Paracoccidioidomycosis manifested by sarcoidosis-like cutaneous lesions and caused by Paracoccidioides brasiliensis sensu stricto (S1a)

Abstract: Molecular studies have shown more than one species of the genus Paracoccidioides to be the causal agent of paracoccidioidomycosis. Efforts have been made to correlate the identified species with epidemiological and clinical data of patients, aiming to determine the real meaning and impact of new species. Bearing this objective in mind, the authors report a clinical case of paracoccidioidomycosis, from São Paulo state, Brazil, that manifested as uncommon sarcoid-like cutaneous lesions and was caused by Paracoccidioides brasiliensis sensu stricto (S1a). The patient was treated with itraconazole 200mg/day for 12 months, with complete clinical remission.

Paracoccidioidomicosis: crónica de una enfermedad olvidada / Paracoccidioidomycosis: chronicle of a neglected disease

La paracoccidioidomicosis (PCM) es una de las micosis sistémicas que son endémicas exclusivamente en América Latina. Está causada por hongos termodimorfos del género Paracoccidoides: P. brasiliensis (S1), P. americana (PS2), P. restrepiensis (PS3), P. venezuelensis (PS4) y P. lutzii. Paracoccidioides habita y produce conidios infecciosos en suelos de zonas subtropicales húmedas. En Argentina está presente al norte del paralelo 34.5° S. Poco se sabe sobre su nicho ecológico específico, lo que dificulta el diseño de medidas de control de la PCM. La infección ocurre en hospederos susceptibles cuando inhalan conidios aerosolizados. Los trabajadores rurales varones son el grupo más expuesto a contraer PCM. La primoinfección puede ser asintomática o causar un cuadro respiratorio leve; este, a su vez, puede autolimitarse o progresar a enfermedad, ya sea pulmonar o diseminada. Existen dos formas de presentación: (i) aguda/subaguda, en niños, adolescentes y personas con sistemas inmunes comprometidos; y (ii) crónica progresiva, en adultos. La cicatrización de las lesiones resulta en secuelas fibróticas que pueden causar disfagia, disfonía, insuficiencia suprarrenal y obstrucción intestinal. Aunque existen herramientas para su diagnóstico, la PCM es rara vez sospechada precozmente porque sus manifestaciones clínicas iniciales son inespecíficas. Sumados, el diagnóstico tardío y la baja adherencia a los efectivos pero largos tratamientos antimicóticos permiten el avance de la enfermedad y el desarrollo de fibrosis tisular extensa, lo que compromete gravemente la función respiratoria y suprarrenal, altera permanentemente la calidad de vida del paciente y puede causar su muerte.

Paracoccidioidomycosis (PCM) is among the systemic mycoses which are endemic only in Latin America. In Argentina, the vast majority of the cases are reported at north of latitude 34.5° S. The disease is produced by thermodimorphic fungi of the genus Paracoccidoides: P. brasiliensis (S1), P. americana (PS2), P. restrepiensis (PS3), P. venezuelensis (PS4) y P. lutzii. The natural habitat of members of this genus is the soil, where they produce infectious conidia. Little is known, however, about their specific ecologic niche(s), and this knowledge gap hampers the design of measures to control the infection. Rural male workers are the group most at risk of developing PCM. Infection occurs by inhalation of aerosolized conidia and may either be asymptomatic or cause mild respiratory symptoms. In turn, this primary infection may be self-limited or progress to severe pulmonary or disseminated disease. The disease has two clinical presentations: (i) acute or subacute (juvenile), frequent in children, adolescents and people with immunodeficiencies; and (ii) chronic progressive, in adults. Active lesions often resolve into fibrotic scars which can cause dysphagia, dysphonia, adrenal insufficiency, and intestinal obstruction. Although efficient tools are available for diagnosis and treatment, the nonspecific nature of PCM clinical manifestations frequently delay the diagnosis. In addition, the poor adherence to long antifungal treatments allows the advance of the disease and the development of extensive fibrosis compromising severely and permanently respiratory and adrenal functions, thus altering the patient´s quality of life and even causing his/her death.

