Variable gp43 secretion by Paracoccidioides brasiliensis clones obtained by two different culture methods

The main objectives of this study were to obtain clones of Paracoccidioides brasiliensis by two methods (micromanipulation and plating assay) and to determine if the secretion of the 43-kDa glycoprotein (gp43) is dependent on the clonal culture. the results show that the secretion of gp43 is not dependent on clonal cultures. Clones that originally were secretors of this molecule, after subculturing, lost this characteristic; on the other hand, clones that originally did not secrete gp43 began to secrete gp43 after subculturing.Universidade Federal de São Paulo, Disciplina Biol Celular, Dept Microbiol Immunol & Parasitol, BR-04023062 São Paulo, BrazilUniversidade Federal de São Paulo, Disciplina Biol Celular, Dept Microbiol Immunol & Parasitol, BR-04023062 São Paulo, BrazilWeb of Scienc

Serology of Paracoccidioidomycosis Due to Paracoccidioides lutzii

Paracoccidioides lutzii is a new agent of paracoccidioidomycosis (PCM) and has its epicenter localized to the Central-West region of Brazil. Serological diagnosis of PCM caused by P. lutzii has not been established. This study aimed to develop new antigenic preparations from P. lutzii and to apply them in serological techniques to improve the diagnosis of PCM due to P. lutzii. Paracoccidioides lutzii exoantigens, cell free antigen (CFA), and a TCA-precipitated antigen were evaluated in immunodiffusion (ID) tests using a total of 89 patient sera from the Central-West region of Brazil. Seventy-two sera were defined as reactive for P. brasiliensis using traditional antigens (AgPbB339 and gp43). Non-reactive sera for traditional antigens (n = 17) were tested with different P. lutzii preparations and P. lutzii CFA showed 100% reactivity. ELISA was found to be a very useful test to titer anti-P. lutzii antibodies using P. lutzii-CFA preparations. Sera from patients with PCM due to P. lutzii presented with higher antibody titers than PCM due to P. brasiliensis and heterologous sera. in western blot, sera from patients with PCM due to P. lutzii were able to recognize antigenic molecules from the P. lutzii-CFA antigen, but sera from patients with PCM due to P. brasiliensis could not recognize any P. lutzii molecules. Due to the facility of preparing P. lutzii CFA antigens we recommend its use in immunodiffusion tests for the diagnosis of PCM due to P. lutzii. ELISA and western blot can be used as complementary tests.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Universidade Federal de São Paulo UNIFESP, Dept Microbiol Imunol & Parasitol, Disciplina Biol Celular, São Paulo, BrazilUniv Fed Mato Grosso do Sul UFMS, Fac Med FAMED, Campo Grande, MS, BrazilUniv Fed Mato Grosso UFMT, Nucleo Doencas Infecciosas & Trop, Cuiaba, Mato Grosso, BrazilUniversidade Federal de São Paulo UNIFESP, Dept Microbiol Imunol & Parasitol, Disciplina Biol Celular, São Paulo, BrazilFAPESP: 2012/06593-0FAPESP: 2009/54024-2Web of Scienc

Paracoccidioidomycosis in Amerindian populations of the Brazilian Suruí tribe: a clinical and laboratory study of two cases

Paracoccidioidomycosis has been considered the most frequent endemic systemic mycosis in Latin America. Although most cases of paracoccidioidomycosis involve rural workers, this systemic fungal disease has been scarcely reported among Amerindian populations from Brazil. We report two cases of paracoccidioidomycosis in Tupi-Mondé Amerindians from Cacoal, state of Rondônia, Brazil. Both cases exhibited positive serological results by a specific immunodiffusion test only when the assay was performed with antigens obtained from the mycelial form of P. brasiliensis. The authors present a literature review of paracoccidioidomycosis in Brazilian Amerindians and discuss the need for further investigations about the impact of the antigenic diversity of P. brasiliensis from different geographic areas on the serological diagnosis of PCM.Paracoccidioidomicose é considerada a micose sistêmica endêmica mais prevalente na América Latina. Apesar da maior parte da casuística de paracoccidioidomicose ocorrer entre trabalhadores rurais, há poucos casos documentados de ocorrência dessa micose entre índios brasileiros. São apresentados 2 casos de paracoccidioidomicose em índios Suruí, família linguística Tupi-Mondé, procedentes de Cacoal, Rondônia. Ambos apresentaram sorologia positiva à imunodifusão apenas com antígenos da fase miceliana do P. brasiliensis. Os autores apresentam revisão de literatura sobre a ocorrência dessa micose entre índios brasileiros e discutem a necessidade de futuras investigações buscando caracterizar as diferenças regionais de cepas de P. brasiliensis e seu impacto no diagnóstico sorológico dessa micose.Universidade Federal de São Paulo (UNIFESP)UNIFESPSciEL

