Fungal contamination in two Portuguese wastewater treatment plants

The presence of filamentous fungi was detected in wastewater and air collected at wastewater treatment plants (WWTP) from several European countries. The aim of the present study was to assess fungal contamination in two WWTP operating in Lisbon. In addition, particulate matter (PM) contamination data was analyzed. To apply conventional methods, air samples from the two plants were collected through impaction using an air sampler with a velocity air rate of 140 L/min. Surfaces samples were collected by swabbing the surfaces of the same indoor sites. All collected samples were incubated at 27°C for 5 to 7 d. After lab processing and incubation of collected samples, quantitative and qualitative results were obtained with identification of the isolated fungal species. For molecular methods, air samples of 250 L were also collected using the impinger method at 300 L/min airflow rate. Samples were collected into 10 ml sterile phosphate-buffered saline with 0.05% Triton X-100, and the collection liquid was subsequently used for DNA extraction. Molecular identification of Aspergillus fumigatus and Stachybotrys chartarum was achieved by real-time polymerase chain reaction (RT-PCR) using the Rotor-Gene 6000 qPCR Detection System (Corbett). Assessment of PM was also conducted with portable direct-reading equipment (Lighthouse, model 3016 IAQ). Particles concentration measurement was performed at five different sizes: PM0.5, PM1, PM2.5, PM5, and PM10. Sixteen different fungal species were detected in indoor air in a total of 5400 isolates in both plants. Penicillium sp. was the most frequently isolated fungal genus (58.9%), followed by Aspergillus sp. (21.2%) and Acremonium sp. (8.2%), in the total underground area. In a partially underground plant, Penicillium sp. (39.5%) was also the most frequently isolated, also followed by Aspergillus sp. (38.7%) and Acremonium sp. (9.7%). Using RT-PCR, only A. fumigatus was detected in air samples collected, and only from partial underground plant. Stachybotrys chartarum was not detected in any of the samples analyzed. The distribution of particle sizes showed the same tendency in both plants; however, the partially underground plant presented higher levels of contamination, except for PM2.5. Fungal contamination assessment is crucial to evaluating the potential health risks to exposed workers in these settings. In order to achieve an evaluation of potential health risks to exposed workers, it is essential to combine conventional and molecular methods for fungal detection. Protective measures to minimize worker exposure to fungi need to be adopted since wastewater is the predominant internal fungal source in this setting.

Characterizing the fungal and bacterial microflora and concentrations in fitness centres

Fitness centres are special places where conditions for microbiological proliferation should be considered. Moisture due to human perspiration and water condensation as a result of human physical activities are prevalent in this type of buildings. Exposure to microbial contaminants is clinically associated with respiratory disorders and people who work out in polluted environments would be susceptible to contaminants. This work studied the indoor air contamination in three gymnasiums in Lisbon. The sampling was performed at two periods: at the opening (morning) and closing (night) of the three gymnasiums. The airborne bacterial and fungal populations were sampled by impaction directly onto Tryptic Soy Agar (for bacteria) and Malt Extract Agar (for fungi) plates, using a Merck MAS-100 air sampler. Higher bacterial concentrations were found at night as compared to the morning but the same behaviour was not found for fungal concentrations. Gram-negative catalase positive cocci were the dominant bacteria in indoor air samples of the studied gymnasiums. In this study, 21 genera/species of fungal colonies were identified. Chrysosporium sp., Chrysonilia sp., Neoscytalidium hialinum, Sepedonium sp. and Penicillium sp. were the most prevalent species identified in the morning, while Cladosporium sp., Penicillium sp., Chrysosporium sp., Acremonium sp. and Chrysonilia sp. were more prevalent at night. A well-designed sanitation and maintenance program for gymnasiums is needed to ensure healthier space for indoor physical activity.

Fungi distribution in poultry feed

Feed can easily be contaminated and colonized by fungi that use up the nutrients for their own metabolism and growth, producing secondary metabolites such as mycotoxins that are not eliminated throughout the feed processing. The major problems associated with mycotoxin contaminated animal feed are metabolic disturbances resulting in poor animal productivity. In addition, handling contaminated animal feed can also raise health issues regarding workers exposure to fungi and mycotoxins. The scope of this work was to characterize fungal distribution in 11 poultry feed samples. Twenty grams of feed were suspended in 180 mL of distilled water and homogenized during 20 minutes at 200 rpm. The washed supernatant was plated in malt extract agar (MEA) and dichloran glycerol agar base (DG18) media for morphological identification of the mycobiota present. Using macro- and microscopic analysis of the colonies, fungal contamination was evident in 72.7% of the analyzed poultry feed samples. Fungal load ranged from 0 to 13140 CFU/g, and the most prevalent species/genera were F. graminearum complex (71.1%), Penicillium sp. (11.6%), Cladosporium sp. (8.8%), and Fusarium poae (3.6%). In addition to these species, we also isolated Aspergillus sections Circumdati, Nigri and Aspergilli, and Mucor and Rhizopus genus albeit at a lower abundance. The data obtained showed that, besides high fungal contamination, mycotoxins contamination is probably a reality, particularly in the final product since mycotoxins resist to all the processing operations including thermal treatment. Additionally, data claimed attention for the probable co-exposure to fungi and mycotoxins of the workers in feed industries.

