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Summary. This study was carried out to evaluate the utility of serological tests in the diagnosis of candidal infections in non-neutropenic critically ill patients. A prospective study was carried out in a 10-bed general intensive care unit; all patients with at least one organic sample with Candida spp. were included. A therapeutic–diagnostic algorithm was designed, and patients were treated or not, according to a classification. Blood samples were taken, and serological tests included: antigenaemia detection using two different commercial latex kits (Cand-Tec® and Pastorex®) and antibody detection by indirect haemagglutination (IHA) and indirect germ tube immunofluorescence (IFA). A total of 56% of antibody tests (IHA 45% and IFA 64%) and 26% of antigen tests (Cand-Tec 36% and Pastorex 17%) were positive. The sensitivity and specificity of these tests with respect to systemic candidosis were 37% and 78%, respectively, for antibodies, and 0% and 90% for antigens. There was statistical significance for mortality and low levels of antibodies; Candida glabrata was detected by IFA and Candida tropicalis by Cand-Tec. Serological tests may help to define the prognosis of these patients and to support the detection of specific Candida species. [ABSTRACT FROM AUTHOR]

Background: The importance of hypersensitivity to fungal allergens is a relatively unknown and somewhat controversial subject., Methods: An open prospective study was carried out in just one centre to determine the clinical and epidemiological characteristics as well as the diagnostic usefulness of skin prick and conjunctiva provocation tests, associated with total and specific IgE determination in two groups of patients, one of which was monosensitised to fungi and the other of which had multiple sensitisations, including fungi., Results: Rhinitis, exclusive or associated with asthma, was the main consultation cause (88% in monosensitised patients). Severe asthma was rarely found. In the polysensitised group, 64% were simultaneously allergic to moulds and mites. Alternaria alternata was the most common sensitising fungus, although a considerable number of cases were associated with other species such as Cladosporium, Penicillium and/or Aspergillus. The skin prick test gave the highest sensitivity and specificity. In 67% of the cases, the specific IgE was found between classes 3 and 4. The conjunctival provocation test was an innocuous and highly useful method for verifying the diagnosis and determining the degree of clinical sensitisation. A large number of patients exclusively allergic to fungi received specific immunotherapy, and it was generally well tolerated., Conclusions: This protocolised study shows the importance of Alternaria and other fungi sensitisations in rhinitis alone or associated with asthma. Combined diagnosis of prick test, specific IgE and conjunctiva provocation test is very useful for deciding specific immunotherapy., (Copyright © 2011 SEICAP. Published by Elsevier Espana. All rights reserved.)

Background: Yeasts represent the second cause of nail fungal infection in the world, and Candida albicans and Candida parapsilosis are the two most common species., Objectives: To determine the yeast species frequency and their in vitro antifungal susceptibility, obtained from patients with clinical features suggestive of onychomycosis., Methods: A prospective study was carried out in four dermatological care centers in Mexico from 2004 to 2007. Clinical diagnosis was corroborated by direct examination and culture. The yeast species was determined by morphological and biochemical tests. An antifungal susceptibility test to ketoconazole, itraconazole and fluconazole by the broth microdilution method was performed on each isolate (document M27-A2)., Results: One hundred sixty-six yeast isolates were obtained; the most frequently found species were C. parapsilosis (31.9%), C. albicans (22.4%) and Candida guilliermondii (12.7%). Of all isolates, 51 showed resistance to one or several of the azole compounds: 33 to itraconazole, 12 to ketoconazole and 6 to fluconazole. It was remarkable that the four Candida glabrata isolates were resistant to the three azole compounds; C. guilliermondii and Candida famata were resistant to itraconazole in 42.9% and 54.5%, respectively., Conclusion: The results obtained show the importance of identifying the aetiological agent and antifungal susceptibility testing in order to avoid therapeutic failures in onychomycosis., (Copyright © 2010 Revista Iberoamericana de Micología. Published by Elsevier Espana. All rights reserved.)

Mueller-Hinton modified agar (MH-GMB) was compared with RPMI + 2% glucose-agar to determine the MICs of 80 isolates of Cryptococcus neoformans and C. gattii to posaconazole with Etest. MH-GMB minimised trailing and agreement between both media was 94%. Agreement of M27-A2 microbroth reference method was 98% with RPMI and 94% with MB-GMB.

We report a case of a lung reactivation of a latent histoplasmosis in a 68-year-old patient without immunologic dysfunction living in Barcelona (Spain). The Histoplasma capsulatum var. capsulatum infection was probably acquired in a previous stay in Equatorial Guinea 45 years before. Diagnosis of mild chronic pulmonary histoplasmosis was performed by histopathology of lung biopsy plus antibodies detection against H. capsulatum. Treatment with oral itraconazol during three months leaded to a complete clinical cure, and antibodies disappeared after four years of follow up.