Use of fluorescent oligonucleotide probes for differentiation between Paracoccidioides brasiliensis and Paracoccidioides lutzii in yeast and mycelial phase

BACKGROUND Fluorescence in situ hybridisation (FISH) associated with Tyramide Signal Amplification (TSA) using oligonucleotides labeled with non-radioactive fluorophores is a promising technique for detection and differentiation of fungal species in environmental or clinical samples, being suitable for microorganisms which are difficult or even impossible to culture. OBJECTIVE In this study, we aimed to standardise an in situ hybridisation technique for the differentiation between the pathogenic species Paracoccidioides brasiliensis and Paracoccidioides lutzii, by using species-specific DNA probes targeting the internal transcribed spacer-1 (ITS-1) of the rRNA gene. METHODS Yeast and mycelial phase of each Paracoccidioides species, were tested by two different detection/differentiation techniques: TSA-FISH for P. brasiliensis with HRP (Horseradish Peroxidase) linked to the probe 5’ end; and FISH for P. lutzii with the fluorophore TEXAS RED-X® also linked to the probe 5’ end. After testing different protocols, the optimised procedure for both techniques was accomplished without cross-positivity with other pathogenic fungi. FINDINGS The in silico and in vitro tests show no reaction with controls, like Candida and Cryptococcus (in silico) and Histoplasma capsulatum and Aspergillus spp. (in vitro). For both phases (mycelial and yeast) the in situ hybridisation showed dots of hybridisation, with no cross-reaction between them, with a lower signal for Texas Red probe than HRP-TSA probe. The dots of hybridisation was confirmed with genetic material marked with 4’,6-diamidino-2-phenylindole (DAPI), visualised in a different filter (WU) on fluorescent microscopic. MAIN CONCLUSION Our results indicated that TSA-FISH and/or FISH are suitable for in situ detection and differentiation of Paracoccidioides species. This approach has the potential for future application in clinical samples for the improvement of paracoccidioidomycosis patients prognosis.

Paracoccidioides brasiliensis AND Paracoccidioides lutzii, A SECRET LOVE AFFAIR / Paracoccidioides brasiliensis e Paracoccidioides lutzii, um caso secreto de amor

SUMMARYTo commemorate Prof. Carlos da Silva Lacaz's centennial anniversary, the authors have written a brief account of a few, out of hundreds, biological, ecological, molecular and phylogenetic studies that led to the arrival of Paracoccidioides lutzii, hidden for more than a century within Paracoccidioides brasiliensis. Lacaz's permanent interest in this fungus, and particularly his conviction on the benefits that research on paracoccidioidomycosis would bring to patients, were pivotal in the development of the field.

RESUMOPara comemorar o centenário de aniversário do Prof. Dr. Carlos da Silva Lacaz, os autores fazem um breve relato dos estudos sobre a biologia, ecologia e filogenia molecular que culminaram na revelação da espécie Paracoccidioides lutzii, que havia permanecido escondida por mais de um século ao lado de Paracoccidioides brasiliensis. O professor Lacaz exerceu papel central no desenvolvimento desta área do conhecimento, pois manteve interesse permanente nas pesquisas deste fungo e da paracoccidioidomicose, visando principalmente proporcionar benefícios aos pacientes acometidos por esta micose.

Cryptic species of Paracoccidioides brasiliensis: impact on paracoccidioidomycosis immunodiagnosis

We aimed to evaluate whether the occurrence of cryptic species of Paracoccidioides brasiliensis, S1, PS2, PS3 and Paracoccidioides lutzii, has implications in the immunodiagnosis of paracoccidioidomycosis (PCM). Small quantities of the antigen gp43 were found in culture filtrates of P. lutzii strains and this molecule appeared to be more variable within P. lutzii because the synonymous-nonsynonymous mutation rate was lower, indicating an evolutionary process different from that of the remaining genotypes. The production of gp43 also varied between isolates belonging to the same species, indicating that speciation events are important, but not sufficient to fully explain the diversity in the production of this antigen. The culture filtrate antigen AgEpm83, which was obtained from a PS3 isolate, showed large quantities of gp43 and reactivity by immunodiffusion assays, similar to the standard antigen (AgB-339) from an S1 isolate. Furthermore, AgEpm83 was capable of serologically differentiating five serum samples from patients from the Botucatu and Jundiaí regions. These patients had confirmed PCM but, were non-reactive to the standard antigen, thus demonstrating an alternative for serological diagnosis in regions in which S1 and PS2 occur. We also emphasise that it is not advisable to use a single antigen preparation to diagnose PCM, a disease that is caused by highly diverse pathogens.

