Antifungal susceptibility profile of diferent yeasts isolates from wild animals, cow's milk with subclinical mastitis and hospital environment / Perfil de suscetibilidade antifúngica de diferentes leveduras isoladas da microbiota de animais silvestres, leite com mastite subclínica e ambiente hospitalar

Abstract Yeast infections have acquired great importance due to increasing frequency in immunocompromised patients or patients undergoing invasive diagnostic and therapeutic techniques, and also because of its high morbidity and mortality. At the same time, it has been seen an increase in the emergence of new pathogenic species difficult to diagnose and treat. The aim of this study was to determine the in vitro susceptibility of 89 yeasts from different sources against the antifungals amphotericin B, voriconazole, fluconazole and flucytosine, using the VITEK® 2 Compact system. The antifungal susceptibility was performed automatically by the Vitek® 2 Compact system. The origin of the yeasts was: Group 1 - microbiota of wild animals (W) (26/89), 2 - cow's milk with subclinical mastitis (M) (27/89) and 3 - hospital enviorment (H) (36/89). Of the 89 yeasts submitted to the Vitek® 2 test, 25 (20.9%) were resistant to fluconazole, 11 (12.36%) to amphotericin B, 3 (3.37%) to voriconazole, and no sample was resistant to flucytosine. Regarding the minimum inhibitory concentration (MIC), fluconazole showed an MIC between 1 and 64 mg/mL for the three groups, voriconazole had an MIC between 0.12 and 8 mg/mL, amphotericin B had an MIC between 0.25 and 4 mg/mL for group H and group W respectively, between 0.25 and 16 mg/mL for group M and flucytosine had an MIC equal to 1μg/mL for all groups. The yeasts isolated from the H group showed the highest resistance to fluconazole 12/89 (13.49%), followed by group W (7.87%) and group M (5.62%). The more resistant group to voriconazole was followed by the M and H groups, the W group showed no resistance to this antifungal. Group H was the least resistant (2.25%) to amphotericin.

Resumo As infecções por leveduras têm adquirido grande importância, devido ao aumento da sua frequência em pacientes imunocomprometidos ou pacientes submetidos a técnicas diagnosticas e terapêuticas agressivas, e devido sua alta morbidade e mortalidade. Paralelamente tem-se observado um incremento na aparição de novas espécies patógenas difíceis de diagnosticar e tratar. O objetivo desse estudo foi avaliar a suscetibilidade in vitro de 89 leveduras de diferentes origens frente aos antifúngicos Anfotericina B, Voriconazol, Fluconazol e Fluocitocina pelo Sistema Vitek® 2. O antifungigrama foi realizado automaticamente pelo Vitek® 2 Compact. A origem das leveduras foi: Grupo 1- Microbiota de Animais Silvestres (S) (26/89), 2- Leite com mastite bovina subclínica (L) (27/89) e 3- Ambiente Hospitalar (H) (36/89). Das 89 leveduras submetidas à carta Vitek®, 25 (20.09%) foram resistentes ao fluconazol, oito (8.99%) à anfotericina B, três (3.37%) ao voriconazol, e nenhuma amostra mostrou-se resistente a fluocitosina. O grupo três (H) foi mais resistente ao fluconazol que os demais, já o dois (L) foi mais resistente ao voriconazol e a anfotericina B que os outros dois. O fluconazol pode ter apresentado maior número de resistências devido ser um fármaco comumente usado principalmente em humanos. As leveduras isoladas de humanos apresentaram maior número de resistências aos fármacos testados do que as leveduras isoladas de animais silvestres. O que pode ocorrer devido a uma maior exposição dos humanos aos fármacos em relação aos animais que vivem isolados em ambientes selvagens e na maioria dos casos nunca teve contato com fármacos de qualquer origem.

Anti-fungal potentials of extracellular metabolites of Western Ghats isolated Streptomyces sp. NII 1006 against moulds and yeasts.

Realization of hazardious effects of chemical fungicides has led to an interest in the usage of biocontrol agents. The present study, therefore, evaluates the biocontrol efficacy of Western Ghats (India) soil bacterial isolates. A potential strain NII 1006 was evaluated for its antagonistic property against a diverse range of moulds and yeasts. The strain was characterized morphologically, biochemically and molecularly, which revealed the isolate belonged to Streptomyces genus. Organic solvent extracts of NII 1006 culture filtrates inhibited the growth of the test pathogens indicating that growth suppression was due to extracellular anti-fungal metabolites present in the culture filtrates. The strain produced extracellular chitinase enzyme in addition to some stable partially purified anti-fungal compounds. Morphological changes such as hyphae degradation into debris and abnormal shapes were observed in test fungi and yeast grown on potato dextrose broth that contained the NII 1006 culture filtrate. The cell free supernatant has a tolerance to wide range of pH, temperature and enzymes such as lipase and protease. The biocontrol potential of NII 1006 strain may be correlated significantly with their ability to produce antibiotics as well as extracellular hydrolytic enzymes particularly chitinolytic enzyme.

