Serological diagnosis of paracoccidioidomycosis using a Paracoccidioides spp. comercial antigen and the counterimmunoelectrophoresis method

ABSTRACT Purpose: In-house Paracoccidioides spp. antigens are commonly used in the serological diagnosis of paracoccidioidomycosis (PCM). The sensitivity and specificity of a commercial Paracoccidioides spp. antigen was assessed for PCM serological testing. Method: Counterimmunoelectrophoresis and double immunodiffusion were used to evaluate the Paracoccidioides ID Antigen reagent in sera from PCM cases and patients with other diseases. Results: All active PCM sera (n=24) were reactive using counterimmunoelectrophoresis (sensitivity = 100%), including 11 cases of infection by P. brasiliensis sensu stricto and one by P. americana. Fifteen (88%) out of 17 sera from patients on treatment or cured were reactive, including one case of P. lutzii infection. One to three bands of antigen-antibody precipitate were observed on the agarose gel, with a predominance of two to three bands in the test with untreated PCM sera or at the beginning of antifungal therapy. All sera from patients with histoplasmosis (n=7), aspergillosis (n=5), and other diseases (n=27) tested negative (specificity = 100%). The overall sensitivity and specificity using the commercial antigen and double diffusion test were 75% and 100%, respectively. Conclusion: The commercial antigen performed satisfactorily and may contribute to the dissemination of the use of serological tests for the PCM diagnosis.

Impacto do uso da reação em cadeia da polimerase para o diagnóstico de meningite bacteriana em substituição a contraimunoeletroforese / Impact of using polymerase chain reaction for the diagnosis of bacterial meningitis in substitution for counterimmunoelectrophoresis

Introdução: a meningite bacteriana é um grave problema de Saúde Pública mundial, tendo como principais agentes: Neisseria meningitidis, Streptococcus pneumoniae e Haemophilus influenzae. A metodologia de diagnóstico empregada no Instituto Adolfo Lutz ­ Centro de Laboratório Regional Santo André até o ano de 2011 era a contraimunoeletroforese (CIE), depois foi substituída pela reação em cadeia da polimerase em tempo real (qPCR), que apresenta maior sensibilidade. Objetivo: este trabalho objetivou comparar ambas as metodologias no período de 2009 a 2018, para avaliação do impacto da introdução da qPCR no diagnóstico das meningites bacterianas nos 7 municípios da região do ABC do Estado de São Paulo. Metodologia: foram avaliadas a quantidade total de exames realizados, a média mensal, a positividade no período, os municípios requisitantes e a prevalência das bactérias causadoras de meningite, no período de abril/2009 até dezembro/2018. Resultados: Foram 377 exames de CIE e 1305 de qPCR, com média anual de 230 exames em 2010-2013 e 130 exames em 2014-2018. Observou-se aumento da positividade entre as técnicas, 17,8% para CIE e 33,8% para qPCR. N. meningitidis foi responsável pela maioria dos casos entre 2011 e 2013, cerca de 61% dos casos positivos, enquanto que entre 2014 e 2018 foi S. pneumoniae, cerca de 53%. Conclusão: os resultados indicaram que a qPCR foi mais eficiente em detectar os agentes causadores de meningite bacteriana na região do que a técnica de CIE. Por fim, este trabalho suporta a implantação da metodologia de qPCR para diagnóstico de meningite em substituição de técnicas menos sensíveis.

Introduction: bacterial meningitis is still a serious worldwide public health problem, and the main etiological agents are: Neisseria meningitidis, Streptococcus pneumoniae and Haemophilus influenzae. The diagnostic methodology employed at the Adolfo Lutz Institute ­ Santo André Regional Laboratory Center until 2011 was the ounterimmunoelectrophoresis (CIE), then it was replaced by the real-time polymerase chain reaction (qPCR), which is more sensitivity. Objective: this study aimed to compare both methodologies from 2009 to 2018 to evaluate the impact of the introduction of qPCR in the diagnosis of bacterial meningitis in the 7 cities of the ABC region of São Paulo State. Methodology: the total number of tests performed, the month average, the positivity in the period, the requesting cities and the prevalence of bacteria causing meningitis were evaluated from April/2009 to December/2018. Results: there were 377 CIE exams and 1305 qPCR exams, with an annual average of 230 exams in 2010-2013 and 130 exams in 2014-2018. There was an increase in positivity between the performed techniques, 17.8% for CIE and 33.8% for qPCR. N. meningitidis accounted for most cases of bacterial meningitis between 2011 and 2013, about 61% of positive cases, whereas between 2014 and 2018 it was S. pneumoniae, with about 53%. Conclusion: the results indicated that qPCR was more efficient in detecting the agents that cause bacterial meningitis in the region than the CIE technique. Finally, this work supports the implementation of qPCR methodology for diagnosis of meningitis in replacement of less sensitive techniques.

