Resumo. Foi realizado um estudo epidemiológico para avaliação dos fatores determinantes para a transmissão da LTA. O estudo foi realizado na área urbana, periurbana e rural da cidade de Itambaracá, incluindo a Vila Rural e as localidades de Porto Almeida e São Joaquim do Pontal, as duas últimas próximas à área impactada pelas hidrelétricas de Canoas I e II, no rio Paranapanema. Capturou-se de fevereiro de 2004 a junho 2006, 3.187 flebotomíneos. As espécies predominantes foram Nyssomyia neivai (34,36%), Pintomyia pessoai (32,57%), Migonemya migonei (11,61%), Nyssomyia whitmani (8,82%) e Pintomyia fischeri (2,73%) todas elas com capacidade de transmissão da doença. Para Ny neivai, houve predominância de machos e para as outras espécies vetoras, as fêmeas prevaleceram, com diferença estatística significante (p<0,001). As espécies mais abundantes, segundo o índice de abundância das espécies padronizado, foram Ny neivai, Pi. pessoai, Ny whitmani, Br. brumpti, Mg migonei e Pi fischeri. As maiores freqüências e diversidade das espécies foram encontradas na localidade de Porto Almeida (PA), seguida por São Joaquim do Pontal (SJP). Ny neivai apresentou médias sazonais mais elevadas e na distribuição mensal, picos em fevereiro, maio e outubro. A maior média horária desta espécie foi registrada das 19:00 às 20:00 h (1,2) mas mostrou-se ativa das 23:00 às 10:00 h da manhã. A prevalência de Acs da classe IgG para LTA em RIFI, na população humana foi de 6,73%, nos caninos de 1,75% e nos equídeos de 16,0 %. Em EIE/ELISA, a prevalência para os caninos foi de 16,49%. Na PCR, todos os cremes leucocitários correspondentes à soros reagentes a Acs da classe IgG (humanos), e Ac totais (eqüinos e caninos) foram negativos para Leishmania sp. No inquérito sócio econômico, verificou-se que a população entrevistada em sua maioria pertence à etnia branca e à faixa etária de 30-50 anos, recebem em torno de dois salários mínimos, possuem ocupações voltadas à agricultura e pecuária, nível de escolaridade fundamental (1º grau), habita casas de alvenaria, possuem animais domésticos com seus abrigos próximos às casas. Realizam atividades de trabalho e lazer em áreas de risco em proximidades às matas, áreas com animais silvestres e domésticos e com presença de matéria orgânica, que são fatores atrativos aos flebotomíneos. A população entrevistada, de uma forma geral, desconhece os aspectos da transmissão da LTA. Observou-se em todas as localidades amostradas a presença de flebotomíneos, com predominância de cinco espécies com capacidade vetorial de agentes da LTA, destacando-se Ny. Neivai , incluindo os peridomicílios. Houve transmissão ativa da doença, ocorrendo dois casos notificados e autóctones, em área de baixa freqüência de espécimes. A soroprevalência na população humana (6,73%) foi considerável, embora seja uma área de baixa ocorrência da doença. Nas localidades onde havia um foco mais recente da doença, em SJP, os cães e eqüinos apresentaram sorologia positiva. A alta prevalência de reagentes na sorologia observada em eqüídeos sugere que estes animais poderiam ser usados como sentinela para a detecção precoce da circulação do agente da LTA nas áreas estudadas. / Abstract. An epidemiological study was carried out in the urban area of Itambaracá and rural areas of the same municipality, including Vila Rural, Porto Almeida and São Joaquim do Pontal, to investigate the phlebotomine fauna and behavioral aspects of its species and also some determining factors of the transmission of American cutaneous leishmaniasis (ACL). The last two localities mentioned are situated close to the areas impacted by the Canoas I and II hydroelectric plants on the Paranapanema river. A total of 3,187 phlebotomines were captured between February 2004 and June 2006. Nyssomyia neivai predominated (34.36%), followed by Pintomyia pessoai (32.6%), Migonemyia migonei (11.6%), Nyssomyia whitmani (8.8%) and Pintomyia fischeri (2.7%), all of them implicated in the transmission of the disease. Regarding Ny. neivai, males predominated over females which may indicate the proximity of its breeding places to the capture sites. As for all the other vector species, the females were predominant, with significant statistical differences (p < 0,001). Ny neivai, Pi. pessoai, Ny. whitmani, Br. brumpti, Mg. migonei and Pi. fischeri presented the highest values for the species standardized abundance index. The greatest frequencies and diversity of the species were found at Porto Almeida (PA), a locality situated close to a well preserved forest fragment and secondly at São Joaquim do Pontal (SJP), with degraded forest. Of all the species, Ny. neivai presented the highest monthly averages with the main peak occurring in successive months of April. Its greatest hourly average was registered at 19:00-20:00 hours, but the species presented activity from 23:00 p.m. to 10:00 a.m. The prevalence of Ac IgG class, serum-reagents for Leishmania obtained with RIFI from the human, canine and equine populations was 6.73% (20/297), 1.75% (5/285) and 16% (8/50), respectively. A prevalence of 16.4% was observed by the EIE/ELISA test only for the canine population. The buffy-coat of all the reagent human and animal blood samples was tested by the polymerase chain reaction (PCR) to investigate the presence of Leishmania, with negative results. In the socioeconomic survey, it was found that the majority of the people interviewed belonged to the white ethnic and 30-50 year-old groups, earning two minimum salaries, exercising occupations associated with agriculture and livestock breeding, of fundamental school level, inhabiting brick houses and keeping pets and domestic animals in and around them. They exercise their work and leisure activities in risk areas close to forest fragments, thus in areas containing both wild and domestic animals with the presence of organic matter, which attract the sandflies. The population interviewed was, in general, ignorant of the facts involved in ACL transmission. Sandflies were observed at all the sites sampled, with the predominance of five vectors of the ACL agents, with Ny. neivai, the main vector in all three areas, also being captured in the peridomiciles. Active transmission of the disease, with two cases registered, was observed at sites with very low frequency of this sandfly. The serum prevalence in the human population (6.73%) was considerable, despite this being an area of few registered ACL cases. In the localities in which there was a more recent focus of the disease, in SJP, the dogs and equines presented positive serology. The high prevalence of equine serum reagents suggests that these animals could be used as sentinels for the early detection of the presence of the ACL agent in the area studied.

