New approach methodologies to enhance human health risk assessment of immunotoxic properties of chemicals - a PARC (Partnership for the Assessment of Risk from Chemicals) project.

As a complex system governing and interconnecting numerous functions within the human body, the immune system is unsurprisingly susceptible to the impact of toxic chemicals. Toxicants can influence the immune system through a multitude of mechanisms, resulting in immunosuppression, hypersensitivity, increased risk of autoimmune diseases and cancer development. At present, the regulatory assessment of the immunotoxicity of chemicals relies heavily on rodent models and a limited number of Organisation for Economic Co-operation and Development (OECD) test guidelines, which only capture a fraction of potential toxic properties. Due to this limitation, various authorities, including the World Health Organization and the European Food Safety Authority have highlighted the need for the development of novel approaches without the use of animals for immunotoxicity testing of chemicals. In this paper, we present a concise overview of ongoing efforts dedicated to developing and standardizing methodologies for a comprehensive characterization of the immunotoxic effects of chemicals, which are performed under the EU-funded Partnership for the Assessment of Risk from Chemicals (PARC).

Occupational exposure to inhalable pathogenic microorganisms in waste sorting.

This study assessed microorganisms in personal inhalable work air samples aiming to identify potential human pathogens, and correlate exposure to adverse health outcomes in waste workers. Full-shift personal exposure was measured in six different waste sorting plants. Microbial concentrations in inhalable air samples were analysed using MALDI-TOF MS for cultivable, and next generation sequencing (NGS) for non-cultivable microorganisms. Concentrations of bacterial and fungal CFUs varied substantially within and between waste sorting plants, ranging from no identifiable organisms to a maximum concentration in the order of 105 CFU/m3. Bacillus and Staphylococcus were among the most abundant bacterial genera, whilst fungal genera were dominated by Aspergillus and Penicillium. Approximately 15% of all identified species were human pathogens classified in risk group 2, whereas 7% belonged to risk group 1. Furthermore, significant correlations between concentrations of fungi in risk group 1 and self-reported adverse symptoms, such as wheezing were identified in exposed workers. The combination of culture-based methods and NGS facilitated the investigation of infectious microbial species with potential pathophysiological properties as well as non-infectious biological agents in inhalable work air samples and thereby contributed to the risk assessment of occupational exposure in waste sorting.

Bioaerosol-induced in vitro activation of toll-like receptors and inflammatory biomarker expression in waste workers.

PURPOSE: Occupational exposure to bioaerosols during waste handling remains a health concern for exposed workers. However, exposure-related health effects and underlying immunological mechanisms are still poorly described. METHODS: The present study assessed the inflammatory potential of work-air samples (n = 56) in vitro and investigated biomarker expression in exposed workers (n = 69) compared to unexposed controls (n = 25). These quantitative results were compared to self-reported health conditions. RESULTS: Personal air samples provoked an activation of TLR2 and TLR4 HEK reporter cells in one-third of all samples, indicating that the work environment contained ligands capable of inducing an immune response in vitro. Monocyte levels, as well as plasma biomarker levels, such as IL-1Ra, IL-18 and TNFα were significantly higher in exposed workers, compared to the control group when confounding factors such as BMI, sex, age and smoking habits were accounted for. Furthermore, a significant exposure-related increase in midweek IL-8 levels was measured among exposed workers. Tendencies of increased prevalence of health effects of the respiratory tract were identified in exposed workers. CONCLUSION: Inhalable dust provoked TLR activation in vitro, indicating that an exposure-related immune response may be expected in susceptible workers. However, despite significant differences in inflammatory plasma biomarker levels between exposed and unexposed workers, prevalence of self-reported health effects did not differ between the groups. This may be due to the healthy worker effect, or other factors such as adequate use of personal protective respiratory devices or adaptation to the work environment with reduced activation of the immune system.

An assessment of occupational exposure to bioaerosols in automated versus manual waste sorting plants.

