Multiplex polymerase chain reaction as an improved method for screening Histoplasma capsulatum mating types

Histoplasmosis is a systemic mycosis infection caused by Histoplasma capsulatum, a heterothallic ascomycete. The sexual reproduction of this fungus is regulated by the mating type (MAT1) locus that contains MAT1-1 and MAT1-2 idiomorphs, which were identified by uniplex polymerase chain reaction (PCR). This study aimed to optimise single-step multiplex PCR for the accurate detection of the distinct mating types of H. capsulatum. Among the 26 isolates tested, 20 had MAT1-1 genotype, while six showed MAT1-2 genotype, in agreement with the uniplex PCR results. These results suggest that multiplex PCR is a fast and specific tool for screening H. capsulatum mating types.

Brote de histoplasmosis en la Escuela de Cadetes de la Base Aérea de Morón, Provincia de Buenos Aires, República Argentina / Histoplasmosis outbreak in Morón, Buenos Aires Province, Argentina

Se describe un brote de histoplasmosis que afectó a 6 cadetes de la Fuerza Aérea Argentina, sin antecedentes patológicos previos. Todos consultaron por problemas respiratorios después de haber limpiado un hangar. En ese recinto se encontraron abundantes deyecciones de animales, presuntamente de palomas y murciélagos. Los pacientes sufrieron fiebre, mialgias, taquipnea y tos no productiva. Las radiografías y tomografías de tórax mostraron imágenes pulmonares micronodulares, engrosamiento de los tabiques interalveolares y adenopatías hiliares. Todos tuvieron una evolución favorable y no requirieron tratamiento antifúngico. Las pruebas de inmunodifusión y contrainmunoelectroforesis con antígenos de Histoplasma capsulatum fueron positivas, al igual que las intradermorreacciones con histoplasmina. Se recogieron 5 muestras de tierra del lugar, las que fueron inoculadas por vía intraperitoneal a 20 hámsteres. De los cultivos de hígado y bazo de dichos animales se consiguió aislar la fase micelial de H. capsulatum. La cepa aislada se comparó con las obtenidas de 12 pacientes argentinos utilizando perfiles genéticos y se observó un clado único con más de 96% de similitud, lo que confirma la homogeneidad de las cepas argentinas. Si bien la histoplasmosis es endémica en la Pampa húmeda, este es el primer brote totalmente documentado al sur del paralelo 34°.

An histoplasmosis outbreak affecting 6 previously healthy Air Force cadets is herein presented. The patients suffered from fever and respiratory symptoms after having cleaned an abandoned hangar soiled with pigeons and bat droppings. They all presented fever, myalgia, tachypnea, and nonproductive cough. Chest X-ray and CT scan studies showed disseminated reticulonodular images affecting both lungs. Hiliar adenomegalies were also observed. All patients achieved a favourable outcome without antifungal treatment. Both serologic tests searching for specificic antibodies (immunodiffusion and counterimmunoelectrophoresis) and histoplasmin skin tests were positive in all cases. Five soil samples mixed with pigeons and bat droppings were collected from the hangar. Suspensions of these samples were inoculated into 20 hamsters by intraperitoneal injection; mycelial phase of H. capsulatum was isolated from liver and spleen cultures. The genetic profile of this strain was compared with 12 isolates obtained from Argentinean patients, and a great degree of homogeneity was observed (> 96% similarity). Although histoplasmosis is endemic in the wet Pampas, this is the first epidemic outbreak reported south of the 34th parallel.