Objective: To evaluate the prevalence of tinea capitis, tinea pedis, and tinea unguium in children from several schools of Barcelona city., Methods: During the period of 2003-2004, a prospective cross-sectional study was carried out in 1,305 children (9% immigrant population) between the ages 3 and 15 in 17 schools in Barcelona. A systematic examination of the feet, (including nails and scalp), was performed to identify lesions compatible with tinea. Cultures of scalp and feet samples were done and analysis of environmental samples was performed for dermatophyte isolation., Results: Dermatophytes were isolated in 2.9% of the samples with a prevalence of 2.5% in feet, 0.23% in scalp, and 0.15% in nails of the feet. The predominant etiologic agents in feet were Trichophyton mentagrophytes in 45.7% of the cases and Trichophyton rubrum in 31.4%. In the nails, T. rubrum and Trichophyton tonsurans were isolated, while T. mentagrophytes (2 cases) and Trichophyton violaceum (1 case) were identified in scalp samples. Forty-five per cent of dermatophytes were isolated from healthy feet, the majority of cases in children 13- 15-years-old (p < 0.05). Microsporum gypseum was the only agent identified in the environmental samples, and was also found in one of the cases of tinea pedis., Conclusion: The results of this study demonstrate a low prevalence of tinea capitis and tinea unguium in school children of Barcelona. On the contrary, high prevalence of dermatophytes in feet was found. It highlights the high prevalence of healthy carriers of dermatophytes in feet.

The in vitro susceptibility of 62 isolates of Sporothrix schenckii in its mycelial form, from Latin-American countries (Peru, Venezuela, Brazil and Uruguay) and Spain, to amphotericin B (AB), itraconazole (IZ), posaconazole (PZ) and terbinafine (TB) was determined by measuring the MICs and minimum fungicidal concentrations (MFCs) using a standardized Clinical and Laboratory Standards Institute method. In general, TB was the most active drug, with the lowest geometric mean (GM) MIC and MFC values amongst isolates from the five countries tested. IZ and PZ showed almost the same activity against all strains tested, except for isolates from Uruguay where IZ gave the highest GM MIC (10.68 mg l(-1)). AB showed the widest MIC range (0.03-16.0 mg l(-1)); however, this drug was less active against 79 % of isolates (MICs above 1 mg l(-1)). MFCs were 5 to 20 times higher than the MICs, but the lowest GM MFC and range values were found for TB. IZ and PZ gave the highest GM MFC. MFC may be a better predictor of therapeutic response than MIC, especially in immunosuppressed patients, making the use of IZ and PZ an inappropriate treatment. There were some differences in susceptibility according to the geographical source of the isolates, with the MIC being lower for TB in Venezuelan strains (P=0.066) and the MFC higher for PZ in Peruvian strains (P=0.02). Thus, geographical origin may be important for appropriate treatment, and may relate to the identification of species of the S. schenckii complex.

Histoplasmosis caused by Histoplasma capsulatum var. duboisii is an endemic mycosis of sub-Saharan Africa that usually affects the skin, subcutaneous tissue, lymph nodes and bones. We present a case of a 10-year-old immunocompetent girl with severe cutaneous and subcutaneous abscesses affecting the head and upper body. Microscopic examination showed polar budding yeasts and short mycelium compatible with H. capsulatum var. duboisii. Cultures were not possible but serology showed antibodies against both H. capsulatum var. duboisii and H. capsulatum var. capsulatum antigens. Presumptive diagnosis of histoplasmosis was done but treatment with itraconazole was inefficacious. After 15 days of treatment with Amphotericin B i/v, improvement was evident and, three months later, the patient was discharged with only residual lesions. Seven months later, no relapses were observed.

Background: Candida is frequently seen in urinalysis studies in patients with diabetes mellitus. The objective was to determine the presence and clinical significance of candiduria, and to identify the different isolated Candida species and their in vitro susceptibility pattern to different antifungal agents by means of the broth microdilution method., Methods: We studied the urine from 50 type 2 diabetes mellitus (DMT2) patients. 24 patients had controlled DMT2 and 26 non-controlled DMT2., Results: Twenty-three Candida spp. positive cultures were obtained, of which 17 were obtained from the non-controlled DMT2 patients; 30.7% of the isolates were caused by Candida infection. The main isolated species were C. glabrata (48%) and C. albicans (35%). Itraconazole, amphotericine B, and ketoconazole showed less antifungal activity in C. glabrata isolates, whereas fluconazole and voriconazole displayed higher antifungal activity., Conclusions: It is important to search routinely for yeast in the urine of DMT2 patients to detect candidiasis, and to perform antifungal susceptibility tests to Candida isolates in order to establish antifungal therapy for these patients.

Cryptococcosis is a life-threatening infection in humans and animals caused by encapsulated yeasts of the genus Cryptococcus. Cryptococcus neoformans and Cryptococcus gattii are the main agents of this mycosis. Until 2002 C. gattii was classified as a variety of C. neoformans but now is accepted as an independent species. The laccase (phenoloxydase) enzyme produced by these yeasts is considered one of the main pathogenic factors for its ability to induce melanin from dihydroxyphenolic compounds. The vast majority of the studies in laccase and melanin synthesis have been developed using isolates of C. neoformans. The main objective of this study was to evaluate laccase activity in strains of C. gattii, serotype B isolated from immunocompetent goats that died of lung and disseminated cryptococcosis, in several outbreaks occurring in Spain. The laccase activities of these isolates were compared with those of other strains of C. gattii and C. neoformans. After fungal cell rupture, the supernatant of each isolate was analyzed for its laccase activity using as substrate an L-dopa 20 mM solution. The degree of enzymatic activity was assessed according to its absorbance at 450 nm and scored using Enzymatic Units (EU). The maximum values were observed in three strains of C. gattii from goats (EU > 12). The smallest values were observed in one environmental isolate of C. gattii serotype C (EU = 0.7). The highest recorded value for C. neoformans was 6.3 EU in a serotype A isolate from one human case of meningitis. C. gattii serotype B obtained from goats showed different degrees of laccase activity, being the highest in those isolated from severe outbreaks of cryptococcosis. This enzyme appears to represent a major, though nonexclusive, pathogenic factor for Cryptococcus gattii.

