Epidemiological and TNFα polymorphism evaluation in patients with cryptococcal meningitis treated at a referral hospital in North Brazil

Abstract INTRODUCTION: The present study evaluated the epidemiology of cryptococcal meningitis and TNFα gene polymorphisms in patients at a reference hospital in northern Brazil. METHODS: Samples from 25 patients infected with Cryptococcus spp. were collected to confirm the infection and to analyze the TNFα gene polymorphisms. RESULTS: Cryptococcus neoformans was detected as the predominant etiological agent (100%) in HIV-positive patients. No genetic polymorphic changes were found. CONCLUSIONS: No correlation was observed between the analyzed TNFα polymorphisms and cryptococcal meningitis.

Cryptococcal meningitis epidemiology: 17 years of experience in a State of the Brazilian Pantanal

Abstract INTRODUCTION: This study aimed to describe cryptococcal meningitis (CM) cases and the associated demographic, clinical, and microbiological data obtained from cities in the State of Mato Grosso do Sul in the Midwestern region of Brazil. METHODS: The data from 129 patients with laboratory-confirmed CM admitted from 1997 to 2014 were retrospectively reviewed. The molecular types of Cryptococcus neoformans and Cryptococcus gattii isolated from cerebrospinal fluid were analyzed to determine their geographic distribution. RESULTS: The patients had a mean age of 37 years and consisted mostly of men (76.7%). Most of the Cryptococcus isolates were obtained from patients infected with human immunodeficiency virus (HIV) and included 105 (87.5%) and 5 (55.6%) isolates of C. neoformans and C. gattii complexes, respectively. A restriction fragment length polymorphism (RFLP) analysis of URA5 revealed that most of the isolates were C. neoformans molecular type VNI (89.1%), whereas the molecular types VGII (7%) and VNII (3.9%) were observed less frequently. Notably, 65% of the cases with a time from symptom onset to laboratory diagnosis of more than 60 days resulted in fatalities, and sequelae were observed among the patients who survived. CONCLUSIONS: The present study documents the occurrence of neurocryptococcosis, which is mainly caused by C. neoformans VNI, in Mato Grosso do Sul, Brazil, with probable autochthonous cases in the Brazilian Pantanal, the world's largest tropical wetland, and a biome where cryptococcosis has not yet been explored.

Caracterización molecular de los aislamientos de Cryptococcus neoformans de pacientes con HIV, Guayaquil, Ecuador / Molecular characterization of Cryptococcus neoformans isolates from HIV patients, Guayaquil, Ecuador

Resumen Introducción. La neurocriptococosis es una infección fúngica oportunista que representa un alto costo en vidas humanas y para la economía de los países. Sus agentes causales, las especies del complejo Cryptococcus neoformans/Cryptococcus gattii, tienen una fase sexuada y otra asexuada, cuatro serotipos principales y siete variedades moleculares con diferencias clínico-epidemiológicas, fenotípicas y de sensibilidad a los antifúngicos. Objetivo. Caracterizar molecularmente los aislamientos clínicos de C. neoformans de Guayaquil, Ecuador. Materiales y métodos. Se determinó el tipo de apareamiento, el serotipo y la variedad molecular mediante reacción en cadena de la polimerasa y análisis del polimorfismo de los fragmentos de restricción de 27 aislamientos levaduriformes previamente identificados como C. neoformans mediante métodos convencionales. Los aislamientos fueron recuperados del líquido cefalorraquídeo de pacientes con síndrome neurológico seropositivos para HIV, internados en el Hospital de Infectología "Dr. José Daniel Rodríguez Maridueña", entre diciembre de 2013 y enero de 2015. Resultados. Se demostró el amplio predominio de C. neoformans del serotipo A, MATα y el genotipo VNI entre los aislamientos estudiados. Conclusiones. Estos datos son similares a los obtenidos en otros países y son los primeros de su tipo en Guayaquil, Ecuador, por lo cual constituyen un aporte importante al conocimiento de la criptococosisen esta ciudad.