Use of the VITEK 2 system to identify and test the antifungal susceptibility of clinically relevant yeast species

Eleven quality control isolates (Candida albicans ATCC 64548, C. tropicalis ATCC 200956, C. glabrata ATCC 90030, C. lusitaniae ATCC 200951, C. parapsilosis ATCC 22019, C. krusei ATCC 6258, C. dubliniensis ATCC 6330, Saccharomyces cerevisiae ATCC 9763, Cryptococcus neoformans ATCC 90012, C. gattii FIOCRUZ-CPF 60, and Trichosporon mucoides ATCC 204094) and 32 bloodstream isolates, including C. albicans, C. tropicalis, C. parapsilosis, C. glabrata, C. krusei, C. guilliermondii, C. pelliculosa (Pichia anomala), C. haemulonii, C. lusitaniae, and C. kefyr were identified at the species level by the VITEK 2 system. A set of clinical isolates (32 total) were used as challenge strains to evaluate the ability of the VITEK 2 system to determine the antifungal susceptibility of yeasts compared with the CLSI and EUCAST BMD reference standards. The VITEK 2 system correctly identified 100% of the challenge strains. The identification of yeast species and the evaluation of their susceptibility profiles were performed in an automated manner by the VITEK 2 system after approximately 15 h of growth for most species of Candida. The VITEK 2 system ensures that each test is performed in a standardized manner and provides quantitative MIC results that are reproducible and accurate when compared with the BMD reference methods. This system was able to determine the MICs of amphotericin B, flucytosine, voriconazole, and fluconazole in 15 h or less for the most common clinically relevant Candida species. In addition, the VITEK 2 system could reliably identify resistance to flucytosine, voriconazole, and fluconazole and exhibits excellent quantitative and qualitative agreement with the CLSI or EUCAST broth microdilution reference methods.

In vitro susceptibility of oral Candida albicans strains to different pH levels and calcium hydroxide saturated aqueous solution

Candida albicans is present in the oral cavity and in the whole digestive tract of humans and other animals, being frequently related to endodontic treatment failure. The present study determined the incidence of C. albicans in the oral cavity and the susceptibility of isolates to different pH values and saturated calcium hydroxide aqueous solution at pH 12.5. Sixty-five patients attending the Endodontic Clinic at the Sagrado Coração University participated in the study. The collected samples were cultivated in selective media for C. albicans and the isolates were tested in terms of resistance to both alkaline pH and saturated aqueous solution of calcium hydroxide. In relation to time variables, yeast viability was assessed by the Sabouraud's agar culture and fluorescein diacetate and ethidium bromide fluorescent staining method. Results from the different pHs and experimental times, including those from different techniques measuring fungal viability, were compared using the chi-square and Fisher's exact tests (α=0.05). The yeasts became completely inviable after 48 h of contact with the calcium hydroxide solution. On the other hand, when exposed to the alkaline culture broth, the yeasts were found to be viable at pHs 9.5 and 10.5 for up to 7 days. In conclusion, C. albicans can only be completely inhibited by direct contact with saturated calcium hydroxide aqueous solution after 48 h of exposure.

Candida albicans está presente na cavidade oral e em todo trato digestivo de humanos e outros animais, podendo estar relacionada a fracassos do tratamento endodôntico. O presente trabalho determinou a incidência de C. albicans na cavidade oral e a susceptibilidade das amostras isoladas frente a diferentes valores de pH e à solução aquosa saturada de hidróxido de cálcio em pH 12,5. Sessenta e cinco pacientes atendidos na Clínica de Endodontia da Universidade Sagrado Coração participaram da pesquisa. As amostras coletadas foram cultivadas em meios seletivos para C. albicans. As cepas isoladas foram testadas quanto a resistência ao pH alcalino e à solução saturada de hidróxido de cálcio. Frente às variáveis de tempo, a viabilidade das leveduras foi avaliada pelos métodos da cultura sobre ágar Sabouraud e de fluorescência pela técnica do diacetato de fluoresceína e brometo de etídio. Foram isoladas 30 linhagens de C. albicans coletadas da cavidade oral dos pacientes. Quando em contato com a solução de hidróxido de cálcio as leveduras foram totalmente inviabilizadas após 48 h de exposição. Quando expostas ao caldo de cultura alcalinizado as leveduras mantiveram-se viáveis em pH 9,5 e 10,5 por até 7 dias. Em conclusão, C. albicans pode ser completamente inibida pelo contato direto com solução aquosa saturada de hidróxido de cálcio após 48 h de exposição.