Urease is an enzyme considered one of the main virulence factors in Cryptococcus neoformans. Quantitative differences in urease production between C. neoformans and the new species Cryptococcus gattii have not been so far documented. Using a standardized method, 25 isolates of C. neoformans and 19 of C. gattii were seeded in Christensen urea broth medium for urease activity detection. Approximately, the 50% of activity of one unit of commercial jack beans urease (A550=0.215) was considered as a reference to classified the Cryptococcus in two cathegories, low (A550<0.215) or high (A550=or>0.215) urease producers. After 72 hours of incubation, 76% of C. neoformans and 15.8% of C. gattii strains were high urease producers (p=0.016). Based on these results, the species C. neoformans appeared as the highest urease producer. Other virulence factors should also be investigated to explain C. gattii pathogenicity.

Since the foundation of the "Asociación Española de Micología" thirty years ago, we have consolidated the Spanish mycological community and also witnessed remarkable changes, not only in the Spanish society but in the scientific community as a whole. As it usually happens to human beings, during this time the "Asociación Española de Micología" has matured transforming itself into a solid scientific society. However, the "Asociación Española de Micología" will have to continue its transformation to adapt to new changes. This article shows the most relevant aspects in the history of the "Asociación Española de Micología" as well as new challenges that the society might face in the future.

Objectives: The microbroth ATB FUNGUS 2 (ATBF2) method (bioMérieux, La Balme-les Grottes, France), designed for in vitro determination of the susceptibility of Candida spp. and Cryptococcus neoformans to antifungal agents, was evaluated with 100 yeasts and compared with Sensititre Yeast One (SYO; Trek Diagnostic Systems, UK), considering CLSI M27-A2 as the reference method., Methods: ATBF2 consists of ready-to-use strips including amphotericin B (AMB), 5-flucytosine, fluconazole and itraconazole for MIC determinations. Reproducibility of ATBF2 was determined. Two quality control strains and a panel of eight Candida isolates were tested five different times with the three methods. The essential agreements within +/-2 log2 dilution between the ATBF2, SYO and M27-A2 methods were assessed. The yeast clinical isolates included were nine species of Candida (n = 80) and C. neoformans (n = 20)., Results: Inter- and intra-laboratory reproducibility, tested with the Candida panel, was >or=99%. MICs for the ATCC strains were within the expected ranges with the three methods. Visual and automated readings of ATBF2 presented good concordance, being lower with itraconazole. The overall essential agreements with the M27-A2 method were 94% and 99% for automated ATBF2 and visual ATBF2 readings, respectively. For SYO, the agreement was 91%. Percentages of agreements by drugs (automated ATBF2/visual ATBF2/SYO) were: 5-flucytosine, 97/100/90; AMB, 97/100/85; fluconazole, 93/97/95; and itraconazole, 89/98/95. Disagreement was higher between M27-A2 and SYO than between M27-A2 and ATBF2., Conclusions: ATBF2 is an objective, reproducible and simple method for the accurate determination of MICs of the most common antifungal drugs in yeasts.

The in vitro susceptibility of Sporothrix schenckii to antifungal drugs has been determined with three different methods. Nineteen Peruvian clinical isolates of S. schenckii were tested against amphotericin B (AB), flucytosine (FC), fluconazole (FZ), itraconazole (IZ), voriconazole (VZ), and ketoconazole (KZ). Modified NCCLS M38-A, Sensititre YeastOne (SYO), and ATB Fungus 2 (ATBF2) methods were used to determine the MICs. ATCC isolates of Candida parapsilosis, Candida krusei, and Aspergillus flavus were used for quality control. Sporothrix inocula were prepared with the mycelial form growing on potato dextrose agar at 28 +/- 2 degrees C. MICs of AB, FC, FZ, and IZ were determined with all three methods, VZ with M38-A and SYO, and KZ with only SYO. The three methods showed high MICs of FZ and FC (MIC(90) of 0.5 microg/ml), being homogeneously lower than those of IZ and KZ. The M38-A method showed a variable MIC range of VZ (4.0 to 16 microg/ml); the geometric mean (GM) was 9.3 mug/ml. The MIC range of AB was wide (0.06 to 16 microg/ml), but the GM was 1.2 microg/ml, suggesting that the MIC is strain dependent. Agreement (two log(2) dilutions) between commercial techniques and the modified M38-A method was very high with FZ, IZ, and FC. In AB and VZ, the agreement was lower, being related to the antifungal concentrations of each method. The highest activity against S. schenckii was found with IZ and KZ. Lack of activity was observed with FZ, VZ, and FC. When AB is indicated for sporotrichosis, the susceptibility of the strain must be analyzed. Commercial quantitative antifungal methods have a limited usefulness in S. schenckii.

