Browsing by Author "Rosado, Laura"
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- Diagnosis of Tinea pedis and onychomycosis in patients from Portuguese National Institute of Health Dr. Ricardo Jorge: a four year studyPublication . Viegas, Carla; Sabino, Raquel; Parada, Helena; Brandão, João; Carolino, Elisabete; Rosado, Laura; Verissimo, CristinaABSTRACT - Tinea pedis and onychomycosis are two rather diverse clinical manifestations of superficial fungal infections, and their etiologic agents may be dermatophytes, non-dermatophyte moulds or yeasts. This study was designed to statistically describe the data obtained as results of analysis conducted during a four year period on the frequency of Tinea pedis and onychomycosis and their etiologic agents. A questionnaire was distributed from 2006 to 2010 and answered by 186 patients, who were subjected to skin and/or nail sampling. Frequencies of the isolated fungal species were cross-linked with the data obtained with the questionnaire, seeking associations and predisposing factors. One hundred and sixty three fungal isolates were obtained, 24.2% of which composed by more than one fungal species. Most studies report the two pathologies as caused primarily by dermatophytes, followed by yeasts and lastly by non-dermatophytic moulds. Our study does not challenge this trend. We found a frequency of 15.6% of infections caused by dermatophytes (with a total of 42 isolates) of which T. rubrum was the most frequent species (41.4%). There was no significant association (p >0.05) among visible injury and the independent variables tested, namely age, gender, owning pet, education, swimming pools attendance, sports activity and clinical information. Unlike other studies, the variables considered did not show the expected influence on dermatomycosis of the lower limbs. It is hence necessary to conduct further studies to specifically identify which variables do in fact influence such infections.
- Diagnosis of Tinea pedis and onychomycosis in patients from Portuguese National Institute of Health: a four-year studyPublication . Viegas, Carla; Sabino, Raquel; Parada, Helena; Brandão, João; Carolino, Elisabete; Rosado, Laura; Veríssimo, CristinaTinea pedis and onychomycosis are two rather diverse clinical manifestations of superficial fungal infections, and their etiologic agents may be dermatophytes, non-dermatophyte moulds or yeasts. This study was designed to statistically describe the data obtained as results of analysis conducted during a four year period on the frequency of Tinea pedis and onychomycosis and their etiologic agents. A questionnaire was distributed from 2006 to 2010 and answered by 186 patients, who were subjected to skin and/or nail sampling. Frequencies of the isolated fungal species were cross-linked with the data obtained with the questionnaire, seeking associations and predisposing factors. One hundred and sixty three fungal isolates were obtained, 24.2% of which composed by more than one fungal species. Most studies report the two pathologies as caused primarily by dermatophytes, followed by yeasts and lastly by non-dermatophytic moulds. Our study does not challenge this trend. We found a frequency of 15.6% of infections caused by dermatophytes (with a total of 42 isolates) of which T. rubrum was the most frequent species (41.4%). There was no significant association (p >0.05) among visible injury and the independent variables tested, namely age, gender, owning pet, education, swimming pools attendance, sports activity and clinical information. Unlike other studies, the variables considered did not show the expected influence on dermatomycosis of the lower limbs. It is hence necessary to conduct further studies to specifically identify which variables do in fact influence such infections
- Identification of a fungal community on gilded wood carved heritagePublication . Pinheiro, Ana Catarina; Brandão, João; Veríssimo, Cristina; Jurado, Valme; Rosado, Laura; Oliveira, Bruna; Macedo, Maria filomenaDuring restoration activities, a fungal reddish area was noted on a gilded wood carved ceiling decoration from a sacristy of a church in Aveiro, Portugal. The protocols used to identify the fungal community present in this wooden material included the classical culturing methods and a molecular biology protocol based on the ability of denaturing high performance liquid chromatography (DHPLC) to separate fungal DNA from several species. This last method complemented the results obtained with the traditional culturing method approach and the overall study revealed three fungal species: Serpula lacrymans (Wulfen) P. Karsten, Paecilomyces lilacinus (Thom) Samson and Penicillium chrysogenum Thom. This is, to the author's knowledge, the first documented case of S. lacrymans in Portuguese built cultural heritage. Despite the antifungal solution applied and the dryer environment recommended, the water activity (Aw) levels recorded on location are still dangerously high and may allow fungal regrowth. Periodic inspections were advised.
