ABSTRACT Background: American cutaneous leishmaniasis is a commonly neglected, vector-borne tropical parasitic disease that is a major public health concern in Brazil. Leishmania (Viannia) braziliensis is the main species associated with the disease. Accurate diagnosis is based on epidemiological surveillance, clinical assessment, and laboratory testing. Leishmania (V.) braziliensis has been detected in several wild and synanthropic mammals. Their epidemiological role has not been entirely elucidated. This study aimed to assess potential L. braziliensis infections in asymptomatic domestic animals, by molecular and serological testing in endemic areas, in the metropolitan region of Recife. Methods: Blood samples and conjunctival fluids were collected from 232 animals (canids, felids, equines, and caprines) for the detection of L. braziliensis using molecular tests (conventional and real-time polymerase chain reaction [PCR and qPCR]). For immunological detection, blood samples from 115 dogs were assessed using enzyme-linked immunosorbent assay. Results: Real-time quantitative PCR showed positive results for blood and conjunctival samples in all investigated species. The results of the blood and conjunctival samples were 68.2% and 26.9% in Canis familiaris, 100% and 41.7% in Felis catus, 77.3% and 30.8% in Equus caballus/Equus asinus, and 50% and 33.3% in Capra hircus samples, respectively. Conclusions: Results from this study adds valuable information to our understanding of the role of asymptomatic domestic animals, L. braziliensis life cycle, and American cutaneous leishmaniasis in Northeast Brazil.

ABSTRACT This study was conducted to characterize the transmission cycle of the tegumentary leishmaniasis (TL) in an old colonization area at Pernambuco State, Brazil. The aims were to identify autochthonous cases, sandflies fauna, domestic animals as possible reservoir hosts and the Leishmania species involved in this endemic area. A total of 168 suspected human cases of TL and 272 domestic animals (canine, feline, equine, goat, and sheep) were included. The sandflies were captured and identified by species. Patients were predominantly male and the average age was 37+18.1 years old. Of 85 patients who had skin lesions, 25.6% of them had direct positive smears for TL and 34 isolates were identified as Leishmania (Viannia) braziliensis. The confirmation for TL diagnosed by molecular detection (PCR) was almost three times more sensitive than the direct test [p < 0.001; PR = 2.72] associated with clinical examination. The Kappa test on PCR between two different specimens, biopsy, and skin lesion swab was 60.8% (p < 0.001). More than 200 specimens of sandflies (80 males and 159 females) were captured and identified as Lutzomyia whitmani (99.6%) and Lu. evandroi (0.4%). The detection of L. (V.) braziliensis by Real-Time PCR in the blood of a captured fed female was positive in 59.3% of Lu. whitmani. Of the 272 domestic animals included, 61.76% were male (n = 168). Thirty-six animals (13.2%) had lesions compatible with TL (34 dogs, 1 cat and 1 sheep) and 3 of them, all dogs, had lesions on the snout, showing destruction of cartilage and mucosa. The study suggests the participation of domestic animals as possible reservoirs. However, further studies are necessary to better understand the transmission cycle and take recommended measures in order to control the disease.

Abstract INTRODUCTION: Brazil has a high number of cases of American cutaneous leishmaniasis (ACL) in the north and northeast regions. Therefore, continuous surveillance of environmental and socioeconomic factors in endemic areas is needed to develop strategic control measures. This study aimed to describe the clinical and epidemiological profiles of patients with ACL. METHODS: All patients were from the states of Amazonas and Pernambuco, and examinations were carried out between 2015 and 2018. All patients had a clinical and epidemiological history compatible with ACL after positive diagnostic tests. Information obtained from medical records included gender, employment activity, level of education, age, and number and sites of lesions. RESULTS: A total of 213 patients were included, of whom 30.98% were female and 69.02% were male. The main employment activity was agriculture (27.56%). The most common level of education was elementary (62.42%). The average age was approximately 39 years. The majority of the patients presented only with one lesion (54.87%), and legs/feet were the most commonly affected area (48.25%), followed by the arms/hands (44.75%). CONCLUSIONS: These data demonstrated that irrespective of the patients’ places of origin, interventions need to be focused on men of economically productive age, in view of the high risk of exposure to the vector in this group. Education activities need to be directed to farmers about the importance of protection against ACL vectors during work. Such information must also be directed to employers as a way of implementing and maintaining appropriate working conditions and stepping up vector control.