Species distribution and susceptibility profile of yeasts isolated from blood cultures: results of a multicenter active laboratory-based surveillance study in Argentina

The Mycology Department of the Instituto Nacional de Enfermedades Infecciosas "Dr. C. Malbrán", conducted the Second National Multicenter Survey on Fungemia due to Yeasts in Argentina. The aim was to obtain updated data of the frequency of the causative species encountered and their in vitro susceptibility to seven antifungal agents. Yeast species were identified by micromorphological and biochemical studies. Antifungal susceptibility testing was performed by the reference microdilution method E.Def 7.1 of the European Committee on Antibiotic Susceptibility Testing (EUCAST). A total of 461 viable yeasts were identified. The most frequent species were: Candida albicans (38.4 %), Candida parapsilosis (26 %), Candida tropicalis (15.4 %) and Candida glabrata (4.3 %). Other uncommon species, such as Candida viswanathii (0.6 %), Candida haemulonii (0.4 %), Candida inconspicua (0.2 %) and Candida fermentati (0.2 %) were also isolated. Among the Candida spp., 5.4 % and 1.6 % were resistant to fluconazole and voriconazole, respectively. Itraconazole and caspofungin were the most efficient agents against all Candida spp. tested (MIC < 1 mg/l). For anidulafungin, 21.6 % of C. parapsilosis showed a MIC value of 4 mg/l. Fluconazole was less active against 53.1 % of Cryptococcus neoformans (MIC > 8 mg/l), 75 % of Trichosporon spp., and 100 % of Rhodotorula spp., Geotrichum candidum, Saccharomyces cerevisiae. The global percentage of mortality was 20 %. The presence of uncommon species reinforces the need for performing continuous laboratory surveillance in order to monitor possible changes, not only in the epidemiological distribution of species, but also in the resistance to antifungal drugs.

Distribución de especies y perfil de sensibilidad de levaduras aisladas de hemocultivos: resultados de un estudio multicéntrico de vigilancia de laboratorio en Argentina. El Departamento Micología del Instituto Nacional de Enfermedades Infecciosas "Dr. Carlos G. Malbrán" condujo el segundo estudio multicéntrico nacional sobre funge- mias debidas a levaduras. El objetivo fue obtener datos actualizados sobre la distribución de especies y la sensibilidad in vitro frente a siete antifúngicos. Las levaduras fueron identificadas mediante el estudio de la micromorfología y la realización de pruebas bioquímicas. La determinación de la sensibilidad se realizó según el método de referencia E.Def 7.1 del European Committee on Antibiotic Susceptibility Testing (EUCAST). Se identificaron 461 levaduras. Las especies más frecuentes fueron Candida albicans (38,4 %), Candida parapsilosis (26 %), Candida tropicalis (15,4 %) y Candida glabrata (4,3 %). Se aislaron otras especies menos comunes, como Candida viswanathii (0,6 %), Candida haemulonii (0,4 %), Candida inconspicua (0,2 %) y Candida fermentati (0,2 %). Entre las especies del género Candida, el 5,4 % y el 1,6 % fueron resistentes al fluconazol y al voriconazol, respectivamente. El itraconazol y la caspofungina fueron los antifúngicos más eficaces in vitro frente a las especies de Candida evaluadas (CIM < 1 mg/l). Para la anidulafungina, el 21,6 % de los aislamientos de C. parapsilosis mostraron una CIM de 4 mg/l. El fluconazol fue menos activo para el 53,1 % de los aislamientos de Cryptococcus neoformans (CIM > 8 mg/l), el 75 % de los aislamientos de Trichosporon spp. y el 100 % de los aislamientos de Rhodotorula spp., Geotrichum candidum y Saccharomyces cerevisiae. El porcentaje de mortalidad fue del 20 %. La presencia de especies infrecuentes refuerza la necesidad de realizar la continua vigilancia de laboratorio con el fin de monitorear posibles cambios, no solo en la epidemiología de las especies causantes de fungemia, sino también en la resistencia a los antifúngicos.