Introdução: Micose fungoide poiquilodérmica (MFp) é uma variante clínica de micose fungoide (MF). É mais indolente e caracterizada pela presença da poiquilodermia. As metaloproteinases (MMP) e seus inibidores específicos TIMP (Tissue Inhibitors of Metaloproteinases) estão envolvidos na oncogênese. Especificamente as MMP2 e MMP9 e seus inibidores, TIMP-2 e TIMP-1, respectivamente, foram relacionados ao prognóstico em tumores. Poucos trabalhos estudaram MMP e nenhum estudou a ação dos TIMP na MF. Objetivos: avaliar a relação entre MMP2 e MMP9 e seus inibidores TIMP2 e TIMP1 e a agressividade da MF e descrever a casuística de micose fungoide poiquilodérmica no ambulatório de linfomas cutâneos da Divisão de Clínica Dermatológica do Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo. Métodos: análise retrospectiva de 54 casos de MFp, sendo 25 de MFp localizada 14 de MFp generalizada e 15 de MFp mista. Para análise das MMP e TIMP, os grupos de MFp foram comparados com 7 amostras de pele normal (PN), 10 casos de MF clássica inicial (MFi), 9 casos de MF tumoral não-transformada (MFT nt) e 10 de MF tumoral transformada (MFT t). Resultados: A proporção de mulheres: homens foi 2,44. MFp apresentou maior tempo entre os primeiros sintomas e o diagnóstico. MFpG apresentou maior prevalência de lesões do tipo pitiríase liquenoide crônica (PLC) (79%). Houve alta prevalência de MF hipocromiante (62%) no grupo MFp mista. A histologia da MFp apresentou características típicas de MF e, adicionalmente, atrofia, telangectasias e derrame pigmentar, específicos da forma poiquilodérmica. Na imuno-histoquímica predominou o fenótipo CD3+, CD4+, CD7-, CD8- em todos os grupos, e MFp apresentou significantemente menor predomínio do fenótipo CD8+ que o grupo MFi. O grupo MFpG apresentou baixa positividade para pesquisa de clonalidade T da pele (12,5%). A MMP2 esteve mais presente na epiderme em MFi e MFp relativamente a MFT. Na derme superficial, os grupos MFi e MFp marcaram mais MMP2 que a pele normal, mas sem diferença estatística entre eles. Não houve diferença estatística em MMP2 na derme profunda entre os grupos. À zimografia, houve maior atividade de MMP2 ativa no grupo MFTt. Não houve expressão de TIMP-2 pela epiderme da pele normal. Os grupos MFi e MFp marcaram TIMP-2 na epiderme de forma semelhante, porém menos que os grupos MFT. Na derme superficial, não houve diferença estatística entre os grupos MFi e MFp. TIMP-2 foi mais expresso na derme profunda dos dois grupos de MFT comparativamente a todos os outros grupos. Na epiderme e na derme superficial, MMP9 foi mais expressa no grupo MFi comparativamente a MFp. Na derme profunda, a expressão de MMP9 foi maior nos grupos MFT, seguido por MFi e, por último, MFp. A atividade de MMP9 foi maior no grupo MFT não transformada comparativamente aos outros grupos. TIMP-1, ne epiderme e na derme superficial e na derme profunda foi mais expresso no grupo MFi, comparativamente aos outros grupos. Discussão: MFpG apresentou mais lesões tipo PLC e a forma mista, lesões hipocrômicas. A histologia da MFp foi semelhante à descrita previamente na literatura, mas a baixa positividade de CD8 difere de relatos prévios. A MMP2 pareceu ser um marcador de atividade para MF, principalmente quando a sua presença por imunohistoquímica foi associada a dados de zimografia. A expressão de MMP9 nas amostras foi compatível com os dados prévios de literatura, tendo sido mais expressa nas formas mais agressivas de MF e, histologicamente, mais localizada nos locais de maior atividade do tumor. TIMP-1 foi expresso de forma análoga à MMP9, conforme descrito previamente na literatura. TIMP-2, por sua vez, seguiu o padrão de distribuição de MMP2. No entanto, não foi expresso pela pele normal e foi mais expresso pelos grupos de MFT, o que não ocorreu com a MMP2 na imuno-histoquímica. Conclusões: A expressão de MMP e TIMP correlacionou-se com o local de maior atividade linfocitária e com a agressividade da MF. A atividade da MMP2 e MMP9 foi maior nos grupos MFT comparativamente aos grupos mais indolentes. Separar os casos de MFp de acordo com suas apresentações localizadas, generalizada e mista foi relevante do ponto de vista clínico, laboratorial e evolutivo / Introduction: poikilodermatous mycosis fungoides (pMF) is a clinical variant of mycosis fungoides (MF). It is more indolent than classic MF and is characterized by the presence of poikiloderma. The matrix metalloproteinases (MMPs) and their specific inhibitors TIMP (Tissue Inhibitors of Metalloproteinases) are involved in oncogenesis. Specifically, MMP2 and MMP9 and their inhibitors, TIMP-2 and TIMP-1, respectively, have been related to prognosis in tumors. There are few studies on MMP and none on the role of TIMPs in MF. Objectives: To evaluate if there is a relationship between the presence and activity of MMP2 and MMP9 and their inhibitors TIMP2 and TIMP1, and the aggressiveness of MF. To describe a casuistic of poikilodermatous mycosis fungoides in an outpatient clinic in the Dermatological Division of Hospital das Clinicas of University of Sao Paulo Medical School. Methods: Retrospective analysis of 54 cases of pMF, this included 25 localized pMF (LpMF), 14 generalized pMF (GpMF) and 15 mixed pMF. For the analysis of MMPs and TIMPs, the pMF groups were compared with 7 normal skin samples (NS), 10 cases of initial classical MF (cMF), 9 cases of non-transformed tumor MF (nt MFT) and 10 transformed tumor MF (t MFT). Results: The proportion of women : men was 2.44. The pMFs groups showed a longer period of time from the first symptoms to the diagnosis than the cMF group. The GpMF group had a higher incidence of pityriasis lichenoides chronica-like lesions (PLC) (79%) than the other groups. There was a high incidence of hypopigmented MF (62%) in the mixed pMF group. Histology showed typical characteristics of MF and, additionally, atrophy, telangiectasia and pigmentary alterations compatible with pMF. At immunohistochemistry the cases were predominantly CD3+, CD4+, CD7-, CD8- phenotype in all groups, and the pMF groups had a significantly lower prevalence of CD8+ phenotype than the cMF group. The GPMF group showed low positivity for clonality of the T-cell receptor at the T skin (12.5%) compared to the other groups. The MMP2 was more present in the epidermis for the cMF and pMF groups compared to MFT. In the superficial dermis, the cMF, LpMF and GpMF groups showed more MMP2 than normal skin, however there was no statistical difference between the three groups. There was no statistical difference in the presence of MMP2 in the deep dermis between the groups. The zymography showed higher MMP2 activity in the MFT group. There was no TIMP-2 expression by the normal epidermis. The epidermis of cMF and pMFs groups marked TIMP-2 in a similar way, but at a lower intensity than the MFT groups. In the superficial dermis, there was no statistical difference between the cMF and pMFs groups. TIMP-2 was more expressed in the deep dermis of the two MFT groups compared to all of the other groups. In the epidermis and superficial dermis, the MMP9 was more expressed in cMF compared to pMF groups. In the deep dermis, MMP9 expression was higher in the MFT groups, followed by cMF and finally pMF. The MMP9 activity was higher in the nt MFT group compared to other groups. TIMP-1, in epidermis, superficial dermis and deep dermis was more expressed in the cMF group compared to other groups. Discussion: The study confirmed that the pMF is an indolent form of MF and the time period between the symptoms and the diagnosis in pMF was longer than in classical MF. There were