Several studies have described the use of non-invasive collection methods, mostly based on the detection of parasite DNA, for diagnosis. However, no Leishmania specimens have been isolated from saliva. Here, we report the first isolation of Leishmania braziliensis from the saliva of humans with cutaneous leishmaniasis but without lesions on their mucosa. The isolates were obtained from salivary fluid inoculated in hamsters and were tested by multilocus enzyme electrophoresis. Seven samples from 43 patients suspected of having the disease were identified for in vivo culture. These findings suggest that saliva is a clinical sample that allows the isolation of Leishmania sp.

ABSTRACT Visceral leishmaniasis, associated with HIV/AIDS coinfection, is becoming a more aggressive disease, complicating an accurate prognosis. A 21-year-old HIV-positive female presenting with clinical features of visceral leishmaniasis was enrolled in this study. Bone marrow cytology, Novy-MacNeal-Nicolle culture and kDNA PCR of peripheral blood were all positive. Typing methods, multilocus enzyme electrophoresis and ITS1-RFLP PCR of peripheral blood confirmed infection by Leishmania (L.) infantum chagasi . PCR has proved to be safer and more affordable than other characterization methods; ITS1-RFLP PCR can diagnose and type Leishmania spp. in both endemic and non-endemic areas, favoring the prognosis and allowing the appropriate treatment of patients.

Abstract In Brazil, visceral Leishmaniasis is caused by Leishmania chagasi. The development of cutaneous lesions in visceral leishmaniasis patients has been described in two different clinical contexts. Patients with compromised immunity can develop skin lesions as a direct consequence of a current visceral disease. Equally, patients with a history of kala-azar and progressive, immune improvement occasionally develop skin lesions as a consequence of immune reconstitution infl ammatory syndrome. These cases manifest in similar fashion to the classic form of post-kala-azar dermal Leishmaniasis. We describe different cases that exemplify these two clinical presentations.

A leishmaniose cutânea americana (LCA) é uma doença complexa com características clínicas e epidemiológicas que podem variar de região para região. De fato, pelo menos, sete diferentes espécies de Leishmania, incluindo Leishmania (Viannia) braziliensis, Leishmania (Viannia) guyanensis, Leishmania (Viannia) lainsoni, Leishmania (Viannia) naiffi, Leishmania (Viannia) shawi, Leishmania (Viannia) lindenbergi e Leishmania (Leishmania) amazonensis, têm sido incriminadas na etiologia da LCA no Brasil, e numerosas espécies de flebotomíneos do gênero Lutzomyia, foram considerados vetores suspeitos ou comprovados. Devido ao seu caráter focal, a compreensão da dinâmica da LCA a nível local é imprescindível para a implementação de medidas de controle eficazes. Este trabalho consiste de uma revisão narrativa sobre a epidemiologia da LCA em Pernambuco, nordeste do Brasil. Além disso, se enfatiza a necessidade de maior efetividade no diagnóstico, tratamento, controle e estratégias de prevenção para as populações afetadas. Este artigo fornecerá aos pesquisadores uma avaliação crítica da LCA em Pernambuco. Espera-se também contribuir com as autoridades de saúde pública no aprimoramento das estratégias atuais de controle da LCA nos níveis estaduais e nacional.

American cutaneous leishmaniasis (ACL) is a complex disease with clinical and epidemiological features that may vary from region to region. In fact, at least seven different Leishmania species, including Leishmania (Viannia) braziliensis, Leishmania (Viannia) guyanensis, Leishmania (Viannia) lainsoni, Leishmania (Viannia) naiffi, Leishmania (Viannia) shawi, Leishmania (Viannia) lindenbergi, and Leishmania (Leishmania) amazonensis, have been implicated in the etiology of ACL in Brazil, and numerous phlebotomine sandfly species of the genus Lutzomyia have been regarded as putative or proven vectors. Because ACL is a focal disease, understanding the disease dynamics at the local level is essential for the implementation of more effective control measures. The present paper is a narrative review about the ACL epidemiology in Pernambuco, northeastern Brazil. Furthermore, the need for more effective diagnosis, treatment, control and prevention strategies for the affected populations is highlighted. This paper will provide researchers with a critical appraisal of ACL in Pernambuco. Hopefully, it will also be helpful for public health authorities to improve current control strategies against ACL at the state and country levels.