Optimization of killer assays for yeast selection protocols / Optimización de la actividad killer para protocolos de selección de levaduras

A new optimized semiquantitative yeast killer assay is reported for the first time. The killer activity of 36 yeast isolates belonging to three species, namely, Metschnikowia pulcherrima, Wickerhamomyces anomala and Torulaspora delbrueckii, was tested with a view to potentially using these yeasts as biocontrol agents against the wine spoilage species Pichia guilliermondii and Pichia membranifaciens. The effectiveness of the classical streak-based (qualitative method) and the new semiquantitative techniques was compared. The percentage of yeasts showing killer activity was found to be higher by the semiquantitative technique (60%) than by the qualitative method (45%). In all cases, the addition of 1% NaCl into the medium allowed a better observation of the killer phenomenon. Important differences were observed in the killer capacity of different isolates belonging to a same killer species. The broadest spectrum of action was detected in isolates of W. anomala NPCC 1023 and 1025, and M. pulcherrima NPCC 1009 and 1013. We also brought experimental evidence supporting the importance of the adequate selection of the sensitive isolate to be used in killer evaluation. The new semiquantitative method proposed in this work enables to visualize the relationship between the number of yeasts tested and the growth of the inhibition halo (specific productivity). Hence, this experimental approach could become an interesting tool to be taken into account for killer yeast selection protocols.

En este trabajo se presenta un nuevo ensayo semicuantitativo que optimiza la detección de actividad killer en levaduras. Se evaluó la actividad killer de 36 cepas pertenecientes a las especies Metschnikowia pulcherrima, Wickerhamomyces anomala y Torulaspora delbrueckii, en vista del potencial uso de estas levaduras como agentes de biocontrol frente a las especies contaminantes de vinos Pichia guilliermondii y Pichia membranifaciens. Se comparó la efectividad de la técnica clásica basada en estrías (método cualitativo) con la del nuevo método semicuantitativo. El porcentaje de levaduras que mostraron actividad killer fue más alto cuando se utilizó el método semicuantitativo (60%) que con el método cualitativo (45%). En todos los casos, el agregado de 1% de NaCl en el medio permitió una mejor observación del fenómeno killer. Se observaron importantes diferencias en la capacidad killer de diferentes cepas dentro de la misma especie. Se detectaron dos cepas de W. anomala (NPCC 1023 y 1025) y dos cepas de M. pulcherrima (NPCC 1009 y 1013) con un amplio espectro de acción, ya que fueron capaces de inhibir el desarrollo de las tres levaduras sensibles evaluadas. La evidencia experimental demuestra la importancia de una adecuada selección de la cepa sensible al evaluar la actividad killer. El nuevo método semicuantitativo propuesto en este trabajo permite visualizar la relación entre el número de levaduras sembradas y el halo de inhibición del crecimiento (productividad específica). En conclusión, este método resulta una herramienta interesante para ser tenida en cuenta en los protocolos de selección de levaduras killer.

Species Distribution and Antifungal Susceptibilities of Yeast Clinical Isolates from Three Hospitals in Korea, 2001 to 2007 / 대한진단검사의학회지

BACKGROUND: We utilized results from the ARTEMIS DISK Global Antifungal Surveillance Program to evaluate the species distribution and fluconazole and voriconazole susceptibilities of yeast isolates from clinical specimens in South Korea from 2001 to 2007. METHODS: Data were collected on 5,665 yeast isolates from all body sites at three locations. All investigators tested clinical yeast isolates using the CLSI M44-A disk diffusion method. Test plates were automatically read and results were recorded using the BIOMIC image analysis plate reader system (Giles Scientific, USA). Species, drug, zone diameter, susceptibility category, and quality control results were collected quarterly via e-mail for analysis. RESULTS: Candida albicans was the most common isolate, but a progressive increase in non-C. albicans Candida and noncandidal yeast species has been observed in recent years. The overall percentages of isolates in each category (susceptible, susceptible dose dependent, and resistant) were 98.8%, 0.5%, and 0.7% and 99.2%, 0.2%, and 0.6% for fluconazole and voriconazole, respectively. Candida of 3 species exhibited decreased susceptibility to fluconazole (<90% S) in the order of that seen with the resistant (R) species: C. krusei, C. guilliermondii, and C. glabrata. Emerging resistance to fluconazole or voriconazole was documented among isolates of Cryptococcus neoformans, Trichosporon spp., and Rhodotorula spp. CONCLUSIONS: The species distribution and antifungal susceptibilities of yeasts may differ according to specimen type, testing method, hospital, and geographic region. Therefore, further large-scaled, long-term surveillance studies are needed to isolate yeasts and to confirm the species distribution and antifungal susceptibilities of yeast isolates from clinical specimens in Korea.