Environmental fungi, moulds and yeasts could reach the nasal cavity with the inhaled air causing respiratory symptoms in atopic subjects, but little is known about the fungal flora of this site. In the present study samples of the nasal cavities of 135 subjects aged 18-35 years (48 allergic patients to fungi, mites and/or cat fur and from 87 normal subjects--healthy, control group) were cultured. All of them lived in the metropolitan area of Barcelona. Fungi were isolated from 41.5% of healthy people and in 14.8% of allergy patients (p = 0.011). Morphologically, 50.4% of the isolates were located within 4 genera: Cladosporium, Penicillium, Aspergillus and Alternaria, fungi which are considered the most allergenic. The most prevalent species were: Cladosporium herbarum and C. cladosporioides (23.6%). Alternaria alternata was isolated only in 8.8% of samples from the allergic group, although most subjects were sensitive to this species. There were not differences in the isolation rate between genera and smoking-no-smoking groups. The lower prevalence of nasal fungi from allergic patients could be related to the nasal insufficiency, the hypersecretion and the larger use of handkerchiefs.

We report a case of proximal fold cellulitis in both big toes, associated with a bilateral proximal onychomycosis and an intertrigo of the fourth space due to Fusarium solani. The infection occurred in an immunocompetent man with diabetes mellitus type II. Apparently, the infection was acquired in a tropical country and once the patient was in Spain the infection progressed causing nail detachment (onychomadesis). Seven months later a relapse that affected the left toenail occurred. The patient was treated topically with chemical toenail avulsion contained 40% urea associated with bifonazole followed by ciclopirox-olamine nail lacquer for 12 months. Complete cure without relapse was observed after 10 years of follow-up. In vitro antifungal susceptibility study demonstrated that two of the recovered isolates were both resistant to itraconazole and voriconazole.

Background: Allergy to airborne fungi can cause rhinitis and severe asthma, hence the exposure to spores inside home is an important factor of sensitization. The aim of this study was to determine the distribution and prevalence of species of Alternaria, Aspergillus, Cladosporium and Penicillium inside and outside of homes of patients allergic to fungi and to evaluate seasonal variations., Methods: Air samples were collected in 22 selected homes of patients with allergy to fungi using a volumetric method of impacting plates with culture media. The isolated species were identified and statistical analysis of the presence of the four fungi was carried out., Results: A total of 431 indoor and 150 outdoor exposed plates were cultured, leading to isolation of 11,843 colonies of fungi (range 0- 1 666 colony-forming units per cubic meter (CFUs/m(3)). 85.5% of total colonies belonged to the four genera considered. The highest presence of Aspergillus, Cladosporium and Penicillium in indoor environment was registered in autumn. Alternaria was more frequent in summer. In the outdoor environment, Penicillium was more abundant in winter and Aspergillus in summer (P= .002). The largest numbers of isolations were of Cladosporium and Penicillium during all four seasons, indoors as well as outdoors. Alternaria was present in all the homes studied both in summer and in autumn. The most prevalent species were: Alternaria alternata, Cladosporium herbarum, Cladosporium cladosporioides, Aspergillus niger and Penicillium chrysogenum., Conclusions: The quantitative analysis of the four taxa related with respiratory allergies demonstrated considerable seasonal variability. Statistical differences between the indoor and outdoor prevalence were detected only in Alternaria. In summer and autumn, the greater level of exposure to the four studied taxas occurred inside homes.

Minimal inhibitory concentrations (MIC) and minimal fungicidal activity of albaconazole, voriconazole and fluconazole against 55 strains of Cryptococcus gattii, clinically or environmentally isolated in Spain and some Latin American countries, were assessed. By means of the microbroth method (National Committee for Clinical Laboratory Standards; document M27-A2), the geometric mean value for fluconazole was 5.01 microg/ml; however, MIC for 12.7% of isolates ranged from 16 to 32 microg/ml, suggesting increased resistance against fluconazole. Geometric mean values of 0.02 and 0.03 microg/ml for albaconazole and voriconazole, respectively, were found, indicating not only a higher susceptibility to these new azoles but also a better performance of albaconazole (P = 0.003). Minimal fungicidal concentrations were also very low for albaconazole and voriconazole (P<0.001; geometric mean values of 0.023 microg/ml and 0.07 microg/ml, respectively). Both azoles may be good alternatives for the treatment of C. gattii cryptococcosis.

Cryptococcus neoformans var. grubii (serotype A) was isolated from 12 soil samples mixed with pigeon droppings (16.9%) from 71 soil samples in Barcelona and rural areas of Catalonia. C. neoformans was not isolated from indoor dust and Eucalyptus debris. PCR fingerprinting was performed in 22 representative isolates and all of them corresponded to the VNI pattern. Susceptibility testing for the 22 isolates of C. neoformans var. grubii showed that all of them were susceptible to amphotericin B. Three isolates presented MICs (Minimal Inhibitory Concentrations) > or = 1 microg/ml to Itraconazole, five MICs > or = 1 microg/ml to ketoconazole and four were fluconazole resistant, (MICs > or = 64 microg/ml), while three of them were shown to have MICs > or = 1 microg/ml to voriconazole. In spite that all isolates presented the same DNA fingerprinting pattern, the susceptibility to antifungals is very variable. The possibility of acquiring cryptococcosis infection with primarily resistant environment strains is feasible.

We report the first case of human cryptococcosis due to Cryptococcus neoformans var. gattii described in our country, which was presented as brain cryptococcoma in an immunocompetent patient. An extensive sampling of the patient's environment was carried out to find the source of infection.