INTRODUÇÃO: Desconhecida a realidade da leishmaniose visceral canina em Garanhuns, objetivou-se investigar a ocorrência de anticorpos antileishmania spp em cães domiciliados e semidomiciliados e os possíveis fatores de risco envolvidos. MÉTODOS: Em uma primeira etapa foram coletadas 256 amostras de sangue de cães que foram submetidas à reação de imunofluorescência indireta (RIFI) na diluição 1:40. Adicionalmente, 23 amostras positivas na RIFI foram testadas com um teste rápido imunocromatográfico. Em uma segunda etapa, novas amostras de sangue de 18 cães positivos na RIFI na primeira fase do estudo foram coletadas, retestadas pela RIFI (1:40 e 1:80) e, adicionalmente, pela reação em cadeia da polimerase para pesquisa de DNA de Leishmania infantum. Ademais, 16 dessas amostras foram retestadas pelo teste rápido imunocromatográfico. RESULTADOS: Na primeira etapa, 16% das amostras foram positivas na RIFI (1:40) e apenas três (13%) foram positivas no teste rápido imunocromatográfico. Na segunda etapa, 12 amostras foram positivas na RIFI na diluição 1:40 e sete também na diluição 1:80. Nenhuma amostra foi positiva na reação em cadeia da polimerase e no teste rápido imunocromatográfico. Sinais clínicos de leishmaniose visceral ocorreram em 4,9% dos cães positivos. Não houve diferença estatística entre idade, sexo e status clínico dos cães, porém entre seus locais de origem. CONCLUSÕES: Os cães domiciliados e semidomiciliados de Garanhuns apresentam anticorpos antileishmania spp, sendo, em sua grande maioria, assintomáticos.

INTRODUCTION: Considering the unknown situation regarding canine visceral leishmaniasis in Garanhuns, this study had the aim of investigating occurrences of anti-Leishmania spp antibodies in domesticated and partially domesticated dogs, and the possible risk factors involved. METHODS: In the first phase of the study, 256 blood samples were collected from dogs and subjected to the indirect fluorescent antibody test (IFAT) reaction at a dilution of 1:40. Additionally, 23 IFAT-positive samples were tested using an immunochromatographic dipstick test. In the second phase, new blood samples were collected from 18 dogs that were IFAT-positive in the first phase. These animals were retested using IFAT (1:40 and 1:80) and, additionally, by means of the polymerase chain reaction to investigate the Leishmania infantum DNA. Furthermore, 16 of these samples were retested using the immunochromatographic dipstick test. RESULTS: In the first phase of the study, 16% of the samples were IFAT-positive (1:40) and only three (13%) were positive in the immunochromatographic dipstick test. In the second phase, 12 samples were IFAT-positive at the dilution of 1:40, and seven were also positive at 1:80. None of the samples were positive in the polymerase chain reaction testing or in the immunochromatographic dipstick test. Clinical signs suggestive of visceral leishmaniasis were observed in 4.9% of the IFAT-positive dogs. There were no statistical differences in relation to age, sex or clinical status of the dogs, but there was a difference in relation to place of origin. CONCLUSIONS: The domesticated and partially domesticated dogs living in Garanhuns present anti-Leishmania spp antibodies, but are mostly asymptomatic.

