A NEW RECORD OF Leishmania (Viannia) guyanensis (Trypanosomatidae) FROM THE PACIFIC COAST OF COLOMBIA / Un nuevo registro de Leishmania (Viannia) guyanensis (Trypanosomatidae) de la costa pacífica de Colombia

ABSTRACT The aim of this study was the identification of Leishmania species that causes cutaneous leishmaniasis in a patient from Buenaventura, Valle del Cauca, on the Pacific coast of Colombia. Clinical samples were obtained from a 29 years-old male who presented a distinct ulcer with raised borders on his neck. Samples were taken for direct microscopic examination, parasite culture, and molecular identification of the infecting Leishmania species by sequencing of the cytochrome b gene. Direct examination was positive for amastigotes of Leishmania but the culture was negative. The infecting parasite species was identified as L. (V.) guyanensis by means of the nucleotide sequence of a 509 bp fragment of the cytochrome b gene. We report the presence of L. (V.) guyanensis in rural areas of Buenaventura in Valle del Cauca, and the expansion of the geographical distribution of this species in the Pacific region of Colombia.

RESUMEN El objetivo de este estudio fue identificar la especie de Leishmania causante de la leishmaniasis cutánea en un paciente de Buenaventura, Valle del Cauca, en la costa Pacífica de Colombia. Se obtuvieron muestras clínicas de un varón de 29 años de edad que presentó una úlcera distintiva con bordes levantados en el cuello. Se tomaron muestras para examen microscópico directo, cultivo de parásitos e identificación molecular de la especie infectante de Leishmania mediante secuenciación del gen del citocromo b. El examen directo fue positivo para amastigotes de Leishmania pero el cultivo fue negativo. La especie parasitaria infectante se identificó como L. (V.) guyanensis por medio de la secuencia de nucleótidos de un fragmento de 509 pb del gen citocromo b. Con este reporte notificamos la presencia de L. (V.) guyanensis en zona rural del municipio de Buenaventura en el Valle del Cauca y la expansión de la distribución geográfica de esta especie en la región Pacífica de Colombia.

Vetor ou vetores? Capacidade vetorial e estratégias de controle / Vector or vectors? Vectorial capacity and control strategies

Variáveis relacionadas a fatores epidemiológicos, ao ambiente, ao agente etiológico, ao vetor e aos reservatórios parecem atuar na determinação dos diferentes cenários de transmissão da leishmaniose visceral no Brasil. No estado de São Paulo a leishmaniose visceral (LV) não apresenta um padrão epidemiológico homogêneo por todas suas regiões, parecendo refletir uma multitude de cenários propícios para a ocorrência da transmissão dentro do território Paulista. Lutzomyia longipalpis é composta por um complexo de espécies das quais duas são encontradas no Estado de São Paulo e parecem possuir diferença na capacidade vetorial. É provável que essa diferença seja o fator determinante na caracterização dos diferentes padrões epidemiológicos observados nas diferentes regiões do Estado. No presente estudo, procuramos determinar a distribuição temporal e geográfica das espécies do complexo Lu. longipalpis, dos casos caninos e dos casos humanos de LV como elementos chave para ajudar na caracterização de alguns cenários de transmissão da doença e apontar áreas de maior risco para a aquisição da doença. Por outro lado, a recente e inesperada ocorrência de transmissão da LV em localidades sem a presença da Lu. longipalpis, caracteriza mais um novo cenário, onde a transmissão da Leishmania infantum ao homem se mostrou possível, configurando um novo desafio para as autoridades da saúde pública.