The interest on the in vitro susceptibility to itraconazole has recently increased due the availability of the intravenous formulation. In this study, comparative MICs of this antifungal with voriconazole were carried out in 62 clinical isolates of filamentous fungi and 100 yeasts isolates using the NCCLS microbroth methods described in M38-A and M27-A2 documents. A MIC90 of 0.125 micrograms per ml was observed for itraconazole and voriconazole against Aspergillus fumigatus. Higher susceptibility to itraconazole was found for the filamentous form of Sporotrhix schenckii (p = 0.001). Voriconazole was more effective against Scedosporium apiospermium while Scedosporium prolificans isolates were resistant to both azoles. Some isolates of Rhizopus stolonifer were susceptible to itraconazole and resistant to voriconazole, but without statistical significance. Susceptibility of nine species of Candida was similar for both triazoles used in this study. However, Candida glabrata was more susceptible to voriconazole. Some fluconazole-resistant Candida albicans isolates were susceptible to itraconazole and / or voriconazole. Cryptococcus neoformans was more susceptible to itraconazole than to voriconazole. Itraconazole and voriconazole showed very close in vitro activity against the tested fungal isolated, except against S. schenckii. In spite of this, there were some differences in susceptibility among isolates within the same fungal species.

Objective: To analyze the epidemiological and clinical characteristics of Histoplasma capsulatum infection in a group of volunteers who had traveled to Guatemala., Methods: A review was performed of the clinical records of nine patients who consulted at the Tropical Medicine Outpatient Unit of Hospital Clinic in Barcelona, Spain. RESULTS. All patients were attended for travelers' diarrhea. In addition, one student had also presented with respiratory symptoms and fever while in Guatemala and her chest X-ray showed interstitial infiltrates. The epidemiological history recognized risk factors for histoplasmosis in all patients (all of them had been working in the reconstruction of an old school building). The intradermal histoplasmin test was performed and was positive in all nine patients. None of the students required treatment., Conclusions: Histoplasmosis should be included in the differential diagnosis of returning travelers' fever. The diagnosis would have been missed in all of our patients if not for a directed clinical history taking. It is important to reinforce prevention measures, e.g. the use of face masks, when working in an environment with a risk for acquiring histoplasmosis.

Objective: To determine a method for the early diagnosis of candidiasis in non-neutropenic critically ill patients in order to reduce mortality., Methods: A prospective study in non-neutropenic critically patients in whom Candida spp. were detected, was made in an intensive care unit (ICU) during an 8-year period from 3389 patients admitted. A diagnostic and therapeutic protocol was designed. Invasive candidiasis was defined according to dissemination and multifocality., Results: Candida spp. were found in 145 cases (4.3%): 120 (83%) were considered as invasive candidiasis and 25 as colonisation (17%). The hospital mortality was 46% (67/145). A post-mortem study was carried out in 54% (36/67) of hospital deaths. Candida albicans was the most frequently isolated species (87%), followed by Candida glabrata (18%). There were 24 candidemias and three cases of endophtalmitis. Digestive and respiratory samples and non-C. albicans yeasts were risk factors for invasive candidiasis. The mortality rate was related statistically to invasive candidiasis and inversely to the appropriate antifungal treatment., Conclusions: Invasive candidiasis is related to digestive and respiratory samples and to the presence of non-C. albicans species. A simpler definition of invasive candidiasis in non-neutropenic critically ill patients will permit more rapid and accurate specific antifungal therapy.

Basis: Description of a situation of incidence increase of bronchial secretions with positive cultures for Aspergillus fumigatus, and analysis of the related risk factors in the invasive aspergillosis., Methods: Between January 1999 and February 2000, a prospective study of the patients was conducted with culture of bronchial secretions and with positive result for A. fumigatus., Variables Studied: age, sex, primary diagnosis, type of cultivated sample, clinical interpretation (colonization/infection), probable source (community/nosocomial), situation of the patient after discharge, and risk factors for opportunistic infection. The results were compared among the colonized and infected patients., Results: Fifty-two patients showed positive cultures of bronchial secretions to A. fumigatus, 43 (82.6%) colonized and 9 (17.3%) infected. Cultivated sputum sample on 30 occasions (57.6%) and bronchial aspiration in 22 (42.3%). Median age: 70 years (31-84). Sex: 40 men (76.9%). Probable source of infection/colonization: nosocomial in 18 cases (34.6%), community in 3 (5.7%) and unknown in 31 (59.6%). Mortality: 15 patient colonized (34.8%) and 8 infected (88.8%). Risk factors with statistical significance for invasive infection by A. fumigatus: diagnosis of chronic bronchopathy (COPD) (p=0.007) and treatment with prednisone in dose higher than 60 mg/day (p=0.0005)., Conclusions: The patients with positive culture of bronchial secretions to A. fumigatus with COPD and treatment with prednisone in dose higher than 60 mg/day should be considered with a greater risk for infection by this pathogen. A more restricted use and adequate of the corticoids in these patients, and an early diagnosis and treatment in light of the suspicion of infection by A. fumigatus in patients with COPD, it could imply a reduction of morbidity and mortality.