clinical differences amongst the groups of pMF. The GpMF group had a higher prevalence of PLC-like lesions than the mixed form of pMF, which had more hypochromic lesions. Histology of pMF was similar to descriptions provided in other case studies. However, the low CD8 positivity differs from previous reports. The MMP2 appeared to be a marker of activity for MF in our work, especially when their presence by immunohistochemistry was associated with the enzyme activity. The expression of MMP9 in our samples was consistent with previous data from other case studies, being more expressed in the most aggressive forms of MF and histologically more localized in most active sites of the tumor. TIMP-1 was expressed in an analogous manner to MMP9, as previously described in the literature. TIMP-2, in turn, followed the distribution pattern of MMP2. However, it was not expressed by normal skin and was more expressed by the MFT group, which did not occur with the MMP2 in immunohistochemistry. Conclusions: The expression of MMP and TIMP was correlated with the location of higher lymphocyte activity and with the aggressiveness of MF. The activity of MMP2 and MMP9 was higher in the MFT groups than the more indolent groups. It was important to split the pMF cases according to their presentation (GpMF, LpMF and mix pMF) from a clinical, laboratory and prognostic point of view

Introdução: As leishmanioses são doenças que se apresentam como um sério obstáculo ao desenvolvimento socioeconômico. A leishmaniose visceral americana (LVA) tem como agente etiológico a Leishmania (Leishmania) infantum chagasi, como reservatório doméstico principal o cão e os vetores são flebotomíneos (Diptera:Psychodidae). Objetivo: investigar a vulnerabilidade do município de São Paulo para a ocorrência de LVA, utilizando os parâmetros de casos humanos notificados, presença do vetor e infecção em reservatórios domésticos. Método: A área de estudo compreende a região dos distritos administrativos do Anhanguera, Jaraguá e Perus (município de São Paulo) que incluem o Parque do Anhanguera e o Parque estadual do Jaraguá. A justificativa para o estudo nessa área é o eixo viário oeste-leste de expansão da LVA no estado de São Paulo. Foram levantados notificações humanas de LVA entre os anos de 2007 a 2014, para análise espacial pelos programas SaTScan e Terra View, num total de 140 e nenhuma dessas é considerada autóctone. Realizamos levantamento entomológico nos parques Anhanguera e Jaraguá. A análise estatística foi realizada pelos cálculos da Média geométrica de Williams, pelo índice de diversidade de Shannon-Weaver (H) e a dominância pelo índice de Pielou (J) também foi calculado o índice de Abundância de espécies padronizado (IAEP). Realizamos inquérito canino amostral e obtivemos 126 amostras de sangue canino que foram submetidas aos testes EIE. Foram colhidas amostras de Swab bucal do cão sororreagente e contactantes, as quais foram analisadas por PCR (RFLP), juntamente ás amostras de coágulos sanguíneos dos cães dos parques Anhanguera e Jaraguá num total de 31 amostras. Resultado: A análise espacial mostrou Clusters nos distritos do Brás e Guaianases (p<0,05) e, a espaço temporal em Raposo Tavares e Parelheiros (p<0,05). Os distritos do Carrão, Cachoeirinha e José Bonifácio formaram clusters secundários (p>0,05). Foram capturados 3660 espécimes de flebotomíneos identificadas 16 espécies de flebotomíneos e as mais frequentes na amostra foram: Pintomyia fischeri (82,46 por cento ), Psathyromyia pascalei (5,57 por cento ), Migonemyia migonei (5.25 por cento ) e Expapillata firmatoi (3.22 por cento ). Dentre as espécies capturadas, está uma fêmea de espécie similar à Lutzomyia longipalpis. O inquérito canino sorológico apontou um sororreagente no distrito de Perus. Do total de 31 amostras analisadas por PCR, 10 foram positivas para Leishmania sp. Foi possível chegar á espécie em seis delas, três positivas para a espécie L. (V.) shawi ou L (V.) lainsoni e três para L. (L.) infantum chagasi. Conclusões: A análise espacial das notificações de LVA humana dá direcionamento para investigações de uma possível transmissão autóctone no município. Há elos epidemiológicos que sugerem que as leishmanioses tegumentar e visceral caninas se tratam de doenças emergentes no município. Possivelmente, os casos caninos encontrados na região de estudo sejam parte da disseminação do eixo oeste-leste da leishmaniose visceral no Estado de São Paulo, pelo eixo viário. Palavras-chave: Leishmaniose visceral canina; Leishmaniose tegumentar. / Introduction: The leishmaniases are diseases which are a serious obstacle to social economic development. American visceral leishmaniasis (AVL) has Leishmania (Leishmania) infantum chagasi as its etiological agent, the domestic dog is its main reservoir and sand flies (Diptera, Psychodidae) its vectors. Objective: to investigate the vulnerability of São Paulo municipality to the occurrence of AVL, using the parameters of notified human cases, the presence of the vector and infection in domestic reservoirs. Method: The study area covers the region of the distinct administrative districts of Anhanguera, Jaraguá and Perus (São Paulo municipality) which include the Parque do Anhanguera and the Parque Estadual do Jaraguá. The reason for undertaking the study in this area is the existence of the roads linking the west and east of São Paulo state along which AVL has expanded eastwards. A survey was made of the 140 human cases (none of which was considered autochthonous) of AVL notified from 2007 to 2014, for the spatial analysis of the SaTScan and Terra View programs. An entomological survey was conducted in the two parks mentioned. The statistical analyses of the insects were undertaken using Williams geometrical means, Shannon-Weavers diversity index (H), Pielous evenness index (J) and the standardized species abundance index (SSAI). A canine sampling inquiry was conducted and 126 blood samples were submitted to the Enzyme-Linked Immunosorbent Assay (ELISA) test. A total of five samples of mouth swabs of the serum positive dog and four of its contacts and also blood clot samples of a total of 26 dogs from the two parks were analyzed by PCR (RFLP). Results: The spatial analysis showed clusters in the Brás and Guaianases (p<0.05) districts and the temporal/spatial analysis in Raposo Tavares and Parelheiros (p<0.05) districts. The Carrão, Cachoeirinha and José Bonifácio districts formed secondary clusters (p>0.05). A total of 3,660 specimens of sand flies were collected, representing 16 species, Pintomyia fischeri (82.46 per cent ), Psathyromyia pascalei (5.57 per cent ), Migonemyia migonei (5.25 per cent ) and Expapillata firmatoi (3.22 per cent ) being the most frequent. One female of a species similar to Lutzomyia longipalpis was captured. The canine serological enquiry showed one reagent in the Perus district. Of the total of 31 samples analyzed by PCR, 10 were positive for Leishmania sp., among which six were identified, three as species L. (V.) shawi/L (V.) lainsoni and three as L. (L.) infantum chagasi. Conclusions: The spatial analysis of the AVL human cases notified suggests the direction to be followed for future investigation of a possible autochthonous transmission in the municipality. There are epidemiological links which suggest that the canine cutaneous and visceral leishmaniases are emergent diseases in the municipality. Possibly the canine cases of visceral leishmaniasis found in this study region are part of its dissemination along the west-east axis of the diseases expansion in the state of São Paulo along the highways.

