Pulmonary mycobiome of patients with suspicion of respiratory fungal infection – an exploratory study

Objective: This pilot study aimed to characterize the pulmonary mycobiome of patients with suspicion of fungal infection of the respiratory tract as well as to identify potentially pathogenic fungi colonizing/infecting their lungs. Methods: A cohort of 10 patients was analyzed, including HIV+ patients and patients with active infection caused by Mycobacterium species. Their respiratory samples (bronchoalveolar lavage fluid/ bronchial secretions) were pre-treated with lyticase and proteinase K; DNA was extracted using the High Pure PCR Template Preparation kit following the manufacturer’s instructions. The internal transcribed spacer region 1 (ITS1) and calmodulin gene were amplified by PCR and the resulting amplicons were sequenced using the Illumina MiSeq platform with pair-end reads of 150 bp. The obtained results were analyzed using the PIPITS pipeline as described by Gweon et al. [1]. Operational taxonomic units (OTU) to which less than 0.1% of the total reads attributed were disregarded. Results: Thirty-seven different OTU were identified from which two belonged to the Plantae kingdom, 11 had less than the 0.1% threshold of the total reads and were therefore disregarded. The remaining 24 different OTU (grouped in 17 phylotypes), were considered as part of the pulmonary mycobiome of patients. Two phyla were identified: Basidiomycota (33.3%) and Ascomycota (54.2%). Regarding the Basidiomycota phylum, reads were classified in three classes (Agaricomycetes, Tremellomycetes and Walleomycetes), while for the Ascomycota phylum four different taxonomical classes were identified: Pneumocystidomycetes, Dothideomycetes, Eurotiomycetes and Saccharomycetes, with the latter being the most frequent class. Twelve fungal genera were identified, being Candida the most frequently detected. The median number of fungal genera detected in patients’ pulmonary mycobiome was six (ranging from two up to nine). The genus Papilotrema and the potentially pathogenic genera Cryptococcus and Pneumocystis were exclusively found in the pulmonary mycobiome of HIV+ + patients. Other potentially pathogenic fungi such as Aspergillus spp., Trichosporon spp., Saccharomyces spp. and Schizophyllum spp. were also detected. Conclusion: This pilot study illustrates how the pulmonary mycobiome is rich and highly variable in patients with fungal infections. The obtained results suggest that the described metagenomic analysis may possess a great ability to quickly and effectively detect potentially pathogenic fungi in the mycobiome of patients, making it a promising future diagnostic tool. Thus, further optimization, standardization and clinical validation of these NGS methodologies should be warranted in the future.info:eu-repo/semantics/publishedVersio

Novel Clinical and Laboratorial Challenges in Aspergillosis

This article belongs to the Special Issue Aspergillus and Health 2.0In recent years, research in the areas of Aspergillus and aspergillosis has continued to ad vance rapidly, including advancements in genomics, immunological studies, clinical areas, and diag nostic areas. Recently, new risk groups for the development of aspergillosis have emerged—patients with influenza- or COVID-19-ssociated pulmonary aspergillosis. The rise and spread of antifungal re sistances have also become a clinical concern in some geographic areas and have drawn the attention of clinicians due to difficulties in treating these infections. In this paper, a snapshot of these issues is presented, emphasizing these novel clinical and laboratorial challenges in the aspergillosis field and focusing on their actual relevance.info:eu-repo/semantics/publishedVersio

Assessing the Potential of FT-IR to Identify Clinical Strains of Candida spp

Produto de um estágio curricular21 clinical isolates of Candida spp were examined by FT-IR in order to compare identification using FT-IR with biochemical identification (bioMerieux®). FT-IR revealed itself as a fast and effective method for identification of the strains tested as all were placed into the appropriate species, including one which was misidentified by the biochemical method.Instituto Nacional de Saúde Doutor Ricardo Jorg

Histoplasmosis in Portugal: rare infection?