Meningite aguda bacteriana na população infantojuvenil: principais agentes e métodos para diagnóstico - uma revisão da literatura / Acute bacterial meningitis in children and young population: main agents and methods for diagnosis - a literature review

A meningite bacteriana é uma afecção de grande significância devido a sua relação com alta mortalidade e morbidade na população neonatal a jovem. Devido a este fato é importante o conhecimento sobre esta doença e os seus principais agentes etiológicos. Com o objetivo de relatar os principais métodos de diagnóstico, assim como os principais agentes etiológicos envolvidos na fisiopatologia da meningite bacteriana em população jovem foi realizada uma busca por artigos publicados nos últimos 5 anos nas bases de dados Pubmed, Scielo, Bireme e Lilacs. A literatura atual aponta como microrganismos predominantes na incidência dessa doença a N. meningitidis S. pneumoniae, sendo as mais recorrentes na população de faixa etária entre 29 dias e 17 anos. Os fatores relacionados ao prognóstico estão intimamente relacionados com a distinção da classificação do agente etiológico em bacteriano ou viral, importante para a determinação da terapia adequada.

Bacterial meningitis is a highly significant disease due to its relationship with high mortality and morbidity in neonatal and young population. Due to this is important to know about this disease and its main etiological agents. In purpose to report the main diagnostic methods, as well as the main etiological agents involved in the pathophysiology of bacterial meningitis was conducted a search for articles published in the last five years in Pubmed, Scielo, Bireme and Lilacs. The current literature indicates N. meningitidis S. pneumoniae the most predominant microorganisms in the incidence of this disease between population aged between 29 days and 17 years. Factors related to prognosis are closely related to the classification distinction of the etiologic agent in bacterial or viral, important to determine the ap- propriate therapy.

Diagnóstico serológico de la esporotricosis mediante el empleo del antígeno de micelio de Sporothrix schenckii sensu stricto / Serological diagnosis of sporotrichosis using an antigen of Sporothrix schenckii sensu stricto mycelium

En este estudio se desarrolló y se evaluó el ensayo por inmunoabsorción ligado a enzimas (ELISA), para la detección de anticuerpos en sueros de pacientes con esporotricosis, para lo cual se empleó un antígeno crudo de Sporothrix schenckii sensu stricto obtenido a partir de la forma micelial. Los sueros positivos para esporotricosis fueron ensayados por otras técnicas serológicas: inmunodifusión doble (IDD) y contrainmunoelectroforesis (CIE). El ensayo fue validado utilizando sueros de otras patologías como histoplasmosis, paracoccidioidomicosis, tuberculosis, leishmaniasis, lupus y sueros de individuos sanos como controles negativos. Se encontró una especificidad de 100 % con las técnicas utilizadas y una sensibilidad del antígeno de S.schenckii sensu stricto, por encima del 98% para IDD, CIE y ELISA. Estos resultados demuestran la alta sensibilidad y especificidad del antígeno de S. schenckii sensu stricto, para el diagnóstico de la esporotricosis, empleando las técnicas de IDD, CIE y ELISA. Los resultados sugieren, que este antígeno podría ser usado en conjunto con otras pruebas convencionales para el diagnóstico diferencial y puede ser útil para monitorizar la evolución de la enfermedad y respuesta al tratamiento.

We developed and analyzed an Enzyme-Linked Immunosorbent Assay (ELISA) in order to detect antibodies in sera from sporotrichosis patients. We used a crude antigen of Sporothrix schenckii sensu stricto, obtained from the mycelial phase of the fungi. Positive sera were analyzed by other serological techniques such as double immunodiffusion (IGG) and counterimmunoelectrophoresis (CIE). The assay was validated by using sera from patients with other pathologies such as: histoplasmosis, paracoccidioidomycosis, tuberculosis, leishmaniasis, lupus and healthy individuals as negative controls. For the Sporothrix schenckii sensu stricto antigen, we found a 100% of specificity by every technique and sensitivity higher than 98% with IDD, CIE and ELISA. Our results show a high sensitivity and specificity for the Sporothrix schenckii sensu stricto antigen, so it can be used for IDD, CIE and ELISA. The results suggest that this antigen could be used in conjunction with other conventional tests for differential diagnosis and may be useful for monitoring the disease progression and response to treatment.