In Vitro Amphotericin B Susceptibility of Korean Bloodstream Yeast Isolates Assessed by the CLSI Broth Microdilution Method, Etest, and Minimum Fungicidal Concentration Test / 대한진단검사의학회지

BACKGROUND: Although amphotericin B (AMB) has a wide spectrum of activity that encompasses the majority of yeast isolates, there have been recent reports suggesting that some yeast isolates exhibit decreased susceptibility to AMB. However, in vitro AMB susceptibility of yeast species isolates from blood cultures in Korea has not been fully surveyed. METHODS: A total of 92 bloodstream yeast isolates from four Korean hospitals, representing 10 Candida species (69 isolates) and 4 non-Candida yeast species (23 isolates) were evaluated. AMB minimum inhibitory concentrations (MICs) were determined by two methods: the CLSI method and Etest. AMB minimum fungicidal concentrations (MFCs) were also determined. RESULTS: For all 92 yeast isolates, the CLSI method generated a restricted range of MICs (0.125 to 4 microgram/mL) with 3.3% exhibiting MICs > or =2 microgram/mL, and the corresponding MFC values ranged from 0.25 to 8 microgram/mL with 26.1% showing MFCs > or =2 microgram/mL. Etest produced the widest distribution of MICs, ranging from 0.03 to 32 microgram/mL. High AMB MICs (> or =0.38 microgram/mL) by Etest was observed in 34.8% of the isolates: Candida krusei (100%), Candida rugosa (100%), Trichosporon asashii (100%), Candida glabrata (82%), and Yarrowia lipolytica (75%). Etest disclosed that all isolates of Candida guilliermondii, Candida lusitaniae, Candida pelliculosa and Kodamaea ohmeri were highly susceptible to AMB (MIC < or =0.19 microgram/mL). CONCLUSIONS: Our study showed that Etest may be more useful to discriminate yeast isolates with reduced susceptibility to AMB, and some isolates of less common yeast species from Korea may have decreased AMB susceptibilities.

Identificación rápida y sensibilidad a toxinas killer de levaduras aisladas de micosis no sistémicas / Rapid identification and susceptibility to killer toxins of yeasts isolated from non-systemic mycoses

La identificación rápida y segura de los agentes etiológicos y el desarrollo de nuevos antifúngicos con blancos de acción más específicos resultarán en tratamientos de las micosis más efectivos y menos lesivos. Mediante un método molecular rápido (ITS1-5.8S ADNr-ITS2 PCR-RFLP) se identificaron 53 aislamientos de levaduras provenientes de infecciones no sistémicas registradas en hospitales públicos de la ciudad de Neuquén y en un centro oftalmológico de Buenos Aires durante el año 2005. Adicionalmente y utilizando el método de inhibición del crecimiento en placa, se evaluó la sensibilidad de estas levaduras a toxinas killer producidas por levaduras indígenas de la Patagonia y por cepas de referencia. Ocho especies de levaduras fueron identificadas entre los aislamientos clínicos: Candida albicans (52%) , Candida parapsilosis (17%) , Candida tropicalis (10%) , Candida krusei (5%) , Candida glabrata (4%) , Candida guilliermondii (4%) , Kluyveromyces lactis (4%) y Saccharomyces cerevisiae (4%) . El 69% de los aislamientos de la especie mayoritaria, C. albicans, se relacionó con infecciones vaginales. Por otra parte, el 61% de las levaduras provenientes de infecciones oculares correspondió a la especie C. parapsilosis. En las condiciones de ensayo, las toxinas producidas por las levaduras killer indígenas DVMais5 y HCMeiss5 pertenecientes a las especies Pichia anomala y P. kluyveri, respectivamente, exhibieron el mayor espectro de acción sobre las levaduras aisladas de materiales clínicos.

The use of quick and reliable yeast identification methods, as well as the development of new antifungal agents with more specific targets, will enable a more efficient treatment of mycoses. In the present work, a total of 53 clinical isolates obtained from non-systemic infections in Neuquén Hospitals and an ophthalmologic clinic in Buenos Aires during 2005, were identified by means of a rapid molecular method (ITS1-5.8S ADNr-ITS2 PCR-RFLP). Additionally, the killer susceptibility of the isolates was tested against reference and indigenous killer yeasts on plate tests. Eight yeast species were identified among the clinical isolates: Candida albicans (52%), Candida parapsilosis (17%), Candida tropicalis (10%), Candida krusei (5%), Candida glabrata (4%) , Candida guilliermondii (4%) , Kluyveromyces lactis (4%) and Saccharomyces cerevisiae (4%) . Sixty-nine percent of the isolates corresponding to the predominant species ( C. albicans) were related to vaginal infections. On the other hand, 61% of the yeasts associated with ocular infections were identified as C. parapsilosis. Two indigenous killer isolates DVMais5 and HCMeiss5, belonging to Pichia anomala and P. kluyveri respectively, exhibited the broadest killer spectrum against clinical isolates.