Cyanobacteria are common inhabitants of freshwater lakes and reservoirs throughout the world. Under favourable conditions, certain cyanobacteria can dominate the phytoplankton within a waterbody and form nuisance blooms. Case reports and anecdotal references dating from 1949 describe a range of illnesses associated with recreational exposure to cyanobacteria: hay fever-like symptoms, pruritic skin rashes and gastro-intestinal symptoms (the latter probably related to ingestion of water) are most frequently reported. Some papers give convincing descriptions of allergic responses to cyanobacteria; others describe more serious acute illnesses, with symptoms such as severe headache, pneumonia, fever, myalgia, vertigo and blistering in the mouth. A U.S. coroner recently found that a teenage boy died as a result of accidentally ingesting a neurotoxic cyanotoxin from a golf course pond; this is the first recorded human fatality attributed to recreational exposure to cyanobacteria. One of the main public health concerns with exposure to freshwater cyanobacteria relates to the understanding that some blooms produce toxins that specifically affect the liver or the central nervous system. The route of exposure for these toxins is oral, from accidental or deliberate ingestion of recreational water, and possibly by inhalation. Cyanobacterial lipopolysaccharides (LPS) are also reported to be putative cutaneous, gastrointestinal, respiratory and pyrogenic toxins. The aims of this project were to enhance the understanding of public health issues relating to recreational exposure to cyanobacteria by conducting epidemiological and laboratory-based toxicology studies. A prospective cohort study of 1,331 recreational water users was conducted at various sites in southern Queensland, the Myall Lakes area of New South Wales, and central Florida. The study design sought to make improvements over previously published epidemiological studies, in that an unexposed group was recruited from cyanobacteria-free waters, cyanobacterial toxins were measured in site water samples, and respondents were asked to rate the severity of reported symptoms. This study has shown an increased likelihood of symptom reporting amongst bathers exposed to high cyanobacterial cell density (measured by total cell surface area) compared to those exposed to low cyanobacteria-affected waters. Mild respiratory symptoms appear to be the predominant symptom category. A clinical dermatology study to examine delayed-contact hypersensitivity reactions to cyanobacterial extracts was conducted. The study groups were 20 patients presenting for diagnostic skin patch testing at the Royal Brisbane Hospital's dermatology outpatient clinic; a convenience sample of 20 individuals was recruited from outside the hospital as a control group. One patient developed unequivocal reactions to several cyanobacteria extracts, with no dose-response pattern seen, indicating that the reactions were allergic in nature. A mouse model of delayed-contact hypersensitivity, the mouse ear swelling test, has demonstrated that the purified toxin cylindrospermopsin, a highly water-soluble compound, is capable of producing cutaneous injury. Encrusting lesions were seen on abdominal skin during the induction phase of these experiments. Delayed-contact hypersensitivity reactions were also demonstrated with this toxin. LPS from two non-axenic cyanobacterial samples - Cylindrospermopsis raciborskii and Microcystis aeruginosa - were extracted and purified. Thermoregulation studies were performed using a mouse model of rectal temperature measurement. Separate groups of mice were injected with these LPS extracts at two dose levels. Thermoregulation studies were also conducted with purified cyanobacterial toxins: microcystin-LR and cylindrospermopsin and anatoxin-a; cyanobacterial LPS samples purified by collaborators in Adelaide were also investigated for thermoregulation potential. These experiments have shown that the LPS extracts are weakly active when compared with responses to much lower doses of Escherichia coli LPS. Cylindrospermopsin also produces hypothermic responses in mice - comparable to pyrexia in larger mammals - although at a later stage than was seen with LPS extracts. Supplementing the experimental observations of the toxicology and immunotoxicology of cyanobacterial LPS are insights gained from beyond the cyanobacteria literature on the mechanisms of toxicity of different LPS structures. Cyanobacterial LPS was initially suggested to be toxic in the 1970s, at a time when lipid A, the active moiety of LPS, was thought to be identical across all Gram-negative bacteria. More recent work raises questions about the attribution of cutaneous and gastrointestinal illness to cyanobacterial LPS, with the understanding that some bacterial lipid A structures are LPS antagonists, LPS is not toxic by the oral route, and LPS is not reported as a toxin in the clinical dermatology literature. Gut-derived lipopolysaccharides, however, exert potent synergistic effects with a variety of xenobiotic hepatotoxins, and the well-known shock-like syndromes associated with severe cylindrospermopsin and microcystin poisoning deserve further scrutiny from the perspective of immunotoxicology.