Variables related to epidemiological factors, etiologic agent, environment, vector and reservoirs seem to act in determining the different scenarios of transmission of visceral leishmaniasis (VL) in Brazil. In the state of São Paulo, VL does not present a homogeneous epidemiological pattern across all its regions, seeming to reflect a multitude of scenarios conducive to the occurrence of transmission within Paulista territory. Lutzomyia longipalpis is composed of a complex of species of which two are found in the state of São Paulo and appear to have a difference in vector capacity. It is likely that this difference is the determining factor in the characterization of the different epidemiological patterns observed in the different regions of the State. In the present study, we sought to determine the temporal and geographic distribution of Lu. longipalpis species, canine cases and human cases of VL as key elements to help characterize some scenarios of transmission of the disease and to indicate areas of greater risk for disease acquisition. The recent and unexpected occurrence of VL transmission in localities without the presence of Lu. longipalpis characterizes another new scenario, where other species of san flies can transmit Leishmania infantum to man and configuring a new challenge for public health authorities.

Buparvaquone nanostructured lipid carrier development: physicochemical and in vitro leishmanicidal performances / Desenvolvimento de carreadores lipídicos nanoestruturados contendo buparvaquona: caracterização físico-química e avaliação da atividade leishmanicida in vitro

Leishmaniases is a group of diseases caused by parasites of the genus Leishmania. The estimated number of deaths from visceral leishmaniases ranges from 20,000 to 50,000 annually. The most common treatment over the past 60 years has been pentavalent antimonials. Besides the doubtful effectiveness, they present several disadvantages such as the need for parenteral administration, large doses, long treatment, severe toxicity and parasite resistance. Buparvaquone (BPQ), a drug used for veterinary treatment of theileriosis, showed promising activity against Leishmania spp. However, due to its low aqueous solubility and bioavailability, it has failed in in vivo tests. The use of nanotechnologies has the potential to overcome these drawbacks due to the following advantages: increase in drug water-solubility, increase in therapeutic efficacy and treatment toxicity reduction. Therefore, the present work aimed the development, optimization, physical-chemical evaluation and in vitro performances of nanostructured lipid carriers (NLC) for BPQ encapsulation. The NLC preparation was performed by high pressure homogenization, and surface response and factorial design were applied to formulation optimization. In vitro dissolution profiles were evaluated in phosphate buffer pH 7.4 with tween 80 0.07% w/v or sodium dodecyl sulfate 1% w/v and simulated body fluid pH 7.4. Cytotoxicity was evaluated in mouse peritoneal macrophages and leishmanicidal activity in L. infantum amastigotes. Six optimized NCL were prepared and they showed solubility improvement from 1.5- fold to 611-fold when compared with free BPQ, depending on the formulation and medium. Dissolution profiles showed the NLC formulation suitability for BPQ regarding oral administration, the release could reach 83.29% of a 4mg dose in 30 minutes for formulation of 175.1 nm, while the free drug could be dissolved only 2.89% of the same dose after 4 hours. Moreover, formulation of 230.7 nm showed 81.42% of drug release in in phosphate buffer pH 7.4 with dodecyl sulfate 1.0% w/v after 30 minutes, while BPQ did not dissolved. Cytotoxicity assay showed the safety of all formulations. The iv CC50 values were close to 500 µM, while the IC50 against amastigotes was only 456.5 nM for free BPQ. Developed NLCs showed an increase in IC50 from 2.0 to 3.1-fold when compared to free drug in the in vitro leishmanicidal evaluation. Therefore, the NLC containing BPQ are a promising alternative for the treatment of leishmaniases as oral and parenteral drug dosage forms. Additionally, they have a potential use for lymphatic targeted drug delivery, which can be an innovative approach for this neglected disease.

