Cryptococcus and cryptococcosis in Cuba. A minireview.

Cryptococcosis has emerged as an important public health problem in Africa, Asia and the Americas due to the increasing numbers of persons at risk of this infection and the adaptation of its aetiological agents to new environments. The proper management requires early recognition of Cryptococcus neoformans/C. gattii species complex infection, familiarity with the use and limitations of diagnostic tests and knowledge of the available treatment options. This review will address these issues with the goal of providing sufficient information to suspect, diagnose and treat patients with cryptococcosis based on Cuban data and review of the literature.

Microsatellite typing and susceptibilities of serial Cryptococcus neoformans isolates from Cuban patients with recurrent cryptococcal meningitis.

BACKGROUND: Cryptococcus neoformans is commonly associated with meningoencephalitis in immunocompromised patients and occasionally in apparently healthy individuals. Recurrence of infection after initial treatment is not uncommon. We studied C. neoformans isolates from 7 Cuban patients with recurrent cryptococcal meningitis. Antifungal susceptibility and genotyping with microsatellite molecular typing were carried out. METHODS: Isolates (n = 19) were recovered from cerebrospinal fluid, blood, urine and semen. Antifungal susceptibilities for amphotericin B, fluconazole, flucytosine, itraconazole, voriconazole, posaconazole and isavuconazole were tested by CLSI M27A3 broth microdilution method. Genotyping was done using a panel of 9 microsatellite (STR) markers: (CT)n, (TG)n, (TA)n, (CTA)n, (TCT)n, (CCA)n, (TTAT)n, (ATCC)n and (TATT)n. RESULTS: The average number of isolates/patient was 2.71. The mean time interval between the collection of any two isolates was 52.5 days. All strains were identified as C. neoformans var. grubii (serotype Aα). Although none of the strains were resistant to the studied drugs, in serial isolates from two patients, MICs values of triazoles increased 4-5 log2 dilutions over time. STR patterns showed 14 distinctive profiles. In three patients the recurrent infection was associated with genotypically identical isolates. The four other patients had relapse isolates which were genotypically different from the initial infecting strain. CONCLUSION: Recurrences of cryptococcal meningitis in our series of patients was not associated with development of drug resistance of the original strain but by an initial infection with different strains or a reinfection with a new strain.

Chronic Pulmonary Aspergillosis in Patients with Underlying Respiratory Disorders in Cuba-A Pilot Study.

Chronic pulmonary aspergillosis (CPA) is a fungal infection with high mortality and morbidity rates. This disease is caused by several Aspergillus species and affects patients with an underlying respiratory condition. This pilot study aims to recognize CPA among patients with different respiratory diseases. Twenty-one out of 47 patients were classified as CPA based on the examination of clinical signs and symptoms, radiological findings, mycological culture of respiratory samples and analysis of Aspergillus IgG antibodies. There was a close association between high levels of Aspergillus IgG antibodies and the presence of cavities. Although Aspergillus flavus was the predominant species among clinical isolates, the number of isolates was small to reach conclusions on the prevalence of this species as main cause of CPA in Cuba. From the eleven evaluable patients for the treatment with itraconazole (Lozartil®), nine improved their health status while two did not show any recovery. This drug is included in the therapy schemes for aspergillosis in Cuba.

Determinación de la concentración mínima inhibitoria de lotes de OLEOZON® tópico frente a Trichophyton rubrum, Trichophyton mentagrophytes y Epidermophyton floccosum

Introducción: En la actualidad las infecciones por hongos afectan entre el 20 y el 25 % de la población. Objetivo: Determinar la concentración mínima inhibitoria de dos lotes de OLEOZON® almacenados a temperaturas de 5 y 30°C en envases de vidrio y polietileno de alta densidad, durante 24 meses como parte del estudio de estabilidad del OLEOZON® tópico. Métodos: Mediante el método dilución en agar fueron evaluadas cinco concentraciones del producto frente a los dermatofitos trichophyton rubrum, trichophyton mentagrophytes y epidermophyton floccosum. Resultados: Se obtuvo que el 8,9 mg/mL fue el valor de la CMI para las cepas evaluadas en el estudio "vida de estante"; se observó en el estudio acelerado el mismo valor frente a las cepas de trichophyton mientras que para epidermophyton floccosum fue de 17,8 mg/mL a excepción del envase frasco de vidrio del lote 803295 donde se obtuvo 8,9 mg/mL. El análisis estadístico, tanto del estudio acelerado como en vida de estante, mostró que existe diferencia estadísticamente significativa entre el primer y el último mes de ensayo, estas son las diferencias más apreciables en los lotes almacenados en frasco de vidrio. El OLEOZON® tópico almacenado en frasco de polietileno de alta densidad presentó mejores valores de actividad frente a los dermatofitos. Conclusiones: Todos los valores de la concentración mínima inhibitoria encontrados, independiente del tipo de envase, el tiempo o la temperatura de almacenamiento, muestran que el producto mantiene su actividad antifúngica. Se evidenció una similitud entre las cepas del género trichophyton en comparación con el género epidermophyton.

