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- Antifungal-resistant mucorales in different indoor environmentsPublication . Aranha Caetano, Liliana; Faria, Tiago; Springer, Jan; Loeffler, Juergen; Viegas, CarlaThis paper sought to address the prevalence of Mucorales in different indoor environments in Portugal. Environmental samples (183 in total) were collected at dwellings (n = 79) and workplaces (bakeries, swine farms, taxis, waste-sorting plants) (n = 93) by passive sampling using electrostatic dust collector (EDC), air-conditioning filters, litter, and/or raw materials. Samples were inoculated onto non-selective MEA and DG18 media and were screened for antifungal drug-resistance in azole-supplemented agar Sabouraud media. A probe-based Mucorales-specific real-time PCR assay (Muc18S) was used to detect Mucorales in complement to conventional culture-based methods. Mucorales order was found as more prevalent in air-conditioning filters from waste-sorting fork lifters (35.7%). Amongst Mucorales isolates able to grow in azole-supplemented media, 16 isolates of Mucor sp., Rhizopus sp. or Rhizomucor sp. were not susceptible to 1 mg/L voriconazole, and four isolates of Mucor sp. or Rhizopus sp. were not susceptible to 4 mg/L itraconazole. In conclusion, a combination of the culture-based and molecular methods proved to be reliable for Mucorales order identification in complex environmental samples.
- Worker’s nasal swab: a tool for occupational exposure assessment to bioburden?Publication . Viegas, Carla; Santos, V.; Moreira, R.; Faria, Tiago; Ribeiro, Edna; Caetano, Liliana Aranha; Viegas, SusanaThe nose cavity is the primary portal of entry for inspired air and the first region of the respiratory tract to be in contst with bioaerosols. Nasal swab allows measurement of bioburden presence in the nose cavity and the collection is easy and painless. We intended to identify scientific papers reporting this tool as a surrogate to access exposure to microbiologic agents in occupational environments. Literature research was performed using scientific and academic databases. In 5 from 11 articles studied only one parameter was analysed, being the most common Methicillin-resistant S. aureus. Sevem studies applied cutured based-methods coupled with molecular tolls essay. Findings from two studies coroborate the use of nasal swab as a tool to complement the occupational exposure assessements, since was found association between the nasal swabs results and the occupational microbiota also assessed. Nasal swabs analyses should comprehend culture based-methods and molecular tools assay.
- Cytotoxic and inflammatory potential of air samples from occupational settings with exposure to organic dustPublication . Viegas, Susana; Aranha Caetano, Liliana; Korkalainen, Merja; Faria, Tiago; Pacífico, Cátia; Carolino, Elisabete; Gomes, Anita Quintal; Viegas, CarlaOrganic dust and related microbial exposures are the main inducers of several respiratory symptoms. Occupational exposure to organic dust is very common and has been reported in diverse settings. In vitro tests using relevant cell cultures can be very useful for characterizing the toxicity of complex mixtures present in the air of occupational environments such as organic dust. In this study, the cell viability and the inflammatory response, as measured by the production of pro-inflammatory cytokines tumor necrosis factor-α (TNFα) and interleukin-1 β (IL-1β), were determined in human macrophages derived from THP-1 monocytic cells. These cells were exposed to air samples from five occupational settings known to possess high levels of contamination of organic dust: poultry and swine feed industries, waste sorting, poultry production and slaughterhouses. Additionally, fungi and particle contamination of those settings was studied to better characterize the organic dust composition. All air samples collected from the assessed workplaces caused both cytotoxic and pro-inflammatory effects. The highest responses were observed in the feed industry, particularly in swine feed production. This study emphasizes the importance of measuring the organic dust/mixture effects in occupational settings and suggests that differences in the organic dust content may result in differences in health effects for exposed workers.
