Evaluation of larval surface antigens from infective larvae of Strongyloides venezuelensis for the serodiagnosis of human strongyloidiasis

ABSTRACT Serodiagnosis of strongyloidiasis is usually performed by ELISA for the detection of IgG antibodies due to its high sensitivity and practicality, but its main limitation is a constant source of S. stercoralis antigens. The use of S. venezuelensis as a heterologous source of antigens has facilitated several published studies on the serodiagnosis and epidemiology of human strongyloidiasis. The main objective of this study was to evaluate the diagnostic accuracy of surface cuticle antigens of infective larvae of S. venezuelensis extracted with CTAB detergent (L3-CTAB) in comparison with soluble somatic extracts (L3-SSE) using a panel of sera from immunocompetent and immunocompromised individuals, at three different cut-offs. ROC curve analysis showed that L3-CTAB had an AUC of 0.9926. At the first cut-off value (OD 450 nm = 0.214), sensitivity and specificity were 100% and 90.11%, respectively, with a diagnostic accuracy of 0.93. At a second cut-off value (OD 450 nm = 0.286), sensitivity and specificity were 70% and 100%, respectively, with a diagnostic accuracy of 0.91. However, at an alternative third cut-off value (OD 450 nm = 0.589), sensitivity and specificity were 95% and 97.8%, respectively, with a diagnostic accuracy of 0.97. Using L3-CTAB as an antigenic source, the seropositivity rate in immunocompromised patients was 28.13% (9/32) whereas a seropositivity rate of 34.38% (11/32) was found when L3-SSE was used in ELISA. Therefore, the L3-CTAB is simple and practical to obtain and was found to be highly sensitive and specific.

Toxocara canis 30-35 kDa excretory-secretory antigen is an important marker in mice challenged by inocula containing different parasite load levels

ABSTRACT The Western-blotting technique was applied to identify antigenic fractions of excretory-secretory Toxocara canis antigen recognized by IgG antibodies throughout an experimental infection in mice challenged by different inocula. Mice were inoculated with 5, 50 and 500 embryonated eggs and serum samples were collected 15, 30, 60, 90 and 120 days post-infection. Serum samples were analyzed using an excretory-secretory Toxocara antigen. Antibodies recognized antigenic fractions from 30 to 90 kDa. The protein fraction of 30-35 kDa was the most frequently recognized regardless of the size of inoculum and the stage of infection represented by the different collection times, but the antigenic recognition was more evident in groups infected with 50 and 500 eggs. This study presents an antigenic panel of the excretory-secretory antigen of T. canis and suggests that the 30-35 kDa antigenic fraction is a promising marker of the infection and should be further explored in future studies on experimental toxocariasis.

Importance of detection of Strongyloides stercoralis DNA in fecal samples from patients with type 2 diabetes mellitus

Abstract Objective: The association between diabetes and Strongyloides infection remains controversial. This study aimed to detect Strongyloides stercoralis DNA in the feces of patients with Diabetes Mellitus type 2 (DM2). Methods: Fecal samples were analyzed via the Lutz, Rugai, and agar plate culture methods. PCR amplification was performed using two targets (PCR-genus and PCR-species) located on the S. stercoralis 18S ribosomal. Results: The positivity for S. stercoralis using parasitological methods was 1.1%. PCR-genus (14.13%) demonstrated a higher positivity than PCR-species (9.78%). Conclusion: The results confirm the greater positivity of the molecular diagnosis in relation to parasitological methods, reinforcing its use as an additional tool for the diagnosis of S. stercoralis infection in patients with DM2 living in endemic areas for this helminthiasis. HIGHLIGHTS Positivity for strongyloidiasis in coproscopic exam was low in diabetic patients. PCR is more sensitive for detecting S. stercoralis infection in diabetic patients. Molecular diagnosis is an important tool for the detection of S. stercoralis.