Abstract Introduction: Neurocryptococcosis is an opportunistic fungal infection that represents a high cost in human lives and for the economy of countries. Its causative agent, the Cryptococcus neoformans/Cryptococcus gattiispecies complex, has a sexual and an asexual phase, four major serotypes and seven molecular varieties with phenotypic, clinical-epidemiological and antifungal susceptibility differences. Objective: To characterize by molecular methods clinicalisolates of C. neoformans from Guayaquil, Ecuador. Materials and methods: We determined mating types, serotypes and molecular varieties by PCR and RFLP in 27 yeast isolates previously identified as C. neoformans by conventional methods. The isolates were recovered from cerebrospinal fluid of HIV seropositive patients with neurological syndrome admitted at "Dr. José Daniel Rodríguez Maridueña" Hospital from December, 2013, to January, 2015. Results: We established a wide prevalence of C. neoformans serotype A, MATαand genotype VNI among the studied isolates. Conclusions: These data are similar to those obtained in other countries and the first identified by molecular characterization in Guayaquil, Ecuador. Therefore, they constitute an important contribution to the knowledge on cryptococcosis in this country.

Molecular diagnosis of cryptococcal meningitis in cerebrospinal fluid: comparison of primer sets for Cryptococcus neoformans and Cryptococcus gattii species complex

Aim: This study evaluated the use of polymerase chain reaction for cryptococcal meningitis diagnosis in clinical samples. Materials and methods: The sensitivity and specificity of the methodology were evaluated using eight Cryptococcus neoformans/C. gattii species complex reference strains and 165 cere- brospinal fluid samples from patients with neurological diseases divided into two groups: 96 patients with cryptococcal meningitis and AIDS; and 69 patients with other neurological opportunistic diseases (CRL/AIDS). Two primer sets were tested (CN4-CN5 and the multiplex CNa70S-CNa70A/CNb49S-CNb-49A that amplify a specific product for C. neoformans and another for C. gattii). Results: CN4-CN5 primer set was positive in all Cryptococcus standard strains and in 94.8% in DNA samples from cryptococcal meningitis and AIDS group. With the multiplex, no 448-bp product of C. gattii was observed in the clinical samples of either group. The 695 bp products of C. neoformans were observed only in 64.6% of the cryptococcal meningitis and AIDS group. This primer set was negative for two standard strains. The specificity based on the negative samples from the CTL/AIDS group was 98.5% in both primer sets. Conclusions: These data suggest that the CN4/CN5 primer set was highly sensitive for the identification of C. neoformans/C. gattii species complex in cerebrospinal fluid samples from patients with clinical suspicion of cryptococcal meningitis. .

Criptococosis meníngea asociada al SIDA. Análisis de los pacientes varones HIV (+) con criptococosis meníngea internados en la Sala 11 del Hospital Francisco J Muñiz / Meningeal cryptococcosis associated with AIDS. Analysis of male HIV (+) patients with internal meningeal cryptococcosis in Room 11 of Francisco J Muñiz Hospital

La criptococosis es una micosis sistémica oportunista con distribución universal causada por una levadura capsulada. El Cryptococcus neoformans (C. neoformans). C. neoformans infecta al hombre y a animales susceptibles por vía inhalatoria, provocando en huéspedes inmunocompetentes una primoinfección asintomática. Crece preferentemente en ambientes ricos con componentes nitrogenados, principalmente a partir de las excretas desecadas de las palomas que son ricas en nitrógeno y creatinina, con alto contenido en sales y pH alcalino, cualidades que facilitan la supervivencia del hongo. La diseminación hematógena del agente causal provoca múltiples localizaciones, las cuales se hacen clínicamente evidentes en pacientes con deterioro de la inmunidad mediada por células. A partir de la eclosión del SIDA se produjo un aumento significativo del número de casos, transformándose esta última condición en la causa predisponente más importante de esta micosis. La mayoría de los pacientes con SIDA y criptococosis del SNC presentan signos y síntomas de meningitis o meningoencefalitis subagudas como cefalea, fiebre, parálisis de nervios craneales, letargo, coma o amnesia de varias semanas de evolución. La anfotericina B es el pilar terapéutico del tratamiento. En nuestro estudio analizamos las características demográficas, epidemiológicas, clínicas, el líquido cefalorraquídeo, el diagnóstico micológico y el tratamiento empleado en los pacientes hospitalizados en Sala 11 con diagnóstico de criptococosis meníngea. En la criptococosis meníngea asociada al SIDA, el diagnóstico puede realizarse por el análisis de distintos tipos de especímenes biológicos, siendo los principales el examen microscópico directo del LCR con tinción de tinta china y el cultivo.