O Histoplasma capsulatum é o agente etiológico da histoplasmose, apresenta duas variedades com caraterísticas epidemiológicas diferentes var. capsulatum e var. duboisii endémicas no continente americano e no continente africano, respetivamente. Nas últimas décadas têm sido descritos casos de histoplasmose na Europa e em países asiáticos como a China, onde esta infeção não é considerada endémica. A facilidade de movimentação das populações tem contribuído para alterar o padrão epidemiológico desta infeção. Este trabalho tem como objetivo analisar o número de casos registados em Portugal e chamar a atenção para a importância de melhor se conhecer a epidemiologia desta infeção no nosso país. A histoplasmose não é uma doença de declaração obrigatória, os casos de histoplasmose existentes resultam de diagnósticos clínicos observados no âmbito dos internamentos hospitalares. O número médio de episódios de internamento hospitalar referenciados nos Base de dados de Grupos de Diagnóstico Homogéneo durante o período de 2009 a 2014 foi de 23 episódios/ano. No mesmo período foram descritos na literatura dez casos de Histoplasmose em Portugal, tratando-se sobretudo de apresentações clínicas de interesse científico em que algumas se referem a casos com período de latência de 40 anos após exposição. Apesar de ser considerada uma doença rara na Europa, clínicos e microbiologistas devem estar em alerta e aumentar o seu conhecimento sobre a patogenicidade, os métodos de diagnóstico diferencial, o tratamento e a evolução do padrão epidemiológico desta e de outras infeções fúngicas.Histoplasma capsulatum is the etiologic agent of histoplasmosis; it has two varieties with different epidemiological features: var. capsulatum and var. duboisii, endemic in America and Africa, respectively. In the last decades, several cases of histoplasmosis have been reported in European and Asian countries like China, where this infection is not considered endemic. The continuous flow of populations around the world has contributed to the change the epidemiological pattern of this infection. This work aims to analyze the number of cases registered in Portugal and raise attention to the importance of a better understanding of the epidemiology of this infection in our country. Histoplasmosis is not a notifiable disease and therefore, the existing cases of histoplasmosis in our country result of clinical diagnosis in the context of hospital admissions. The average number of hospitalization episodes referenced in the Homogeneous Diagnostic Group Database during the period 2009-2014 was of 23 episodes/year. In the same period, ten cases of histoplasmosis were published. These publications are mostly clinical presentations of scientific interest, some of which describe histoplasmosis cases with a latent period of 40 years after exposure. Although considered as a rare disease in Europe, physicians and clinical microbiologists should be aware of this infection. An increased knowledge on the pathogenicity, methods of differential diagnosis, and treatment is, therefore, mandatory to understand the evolution of the epidemiological pattern of this and other fungal infections

Algorithm for the diagnosis of deep fungal infections

Amostras de tecidos de doentes com suspeita de infeções fúngicas profundas ou subcutâneas foram analisadas no Laboratório Nacional de Referência de Infeções Parasitárias e Fúngicas do INSA, de acordo com um algoritmo de diagnóstico desenvolvido que apresenta uma abordagem polifásica, que tem por objetivo permitir um diagnóstico laboratorial mais rápido das infeções fúngicas profundas. Quarenta e seis amostras de tecido de 39 doentes com suspeita de infeções fúngicas profundas ou subcutâneas foram analisadas durante um período de 26 meses ( janeiro de 2015-fevereiro de 2017) pelo laboratório do INSA, usando uma abordagem laboratorial que incluiu cultura, PCR panfúngica e PCR dirigida a Aspergillus. Do total de amostras estudadas, 23 foram positivas para fungos (PCR, cultura e/ou histologia). Das 46 amostras, 16 apresentaram resultados positivos para DNA fúngico. Em 12 amostras foi detetado um sinal positivo por PCR panfúngica e em 6 por PCR dirigida a Aspergillus (em 2 das amostras ocorreu deteção pelas duas metodologias). Em 61% (22/36) das amostras estudadas, houve concordância entre os métodos moleculares e culturais. Os agentes etiológicos identificados foram Candida albicans, C. glabrata, C. tropicalis, Trichosporon montevideense, Alternaria spp., Exophiala sp., Trichoderma sp., Histoplasma spp., Aspergillus fumigatus, Trichophyton rubrum e Paracoccidioides brasiliensis. Os resultados obtidos mostraram que a abordagem polifásica proposta parece ser uma boa estratégia na deteção de fungos em amostras de tecidos, permitindo eventualmente um melhor prognóstico. Em estudos posteriores, pretende-se estudar um maior número de amostras clinicas e implementar mais metodologias moleculares que permitam a deteção dirigida a determinados géneros fúngicos.Tissue specimens from patients with suspicion of having deep or subcutaneous fungal infections were analyzed at the National Reference Laboratory for Parasitic and Fungal Infections of INSA, according to a developed diagnostic algorithm comprising a polyphasic approach. This algorithm might allow a faster laboratory diagnosis of deep fungal infections. Fortysix tissue samples from 39 patients suspected of having deep or subcutaneous fungal infections were analyzed during a 26-month period (January 2015-February 2017) by the laboratory of INSA, using a laboratory approach that includes culture, panfungal PCR and Aspergillus-directed PCR. From the total samples studied, 23 were positive for fungi (PCR, culture and/or histology). From the 46 samples, 16 showed positive results for fungal DNA. In 12 samples a positive signal was detected by panfungal PCR and in 6 by Aspergillus-directed PCR (in 2 samples, there was positive detection using both methods). In 61% (22/36) of the samples studied, there was concordance between molecular and cultural methods. The etiological agents identified were Candida albicans, C. glabrata, C. tropicalis, Trichosporon montevideense, Alternaria spp., Exophiala sp., Trichoderma sp., Histoplasma spp., Aspergillus fumigatus, Trichophyton rubrum and Paracoccidioides brasiliensis. The results showed that the proposed polyphasic approach seems to be a good strategy for the detection of fungi in tissue samples, possibly allowing a better prognosis. In subsequent studies, it is intended to study a higher number of clinical samples and to establish more directed PCRs, allowing the detection of specific fungal genera.info:eu-repo/semantics/publishedVersio