Evolution of bacterial meningitis diagnosis in Sao Paulo State-Brazil and future challenges / Evolucao do diagnostico das meningites bacterianas no Estado de Sao Paulo-Brasil e os futuros desafios

Bacterial meningitis (BM) is a severe disease and still represents a serious public health problem with high rates of morbidity and mortality. The most common cases of BM around the world, mainly in Brazil, have been caused by Neisseria meningitidis, Streptococcus pneumoniae, and Haemophilus influenzae type b. Bacterial culture is the gold-standard technique for BM confirmation, but approximately 50% of suspected cases are not culture-confirmed, due to problems related to improper transportation and seeding or previous antibiotic treatment. Immunological methods present low sensitivity and have possibility of cross-reactions. Real time PCR (qPCR) is a molecular technique and has been successful used for BM diagnosis at Instituto Adolfo Lutz in São Paulo State, Brazil, since 2007. The incorporation of qPCR in the Public Health surveillance routine in our state resulted in diminishing 50% of undetermined BM cases. Our efforts are focused on qPCR implementation in the BM diagnostic routine throughout Brazil.

A meningite bacteriana (MB) é uma doença grave e ainda representa um sério problema de saúde pública, com altas taxas de morbidade e mortalidade. Os casos mais comuns de MB em todo o mundo, principalmente no Brasil, tem sido causados por Neisseria meningitidis, Streptococcus pneumoniae e Haemophilus influenzae tipo b. Cultura bacteriana é a técnica padrão-ouro para a confirmação de MB, mas cerca de 50% dos casos suspeitos não são confirmados por cultura, devido a problemas relacionados ao transporte inadequado e semeadura ou antibioticoterapia prévia. Métodos imunológicos apresentam baixa sensibilidade e têm possibilidade de reações cruzadas. PCR em tempo real (qPCR) é uma técnica molecular e tem sido utilizada com êxito para o diagnóstico de MB no Instituto Adolfo Lutz, em São Paulo, Brasil, desde 2007. A incorporação da qPCR na rotina de vigilância em Saúde Pública em nosso estado resultou na diminuição de 50% dos casos de MB indeterminadas. Nossos esforços estão focados na implementação da qPCR na rotina diagnóstica de MB em todo o Brasil.

Comparación de tres pruebas serológicas para detectar infecciones por Paragonimus mexicanus en gatos infectados / Comparison of three serological tests to detect Paragonimus mexicanus infections in infected cats

Introducción: La paragonimiasis es una enfermedad pulmonar causada por tremátodes del género Paragonimus. Generalmente, el diagnóstico de laboratorio de la infección en humanos se realiza mediante la detección de huevos del parásito en esputo o heces; sin embargo, los resultados pueden ser negativos, necesitándose métodos diagnósticos alternativos. Objetivos: Comparar tres pruebas serológicas para la detección de anticuerpos contra el antígeno somático del estadio adulto de Paragonimus mexicanus (PmAS) en muestras de suero de gatos infectados experimentalmente. Diseño: Estudio experimental. Lugar: Bioterio del Instituto de Medicina Tropical ôDaniel A. Carriónõ de la Facultad de Medicina, Universidad Nacional Mayor de San Marcos. Material biológico: Gatos domésticos de 4 a 6 meses de edad y antígeno somático crudo de P. mexicanus (PmAS). Intervenciones: Se obtuvo muestras de sangre de 12 gatos domésticos infectados experimentalmente con P. mexicanus. Los sueros fueron evaluados por el inmunoensayo enzimático (ELISA), la doble difusión (DD) y contrainmunoelectroforesis (CIEF). Principales medidas de resultados: Se determinó el porcentaje de gatos que revelaron resultados positivos de acuerdo a los criterios establecidos. Resultados: Se encontró una buena correlación entre los resultados obtenidos por ELISA y aquellos obtenidos por CIEF o DD. La CIEF y el ELISA fueron más sensibles que la doble difusión. Con la CIEF y el ELISA, 81,8 por ciento (9/11) y 75,0 por ciento (9/12) de los sueros evaluados dieron resultados positivos, respectivamente, mientras que 58,3 por ciento (7/12) de ellos fue positivo para la DD. Los sueros de 2 conejos con fasciolosis experimental dieron una reacción positiva con el antígeno PmAS por ELISA y CIEF. No se observó reacción cruzada cuando se usó sueros de gatos con dipilidiasis o de gatos sanos. Conclusiones: Estos resultados indican que la CIEF y el ELISA, que emplean antígeno somático crudo, son pruebas de similar sensibilidad para el diagnóstico de paragonimiosis, pero no pueden ser usadas para el diagnóstico diferencial de las infecciones por P. mexicanus y F. hepatica, debido a la ocurrencia de reacciones cruzadas.