Introduction: Currently fungal infections affect between 20 and 25% of the population. Objective: To determine the minimum inhibitory concentration of two batches of OLEOZON® stored at temperatures of 5 and 30 0C in glass and high-density polyethylene containers, for 24 months as part of the stability study of topical OLEOZON®. Methods: Using the agar dilution method, five concentrations of the product were evaluated against the dermatophytes Trichophyton rubrum, Trichophyton mentagrophytes and Epidermophyton floccosum. Results: It was found that 8.9 mg/mL was the MIC value for the strains evaluated in this shelf life study; and the same value was observed in the accelerated study against the trichophyton strains, while for epidermophyton floccosum it was 17.8 mg/mL with the exception of the glass bottle container of batch 803295 where 8.9 mg/mL was obtained. The statistical analysis, both in the accelerated study and in shelf life, showed that there is a statistically significant difference between the first and the last month of the trial, these are the most appreciable differences in the batches stored in glass jars. Topical OLEOZON® stored in a high-density polyethylene bottle presented better activity values against dermatophytes. Conclusions: All the values of the minimum inhibitory concentration found, regardless of the type of container, the time or the storage temperature, show that the product maintains its antifungal activity during the months of study. A similarity was apparent between the strains of the genus trichophyton compared to the genus epidermophyton.

Aportes del Laboratorio de Micología del Instituto de Medicina Tropical Pedro Kourí al desarrollo de la especialidad en Cuba / Contributions of the Mycology Laboratory at Pedro Kouri Tropical Medicine Institute to the development of the specialty in Cuba

En el marco del 80 aniversario del Instituto de Medicina Tropical Pedro Kourí, se exponen en apretada síntesis, las actividades más relevantes en el campo del diagnóstico, la investigación y la docencia desarrolladas hasta el momento por el Laboratorio de Micología de la referida institución. A 38 años de su creación, este sigue siendo uno de los pocos lugares en Cuba que se desempeña en este campo y que se reconoce por los logros científico-técnicos alcanzados. Sin lugar a duda, estos responden básicamente al cuadro de salud nacional, sin descuidar el entorno regional y mundial. Todos los resultados que se exponen, se encuentran sustentados por más de 100 publicaciones y otras obras científicas, lo que avala la calidad y rigor del trabajo desplegado. Sirva este artículo para homenajear al fundador del Instituto de Medicina Tropical Pedro Kourí y a todos los científicos que en el transcurso de estos años contribuyeron a engrandecer su nombre(AU)

In the framework of the 80th anniversary of Pedro Kouri Tropical Medicine Institute, a succinct presentation is provided of the most relevant activities in the fields of diagnosis, research and teaching so far performed by the Mycology Laboratory at the Institute. Thirty-eight years after its founding, the Laboratory continues to be one of the few places in Cuba devoted to this field and recognized for its scientific and technical achievements, which no doubt essentially respond to the national health status, without neglecting the regional and global environment. All the results presented are endorsed by over 100 publications and scientific works, which is evidence of the quality and rigor of the work done. May this paper be a tribute to the founder of Pedro Kouri Tropical Medicine Institute and all the scientists who have brought prestige to its name throughout these years(AU)

Susceptibilidad antifúngica de aislados vaginales de Candida spp / Antifungal susceptibility of Candida spp. vaginal isolates

Introducción: la vulvovaginitis constituye una de las principales afecciones ginecológicas, y su causa más frecuente es la candidiasis. Candida albicans se considera el agente etiológico más importante de esta entidad; sin embargo, estudios recientes revelan un incremento en la incidencia de otras especies del género. Algunas de estas tienen la particularidad de presentar resistencia a los tratamientos usuales con antimicóticos. Objetivo: evaluar la susceptibilidad antifúngica de aislados vaginales de pacientes cubanas con sospecha de candidiasis vulvovaginal que se obtuvieron en el 2015. Métodos: a 28 aislados pertenecientes al género Candida, se les realizó las pruebas de susceptibilidad in vitro con la galería ATBTM Fungus 3 frente a diferentes antifúngicos (5-fluorocitosina, anfotericina B, fluconazol, itraconazol y voriconazol). Resultados: todos los aislados fueron sensibles a la anfotericina B y uno de C. albicans se informó resistente a los azoles estudiados. Todas las especies diferentes de C. albicans fueron susceptibles al voriconazol (CMI≤ 1 mg/L). Conclusiones: el estudio de patrones de susceptibilidad en aislados de Candida provenientes de mujeres con vulvovaginitis permite profundizar en cómo abordar la terapéutica de esta afección; el fluconazol resultó el tratamiento de elección. Los resultados alertan sobre la emergencia de C. glabrata, C. krusei, C. parapsilosis, C. tropicalis, C. inconspicua y C. lusitaniae como agentes causales de la candidiasis vulvovaginal(AU)