- Isolates from hospital environments are the most virulent of the Candida parapsilosis complexPublication . Sabino, Raquel; Sampaio, Paula; Carneiro, Catarina; Rosado, Laura; Pais, CéliaBackground: Candida parapsilosis is frequently isolated from hospital environments, like air and surfaces, and causes serious nosocomial infections. Molecular studies provided evidence of great genetic diversity within the C. parapsilosis species complex but, despite their growing importance as pathogens, little is known about their potential to cause disease, particularly their interactions with phagocytes. In this study, clinical and environmental C. parapsilosis isolates, and strains of the related species C. orthopsilosis and C. metapsilosis were assayed for their ability to induce macrophage cytotocixity and secretion of the pro-inflammatory cytokine TNF-a, to produce pseudo-hyphae and to secrete hydrolytic enzymes. Results: Environmental C. parapsilosis isolates caused a statistically significant (p = 0.0002) higher cell damage compared with the clinical strains, while C. orthopsilosis and C. metapsilosis were less cytotoxic. On the other hand, clinical isolates induced a higher TNF-a production compared with environmental strains (p < 0.0001). Whereas the amount of TNF-a produced in response to C. orthopsilosis strains was similar to the obtained with C. parapsilosis environmental isolates, it was lower for C. metapsilosis strains. No correlation between pseudo-hyphae formation or proteolytic enzymes secretion and macrophage death was detected (p > 0.05). However, a positive correlation between pseudo-hyphae formation and TNF-a secretion was observed (p = 0.0119). Conclusions: We show that environmental C. parapsilosis strains are more resistant to phagocytic host defences than bloodstream isolates, being potentially more deleterious in the course of infection than strains from a clinical source. Thus, active environmental surveillance and application of strict cleaning procedures should be implemented in order to prevent cross-infection and hospital outbreaks.
- Monitorização da qualidade das areias em zonas balneares: relatórioPublication . Brandão, João; Rosado, Carmen; Silva, Cecília; Alves, Célia; Almeida, Cristina; Carrola, Cristina; Veríssimo, Cristina; Noronha, Graça; Parada, Helena; Barroso, Manuela; Falcão, Leonor; Rodrigues, Raquel; Sabino, Raquel; Rosado, LauraA qualidade ambiental das praias tem vindo a adquirir uma importância crescente entre os critérios de escolha de destino turístico. Apesar de nos últimos anos a legislação ter evoluído no sentido do ordenamento da zona costeira e do tratamento de águas residuais, o único indicador da qualidade relacionado com a saúde publica que pode permitir aos utentes uma escolha orientada, é a qualidade da água balnear. Justifica-se o estudo da qualidade microbiológica da areia, tendo em consideração que a actual directiva 2006/7/EC defende toda uma estrutura de protecção de qualidade da água balnear e zona envolvente bem como a saúde dos seus utilizadores. Também a Organização Mundial de Saúde no guia “Guidelines for safe recreational waters Volume 1 - Coastal and fresh waters” (publicado em 2003) exprime a preocupação com a qualidade das areias da praia por estas poderem constituir um reservatório de agentes de infecção, sobretudo em zonas balneares onde a utilização da areia apresenta maior relevância. Perante esta motivação, a Associação das Bandeiras Azuis para a Europa, o Instituto do Nacional de Saúde Dr. Ricardo Jorge e o Instituto do Ambiente reuniram esforços no sentido de desenvolver mais este projecto para validar a metodologia desenvolvida em estudos anteriores e alargar a sua aplicação ao território insular.