Background: Paragonimiasis is a lung disease caused by Paragonimus trematodes genus. Human infection laboratory diagnosis is usually done by detection of parasite eggs in sputum or feces; however, findings may be negative and alternative diagnostic methods are required. Objectives: To compare three serologic tests for detection of Paragonimus mexicanus somatic antigen (PmAS) antibodies in serum samples of cats infected experimentally. Design: Experimental study. Setting: Instituto de Medicina Tropical ôDaniel A. Carriónõ bioterium, Faculty of Medicine, Universidad Nacional Mayor de San Marcos, Lima, Peru. Biologic material: Domestic cats 4-6 years old and P. mexicanus (PmAS) crude somatic antigen. Interventions: Blood samples were collected from twelve domestic cats infected with P. mexicanus. Serum samples were evaluated by enzyme-linked immunosorbent assay (ELISA), double diffusion (DD) and counterimmunoelectrophoresis (CIE). Results: Good correlation was found between ELISA results and those of CIE or DD. Both CIE and ELISA were more sensitive than DD. With CIE and ELISA sera were respectively positive 81.8 per cent (9/11) and 75.0 per cent (9/12), whereas 58.3 per cent (7/12) were positive for DD test. Sera from 2 rabbits with experimental fascioliosis had positive reaction with PmAS antigen by both ELISA and CIE tests. No cross-reactivity was observed using sera from cats with dipylidiasis or from healthy cats. Conclusions: Results showed that both ELISA and CIE were equally sensitive tests for paragonimiosis diagnosis but could not be used for differential diagnosis of P. mexicanus and F. hepatica infections due to cross-reaction.

Serological diagnosis of paracoccidioidomycosis in HIV-coinfected patients

Paracoccidioidomycosis should be differentiated from other opportunistic diseases in human immunodeficiency virus (HIV)/acquired immunodeficiency syndrome (AIDS) patients who live in Latin America. Laboratory investigation can begin with serological tests, which are rapid and efficient. In the present study, double immunodiffusion (DID), counterimmunoelectrophoresis (CIEP) and an enzyme linked immunosorbent assay (ELISA) tests were assessed for the detection of anti-Paracoccidioides brasiliensis antibodies in 40 patients coinfected with HIV. The results were compared to those obtained for 75 non-HIV-infected patients with endemic paracoccidioidomycosis. Anti-P. brasiliensis antibodies were detected in 65 percent (DID), 79 percent (CIEP) and 95 percent (ELISA) of the patients with HIV/AIDS, significantly lower rates than those detected in cases of endemic paracoccidioidomycosis, which were 89 percent, 99 percent and 100 percent, respectively. The reactive sera of HIV-infected patients also showed lower anti-P. brasiliensis antibody titres than those of non-HIV-infected patients. Despite the lower intensity of the specific humoral response, serological tests are useful for the diagnosis of opportunistic paracoccidioidomycosis in the HIV/AIDS population. We suggest optimization of the laboratory diagnosis by combining the ELISA test with CIEP or DID.

Community-acquired pneumonia in the childhood: analysis of the diagnostic methods

Immunological assays such as CIE, LA, and Dot-ELISA were compared in order to diagnose community-acquired pneumonia. Serum, pleural fluid and urine samples were comparatively employed for bacterial antigen detection. Dot-ELISA proved to be an original and practical alternative procedure for detecting bacterial polysaccharide antigens from pleural fluid and/or concentrated urine samples, providing a rapid diagnosis for pediatric patients with community-acquired pneumonia.

Antinuclear antibodies and HLA class II alleles in Jamaican patients with systemic lupus erythematosus / Anticuerpos antinucleares y aleles HLA de clase II en pacientes Jamaicanos con lupus eritematoso sistémico