Leishmanioses são um grupo de doenças causadas por parasitas do gênero Leishmania. O número estimado de óbitos por leishmaniose visceral varia entre 20.000 e 50.000 por ano. O tratamento mais comum nos últimos 60 anos tem sido os antimônios pentavalentes. Além da eficácia duvidosa, eles apresentam várias desvantagens, como a necessidade de administração parenteral, altas doses, tratamento prolongado, toxicidade severa e resistência parasitária. Buparvaquona (BPQ), um fármaco usado para tratamento veterinário da teileriose, mostrou atividade promissora contra Leishmania donovani. No entanto, devido à sua baixa solubilidade e biodisponibilidade aquosa, falhou em testes in vivo. O uso das nanotecnologias tem o potencial de superar esses obstáculos devido às seguintes vantagens: aumento da solubilidade em água, aumento da eficácia terapêutica e redução da toxicidade do tratamento. Portanto, o presente trabalho objetivou o desenvolvimento, otimização, avaliação físico-química e avaliação do desempenho in vitro de carreadores lipídicos nanoestruturados (NLC) para o encapsulação da BPQ. A preparação do NLC foi realizada por homogeneização de alta pressão e superfície de resposta e planejamento fatorial foram aplicados à otimização das formulações. Os perfis de dissolução in vitro foram avaliados em tampão fosfato pH 7.4 com tween 80 a 0.07% p/v ou dodecilsulfato de sódio 1.0% p/v e fluido corporal simulado pH 7.4. A citotoxicidade foi avaliada em macrófagos peritoneais de camundongos e atividade leishmanicida em amastigotas de L. infantum. Foram preparados quatro NCL otimizados e mostraram melhora da solubilidade de 1,5 a 611 vezes quando comparado com a BPQ livre, dependendo da formulação e do meio. Os perfis de dissolução mostraram a adequação da formulação NLC para BPQ em relação à administração oral. A dissolução pode atingir 83,29% de uma dose de 4.0 mg em 30 minutos para a formulação de 175,1 nm, enquanto o fármaco livre dissolveu apenas vi 2,89% da mesma dose após 4 horas. Além disso, a formulação de 230,7 nm mostrou 81,42% de liberação do fármaco em tampão fosfato pH 7.4 com dodecil sulfato de sódio 1.0% p/v após 30 minutos, enquanto o BPQ não se dissolveu. O teste de citotoxicidade mostrou a segurança de todas as formulações. Os valores CC50 foram próximos de 500 µM, enquanto o IC50 em amastigotas foi de apenas 456,5 nM para BPQ livre. Os NLC desenvolvidos mostraram um aumento no IC50 de 2,0 a 3,1 vezes quando comparado ao;fármaco livre na avaliação leishmanicida in vitro. Logo, as NLC contendo BPQ são uma alternativa promissora para o tratamento de leishmanioses como formas farmacêuticas oral e parenteral. Além disso, eles têm um uso potencial para a sítio-específico ao sistema linfático, o que pode ser uma abordagem inovadora para esta doença negligenciada.

Tissue distribution of residual antimony in rats treated with multiple doses of meglumine antimoniate

Meglumine antimoniate (MA) and sodium stibogluconate are pentavalent antimony (SbV) drugs used since the mid-1940s. Notwithstanding the fact that they are first-choice drugs for the treatment of leishmaniases, there are gaps in our knowledge of their toxicological profile, mode of action and kinetics. Little is known about the distribution of antimony in tissues after SbV administration. In this study, we evaluated the Sb content of tissues from male rats 24 h and three weeks after a 21-day course of treatment with MA (300 mg SbV/kg body wt/d, subcutaneous). Sb concentrations in the blood and organs were determined by inductively coupled plasma-mass spectrometry. In rats, as with in humans, the Sb blood levels after MA dosing can be described by a two-compartment model with a fast (t1/2 = 0.6 h) and a slow (t1/2 >> 24 h) elimination phase. The spleen was the organ that accumulated the highest amount of Sb, while bone and thyroid ranked second in descending order of tissues according to Sb levels (spleen >> bone, thyroid, kidneys > liver, epididymis, lungs, adrenals > prostate > thymus, pancreas, heart, small intestines > skeletal muscle, testes, stomach > brain). The pathophysiological consequences of Sb accumulation in the thyroid and Sb speciation in the liver, thyroid, spleen and bone warrant further studies.

