First report on mycetoma in Turkana County-North-western Kenya.

Mycetoma is one of the six Neglected Tropical Diseases that are prevalent in Turkana County (northwest Kenya). The aim of the study was to estimate the prevalence of mycetoma in the county, as well as to describe the main causative agents involved in the disease using methods affordable locally. Based on the data collected by the team of cooperative medicine Cirugia en Turkana (Surgery in Turkana), a specific study for mycetoma was started during the 16th humanitarian medicine campaign in February 2019. Patients with suspected mycetoma were studied at the Lodwar County Referral Hospital (LCRH). After informing the patient and getting their consent, the lesions were examined and sampled (mainly by biopsy) and clinical data were recorded. Samples were washed in sterile saline solution and cut in fragments. Some of these were inoculated on Sabouraud Dextrose Agar, Malt Extract Agar, and diluted Nutrient Agar plates. One fragment of each sample was used for DNA extraction. The DNA and the rest of the fragments of samples were kept at -20°C. All cultures were incubated at room temperature at the LCRH laboratory. The DNA obtained from clinical samples was submitted to PCR amplification of the ITS-5.8S and the V4-V5 16S rRNA gene region, for the detection and identification of fungi and bacteria respectively. From February 2019 till February 2022, 60 patients were studied. Most of them were men (43, 74,1%) between 13 and 78 y.o. (mean age 37). Half of the patients were herdsmen but, among women 40% (6) were housewives and 26.7% (4) charcoal burners. Lesions were mainly located at the feet (87.9%) and most of the patients (54; 93.1%) reported discharge of grains in the exudate, being 27 (46.6%) yellow or pale colored and 19 (32.8%) of them dark grains. Culture of clinical samples yielded 35 fungal and bacterial putative causative agents. Culture and molecular methods allowed the identification of a total of 21 causative agents of mycetoma (39.6% of cases studied). Most of them (17) corresponded to fungi causing eumycetoma (80.9%) being the most prevalent the genus Madurella (7; 41.2%), with two species involved (M. mycetomatis and M. fahalii), followed by Aspergillus (2; 11.8%). Other minority genera detected were Cladosporium, Fusarium, Acremonium, Penicillium, and Trichophyton (5.9% each of them). Actinobacteria were detected in 19.1% of samples, but only Streptomyces somaliensis was identified as a known agent of mycetoma, the rest being actinobacteria not previously described as causative agents of the disease, such as Cellulosimicrobium cellulans detected in two of the patients. Although Kenya is geographically located in the mycetoma belt, to our knowledge this is the first report on mycetoma in this country from 1973, and the first one for Turkana County.

The Domestic Environment and the Lung Mycobiome.

This study analyzes the relationship between the mycobiome of the Lower Respiratory Tract (LRT) and the fungi in the domestic environment. Samples studied consisted of Broncho-Alveolar Lavage (BAL) from 45 patients who underwent bronchoscopy for different diagnostic purposes, and dust and air from the houses (ENV) of 20 of them (44.4%). Additionally, five bronchoscopes (BS) were also analyzed and negative controls were included for every procedure. All samples were processed for DNA extraction and cultures, which were performed in Sabouraud Dextrose and Potato Dextrose Agar. The fungal Internal Transcribed Spacer (ITS2) was sequenced by the Solexa/Illumina system and sequences were analyzed by QIIME 1.8.0 and compared with the UNITE Database for identification. The similarity between the two fungal communities (BAL and ENV) for a specific patient was assessed via the percentage of coincidence in the detection of specific operational taxonomic units (OTUs), and about 75% of co-occurrence was detected between the mycobiome of the LRT and the houses. Cultures confirmed the presence of the core mycobiome species. However, the low rate of isolation from BAL suggests that most of its mycobiome corresponds to non-culturable cells. This likely depends on the patient's immune system activity and inflammatory status.