Cryptococcosis is a fungal opportunistic infection systemic who has universal distribution, caused by capsulated yeast, Cryptococcus neoformans. C. neoformans infects humans and susceptible animals by inhalation, causing an asymptomatic primary infection in immunocompetent hosts. It grows preterentially in rich environments with nitrogen components, mainly from dried excreta of pigeons that are rich in nitrogen and creatinine, with a high salt content and alkaline pH, qualities which facilitate the survival of the fungus. Haematogenous dissemination of the causative agent leads to multiple locations, which are clinically evident in patients with impairment of cell-mediated immunity. Since the emergence of AIDS, it was a significant in crease in the number of cases, who becomes in the most important predisposing cause of this mycosis. Most patients with AIDS and CNS cryptococcosis have signs and symptoms of subacute meningitis or meningoencephalitis as headache, fever, cranial nerve palsy, lethargy, coma or amnesia of several weeks of evolution. Amphotericin B therapy is the mainstay of treatment. We analyzed the demographic, epidemiological, clinical, cerebrospinal fluid, mycological diagnosis and treatment used for hospitalized patients in the unit 11 of the Infectious Disease Hospital Muñiz with the diagnosis of meningeal cryptococcosis. In meningeal cryptococcosis associated with AIDS, the diagnosis can be performed by the analysis of different types of biological specimens, mainly direct microscopic examination of CSF with India ink staining and culture.

Síndrome de restauración inmune asociado a tratamiento antirretroviral y criptococosis meníngea: Caso clínico / Inflammatory reconstitution immune syndrome associated to antiretroviral therapy and meningeal cryptococcosis

The objective of high activity antiretroviral therapy (HAART) in patients with AIDS, is to obtain immune restoration. This means a reduction of the viral load and restitution of the CD4 cell count. A decreased rate of HIV replication improves both the number and function of CD4 cells. Nevertheless, this treatment sometimes results in the reappearance of previous symptoms from treated conditions due to opportunistic infections (ie: tuberculosis, criptococcosis, hepatitis, Pneumocystis jirovesi, toxoplasmosis, etc) or non infectious condition such as sarcoidosis, Graves disease or Kaposi sarcoma. This is known as Inflammatory Reconstitution Immune Syndrome (IRIS). We report a 37 year-old woman in stage C3-AIDS with a previous criptococcal meningitis. She was treated, achieving a marked improvement with treatment and subsequent suppressive therapy with fluconazole 200 mg/day. IRIS appeared after 8 months of ongoing antiretroviral therapy with immune restoration with the development of aseptic meningitis and intracranial hypertension. The opportunistic agent could not be identified by cultures. Additional laboratory tests excluded toxoplasmosis, tuberculosis, bacterial cerebral abscesses, syphilitic cerebral gummas, and lymphoma. Brain CT and magnetic resonance studies were compatible with brain vasculitis and leptomeningitis. The patient condition improved with general measures, such as a repeated lumbar punctures and non steroidal anti-inflammatory drugs. We conclude that this patient had an IRIS due to a Cryptococcus neoformans antigen.

Cryptococcus neoformans causing meningitis in AIDS patients

Cryptococcus neoformans causes meningitis in AIDS patients. In the Säo José Hospital, reference center for patients with AIDS in Fortaleza - Ceará State - Brazil, 54 samples of the cerebrospinal fluid were analyzed from January 1st through December 31st 2000. Direct examination with India ink and culture in Sabouraud's agar at 37°C were done. Of the specimens studied 5(9.25 percent) were positive

Criptococosis diseminada en pacientes con SIDA. Análisis clÝnico, microbiológico e inmunológico de 51 pacientes / Disseminated cryptococcosis in patients with AIDS. Clinical, microbiological, and immunological analysis of 51 patients