Species composition and seasonal abundance of sandflies (Diptera: Psychodidae: Phlebotominae) in coffee agroecosystems

The composition and seasonal occurrence of sandflies were investigated in coffee agroecosystems in the Soconusco region of Chiapas, Mexico. Insect sampling was performed on three plantations located at different altitudes: Finca Guadalupe Zajú [1,000 m above sea level (a.s.l.)], Finca Argovia (613 m a.s.l.) and Teotihuacán del Valle (429 m a.s.l.). Sandflies were sampled monthly from August 2007-July 2008 using three sampling methods: Shannon traps, CDC miniature light traps and Disney traps. Sampling was conducted for 3 h during three consecutive nights, beginning at sunset. A total of 4,387 sandflies were collected during the course of the study: 2,718 individuals in Finca Guadalupe Zajú, 605 in Finca Argovia and 1,064 in Teotihuacán del Valle. The Shannon traps captured 94.3% of the total sandflies, while the CDC light traps and Disney traps captured 4.9% and 0.8%, respectively. More females than males were collected at all sites. While the number of sandflies captured was positively correlated with temperature and relative humidity, a negative correlation was observed between sandfly numbers and rainfall. Five species of sandflies were captured: Lutzomyia cruciata , Lutzomyia texana , Lutzomyia ovallesi , Lutzomyia cratifer / undulata and Brumptomyia sp. Lu. cruciata , constituting 98.8% of the total, was the most abundant species. None of the captured sandflies was infected with Leishmania spp.

Phlebotomine sandflies (Diptera: Psychodidae) in São Vicente Férrer, a sympatric area to cutaneous and visceral leishmaniasis in the state of Pernambuco, Brazil / Flebotomíneos (Diptera: Psychodidae) em São Vicente Férrer, uma área simpátrica para leishmaniose tegumentar e visceral, no estado de Pernambuco, Brasil

INTRODUCTION: In the last decades, a considerable geographic expansion of the leishmaniases in all regions of Brazil has been observed. The present study was carried out to identify the composition of the phlebotomine sandfly fauna and verify the seasonal variation of the main species after environmental changes occurred in São Vicente Férrer Municipality, State of Pernambuco, Brazil. METHODS: Captures were carried out during four consecutive nights of each month using Centers for Disease Control and Prevention light traps from September 2009 to September 2010. The correlation between the number of phlebotomine sandflies captured and climatic factors (temperature and rainfall) was evaluated. RESULTS: A total of 13,872 specimens belonging to 20 species were captured, of which, 6,247 (45 percent) were females, and 7,625 (55 percent) were males. Lutzomyia migonei was the most abundant species with 9,964 (71.8 percent) specimens, being predominant in the intradomicile and peridomicile areas with 108 (86.4 percent) and 9,746 (97 percent), respectively. In the forest remnants, Lutzomyia complexa 2,395 (65 percent) and Lutzomyia sordellii 770 (20.8 percent) predominated. The correlation analysis between the total number of sandflies captured and climatic factors did not show a significant influence on population density. CONCLUSIONS: The high abundance of Lutzomyia migonei and Lutzomyia complexa indicates the possibility of new cases of cutaneous leishmaniasis (CL).

INTRODUÇÃO: Nas últimas décadas tem se observado uma considerável expansão geográfica das leishmanioses em todas as regiões do Brasil. O presente estudo foi realizado para identificar a fauna de flebotomíneos e verificar a sazonalidade das principais espécies após mudanças ambientais ocorridas no município de São Vicente Férrer, Estado de Pernambuco, Brasil. MÉTODOS: As capturas foram realizadas durante quatro noites consecutivas por mês usando armadilhas luminosas CDC, no período de setembro de 2009 a setembro de 2010. A correlação entre o número de flebotomíneos capturados e fatores climáticos (temperatura e umidade) foi avaliada. RESULTADOS: Foi capturado um total de 13.872 espécimes pertencentes a 20 espécies, sendo 6.247 (45 por cento) fêmeas e 7.625 (55 por cento) machos. Lutzomyia migonei foi a espécie mais abundante com 9.964 (71,8 por cento) espécimes, sendo predominante no intradomicílio 108 (86,4 por cento) e peridomicílio 9.746 (97 por cento). Nos resquícios de mata primária, predominaram Lutzomyia complexa 2.395 (65 por cento) e Lutzomyia sordellii 770 (20,8 por cento). A análise de correlação entre o número total de flebotomíneos capturados e os fatores climáticos não mostrou uma influência significativa na densidade da população. CONCLUSÕES: A elevada abundância de Lutzomyia migonei e Lutzomyia complexa indica a possibilidade de novos casos de leishmaniose cutânea (LC).