Fusariosis cutánea por una especie del complejo Fusarium dimerum en una paciente con leucemia mieloblástica aguda / Cutaneous fusariosis by a species of the Fusarium dimerum species complex in a patient with acute mieloblastic leukemia

Antecedentes. La fusariosis es una hialohifomicosis oportunista, emergente, producida por hongos pertenecientes al género Fusarium. Estos hongos pueden provocar enfermedades que amenazan la vida en pacientes inmunodeficientes, en especial en portadores de leucemia. También se ha descrito en individuos inmunocompetentes, en los que induce lesiones localizadas, no invasivas. En pacientes inmunodeficientes, la fusariosis se asocia a una elevada morbimortalidad, sobre todo debido a la falta de sensibilidad de estos hongos a los antimicóticos disponibles. Caso clínico. Describimos el caso de una paciente con leucemia mieloblástica aguda que experimentó una fusariosis por una especie del complejo Fusarium dimerum. El diagnóstico precoz se estableció en función de la observación microscópica de muestras de las lesiones cutáneas y se prescribió tratamiento con voriconazol. Más tarde, un estudio completo del aislamiento fúngico por cultivo y métodos moleculares permitió la identificación de F. dimerum, una especie apenas descrita como patógeno en el ser humano. La sensibilidad de la cepa se examinó con el método comercializado Sensititre YeastOne®, que reveló su sensibilidad a voriconazol y posaconazol, al igual que a anfotericina B. La paciente falleció a los 7 días del ingreso debido a una insuficiencia hemodinámica. Conclusiones. La identificación completa de nuevos aislamientos de Fusarium y su patrón de sensibilidad a los antimicóticos suscita un gran interés para incrementar nuestros conocimientos sobre la epidemiología de la enfermedad y el tratamiento óptimo de los pacientes(AU)

Fusariosis is an emergent opportunistic hyalohyphomycosis produced by fungi belonging to the genus Fusarium. These molds are capable of producing life-threatening diseases in immunocompromised hosts, especially in those suffering from leukemia. It has also been described in immunocompetent patients, where it usually causes non-invasive localized lesions. Fusariosis in immunocompromised individuals has a high morbidity and mortality mainly because of the low sensitivity of these fungi to the antifungal drugs available. Case report. We describe here the case of a patient with acute mieloblastic leukemia who developed fusariosis by a species of the Fusarium dimerum species complex. The early diagnosis was made on the basis of microscopic observation of samples from cutaneous lesions, and voriconazole treatment was prescribed. A subsequent complete study of the fungal isolate by culture and molecular methods allowed the identification of F. dimerum, a species rarely described as a human pathogen. The sensitivity of the strain was tested using the Sensititre YeastOne® commercial system, which showed sensitivity to voriconazole and posaconazole, as well as to amphotericin B. The patient died after 7 days at hospital due to an hemodynamic failure. Conclusions. Complete identification of new isolates of Fusarium and their antifungal susceptibility patterns is of high interest to improve our knowledge about the epidemiology of the disease and how to best manage patients(AU)

Cutaneous fusariosis by a species of the Fusarium dimerum species complex in a patient with acute myeloblastic leukemia.

BACKGROUND: Fusariosis is an emergent opportunistic hyalohyphomycosis produced by fungi belonging to the genus Fusarium. These molds are capable of producing life-threatening diseases in immunocompromised hosts, especially in those suffering from leukemia. It has also been described in immunocompetent patients, where it usually causes non-invasive localized lesions. Fusariosis in immunocompromised individuals has a high morbidity and mortality mainly because of the low sensitivity of these fungi to the antifungal drugs available. CASE REPORT: We describe here the case of a patient with acute mieloblastic leukemia who developed fusariosis by a species of the Fusarium dimerum species complex. The early diagnosis was made on the basis of microscopic observation of samples from cutaneous lesions, and voriconazole treatment was prescribed. A subsequent complete study of the fungal isolate by culture and molecular methods allowed the identification of F. dimerum, a species rarely described as a human pathogen. The sensitivity of the strain was tested using the Sensititre YeastOne(®) commercial system, which showed sensitivity to voriconazole and posaconazole, as well as to amphotericin B. The patient died after 7 days at hospital due to an hemodynamic failure. CONCLUSIONS: Complete identification of new isolates of Fusarium and their antifungal susceptibility patterns is of high interest to improve our knowledge about the epidemiology of the disease and how to best manage patients.

Pseudomonas keratitis 4 years after laser in situ keratomileusis.

PURPOSE: To report a patient who presented an infectious keratitis 4 years after laser in situ keratomileusis (LASIK) without any other predisposing risk factor than the LASIK procedure itself. CASE REPORT: We report a 32-year-old man operated by LASIK in January 2006 who presented with infectious keratitis in the OD in April 2010. Clinical examination showed a corneal abscess at 10-o'clock position in the interface and fibrin and Tyndall 4+ in the anterior chamber. Microbiological analysis identified Pseudomonas aeruginosa as the cause of infection. The patient was given ofloxacin, sulfate neomycin, polymyxin B, and prednisolone acetate to be used every 2 h. Treatment led to clinical improvement with resolution of corneal infiltrate. Keratitis with intact epithelium by Pseudomonas can occur up to 4 years after LASIK. CONCLUSIONS: LASIK treatment is a predisposing factor for bacterial keratitis even years after surgery. This report demonstrates the importance of continued postoperative vigilance by patient and his/her clinician.

