marker for protective immunity instead of the DTH response. In addition, differences in innate immune response may determine whether individuals maintain or eliminate the infection by Leishmania infantum chagasi in asymptomatic patients

Estudo do Cultivo de dois clones de Escherichia coli recombinantes (eIF, LACK) para a Expressão de Antígenos da Leishmania chagasi

With advent of the technology of the recombinant DNA, the recombinant protein expression becomes an important tool in the studies of the structure, function and identification of new proteins, mainly with therapeutical purposes. The Escherichia coli has been procarioto predominant in the studies of genetic engineering due to wealth of information regarding its metabolism. Despite the expressivo advance of the studies of molecular biology and the immunology of the infections, it does not exist, currently, no prophylactic drug capable to prevent calazar. Of this form, it exists a great necessity of specific antigen identification for the vaccine development and kits for disgnostic against the visceral Leishmaniose. In this context, this work objectified to study the recombinant antigen expression of the Leishmania chagasi during the culture of Escherichia coli in shaker. A first set of assays was carried through with the objective of if knowing the kinetic behavior of the growth of two clones recombinant proteins (eIF, LACK) in two different compositions of culture medium (2xTY, TB) supplemented by antibiotics, without IPTG addition. In the second stage of the assays, the procedure of induction for IPTG was carried through, in order to verify the influence of the composition of the ways tested in the expression them recombinant proteins. On the basis of the gotten results, can be observed that the high complexity of culture medium favored the kinetic one of growth of clones recombinant (eIF, LACK), however, to if to deal with the assays submitted to the procedure of induction for IPTG, the raised complexity of culture medium did not favor the expression of recombinant proteins. On the other hand, they had been gotten resulted positive for all clones recombinant (eIF, LACK) tested, confirmed through the eletroforético profile

Influência das condições de cultivo na produção de antígenos recombinantes de Leishmania i. chagasi utilizando Escherichia coli M15 cultivada em incubador rotativo e biorreator

Escherichia coli has been one of the most widely used hosts in recombinant protein production, in both laboratory and industrial scale since the advent of recombinant DNA technology. Despite the substantial progress of studies on the molecular biology and immunology of infections, there is currently no medication-based prophylaxis capable of preventing leishmaniasis. As such, there is a great need to identify specific antigens for the development of vaccines and diagnostic kits against visceral leishmaniasis. Thus, the primary goal of the present study is to assess the influence of cultivation conditions on the production of Leishmania chagasi antigens, carried out in a rotating incubator and bioreactor. To that end, several assays were conducted to evaluate the kinetic behavior of antigens (648, 503) of Leishmania. i. chagasi in two different compositions of media (2xTY, TB), with and without an inducer. In order to improve expression, assays were performed in a benchtop bioreactor using the best conditions obtained in a rotating incubator, in addition to assessing the influence of stirring speed. Results show that high complexity of the cultivation medium favored kinetic growth of clones (648, 503). However, in assays submitted to induction by IPTG, this elevated complexity did not promote the expression of recombinant proteins. Expression of antigens 648 and 503 exhibited behavior associated with growth and, in terms of location, proteins 648 and 503 are intracellularly stored. Lactose may be the most adequate inducer in protein expression, when considering factors, cost, toxicity and stability. Elevated stirring may increase cell growth in clone 53, although it may not result in high concentrations for the protein of interest. On the other hand, positive results were obtained for all recombinant clones (648, 503) tested, confirmed by the electrophoretic profile

Perfil imunológico associado à susceptibilidade, resistência e cura na infecção por Leishmania ( Leishmania ) infantum

Visceral Leishmaniasis (VL) is endemic in Brazil and the northeast region had the highest incidence of the disease , despite, in the last 30 years, it has spread to all geographic regions of the country. Leishmania infantum is the m ain etiological agent of VL in Latin America, Europe and North Africa. However, not all infected individuals develop the disease; in fact, the majority present spontaneous re solution of infection without symptoms. The evaluation of the immunological profil e has been mostly conducted stimulating, with Leishmania spp. antigen, peripheral blood mononuclear cells isolated from subjects with VL. These studies showed that VL patients had an inhibition of both, lymphocyte proliferation and proinflammatory response to Leishmania spp. antigen. Our study aimed to evaluate the immune response in active LV, cured post treatment and asymptomatic infection. To reach this aim, we analyzed immunophenotypic features related to activation, Treg and memory lymphocytes, by flow cytometry, as well as, evaluation of cytokine production, in ex vivo or in whole blood culture. In active VL volunteers, a longitu dinal study was conducted with reassessment at 4 and 14 months after clinical cure. The control group included individuals th at live d in endemic region and were either Positive Control, consisting of individuals with positive anti - L eishmania spp. serology and/or positive PCR for Leishmania  spp. and Negative Control composed by individuals with negative anti - Leishmania antibodie s serology and negative PCR for Leishmania . During VL, CD4 lymphocytes showed greater activation and memory profile s and were the major source of cytokines in culture when compared to CD8 lymphocytes , and these were not Leishmania specific. There were act ivated lymphocytes during VL (CD4 + CD69 + :4.9%) when compared to control groups, Positive (CD4 + CD69 + :1.96%, p=0.0045) and Negative (CD4 + CD69 + :1.35%, p=0.006), on the other hand, this was non - specific activation. The lymphocyte activation profile remain ed el evated even 14 months post treatmen t. A fter clinical cure , the activation was Leishmania specific (CD4 + CD25 + absence of SLA: 8.4%, and presence of SLA: 10.7% p=0.0279). CD8 + CD25 + lymphocytes were able to produce Leishmania specific IFN - γ in both, Positive Controls (absence of SLA 5.2% and presence of SLA: 9.5%, p=0.0391) and Cured 4 month (absence of SLA: 3.9%; presence of SLA: 10.7% p=0.0098). Whole blood culture cells, of VL patients, were able to produce IFN - γ, by SLA stimulation (absence of SLA: 28.0 pg ∕mL, and presence: 44.3 pg∕mL p=0.0020) as well as recovered groups (absence of SLA 2.3 pg∕mL and presence of SLA 139.8 pg∕mL, p=0.0005). However, the high level of IL - 10 seem ed to inhibit pro - inflammatory activity of IFN - γ and TNF - α during symptomatic dis ease . Unlike other pro - inflammatory cytokines, active VL group d id not produce Leishmania specific IL - 2 (absence of SLA 2.4 pg∕mL and presence of SLA: 2.6 pg∕mL). Based on these data we conclude that the restoration of lymphocyte activation and decreased i n IL - 10 Leishmania specific production were related to a protective immune profile.