Phlebotomine sand flies (Diptera: Psychodidae: Phlebotominae) in the State of Pernambuco / Flebotomíneos (Diptera: Psychodidae: Phlebotominae) no Estado de Pernambuco

INTRODUCTION: Phlebotomine sand flies are small insects of great medical relevance. This article presents an updated list of the phlebotomine sand flies occurring in the State of Pernambuco, Brazil. METHODS: A review of literature published since the 1940s up to May 2010 was conducted and voucher material deposited in reference collections were studied. RESULTS: A total of 37 phlebotomine species have previously been reported as occurring in Pernambuco, but the record of six species needs confirmation. CONCLUSIONS: This work provides an updated list of phlebotomine sand flies of Pernambuco, with some notes on their taxonomy, ecology, distribution and epidemiological relevance.

INTRODUÇÃO: Flebotomíneos são pequenos insetos de grande importância médica e veterinária. Este artigo apresenta uma lista atualizada dos flebotomíneos que ocorrem no Estado de Pernambuco, Brasil. MÉTODOS: Uma revisão da literatura publicada desde a década de 40 até maio de 2010 foi realizada e material testemunho depositado em coleções de referência foi estudado. RESULTADOS: Um total de 37 espécies de flebotomíneos foram previamente relatadas como presentes em Pernambuco, mas o registro de seis delas necessita de confirmação. CONCLUSÕES: Esse trabalho fornece uma lista atualizada de flebotomíneos de Pernambuco, com algumas notas sobre a sua taxonomia, ecologia, distribuição e relevância epidemiológica.

The potential of the extracts of Tagetes minuta Linnaeus (Asteraceae), Acalypha fruticosa Forssk (Euphorbiaceae) and Tarchonanthus camphoratus L. (Compositae) against Phlebotomus duboscqi Neveu Lemaire (Diptera: Psychodidae), the vector for Leishmania major Yakimoff and Schokhor.

Background & objectives: Harmful effects of synthetic chemical insecticides including vector resistance, environmental pollution and health hazards have necessitated the current significance in the search for plant-based insecticide products that are environmentally safe and effective to leishmaniases control. The insecticidal activity of Tagetes minuta Linnaeus (Asteraceae), Acalypha fruticosa Forssk (Euphorbiaceae) and Tarchonanthus camphoratus L. (Compositae) extracts were investigated against Phlebotomus duboscqi Neveu Lemaire (Diptera: Psychodidae). Methods: The extracts were prepared from dried aerial parts soaked in methanol and ethyl acetate twice until the filtrates became clear, filtered and dried out by rotary evaporation at 30–35oC. The solid extracts obtained were later prepared into 2.5, 5 and 10 mg/ml. Two millilitres of the solutions were blotted on filter papers, which were dried overnight and placed into jars where adult sandflies were aspirated. Males and females were assayed separately. Results & conclusion: The extracts had significant mortality (p<0.05) in both males and females bioassays but were not significantly different between sexes. The extracts of Acalypha fruticosa and Tagetes minuta had significantly higher mortality rates than those of Tarchonanthus camphoratus and the different concentrations used showed significantly different mortality rates and 10 mg/ml was the most effective concentration. Cent percent mortality was obtained at 96 h of exposure to 5 and 10 mg/ml concentrations except for Tarchonanthus camphoratus which had a mortality of only 46.7% in 10 mg/ml bioassay. These extracts were found to be insecticidal to adult sandflies.