Current status of research regarding Blastocystis sp, an enigmatic protist, in Brazil

The present study aimed to evaluate the occurrence of Blastocystis sp. in Brazilian studies over a period of years (2000-2020), as well as point out relevant aspects of this enigmatic organism. We performed a literature search using six sources of international databases. The data were divided into diagnostic by parasitological and molecular techniques, and relevant aspects. After applying the inclusion and exclusion criteria, 52 studies were included in the final analysis. The occurrence of Blastocystis sp. in Brazil ranged from 0.5% to 86.6%, as determined using parasitological techniques. The highest occurrence was in the North (27.3%) and the lowest, in the Midwest region (13.4%). In Brazil, most studies have employed molecular techniques and are concentrated in the Southeast region. The Blastocystis sp. subtype ST3 had the highest average positivity, followed by ST1 and ST2. These findings represent a panorama that reflects the reality of Brazil; thus, we believe that the effectiveness of parasitological diagnosis should be considered with regard to making an appropriate choice of technique for detecting Blastocystis sp. Additionally, we emphasize the importance of further studies in the context of molecular epidemiology with regard to this genus. Blastocystis sp. is not well understood yet, and very little information regarding this genus is available; hence, further research regarding this genus is urgently needed.

Four Corners: an educational strategy for learning infectious diseases in medical school / Four Corners: uma estratégia educacional para o ensino de infectologia para estudantes de Medicina

Abstract: Introduction: An extensive literature has demonstrated the benefits of active learning in medical education and has revealed the need for updating the teaching methodologies. Active learning strategies consolidate the theoretical knowledge, while simultaneously encouraging the student´s concrete abilities. Nonetheless, remodeling the pedagogical practice is challenging. Several active learning strategies are available and selecting the most suitable one is often a matter of format, not of content. In this article, we discuss the experience of the 'Four Corners' technique in the teaching of Infectious Diseases during the internship at our institution. Experience report: The four corners strategy can be adapted to different topics and should be implemented as follows: 1. pre-activity learning: students receive the supportive material for previous analysis; 2. Preparation of the room: each of the four corners is labeled with a different case vignette; 3. Dividing tasks: the specific assignments are divided among the students; 4. Activity: the students read the case vignette and discuss the questions, while the mediator has an observer's role, and 5. Debriefing: final discussion with all participants led by the mediator. The entire process takes from 2h30 to 3 hours. Discussion: The educational process emerges from the learner´s experience. Many instructors working in higher education believe they promote critical thinking and active teaching during their classes; however, the percentage of teachers who regularly use active teaching strategies is still low. A range of different active learning strategies have been described and many can be adapted to different scenarios; most approaches change the nature of the learning experience but do not always require a change in the given subject. The Four Corners technique is an active learning strategy that promotes debate and exchange of ideas among students. Conclusion: This experience report describes an active learning strategy for the teaching of Infectious Diseases to medical students. The medical education improvement goes through highlighting the teacher's role as a mediator of the learning process; proposing new teaching strategies is challenging and requires frequent adjustments. The Four Corners strategy improves student's engagement with the learning process, and it is an efficient strategy to comprise an extensive theoretical content in a relatively short discussion time.

Resumo: Introdução: Uma extensa literatura demonstra os benefícios do ensino ativo na educação médica e a necessidade de atualização das metodologias de ensino. As estratégias de aprendizagem ativa consolidam o conhecimento teórico e estimulam as habilidades conceituais do aluno. No entanto, reformar a prática pedagógica é desafiador. Existem diversas estratégias de aprendizagem ativa que sugerem uma adaptação do formato de entrega do conteúdo, de modo a adequá-las a diversos contextos. Neste artigo, discutimos a experiência da aplicação da técnica Four Corners no ensino de doenças infecciosas em nossa instituição. Relato de experiência: A estratégia Four Corners foi implementada da seguinte maneira: 1. preparação: os estudantes recebem material de estudo para análise prévia; 2. preparação da sala: cada um dos quatro cantos recebe cartelas com os casos clínicos escolhidos; 3. divisão de tarefas: as tarefas são divididas entre os estudantes; 4. atividade: os estudantes leem e discutem cada um dos casos propostos com mínima interferência do moderador; 5. finalização: discussão final com todos os participantes liderados pelo moderador. A duração de toda a tarefa varia de duas horas e meia a três horas. Discussão: O processo educacional emerge da experiência do aprendiz. Muitos professores do ensino superior acreditam que promovem o pensamento crítico por meio do ensino ativo em suas aulas, entretanto a porcentagem dos que aplicam essas técnicas ainda é baixa. Existe uma ampla variedade de técnicas ativas de ensino, e muitas delas se adaptam a diferentes cenários. A técnica Four Corners é uma estratégia de aprendizagem ativa que promove debate e troca de conceitos entre os estudantes. Conclusão: Este relato de experiência descreve uma estratégia de aprendizagem ativa para o ensino de doenças infecciosas para estudantes de Medicina. O aprimoramento da educação médica passa por entender o papel do professor como mediador do processo de aprendizagem. Propor novas estratégias de ensino é desafiador e requer ajustes frequentes. A estratégia Four Corners melhora o envolvimento do aluno com o processo de aprendizagem e é eficiente para compreender um conteúdo teórico extenso em um tempo de discussão relativamente curto.