O presente trabalho se propõe avaliar a atividade citotóxica do extrato etanólico bruto e da fração dos alcalóides totais (FAT) da planta Crotalaria retusa para células promastigotas metacíclicas de Leishmania chagasi. O estudo da cinética de extração por ultra-som para os alcalóides totais da Crotalaria retusa, tornou possível a otimização dos parâmetros de extração. Foi avaliada a ação leishmanicida da FAT da planta em estudo, a qual não mostrou atividade citotóxica em altas concentrações. Foi observado uma potente ação leishmanicida para os extratos etanólicos (10 e 30%) após a concentração de 5,6 mg/mL de Crotalaria retusa e do etanol presente na solução extrativa (10 e 30%) nas concentrações de 70 e 210 x 10-4 %, respectivamente. Estes resultados sugerem a citotoxicidade do extrato etanólico da Crotalaria retusa de 10 a 30% para células de Leishmania chagasi, associada possivelmente à concentração do etanol presente no extrato.

The purpose of the present work is to conduct an evaluation of the cytotoxicity of ethanol extracts and the total alkaloid fraction (TAF) from Crotalaria retusa for procyclic promastigotes cells of Leishmania chagasi. The kinetic study of extraction assisted by ultrasound of the total alkaloids present in Crotalaria retusa made it possible the optimization of the extraction parameters. It was evaluated the leishmanicide action of the TAF which did not show toxic activity for cells of the parasite in high concentrations. It was observed a powerful leishmanicide action of the ethanol extracts (10 and 30%) after the concentration of 5.6 mg/mL of Crotalaria retusa, and the ethanol present in the extractive solution (10 and 30%) in the concentration from 70 and 210 x 10-4 %, respectively. These results suggest that the cytotoxicity of the ethanol extract of Crotalaria retusa at 10 and 30% for cells of Leishmania chagasi, can be associated only to the concentration of the alcohol present in the extract.

A leishmaniose tegumentar americana incide em todas as regiões do Estado de Pernambuco, Brasil, onde se apresenta em expansão, com o surgimento de focos em novas áreas, como o Município de Moreno, na região metropolitana do Recife. O objetivo deste estudo foi avaliar a prevalência da infecção e identificar os casos autóctones de leishmaniose tegumentar americana. Em abril de 2006, realizou-se um inquérito populacional por meio da aplicação de um questionário e do teste de intradermoreação de Montenegro a 481 indivíduos do Engenho Pinto, dos quais 47% eram do sexo masculino e 74% menores de 30 anos. Cento e quarenta e quatro (30%) indivíduos apresentaram reação positiva ao teste. Dentre estes, cinco apresentaram lesões ativas, 41 foram curados após quimioterapia e um curado espontaneamente. A RIFI realizada nos soros de 89 indivíduos apresentou 36% de positividade. A alta prevalência aponta para a expansão epidemiológica da doença nessa área da região metropolitana, fato preocupante, pois não há medidas eficazes para o controle da leishmaniose tegumentar americana, a não ser uma conscientização ecológica para minimizar o risco de infecção.

American tegumentary leishmaniasis (ATL) is present in all regions of Pernambuco State, Brazil, where it is spreading, with the emergence of foci in new areas like the Municipality (County) of Moreno in Greater Metropolitan Recife. The objective of the current study was to assess the prevalence of infection and identify autochthonous cases of ATL. In April 2006 a population survey was performed using a questionnaire and the Montenegro skin test (MST) in 481 individuals from the Pinto Sugar Mill, of whom 47% were males and 74% were younger than 30 years. One hundred forty-four individuals (30%) showed a positive MST. Of these, five presented active lesions, 41 were cured after meglumine antimoniate treatment, and one showed a spontaneous cure. Indirect immunofluorescence (IIF) was performed on sera from 89 individuals and showed a 36% positive rate. The high ATL prevalence demonstrates the epidemiological spread of the disease in Greater Metropolitan Recife, a worrisome development since there are no effective measures for ATL control except ecological awareness raising to minimize the risk of infection.