OBJECTIVE: The relationship between human leukocyte antigens class II (HLA) and antinuclear antibodies was investigated in Jamaican patients with Systemic Lupus Erythematosus (SLE). METHODS: Samples of blood of 82 patients with SLE and 75 healthy controls were tested for antinuclear antibodies using the fluorescent antinuclear antibody (FANA) test, counterimmunoelectrophoresis (CIEP) and the Crithidia luciliae immunofluorescence test (CL-IFT). A DNA-based HLA typing method was used to determine the frequencies of alleles of HLA-DRB1, DRB3, DRB4 and DRB5 in patients and healthy controls. RESULTS: The FANA test was positive in all of the sera from patients with SLE. Anti-dsDNA antibodies were present in 49% (40/82), anti-Sm/RNP 44% (36/82) and anti-Ro/La 43% (35/82) of the sera from SLE patients. The frequency of HLA-DR4 was significantly lower in SLE patients than in healthy controls (2/82, 2% vs 15/75, 20%; RR = 0.12; p = 0.0004; CP = 0.005) but no other HLA-DRB1 SLE associations were found. A positive HLA-DR3 anti-Ro/La antibody association was found in the patients with SLE (9/21, 43% vs 5/55, 9%; odds ratio (OR) = 7.5; CP = 0.01). In contrast, possession of HLA-DR6 was negatively associated with the absence of anti-dsDNA antibodies (9/32, 28% vs 27/44, 61%; OR = 0.2; CP = 0.05). CONCLUSION: The HLA-DR6 allele is associated with the absence of antinuclear antibodies and HLA-DR3 with the presence of anti-Ro/La antibodies in Jamaican patients with SLE. However, these results and those of previous studies of Jamaican patients suggest that the HLA-DR3 association with the development of SLE reported in other populations might in fact reflect the association of HLA-DR3 with anti-Ro/La antibodies. Further investigations are needed to determine whether HLA-DRB antinuclear antibody associations define clinical subsets of SLE in Jamaican patients.

OBJETIVO Se investigó la relación entre los antígenos de leucocito humano (human leukocyte antigens o HLAs). Clase II y los anticuerpos antinucleares en pacientes jamaicanos con lupus eritematoso sistémico (LES). MÉTODOS: Se examinaron muestras de sangre de 82 pacientes con LES y 75 controles saludables para determinar la presencia de anticuerpos antinucleares, usando la prueba del anticuerpo antinuclear fluorescente (FANA), la contrainmunoelectroforesis (CIEP) y el test de inmunofluorescencia con Crithidia luciliae (CL-IFT). Un método de tipificación HLA basado en el ADN fue usado para determinar las frecuencias de aleles de HLA-DRB1, DRB3, DRB4 y DRB5 tanto en los pacientes como en los controles saludables. RESULTADOS: La prueba FANA fue positiva en todos los sueros de pacientes con LES. Anticuerpos anti-dsADN se hallaban presentes en 49% (40/82), anti-Sm/RNP en 44% (36/82) y anti-Ro/La en 43% (35/82) de los sueros de los pacientes de LES. La frecuencia de HLA-DR4 fue significativamente más baja en los pacientes con LES que en los controles saludables (2/82, 2% vs 15/75, 20%; RR = 0.12; p = 0.0004; CP = 0.005) pero no se hallaron otras asociaciones de LES con HLA-DRB1. Se halló una asociación positiva de anticuerpos HLA-DR3 anti-Ro/La en los pacientes con LES (9/21, 43% vs 5/55, 9%; odds ratio (OR) = 7.5; CP = 0.01). En contraste con ello, la posesión de HLA-DR6m estuvo asociada negativamente con la ausencia de anticuerpos anti-dsADN (9/32, 28% vs 27/44, 61%; OR = 0.2; CP = 0.05). CONCLUSIÓN: El alele HLA-DR6 está asociado con la ausencia de anticuerpos antinucleares y el de HLA-DR3 con la presencia de anticuerpos anti-Ro/La en pacientes jamaicanos con LES. Sin embargo, estos resultados al igual que los de los previos estudios de pacientes jamaicanos, sugieren que la asociación HLA-DR3 con el desarrollo de LES reportado en otras poblaciones podría de hecho reflejar la asociación de HLA-DR3 con anticuerpos anti-Ro/La. Se requieren investigaciones ulteriores a fin de determinar si las asociaciones de anticuerpo antinuclear HLA-DRB definen subconjuntos de LES en pacientes jamaicanos.