Novel clinical and laboratorial challenges in Aspergillosis

© 2022 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).In recent years, research in the areas of Aspergillus and aspergillosis has continued to advance rapidly, including advancements in genomics, immunological studies, clinical areas, and diagnostic areas. Recently, new risk groups for the development of aspergillosis have emerged—patients with influenza- or COVID-19-ssociated pulmonary aspergillosis. The rise and spread of antifungal resistances have also become a clinical concern in some geographic areas and have drawn the attention of clinicians due to difficulties in treating these infections. In this paper, a snapshot of these issues is presented, emphasizing these novel clinical and laboratorial challenges in the aspergillosis field and focusing on their actual relevance.info:eu-repo/semantics/publishedVersio

The burden of serious fungal infections in Portugal

Using published data, we were able to estimate the incidence or prevalence of the above referred fungal infections and ~194 293 (1.8%) people in Portugal suffer from those fungal infections each year

Tonsillar ulceration as manifestation of disseminated African histoplasmosis in an immunocompetent Portuguese host

Free PMC Article: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5466580/Histoplasmosis is a systemic mycosis caused by Histoplasma capsulatum. Rare in Europe but endemic in some regions of Brazil, United States, Africa and Asia. Most of the cases are asymptomatic. Disseminated form is defined by the presence of an extra-pulmonary focus, particularly associated with immunosuppression. We report a case of an unilateral persisted tonsillar ulceration, in an immunocompetent Portuguese host, as manifestation of disseminated African histoplasmosis 45 years later after living 3 years in Africa.info:eu-repo/semantics/publishedVersio

Possible aflatoxin presence in Portuguese poultry units

Introduction: Aflatoxins are known to be human carcinogens based on sufficient evidence of carcinogenicity in humans (hepatocellular carcinoma, or primary liver-cell cancer). Aflatoxin B1 is one of the most deeply studied mycotoxins, known for a long time as belonging to the group of toxins produced by the genus Aspergillus (A. flavus, A. parasiticus, A. nominus). The presence in food stuffs depends on their geographical origin and production methods. Occupational exposure to aflatoxins can occur by inhalation of dust generated during the handling and processing of contaminated crops and feeds. Therefore, farmers and other agricultural workers have one of the greatest risks of occupational exposure to these mycotoxins. Objective: To characterize A. flavus prevalence in seven poultry units, with emphasis to the possible presence of aflatoxin in the air. Methods: A descriptive study was developed to monitor air fungal contamination in seven poultry units. Nineteen interior air samples of 25 litres were collected through impaction method. Results: From the seven poultry units analyzed, A. flavus was found in three of them. From all fungal genus identified in the referred units, A. flavus was the third species most frequently found in air samples (7.23%). Moreover, in those units, and from the Aspergillus genus, A. flavus was the most frequently isolated species in air samples (74.5%). Conclusions: Regarding the observed results and considering the high number of units contaminated by fungi known as possible aflatoxin producers, we have to believe that exposition can occur by inhalation (workers) and ingestion (consumers). This situation might represent a public health problem considering that aflatoxin is a known cancer agent