A leishmaniose tegumentar americana incide em todas as regiões do Estado de Pernambuco, Brasil, onde se apresenta em expansão, com o surgimento de focos em novas áreas, como o Município de Moreno, na região metropolitana do Recife. O objetivo deste estudo foi avaliar a prevalência da infecção e identificar os casos autóctones de leishmaniose tegumentar americana. Em abril de 2006, realizou-se um inquérito populacional por meio da aplicação de um questionário e do teste de intradermoreação de Montenegro a 481 indivíduos do Engenho Pinto, dos quais 47% eram do sexo masculino e 74% menores de 30 anos. Cento e quarenta e quatro (30%) indivíduos apresentaram reação positiva ao teste. Dentre estes, cinco apresentaram lesões ativas, 41 foram curados após quimioterapia e um curado espontaneamente. A RIFI realizada nos soros de 89 indivíduos apresentou 36% de positividade. A alta prevalência aponta para a expansão epidemiológica da doença nessa área da região metropolitana, fato preocupante, pois não há medidas eficazes para o controle da leishmaniose tegumentar americana, a não ser uma conscientização ecológica para minimizar o risco de infecção.

American tegumentary leishmaniasis (ATL) is present in all regions of Pernambuco State, Brazil, where it is spreading, with the emergence of foci in new areas like the Municipality (County) of Moreno in Greater Metropolitan Recife. The objective of the current study was to assess the prevalence of infection and identify autochthonous cases of ATL. In April 2006 a population survey was performed using a questionnaire and the Montenegro skin test (MST) in 481 individuals from the Pinto Sugar Mill, of whom 47% were males and 74% were younger than 30 years. One hundred forty-four individuals (30%) showed a positive MST. Of these, five presented active lesions, 41 were cured after meglumine antimoniate treatment, and one showed a spontaneous cure. Indirect immunofluorescence (IIF) was performed on sera from 89 individuals and showed a 36% positive rate. The high ATL prevalence demonstrates the epidemiological spread of the disease in Greater Metropolitan Recife, a worrisome development since there are no effective measures for ATL control except ecological awareness raising to minimize the risk of infection.

O diagnóstico da leishmaniose cutânea americana é baseado na associação dos aspectos clínicos, epidemiológicos e laboratoriais. O presente estudo identificou a espécie de Leishmania circulante no Estado de Pernambuco, descreveu os aspectos clínico-epidemiológicos e diagnosticou a doença. Foram selecionados 19 pacientes apresentando lesão ativa e diagnosticados através de avaliação clínica e pelos exames laboratoriais que incluíram a pesquisa direta, cultivo in vitro, intradermorreação de Montenegro, imunofluorescência indireta e reação em cadeia de polymerase. A intradermorreação de Montenegro apresentou resultado positivo em 89% dos pacientes. A imunofluorescência indireta apresentou 79% de positividade, a pesquisa direta apresentou 58% e a reação em cadeia de polymerase 75% de positividade. Sete amostras de Leishmania (Viannia) braziliensis foram isoladas desses pacientes e caracterizadas através de anticorpos monoclonais específicos. Esses dados confirmam que é necessário a combinação de diferentes técnicas diagnósticas para se obter resultados eficientes e que, até o momento, a Leishmania (Viannia) braziliensis é a única espécie responsável pela infecção da leishmaniose cutânea americana em Pernambuco. Portanto, a identificação da espécie do parasito envolvido nos casos da doença no homem em uma área endêmica é essencial para o conhecimento dos aspectos clínicos e epidemiológicos, sobretudo para o diagnóstico e para o desenvolvimento da terapia e prognóstico da doença.

The diagnosis for American cutaneous leishmaniasis is based on an association of clinical, epidemiological and laboratory characteristics. The present study identified the circulating species of Leishmania in the State of Pernambuco, described its clinical-epidemiological characteristics and diagnosed the disease. Nineteen patients presenting active lesions who had been diagnosed through clinical evaluation and laboratory tests were selected. The tests included direct investigation, in vitro culturing, Montenegro skin test, indirect immunofluorescence and polymerase chain reaction. The Montenegro Skin Test showed positive results in 89% of the patients; indirect immunofluorescence, in 79%; direct investigation, in 58%; and polymerase chain reaction in 75%. Seven Leishmania (Viannia) braziliensis samples were isolated from these patients and were characterized by means of specific monoclonal antibodies. These data confirm that a combination of different diagnosis techniques is needed in order to obtain efficient results and that, so far, Leishmania (Viannia) braziliensis is the only species responsible for American cutaneous leishmaniasis infection in Pernambuco. Thus, it is essential to identify the parasite species involved in cases of human disease in an endemic area in order to determine the clinical and epidemiological characteristics, especially with regard to diagnosis, therapy development and disease prognosis.