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 percent 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.

Genetic characterization of morphologically variant strains of Paracoccidioides brasiliensis

Molecular characterization of Paracoccidioides brasiliensis variant strains that had been preserved under mineral oil for decades was carried out by random amplified polymorphic DNA analysis (RAPD). On P. brasiliensis variants in the transitional phase and strains with typical morphology, RAPD produced reproducible polymorphic amplification products that differentiated them. A dendrogram based on the generated RAPD patterns placed the 14 P. brasiliensis strains into five groups with similarity coefficients of 72 percent. A high correlation between the genotypic and phenotypic characteristics of the strains was observed. A 750 bp-RAPD fragment found only in the wild-type phenotype strains was cloned and sequenced. Genetic similarity analysis using BLASTx suggested that this RAPD marker represents a putative domain of a hypothetical flavin-binding monooxygenase (FMO)-like protein of Neurospora crassa.

Analysis of the genetic polymorphism of Paracoccidioides brasiliensis and Paracoccidioides cerebriformis "Moore" by random amplified polymorphic DNA (RAPD) and 28S ribosomal DNA sequencing: Paracoccidioides cerebriformis revisited

Nosso propósito foi comparar o polimorfismo genético de seis amostras de P. brasiliensis (113, 339, BAT, T1F1, T3B6, T5LN1), com quatro amostras de P. cerebriformis (735, 741, 750, 361) do laboratório de micologia do Instituto de Medicina Tropical de São Paulo, utilizando a técnica de Amplificação Aleatória do Polimorfismo de DNA (RAPD). O perfil de bandas do RAPD diferenciou claramente os isolados de P. brasiliensis de P. cerebriformis. Entretanto, ocorreu uma variação significativa no padrão de bandas das amostras de P. cerebriformis. O sequenciamento do gene ribossomal 28S revelou seqüências de nucleotídeos bastante conservadas entre os isolados de P. cerebriformis, fornecendo subsídio para o agrupamento taxonômico destas amostras, diferenciando estas de P. brasiliensis e mostrando que de fato são espécies distintas. A seqüência de DNA sugere que P. cerebriformis pertence ao gênero Aspergillus.

Tomografia computadorizada de torax de alta resolucao em paracoccidioidomicose (blastomicose sul-americana) / Resolution thoracic computed tomography for the diagnosis of south american blastomycosis

A blastomicose sul americana (BSB) e doenca causada pelo fungo Paracoccidioides brasiliensis.Ela acomete principalmente lavradores do sexo masculino, por via inalatoria,acometendo os pulmoes em mais de 80 por cento dos casos na forma cronica do adulto.Foram estudados 30 pacientes com DSA durante tes anos com radiogramas de torax e tomografia computadorizada de torax de alta resolucao(TCTAR) para correlacionar seus achados no sentido de auxiliar o diagnostico.O radiograma confirmou os achados mais comuns:infiltrados reticular e nodular,mais nos campos medios,alguns com consolidacao(19=63 por cento)hiperinsuflacao(20=66 pr cento)e bolhas subpleurais(10=33por cento).A TCTAR mostrou a historia natural da doenca: 1. Via inalatoria:com multiplas areas parenquimatosas semelhates a vidro fosco que podem progredir ou regredir(3=10 por cento) 2. Lesoes parenquimatosas progressivas com nodulos(25=83,3 por cento) consolidacoes irregulares(20=66 por cento, as vezes com insuflacao(22=73 por cento e bronquiectasias de tracao( 4=13 por cento)cavitacoes podem ocorrer por necrose(9=30 por cento) 3.Via linfatica com espessamento intersticial axial na zona medular com distorcao(25=83,3 por cento)4.Doenca progressiva com componente fibrotico e insuflacao(4=13 por cento).Conclusoes:As lesoes podem predominar em uma das duas regioes pulmonares.Temos:a padrao predominando lesoes parenquimatosas;padra predominando lesoes interticiais axiais;padrao predominando lesoes escavadas;e padrao combinado das lesoes