Clinical-epidemiological and laboratory profiles of severe Schistosomiasis mansoni infections at a university hospital

OBJECTIVE: Schistosomiasis remains a public health problem. This was a descriptive and retrospective study of 42 patients with a severe form of schistosomiasis who were admitted to the outpatient clinic of the Faculdade de Medicina da Universidade de São Paulo, São Paulo, in São Paulo, Brazil. METHODS: A data collection questionnaire was designed from the patient charts, and the following variables were evaluated: age, sex, place of birth, occupation, signs and symptoms of schistosomiasis, data from laboratory and imaging examinations, data regarding treatment outcomes, and the existence of comorbidities. Statistical analysis was carried out using Statistical Package for the Social Sciences 15.0 and Microsoft Excel 2003 software. The significance levels of the tests were fixed, accepting 5% type 1 error (α=0.05). Since this was a retrospective observational study, not all data were available for analysis. RESULTS: The mean age of the patients was 48.24 years; 57.1% were male. Statistically significant associations were observed between splenomegaly and thrombocytopenia (p=0.004) and between splenomegaly and leukopenia (p=0.046); however, only 4.5% of the patients had esophageal hemorrhage. Some patients received a specific treatment; of those, 42% took praziquantel, and 35.4% took oxamniquine. Nonspecific drug therapy was given as follows: 65% received propranolol, 90% omeprazole, and 43.6% aluminum hydroxide. The other treatments were as follows: 92.9% of patients underwent endoscopic treatment, 85% received sclerotherapy, and 62.5% used elastic bandages. CONCLUSION: This preliminary study presents a multidisciplinary outpatient follow-up associated with endoscopic and drug treatments that may be effective at preventing bleeding.

Experimental toxocariasis in BALB/c mice: relationship between parasite inoculum and the IgG immune response

BALB/c mice were inoculated with 5-500 Toxocara canis infective eggs, and bled at 15-120 days post infection (dpi) to evaluate the dynamics of IgG antibody response and larvae distribution. Positive results were observed in all occasions for every inoculum, and a direct proportional relationship between antibody detection and the parasitic load was observed. In samples collected at 60 dpi, detection of IgG was more intense, especially with the 50 and 500 egg doses; also, a correlation between antibody level and egg count was observed with these two inocula. At 120 dpi, a decrease in antibody titer was observed for all groups; and at the end of the experiment, larvae were recovered from carcass, liver and brain. In the liver, larvae were only found in mice inoculated with 500 T. canis eggs. In carcasses, these were recovered in all groups, and the group inoculated with 50 eggs showed the highest percentage of larvae in the brain.

Membrane fractions from Strongyloides venezuelensis in the immunodiagnosis of human strongyloidiasis / Frações de membrana de Strongyloides venezuelensis para o imunodiagnóstico da estrongiloidíase humana

Strongyloides venezuelensis is a parasitic nematode of rodents frequently used to obtain heterologous antigens for the immunological diagnosis of human strongyloidiasis. The aim of this study was to evaluate membrane fractions from S. venezuelensis for human strongyloidiasis immunodiagnosis. Soluble and membrane fractions were obtained in phosphate saline (SS and SM) and Tris-HCl (TS and TM) from filariform larvae of S. venezuelensis. Ninety-two serum samples (n = 92) were obtained from 20 strongyloidiasis patients (Group I), 32 from patients with other parasitic diseases (Group II), and 40 from healthy individuals (Group III), and were analyzed by enzyme-linked immunosorbent assay (ELISA). Soluble fractions (SS and TS) showed 90.0% sensitivity and 88.9% specificity, whereas the membrane fractions (SM and TM) showed 95.0% sensitivity and 94.4% specificity. The present results suggest the possible use of membrane fractions of S. venezuelensis as an alternative antigen for human strongyloidiasis immunodiagnosis.