Immunodiagnosis of Paracoccidioidomycosis due to Paracoccidioides brasiliensis Using a Latex Test: Detection of Specific Antibody Anti-gp43 and Specific Antigen gp43

BackgroundParacoccidioidomycosis (PCM) is a life-threatening systemic disease and is a neglected public health problem in many endemic regions of Latin America. Though several diagnostic methods are available, almost all of them present with some limitations.Method/Principle FindingsA latex immunoassay using sensitized latex particles (SLPs) with gp43 antigen, the immunodominant antigen of Paracoccidioides brasiliensis, or the monoclonal antibody mAb17c (anti-gp43) was evaluated for antibody or antigen detection in sera, cerebrospinal fluid (CSF), and bronchoalveolar lavage (BAL) from patients with PCM due to P. brasiliensis. the gp43-SLPs performed optimally to detect specific antibodies with high levels of sensitivity (98.46%, 95% CI 91.7-100.0), specificity (93.94%, 95% CI 87.3-97.7), and positive (91.4%) and negative (98.9%) predictive values. in addition, we propose the use of mAb17c-SLPs to detect circulating gp43, which would be particularly important in patients with immune deficiencies who fail to produce normal levels of immunoglobulins, achieving good levels of sensitivity (96.92%, 95% CI 89.3-99.6), specificity (88.89%, 95% CI 81.0-94.3), and positive (85.1%) and negative (97.8%) predictive values. Very good agreement between latex tests and double immune diffusion was observed for gp43-SLPs (k = 0.924) and mAb17c-SLPs (k = 0.850), which reinforces the usefulness of our tests for the rapid diagnosis of PCM in less than 10 minutes. Minor cross-reactivity occurred with sera from patients with other fungal infections. We successfully detected antigens and antibodies from CSF and BAL samples. in addition, the latex test was useful for monitoring PCM patients receiving therapy.Conclusions/SignificanceThe high diagnostic accuracy, low cost, reduced assay time, and simplicity of this new latex test offer the potential to be commercialized and makes it an attractive diagnostic assay for use not only in clinics and medical mycology laboratories, but mainly in remote locations with limited laboratory infrastructure and/or minimally trained community health workers.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Universidade Federal de São Paulo, Dept Microbiol Immunol & Parasitol, Div Cell Biol, São Paulo, BrazilEvandro Chagas Inst, Bacteriol & Mycol Div, Ananindeua, Para, BrazilUniv Fed Alfenas, Inst Biomed Sci, Alfenas, MG, BrazilUniversidade Federal de São Paulo, Dept Microbiol Immunol & Parasitol, Div Cell Biol, São Paulo, BrazilFAPESP: FAPESP 2011/07350-1FAPESP: 2011/01628-8FAPESP: 2009/54181-0FAPESP: 2009/54024-2Web of Scienc

Simultaneous infection of human host with genetically distinct isolates of Paracoccidioides brasiliensis

This study is the first report on genetic differences between isolates of Paracoccidioides brasiliensis from a single patient. We describe a simultaneous infection with genetically distinct isolates of P. brasiliensis in a patient with chronic paracoccidioidomycosis. The clinical isolates were obtained from lesions in different anatomical sites and were characterised by random amplified polymorphic DNA (RAPD) analysis. The RAPD technique can be helpful for distinguishing between clinical isolates. Different random primers were used to characterise these clinical isolates. The RAPD patterns allowed for differentiation between isolates and the construction of a phenetic tree, which showed more than 28% genetic variability in this fungal species, opening new possibilities for clinical studies of P. brasiliensis. Based on these results and preliminary clinical findings, we suggest that different genotypes of P. brasiliensis might infect the same patient, inducing the active form of the disease.Universidade Federal de Mato Grosso Faculdade de Ciências Médicas Laboratório de MicologiaUniversidade Federal de São Paulo (UNIFESP) Departamento de Microbiologia, Imunologia e ParasitologiaUniversidade de Cuiabá Hospital Geral Universitário Laboratório de MicologiaUNIFESP, Depto. de Microbiologia, Imunologia e ParasitologiaSciEL