In vitro antimicrobial activity and antagonistic effect of essential oils from plant species.

Kahramanmaras, is a developing city located in the southern part of Turkey Thymus eigii (M. Zohary and RH. Davis) Jalas, Pinus nigraAm. sub sp pallasiana and Cupressus sempervirens L. are the useful plants of the Kahramanmaras province and have been understudy since 2004 for the traditional uses of plants empiric drug, spice, herbal tea industry herbal gum and fuel. The study was designed to examine the antimicrobial activities of essential oils of these plants by the disc diffusion and minimum inhibitory concentration (MIC) methods. In addition, antimicrobial activity of Thymus eigii was researched by effects when it was used together with antibiotics and even when it was combined with other essential oils. When the results of this study were compared with vancomycin (30 mcg) and erytromycin (15 mcg) standards, it was found that Thymus eigii essential oil was particularly found to possess strongerantimicrobial activity whereas other essential oils showed susceptible or moderate activity However, antimicrobial activity changed also by in vitro interactions between antibiotics and Thymus eigii essential oil, also between essential oils of these plants and that of Thymus eigii causing synergic, additive, antagonist effect.

Antifungal activity of propolis extract against yeasts isolated from onychomycosis lesions

The aim of this study was to determine the in vitro activity of propolis extract against 67 yeasts isolated from onychomycosis in patients attending at the Teaching and Research Laboratory of Clinical Analysis of the State University of Maringá. The method used was an adaptation made from the protocol approved by the National Committee for Clinical Laboratory Standards. The yeasts tested were: Candida parapsilosis 35 percent, C. tropicalis 23 percent, C. albicans 13 percent, and other species 29 percent. The propolis extract showed excellent performance regarding its antifungal activity: the concentration capable of inhibiting the all of the yeasts was 5 Î 10-2 mg/ml of flavonoids and 2 Î 10-2 mg/ml of flavonoids stimulated their cellular death. Trichosporon sp. were the most sensitive species, showing MIC50 and MIC90 of 1.25 Î 10-2 mg/ml of flavonoids, and C. tropicalis was the most resistant, with CFM50 of 5 Î 10-2 mg/ml of flavonoids and MFC90 of 10 Î 10-2 mg/ml. In view of the fact that propolis is a natural, low cost, non-toxic product with proven antifungal activity, it should be considered as another option in the onychomycosis treatment.

Estudio multicéntrico de fungemias por levaduras en la República Argentina / Multicenter study of bloodstream infections due to yeasts in Argentina

La incidencia de candidemias aumentó aproximadamente en un 500% en hospitales de alta complejidad y se observó un cambio en la distribución de especies del género Candida, con un incremento de las levaduras no Candida albicans. Con el objeto de conocer la distribución de especies asociadas a fungemias por levaduras en Argentina y determinar su sensibilidad a los antifúngicos de uso convencional, se realizó un estudio multicéntrico durante el período abril 1999 a abril 2000. Participaron 36 instituciones del país. Se colectaron 265 aislamientos de levaduras provenientes de hemocultivos, que se identificaron utilizando pruebas morfológicas, fisiológicas y bioquímicas y la determinación de la concentración inhibitoria mínima se realizó en base al estándar del NCCLS. La distribución de especies fue: Candida albicans (40,75%), Candida parapsilosis (28,67%), Candida tropicalis (15,84%), Candida famata (3,77%), Cryptococcus neoformans (3,77%), Candida glabrata (2,64%) y otras (4,53%). La mayoría de los aislamientos fueron sensibles a anfotericina B, fluconazol e itraconazol. La mortalidad asociada a las fungemias por levaduras estudiadas (n=265) fue del 30%, siendo más baja a lo descrito (33-54%) y fue menor en los pacientes que recibieron tratamiento antifúngico (26,3%), que en los no tratados (47%).