In the context of HIV infection, cryptococcal meningitis is the most common mycosis threatening the patient's life. We conducted a retrospective evaluation to determine the epidemiological, microbiological, immunological and clinical characteristics of disseminated cryptococcosis in 51 hospitalised HIV seropositive patients. All the individuals (n = 51) presented reactive serology for HIV (ELISA and/or Western blot) and none fulfilled strict HAART treatment, previous to the opportunistic infection. CD4+ lymphocyte T counts showed levels between 361 and 0 cells/microliter (mean = 45). All patients but one had counts lower than 100 cells/microliter. Cryptococcosis presented as unique episode in 35 patients (68.6) and in 16 as relapse (31.3). In all of them we detected central nervous system involvement. The induction treatment was carried out with amphotericin B (AMB), continued with maintenance therapy with fluconazole. Lethality rate was 36.7, slightly superior among patients in relapse (40) compared to those who presented a first episode of the mycosis (35.2). In those individuals for whom data were available, 65.2 of blood cultures, 94.1 of CSF cultures and 79.06 of microscopic CSF examination with India ink were positive. Titers of Cryptococcus neoformans capsular antigen in CSF > or = 1/1000 were found in 36.1 and > or = 1/1000 in 73.6 of serum samples. In conclusion, manifestations and severity of disseminated cryptococcosis continue maintaining the characteristics of half a decade behind, in those patients who are not treated with HAART. Neurological involvement existed in all patients of this cohort. Treatment is not able to modify the parameters of mortality seen in previous communications. Diagnostic methods applied in this study are in accordance with those in the bibliography.

Criptococosis meníngea asociada al SIDA en el Hospital Diego Paroissien en el período 1996/1999 / AIDS-associated meningeal cryptococcosis in the Hospital Diego Paroissien from 1996-1999

A total of 148 patients with a diagnosis of HIV infection were studied in order to evaluate the incidence of meningeal cryptococcosis, including epidemic, immunologic and diagnostic characteristics. The diagnosis of cryptococcosis was carried out by direct examination with India ink and culture in Sabouraud agar of CSF in 28 patients (93.3) and by blood cultures (lysis-centrifugation) in 2 patients (6.6). All the isolated strains were identified as Cryptococcus neoformans. The incidence was 20.3 (30 patients). The preponderant risk behavior was endovenous drug addiction, and it was observed in 18 patients (60). The symptomatology that prevailed was headache (87). The median age was 28 years. At diagnosis, the immunologic impairment was severe (CD4+ lymphocyte count < 200) in 90 of patients. We found that 86.7 of patients had not completed their primary studies and only 13.3 had completed secondary studies. Although the acute mortality was high (36.7), it was observed that all the patients who survived (24) had been treated with anti-retroviral drugs.

Meningite criptocócica em pacientes imunocompetentes: relato de dois casos e revisäo da literatura / Cryptococcal meningitis in immunocompetent patients: report of two cases and literature review

A criptococose é uma doença sistêmica causada pelo fungo Cryptococcus neoformans, que acomete com mais freqüência os pulmões e o sistema nervoso central e, menos freqüentemente, a pele, o sistema esquelético e a próstata. Até a década de 80, antes do surgimento da AIDS, era relativamente rara. Apesar dela ocorrer com freqüência no hospedeiro imunodeprimido, cerca de um terço dos pacientes com a doença näo apresenta uma condiçäo de base, nem fatores predisponentes evidentes. Relatamos dois casos de pacientes imunocompetentes que apresentaram meningite criptocócica, enfatizando epidemiologia, etiopatogenia, clínica, diagnóstico e conduta terapêutica

Relato de cinco casos de meningite criptocócica em crianças com a síndrome da imunodeficiência adquirida (AIDS) / Report on five cases of cryptococcal meningitis in children with acquired immunodeficiency syndrome (AIDS)

Este trabalho relata cinco casos de meningite criptocócica, em crianças com AIDS, atendidas em 1995, no Hospital Municipal Jesus - Rio de Janeiro. A idade variou de quatro a oito anos. A transmissäo do vírus HIV foi vertical em quatro pacientes e provavelmente, também no outro. Quantro estavam em uso de antiretrovirais. A cefaléia foi o sintoma predominante (100 porcento). A febre esteve presente em dois casos. Os sintomas de irritaçäo meníngea estavam ausentes nos cincos pacientes. O líquor apresentava alteraçöes mínimas ou ausentes. A pesquisa do criptococo no líquor, pela Tinta da China, foi positiva em quatro crianças. Três faleceram e as outras duas, atualmente, apresentam sobrevida de 11 meses e 1 ano e 9 meses respectivamente, após o início da terapêutica. Foi empregada a anfotericina B intravenosa, numa fase inicial, seguida do fluconazol, por via oral, por tempo indeterminado. Comentários säo feitos sobre os achados clínicos, laboratoriais e a terapêutica empregada, comparando-se com trabalhos nacionais e internacionais. É importante ficarmos alerta aos sinais desta meningite, que apresenta um quadro clínico e liquórico com características próprias, diferentes das clássicas meningites observadas na infância