Strongyloides venezuelensis é um nematódeo parasita de roedores, frequentemente usado como antígeno heterólogo para o diagnóstico imunológico da estrongiloidíase humana. O objetivo deste estudo foi avaliar frações de membrana de S. venezuelensis para o imunodiagnóstico da estrongiloidíase humana. Para tanto, frações solúveis e de membrana foram obtidas em solução salina fosfato (SS e MS) e Tris-HCl (ST e MT) de larvas filarioides de S. venezuelensis. Amostras de soro de 92 indivíduos, sendo 20 com estrongiloidíase (Grupo I); 32 com outras parasitoses (Grupo II), e 40 indivíduos saudáveis (Grupo III), foram analisadas pelo teste Imunoenzimático (ELISA). As frações solúveis (SS e ST) apresentaram 90,0% e 88,9%, enquanto que as frações de membrana (MS e MT) demonstraram 95,0% e 94,4%, de sensibilidade e especificidade, respectivamente. Os resultados obtidos permitem indicar as frações de membranas como antígeno alternativo para o diagnóstico da estrongiloidíase humana.

Two sequential PCR amplifications for detection of Schistosoma mansoni in stool samples with low parasite load / Duas amplificações sequenciais por PCR para detecção de Schistosoma mansoni em amostras de fezes com baixa carga parasitária

Schistosomiasis constitutes a major public health problem, with an estimated 200 million individuals infected worldwide and 700 million people living in risk areas. In Brazil there are areas of high, medium and low endemicity. Studies have shown that in endemic areas with a low prevalence of Schistosoma infection the sensitivity of parasitological methods is clearly reduced. Consequently diagnosis is often impeded due to the presence of false-negative results. The aim of this study is to present the PCR reamplification (Re-PCR) protocol for the detection of Schistosoma mansoni in samples with low parasite load (with less than 100 eggs per gram (epg) of feces). Three methods were used for the lysis of the envelopes of the S. mansoni eggs and two techniques of DNA extraction were carried out. Extracted DNA was quantified, and the results suggested that the extraction technique, which mixed glass beads with a guanidine isothiocyanate/phenol/chloroform (GT) solution, produced good results. PCR reamplification was conducted and detection sensitivity was found to be five eggs per 500 mg of artificially marked feces. The results achieved using these methods suggest that they are potentially viable for the detection of Schistosoma infection with low parasite load.

A esquistossomose constitui grande problema de saúde pública, sendo que estimativas apontam para 200 milhões de pessoas infectadas no mundo e 700 milhões de pessoas em áreas de risco. No Brasil, existem áreas de alta, média e baixa endemicidade. Estudos demonstram que nas áreas endêmicas de baixa prevalência da infecção, a reduzida sensibilidade dos métodos parasitológicos torna-se evidente. Isto dificulta o diagnóstico, pela presença de resultados falso-negativos. O objetivo deste estudo foi a padronização de um protocolo de reamplificação da PCR (Re-PCR) para a detecção de Schistosoma mansoni em amostras com menos de 100 ovos por grama (opg) de fezes. Foram utilizados três métodos para ruptura dos envoltórios dos ovos de S. mansoni e duas técnicas de extração de DNA foram aplicadas. O DNA extraído foi quantificado e os resultados sugerem que a técnica de extração de melhor produtividade foi a que associa esferas de vidro a uma solução de isotiocianato de guanidina/fenol/clorofórmio (GT). Aplicou-se a Re-PCR, que demonstrou sensibilidade para a detecção de cinco ovos/500 mg de fezes artificialmente marcadas. Assim, essas novas ferramentas são potencialmente aplicáveis nas infecções por S. mansoni com baixa carga parasitária.