Cyanobacteria are common inhabitants of freshwater lakes and reservoirs throughout the world. Under favourable conditions, certain cyanobacteria can dominate the phytoplankton within a waterbody and form nuisance blooms. Case reports and anecdotal references dating from 1949 describe a range of illnesses associated with recreational exposure to cyanobacteria: hay fever-like symptoms, pruritic skin rashes and gastro-intestinal symptoms (the latter probably related to ingestion of water) are most frequently reported. Some papers give convincing descriptions of allergic responses to cyanobacteria; others describe more serious acute illnesses, with symptoms such as severe headache, pneumonia, fever, myalgia, vertigo and blistering in the mouth. A U.S. coroner recently found that a teenage boy died as a result of accidentally ingesting a neurotoxic cyanotoxin from a golf course pond; this is the first recorded human fatality attributed to recreational exposure to cyanobacteria. One of the main public health concerns with exposure to freshwater cyanobacteria relates to the understanding that some blooms produce toxins that specifically affect the liver or the central nervous system. The route of exposure for these toxins is oral, from accidental or deliberate ingestion of recreational water, and possibly by inhalation. Cyanobacterial lipopolysaccharides (LPS) are also reported to be putative cutaneous, gastrointestinal, respiratory and pyrogenic toxins. The aims of this project were to enhance the understanding of public health issues relating to recreational exposure to cyanobacteria by conducting epidemiological and laboratory-based toxicology studies. A prospective cohort study of 1,331 recreational water users was conducted at various sites in southern Queensland, the Myall Lakes area of New South Wales, and central Florida. The study design sought to make improvements over previously published epidemiological studies, in that an unexposed group was recruited from cyanobacteria-free waters, cyanobacterial toxins were measured in site water samples, and respondents were asked to rate the severity of reported symptoms. This study has shown an increased likelihood of symptom reporting amongst bathers exposed to high cyanobacterial cell density (measured by total cell surface area) compared to those exposed to low cyanobacteria-affected waters. Mild respiratory symptoms appear to be the predominant symptom category. A clinical dermatology study to examine delayed-contact hypersensitivity reactions to cyanobacterial extracts was conducted. The study groups were 20 patients presenting for diagnostic skin patch testing at the Royal Brisbane Hospital's dermatology outpatient clinic; a convenience sample of 20 individuals was recruited from outside the hospital as a control group. One patient developed unequivocal reactions to several cyanobacteria extracts, with no dose-response pattern seen, indicating that the reactions were allergic in nature. A mouse model of delayed-contact hypersensitivity, the mouse ear swelling test, has demonstrated that the purified toxin cylindrospermopsin, a highly water-soluble compound, is capable of producing cutaneous injury. Encrusting lesions were seen on abdominal skin during the induction phase of these experiments. Delayed-contact hypersensitivity reactions were also demonstrated with this toxin. LPS from two non-axenic cyanobacterial samples - Cylindrospermopsis raciborskii and Microcystis aeruginosa - were extracted and purified. Thermoregulation studies were performed using a mouse model of rectal temperature measurement. Separate groups of mice were injected with these LPS extracts at two dose levels. Thermoregulation studies were also conducted with purified cyanobacterial toxins: microcystin-LR and cylindrospermopsin and anatoxin-a; cyanobacterial LPS samples purified by collaborators in Adelaide were also investigated for thermoregulation potential. These experiments have shown that the LPS extracts are weakly active when compared with responses to much lower doses of Escherichia coli LPS. Cylindrospermopsin also produces hypothermic responses in mice - comparable to pyrexia in larger mammals - although at a later stage than was seen with LPS extracts. Supplementing the experimental observations of the toxicology and immunotoxicology of cyanobacterial LPS are insights gained from beyond the cyanobacteria literature on the mechanisms of toxicity of different LPS structures. Cyanobacterial LPS was initially suggested to be toxic in the 1970s, at a time when lipid A, the active moiety of LPS, was thought to be identical across all Gram-negative bacteria. More recent work raises questions about the attribution of cutaneous and gastrointestinal illness to cyanobacterial LPS, with the understanding that some bacterial lipid A structures are LPS antagonists, LPS is not toxic by the oral route, and LPS is not reported as a toxin in the clinical dermatology literature. Gut-derived lipopolysaccharides, however, exert potent synergistic effects with a variety of xenobiotic hepatotoxins, and the well-known shock-like syndromes associated with severe cylindrospermopsin and microcystin poisoning deserve further scrutiny from the perspective of immunotoxicology.

Cyanobacteria are common inhabitants of freshwater lakes and reservoirs throughout the world. Under favourable conditions, certain cyanobacteria can dominate the phytoplankton within a waterbody and form nuisance blooms. Case reports and anecdotal references dating from 1949 describe a range of illnesses associated with recreational exposure to cyanobacteria: hay fever-like symptoms, pruritic skin rashes and gastro-intestinal symptoms (the latter probably related to ingestion of water) are most frequently reported. Some papers give convincing descriptions of allergic responses to cyanobacteria; others describe more serious acute illnesses, with symptoms such as severe headache, pneumonia, fever, myalgia, vertigo and blistering in the mouth. A U.S. coroner recently found that a teenage boy died as a result of accidentally ingesting a neurotoxic cyanotoxin from a golf course pond; this is the first recorded human fatality attributed to recreational exposure to cyanobacteria. One of the main public health concerns with exposure to freshwater cyanobacteria relates to the understanding that some blooms produce toxins that specifically affect the liver or the central nervous system. The route of exposure for these toxins is oral, from accidental or deliberate ingestion of recreational water, and possibly by inhalation. Cyanobacterial lipopolysaccharides (LPS) are also reported to be putative cutaneous, gastrointestinal, respiratory and pyrogenic toxins. The aims of this project were to enhance the understanding of public health issues relating to recreational exposure to cyanobacteria by conducting epidemiological and laboratory-based toxicology studies. A prospective cohort study of 1,331 recreational water users was conducted at various sites in southern Queensland, the Myall Lakes area of New South Wales, and central Florida. The study design sought to make improvements over previously published epidemiological studies, in that an unexposed group was recruited from cyanobacteria-free waters, cyanobacterial toxins were measured in site water samples, and respondents were asked to rate the severity of reported symptoms. This study has shown an increased likelihood of symptom reporting amongst bathers exposed to high cyanobacterial cell density (measured by total cell surface area) compared to those exposed to low cyanobacteria-affected waters. Mild respiratory symptoms appear to be the predominant symptom category. A clinical dermatology study to examine delayed-contact hypersensitivity reactions to cyanobacterial extracts was conducted. The study groups were 20 patients presenting for diagnostic skin patch testing at the Royal Brisbane Hospital's dermatology outpatient clinic; a convenience sample of 20 individuals was recruited from outside the hospital as a control group. One patient developed unequivocal reactions to several cyanobacteria extracts, with no dose-response pattern seen, indicating that the reactions were allergic in nature. A mouse model of delayed-contact hypersensitivity, the mouse ear swelling test, has demonstrated that the purified toxin cylindrospermopsin, a highly water-soluble compound, is capable of producing cutaneous injury. Encrusting lesions were seen on abdominal skin during the induction phase of these experiments. Delayed-contact hypersensitivity reactions were also demonstrated with this toxin. LPS from two non-axenic cyanobacterial samples - Cylindrospermopsis raciborskii and Microcystis aeruginosa - were extracted and purified. Thermoregulation studies were performed using a mouse model of rectal temperature measurement. Separate groups of mice were injected with these LPS extracts at two dose levels. Thermoregulation studies were also conducted with purified cyanobacterial toxins: microcystin-LR and cylindrospermopsin and anatoxin-a; cyanobacterial LPS samples purified by collaborators in Adelaide were also investigated for thermoregulation potential. These experiments have shown that the LPS extracts are weakly active when compared with responses to much lower doses of Escherichia coli LPS. Cylindrospermopsin also produces hypothermic responses in mice - comparable to pyrexia in larger mammals - although at a later stage than was seen with LPS extracts. Supplementing the experimental observations of the toxicology and immunotoxicology of cyanobacterial LPS are insights gained from beyond the cyanobacteria literature on the mechanisms of toxicity of different LPS structures. Cyanobacterial LPS was initially suggested to be toxic in the 1970s, at a time when lipid A, the active moiety of LPS, was thought to be identical across all Gram-negative bacteria. More recent work raises questions about the attribution of cutaneous and gastrointestinal illness to cyanobacterial LPS, with the understanding that some bacterial lipid A structures are LPS antagonists, LPS is not toxic by the oral route, and LPS is not reported as a toxin in the clinical dermatology literature. Gut-derived lipopolysaccharides, however, exert potent synergistic effects with a variety of xenobiotic hepatotoxins, and the well-known shock-like syndromes associated with severe cylindrospermopsin and microcystin poisoning deserve further scrutiny from the perspective of immunotoxicology.

