Waste Sorting Industry Research Articles

Are In Vitro Cytotoxicity Assessments of Environmental Samples Useful for Characterizing the Risk of Exposure to Multiple Contaminants at the Workplace? A Systematic Review.

In some occupational environments risk characterization is challenging or impossible to achieve due to the presence of multiple pollutants and contaminants. Thus, in vitro testing using the most relevant cell lines will provide information concerning health effects due to the co-exposure to multiple stressors. The aim of this review article is to identify studies where the cytotoxicity assessment was performed in environmental samples, as well as to describe the main outputs and challenges regarding risk characterization and management. This study is based on a study of the available information/data on cytotoxicity assessment performed on environmental samples following the PRISMA methodology. Different cell lines were used depending on the environment assessed and exposure routes implicated. The A549 alveolar epithelial cell line was applied in four studies for occupational exposure in the waste sorting industry and for outdoor environments; lymphocytes were used in two studies for occupational and outdoor environments; swine kidney cells were used in three studies performed in the waste industry and hepatocellular/Hep G2 in one study in the waste industry. Cytotoxicity assessments in environmental samples should have a more prominent role due to their contribution for identifying and better understanding the associations between co-exposure to environmental contaminants and adverse human health effects as a prioritization for risk management.

Cytotoxicity of Aspergillus Section Fumigati Isolates Recovered from Protection Devices Used on Waste Sorting Industry.

Safe working conditions must be guaranteed during waste sorting, which is crucial to maximizing recycling and reuse, in order to minimize workers’ exposure to chemical and biological hazards. This study determines the contribution of Aspergillus section Fumigati to the overall cytotoxicity of filtering respiratory protection devices (FRPD) and mechanic protection gloves (MPG) collected in 2019 from different workstations in one waste sorting industry in Portugal. The cytotoxicity of 133 Aspergillus section Fumigati isolates was determined as IC50 in human A549 epithelial lung cells and swine kidney cells, using the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. Aspergillus section Fumigati cytotoxicity results were compared with previous total cytotoxicity data from FRPD and MPG samples. A significant correlation was detected between the total cytotoxicity of samples and cytotoxicity of Aspergillus section Fumigati isolates in A549 cells (rS = −0.339, p = 0.030). The cytotoxicity of Aspergillus section Fumigati isolates explained 10.7% of the total cytotoxicity of the sample. On the basis of the comparison of cytotoxicity levels, it was possible to determine the contribution of Aspergillus section Fumigati isolates for the total cytotoxicity of protection devices used in the waste sorting industry. The results support in vitro toxicology as a relevant approach in risk assessments regarding cytotoxicity in passive sampling, and thus, useful in determining the contribution of relevant microbial contaminants to overall cytotoxicity. This approach can provide valuable answers in dose/response studies, and support innovations in risk characterization and their translation into occupational policies.

Cytotoxicity of filtering respiratory protective devices from the waste sorting industry: A comparative study between interior layer and exhalation valve

Filtering respiratory protection devices (FRPD) are mandatory for workers to wear in the Portuguese waste-sorting industry. Previous results regarding microbial contamination found on FRPD interior layer raised the question of whether microbial contamination from the exhalation valve would also have cytotoxicity effects. Since the FRPD exhalation valves are very close to workers’ nose and mouth, they represent a source of exposure to bioburden by inhalation. This study aimed to evaluate the cytotoxicity of the microbial contamination present in the FRPD exhalation valves. For this purpose, the cytotoxicity effects were determined through the MTT assay in two different cell lines (human A549 epithelial lung cells, and swine kidney cells) and compared with previous results obtained with FRPD interior layers. The contamination present in the FRPD exhalation valves presented some cytotoxicity on epithelial lung cells, suggesting the inhalation route as a potential route of exposure through the use of FRPD in the waste-sorting industry. Half-maximal (50%) inhibitory concentration (IC50) values were lower for FRPD interior layer than exhalation valves in lung cells, with overall cytotoxicity lower in exhalation valves when compared to interior layer (z = −4.455, p = 0.000). Higher bacterial counts in TSA were correlated with lower IC50 values, thus, higher cytotoxicity effect in lung cells. No statistically significant differences were detected among different workplaces.

Azole-Resistant Aspergillus fumigatus Harboring the TR34/L98H Mutation: First Report in Portugal in Environmental Samples.