The pathogenicity of two different genomic profiles of Cryptococcus neoformans var. gattii serotype B isolated from goats that died from cryptococcal pneumonia was assessed in an experimental model of immunocompetent mice. One strain of each randomly amplified polymorphic DNA (RAPD) profile (GR52 and GR56) and three reference C. neoformans isolates representing serotypes B, D and C were used. BALB/c male mice were inoculated by the intraperitoneal route with each strain. After 4 weeks of follow-up, the animals were sacrificed and autopsy specimens of testes, liver, spleen, kidney, lungs and brain were cultured and stained for histopathology. Although spontaneous mortality was only 2% (one animal), all mice except for those inoculated with serotype C showed positive cultures in almost one organ. The strain GR52 isolated from goat showed the highest rate of positive cultures (80%) followed by serotype D (77%). Serotype B reference strain and second goat strain GR56 were both isolated from 70% of samples. Serotype C was recovered in only 33% of organs, and never from brain or lung specimens. GR52 grew abundantly from all lung cultures, and yeast cells with large capsules were seen in histopathology inside the alveoli, peribronchial vessels and interalveolar spaces. They appeared to elicit no inflammatory response. We conclude that intraperitoneally inoculated C. neoformans var. gattii shows high virulence in this immunocompetent mouse model. Strain GR52 was highest in pathogenicity and had marked lung tropism. In contrast, the serotype C reference strain showed the lowest pathogenicity and seemed not to spread outside the abdominal viscera.

In vitro susceptibility to the sordarin derivative GM 237354 and amphotericin B were tested in a total of 190 Cryptococcus neoformans clinical isolates from different geographical areas of Spain and South American countries. Minimal inhibitory concentrations (MICs) were obtained using the NCCLS reference microbroth dilution method and analysed according the serotypes of Cr. neoformans. The MICs for amphotericin B were lower than 1.0 microg ml(-1) (MIC90% 0.5 microg ml(-1) , MIC50% 0.125 microg ml(-1)) but five isolates showed MICs of 2.0 microg ml(-1) to GM 237354 (MIC90% 1.0 microg ml(-1), MIC50% 0.5 microg ml(-1)). Cryptococcus neoformans var. gattii serotype B, was significantly less susceptible than A and AD serotypes (P = 0.047 and P = 0.022, respectively).

Background: The isolation of Candida species in urine in patients admitted in intensive care units who bear a vesical catheter is considered as a sign of bad prognosis. The etiological mechanisms and the appropriateness of administering an antifungal treatment in these patients is controversial., Patients and Method: The prevalence of Candida species in urine and the in vitro susceptibility to fluconazole by means of the E-test were determined in 560 patients admitted in an intensive care unit., Results: In 60 patients (11%) Candida species was isolated in urine after catheter replacement. 56.7% of isolates corresponded toC. albicans, 21.7% to C. glabrata, 15% to C.tropicalis, 5% to C. parapsilosis and 1.7% to C.kefyr. Only two isolates (C. albicans and C. glabrata)had a minimal inhibitory concentration (MIC) >= 64 microgram/ml (which was regarded as resistant) and 91% of strains had a MIC between 0.125 and 8 microgram/ml. MIC 90% for C. albicans and C. tropicalis was 1 microgram/ml,while it was 16 microgram/ml for C. glabrata. C. albicans is the most frequent species isolated in urine in these patients. However,over 40% of cases are found to have non-albicans species,especially C. glabrata and C. tropicalis. The majority of Candida isolates (91%) are sensitive in vitro to fluconazole.

The study constitutes an approach to the knowledge of the epidemiology of cryptococosis in Spain. For detection of cases 167 Spanish hospitals were contacted. All cases included were accompanied by the correspondent isolate of Cryptococcus neoformans, together with clinical, demographic and mycological data. Results obtained from January 1998 to end of December 1999 are analysed and presented here. Fifty-six Spanish hospitals reported 58 cases of cryptococcosis; only 43 of them were adequately documented and accompanied by the clinical isolate. The results showed a higher incidence in males (88.4%) than in females (11.6%); being most frequently affected those between 30 and 40 years old (48.8%). The 84.6% (33) corresponded to new cases and 15.4% (6) to relapses of the disease. The HIV infection was the most frequent risk factor reported (86%) and, for 29.7% (11) of them, cryptococcosis was the AIDS defining disease. For the diagnosis, CSF analysis showed the best results (India ink; culture and antigen detection). All strains collected (100%) corresponded to C. neoformans variety neoformans. Serotypes distribution was 45.5% for serotype A and 22.7% for each of serotypes D and AD.

The usefulness of the E-test((R) )has been studied to determine the in vitro susceptibility of 52 isolates of 17 species to five antifungal drugs: amphotericin B, 5-flucytosine, ketoconazole, itraconazole and fluconazole. Minimal inhibitory concentrations were determined following the manufacturers' instructions, except in the preparation of the inoculum. In this case a spectrophotometric method was used to obtain 1-5 ' 10(6) CFU/ml. Two different culture media were included: casitone-agar and RPMI 1640 agar. Most isolates showed clear growth in both media after 96 h of incubation at 37 degrees C. The species showed low MIC concentrations to ketoconazole and itraconazole. Only 55.8% of isolates showed MICs