Paracoccidioidomycosis and AIDS: report of the first two colombian cases

Relato de dois primeiros casos de pacientes colombianos com AIDS e paracoccidioidomicose. Os pacientes, ambos masculinos, nao tinham conhecimento do fator de risco por HIV, embora tivessem no passado trabalhado no campo onde poderiam ter sido infectados por fungos. Eles tiveram o tipo juvenil da doenca em varios orgaos com sintomas de curta duracao. Eles estavam profundamente imunodeprimidos, com menos de 100 CD4 T linfocitos por mL; todavia, os testes sorologicos revelaram anticorpos circulantes anti-Paracoccidioides brasiliensis em um dos pacientes e os primeiros indicios diagnosticos vieram destes testes. Em um caso, a micose precedeu o diagnostico da AIDS enquanto que no outro, ambas patologias foram descobertas simultaneamente. A terapia antimicotica com itraconazole, foi dada por 10 meses, comecando com 200mg/dia e seguida por 100mg/dia; foi notada pronunciada melhora nos sintomas e sinais da micose, sem recorrencia...

Paracoccidioides cerebriformis Moore, 1935: mycologic and immunochemical study

O presente estudo trata dos resultados obtidos, do ponto de vista micologico e imunoquimico, de duas amostras de Paracoccidioides consideradas como pertencentes a espécie cerebriformis, criada por MOORE em 1935 e mantidas desde aquela época, através de repiques em agar-Sabouraud, na Micoteca do Instituto de Medicina Tropical de Säo Paulo. Após cerca de 60 anos, tais amostras conservavam as mesmas características descritas por MOORE (1935). Näo foram registradas lesöes experimentais em cobaias inoculadas por via intratesticular, näo se demonstrando também, pelas técnicas SDS PAGE e Western blotting...

Reactivity of anti-gp43 antibodies from Paracoccidioides brasiliensis antiserum with extracts from cutaneous lesions of Lobo's disease: preliminary note

Atraves de varias provas imunoquimicas foi demonstrada a presenca da gp43 em extratos de lesoes cutaneas da doenca de Jorge Lobo. A glicoproteina de 43 kDa e um dos antigenos dominantes do Paracoccidioides brasiliensis. permitindo a identificacao desta especie fungica

Cyclophosphamide effect on Paracoccidioidomycosis in the rat

La paracoccidioidomicosis es una enfermedad endemica fungica ampliamente distribuida en Latino-America. La ciclofosfamida ha sido usada como potente imunosupresor para modular la respuesta inmune, en um modelo experimental infectado con Paracocidioides brasiliensis. Ratas macho buffalo/sim endocriadas de 250-300 gr. de peso, fueron inoculadas por via intracardiaca con 5.10 elevado a sexta potencia celalas de P. brasiliensis en fase levaduriforme...

Paracoccidioides brasiliensis: a mycologic and immunochemical study of a sample isolated from a armadillo (Dasipus novencinctus)

Amostra de Paracoccidioides brasiliensis isolada de visceras (baco e figado) de um tatu (Dasipus novencintus)foi estudada no ponto de vista micologico e imunoquimico. O tatu havia sido capturado em area da usina hidroeletrica de Tucurui (Estado do Para). Este ja havia sido considerado como reservatorio enzootico do Paracoccidioides brasiliensis naquela regiao. Esta amostra, conservada na Micoteca do Instituto de Medicina Tropical de Sao Paulo sob o numero 135, apresenta todas as caracteristicas de Paracoccidioides brasiliensis, com elevado poder antigenico e baixa virulencia para cobaios e ratos Wistar. A demonstracao do exo-antigeno especifico do P. brasiliensis, representado pela glicoproteina de peso molecular 43 KDa, foi evidente atraves das tecnicas de Imunodifusao Dupla, Imunoeletroforese, SDS-PAGE e Imunoblotting.