- Characterization of occupational exposure to fungal burden in Portuguese bakeriesPublication . Viegas, Carla; Faria, Tiago; Aranha Caetano, Liliana; Carolino, Elisabete; Quintal-Gomes, Anita; Twarużek, Magdalena; Kosicki, Robert; Viegas, SusanaSeveral studies reported adverse respiratory health effects in workers exposed to ambient contaminants in bakeries. The aim of this study was to examine worker exposure to fungi and mycotoxins in Portuguese bakeries in order to develop new policies in occupational health. Environmental samples such as air, surfaces, settled dust and electrostatic dust collector (EDC) were collected in 13 bakeries for fungal and mycotoxins assessment. Air samples obtained by impaction were performed applying malt extract agar (MEA) supplemented with chloramphenicol (0.05%) and dichloran glycerol (DG18) agar-based media. Air samples collected through impinger method were determined as well for fungal detection by molecular tools of Aspergillus sections and mycotoxins. The highest median value for fungal load was 1053 CFU·m-3 and 65.3% (32 out of 49) of the sampling sites displayed higher fungal load than limits imposed by the World Health Organization. Aspergillus genera was found in air, surface swabs and EDC. Molecular tools were effective in measuring Aspergillus section Fumigati in 22.4% on air, 27.8% on surface swabs and in 7.4% in EDC and Aspergillus section Versicolores in one air sample. All settled dust samples showed contamination with six to eight mycotoxins in each sample. The mycotoxins detected were deoxynivalenol-3-glucoside, deoxynivalenol, zearalenone, 15-acetyldeoxynivalenol, monoacetoxyscirpenol, diacetoxyscirpenol, fumonisin B1, fumonisin B2, griseofulvin, HT2, ochratoxin A, ochratoxin B and mycophenolic acid. Industrial hygienists and exposure assessors should rely on different sampling methods (active and passive) and different assays (culture based and molecular methods) to obtain an accurate risk characterization regarding fungal burden (fungi and mycotoxins). Additionally, the awareness for the raw material as a potential mycotoxins indoor contamination source is important.
- Nasal swab as a tool to access occupational exposure to fungi in a cork industryPublication . Viegas, Carla; Pacífico, Cátia; Faria, Tiago; Aranha Caetano, Liliana; Gomes, Anita Quintal; Viegas, SusanaApproximately 49% of all cork produced worldwide in 2016 was from Portugal, where there are 650 companies working in this production sector. Additionally, two thirds of the world exportation comes from Portugal, with Portuguese cork industry employing about ten thousands workers (MTSS 2009). The presence of the Penicillium section Aspergilloides (formerly known as Penicillium glabrum) in this industry involves the risk of respiratory diseases such as suberosis, a type of hypersensitivity pneumonitis that is one of the most prevalent diseases among cork workers. Besides Penicillium section Aspergilloides, Aspergillus section Fumigati was also reported in cork industries. This is one of the most ubiquitous saprophytic fungi and is also considered one of the potentially pathogenic species with the highest clinical relevance. The use of the nasal swab procedure is of particular importance since it allows to determine the fungal presence in the nose cavity, being an easy and painless collection method. The aim of this study was to determine the exposure to the dominant mycobiota of this occupational environment through the mycological analysis of nasal exudates from cork industry workers. Nasal mucus samples from 360 workers from 3 companies (plant A - 41workers; plant B – 165 workers; and plant C – 154 workers) and also from a control group (38 individuals) with administrative tasks were performed. Duplicate samples were taken with sterilized cotton swabs from one nostril of each worker. The swabs were rotated against the internal anterior walls of the nostril and then placed in the provided transport tube. One of the swab samples was then plated onto malt extract agar (MEA) supplemented with chloramphenicol (0.05 %). All the collected samples were incubated at 27 °C for 5 to 7 days. The other swab sample was used for DNA extraction following molecular identification of Penicillium section Aspergilloides and Aspergillus section Fumigati by Real-Time PCR (qPCR). Among the 360 workers subjected to the nasal swab assay only 50 (13.9%) did not present fungal contamination. Around 36.6% of the workers' nasal swabs presented Penicillium genus contamination, 9.9% with Aspergillus sp. and 29.1% with more than one fungal genera. Among the 38 subjects from the control group, 16 (42.1%) did not present fungal contamination, 44.7% present Penicillium sp. and 18.4% Cladosporium sp. One subject presented Mucor sp. and other Geotrichum sp. contamination. DNA from Penicillium section Aspergilloides was successfully amplified by qPCR in 37 cork workers. From those, it was only possible to identify in 12 samples the genus Penicillium by culture based-methods. Aspergillus section Fumigati was also co-amplified with Penicillium section Aspergilloides in one worker, while in another one was detected singularly. As expected, in the 38 controls analyzed none were positive for Penicillium section Aspergilloides nor Aspergillus section Fumigati. The fungal species identified and detected in the collected nose swabs presented the same trend described for this very specific occupational environment. This approach allowed us to estimate the risk associated with the performance of the tasks since high dust contamination is expected to promote the exposure to fungi playing a role as carriers to the worker’s nose. As observed in previous environmental assessments, culture-based methods coupled with molecular tools allowed to obtain a wider spectrum of the workers' nasal mycobiota.