Microbiota assessment in optical shops: an ignored concern to public health

The presence of microorganisms in ophthalmic instruments and surfaces can lead to the exposure of patients to several infections. However, there is no information regarding fungal and bacteria contamination in optical shops. This study aims to characterize fungi and bacteria contamination in air and surfaces from 10 optical shops covering also ophthalmic instruments. Air samples were collected through an impaction method onto malt extract agar (MEA) supplemented with chloramphenicol (0.05%) used for fungi and Tryptic Soy Agar (TSA) supplemented with nystatin (0.2%) used for bacteria. Outdoor samples were also performed to be used as reference. Surface and equipment’s swab samples were also collected side-by-side. All the collected samples were incubated at 27ºC for 5 to 7 days (fungi) or at 30º for 7 days (bacteria). Regarding fungal distribution, thirteen different species/genera were found in the air, being the most common Alternaria sp. (62.0%). Eight different species/genera were identified in the surfaces, ranging from 2 to 5x104 CFU/m2, being the most common A. versicolor complex and Penicillium sp. (40.0%). The trial frames were the most contaminated equipment, since 50.0% of the collected samples were with countless colonies. The airborne bacterial population indicated higher concentrations in the contactology office (average: 133 CFU/m3) than in the client’s waiting rooms (average: 126 CFU/m3). The surface samples indicated bacterial concentrations ranging from 2x104 to 1x106 CFU/m2, pointing out the automatic refractometer as the surface with higher bacterial load.

Exposição ocupacional a bioaerossóis durante atividades de limpeza de quartos de hotel

O presente estudo pretende avaliar a exposição ocupacional a contaminação fúngica e bacteriana em quartos de hotel, mais precisamente em dois quartos com características diferentes, nomeadamente, com pavimento em alcatifa e outro sem alcatifa. Doze amostras de ar de 250L foram colhidas pelo método de impacto, em meio agar de extracto de malte (MEA) suplementado com cloranfenicol (0,05%) para fungos e em meio de TSA (agar de soja tríptica) com nistatina (0,2%) para bactérias. Foram também realizadas amostras de superfície nos mesmos locais. Em ambos os quartos apenas uma amostra de ar, no quarto sem alcatifa, apresentou contagens de fungos mais elevadas do que no exterior. No entanto, as concentrações de bactérias no ar interior foram superiores às do ar exterior. Em relação às superfícies, o quarto sem alcatifa apresentou diferenças estatisticamente significativas em relação ao quarto com alcatifa, sendo que o primeiro apresentou concentrações mais elevadas de fungos. Todas as superfícies analisadas apresentaram contaminação bacteriana, mas não houve diferenças estatisticamente significativas entre os quartos. Os géneros de fungos mais prevalentes no ar foram idênticos em ambos os quartos (Penicillium sp. 40,7% - 12,3% e Cladosporium sp. 43,5% - 55,4%). Nas superfícies analisadas, os isolados pertencentes ao complexo Aspergillus fumigatus foram os únicos encontrados no quarto com alcatifa, enquanto no outro quarto os géneros mais prevalentes foram Penicillium sp. (63,6%) e Aspergillus sp. (13,6%).

Fungi spores dimension matters in health effects: a methodology for more detail fungi exposure assessment

Health effects resulting from dust inhalation in occupational environments may be more strongly associated with specific microbial components, such as fungi, than to the particles. The aim of the present study is to characterize the occupational exposure to the fungal burden in four different occupational settings (two feed industries, one poultry and one waste sorting industry), presenting results from two air sampling methods – the impinger collector and the use of filters. In addition, the equipment used for the filter sampling method allowed a more accurate characterization regarding the dimension of the collected fungal particles (less than 2.5 μm size). Air samples of 300L were collected using the impinger Coriolis μ air sampler. Simultaneously, the aerosol monitor (DustTrak II model 8532, TSI®) allowed assessing viable microbiological material below the 2.5 μm size. After sampling, filters were immersed in 300 mL of sterilized distilled water and agitated for 30 min at 100 rpm. 150 μl from the sterilized distilled water were subsequently spread onto malt extract agar (2%) with chloramphenicol (0.05 g/L). All plates were incubated at 27.5 ºC during 5–7 days. With the impinger method, the fungal load ranged from 0 to 413 CFU.m-3 and with the filter method, ranged from 0 to 64 CFU.m-3. In one feed industry, Penicillium genus was the most frequently found genus (66.7%) using the impinger method and three more fungi species/genera/complex were found. The filter assay allowed the detection of only two species/genera/complex in the same industry. In the other feed industry, Cladosporium sp. was the most found (33.3%) with impinger method and three more species/genera/complex were also found. Through the filter assay four fungi species/genera/complex were found. In the assessed poultry, Rhyzopus sp. was the most frequently detected (61.2%) and more three species/genera/complex were isolated. Through the filter assay, only two fungal species/genera/complex were found. In the waste sorting industry Penicillium sp. was the most prevalent (73.6%) with the impinger method, being isolated two more different fungi species/genera/complex. Through the filter assay only Penicillium sp. was found. A more precise determination of occupational fungal exposure was ensured, since it was possible to obtain information regarding not only the characterization of fungal contamination (impinger method), but also the size of dust particles, and viable fungal particles, that can reach the worker ́s respiratory tract (filters method). Both methods should be used in parallel to enrich discussion regarding potential health effects of occupational exposure to fungi.

Tratamento de cor em pós-produção televisiva: o caso da produtora de conteúdos SP Televisão

Dissertação apresentada à Escola Superior de Comunicação Social como parte dos requisitos para obtenção de grau de mestre em Audiovisual e Multimédia.