Urine contamination by microorganisms may affect the interpretation of urinalysis in different areas of clinical diagnosis. This is particularly relevant in doping control. A prospective study was designed to assess the effects of urine contamination by selected pathogens on the endogenous androgenic steroid profile. Pooled urine from a healthy male volunteer with standard steroid profile compared with reference values for the Caucasian population was sterilized by filtration and stored in sterile glass tubes. Aliquots were inoculated with known amounts of 15 different organisms (bacteria, fungi, and moulds) and incubated at 37 degrees C for 2 weeks. Different markers of urine contamination, such as pH, deconjugation of steroids, and metabolic by-products, were determined. Alkalization of urinary pH was not a reliable indicator of urine contamination as several organisms grew in this medium and no alteration of this parameter was found. In uncontaminated urine, less than 10% of steroid glucuronide conjugates were spontaneously hydrolyzed. Higher rates of hydrolysis for sulfate conjugates were found. An unconjugated fraction higher than 10% of the total amount of testosterone was a reliable indicator of urine contamination. However, microbial production of testosterone or epitestosterone was not detected. In contrast, a few organisms were able to synthesize 5alpha-androstanedione, 5beta-androstanedione, and androstenedione using endogenous steroids as substrates., (Copyright 2001 Academic Press.)

Background: There are no established methods for the preparation and standardization of Fusarium solani antigens. This lack of standardization makes it difficult to use these antigens in allergenic diagnostic tests., Objective: To obtain an appropriate standardized method for the preparation of the different antigen types of F. solani., Methods: Production of fungal extracts, followed by biochemical and immunological characterization., Results: The somatic antigens presented the greatest protein content most of these proteins are common to the metabolic and hydrosoluble antigens, particularly those proteins at 35-39 kDa, 29-32 kDa and 15-16 kDa, as detected by electrophoresis and immunoblotting. The hydrosoluble antigens presented the highest protein diversity; these proteins were the most specific, showing minor determinants in common with the other antigens., Conclusions: It is recommended that a mixture of the different antigen sources be used in order to obtain extracts which would cover the maximum number of diagnostic possibilities.

The histoplasmosis in Spain is an imported disease presenting in most of case diagnostic difficulties. In this paper, the intradermal skin test with Histoplasma capsulatum antigen as diagnostic method in immunocompetent patients with clinical and radiological signs compatible with histoplasmosis after being visited Central and South American endemic counties, in which this mycosis is endemic. Nine Spanish patients coming from different countries of Latin America with fever and acute respiratory symptoms compatible with histoplasmosis were studied. Other nine accompanying subjects and five controls were also evaluated. Patients underwent mycological cultures and and serological tests for H. capsulatum. Intradermal test with 1% histoplasmine were done in all patients. Serology and skin tests tests were also performed in accompanying people. Intradermal were done in healthy controls. Skin test with histoplasmine were positive in seven of the nine patients. Six of these showed precipitating antibodies against the same antigen. H. capsulatum was only isolated from bone marrow biopsy samples in one patient. The seven patients were given itraconazole by oral route and all symptoms improved after 2 and 4 weeks. In five accompanying subjects the skin test were also positive so that a subclinical histoplasmosis was diagnosed. In the remaining patients and healthy accompanying subjects histoplasmosis infection was excluded. In non endemic geographical areas of histoplasmosis intradermal skin test with histoplasmin when used in immunocompetent individuals is an easy and reliable method for the diagnosis of this mycosis as well as for epidemiological studies.

Minimum inhibitory concentrations (MICs) of amphotericin B, 5-flucytosine, fluconazole, itraconazole and ketoconazole were determined against 42 clinical isolates of Cryptococcus neoformans var. neoformans using the Alamar YeastOne colorimetric method and the NCCLS reference microdilution method. No strains with resistance to amphotericin B, itraconazole or ketoconazole were detected with either method. Using the reference method, the MICs of fluconazole were >/= 64 mg/L, whereas using the colorimetric method all MICs were >/=16 mg/L. The MIC values of 5-flucytosine were also higher using the reference method (8-16 mg/L for 32% of isolates) compared with the colorimetric method. The percentage of agreement between the methods, using a difference of two dilutions, was 70.7% for itraconazole, 73.2% for amphotericin B, 80% for fluconazole, 88% for 5-flucytosine and 95% for ketoconazole. Overall, we conclude that for fluconazole and 5-flucytosine, in a low but not insignificant number of isolates, results with the two methods are discordant, some isolates being found sensitive with the colorimetric test, but resistant with the reference method.

Prevalence of candidemia has increased during the last decade, numerous predisposing factors are invoked, like parenteral drug abuse in the case of brown heroin syndrome, neutropenia, prolonged antibiotic therapeutics and immunosupression. Only a few articles are published about candidemia. Candidemia in AIDS patients seems very low; its incidence is estimated approximately one candidemia per 120 patients.

There are few reports of severe infections caused by Candida norvegensis. We here describe a case of C. norvegensis-associated intraabdominal abscess and Gore-Tex mesh-associated infection, successfully treated with liposomal amphotericin B and removal of the mesh. This is, to our knowledge, the first report of C. norvegensis causing this type of infection.

A new session in this Congress was the development of a discussion panel regarding controversies and queries about the main topics of treatment and prophylaxis of severe systemic mycoses. Experts presenting each side of three controversial areas provided an interchange of ideas and clarified those areas where there remain substantial disagreements. Some common recommendations have been made, and some differences persist. The contents of this session are compiled in this paper. Due to the limitations of space, this paper presents the most relevant parts of each presentation. The large and up-to-date list of references should be useful to gain a better understanding of these subjects.