Identification of fungemia agents using the polymerase chain reaction and restriction fragment length polymorphism analysis

Prompt and specific identification of fungemia agents is important in order to define clinical treatment. However, in most cases conventional culture identification can be considered to be time-consuming and not without errors. The aim of the present study was to identify the following fungemia agents: Candida albicans, Candida parapsilosis, Candida tropicalis, Candida glabrata, Cryptococcus neoformans, Cryptococcus gattii, and Histoplasma capsulatum using the polymerase chain reaction and restriction fragment length polymorphism analysis (PCR/RFLP). More specifically: a) to evaluate 3 different amplification regions, b) to investigate 3 different restriction enzymes, and c) to use the best PCR/RFLP procedure to indentify 60 fungemia agents from a culture collection. All 3 pairs of primers (ITS1/ITS4, NL4/ITS5 and Primer1/Primer2) were able to amplify DNA from the reference strains. However, the size of these PCR products did not permit the identification of all the species studied. Three restriction enzymes were used to digest the PCR products: HaeIII, Ddel and Bfal. Among the combinations of pairs of primers and restriction enzymes, only one (primer pair NL4/ITS5 and restriction enzyme Ddel) produced a specific RFLP pattern for each microorganism studied. Sixty cultures of fungemia agents (selected from the culture collection of Fundação de Medicina Tropical do Amazonas - FMTAM) were correctly identified by PCR/RFLP using the prime pair NL4/ITS5 and Ddel. We conclude that the method proved to be both simple and reproducible, and may offer potential advantages over phenotyping methods.

Histoplasma capsulatum fungemia in patients with acquired immunodeficiency syndrome: detection by lysis-centrifugation blood-culturing technique

Progressive disseminated histoplasmosis (PDH) is an increasingly common cause of infection in patients with acquired immune deficiency syndrome (AIDS). We report 21 cases of PDH associated with AIDS diagnosed by lysis-centrifugation blood culture method. The most prevalent clinical findings were fever, weight loss, respiratory symptoms, and mucocutaneous lesions. Chest roentgenogram showed diffuse pulmonary infiltrates in 13 of 21 patients (62 percent). Brochoalveolar fluid has yelded positive culture in four patients only in medium with cycloheximide.

Histoplasmose progressiva disseminada (HPD) tem aumentado e é causa comum de infecção em pacientes com síndrome da imunodeficiência adquirida (Aids). Relatamos 21 casos de HPD associado com Aids diagnosticada pela técnica de hemocultivo por lise-centrifugação. Os achados clínicos mais prevalentes foram febre, perda de peso, sintomas respiratórios e lesões mucocutâneas. Raios X de tórax mostrou infiltrados pulmonares difusos em 13 dos 21 pacientes (62 por cento). Amostras de lavado broncoalveolar foram positivos em apenas 4 pacientes através de meio com cicloheximida.

Disseminated histoplamosis.

A case of disseminated histoplasmosis in a 45-year-old male patient with acquired immunodeficiency syndrome (AIDS) from Pune is reported. The patient presented with high-grade fever and pain in hypochondrium. Clinical signs were pallor and hepatosplenomegaly. Bone marrow and splenic aspirate revealed numerous intracellular oval shaped yeast forms. Histoplasma capsulatum was isolated from the bone marrow and splenic aspirate. H. capsulatum infection is an opportunistic infection usually reported from patient with AIDS in areas endemic for H. capsulatum. The present case highlights the fact that histoplasmosis could be an emerging opportunistic infection in India.

Study on Histoplasmosis (Epizootic Lymphangitis) in Cart-Horses in Ethiopia

A cross-sectional study was conducted to determine the prevalence of Histoplasma farciminosum in 2907 carthorses using clinical and microbiological examinations at three towns (Debre Zeit, Mojo and Nazareth), Central Ethiopia, between December 1999 and January 2001. An overall prevalence of 26.2% (762/2907) was recorded; the highest prevalence (39.1%) being recorded at Mojo whereas the lowest (21.1%) was recorded at Nazareth. The difference in prevalence among the three towns was highly significant (chi2 = 76.92, P<0.0001). Carthorses found at Mojo [OR =2.4, CI=(1.9-3.0), P<0.0001] and Debre Zeit [OR=1.9, CI=(1.5-2.3), P<0.0001] were at higher risk of infection than those found at Nazareth. The mycelial and yeast forms of the Histoplasma capsulatum variety farciminosum were isolated on the Sabouraud's dextrose agar. The results of the present study showed the rampant occurrence of histoplasmosis farciminosi at the three towns and indicates the need for further nationwide investigation into the disease to design sound control strategy.