Introduction: vulvovaginitis is one of the main gynecological diseases frequently caused by candidiasis. Candida albicans is considered as the most important etiological agent for the disease; however, recent students have revealed an increased incidence of other species of the genus. Some of them may show particular resistence to usual antimycotic treatments. Objective: to evaluate the antifungal susceptibility of vaginal isolates from Cuban female patients suspected of vulvovaginal candidiasis in 2015. Methods: twenty eight Candida genus isolates underwent in vitro susceptibility tests with ATBTM Fungus 3 using several antifungal agents (5 fluorocytosine, anphotericin B, fluconazole, itraconazole and vorixonazole). Results: all isolates were susceptible to B anphotericin and one C. albicans isolate was reported as resistant to the studied azoles. All the species other thanC. albicans were susceptible to voriconazole (CMI≤ 1mg/L). Conclusions: the study of susceptibility patterns in Candida isolates from women with vulvovaginitis allow delving into the different ways of approaching the therapeutics of this disease; fluconazole was the treatment of choice. The results show emergence of C. glabrata,C. krusei, C. parapsilosis, C. tropicalis, C. inconspicua and C. lusitaniae as causative agents of vulvovaginal candidiasis(AU)

Neurocriptococosis en el contexto de la infección con el VIH en Guayaquil, Ecuador / Neurocryptococcosis in the context of HIV infection in Guayaquil, Ecuador

Introducción: la neurocriptococosis resulta de la inhalación de levaduras del complejo de especies Cryptococcus neoformans y afecta principalmente a pacientes inmunocomprometidos, provocando altas tasas de mortalidad. Objetivo: describir la infección por Cryptococcus spp. en pacientes con VIH/sida de Guayaquil, Ecuador. Métodos: estudio descriptivo, transversal y prospectivo, entre diciembre/2013-enero/2015. Se recopilaron 82 muestras de líquido cefalorraquídeo así como los datos demográficos, clínicos y de laboratorio de igual cantidad de pacientes seropositivos al VIH ingresados en el Hospital de Infectología Dr. José Daniel Rodríguez Maridueña. La infección criptococósica se confirmó mediante examen microscópico directo del líquido cefalorraquídeo con tinta china, cultivo en agar Sabouraud y pruebas bioquímicas convencionales. El estudio cumplió con los requerimientos éticos establecidos. Resultados: el 89,02 por ciento de los pacientes incluidos en el estudio fueron de género masculino y el 45,12 por ciento del grupo etario de 20-30 años. El 33 por ciento de los pacientes presentaron la infección por C. neoformans, y sus características clínicas más frecuentes fueron: impresión diagnóstica de neuroinfección (41 por ciento), cefalea (78 por ciento; 21/27), vómitos (85 por ciento; 23/27) y pérdida de peso (89 por ciento; 24/27); niveles de CD4 < 200/uL (26 por ciento; 7/27), leucocitos 5 000-10 000 cél/mm3 (63 por ciento; 17/27), hemoglobina 11-15 g/dL (44 por ciento; 12/27) y hematócrito < 35 por ciento (78 por ciento; 21/27). Se demostró además, la existencia de una asociación entre la infección y la presencia de vómitos, pérdida de peso y adenopatías (razón de prevalencias > 1). Conclusiones: La infección criptococósica es una importante micosis oportunista en pacientes VIH-SIDA, que puede ser asociada a determinadas características, lo que permite definir mecanismos de control y prevención(AU)

Introduction: neurocryptococcosis results from inhalation of yeasts from the Cryptococcus neoformans species complex. The disease mainly affects immunocompromised patients, causing high mortality rates. Objective: describe infection due to Cryptococcus spp. in patients with HIV/AIDS from Guayaquil, Ecuador. Methods: a descriptive cross-sectional prospective study was conducted from December 2013 to January 2015. Eighty-two cerebrospinal fluid samples were collected, as well as the demographic, clinical and laboratory data of an equal number of HIV seropositive inpatients from Dr. José Daniel Rodríguez Maridueña infectious diseases hospital. Cryptococcal infection was confirmed by India ink direct microscopic examination of cerebrospinal fluid, Sabouraud agar culture and conventional biochemical tests. The study met the ethical requirements established. Results: 89.02 percent of the patients included in the study were male and 45.12 percent were in the 20-30 years age group. 33 percent had infection with C. neoformans, and their most common clinical features were diagnostic impression of neuroinfection (41 percent), headache (78 percent; 21/27), vomiting (85 percent; 23/27), weight loss (89 percent; 24/27); CD4 counts < 200/uL (26 percent; 7/27), leucocytes 5 000-10 000 cells/mm3 (63 percent; 17/27), hemoglobin 11-15 g/dL (44 percent; 12/27) and hematocrit < 35 percent (78 percent; 21/27). An association was also found between infection and the presence of vomiting, weight loss and adenopathies (prevalence ratio >1). Conclusions: cryptococcal infection is an important opportunistic mycosis in HIV/AIDS patients. It may be associated with certain features, which makes it possible to define control and prevention mechanisms(AU)