- Mould and yeast identification in archival settings: Preliminary results on the use of traditional methods and molecular biology options in Portuguese archivesPublication . Pinheiro, Ana Catarina; Macedo, M.F.; Jurado, V.; Saiz-Jimenez, C.; Viegas, Carla; Brandão, João; Rosado, LauraThis project was developed to fully assess the indoor air quality in archives and libraries from a fungal flora point of view. It uses classical methodologies such as traditional culture media e for the viable fungi e and modern molecular biology protocols, especially relevant to assess the non-viable fraction of the biological contaminants. Denaturing high-performance liquid chromatography (DHPLC) has emerged as an alternative to denaturing gradient gel electrophoresis (DGGE) and has already been applied to the study of a few bacterial communities. We propose the application of DHPLC to the study of fungal colonization on paper-based archive materials. This technology allows for the identification of each component of a mixture of fungi based on their genetic variation. In a highly complex mixture of microbial DNA this method can be used simply to study the population dynamics, and it also allows for sample fraction collection, which can, in many cases, be immediately sequenced, circumventing the need for cloning. Some examples of the methodological application are shown. Also applied is fragment length analysis for the study of mixed Candida samples. Both of these methods can later be applied in various fields, such as clinical and sand sample analysis. So far, the environmental analyses have been extremely useful to determine potentially pathogenic/toxinogenic fungi such as Stachybotrys sp., Aspergillus niger, Aspergillus fumigatus, and Fusarium sp. This work will hopefully lead to more accurate evaluation of environmental conditions for both human health and the preservation of documents
- New polymorphic microsatellite markers able to distinguish among Candida parapsilosis sensu stricto isolatesPublication . Sabino, Raquel; Sampaio, Paula; Rosado, Laura; Stevens, David A.; Clemons, Karl V.; Pais, CéliaAmong the Candida species causing bloodstream infections, Candida parapsilosis is one of the most frequently isolated. The objective of the present work was the identification of new microsatellite loci able to distinguish among C. parapsilosis isolates. DNA sequences with trinucleotide repeats were selected from the C. parapsilosis genome database. PCR primer sets flanking the microsatellite repeats were designed and tested with 20 independent isolates. On the basis of the amplification efficiency, specificity, and observed polymorphism, four of the sequences were selected for strain typing. Two hundred thirty-three independent C. parapsilosis sensu stricto isolates were genotyped by using these markers. The polymorphic loci exhibited from 20 to 42 alleles and 39 to 92 genotypes. In a multiplex analysis, 192 genotypes were obtained and the combined discriminatory power of the four microsatellites was 0.99. Reproducibility was demonstrated by submission of subcultures of 4 isolates each, in triplicate, interspersed with unique numbers among a group of 30 isolates for blind testing. Comparison of the genotypes obtained by microsatellite analysis and those obtained by randomly amplified polymorphic DNA analysis, restriction fragment length polymorphism analysis, and internal transcribed sequence grouping was performed and showed that the microsatellite method could distinguish individual isolates; none of the other methods could do that. Related species, C. orthopsilosis and C. metapsilosis, were not confused with C. parapsilosis sensu stricto. These new microsatellites are a valuable tool for use for the differentiation of C. parapsilosis sensu stricto strains, vital in epidemiology to answer questions of strain relatedness and determine pathways of transmission.
