Subcutaneous phaeohyphomycosis due to Phaeoacremonium venezuelense: The first clinical case report in Costa Rica.

Phaeoacremonium is a genus of dematiaceous fungi that rarely causes human infections. We describe a case of subcutaneous infection in a 70-year-old diabetic man with lesions on the dorsum of the one foot. The agent was isolated, and for the final identification we performed matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) and DNA sequencing. After diagnosis, the patient underwent curettage of the cyst and received 100mg of Itraconazole, twice daily for 6 months. Clinical resolution of the lesion was observed after treatment. This is the first case of infection by Phaeoacremonium venezuelense reported in Costa Rica.

Application of Real-Time PCR Assays for the Diagnosis of Histoplasmosis in Human FFPE Tissues Using Three Molecular Targets.

Histoplasmosis is a fungal infection caused by the thermally dimorphic fungus Histoplasma capsulatum. This infection causes significant morbidity and mortality in people living with HIV/AIDS, especially in countries with limited resources. Currently used diagnostic tests rely on culture and serology but with some limitations. No molecular assays are commercially available and the results from different reports have been variable. We aimed to evaluate quantitative real-time PCR (qPCR) targeting three protein-coding genes of Histoplasma capsulatum (100-kDa, H and M antigens) for detection of this fungus in formalin-fixed paraffin-embedded (FFPE) samples from patients with proven histoplasmosis. The sensitivity of 100-kDa, H and M qPCR assays were 93.9%, 91% and 57%, respectively. The specificity of 100-kDa qPCR was 93% when compared against samples from patients with other mycoses and other infections, and 100% when samples from patients with non-infectious diseases were used as controls. Our findings demonstrate that real-time PCR assays targeting 100-kDa and H antigen showed the most reliable results and can be successfully used for diagnosing this mycosis when testing FFPE samples.

Standardization and validation of real time PCR assays for the diagnosis of histoplasmosis using three molecular targets in an animal model.

Histoplasmosis is considered one of the most important endemic and systemic mycoses worldwide. Until now few molecular techniques have been developed for its diagnosis. The aim of this study was to develop and evaluate three real time PCR (qPCR) protocols for different protein-coding genes (100-kDa, H and M antigens) using an animal model. Fresh and formalin-fixed and paraffin-embedded (FFPE) lung tissues from BALB/c mice inoculated i.n. with 2.5x106 Histoplasma capsulatum yeast or PBS were obtained at 1, 2, 3, 4, 8, 12 and 16 weeks post-infection. A collection of DNA from cultures representing different clades of H. capsulatum (30 strains) and other medically relevant pathogens (36 strains of related fungi and Mycobacterium tuberculosis) were used to analyze sensitivity and specificity. Analytical sensitivity and specificity were 100% when DNAs from the different strains were tested. The highest fungal burden occurred at first week post-infection and complete fungal clearance was observed after the third week; similar results were obtained when the presence of H. capsulatum yeast cells was demonstrated in histopathological analysis. In the first week post-infection, all fresh and FFPE lung tissues from H. capsulatum-infected animals were positive for the qPCR protocols tested except for the M antigen protocol, which gave variable results when fresh lung tissue samples were analyzed. In the second week, all qPCR protocols showed variable results for both fresh and FFPE tissues. Samples from the infected mice at the remaining times post-infection and uninfected mice (controls) were negative for all protocols. Good agreement was observed between CFUs, histopathological analysis and qPCR results for the 100-kDa and H antigen protocols. We successfully standardized and validated three qPCR assays for detecting H. capsulatum DNA in fresh and FFPE tissues, and conclude that the 100-kDa and H antigen molecular assays are promising tests for diagnosing this mycosis.

Childhood histoplasmosis in Colombia: Clinical and laboratory observations of 45 patients.