Dermatomycoses caused by non-dermatophytic mycelial fungi are very rare with the exception of onychomycoses. Controversies regarding the pathogenic role often arise when these species and yeasts are isolated from nail scrapings. Candida albicans and Candida parapsilosis are the species identified more frequently from nails, particularly from finger nails. Because they could be resident flora of the skin, cultures should be interpreted according to clinical data, direct microscopic observation of clinical samples, and quantification of colonies. The recognition of other yeasts, such as Candida guillermondii, Candida famata or Candida krusei is more problematic. Isolation of moulds form toe nails accounts for 2 to 12% in different studies, with a prevalence of 7.6% in Barcelona being Scopulariopsis brevicaulis the most frequent species, but Aspergillus versicolor is also a particular etiologic agent of onychomycosis (2.5%). To confirm the etiology of any onychomycosis, standard criteria for mycological diagnosis and identification of moulds should be strictly applied. Experience in the diagnosis of superficial mycosis due to new species of fungi is required for improving current knowledge on the prevalence and clinical importance of this type of infections.

Twelve Spanish laboratories collected 325 yeast clinical isolates during a 30 day's period, among them 224 Candida albicans, 30 Candida glabrata, and 27 Candida parapsilosis. In vitro antifungal susceptibility to amphotericin B, ketoconazole, fluconazole and itraconazole was determined by an agar diffusion test (Neo-Sensitabs, Rosco, Denmark). All the isolates tested were susceptible in vitroto amphotericin B and nearly all (97.2%) to itraconazole. In vitrosusceptibility to fluconazole and ketoconazole was high (90.2% and 91.4% of isolates, respectively) but showed variations depending on the species tested. Resistance to fluconazole and ketoconazole was low in C. albicans (4% and 3%, respectively), but 30% of Candida guilliermondii and 36% of C. glabrata isolates were resistant to fluconazole. Ketoconazole resistance was observed in 40% of C. glabrata, and 17% of Candida tropicalis. Resistance to antifungal drugs is very low in Spain and it is related to non-C. albicans isolates.

The antifungal activity of eberconazole, a new imidazole derivative, against 124 clinical isolates of Candida comprising eight different species and to 34 isolates of Cryptococcus neoformans was compared to those of clotrimazole and ketoconazole. MICs of eberconazole, determined by the National Committee for Clinical Laboratory Standards standardized microbroth method, were equal to or lower than those of other azoles, especially for Candida krusei and Candida glabrata, which are usually resistant to triazoles.

We determined biovars and serotypes of 154 isolates of Cryptococcus neoformans from clinical and environmental sources from different areas of Spain. All clinical isolates belonged to C. neoformans var. neoformans. Serotypes showed an irregular distribution. C. neoformans var. gattii serotype B was isolated from necropsy specimens from goats with pulmonary disease.

Eberconazole is a new azole antifungal drug for topical treatment of superficial mycoses. The usefulness of this drug was evaluated in an experimental model of cutaneous candidosis in guinea pigs comparing with the classical clotrimazole in a single blind trial. Twenty-five animals were inoculated in two symmetrical areas of the back with Candida albicans developing skin infection. One group of 10 animals were treated once per day with clotrimazole 1% cream in one side and with excipient in the other. Other group of 10 guinea pigs received eberconazole 1% cream and excipient. Five animals did not receive any treatment and were used as controls. After five days of treatment most lesions cured or improved and cultures were negative when clotrimazole or eberconazole were applied. Seventy per cent of lesions treated with excipient were clinically improved and 10% cured, but 85% of cultures remained positive for C. albicans. The therapeutic efficacy of eberconazole 1% cream was similar to clotrimazole 1% cream in the guinea ping model of cutaneous candidosis. Tolerance of both drugs was excellent. These results suggest the usefulness of eberconazole in human cutaneous infections due to C. albicans.

Terbinafine 250 mg po daily was administered to 21 patients affected by tinea unguium of the toenails. In 14 cases Trichophyton rubrum was the aetiological agent while Trichophyton mentagrophytes var. interdigitale affected the rest. The treatment was administered randomized during 12 or 24 weeks in two groups of 11 (group A) and 10 (group B) patients. The clinical and mycological response was evaluated at week 12, 24 and 48. Adverse events were registered during the therapeutic period. At week 24, 55.5% of patients from group A were cured, only one patient did not improve and cultures were positive. In group B 66.6% were cured and three were clinically improved but cultures were positive. In the last control after 48 weeks the results were similar except one patient of the group B who worsened of his nails lesions. Only one case of acute urticaria was noted, the symptoms disappeared after the withdrawal of terbinafine. Terbinafine per os is a safe and effective antifungal for the treatment of the tinea unguium of the toenails. A period of 12 weeks of administration is enough in most of cases but when T. rubrum is the aetiology relapse could be possible.

The aim of this study was to adapt a spectrophotometric method for preparing the inocula of dematiaceous fungi used for in vitro susceptibility tests. Fifty-two isolates of 17 different species of dematiaceous fungi were used for this purpose. Homogeneous suspensions of conidia and hyphae of these isolates were obtained and adjusted for reading at 530 and 550 nm at 40% and 50% of transmittance. The suspensions were standardised to 1-5 x 10 e6 CFU/ml. Quality controls of the inocula were done by quantitative cultures on agar-Sabouraud plates. The inocula obtained by spectrophotometry showed little variability within all the isolates. This method can be useful for in vitro antifungal evaluation of dematiaceous fungi.