Comparative efficacy of standard AGID, CCIE and competitive ELISA for detecting bluetongue virus antibodies in indigenous breeds of sheep and goats in Rajasthan, India

The sero-prevalence of antibodies against blue tongue virus (BTV) in 408 local breeds of sheep in Rajasthan state in India was investigated using standard agar gel immunodiffusion (AGID) test. Maximum seropositivities of 11.3% (13/115), 10.7% (13/121), 7.1% (11/155) and 5.9% (1/17) were recorded in the Chokla, Magra, Nali and Pugal breeds, respectively. Out of 107 goat serum samples, 6 (5.6%) were AGID positive. The performance of the standard AGID, counter current immuno-electrophoresis (CCIE) and the competitive enzyme-linked immunosorbent assay (cELISA) for the detection of serum antibody against BTV in indigenous breeds of sheep were compared. Out of 178 sheep serum samples tested, 17 (9.5%), 22 (12.3%) and 54 (30.3%) were positive for group-specific bluetongue antibodies by AGID, CCIE and cELISA, respectively. There was appreciable difference in the seroprevalence detected by AGID, CCIE and cELISA in clinically healthy and diseased sheep with regard to relative sensitivities and specificities of the tests with cELISA being highly sensitive and specific followed by CCIE and AGID test. It was concluded that these indigenous breeds of sheep may be a potential reservoir of BTV infection and cELISA should be routinely used for the detection of antibodies against BTV in these local breeds of sheep.

Cross- reaction of sera from patients with various infectious diseases with Leishmania infantum

To compare and evaluate the application of indirect fluorescent antibody [IFA] and counterimmunoelectrophoresis [CIEP] for laboratory identification of visceral leishmaniasis. Materials and Serum samples from patients with malaria [Plasmodium vivax, n = 86; Plasmodium falciparum, n = 38], brucellosis [n = 26], tuberculosis [n = 31] and typhoid fever [n = 35] were examined for the presence of antibody to Leishmaniainfantum antigen using IFA and CIEP tests. Using IFA, false-positive results were malaria [P. vivax 19.8%, P. falciparum 13.2%], tuberculosis [6.4%], brucellosis [3.8%], and typhoid fever [2.8%]. Using CIEP, a lower percentage of false-positives was observed only among malaria patients [P. vivax 2.3%, P. falciparum 2.6%]. Serum samples from patients with other infectious diseases were negative in the CIEP test. Based on the results of this study, the CIEP technique is recommended for immunodiagnosis of visceral leishmaniasis, especially in regions where malaria, brucellosis and tuberculosis are prevalent

A importância da contra-imunoeletroforese na detecção de antígenos nucleares extraíveis para o diagnóstico de doenças reumáticas sistêmicas / The importance of counterimmunoelectrophoresis in the detection of extractable nuclear antigens for the diagnosis of systemic rheumatic diseases

Os antígenos nucleares extraíveis (ENAs) são encontrados no soro da maioria dos pacientes com doença reumática sistêmica. Os principais ENAs estudados são SS-A/Ro, SS-B/La, RNP, Sm, Scl-70 e Jo-1. Objetivou-se neste trabalho: a) padronizar a técnica de contra-imunoeletroforese (CIE) para a detecção de ENAs; b) padronizar o substrato antigênico (ENAs) para a CIE a partir de baço de cão; c) comparar os resultados da CIE com as técnicas de imunofluorescência indireta (IFI) e Elisa para esses antígenos. Para tal foram estudados 40 soros de pacientes com doença reumática sistêmica confirmada por exames clínico e laboratorial (sorológico e biópsia). Como controle negativo foram utilizados dez soros de doadores de sangue normais, e, como controles positivos, seis soros-padrão anti-Ro, anti-La, anti-RNP, anti-Sm, anti-Scl-70, anti-Jo-1, para caracterizar os antígenos presentes no extrato de baço. Neste último foram detectados vários ENAs, exceto RNP e Scl-70. A técnica de CIE apresentou boas sensibilidade (70 por cento) e especificidade (100 por cento) em relação às outras técnicas (IFI e Elisa). A titulação dos soros pela CIE revelou positividade até diluições de 1:16 em 32,5 por cento dos casos. Concluímos que a CIE e os antígenos extraídos de baço de cão podem ser utilizados na rotina laboratorial para triagem destes ENAs, com a vantagem, em relação à IFI, de poderem ser titulados.

Immunoprecipitation techniques and Elisa in the detection of anti-Fonsecaea pedrosoi antibodies in chromoblastomycosis

Chromoblastomycosis (CBM) is a chronic subcutaneous infection caused by several dematiaceous fungi. The most commonly etiological agent found in Brazil is Fonsecaea pedrosoi, which appears as thick walled, brownish colored cells with transverse and longitudinal division in the lesions, called "muriform cells". This disease is found worldwide but countries like Madagascar and Brazil have highest incidence. Diagnosis is made by clinical, direct and histopathologic examination and culture of specimens. Serological tests have been used to identify specific antibodies against Fonsecaea pedrosoi antigens, as well as immunotechniques have been used for CBM serological identification and diagnosis. In the present study double immunodiffusion (DID), counterimmunoelectrophoresis (CIE) and immunoenzymatic test (ELISA) have been used to evaluate humoral immune response in patients with CBM caused by F. pedrosoi. Metabolic antigen was used for immunoprecipitation tests (DID and CIE) while somatic antigen for ELISA. Our results demonstrated 53 percent sensitivity and 96 percent specificity for DID, while CIE presented 68 percent sensitivity and 90.5 percent specificity. ELISA demonstrated 78 percent sensibility and 83 percent specificity. Serological tests can be a useful tool to study different aspects of CBM, such as helping differential diagnosis, when culture of the pathogenic agent is impossible.