Histoplasmosis is an important mycosis in the Americas; and in children with no immune system abnormalities, histoplasmosis is typically a self-limited process. In contrast, in children with immune problems, disease manifestations are frequently more severe and include dissemination. From 1984 to 2010, a retrospective study of paediatric patients who had been diagnosed with histoplasmosis was performed. A total of 45 pediatric cases of histoplasmosis were identified. The most important risk factor was malnutrition (37%), followed by environmental exposure (33%). The patients exhibited pulmonary infiltrates (83%), fever (76%), cough, constitutional symptoms (38%), headache (35%), and lymph node hypertrophy (33%). Concerning the clinical forms, 64% of the patients presented with the progressive disseminated form that frequently affected the central nervous system (48%). Diagnostic laboratory tests indicated that the cultures were positive for 80% of the patients, the agar gel immunodiffusion was reactive in 95%, the M band of the precipitate was more commonly observed (81%), and the complement fixation tests were reactive in 88% of the patients. The timely diagnosis of histoplasmosis is important, and for this reason, it is hoped that the results of this study will lead pediatricians toward a better understanding of this mycosis in children.

Whole-Genome Sequencing to Determine Origin of Multinational Outbreak of Sarocladium kiliense Bloodstream Infections.

We used whole-genome sequence typing (WGST) to investigate an outbreak of Sarocladium kiliense bloodstream infections (BSI) associated with receipt of contaminated antinausea medication among oncology patients in Colombia and Chile during 2013-2014. Twenty-five outbreak isolates (18 from patients and 7 from medication vials) and 11 control isolates unrelated to this outbreak were subjected to WGST to elucidate a source of infection. All outbreak isolates were nearly indistinguishable (<5 single-nucleotide polymorphisms), and >21,000 single-nucleotide polymorphisms were identified from unrelated control isolates, suggesting a point source for this outbreak. S. kiliense has been previously implicated in healthcare-related infections; however, the lack of available typing methods has precluded the ability to substantiate point sources. WGST for outbreak investigation caused by eukaryotic pathogens without reference genomes or existing genotyping methods enables accurate source identification to guide implementation of appropriate control and prevention measures.

Characterising atypical Candida albicans clinical isolates from six third-level hospitals in Bogotá, Colombia.

BACKGROUND: Candida species are the most frequently found fungal pathogens causing nosocomial disease in a hospital setting. Such species must be correctly identified to ensure that appropriate control measures are taken and that suitable treatment is given for each species. Candida albicans is causing most fungal disease burden worldwide; the challenge lies in differentiating it from emerging atypical, minor and related species such as Candida dubliniensis and Candida africana. The purpose of this study was to compare identification based on MALDI-TOF MS to standard identification systems using a set of nosocomial isolates. METHODS: Eleven nosocomial samples were collected from 6 third-level hospitals in Bogotá, Colombia. All the samples were identified by combining MALDI-TOF MS with morphological characters, carbohydrate assimilation and molecular markers (D1/D2 and HWP1). RESULTS: The present work describes the first collection of atypical Colombian Candida clinical isolates; these were identified as Candida albicans/Candida africana by their MALDI-TOF MS profile. Phenotypical characteristics showed that they were unable to produce chlamydospores, assimilate trehalose, glucosamine, N- acetyl-glucosamine and barely grew at 42 °C, as would be expected for Candida africana. The molecular identification of the D1/D2 region of large subunit ribosomal RNA and HWP1 hyphal cell wall protein 1 sequences from these isolates was consistent with those for Candida albicans. The mass spectra obtained by MALDI-TOF MS were analysed by multi-dimensional scaling (MDS) and cluster analysis, differences being revealed between Candida albicans, Candida africana, Candida dubliniensis reference spectra and two clinical isolate groups which clustered according to the clinical setting, one of them being clearly related to C. albicans. CONCLUSION: This study highlights the importance of using MALDI-TOF MS in combination with morphology, substrate assimilation and molecular markers for characterising Candida albicans-related and atypical C. albicans species, thereby overcoming conventional identification methods. This is the first report of hospital-obtained isolates of this type in Colombia; the approach followed might be useful for gathering knowledge regarding local epidemiology which could, in turn, have an impact on clinical management. The findings highlight the complexity of distinguishing between typical and atypical Candida albicans isolates in hospitals.