Cyanobacteria are common inhabitants of freshwater lakes and reservoirs throughout the world. Under favourable conditions, certain cyanobacteria can dominate the phytoplankton within a waterbody and form nuisance blooms. Case reports and anecdotal references dating from 1949 describe a range of illnesses associated with recreational exposure to cyanobacteria: hay fever-like symptoms, pruritic skin rashes and gastro-intestinal symptoms (the latter probably related to ingestion of water) are most frequently reported. Some papers give convincing descriptions of allergic responses to cyanobacteria; others describe more serious acute illnesses, with symptoms such as severe headache, pneumonia, fever, myalgia, vertigo and blistering in the mouth. A U.S. coroner recently found that a teenage boy died as a result of accidentally ingesting a neurotoxic cyanotoxin from a golf course pond; this is the first recorded human fatality attributed to recreational exposure to cyanobacteria. One of the main public health concerns with exposure to freshwater cyanobacteria relates to the understanding that some blooms produce toxins that specifically affect the liver or the central nervous system. The route of exposure for these toxins is oral, from accidental or deliberate ingestion of recreational water, and possibly by inhalation. Cyanobacterial lipopolysaccharides (LPS) are also reported to be putative cutaneous, gastrointestinal, respiratory and pyrogenic toxins. The aims of this project were to enhance the understanding of public health issues relating to recreational exposure to cyanobacteria by conducting epidemiological and laboratory-based toxicology studies. A prospective cohort study of 1,331 recreational water users was conducted at various sites in southern Queensland, the Myall Lakes area of New South Wales, and central Florida. The study design sought to make improvements over previously published epidemiological studies, in that an unexposed group was recruited from cyanobacteria-free waters, cyanobacterial toxins were measured in site water samples, and respondents were asked to rate the severity of reported symptoms. This study has shown an increased likelihood of symptom reporting amongst bathers exposed to high cyanobacterial cell density (measured by total cell surface area) compared to those exposed to low cyanobacteria-affected waters. Mild respiratory symptoms appear to be the predominant symptom category. A clinical dermatology study to examine delayed-contact hypersensitivity reactions to cyanobacterial extracts was conducted. The study groups were 20 patients presenting for diagnostic skin patch testing at the Royal Brisbane Hospital's dermatology outpatient clinic; a convenience sample of 20 individuals was recruited from outside the hospital as a control group. One patient developed unequivocal reactions to several cyanobacteria extracts, with no dose-response pattern seen, indicating that the reactions were allergic in nature. A mouse model of delayed-contact hypersensitivity, the mouse ear swelling test, has demonstrated that the purified toxin cylindrospermopsin, a highly water-soluble compound, is capable of producing cutaneous injury. Encrusting lesions were seen on abdominal skin during the induction phase of these experiments. Delayed-contact hypersensitivity reactions were also demonstrated with this toxin. LPS from two non-axenic cyanobacterial samples - Cylindrospermopsis raciborskii and Microcystis aeruginosa - were extracted and purified. Thermoregulation studies were performed using a mouse model of rectal temperature measurement. Separate groups of mice were injected with these LPS extracts at two dose levels. Thermoregulation studies were also conducted with purified cyanobacterial toxins: microcystin-LR and cylindrospermopsin and anatoxin-a; cyanobacterial LPS samples purified by collaborators in Adelaide were also investigated for thermoregulation potential. These experiments have shown that the LPS extracts are weakly active when compared with responses to much lower doses of Escherichia coli LPS. Cylindrospermopsin also produces hypothermic responses in mice - comparable to pyrexia in larger mammals - although at a later stage than was seen with LPS extracts. Supplementing the experimental observations of the toxicology and immunotoxicology of cyanobacterial LPS are insights gained from beyond the cyanobacteria literature on the mechanisms of toxicity of different LPS structures. Cyanobacterial LPS was initially suggested to be toxic in the 1970s, at a time when lipid A, the active moiety of LPS, was thought to be identical across all Gram-negative bacteria. More recent work raises questions about the attribution of cutaneous and gastrointestinal illness to cyanobacterial LPS, with the understanding that some bacterial lipid A structures are LPS antagonists, LPS is not toxic by the oral route, and LPS is not reported as a toxin in the clinical dermatology literature. Gut-derived lipopolysaccharides, however, exert potent synergistic effects with a variety of xenobiotic hepatotoxins, and the well-known shock-like syndromes associated with severe cylindrospermopsin and microcystin poisoning deserve further scrutiny from the perspective of immunotoxicology.

Nous avons étudié le rôle de l'immunité innée de la peau lors de la transmission des Borrelia (agent infectieux de la borréliose de Lyme) par son vecteur, une tique dure du genre Ixodes. Nous avons montré que la salive de tique et la protéine salivaire Salp15 inhibent la réaction inflammatoire (production de chimiokines et de peptides antimicrobiens) des kératinocytes induite par Borrelia. Cet effet anti- « alarmine » de la salive de tique contribue probablement à créer un environnement cutané local favorable à la transmission de Borrelia. Nous avons montré que Borrelia induit également au niveau des fibroblastes cutanés la transcription de nombreux gènes proinflammatoires. Nous avons observé un effet toxique direct de la salive de tique sur les fibroblastes cutanés : cet effet dose-dépendant est de nature protéique mais non lié à la protéine Salp15. Ces résultats indiquent que les fibroblastes jouent un rôle important dans l’inflammation cutanée induite par Borrelia. / We studied the role of the skin innate immunity during the transmission of Borrelia (the infectious agent of Lyme borreliosis) by its vector, a hard tick belonging to the genus Ixodes. We showed that tick saliva and its protein Salp15 both inhibate Borrelia-induced inflammatory reaction of keratinocytes. The antialarmin effect of tick saliva ensure a favorable environment for Borrelia. We also showed that Borrelia induce a strong inflammatory response in dermal fibroblasts. We also demonstrate a dose-dependent lytic effect of tick salivary gland extracts on dermal fibroblasts and that this cytotoxic effect was of proteinaceous nature and not related to Salp15. These results indicate that dermal fibroblasts could be considered as central mediators in immune cell recruitment to the skin site of Borrelia invasion.