- Qualidade Microbiológica de Areias de Praias Litorais : relatório finalPublication . Brandão, João; Wergikosky, Bela; Rosado, Carmen; Noronha, Graça; Rosado, Laura; Veríssimo, Cristina; Falcão, Maria Leonor; Giraldes, Alexandra; Simões, M.; Rebelo, Helena; Veríssimo, CristinaActualmente, não existem quaisquer linhas de orientação quanto à qualidade das areias das praias. A qualidade ambiental das praias tem vindo a adquirir uma importância crescente entre os critérios de escolha do destino turístico. O único indicador da qualidade disponível para os utentes conhecerem o estado do ambiente de uma praia, é a qualidade da sua água balnear. No âmbito da campanha «Areia Limpa, Praia Saudável» da Associação Bandeira Azul da Europa (ABAE) para o ano de 2001, esta Associação convidou o Instituto do Ambiente (IA), o Instituto Nacional de Saúde Dr. Ricardo Jorge (INSA), a Câmara Municipal de Cascais (CMC), a Câmara Municipal de Viana do Castelo (CMVC) e o Instituto da Água (INAG), para conceber um projecto de investigação sobre este tema. Assim, planeou-se um estudo que teve por objectivos seleccionar os indicadores de qualidade que melhor caracterizam a contaminação microbiológica das areias das praias, propor os respectivos valores de referência e os métodos de análise mais adequados para a determinação dos indicadores propostos. Pretende-se que este estudo contribua para o estabelecimento de regras para a monitorização da qualidade microbiológica das areias das praias, de métodos de gestão de zonas balneares e para a elaboração de normas de conduta de utilização destas pelos utentes. Foi então delineado um modelo de estudo, no qual foram comparados três grupos de praias com níveis de qualidade crescente: zonas balneares que segundo a Directiva 76/160/CEE, foram classificadas de má qualidade, outras de boa qualidade (Bandeira Azul) e praias com baixa ocupação humana, de forma a se poder estabelecer os limites de contaminação aceitáveis. As colheitas foram efectuadas ao longo de um ano e analisado o potencial efeito da sazonalidade. Colheram-se amostras de areia das zonas molhada e seca que foram lavadas e o líquido de lavagem analisado para parâmetros microbiológicos e químicos previamente seleccionados. Simultaneamente foram colhidas amostras de água do mar que conjuntamente com as areias, foram analisadas por dois métodos nos Laboratórios de Referência do Ambiente e da Saúde. Tratados os resultados, foram detectadas diferenças significativas entre os três grupos de praias, para a água e areia molhada e uma boa correlação entre estas variáveis para a maioria dos parâmetros bacteriológicos. A areia seca não apresentou diferenças significativas entre os grupos de praias, teve uma boa correlação com a areia molhada mas não com a água, principalmente nos parâmetros micológicos. Os indicadores que melhor definem estas diferenças são os coliformes totais e os enterococos intestinais em bacteriologia e o número total de fungos, leveduras e fungos potencialmente patogénicos em micologia. Os métodos escolhidos foram: o método de espalhamento para os parâmetros micológicos, o método do número mais provável para a determinação de coliformes incluindo E. coli e o método de filtração por membrana para os enterococos intestinais. Esta escolha fundamentou-se nos resultados da análise estatística e no caso de métodos equivalentes na sua operacionalidade. As principais recomendações resultantes deste trabalho são em primeiro lugar a alteração da classificação das zonas balneares com base na qualidade das águas; em seguida acrescentar a qualidade microbiológica das areias como critério obrigatório para a atribuição do galardão da Bandeira Azul; finalmente monitorizar apenas a areia seca das zonas balneares, três vezes ao ano durante a época balnear, para os parâmetros acima descritos pelos métodos aqui estabelecidos.
- The Sensitivity and Resistance of Yeasts Isolated from Women with Vulvovaginal Candidiasis to Common Antifungal drugs Using Disc DiffusionPublication . Moallaie, Hossein; Verissimo, Cristina; Brandão, João; Rosado, LauraBackground and Purpose: Due to the ever-increasing use of antifungal drugs especially those of azole group, the prevalence of recurrent forms of vaginal infections and the number of drug-resistant yeasts are on the rise. Therefore, the rpesent study is conducted to investigate the sensitivity and resistance of yeasts isolated from vaginal infections to antifungal drugs. Methods and Materials: This cross-sectional descriptive analytical study was conducted on 118 yeasts isolated from 436 suspects of vulvovaginal candidiasis; their sensitivity and resisitance to drugs belonging to imidazoles group including clotrimazole (CTR), fluconazole (FCA), ketoconazole (KET), miconazole (MCZ) and econazole (EC) as well as nystatine (NY) belonging to polyene group using the standard disc diffusion technique. To determine their relationship clinical symptoms and the raltion of resistance to one drug with resistance to other drugs, relevant tests were used including chi-square, kappa and linear regression coefficient in SPSS 11. Results: The results showed no resistance to nystatine from polyene group and econazole from azole group; however, 53 cases (%44.9) were resistant to fluconazole, 26 cases (%22) to miconazole, 10 cases (%8.5) to clotrimazole and 2 cases (%1.7) to ketoconazole. Conclusion: The results of linear correlation showed a negative correlation between the sensitivity of yeasts to CTR and KET and clinical symptoms.