[Effect of 4B3 monoclonal antibody on the experimental Cryptococcus neoformans infection]. / Efecto del anticuerpo monoclonal 4B3 en la infección experimental por Cryptococcus neoformans.

INTRODUCTION: frequent relapses and high lethality of criptococcosis has encouraged the search for new therapeutic strategies. OBJECTIVES: to evaluate the effect of the monoclonal antibody 4B3 on the cryptococcal infection in Balb/c mice. METHODS: the kinetics in serum concentration of the studied monoclonal antibody was determined for intraperitoneal administration (500 microg) by quantitative sandwich ELISA. In order to assess its protective capability, were administered 500 microg of 4B3 and innoculated 2 x 10(22) cells/mL of Cryptococcus neoformans. The survival of mice was recorded and the yeast dissemination to the main target organs was evaluated. Macrophages P338.D1 cell lines were used to measure the effect of the monoclonal antibody 4B3 on the phagocytosis and lysis of the microorganism. RESULTS: the used dose helped to keep high values (38 microg/m) of 4B3 in serum for at least 46 days. It was found that the monoclonal antibody does not give protection, which makes the microorganism dissemination possible, along with the reduction in the survival of mice. Finally, the phagocytosis test showed that 4B3 increased the phagocytic activity of macrophages without any fungicidal effect. CONCLUSIONS: the monoclonal antibody 4B3 stimulates C. neoformans phagocytosis by macrophages without fungicidal effect, thus favoring yeast dissemination and decreasing the survival of mice due to cryptococcal infection

[Phenotypic characterization of Aeromonas isolated from patients with acute diarrheal diseases]. / Caracterización fenotípica de cepas de Aeromonas aisladas de pacientes con enfermedad diarreica aguda.

INTRODUCTION: The acute diarrheal diseases are one of the main causes of morbidity and mortality in children aged under 5 years and in the general population; this demands a great deal of care in the healthcare services. The situation worsens due to the overuse of antimicrobials and the development of bacterial resistance, being the latter an emerging health problem in different areas of the world. Among the causative microorganisms of acute diarrheal diseases are those of Aeromonas genus, recognized as second risk emerging pathogens. OBJECTIVES: To determine the most frequently isolated Aeromonas species in fecal samples from acute diarrheal patients and their antimicrobial susceptibility. METHODS: The Bauer-Kirby's method allowed identifying the susceptibility to several antimicrobials of 100 isolated samples coming from the provincial hygiene and epidemiology centers during 2007 and 2008. RESULTS: Identification of the species was accomplished in 67% of isolates, being A. caviae (33%) y A. hydrophila (29%) the predominant species. It was demonstrated that 100% of isolates got resistant to at least one of the studied antimicrobials. Drug resistance to cefalotine, sulfonamides and nalidixic acid showed the highest percentages. CONCLUSIONS: Tetracycline and chloramphenicol are recommended as the drugs of choice for treating intestinal infections caused by these microorganisms in Cuba.

[Outbreaks of occupational acquired histoplasmosis in La Habana province]. / Brotes de histoplasmosis ocupacional en la provincia La Habana.

INTRODUCTION: histoplasmosis, an infection caused by Histoplasma capsulatum fungus, is considered as one of the main endemic infections in the American continent where there are frequent epidemic outbreaks. Most of the outbreaks have been associated to risky activities in bat-inhabited places. This paper presented two outbreaks of occupational acquired histoplasmosis occurred in La Habana province. METHODS: one hundred and sixteen workers, who performed aerosol-generating tasks in caves, were studied. Samples from the cave soils were taken to assess the existence of the causal agent. Clinical and epidemiological data were gathered for the outbreak characterization. RESULTS: the rates of infection for both outbreaks were 60 % and 78 % respectively. The patients aged 16 to 64 years; additionally, the onset of main clinical manifestations (fever, general malaise, cough, headache, loss of weight, and chest pain) ranged from 10 to 20 days. Histoplasmosis was microbiologically and serologically diagnosed whereas its source of infection was confirmed through H. capsulatum isolation in those places where extensive excavation work had been carried out, resulting in abundant aerosols for up to 21d periods. The similarity between the two outbreaks was the performance of aerosol-generating tasks in risky places without using appropriate safety devices. CONCLUSIONS: It is necessary to disseminate the existing preventive measures among the health care staff and the risky groups as well as the general population in order to prevent and take control of possible histoplasmosis outbreaks in the future.