Evaluation of molecular diagnosis in fungal keratitis. Ten years of experience.

PURPOSE: The aims of this study were to assess the utility of polymerase chain reaction (PCR) in diagnosing fungal keratitis in the last decade in our center and to review the molecular diagnosis of mycotic keratitis. METHODS: A retrospective nonrandomized investigation was undertaken at Vissum Corporación Instituto Oftalmologico de Alicante to evaluate 27 corneal samples of 20 patients with proven fungal keratitis from January 2000 to December 2009. Corneal samples (21 corneal scrapings, 5 biopsies, and 1 cornea) were evaluated by Gram stain or calcofluor stain, culture, and PCR. The detection and molecular identification were carried out by DNA amplification and sequencing of the internal transcribed spacer and 5.8S rRNA region from the corneal samples. RESULTS: PCR detected all the samples that were positive by conventional methods. Four samples were positive by PCR and showed negative results by culture and stain. Combination of microscopy and culture gave positive results in 21 of the 27 samples of patients with mycotic keratitis. Stains showed a 66.7% of positive results, culture showed 59.3%, and PCR showed 92.6%. The time taken for PCR assay was 4 to 8 h whereas positive fungal cultures took 1 to 35 days. Identification at species level by molecular methods was possible in all cases except one. Identification at species level by conventional methods only was possible in eight cases. CONCLUSIONS: PCR not only proved to be an effective rapid method for the diagnosis of fungal keratitis but was also more sensitive than stain and culture methods. Fungal PCR must be added as the screening diagnosis test when an early mycotic keratitis is suspected. Molecular identification is the gold standard technique for the identification of corneal fungal pathogens.

Microbial keratitis after intrastromal corneal ring segment implantation.

PURPOSE: To evaluate the incidence of acute microbial keratitis after intrastromal corneal ring segment implantation and to investigate whether microbial keratitis is related to the intrastromal corneal ring type or surgical technique. METHODS: Intrastromal corneal ring segments were implanted in 212 eyes of 149 patients. Two different types of intrastromal corneal ring segments were used during the study, Intacs and Ferrara rings, and two surgical techniques were used for intrastromal corneal ring segment implantation, manual and femtosecond laser. RESULTS: One hundred thirty-four eyes (63.2%) underwent Intacs implantation and 78 (36.8%) eyes received Ferrara rings. Corneal tunnels were created using the manual technique in 119 (56.1%) eyes and by femtosecond laser in 93 (43.9%) eyes. Three cases (1.4%) of acute microbial keratitis were clinically identified. Cultures were positive in 2 eyes and negative in 1 eye, and polymerase chain reaction was positive in all 3 cases. The microorganisms isolated from cultures were Streptococcus mitis and Staphylococcus aureus. In 2 cases, the femtosecond laser technique was used (1 eye Intacs, 1 eye Ferrara), and in 1 case, the manual technique was used (Intacs). No statistically significant difference was noted between techniques (P=.582) or segment type (P=1.000). In all cases, intrastromal corneal ring segments were explanted and intensive topical antibiotics were used with clinical success. CONCLUSIONS: Acute microbial keratitis incidence after intrastromal corneal ring segment implantation was 1.4%. Gram-positive cocci were the organisms isolated more frequently. Proper management of this condition requires intrastromal corneal ring segment explantation and intensive topical antibiotics.

Causes of intrastromal corneal ring segment explantation: clinicopathologic correlation analysis.