C-reactive protein in the diagnosis of community-acquired pneumonia

Qualitative determination of C-reactive protein (CRP) was evaluated as a diagnostic method for community-acquired pneumonia. Paired serum and pleural fluid samples from child patients were examined with a CRP test, compared to bacterial cultures, counterimmunoelectrophoresis and immunoassay. The CRP test gave excellent parameters of sensitivity, specificity and predictive values for the diagnosis of bacterial pneumonia

Inquérito sorológico da infecçäo pelo Echinococcus sp no município de Sena Madureira, AC / A serological survey of the infection by Echinococcus sp. in the municipal ity of Sena Madureira, AC

Foi realizado no município de Sena Madureira, Acre, um inquérito soroepidemiológico para avaliar o contato de indivíduos com o Echinococcus sp. Escolheu-se duas populações distintas, uma residente na área urbana e a outra na área rural do município, distribuída em comunidades ribeirinhas da regiäo. Foram avaliados no total 1.064 indivíduos, dos quais 851 pertencentes à área urbana e 213 à área rural. O estudo foi dividido em duas partes: inquérito sorológico, no qual foram coletadas e enviadas amostras de sangue ao Laboratório de Sorologia do Hospital das Clínicas da Faculdade de Medicina de Ribeirão Preto da Universidade de São Paulo, onde foram realizados os testes imunológicos pela técnica de contraimunoeletroforese; inquérito epidemiológico para avaliaçäo individual, além de as condições de moradia e hábitos dos indivíduos. A prevalência sorológica na área rural foi 6 por cento, enquanto na urbana fomos 3,5 por cento. A prevalência geral foi 4 por cento. Quando analisada a possibilidade da existência de outros hospedeiros intermediários no ciclo do Echinococcus vogeli, foram encontrados resultados que sugerem ser o porco doméstico o mais provável

Inquérito soroepidemiológico para infecções por fungos causadores de micoses sistêmicas na Reserva Indígena Xacriabá, Estado de Minas Gerais / Seroepidemiological survey for infections by fungi causing systemic mycoses in the Xacriabá Indian Reserve, Minas Gerais State, Brazil

Xacriabá Indian Reserve is situated in the north of Minas Gerais State, Brazil, near the municipality of Manga and São Francisco River. The Indian population is miscegenated with caucasian and negroid people and have farming activities. Blood samples were collected from 180 inhabitants of the reserve (85 men and 95 women), 15 to 84 years old. Serum antibodies against antigens of Paracoccidioides brasiliensis, Histoplasma capsulatum, Cryptococcus neoformans, and Candida albicans were tested for by counterimmunoelectrophoresis method. Seropositivity was verified in 5, 3.9, 4.2, and 6.7, respectively of the 180 samples. Those seropositive for antigens of P. brasiliensis and H. capsulatum were predominantly women and had lower age and more elevated mean titers of antibodies than individuals whose sera reacted with antigens of C. neoformans and C. albicans. The results suggest the occurrence of paracoccidioidomycosis-infection and histoplasmosis-infection in the Xacriabá Indian Reserve, particularly in the first decades of life of the inhabitants.

Comparison between the counter immunoelectrophoresis test and mouse neutralization test for the detection of antibodies against rabies virus in dog sera

The detection of rabies antibodies is extremely valuable for epidemiological studies, determination of immune status in man, animals, and for the diagnosis of the disease. Several serological procedures have been described for this purpose. The present study reports a comparison between counterimmunoelectrophoresis test (CIET) and mouse neutralization test (MNT) in the detection of antibodies against rabies virus from 212 serum samples of vaccinated dogs. The agreement between both techniques was 79.7 percent and a significative association was demonstrated. The correlation coefficients between MNT and the CIET titers was determined considering 88 samples showing positive results in both techniques [CIET = 2 and MNT = 5 (0.13 IU/ml)] and resulted r² = 0.7926 (p < 0.001). The performance of CIET system was technically simple, cheap and rapid, and thereby it could be useful for serological monitoring of dog vaccination campaigns as well as for individual analysis