Made available in DSpace on 2011-05-04T12:36:21Z (GMT). No. of bitstreams: 0 Previous issue date: 2009 / Embora seja benigna, a leishmaniose tegumentar pode levar a quadros clínicos mais graves quando não tratada, com o acometimento das mucosas naso-buco-faríngeas e conseqüências psicológicas, sociais e econômicas sérias para esses indivíduos e suas famílias. Ressalta-se a necessidade de se buscar conhecer os fatores que levam pessoas a abandonarem o tratamento disponibilizado gratuitamente pelo Ministério da Saúde, assim como questionar sobre os possíveis fatores associados aos óbitos atribuídos a LT. Objetivo geral: analisar os fatores associados aos desfechos desfavoráveis do tratamento (abandono e óbito) por leishmaniose tegumentar na Região Sudeste, no período de 2002 a 2006. Material e método: O presente estudo é analítico de abordagem quantitativa, utilizou como fonte de dados as bases contendo os casos de leishmaniose tegumentar do Sistema de Informação de Agravos Notificáveis (SINAN).Foram incluídas as variáveis com um mínimo de 70 por cento de preenchimento. O processo de análise seguiu os seguintes passos: consolidação dos dados, descrição das variáveis, análise dos fatores associados aos desfechos desfavoráveis utilizando a regressão logística múltipla e as redes neurais artificiais e por fim a comparação entre essas duas ferramentas de análise. Resultados: Utilizando a regressão logística múltipla, foram associadas ao abandono do tratamento da leishmaniose tegumentar de acordo com a forma clínica: cutânea - deslocamento, raça negra e recidiva; mucosa - deslocamento do paciente em busca de tratamento e pacientes que moram em cidades com grandes populações, tendo sido identificada uma interação entre essas duas variáveis. Quanto aos óbitos atribuídos a LT, o modelo não se apresentou bem ajustado aos dados quando a idade acima dos 50 anos foi incluída, com isso foram associados: sexo e população pequena. A idade acima de 50 anos foi a única variável relacionada aos óbitos dos pacientes com LT que apresentaram lesões mucosas, mas o modelo não se mostrou bem ajustado. O modelo final com as Redes Neurais Artificiais adicionou as seguintes variáveis ao modelo de regressão logística: 1- abandono do tratamento da LT para aforma clínica: cutânea - municípios com população grande; mucosa - superdosagem e etnia negra. Quanto aos óbitos atribuídos a LT adicionou-se: forma cutânea - idade acima de 50 anos; forma mucosa - superdosagem, etnias indígenas e negra e escolaridade. Comparando os modelos, ambos mostram bom desempenho para praticamente todos os desfechos. Discussão e considerações finais: Os fatores encontrados para o abandono do tratamento e dos óbitos mostram nitidamente a presença das questões socioeconômicas, outras variáveis que não foram incluídas por falta de preenchimento adequado (como exemplo a ocupação dos pacientes) podem corroborar essa conclusão. Os modelos finais da regressão logística e das redes neurais mostraram bom desempenho (percentual de acerto acima de 90 por cento), variando a inclusão de algumas variáveis / indicadores. Este estudo indica a confecção de um programa (software) simples que possa ser utilizado pelas vigilâncias epidemiológicas para predizer os pacientes com maiores chances de abandonar o tratamento da LT. Os modelos para os óbitos precisam ser revistos, com a inclusão de novas variáveis na ficha de investigação e a melhoria do preenchimento. / Although usually not severe, cutaneous leishmaniasis (LT) can lead to more serious clinical pictures when it is not treated. The most common complications are mucocutaneous lesions in noses, mouth and pharynx causing serious psychological, social and economic consequences for the individuals and their families. It is necessary to study the factors that lead people to abandon the treatment, given for free Ministry of Health, as well as question about possible factors related to deaths due to LT. General Objective: analyze factors related to treatment unfavorable outcomes (abandonment and death) by cutaneous leishmaniasis on Southeast Region, from 2002 through 2006. Material and Method: The current study is a quantitative analytic approach, using data bases of cutaneous leishmaniasis from SINAN (Sistema de Informação de Agravos Notifiáveis) as data source. Cases with consistent filling of the selected variables were included. The process of analysis followed the given steps: data consolidation, variables description, analysis of the factors related to unfavorable outcomes using multiple logistic regression and artificial neural networks and finally comparison of both analytical tools. Results: Using multiple logistic regression, the clinical form was related to abandonment of cutaneous leishmaniasis treatment: cutaneous – displacement, black race and recurrence; mucosal – patient displacement searching treatment and patients living in huge population cities, with interaction been indentified between these two variables. As for the deaths assigned to LT, the model did not appear well adjusted to the data when the age above 50 years was included, therefore, were associated: gender and small population. Age was the only variable related to deaths of patients with LT that showed mucosal lesions, but the model was not significant. The final model with the Artificial Neural Networks added the following variables to the logistic regression model: 1 - abandonment of the treatment of LT for clinical forms: cutaneous - cities with large populations; mucosal - overdose and black ethnicity. As for deaths attributed to LT, added: cutaneous form - age over 50 years; mucosal form - overdose, black and indigenous ethnicity and education. Comparing the models, both show good performance for almost all outcomes. Discussion and final considerations: The factors found for the abandonment of treatment and deaths show the presence of socioeconomic issues. Other variables that were not included for lack adequate filling (for example the occupation of the patients) could corroborate for this conclusion. The final logistic regression models and neural networks showed good performance (percentage of accuracy above 90%), varying the inclusion of some variables / indicators. This study shows the preparation of a simple program (software) that can be used by the epidemiological surveillance to predict patients more likely to leave the treatment of LT. The models for the deaths need to be reviewed, with the inclusion of new variables in the research form and improvement of fulfillment.

