RESUMO O Plano Global de Eliminação da Filariose Linfática, lançado pela Organização Mundial da Saúde em 2000, propõe o uso de testes de detecção de antígeno circulante filarial como ferramenta diagnóstica para avaliação e monitoramento das ações de controle da parasitose. Entretanto, esses testes, apesar de apresentarem alta sensibilidade, não conseguem detectar com eficiência a infecção em seu estágio inicial, quando ainda não existe a presença de helmintos adultos. Considerando essa limitação, a pesquisa de anticorpos antifilariais tem sido apontada como uma alternativa, uma vez que os anticorpos produzidos contra as larvas infectantes do parasito são detectados antes da presença de antígeno circulante filarial. O objetivo deste estudo foi definir o ponto de corte e avaliar a acurácia do kit Filaria Detect™ IgG4 produzido com o antígeno recombinante Wb123 para diagnóstico da filariose linfática no Brasil. Para isso, foi realizado um estudo de avaliação de teste diagnóstico, no qual foram utilizadas 256 amostras de soro: 79 (30,9%) obtidas de indivíduos microfilarêmicos e 177 (60,1%), de indivíduos amicrofilarêmicos e que testaram negativo para os testes imunológicos Bm14 CELISA e Og4C3 ELISA. A definição do ponto de corte ideal, bem como da acurácia do kit Filaria Detect™ IgG4, foi obtida através da construção de curvas ROC, sendo a densidade óptica de 0,239 aquela na qual o teste obteve melhor desempenho, com sensibilidade de 81,0% e especificidade de 96,6%. Os resultados obtidos demonstraram que o kit Filaria Detect™ IgG4 é uma ferramenta promissora para investigação e monitoramento de áreas submetidas ao tratamento em massa para filariose linfática.

ABSTRACT The Global Programme to Eliminate Lymphatic Filariasis, launched by the World Health Organization in the year 2000, proposes the use of circulating filarial antigen tests as a diagnostic tool to assess and monitor initiatives to control filarial infection. However, despite a high sensitivity, these tests are not efficient to detect infection at early stages, before worms have reached the adult stage. Considering this limitation, anti-filarial antibody testing has been suggested as an alternative, given that the antibodies produced against the larvae are detectable before the presence of circulating filarial antigen. The objective of the present study was to determine the diagnostic cut-off and the accuracy of the Filaria Detect™ IgG4 kit employing recombinant Wb123 antigen for diagnosis of lymphatic filariasis in Brazil. For that, we performed a diagnostic evaluation study in which 256 serum samples were analyzed: 79 (30.9%) obtained from microfilaremic individuals and 177 (60.1%) from amicrofilaremic individuals who tested negative with the Bm14 CELISA and Og4C3 ELISA immunologic tests. The ideal cutoff as well as the Filaria Detect™ IgG4 kit accuracy were determined based on ROC curve analyses, with an optical density of 0.239 identified as the cutoff with the best performance, with 81.0% sensitivity and 96.6% specificity. The results show that the Filaria Detect™ IgG4 kit is a promising tool for investigation and monitoring of areas undergoing mass drug administration for lymphatic filariasis.

RESUMEN En el programa mundial de eliminación de la filariasis linfática, puesto en marcha por la Organización Mundial de la Salud en el año 2000, se propone el uso de pruebas de detección del antígeno filárico circulante como instrumento de diagnóstico para la evaluación y el seguimiento de las medidas de control de la parasitosis. Sin embargo, esas pruebas, a pesar de tener un alto grado de sensibilidad, no permiten detectar con eficiencia la infección en su fase inicial, cuando todavía no existen helmintos adultos. En vista de esa limitación, se ha señalado como una opción el estudio de anticuerpos antifiláricos, puesto que los anticuerpos producidos contra las larvas infectantes del parásito se detectan antes de la existencia de antígeno filárico circulante. El objetivo de este estudio fue definir el punto de corte y evaluar la exactitud del estuche Detect™ para pruebas de anticuerpos antifiláricos IgG4, fabricado con el antígeno recombinante Wb123, para el diagnóstico de la filariasis linfática en Brasil. Para ello, se realizó un estudio de evaluación de la prueba diagnóstica, en el cual se utilizaron 256 muestras de suero, a saber, 79 (30,9%) obtenidas de personas microfilarémicas y 177 (60,1%) de personas amicrofilarémicas, que arrojaron resultados seronegativos en las pruebas inmunológicas CELISA Bm14 y ELISA Og4C3. La definición del punto de corte ideal y de la exactitud del estuche Detect™ se obtuvo con la construcción de curvas de la característica operativa del receptor (ROC); una densidad óptica de 0,239 marcó el mejor nivel de desempeño de la prueba, con una sensibilidad de 81,0% y una especificidad de 96,6%. Los resultados obtenidos demostraron que el estuche Detect™ es un instrumento prometedor para la investigación y el seguimiento de las regiones donde se realiza un tratamiento masivo de la filariasis linfática.