Contribuiçäo de amostras de sangue no diagnóstico laboratorial da doença meningocócica / Contribution of blood samples in laboratorial diagnosis of meningococcal disease

Foram estudados retrospectivamente 73 casos de doença meningocócica (DM), dos quais, analisou-se amostras de líquido céfalorraquidiano (LCR) e de sangue, empregando-se as técnicas de cultura e pesquisa de antígenos polissacarídicos, através da contraimunoeletroforese (CIE) e pela reaçäo de aglutinaçäo de partículas de látex sensibilizadas por antissoros específicos (LA). Os resultados laboratoriais revelaram que 45,2 por cento das amostras foram positivas através da análise do LCR; 41,0 por cento foram positivas pela análise do LCR, hemocultura e/ou soro e 13,6 por cento foram positivas através da análise de hemocultura e/ou soro. Quanto ao tipo de exame realizado com o sangue, notou-se que 35,2 por cento dos casos foram positivos pela hemocultura, e quando se pesquisou o antígeno no soro dos pacientes, obteve-se 24,6 por cento de positividade pela CIE e 46,7 por cento pelo LA. Constatou-se que 13,6 por cento dos casos, só tiveram a confirmaçäo laboratorial, através dos exames realizados em amostras de sangue, destacando-se assim, a importante contribuiçäo desse tipo de material na identificação do agente etiológico e no aumento da positividade do diagnóstico laboratorial da DM. (AU)

73 cases of meningococcal disease (MD) have been studied retrospectively in whichsamples of cerebrospinal fluid (CSF) and blood were analysed using culture and meningococcal antigendetection through counterimmunoelectrophoresis (CIE) and reaction of latex agglutination (LA). Thelaboratory results showed that 45,2% of the samples were found positive in the CSF alone; 41,0% in theCSF, blood culture and/or serum, and only 13,6% in the blood culture and/or serum. As for the type of testcarried out in the blood, it was observed that 35,2% of the cases were positive in the blood culture, whilethe tests of antigen detection in the serum, showed that 24,6% were positive by CIE and 46,7% by LA. Itwas noticed that 13,6% of the cases only had the laboratorial confirmation through blood samples, thusshowing the important contribution of the blood in the identification of the ethiological agent and theincrease of the positivity of MD laboratorial diagnosis. (AU)

Recognition of antigenic components of fasciola gigantica and their use in immunodiagnosis

Antigenicity of the tegumental extract, excretory-secretory products and the whole somatic extract of Fasciola gigantica were evaluated to detect the most sensitive and specific antigen out of them that could be used as an immunodiagnostic tool. Scanning electron microscopic study was carried out to have a full picture for the tegumental structure. The immunohistochemical staining technique was done using the indirect immunoperoxidase method on F. gigantica sections before and after removal of the tegument to recognize the most antigenic parts by using patients sera. Counter immunoelectrophoresis was carried out by using sera from patients with fascioliasis [positive control], patients with schistosomiasis and healthy individuals [control group]. In addition, antibody response against the three types of antigens was detected as ELISA absorbance readings. The results revealed that antigens of F. Gigantica that cause antibody formation in hosts are those generated and released mainly from the tegument and they were the most sensitive and specific

Evaluación de reactivos cubanos elaborados para el diagnóstico rápido de Haemophilus influenzae tipo B / Evaluation of Cuba-made reagents for the rapid diagnosis of Haemophilus influenzae type B Formato PDF

Se probaron en la detección del antígeno capsular específico de haemophilus influenzae tipo b (Hib), un reactivo de látex y un antisuero para contrainmunoelectroforesis (CIE) e hinchazón capsular (HC), producidos en el Laboratorio de Infecciones Respiratorias Agudas Bacterianas del Instituto ®Pedro Kourí¼. Se evaluaron los reactivos en el serotipaje de 786 cepas de Haemophilus influenzae aisladas de procesos invasivos y de portadores nasofaríngeos, y en el diagnóstico rápido a partir de muestras de líquido cefalorraquídeo, suero y orina de 258 pacientes con diagnóstico clínico de meningoencefalitis. Se obtuvieron satisfactorios índices de sensibilidad y especificidad, los que fueron determinados utilizando como referencia el antisuero para Hib comercializado por la DIFCO para la técnica de HC y CIE, y el reactivo de látex Wellcogen para el diagnóstico de Hib. Se propone la validación de ambos reactivos por parte de los organismos nacionales acreditados, para que estos puedan ser introducidos en la red cubana de salud pública