Submitted by Adagilson Silva (adagilson@cpqam.fiocruz.br) on 2015-03-25T18:04:54Z No. of bitstreams: 1 2015morais-rcs.pdf: 1252683 bytes, checksum: 22ecfe946756c7d2dfa99794ad2d2d17 (MD5) / Approved for entry into archive by Adagilson Silva (adagilson@cpqam.fiocruz.br) on 2015-03-25T18:27:04Z (GMT) No. of bitstreams: 1 2015morais-rcs.pdf: 1252683 bytes, checksum: 22ecfe946756c7d2dfa99794ad2d2d17 (MD5) / Approved for entry into archive by Adagilson Silva (adagilson@cpqam.fiocruz.br) on 2016-02-19T13:29:39Z (GMT) No. of bitstreams: 1 2015morais-rcs.pdf: 1252683 bytes, checksum: 22ecfe946756c7d2dfa99794ad2d2d17 (MD5) / Made available in DSpace on 2016-02-19T13:29:39Z (GMT). No. of bitstreams: 1 2015morais-rcs.pdf: 1252683 bytes, checksum: 22ecfe946756c7d2dfa99794ad2d2d17 (MD5) / Fundação Oswaldo Cruz. Centro de Pesquisas Aggeu Magalhães / A Leishmaniose Tegumentar Americana (LTA) é causada por protozoários, do gênero Leishmania, envolvidos em um complexo ciclo biológico. No Brasil, sete espécies estão envolvidas com a etiologia da doença, distribuídas em todas as regiões geográficas e responsáveis por diferentes manifestações clínicas. Diante disso, o diagnóstico em conjunto com a identificação da espécie é de grande importância clínico-terapêutica. Este trabalho tem por objetivo avaliar a aplicabilidade da técnica de PCR quantitativa em tempo real (qPCR) para identificação de espécies de Leishmania envolvidas com a etiologia da LTA. Foram realizados ensaios de qPCR para padronização da Temperatura de melting (Tm) utilizando cepas de referência de diferentes espécies de Leishmania. Após o diagnóstico em amostras de sangue de animais domésticos utilizando a qPCR, as amostras positivas foram analisadas através de suas Tm, e os produtos de qPCR foram purificados e sequenciados. Dez amostras previamente caracterizadas por isoenzimas, também foram analisadas através da Tm. Ainda como teste de referência, foi padronizada uma Restriction Fragment Length Polymorphism (RFLP) utilizando as cepas de referência e testada nas amostras. Através da padronização da Tm das espécies, foram criados dois intervalos de análise: 1 (Tm = 78-79,99°C), que compreende: Leishmania (V.) braziliensis, Leishmania (V.) panamensis, Leishmania (V.) lainsoni, Leishmania (V.) guyanensis e Leishmania (V.) shawi; e 2 (Tm = 80-82,2°C), que compreende: Leishmania (V.) naiffi, Leishmania (L.) amazonensis e Leishmania (L.) mexicana. Um total de 223 amostras positivas foi analisado, destas, 58 incluídas no intervalo 1 e 165 no intervalo 2. O sequencimento de 94 destas amostras foi correspondente à L. (V.) braziliensis, L. (V.) panamensis e L. (V.) guyanensis. A RFLP em 173 amostras identificou 167 L. (V.) braziliensis, 05 L. (L.) mexicana e 01 L. (V.) panamensis. A análise da Tm das dez amostras caracterizadas por isoenzimas demonstrou 80 por cento de concordância (p = 0,6499) entre o padrão-ouro (isoenzimas) e os intervalos desenvolvidos neste estudo. Os resultados do sequenciamento foram concordantes com a qPCR em 43,62 por cento (n= 41) das amostras. A RFLP e qPCR tiveram 27,74 por cento (n= 48) de concordância. Análise estatística mostrou que a qPCR e sequenciamento são testes sem diferenças estatísticas significantes (p = 0,2566). Sendo assim, é possível concluir que a Tm da qPCR pode ser utilizada como um método para direcionamento da espécie de Leishmania presente na amostra em análise, sendo necessário a realização de métodos clássicos de caracterização, quando a precisão na identificação do parasito for crucial para implementação do tratamento, como em casos de resistência e/ou recidiva da doença. A utilização desta tecnologia pode ser ainda mais precisa quando for utilizada a metodologia com sondas desenhadas para cada espécie de Leishmania, sendo necessários estudos futuros para comprovar esta perspectiva (AU) / The American Cutaneous Leishmaniasis (ACL) is caused by protozoa of the genus Leishmania, involved in a complex biological cycle. In Brazil, seven species are involved in the etiology of the disease, distributed in all geographic regions and responsible for different clinical manifestations. Therefore, the diagnosis together with the identification of the species is of great clinical and therapeutic importance. This work aims to evaluate the applicability of the technique of real-time quantitative PCR (qPCR) for the identification of Leishmania species involved in the etiology of ACL. qPCR assays for standardizing the melting temperature (Tm) using reference strains of different species of Leishmania were performed. After the diagnosis on blood samples of domestic animals using the qPCR positive samples were analyzed by their Tm and qPCR products were purified and sequenced. Ten samples previously characterized by isoenzymes were also analyzed by Tm. Also as a reference test was standardized as Restriction Fragment Length Polymorphism (RFLP) using the reference strains and tested on samples. Through standardization of Tm species two ranges of analysis were created: 1 (Tm = 78-79,99°C), comprising: Leishmania (V.) braziliensis, Leishmania (V.) panamensis, Leishmania (V.) lainsoni, Leishmania (V.) guyanensis e Leishmania (V.) shawi; and, 2 (Tm = 80-82,2°C), comprising: Leishmania (V.) naiffi, Leishmania (L.) amazonensis e Leishmania (L.) mexicana. A total of 223 positive samples were analyzed, of these, 58 included in the range 1 and 165 in the range 2 to 94 and the sequence of these samples corresponded to L. (V.) braziliensis, L. (V.) panamensis and L. (V.) guyanensis. By RFLP in 173 samples were identified 167 L. (V.) braziliensis, 05 L. (L.) mexicana and 01 L. (V.) panamensis The analysis of Tm of the ten samples characterized by isoenzymes showed 80 per cent agreement (p = 0.6499) between the gold standard (isoenzymes) and intervals developed in this study. The sequencing results were concordant with qPCR in 43.62 per cent (n= 41) of the samples. The RFLP and qPCR had 27.74 per cent (n= 48) agreement. Statistical analysis showed that qPCR and sequencing tests are no statistically significant differences (p = 0.2566). Thus, we conclude that the Tm of qPCR can be used as a method for direction of Leishmania species present in the sample under analysis, performing classic methods of characterization is necessary when the accuracy in identifying the parasite is crucial for implementation treatment, such as in cases of resistance and / or recurrence of the disease. The use of this technology can be even more precise when the methodology with probes designed for every Leishmania species is used; future studies are needed to confirm this view (AU)