PURPOSE: To determine the main causes of intrastromal corneal ring segment (ICRS) explantation and the relationship with the microscopic findings on the ICRS surface. SETTING: Vissum Corporation-Instituto Oftalmológico de Alicante, Alicante, Spain. METHODS: This study evaluated ICRS that were explanted in centers in Spain from 2000 to 2008. Clinical data (reasons for explantation, date of implantation/explantation, tunnel creation technique, ICRS type) and scanning electron microscopy findings on the ICRS surface (adherent tissue-like material, cell deposits, protein) were documented. RESULTS: Intrastromal corneal ring segments were explanted from 58 eyes (47 patients). The main cause was extrusion (48.2% of explanted segments), followed by refractive failure (ie, poor refractive outcome) (37.9%), keratitis (6.8%; 3.7% culture positive), and corneal melting and perforation (6.8%). Scanning electron microscopy showed cells and cell debris on the ICRS explanted by extrusion, a clean surface on the ICRS explanted for refractive failure, and bacteria (cocci) in the case of proven infectious keratitis. CONCLUSIONS: The main cause of explantation was extrusion of the ICRS followed by refractive failure. There was a clear correlation between the cause of explantation and the microscopic findings on the ICRS. Extrusion was accompanied by inflammatory cells and cell debris on the ICRS surface. No inflammatory reaction was observed on the ICRS explanted for refractive failure.

Pyrenochaeta keratinophila sp. nov., isolated from an ocular infection in Spain

Se describe e ilustra el nuevo celomicete Pyrenochaeta keratinophila, aislado de un raspado corneal procedente de un caso de queratitis en España. Este hongo se caracteriza morfológicamente por tener colonias de color gris oliváceo a verdoso, setas picnidiales escasas principalmente situadas cerca del ostiolo, y producir conidios fialídicos a partir del micelio aéreo. Este último carácter no se ha observado en ninguna otra especie del género Pyrenochaeta. La secuencia de la región ITS de ADNr de esta cepa clínica confirma nuestra propuesta y pone de manifiesto su estrecha relación con la familia Leptosphaeriaceae(AU)

The new coelomycete Pyrenochaeta keratinophila, isolated from corneal scrapings of a case of keratitis in Spain, is described and illustrated. This fungus is morphologically characterized by grey-olivaceous to greenish olivaceous colonies, scarce pycnidial setae placed mainly near the ostiole and production of phialoconidia from the aerial mycelium. The latter feature is unknown in any other species of the genus Pyrenochaeta. Sequencing of the ITS rDNA region of this clinical strain confirmed this proposal and revealed its close genetic relationship with the Leptosphaeriaceae

Pyrenochaeta keratinophila sp. nov., isolated from an ocular infection in Spain.

The new coelomycete Pyrenochaeta keratinophila, isolated from corneal scrapings of a case of keratitis in Spain, is described and illustrated. This fungus is morphologically characterized by grey-olivaceous to greenish olivaceous colonies, scarce pycnidial setae placed mainly near the ostiole and production of phialoconidia from the aerial mycelium. The latter feature is unknown in any other species of the genus Pyrenochaeta. Sequencing of the ITS rDNA region of this clinical strain confirmed this proposal and revealed its close genetic relationship with the Leptosphaeriaceae.

Scanning electron microscopy analysis of a sputnik-like intraocular lens 28 years after implantation.

PURPOSE: To report a pupil-supported, iris-clip intraocular lens (IOL) that was explanted more than 28 years after implantation. METHODS: A pupil-supported, iris-clip, Sputnik-like IOL was implanted in the left eye of a 33-year-old man to correct aphakia after extracapsular cataract extraction due to trauma. RESULTS: Twenty-eight years after implantation, the patient was referred to our center with loss of vision. Clinical examination showed dislocation of the IOL, which was subsequently explanted. Scanning electron microscopic examination showed a transparent, polymethylmethacrylate (PMMA), pupil-supported, iris-clip IOL with melanosomes and cell deposits (foreign-body reaction) on its surface. CONCLUSIONS: This case demonstrates the inertness of PMMA material and reports that a foreign body reaction can be induced following IOL dislocation 28 years after implantation.

Molecular epidemiology of isolates of the Cryptococcus neoformans species complex from Spain.

To study genetic diversity of Cryptococcus neoformans species complex in Spain, 97 isolates of the yeast recovered from human, animal and environmental samples have been analysed using three molecular epidemiological techniques. One of these, URA5 gene fragment length polymorphism (RFLP) analysis, has been previously described as a molecular epidemiology tool. Thus, standard profiles and reference strains have been defined for it. In addition, 5S rDNA/IGS RFLP and [GACA](4) microsatellite PCR fingerprinting were also used. Our results show five of the previously defined URA5 genotypes with a high frequency (33%) of the VNI type, which is in concordance with other studies. The high presence of VNIII pattern (28.9%) among our strains is remarkable and could be a specific feature of the isolates from our country. 5S rDNA/IGS RFLP showed a low intra-species discriminative power. Three different molecular profiles (S1-3), which showed a good correlation with the different species, varieties and genotypes, were obtained. [GACA](4) microsatellite PCR-fingerprinting analysis showed a high variability of patterns among the studied strains. Molecular profiles represented in a dendrogram clustered strains in four main groups related with the source of the yeast and also in concordance with some of the described genotypes (VNI-IV and VGI).