Prevalencia de vaginosis bacteriana, actinomyces spp, candida spp. y trichomonas vaginalis en usuarias del dispositivo intrauterino (DIU), Medellín, Colombia, 2011-2013 / Prevalence of bacterial vaginosis, actinomyces spp, candida spp, and trichomonas vaginalis in intrauterine device (IUD) users, Medellin, Colombia, 2011-2013

Introducción: el uso del dispositivo intrauterino incrementa el riesgo de algunas infecciones genitales. Objetivo: determinar la prevalencia de vaginosis bacteriana, Actinomyces spp., Candida spp. y Trichomonasvaginalis en usuarias del dispositivo intrauterino. Materiales y métodos: se realizó un estudio transversal en usuarias del dispositivo intrauterino, atendidas en un programa de tamización de cáncercérvico-uterino en Medellín, Colombia, entre 2011 y 2013. Se empleó una fuente de información secundaria basada en los registros citológicos del laboratorio clínico, se calculó la prevalencia global de las cuatro infecciones y las prevalencias específicas según grupo etario y sector de residencia, con un intervalo de confianza del 95%. La exploración de asociaciones se hizo con la prueba chi cuadrado de Pearson. Resultados: Se registraron 12.541 usuarias del dispositivo intrauterino de 10 comunas de Medellín. La edad promedio fue 34,0±9,8 años; el 50% de los valores centrales estuvo entre 26 y 41 años. Las prevalencias de infecciones vaginales fueron: vaginosis bacteriana 25,6%, Actinomyces spp. 8,9%, Candida spp. 5,1% y Trichomonas vaginalis 1,2%. La prevalencia de vaginosisbacteriana y Candida spp. fue estadísticamente mayor en adolescentes y jóvenes. La prevalencia de las cuatro infecciones fue estadísticamente diferente según el sector de residencia. Conclusión: en usuarias del dispositivo intrauterino las principales infecciones genitales son vaginosis bacteriana y Actinomyces spp., las adolescentes y jóvenes son los grupos de mayor riesgo para vaginosis bacterianay Candida spp. y la ocurrencia de infecciones varía entre los sectores de la ciudad; información relevante para la planeación de programas de prevención y atención.

Introduction: the use of intrauterine device increases the risk of genital infections. Objective: To determine the prevalence of bacterial vaginosis, Actinomyces spp., Candida spp., and Trichomonas vaginalis in intrauterine device users. Material and methods: A cross-sectional study was performed in intrauterine device users attended in a screening program for cervical cancer in Medellín, Colombia, between 2011 and 2013. A source of information was secondary, based on the records of the clinical laboratory of cytology. The overall prevalence of the four infections and specific prevalence by age group and residence area was calculated, all with confidence intervals of 95%. To explore associationsPearson chi-square test was used. Results: It was registered 12,541 users of the intrauterine device from 10 districts of Medellin. The users mean age was 34.0±9.8 years; 50% of the central values was between 26 and 41 years. The prevalence of vaginal infections were: bacterial vaginosis 25.6%, Actinomyces spp. 8.9%, Candida spp. 5.1% and Trichomonas vaginalis 1.2%. The prevalence of bacterial vaginosis and Candida spp. was statistically higher in adolescents and youth. The prevalenceof the four infections was statistically different according to the residence sector.Conclusion: in intrauterine devices users the major genital infections are bacterial vaginosis and Actinomycesspp. Adolescents and young people are the groups most at risk for bacterial vaginosis and Candida spp. and the occurrence of infections varies between sectors of the city; information that is relevant for planning prevention and care programs.