[Cyptococcus gattii isolated from a cheetah (Acinonyx jubatus) in the National Zoo of Cuba]. / Cryptococcus gattii aislado de un guepardo (Acinonyx jubatus) del Parque Zoológico Nacional de Cuba.

INTRODUCTION: Cryptoccosis--systemic mycosis caused by Cryptococcus species--has considerably raised its incidence in the last years, mainly associated with the human immunodeficiency virus infection. It has also been described in animals, but rare cases. METHODS: a case of a female cheetah (Acinonyx jubatus) kept in the Nacional Zoo of Havana was presented. The animal came from South Africa. She began losing weight, and suffering asthenia, anorexia and breathing problems with abundant nasal secretion. RESULTS: mycological testing of these secretions disclosed the presence of serotype B Cryptococcus gattii. Because of the origin and captive condition of the animal, it was believed that the infection had been latent for 16 months at least. CONCLUSIONS: up to the present, in Cuba, all clinical Cryptococcus isolates were C. neoformans var. grubii, so it is considered that the infection was caught in the country of origin of the female cheetah. This is the first C. gattii isolate in Cuba from an animal coming from South Africa where this fungus is endemic.

Presentación de siete casos de tiña negra en niños / Reports of seven cases of tinea nigra in children

Introducción: la tiña negra es una micosis superficial causada por el hongo Hortaea werneckii. Se considera una micosis benigna que por lo general es observada en países tropicales. Objetivo: reportar siete casos de tiña negra en niños de dos hospitales de La Habana, Cuba. Métodos: se realizó estudio micológico (examen directo y cultivo) a partir de escamas tomadas mediante raspado de las lesiones a siete niños con diagnóstico clínico presuntivo de tiña negra palmar. Se registraron las características de las lesiones, edad, sexo y factores predisponentes de los pacientes, así como la evolución del cuadro con el tratamiento antifúngico. Resultados: se confirmó la sospecha clínica de tiña negra a través del aislamiento e identificación de Hortae werneckii. Las edades de los pacientes oscilaron entre 3 y 6 años y el 57 por ciento era del sexo femenino. La hiperhidrosis se encontró en el 43 por ciento de los casos. El tratamiento específico con antifúngicos azólicos y terbinafina tópicos fue satisfactorio en 21 días como promedio. Conclusiones: todos los casos con sospecha de tiña negra fueron confirmados de manera oportuna en el laboratorio, lo que permitió descartar enfermedades malignas y aplicar tratamiento específico(AU)

Microsatellite typing of clinical and environmental Cryptococcus neoformans var. grubii isolates from Cuba shows multiple genetic lineages.

BACKGROUND: Human cryptococcal infections have been associated with bird droppings as a likely source of infection. Studies toward the local and global epidemiology of Cryptococcus spp. have been hampered by the lack of rapid, discriminatory, and exchangeable molecular typing methods. METHODOLOGY/PRINCIPAL FINDINGS: We selected nine microsatellite markers for high-resolution fingerprinting from the genome of C. neoformans var. grubii. This panel of markers was applied to a collection of clinical (n = 122) and environmental (n = 68; from pigeon guano) C. neoformans var. grubii isolates from Cuba. All markers proved to be polymorphic. The average number of alleles per marker was 9 (range 5-51). A total of 104 genotypes could be distinguished. The discriminatory power of this panel of markers was 0.993. Multiple clusters of related genotypes could be discriminated that differed in only one or two microsatellite markers. These clusters were assigned as microsatellite complexes. The majority of environmental isolates (>70%) fell into 1 microsatellite complex containing only few clinical isolates (49 environmental versus 2 clinical). Clinical isolates were segregated over multiple microsatellite complexes. CONCLUSIONS/SIGNIFICANCE: A large genotypic variation exists in C. neoformans var. grubii. The genotypic segregation between clinical and environmental isolates from pigeon guano suggests additional source(s) of human cryptococcal infections. The selected panel of microsatellite markers is an excellent tool to study the epidemiology of C. neoformans var. grubii.

[Susceptibility to nystatin of oral Candida isolates and its correlation with the response to treatment]. / Susceptibilidad a la nistatina de aislamientos bucales de Candida y su correlación con la respuesta al tratamiento.