Molecular epidemiology of isolates of the Cryptococcus neoformans species complex from Spain

Para ampliar el conocimiento de la diversidad genética que presenta el complejo Cryptococcus neoformansen España, se analizaron 97 aislamientos de esta levadura obtenidos a partir de muestras ambientales, veterinariasy clínicas, utilizando 3 técnicas epidemiológicas moleculares diferentes. Uno de estos marcadoresmoleculares, el análisis del URA5 RFLP, se ha descrito previamente como herramienta epidemiológica,por lo que se dispone de perfiles estándar y cepas de referencia. Además, se utilizaron también el análisisdel RFLP del 5S rDNA/IGS y la huella digital tras amplificación por PCR de la secuencia microsatélite[GACA]4. Nuestros resultados muestran 5 de los genotipos URA5 con una elevada frecuencia del tipo VNI(33%), lo que concuerda con otros estudios. Es de resaltar la elevada presencia del perfil VNIII entre nuestrascepas (28,9%), lo que podría ser el rasgo específico más destacable de los aislamientos de nuestro país.El marcador 5S rDNA/IGS mostró un bajo poder discriminativo intraespecie. Se obtuvieron 3 perfiles molecularesdistintos (S1-3), que presentaron una buena correlación con las especies, variedades y genotipos. Laobtención de perfiles de huella digital con [GACA]4, presentó una gran variabilidad entre las cepas estudiadas.La representación en dendrograma agrupó las cepas en 4 agrupamientos principales relacionados conel origen de las levaduras y también con cierta concordancia con los genotipos descritos (VNI-IV y VGI)(AU)

To study genetic diversity of Cryptococcus neoformans species complex in Spain, 97 isolates of the yeastrecovered from human, animal and environmental samples have been analysed using three molecularepidemiological techniques. One of these, URA5 gene fragment length polymorphism (RFLP) analysis, hasbeen previously described as a molecular epidemiology tool. Thus, standard profiles and reference strainshave been defined for it. In addition, 5S rDNA/IGS RFLP and [GACA]4 microsatellite PCR fingerprinting werealso used. Our results show five of the previously defined URA5 genotypes with a high frequency (33%) ofthe VNI type, which is in concordance with other studies. The high presence of VNIII pattern (28.9%) amongour strains is remarkable and could be a specific feature of the isolates from our country. 5S rDNA/IGS RFLPshowed a low intra-species discriminative power. Three different molecular profiles (S1-3), which showeda good correlation with the different species, varieties and genotypes, were obtained. [GACA]4 microsatellitePCR-fingerprinting analysis showed a high variability of patterns among the studied strains. Molecularprofiles represented in a dendrogram clustered strains in four main groups related with the source of theyeast and also in concordance with some of the described genotypes (VNI-IV and VGI)(AU)

New pyrenochaeta species causing keratitis.

We report a new fungus as an agent of fungal keratitis in a diabetic woman. The fungal etiology was established by classic microbiology and PCR following 3 months of antibacterial therapy. The morphological features of the isolate and sequence analysis of the internal transcribed spacer region indicate a new species of Pyrenochaeta (Coelomycetes).

Laboratory diagnosis of endophthalmitis: comparison of microbiology and molecular methods in the European Society of Cataract & Refractive Surgeons multicenter study and susceptibility testing.

PURPOSE: To investigate and compare the use of molecular biology with the use of traditional Gram stain and organism culture for the laboratory diagnosis of postoperative endophthalmitis. SETTING: Twenty-four ophthalmology units together with 9 microbiology laboratories and 2 European reference molecular biology laboratories. METHODS: A prospective randomized partially masked multicenter cataract surgery study recruited 16 603 patients. This resulted in 29 cases of presumed postoperative endophthalmitis. Gram stain and culture were performed in the local laboratory according to agreed protocols. Samples of aqueous and/or vitreous were transported to the first referenced molecular biology laboratory (Regensburg, Germany) for polymerase chain reaction (PCR) testing, and an extracted aliquot of DNA was then referred to the second laboratory (Alicante, Spain) for PCR. RESULTS: Of the 29 who presented with presumed postoperative endophthalmitis, 20 were classified as proven infective endophthalmitis with positive Gram stain, culture, or PCR. Fourteen patients were culture-positive; all but 1 of these was also positive by PCR. Six patients were positive by PCR but negative by Gram stain or culture. Nine patients were negative by both microbiology and PCR testing. CONCLUSIONS: Use of molecular biology technique increased the laboratory rate of identifying the pathogen by 20%, confirming the technique is very useful for the endophthalmitis specimen. Samples of both aqueous and vitreous should be collected and stored at -20 degrees C for PCR at the time of the diagnostic taps.