Phlebotomines (Diptera, Psychodidae) in the Speleological Province of the Ribeira Valley: 3. Serra district - area of hostels for tourists who visit the Parque Estadual do Alto Ribeira (PETAR), state of São Paulo, Brazil

Phlebotomines (Diptera, Psychodidae) in the Speleological Province of the Ribeira Valley: 3. Area of hostels for tourists who visit the Parque Estadual do Alto Ribeira (PETAR), state of São Paulo, Brazil. The study characterizes some ecological aspects of the phlebotomine fauna in an endemic area of cutaneous leishmaniasis (CL) situated in the Serra district, Iporanga municipality where the hostels for tourists visiting the PETAR are located. Captures were undertaken on a smallholding and a small farm situated near the hostels, monthly between January/2001 and December/2003 with automatic light traps (ALT) in pigsty, hen-house and veranda of a domicile at the two sites, and in peridomicile of the small farm also with black/white Shannon traps. With the ALT a total of 87,224 phlebotomines representing 19 species and also two hybrids of Nyssomyia intermedia (Lutz & Neiva) and Nyssomyia neivai (Pinto) and two anomalous specimens were captured. The standardized index species abundance was for Ny. intermedia = 1.0 and Ny. neivai = 0.935. The highest frequencies of the smallholding occurred in the pigsty, the Williams' mean/capture for Ny. intermedia being 63.7 specimens and for Ny. neivai 29.2, and on the small farm, in the hen-house, Ny. intermedia 402.6 and Ny. neivai 116.2. A total of 863 phlebotomines (Ny. intermedia: 75.4 percent; Ny. neivai: 24.3 percent) were captured with black/white Shannon traps; females of both species being predominant in the white trap. The high frequencies of Ny. intermedia and Ny. neivai, both implicated in CL transmission, indicate the areas presenting risk of the disease.

Phlebotominae (Diptera, Psychodidae) na Província Espeleológica do Vale do Ribeira: 3. Área de pousadas para turistas que visitam o Parque Estadual do Alto Ribeira (PETAR), estado de São Paulo, Brasil. O estudo teve como objetivo caracterizar alguns aspectos ecológicos da fauna flebotomínea em área endêmica de leishmaniose tegumentar situada do Bairro Serra, município de Iporanga, onde se situam pousadas para turistas que visitam o PETAR. As capturas foram realizadas em uma chácara e em um sítio, mensalmente, entre janeiro de 2001 e dezembro de 2003, com armadilhas automáticas luminosas (AAL) em chiqueiro, galinheiro e varanda do domicílio de ambos os locais, e no sítio, também com armadilhas de Shannon branca/preta. Capturou-se com as AALs um total de 87.224 flebotomíneos, representando 19 espécies, mais dois híbridos de Nyssomyia intermedia (Lutz & Neiva, 1912) e Nyssomyia neivai (Pinto, 1926) e dois espécimes anômalos. Ny. intermedia apresentou o índice de abundância das espécies padronizado = 1,0 e Ny. neivai = 0,935. Na chácara, ambas predominaram no chiqueiro, com a média geométrica de Williams/captura para Ny. intermedia = 63,7 espécimes e para Ny. neivai = 29,2, e no sítio, no galinheiro, Ny. intermedia = 402,6 e Ny. neivai = 116,2. Capturou-se nas armadilhas de Shannon branca/preta 863 flebotomíneos: Ny. intermedia (75,4 por cento) e Ny. neivai (24,3 por cento), as fêmeas sendo significativamente predominantes na branca. As altas freqüências de Ny. intermedia e Ny. neivai, ambas implicadas na transmissão de agente da leishmaniose tegumentar, indicam risco de transmissão da doença na área.