Abdominal ultrasound in the evaluation of fibrosis and portal hypertension in an area of schistosomiasis low endemicity

This study was undertaken in the municipality of Bananal, São Paulo, an endemic area for schistosomiasis with a prevalence under 10 percent and low parasite load among infected individuals. Our objective was to identify the clinical forms of schistosomiasis among 109 patients in whom the disease had been diagnosed through direct fecal analysis and who had been medicated with oxamniquine at the time of the Plan for the Intensification of Schistosomiasis Control Actions (1998-2000). These patients were submitted to an abdominal ultrasonography and fecal analysis by Kato-Katz method, four years, on average, after the end of the Plan. Five patients, whose abdominal ultrasound images were compatible with either peripheral or central periportal fibrosis and portal hypertension, were identified. None of the 109 patients presented Schistosoma mansoni eggs at fecal analysis. Ultrasonography is a sensitive, noninvasive diagnostic method that allows a better identification of the extent of liver involvement in schistosomiasis cases.

Este estudo desenvolveu-se no município de Bananal, São Paulo, uma área endêmica para esquistossomose com prevalência menor que 10 por cento e baixa carga parasitária nos infectados. Teve como objetivo a identificação de formas clínicas da esquistossomose mansoni através do exame ultra-sonográfico, em 109 pacientes diagnosticados parasitologicamente e medicados com oxamniquine, durante a realização do Plano de Intensificação das Ações de Controle da Esquistossomose mansônica (1998-2000). Foram utilizadas a ultra-sonografia abdominal e exames de fezes (Kato-Katz) realizados após o término do plano, quatro anos em média. Nesta casuística, foram identificados cinco pacientes com imagens ultra-sonográficas abdominais compatíveis com fibrose periportal periférica ou central e hipertensão portal, além da negatividade de todos os exames parasitológicos nos 109 pacientes. A ultra-sonografia, um método de diagnóstico sensível e não invasivo, possibilitou a identificação de casos com comprometimento hepático em uma área de baixa endemicidade para esquistossomose mansoni.

Eficácia e tolerabilidade da associação oxamniquine com praziquantel no tratamento da infecção humana por Schistosoma mansoni / Efficacy and tolerability of the combination of oxamniquine and praziquantel in the treatment of human Schistosoma mansoni infection

Objetivo: Avaliar a eficácia e a segurança da associação de oxamniquine com praziquantel na quimioterapia da esquistossomose mansoni. Métodos: Foram tratados 131 pacientes com diagnóstico parasitológico de esquistossomose mansoni, divididos em dois grupos: no grupo I (n=61), receberam a associação nas doses de 12,5 mg/kg de peso de ambas as drogas, enquanto que os pacientes do grupo II (n=70) receberam doses de 7,5 mg/kg de peso do oxamniquine e 15 mg/kg de peso de praziquantel. Em ambos os grupos predominaram as formas clínicas mais brandas, bem como baixas cargas parasitárias. Os controles de cura foram realizados através de, no mínimo, seis coproscopias após o tratamento. Resultados: Cumpriram esse controle, 44 pacientes no grupo I, dos quais 26 (59,1 por cento) tiveram a totalidade dos exames de fezes negativos e 49 pacientes no grupo II, no qual apenas 20 (40,8 por cento) tiveram todas as coproscopias de controle negativas. Conclusões: Esses resultados indicam eficácia inferior àquela observada para ambas as drogas administradas isoladamente em doses plenas. A ocorrência de efeitos adversos com o uso dessa associação medicamentosa foi pouco importante, tanto em relação à frequência das reações como à sua gravidade

Diagnóstico e tratamento de parasitoses intestinais / Diagnosis and treatment of intestinal parasitosis

Säo revistas as principais técnicas utilizadas para o diagnóstico laboratorial de rotina das parasitoses intestinais prevalentes em nosso meio, bem como os esquemas terapêuticos habitualmente empregados.

Contribuiçao dos metodos por imagem no diagnostico evolutivo do carbunculo renal / Contribution of image methods in evolutive diagnostic of renal carbuncle

Os autores apresentam um caso de carbunculo renal numa paciente de 29 anos sem fatores debilitantes previos. Amparados pela revisao da literatura ressaltam a importancia a ultra-sonografia e da tomografia computadorizada no diagnostico e no acompanhamento clinico desta enfermidade