Esse estudo teve como objetivo geral caracterizar a fauna de flebotomíneos envolvida em unidade de treinamento militar localizada na Zona da Mata do Estado de Pernambuco, Brasil, onde foram verificados casos humanos autóctones de leishmaniose tegumentar americana (LTA) associados a Leishmania (Viannia) braziliensis. A presença de 16 espécies de flebotomíneos no Centro de Instrução Militar Marechal Newton Cavalcanti indicou uma importante diversidade na área de estudo. Lutzomyia complexa constituiu a espécie mais abundante (51,36%), seguida de L. choti (43,54%). Todas as capturas foram realizadas nas áreas onde foram desenvolvidos treinamentos militares, nas quais foram verificados casos clínicos em indivíduos após realizarem treinamentos. Os dados obtidos sobre a fauna local, associados com achados prévios nessa área da Zona da Mata, apresentam evidências consistentes sobre a manutenção de um ciclo enzoótico, com a ocorrência de surtos periódicos de LTA, posteriores à realização de treinamentos, em áreas correspondentes à Mata Atlântica remanescente e matas secundárias.

The aim of this study was to characterize the sandfly fauna in a military training area situated in the "Zona da Mata" region of Pernambuco State, Brazil, where human cases of American tegumentary leishmaniasis (ATL) had been observed, caused by Leishmania (Viannia) braziliensis. The occurrence of 16 sandfly species in the Marshall Newton Cavalcanti Military Training Camp (CIMNC) indicates an important diversity of sandfly species in this study area. Lutzomyia complexa was the most common species (51.36%) followed by L. choti (43.54%). All the captures were made in areas where training activities were conducted and where clinical cases had been detected in individuals performing training, following this period. The data relating the sandfly behavior to previous data in the same area provides strong evidence for the occurrence of an enzootic cycle of the parasite, and outbreaks followed the period of training activities in the remaining Atlantic Rain Forest and secondary forest areas.

Esse estudo teve como objetivo geral caracterizar a fauna de flebotomíneos envolvida em unidade de treinamento militar localizada na Zona da Mata do Estado de Pernambuco, Brasil, onde foram verificados casos humanos autóctones de leishmaniose tegumentar americana (LTA) associados a Leishmania (Viannia) braziliensis. A presença de 16 espécies de flebotomíneos no Centro de Instrução Militar Marechal Newton Cavalcanti indicou uma importante diversidade na área de estudo. Lutzomyia complexa constituiu a espécie mais abundante (51,36%), seguida de L. choti (43,54%). Todas as capturas foram realizadas nas áreas onde foram desenvolvidos treinamentos militares, nas quais foram verificados casos clínicos em indivíduos após realizarem treinamentos. Os dados obtidos sobre a fauna local, associados com achados prévios nessa área da Zona da Mata, apresentam evidências consistentes sobre a manutenção de um ciclo enzoótico, com a ocorrência de surtos periódicos de LTA, posteriores à realização de treinamentos, em áreas correspondentes à Mata Atlântica remanescente e matas secundárias.

The aim of this study was to characterize the sandfly fauna in a military training area situated in the "Zona da Mata" region of Pernambuco State, Brazil, where human cases of American tegumentary leishmaniasis (ATL) had been observed, caused by Leishmania (Viannia) braziliensis. The occurrence of 16 sandfly species in the Marshall Newton Cavalcanti Military Training Camp (CIMNC) indicates an important diversity of sandfly species in this study area. Lutzomyia complexa was the most common species (51.36%) followed by L. choti (43.54%). All the captures were made in areas where training activities were conducted and where clinical cases had been detected in individuals performing training, following this period. The data relating the sandfly behavior to previous data in the same area provides strong evidence for the occurrence of an enzootic cycle of the parasite, and outbreaks followed the period of training activities in the remaining Atlantic Rain Forest and secondary forest areas.