Introduction Since the launch of the Global Programme to Eliminate Lymphatic Filariasis, more than 70% of the endemic countries have implemented mass drug administration (MDA) to interrupt disease transmission. The monitoring of filarial infection in sentinel populations, particularly schoolchildren, is recommended to assess the impact of MDA. A key issue is choosing the appropriate tools for these initial assessments (to define the best intervention) and for monitoring transmission. Methods This study compared the pre-MDA performance of five diagnostic methods, namely, thick film test, Knott's technique, filtration, Og4C3-ELISA, and the AD12-ICT card test, in schoolchildren from Brazil. Venous and capillary blood samples were collected between 11 pm and 1 am. The microfilarial loads were analyzed with a negative binomial regression, and the prevalence and associated 95% confidence intervals were estimated for all methods. The accuracies of the AD12-ICT card and Og4C3-ELISA tests were assessed against the combination of parasitological test results. Results A total of 805 schoolchildren were examined. The overall and stratified prevalence by age group and gender detected by Og4C3-ELISA and AD12-ICT were markedly higher than the prevalence estimated by the parasitological methods. The sensitivity of the AD12-ICT card and Og4C3-ELISA tests was approximately 100%, and the positive likelihood ratios were above 6. The specificity of the Og4C3-ELISA was higher than that of the AD12-ICT at different prevalence levels. Conclusions The ICT card test should be the recommended tool for monitoring school-age populations living in areas with ongoing or completed MDA.

Introduction Polymerase chain reaction (PCR) may offer an alternative diagnostic option when clinical signs and symptoms suggest visceral leishmaniasis (VL) but microscopic scanning and serological tests provide negative results. PCR using urine is sensitive enough to diagnose human visceral leishmaniasis (VL). However, DNA quality is a crucial factor for successful amplification. Methods A comparative performance evaluation of DNA extraction methods from the urine of patients with VL using two commercially available extraction kits and two phenol-chloroform protocols was conducted to determine which method produces the highest quality DNA suitable for PCR amplification, as well as the most sensitive, fast and inexpensive method. All commercially available kits were able to shorten the duration of DNA extraction. Results With regard to detection limits, both phenol: chloroform extraction and the QIAamp DNA Mini Kit provided good results (0.1 pg of DNA) for the extraction of DNA from a parasite smaller than Leishmania (Leishmania) infantum (< 100fg of DNA). However, among 11 urine samples from subjects with VL, better performance was achieved with the phenol:chloroform method (8/11) relative to the QIAamp DNA Mini Kit (4/11), with a greater number of positive samples detected at a lower cost using PCR. Conclusion Our results demonstrate that phenol:chloroform with an ethanol precipitation prior to extraction is the most efficient method in terms of yield and cost, using urine as a non-invasive source of DNA and providing an alternative diagnostic method at a low cost.

RESUMORelato de caso de paciente masculino de 45 anos, agricultor, residente na zona da mata do Estado de Pernambuco, diagnosticado com HIV em 1999 e em uso de ARV. Em 2005 foi registrada a primeira ocorrência de LV através do diagnóstico parasitológico a partir do aspirado da medula óssea. À admissão no hospital apresentava-se com febre, hepatoesplenomegalia, perda de peso e diarréia. Desde então houve a ocorrência de mais sete episódios de LV, tendo ocorrido em media, um evento a cada ano (2005-2012 exceto em 2008). O paciente apresentou, em 2011, um quadro cutâneo disseminado, sendo realizada biopsia de pele que evidenciou formas amastigotas de Leishmania no exame histopatológico. Em 2005, o tratamento foi realizado com antimoniato de N-metil-glucamina e anfotericina B desoxicolato, mas desde 2006, devido à toxicidade, o medicamento de escolha foi a anfotericina B lipossomal. Como recomendado pelo Ministério da Saúde, esse relato reforça a necessidade de que os casos de HIV residentes em área endêmica de LV deverão ter inserido em seu protocolo de acompanhamento essa parasitose, principalmente após o primeiro episódio.

SUMMARYReport of a 45-year-old male farmer, a resident in the forest zone of Pernambuco, who was diagnosed with human immunodeficiency virus (HIV) in 1999 and treated using antiretroviral (ARV) drugs. In 2005, the first episode of visceral leishmaniasis (VL), as assessed by parasitological diagnosis of bone marrow aspirate, was recorded. When admitted to the hospital, the patient presented fever, hepatosplenomegaly, weight loss, and diarrhea. Since then, six additional episodes of VL occurred, with a frequency rate of one per year (2005-2012, except in 2008). In 2011, the patient presented a disseminated skin lesion caused by the amastigotes of Leishmania, as identified by histopathological assessment of skin biopsy samples. In 2005, he was treated with N-methyl-glucamine-antimony and amphotericin B deoxycholate. However, since 2006 because of a reported toxicity, the drug of choice was liposomal amphotericin B. As recommended by the Ministry of Health, this report emphasizes the need for HIV patients living in VL endemic areas to include this parasitosis in their follow-up protocol, particularly after the first infection of VL.