INTRODUCTION: oropharyngeal candidiasis is an early marker of progression to AIDS in H1V-positive patients and an indicator of non-adherence of treatment or possible failure in patients undergoing anti-retroviral therapy. OBJECTIVE: to determine the in vitro susceptibility to nystatin in Candida strains isolated from the oral cavity of HIV-positive patients, and to correlate the results with the response to treatment. METHODS: a study of in vitro susceptibility to nystatin was conducted in 104 oral isolates from 97 HIV/AIDS patients, who participated as volunteers in a clinical trial to evaluate the response to four antifungal drugs. Fifty-eight of the 104 isolates were obtained before starting treatment and the remaining 46 at the end of therapy. Broth microdilution method was performed according to the document M27-A3 of the Clinical and Laboratory Standards Institute. RESULTS: one hundred percent of the strains were susceptible to nystatin, with minimum inhibitory concentration (MIC) values of <16 microg/mL, regardless of the time of isolation and of the drug administered to patients. In the group of patients treated with nystatin, the MIC80 ranges (1-2 microg/mL), geometric mean and cumulative values of the drug against Candida albicans isolates before starting treatment were similar to those obtained against strains recovered at the end. Among the identified species, C. lusitaniae reached the highest absolute MIC value (4 microg/mL). CONCLUSIONS: the correlation between the clinical evolution and the results of in vitro susceptibility tests was good in patients with a favorable outcome; however, it did not allow predicting possible treatment failure.

Diagnóstico molecular de histoplasmosis diseminada en pacientes cubanos con sida / Molecular diagnosis of disseminated histoplasmosis in Cuban aids patients

Introducción: Histoplasma capsulatum es el agente causal de la histoplasmosis. La forma diseminada de esta micosis de no ser tratada suele ser fatal. Objetivo: evaluar el empleo de sangre periférica para el diagnóstico molecular de histoplasmosis diseminada y comparar sus resultados con el cultivo y la detección de anticuerpos específicos. Métodos: se determinó la presencia de ADN de Histoplasma capsulatum a partir de 12 muestras de sangre de pacientes con sida y sospecha clínica de histoplasmosis diseminada, mediante dos sistemas de reacción en cadena de la polimerasa (una simple y otra anidada, que amplifican un fragmento de la región ITS y del gen Hcp100, respectivamente). Se compararon los resultados con el cultivo y la serología. Resultados: mediante la reacción en cadena de la polimerasa simple y anidada se diagnosticó con histoplasmosis diseminada a los 12 pacientes. Por cultivo resultaron positivos 6 casos (50 por ciento). Mientras que la presencia de anticuerpos anti-Histoplasma capsulatum se determinó en 3 casos (25 por ciento). Los sistemas de reacción en cadena de la polimerasa empleados mostraron elevada sensibilidad, especificidad analítica y diagnóstica en la detección de Histoplasma capsulatum en esta forma clínica. Conclusiones: los resultados del estudio sugieren que las técnicas de reacción en cadena de la polimerasa aumentan la fiabilidad del diagnóstico de la histoplasmosis cuando se utiliza en combinación con los métodos establecidos. Los sistemas de reacción en cadena de la polimerasa aplicados al diagnóstico de histoplasmosis diseminada en pacientes con sida probaron ser sistemas rápidos, específicos y sensibles(AU)

Introduction: Histoplasma capsulatum is the causal agent of histoplasmosis. If not treated, the disseminated form of this mycosis is often fatal. Objective: evaluate the use of peripheral blood for the molecular diagnosis of disseminated histoplasmosis and compare the results with those obtained by culture and detection of specific antibodies. Methods: the presence of Histoplasma capsulatum DNA was determined from 12 blood samples from patients with AIDS and clinical suspicion of disseminated histoplasmosis, using two polymerase chain reaction systems (one simple and one nested, amplifying a fragment of the ITS region and the Hcp100 gene, respectively). The results were compared with those obtained by culture and serology. Results: all 12 patients were diagnosed with disseminated histoplasmosis by simple and nested polymerase chain reaction. Culture results were positive in 6 cases (50 percent), whereas the presence of anti-Histoplasma capsulatum antibodies was determined in 3 cases (25 percent). The polymerase chain reaction systems used showed high sensitivity, and analytical and diagnostic specificity for detection of this clinical form of Histoplasma capsulatum. Conclusions: results suggest that polymerase chain reaction techniques increase the reliability of histoplasmosis diagnosis when used in combination with established methods. When used for the diagnosis of disseminated histoplasmosis in AIDS patients, polymerase chain reaction systems proved to be fast, specific and sensitive(AU)

Aspectos epidemiológicos, clínicos, microbiológicos y terapéuticos de la histoplasmosis / Epidemiological, clinical, microbiological and therapeutic aspects of histoplasmosis