Introduction: The frequency in detection of azole-resistant Aspergillus fumigatus isolates has increased since 2010. In Portugal, the section Fumigati is one of the most frequent, and resistant strains to have been found in clinical and environmental contexts. Although several cryptic species within the Fumigati section show intrinsic resistance to azoles, one factor driving (acquired) resistance is selective pressure deriving from the extensive use of azoles. This is particularly problematic in occupational environments where high fungal loads are expected, and where there is an increased risk of human exposure and infection, with impact on treatment success and disease outcome. The mechanisms of resistance are diverse, but mainly associated with mutations in the cyp51A gene. Despite TR34/L98H being the most frequent mutation described, it has only been detected in clinical specimens in Portugal. Methods: We analyzed 99 A. fumigatus isolates from indoor environments (healthcare facilities, spas, one dairy and one waste sorting unit) collected from January 2018 to February 2019 in different regions of Portugal. Isolates were screened for resistance to itraconazole, voriconazole and posaconazole by culture, and resistance was confirmed by broth microdilution. Sequencing of the cyp51A gene and its promoter was performed to detect mutations associated with resistance. Results: Overall, 8.1% of isolates were able to grow in the presence of at least one azole, and 3% (isolated from the air in a dairy and from filtering respiratory protective devices in a waste sorting industry) were pan-azole-resistant, bearing the TR34/L98H mutation. Conclusion: For the first time in Portugal, we report environmental isolates bearing the TR34/L98H mutation, isolated from occupational environments. Environmental surveillance of the emergence of azole-resistant A. fumigatus sensu stricto strains is needed, to ensure proper and timely implementation of control policies that may have a positive impact on public and occupational health.

Cytotoxic effect of filtering respiratory protective devices from the waste sorting industry: is in vitro toxicology useful for risk characterization?

The use of Filtering Respiratory Protective Devices (FRPD) is mandatory in Portugal to protect workers from the waste industry of harmful exposures. Deleterious health effects of exposure to bioburden via inhalation and/or ingestion include respiratory symptoms and nephrotoxicity. Between January and February 2019, 118 FRPD samples were collected in one waste sorting industry and characterized regarding microbial contamination and cytotoxicity, defined as cell metabolic activity, through the MTT colorimetric assay (3-(4,5-dimethylthiazol-2-yl)-2,5- diphenyltetrazolium bromide). Cytotoxic effect was classified according to percentage of extinction values with respect to the control group, as follows: absent (≥90); low (80%–90%, +); medium (60%–79%, ++); and high (below 60%, +++). For 113 samples the MTT assay revealed a cytotoxic effect in A549 cells, of which 81 presented high cytotoxicity. In SK cells, a cytotoxic effect was observed in 56 samples, of which five displayed a high cytotoxic effect. Several moderate (p < 0.05) to strong (p < 0.01) correlations were found between higher bacterial and fungal counts both in interior layers (fungi and bacteria) and in exhalation valves (fungi) of FRPD samples and reduced cell metabolic activity of SK cells. On the basis of the obtained results for the cytotoxic effect of FRPD samples on two different cells lines, it was determined that A549 cells exhibited a cytotoxic effect for a higher number of FRPD, whereas the SK cells model correlated better with the other assessed parameters, namely, bacterial and fungal counts and conditions of FRPD use. Although the results are not conclusive on the most appropriate cell line to assess FRPD cytotoxicity, they reinforce the importance of in vitro toxicology in exposure assessments to determine the cytotoxicity of mixtures of contaminants, for better risk characterization and selection of appropriate risk management measures.

Assessment of the microbial contamination of mechanical protection gloves used on waste sorting industry: A contribution for the risk characterization