Criptococosis meníngea de 1990 a 1995: hallazgo de laboratorio / Cryptococcosis of 1990 to 1995: laboratory findings

El Laboratorio de Microbiología del Instituto Nacional de Salud ha prestado, durante los últimos 15 años, los servicios de diagnóstico, confirmación de aislamientos y determinación de las variedades y serotipos de Cryptocococcus neoformans, agente de la criptococosis. Entre 1990 y 1995, se recibieron 1.761 muestras de líquido cefalorraquídeo (LCR) y 1.068 sueros para el estudio de esta micosis; adicionalmente, se recibieron 66 aislamientos para su confirmación. A partir de estas muestras se clasificaron en tres grupos según los resultados de la prueba serólógica para VIH: 151 *62,9 por ciento) VIH positivos, 43 (17,9 por ciento) VIH negativos y 46 (19,2 por ciento) sin el dato. El porcentaje de los pacientes VIH positivos con criptococosis se incrementó de 26,7 por ciento, en 1990, a 80,4 por ciento en 1995. La entidad se presentó con mayor frecuencia en hombres (80,4 por ciento) que en mujeres (19,6 por ciento) y en el grupo de 21 a 40 años (55 por ciento), lo que fue más evidente en el grupo VIH positivos (67,5 por ciento). La sensibilidad y el valor predictivo negativo (VPN) del examen directo y de la aglutinación de látex en LCR y en suero presentaron valores más altos en los pacientes VIH positivos. El gran aumento en el número de casos de criptococosis es un reflejo del incremento de casos de SIDA en Colombia, epidemia que ha puesto de manifiesto el carácter oportunista de esta micosis

Estudio comparativo mediante pruebas inmunológicas y micológicas para el diagnóstico de la criptococosis meníngea / Comparative study through immunologic and mycologic test for the diagnosis of meningeal cryptococosis

Se estudió un grupo de 24 pacientes con diagnóstico de criptococosis meníngea. Con el líquido cefalorraquídeo se realizaron métodos tradicionales de diagnóstico, como son cultivos en Sabouraud y tinción de tinta china, y se compararon con un medio de cultivo diferencial de alpiste negro (Guizotia abissinica) de producción casera, y tinción de resaltado capsular. De manera independientes se determinó y tituló antígeno criptococósico en líquido cefalorraquídeo mediante aglutinación con anticuerpos adheridos a látex (reactivo previamente preparado y estandarizado). Los resultados obtenidos se analizaron en base a la prueba de X² y no se encontraron diferencias estadísticamente significtivas entre los métodos tradicionales y los propuestos. El medio de alpiste funcionó como un método diferencial eficaz (95 por ciento). La tinción de resaltado capsular fue ligeramente superior a la tradicional, sin diferencia significativa. La cuantificación de antígeno criptococósico fue una prueba sumamente sensible (83.3 por ciento) sin que se puediera correlacionar clínicamente

Criptococosis y su diagnóstico de laboratorio / Criptococosis and laboratory diagnostic

La criptococosis es una micosis oportunista producida por la levadura capsulada Cryptococcus neoformans, que se observa cada vez con más frecuencia; la variedad clínica más común es la meningitis y se asocia sobre todo con pacientes de SIDA (95 por ciento). El diagnóstico de laboratorio de ser integral, las pruebas más útiles son: examen en fresco con tinta china y tinción de resaltado capsular; los medios de cultivo rutinarios para su aislamiento son el Sabouraud y el agar alpiste negro (Niger o Staib), es necesaria la identificación posterior de la cepa por medios bioquímicos (auxonograma y zimograma). La prueba inmunológica más efectiva para diagnóstico y monitoreo terapéutico es la determinación de antígeno criptococósico por aglutinación directa con partículas de látex (DACAD). El pronóstico del padecimiento es malo, y el mejor exquema de tratamiento es a base de anfotericina B más fluconazol