The incidence of candidemia has increased approximately 500% in high-complexity hospitals. A change in the spectrum of Candida infections due to species other than Candida albicans has also been detected. Between April 1999 and April 2000 a multicenter study was performed in order to determine the species distribution associated to candidemias in Argentina and the susceptibility profile of the isolates to the current antifungal drugs. Thirty six institutions have participated. All the 265 yeast strains isolated from blood cultures were identified by morphological, physiological, and biochemical tests. The antifungal susceptibility testing of isolates was performed based on the reference NCCLS procedure. The distribution of species was: Candida albicans (40.75%), Candida parapsilosis (28.67%), Candida tropicalis (15.84%), Candida famata (3.77%), Cryptococcus neoformans (3.77%), Candida glabrata (2.64%), and others (4.53%). Most of the isolates were susceptible to amphotericin B, fluconazole and itraconazole. Mortality associated to the fungemia by yeasts episodes (n=265) was 30%, lower than results previously determined (33-54%). The mortality percentage in patients who received antifungal therapy versus patients without treatment was 26.3% and 47%, respectively.

Tendencia del perfil de sensibilidad antimicrobiana de los aislamientos de sangre en un hospital oncológico (1998-2003) / Trend of antimicrobial drug-susceptibility of blood isolates at an oncological center (1998-2003)

OBJETIVO: Describir la frecuencia de la resistencia en microrganismos aislados en cultivos de sangre en pacientes de un hospital oncológico de tercer nivel. MATERIAL Y MÉTODOS: De enero de 1998 a diciembre de 2003, en el Instituto Nacional de Cancerología se desarrolló un estudio retrospectivo en el cual se obtuvieron cepas de cultivos de sangre que fueron incluidas y procesadas por sistema Bactec y Microscan, para determinar identificación y sensibilidad antimicrobiana. Se determinó la tendencia anual de la resistencia de cada organismo especificado a los diferentes antibióticos. Se obtuvo la diferencia porcentual (incremento o decremento) comparando la frecuencia de resistencia al inicio y al final del estudio. RESULTADOS: Se detectaron 2 071 cultivos positivos. Se recuperaron Gram negativos en 59.7 por ciento de las muestras, Gram positivos en 35.7 por ciento y levaduras en 4.6 por ciento. Escherichia coli fue el principal germen identificado (18.6 por ciento), seguido de S. epidermidis (12.7 por ciento) y Klebsiella spp (9 por ciento). Durante el periodo de estudio la sensibilidad se mantuvo estable y por arriba de 88 por ciento (excepto para Pseudomonas aeruginosa). La sensibilidad de ciprofloxacina para E. coli se encontró alrededor de 50 por ciento. Amikacina presentó mayor sensibilidad que gentamicina. Staphylococcus aureus presentó una sensibilidad a oxacilina de 96 por ciento y a vancomicina de 100 por ciento. S.epidermidis de 14 por ciento a oxacilina y de 98.6 por ciento a vancomicina. No se encontraron cepas de enterococo resistente a vancomicina. Todas las cepas de S. pneumoniae fueron sensibles a penicilina. CONCLUSIONES: Se considera que los patrones de resistencia encontrados en este hospital son el resultado del control en el uso de antimicrobianos, del programa de vigilancia de infecciones nosocomiales y de la utilización de terapia combinada en todos los pacientes con bacteremia.

Identificación de levaduras de interés médico / Identification of yeasts of medical interest

De distintas muestras clínicas, se aislaron 238 cepas de levaduras. Para identificarlas, se emplearon pruebas convencionales como la emisión de tubos germinativos y producción de clamidosporos para identificar a Candida albicans. Las levaduras que no presentaron los caracteres antes mencionados, se identificaron mediante equipo comercial que mide la actividad enzimática de las levaduras en presencia de cicloheximida y de diferentes azúcares (Candifast de International Mycoplasma S.A.). De las levaduras aisladas, 194 resultaron ser Candida albicans y 44 cepas correspondieron a otras especies de Candida y a otros géneros de levaduras

Pruebas de sensibilidad in vitro para levaduras: evaluación de un micrométodo / In vitro susceptibility tests for yeasts: evaluation of a micromethod

Se evaluó un micrométodo para la realización de pruebas de sensibilidad de levaduras frente a antifúngicos, basado en el macrométodo en medio líquido estandarizado por el National Committee for Clinical Laboratory Standards (NCCLS) Subcommittee on Antifungal Susceptibility Testing. En este trabajo se compararon ambos métodos utilizando 6 cepas de referencia de diferente sensibilidad a los siguientes antifúngicos, anfotericina B (AMB), flucitosina (5FC), fluconazol (FCZ), itraconazol (ITZ), ketoconazol (KTZ) y miconazol (MCZ). Se observaron variaciones de sólo 1 ó 2 diluciones entre los resultados de las concentraciones inhibitorias mínimas (CIM) obtenidas con las dos técnicas. Asimismo, se compararon las lecturas visuales de CIM por micrométodo con las mediciones turbidimétricas del crecimiento en distintas concentraciones de antifúngicos frente a 47 aislamientos de Candida albicans. Existió una correlación significativa (p<0.001) entre CIM visual y la inhibición del 80 por ciento de crecimiento determinada por turbidimetría con AMB, 5FC, FCZ, ITZ y MCZ; en cambio no hubo correlación alguna para KTZ (p=1.00)