Polymerase chain reaction and DNA typing for diagnosis of infectious crystalline keratopathy.

A 63-year-old man developed infectious crystalline keratopathy (ICK) in his right eye 1 year after phacoemulsification. The white peripheral lesion was adjacent to the corneal phacoemulsification incision. Infiltrates in the form of creamy-white, midstromal branching, needle-like opacities without evidence of inflammatory cells were noted. A corneal biopsy by double lamellar flap was done and studied by 3 techniques: microbiological culture, stain, and polymerase chain reaction (PCR). Fungal and bacterial PCR were positive. A second sample was necessary to obtain a positive stain and culture. The DNA sequence analysis showed Candida parapsilosis and Staphylococcus aureus as the causal agents of the crystalline keratopathy. Treatment was started with amphotericin B 1% and cefazolin 6 times a day, and systemic voriconazole was recommended. This is the first reported case of ICK after cataract surgery. Polymerase chain reaction amplification and subsequent DNA typing were useful tools in detecting and identifying the ocular pathogens involved in this case.

[Molecular biology in the diagnosis of deep-seated candidiasis in the critically ill non-neutropenic patient]. / Biología molecular en el diagnóstico de la candidiasis profunda en el paciente crítico no neutropénico.

Candida infections are an important cause of morbidity and mortality in critically ill patients. Rapid detection of the yeast in blood and other tissues by molecular biology methods has been the goal of some recent studies. An analysis of the sensitivity and specificity of these methods assayed in clinical specimens from critically ill and other patients is carried out. PCR amplification of ribosomal genes and their internal spacers showed a higher sensitivity than culture based methods. A standardization of most of the methodological steps in molecular methods is needed. Real time PCR with fluorescent probes seems to be the most interesting proposal. It has the advantage of the possible quantification of fungal presence in tissues and minimizes the samples' contamination risk.

Biología molecular en el diagnóstico de la candidiasis profunda en el paciente crítico no neutropénico / Molecular biology in the diagnosis of deep-seated candidiasis in the critically ill non-neutropenic patient

Las infecciones por Candida son una importante causa de mortalidad y morbilidaden pacientes críticos. La detección rápida de la presencia de la levadura ensangre y otros tejidos es un objetivo que recientemente se intenta abordar aplicandodistintos métodos moleculares de diagnóstico. Analizamos la sensibilidady especificidad de los métodos aplicados a la detección de Candida en muestrasclínicas de pacientes críticos y otros susceptibles de padecer candidiasis profunda.Los métodos basados en la reacción en cadena de la polimerasa (PCR),especialmente los que tienen como diana las secuencias de los genes ribosomalesy sus espaciadores internos, muestran claramente una mayor sensibilidadque el cultivo y una especificidad equiparable a este. Todavía no hay propuestasclaras en cuanto a la posible estandarización de la extracción y purificación delDNA y de los sistemas de lectura de productos, aunque la técnica de PCR atiempo real con emisión de fluorescencia, parece la propuesta más interesantepor la ventaja añadida de permitir la cuantificación de la carga fúngica y minimizarla manipulación de las muestras y por tanto el riesgo de falsos positivos

Candida infections are an important cause of morbidity and mortality in criticallyill patients. Rapid detection of the yeast in blood and other tissues by molecularbiology methods has been the goal of some recent studies. An analysis of thesensitivity and specificity of these methods assayed in clinical specimens fromcritically ill and other patients is carried out. PCR amplification of ribosomalgenes and their internal spacers showed a higher sensitivity than culture basedmethods. A standardization of most of the methodological steps in molecularmethods is needed. Real time PCR with fluorescent probes seems to be themost interesting proposal. It has the advantage of the possible quantification offungal presence in tissues and minimizes the samples’ contamination risk