Histoplasmosis presenting as acute respiratory distress syndrome after exposure to bat feces in a home basement

This report describes the clinical and epidemiological findings of an outbreak of histoplasmosis in Brazil involving four persons of the same family, one adult and three children. Contamination by fecal material of bats in the basement of the house was found to be the most likely cause of this outbreak. None of the patients had evidence of immune deficiency. The adult died of acute respiratory distress syndrome. A more severe presentation of the disease in this patient may be partly explained by a heavier exposure to the feces of bats.

Genotipificación de cepas de Histoplasma Capsulatum aisladas de pacientes con histoplasmosis asociada al SIDA, mediante el polimorfismo en la longitud de los fragmentos de restricción / Genotypic characterization of Histoplasma capsulatum isolates from AIDS associated-histoplasmosis-patients by using restriction fragment langth polymorphysms (RFLP)

Introducción: La histoplasmosis es una de las más importantes micosis asociada al síndrome de inmunodeficiencia adquirida (SIDA) en México. En este trabajo se efectuó la caracterización genotípica por el polimorfismo en la longitud de los fragmentos de restricción (RFLP) del DNA total de 31 cepas de Histoplasma capsulatum de pacientes con histoplasmosis asociada a SIDA, y su comparación con cepas de referencia de México, Panamá y Estados Unidos de America (EUA). Material y métodos: Las cepas fueron plenamente identificadas como H. capsulatum mediante su morfología colonial y microscópica, conversión de micelio a levadura y repoducción de exoantígenos probados por inmunodifusión con sueros específicos. El DNA total de las cepas fue obtenido a partir de micelio a levadura y producción de exoantígenos probados por inmunodifusión con sueros específicos. El DNA total de las cepas fue obtenido a partir de micelio macerado con nitrógeno líquido e incubado en presencia de proteinasa K y dodecilsulfato de sodio. El DNA fue diferido con la enzima HaeIII y los productos separados por electroforesis en agarosa y teñidos con bromuro de etidio. Utilizando el programa SPSS/PC+ se construyó un dendrograma a fin de establecer las relaciones polimórficas entre las cepas. Resultados y discusión: Las cepas formaron 10 grupos según sus perfiles de RFLP, obsevándose dos perfiles principales A y C con 33.3 y 29.1 por ciento de las cepas, respectivamente. Los perfiles restantes estuvieron representados por cepas únicas, a excepción del perfil J con dos cepas que mostraron un patrón similar al de la cepa Downs (clase 1, de EUA), No se encontró ningún perfil semejante a los de las cepas prevalentes en EUA o Panamá. El dendrograma mostró una relación de 85 por ciento entre las cepas de los perfiles de mayor prevalencia (A y C) asimismo del perfil J, además de las cepas Downs y FLs1 de los EUA. Los demás aislados con perfiles únicos se relacionaron en un 75 por ciento con las otras cepas

Organización de la colección de cepas de Histoplasma capsulatum del Laboratorio de Inmunología de Hongos, Facultad de Medicina, UNAM / Organization of the Histoplasma capsulatum strains collection from the laboratorio de Inmunologia de Hongos, Facultad de Medicina, UNAM

Objetivo: El presente trabajo muestra la organización, los lineamientos y las particularidades de la colección de cepas de Histoplasma capsulatum del laboratorio de Inmunología de Hongos del Departamento de Microbiología y parasitología, de la Facultad de medicina, UNAM. Características de la colección: Está formada por primoaislamientos del hongo a partir de diversas fuentes y distintas procedencias geográficas dentro de la República Mexicana, además de cepas de la república Mexicana, además de cepas de referencia de otros países, características que destacan el aspecto especial y selecto de esta colección