In vitro activity of propolis against bacterial and yeast pathogens isolated from human infections

The in vitro activity of propolis against 118 Staphylococcus aureus, 108 Escherichia coli, 60 Salmonella typhimurium, 50 Candida albicans, 23 Candida parapsilosis, 19 Candida tropicalis and 14 Candida guilliermondii isolated from human infections was studied by the agar dilution method. Among the bacteria, the Gram-negative were the least susceptible organisms showing minimal inhibitory concentration (MIC) for 90 per cent of the strains tested ranging from 22.5 mg/ml - 10,2 per cent, 23.1 mg/ml - 10.5 per cent V/V. The MIC's for Candida ranged from 0.80 mg/ml to > 11 mg/ml (0.40 per cent V/V to>5.0 per cent V/V) the strains of C. parapsilosis being the least susceptible. The relative order of susceptibility among all isolates,was: S.aureus > C.tropicalis > C.albicans > C.guilliermondii > C.parapsilosis > S.typhimurium > E.coli.

In vitro activity of the killer toxin from the yeast hansenula anomala against some different microorganisms

The killer toxin of Hansenula anomala showed killer activity toward some different microrganisms. The killer toxin produced by this species was purified. This toxin inhibited completely the growth of 2 species of yeasts,. Candida albicans, and CKefyr; three species of dermatophytes, Microsporum gypseum, M. Canis and Trichophyton mentagrophytes; and 2 species of moulds Fusarium solani and Aspergillus terreus tested at concentrations 100 ul/ml or less. The minimum inhibitory concentration [MIC] of the killer toxin against Candida guilliermondii and C. parapsilosis tested was 20 ul/ml or more. [MIC] of the killer toxin against, M. audoiunii, Sepedomium chrysospermun and Penicillium Coryophilum tested was 60 ul/ml or more. Finally the [MIC] of this killer toxin against the following bacterial strains: Proteus Vulgaris. Staphylococcus aureus, Salmonella typh, E. coli and Rhizobium legumnosarum tested was 25 ul/ml or more. But the [MIC] of this toxin against the two species of: Bacillus subtilus and Pseudomonas areuginosa was 50 ul/ml or more. In this paper we found that the purified H. anomala killer toxin used for the determination the [MIC] of a wide range of yeasts, moulds and bacteria including a number of important species

In vitro study of the antibiotic activity of diterpenes

The antibiotic activity of 16 diterpenes and of 2 phenanthrehene derivatives of diterpene origin, isolated from Brazilian plants was investigated against bacteria, yeasts, dermatophytes and aflatoxin-producing fung. Eight of the compounds were completely inactive. None showed inhibitory activity against the Gram-negative bacteria assaywed or aginst the Aspergillus species. The phenanthrene derivatives, chemically very simular to phytoalexins isolated from orchids, were especially active

Atividade da inulinase em leveduras e fermentaçöes de extratos de Helianthus tuberosus L / Inulinase activity in yeasts and fermentations of Helranthus tuberosus L extracts

Devido às concentraçöes de inulina em H. tuberosus, leveduras com boa produçäo de inulinase e alto potencial fermentativo säo imprescindíveis na viabilizaçäo industrial deste vegetal. Selecionando-se leveduras com estas propriedades, dentre 21 espécies avaliadas, encontram-se 11 linhagens com boa capacidade fermentativa. As leveduras com maior rendimento de produçäo de álcool foram Kluyveromyces marxianus, K. fragilis, Candida pseudotropicalis e Torulopsis coelliculosa. Em fermentaçöes sucessivas conduzidas a 30-C, com 15,6% de ART e por 48 horas, chegou-se a obter o rendimento médio de 85% em relaçäo ao teórico. Nestas condiçöes, após 10 horas de fermentaçäo encontraram-se rendimentos de 74,4% e uma taxa de fermentaçäo de 5,9g/1/h**-1. A atividade máxima da inulinase foi de 3,6micronM (equivalente frutose)/ml/min e verificou-se ocorrer repressäo catabólica da inulinase por frutose em K. marxianus e K. fragilis. Com estes resultados conclui-se ser viável a fermentaçäo de extratos sem prévia hidrólise, o que facilitará o processamento industrial de extratos de H. tuberosus