El Histoplasma capsulatum se considera un hongo dimórfico. Constituye una micosis endémica que presenta un comportamiento oportunista. Su modo de transmisión es principalmente respiratorio, por la inhalación de conidios presentes en la naturaleza. El cuadro clínico puede variar desde infecciones asintomáticas hasta cuadros graves diseminados, sobre todo en pacientes con sida, trasplantados o con neoplasias hematológicas. El diagnóstico del hongo se obtiene de fluidos orgánicos o tejidos, el cultivo de materiales biológicos y la serología. Se utilizaron las bases de datos de Scielo, HINARI, Pubmed-Medline y diferentes textos y artículos donados por especialistas de la materia, los cuales resultaron muy útiles para confeccionar la revisión. Los trabajos encontrados mostraron una información útil y valiosa de la histoplasmosis, entidad considerada la micosis respiratoria más frecuente en pacientes inmunocompetentes y la tercera causa de muerte en pacientes con sida. El objetivo de este trabajo es presentar una actualización sobre los principales aspectos clínicos, epidemiológicos, microbiológicos y terapéuticos de la micosis sistémica más frecuentes en Cuba y en el continente americano.

Histoplasma capsulatum is considered a dimorphic fungus. It is an endemic mycosis presenting opportunistic behavior. Its transmission is mainly by respiratory inhalation of naturally occurring conidia. The clinical conditions may vary from asymptomatic infections to severe disease spread, especially in AIDS patients, transplant or with haematological malignancies patients. The diagnosis of the fungus is obtained from body fluids or tissues, the culture of biological materials and serology. The database used in this work was selected from SciELO, HINARI, PubMed, Medline databases and various texts and articles donated by specialists, which were very useful material for this review. The papers found showed useful and valuable information of histoplasmosis, which is a breathing entity considered the most common fungal infection in immuno-competent patients and the third cause of death in AIDS patients. The objective of this review is to present an update on key clinical, epidemiological, microbiological and therapeutic systemic fungal infection, which are most common in Cuba and America.

Contribución al estudio de la criptococosis infantil en Cuba / Contribution to the study of infant cryptococosis in Cuba

Introducción: a partir de 1998 en el laboratorio de micología del Instituto Pedro Kourí se observó un incremento del número de aislamientos de Cryptococcus obtenidos de niños seronegativos al VIH con meningoencefalitis. Los reportes de criptococosis infantil a nivel mundial son infrecuentes y generalmente se encuentran asociados al sida. Objetivo: estudiar la criptococosis infantil en Cuba a través de la evaluación integral de 7 casos. Métodos: se realizó un estudio retrospectivo integral, se revisaron las historias clínicas de 7 pacientes pediátricos afectados por criptococosis menigoencefálica y se caracterizaron los aislamientos obtenidos (uno de cada paciente). A estos se les determinó la especie, variedad, serotipo, patogenicidad en un modelo murino y susceptibilidad a la anfotericina B. Resultados: la edad media de los niños fue de 5,8 años, con ligero predominio del sexo femenino (4:3). Los síntomas más frecuentes resultaron la cefalea, fotofobia, fiebre y los vómitos. En 6 pacientes el estudio citoquímico del líquido cefalorraquídeo y el examen imaginológico de cráneo evidenciaron anomalías. El diagnóstico microbiológico se realizó mediante examen directo y cultivo. Todos los aislamientos correspondieron con Cryptococcus neoformans serotipo A y la anfotericina B mostró una concentración mínima inhibitoria menor o igual que 0,125 mg/mL. El 85,7 porciento mostró actividad fenoloxidasa y elevada letalidad en los animales infectados (71,4-100 porciento). Conclusiones: el pronóstico de la meningoencefalitis por Cryptococcus está estrechamente ligado a un diagnóstico precoz. Esta realidad hace que deba tenerse siempre presente, por lo que los datos aportados bien pudieran servir de alerta a los médicos de asistencia

Introduction: since 1998, an increasing number of Cryptococcus isolates obtained from HIV-negative children with meningoencephalitis was observed in the mycology laboratory at Pedro Kourí Institute. The global reports of child cryptococcosis are rare and usually associated with AIDS. Objectives: to study child cryptococosis in Cuba through the comprehensive evaluation of 7 cases. Methods: the clinical histories of seven pediatric patients with cryptococcal meningoencephalitis were examined and the obtained isolates (one per patient) were characterized. The species, variety, serotype, pathogenicity in a mouse model and amphotericin B susceptibility of the isolates were determined. Results: average age of the children was 5.8 years, with a slight female predominance (4:3). The most common symptoms were headache, photophobia, fever and vomiting. The cytochemical study of cerebral spinal fluid and skull imaging test showed abnormalities in 6 patients. The microbiological diagnosis was made through direct examination and culture. All the isolates were from Cryptococcus neoformans serotype A and amphotericin B showed a minimum inhibitory concentration £ 0.125 mg/mL. In the group, 85.7 percent showed phenoloxidase activity and high lethality rate in infected animals (71.4 to 100 percent). Conclusions: the prognosis of cryptococcal meningoencephalitis is closely linked to early diagnosis, therefore, this should be always kept in mind and the provided data from this study may serve as warning to attending physicians