In Portugal, mechanical protection gloves (MPG) are of mandatory use and during their use sweat is released and, consequently, the humidity of the material increases leading to conditions favorable to the growth of microorganisms. However, no studies have been conducted in MPG to assess the bioburden. This study intended to determine the bioburden present in MPG and their biological effects, and to discuss the possibility to use MPG as a passive method to assess occupational exposure to microbial contamination. Fungal burden was characterized through molecular tools for fungal toxigenic species, and antifungal resistance and mycotoxins profiles were determined. Cell viability was determined in swine kidney (SK) monolayer and hepatocellular carcinoma (Hep G2) cell lines. All MPG samples presented Gram-negative bacteria. The fungal contamination ranged from 0 CFU.m−2 in both MEA and DG18, to 5.09 × 106 and 2.75 × 106 and the most commonly fungi found was Aspergillus spp. (50.46%). Azole resistant Aspergillus sections were found in azole supplemented media. Aspergillus sections (Circumdati, Flavi, Fumigati and Versicolores) were detected by molecular tools in 66 out of 67 samples. The most reported mycotoxin was mycophenolic acid (89.6%). HepG2 cells appear to be more sensitive to MPG contamination, with high cytotoxicity (IC50 < 0.05 mm2/ml) observed for 18 out of 57 gloves.MPG can be used in passive sampling to assess occupational exposure to bioburden in waste sorting industries and contribute for risk characterization. Some contaminants of MPG had cytotoxic potential and affected the biology of hepatic cells more than renal cells.

Aspergillus spp. burden on filtering respiratory protective devices. Is there an occupational health concern?

There is scientific evidence that associates occupational exposure to fungi in the waste industry to health outcomes, being Aspergillus genera reported as the most prevalent. During Filtering Respiratory Protective Devices (FFR) use, humidity and temperature conditions provide favorable conditions for Aspergillus growth. The aim of this study was to characterize Aspergillus burden retained by FFR interior layer and exhalation valve. The sample consisted of 120 FFR and the correspondent interior layer and exhalation valves collected from different workstations from one waste sorting industry. The fungal content of exhalation valve and from the interior layer of the each FFR was extracted and inoculated on malt extract agar (MEA) supplemented with chloramphenicol (0.05%) and dichloran-glycerol agar (DG18). FFR extracts were also used for the molecular detection of Aspergillus sections Flavi and Fumigati. Valves presented Aspergillus spp. as the most prevalent genera on MEA media (44.4%), unlike what was observed in the FFR interior layer, where Aspergillus spp. was not the most prevalent genera (6.84% MEA; 5.2% DG18). Among the Aspergillus genera, section Fumigati was the one with highest prevalence, both in exhalation valves (76.57% MEA; 87.24% DG18) and interior layer (75.81% MEA; 51.22% DG18). qPCR analysis successfully amplified DNA from the Aspergillus sections Flavi and Fumigati. Interior layers presented statistically significant differences showing higher number of FFR contaminated with higher levels of Aspergillus in the workstations where workers contact directly with waste. This work, revealing strong fungal contamination of FFR used by workers at waste industry, clearly indicates that future trials to test FFR protective efficacy and establishment of times for FFR replacement should be performed.

Are workers from waste sorting industry really protected by wearing Filtering Respiratory Protective Devices? The gap between the myth and reality

One of the solutions for decreasing the workers' exposure to bioburden is the use of Filtering Respiratory Protective Devices (FRPD). As such it is important to determine whether these devices are fulfilling their protective role. This is the basis of the current study, aimed at characterizing bioburden retained by 120 FRPD (both in interior layers and in exhalation valves) through culture based-methods and molecular tools and also via analysis of antifungal resistance and mycotoxins profile. Our results show that Gram – Bacteria are present at a higher prevalence than total bacteria in both matrixes. Regarding fungal identification, Chrysonilia sitophila presented the highest prevalence on interior layers (55.1% on malt extract agar (MEA) supplemented with chloramphenicol (0.05%); 59.6% on dichloran-glycerol agar (DG18)), whereas on exhalation valves Aspergillus sp. presented the highest prevalence on MEA (6.8%) and C. sitophila on DG18 (36.3%). Among Aspergillus genera, section Fumigati was the one with the highest prevalence in both matrices. Aspergillus sp. was the most prevalent on exhalation valves (75.0% ITRA) in the screening of azole resistance. Fumigati section was the most abundant Aspergillus sp. detected on the interior layers (33.33%, 40 samples out of 120) and on the exhalation valves (1.66%, 2 samples out of 120). The interior layers and exhalation valves from workers with more waste contact showed an increased exposure to bioburden. This study showed that FRPD can have high levels of bioburden, toxigenic fungal strains and Aspergillus sections with reduced susceptibility to the tested azoles and can be used as a passive sampling method since it mimics the results obtained by active methods in previous studies. The gathered information will be useful to prioritize multiple interventions on workers’ education or even on FRPD replacement frequency.