Research regarding anti-PGL-I antibodies by ELISA in wild armadillos from Brazil

Armadillos have been involved in leprosy transmission and are considered a source of Mycobacterium leprae in numerous reports. Clinicians from certain areas of the USA consider contact with armadillos a risk factor for leprosy. However, there is a challenge associated with the role of wild armadillos perpetuating human leprosy in the American Continent. The presence of anti-PGL-I antibodies was investigated in wild nine-banded armadillos from leprosy-endemic areas in State of Espirito Santo, Brazil, by ELISA performed on serum samples from 47 armadillos. Positive ELISA was obtained from 5 (10.6%) armadillos. Infected armadillos may play some role in leprosy transmission, disseminating bacilli in the environment, perhaps making it more difficult to interrupt transmission and reduce the number of new leprosy cases. ELISA is an efficient tool for seroepidemiological investigations of Mycobacterium leprae in armadillos.Tatus têm sido envolvidos na transmissão da hanseníase e considerados como fonte de Mycobacterium leprae em muitas publicações. Médicos de partes dos EUA consideram o contato com tatus um fator de risco para hanseníase. Entretanto, há um desafio associado ao papel do tatu na perpetuação da hanseníase no Continente Americano. Foi pesquisada a presença de anticorpos anti-PGL-I em tatus selvagens de áreas endêmicas em hanseníase do Estado do Espírito Santo, Brasil, através de ELISA realizado em amostras de soro de 47 animais. Elisa positivo foi encontrado em 5 (10.6%) tatus. Tatus infectados podem ter algum papel na transmissão da hanseníase disseminando bacilos no meio ambiente, talvez tornando mais difícil a interrupção da cadeia de transmissão e redução do número de casos novos de hanseníase. A técnica de ELISA é um eficiente método para investigação soroepidemiológica da presença do Mycobacterium leprae em tatus.Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Universidade Federal do Espírito Santo Departmento de Medicina SocialLondon School of Hygiene & Tropical Medicine Departmento de Infectologia e Doenças TropicaisEscola de Medicina da Santa Casa de Misericórdia Departmento de PatologiaRoyal Tropical InstituteUniversidade Federal de Goiás Instituto de Patologia Tropical e Saúde PúblicaUniversidade Federal de São Paulo (UNIFESP) Departmento de Biologia CelularInstituto Lauro de Souza LimaUniversidade Federal de São Paulo (UNIFESP) Departmento de DermatologiaUNIFESP, Departmento de Biologia CelularUNIFESP, Departmento de DermatologiaSciEL

Rapid diagnosis of coccidioidomycosis by nested PCR assay of sputum

Coccidioidomycosis is a deep infection caused by two dimorphic fungi, Coccidioides immitis and Coccidioides posadasii. Diagnosis of the disease requires culture of suspicious clinical samples on mycological media. However, as these species are virulent pathogens, handling of their cultures is a high-risk activity, and is limited to Biosafety Level 3 laboratories. This study describes the direct detection of C. posadasii DNA in an inappropriate sputum sample by PCR amplification of the highly specific Ag2/PRA antigen gene. the results obtained suggest that direct detection of the Ag2/PRA sequence in sputum is an excellent method for rapid and specific diagnosis of coccidioidomycosis.Fed Univ Ceara, Med Mycol Specialized Ctr, Fortaleza, Ceara, BrazilFed Univ Ceara, Postgrad Program Med Sci, Fortaleza, Ceara, BrazilState Univ Ceara, Postgrad Program Vet Sci, Fortaleza, Ceara, BrazilState Univ Ceara, Dept Sci Biol, Fortaleza, Ceara, BrazilUniversidade Federal de São Paulo, Dept Microbiol Immunol & Parasitol, São Paulo, BrazilUniversidade Federal de São Paulo, Dept Microbiol Immunol & Parasitol, São Paulo, BrazilWeb of Scienc