Conservación de cultivos de hongos de importancia médica en agua destilada / Preservation of fungal cultures of medical importance in distilled water

Introducción: en la colección de cultivos de hongos patógenos del Instituto de Medicina Tropical Pedro Kourí, como centro de referencia nacional, se conservan los agentes causales de las principales micosis humanas, con fines de diagnóstico, investigaciones y enseñanza de la microbiología médica. La conservación de cultivos fúngicos en agua destilada estéril o método de Castellani ha sido avalada como un método que garantiza porcentajes elevados de viabilidad, pureza y estabilidad de las cepas; esto unido a su bajo costo y sencillez, la convierte en una alternativa ventajosa para el mantenimiento de los microorganismos en muchos laboratorios. Objetivo: mostrar los resultados en la preservación en agua destilada estéril de las principales especies fúngicas que integran esta colección. Métodos: se evaluó la viabilidad, pureza y estabilidad de las principales características morfológicas y fisiológicas de 240 cepas de diferentes especies fúngicas pertenecientes a la colección, conservadas en agua destilada estéril. Resultados: de los cultivos, 80 por ciento se conservó en estado viable y sin contaminaciones. Los mejores resultados se obtuvieron en la preservación de los hongos productores de abundantes conidios (97 por ciento de recuperación) y las levaduras (83 por ciento), mientras que con los dermatofitos y hongos dimórficos fue de 69 y 68 por ciento, respectivamente. En 17 géneros, la recuperación de cepas viables fue superior a 60 por ciento, mientras que en 8 resultó de 100 por ciento. El tiempo de conservación fue de 15 a 20 años. Se implementó una base de datos en formato digital de la colección sobre plataforma web. Conclusiones: el método de Castellani es un método de elección para la conservación de cultivos fúngicos en laboratorios de recursos limitados

Introduction: causal agents of the main human myicoses have been preserved in the culture collection of pathogenic fungi at Pedro Kourí Tropical Medicine Institute, a national reference center, with the purpose of using them for medical microbiology diagnoses, research and teaching. Preservation of fungal cultures in sterile distilled water, or Castellani's method, has been endorsed as a method ensuring high rates of strain viability, purity and stability, low costs and great simplicity. It is therefore an advantageous alternative for the maintenance of microorganisms in many laboratories. Objective: present the results obtained from the preservation in sterile distilled water of the main fungal species included in the collection. Methods: an evaluation was conducted of the viability, purity and stability of the main morphological and physiological characteristics of 240 strains of different fungal species from the collection, which had been preserved in sterile distilled water. Results: 80 percent of the cultures had retained their viable status without any contamination. The best results corresponded to the preservation of fungi producing abundant conidia (97 percent recovery) and yeasts (83 percent), followed by dermatophytes (69 percent) and dimorphic fungi (68 percent). In 8 genera, recovery of viable strains was 100 percent, and in 17 it was above 60 percent. Preservation time was 15-20 years. A web-based digital database was created for the collection. Conclusions: Castellani's is the method of choice for the preservation of fungal cultures in resource-limited laboratories

[In vitro susceptibility of Candida strains to fluconazole and amphotericin B]. / Sensibilidad in vitro de cepas de Candida frente a fluconazol y anfotericina B.

In vitro antifungal susceptibility testing is really necessary at present in medical mycology laboratories. Minimal inhibitory concentration of 210 Candida yeast strains isolated from HIV/AIDS patients with oral lesions was determined to find out susceptibility to fluconazole and amphotericin B. A CLSI's M27A2 protocol-based dilution micromethod was used. C. albicans was predominant (62.4%)followed by C. glabrata, C. parapsilosis, and C. tropicalis. Regarding amphotericin B, the mode value and MIC50 were 0.125 microg/mL, which indicated the effectiveness of this drug in vitro conditions. Only C. krusei strain showed a minimal inhibitory concentration over 1 microg/mL. 8.1% of strains were resistent to fluconazole whereas 8.1% was dose-depending susceptible. The less susceptible species were C. krusei and C. glabrata. The achieved results laid the foundations for wider studies to be made in the Cuban context and indicated the need of keeping surveillance on the behaviour of clinical yeast isolates to antifungal drugs.