Occupational exposure during waste sorting is associated with several health outcomes. This study obtained knowledge about the impact of work in fully automated waste sorting plants (AWSP; n = 3) vs manual waste sorting plants (MWSP; n = 3) on personal exposure (n = 71) to bioaerosols and exposure-related health effects. Personal full-shift air samples were collected using various filter-based active sampling devices that were placed in the workers' breathing zone. Personal exposure to inhalable and thoracic dust, endotoxin and microorganisms varied considerably between and within types of waste sorting plants (WSP). Workers at AWSP were on average exposed to 0.34 mg/m3 inhalable dust, 0.15 mg/m3 thoracic dust, and 51 EU/m3 endotoxins (geometric mean (GM) levels), whereas GM exposure levels at MWSP were 0.66 mg/m3 for inhalable dust, 0.44 mg/m3 for thoracic dust, and 32 EU/m3 for endotoxins. Exposure to submicronic fungal fragments did not differ between types of plants and ranged from levels below the detection limit (limit of detection, LOD) to levels in the order of 106 fragments/m3. Higher levels of fungal fragments and fungal spores were found at AWSP compared to MWSP with a GM of 2.1 × 105 spores/m3and with a GM of 1.2 × 105 spores/m3, respectively. Actinobacterial spores were found in samples from AWSP only, with exposure levels ranging from 1.9 × 104 to 1.1 × 107 spores/m3. Exposure to microbial DNA varied within and between WSP and was on average in the order of 104 copies/m3 for fungi and 105 copies/m3 for bacteria. Health symptoms, such as sneezing, congested nose and runny nose were significantly more common among exposed workers compared to the unexposed control group.

Comprehensive assessment of occupational exposure to microbial contamination in waste sorting facilities from Norway.

Introduction: It is of upmost importance to contribute to fill the knowledge gap concerning the characterization of the occupational exposure to microbial agents in the waste sorting setting (automated and manual sorting). Methods: This study intends to apply a comprehensive field sampling and laboratory protocol (culture based-methods and molecular tools), assess fungal azole resistance, as well as to elucidate on potential exposure related health effects (cytotoxicity analyses). Skin-biota samples (eSwabs) were performed on workers and controls to identify other exposure routes. Results: In personal filter samples the guidelines in one automated industry surpassed the guidelines for fungi. Seasonal influence on viable microbial contamination including fungi with reduced susceptibility to the tested azoles was observed, besides the observed reduced susceptibility of pathogens of critical priority (Mucorales and Fusarium sp.). Aspergillus sections with potential toxigenic effect and with clinical relevance were also detected in all the sampling methods. Discussion: The results regarding skin-biota in both controls´ and workers´ hands claim attention for the possible exposure due to hand to face/mouth contact. This study allowed concluding that working in automated and manual waste sorting plants imply high exposure to microbial agents.

Circulating miRNAs as molecular markers of occupational grain dust exposure.

Dust from grain and feed production may cause adverse health effects in exposed workers. In this study we explored circulating miRNAs as potential biomarkers of occupational grain dust exposure. Twenty-two serum miRNAs were analyzed in 44 grain dust exposed workers and 22 controls. Exposed workers had significantly upregulated miR-18a-5p, miR-124-3p and miR-574-3p, and downregulated miR-19b-3p and miR-146a-5p, compared to controls. Putative target genes for the differentially expressed miRNAs were involved in a range of Kyoto Encyclopedia of Genes and Genomes signaling pathways, and 'Pathways in cancer' and 'Wnt signaling pathway' were common for all the five miRNAs. MiRNA-diseases association analysis showed a link between the five identified miRNAs and several lung diseases terms. A positive correlation between miR-124-3p, miR-18a-5p, and miR-574-3p and IL-6 protein level was shown, while miR-19b-3p was inversely correlated with CC-16 and sCD40L protein levels. Receiver-operating characteristic analysis of the five miRNA showed that three miRNAs (miR-574-3p, miR-124-3p and miR-18a-5p) could distinguish the grain dust exposed group from the control group, with miR-574-3p as the strongest predictor of grain dust exposure. In conclusion, this study identified five signature miRNAs as potential novel biomarkers of grain dust exposure that may have potential as early disease markers.

Pneumoproteins and markers of inflammation and platelet activation in the blood of grain dust exposed workers.

PURPOSE: To investigate if blood biomarkers could indicate early signs of lung damage or cardiovascular risk due to exposure to grain dust. MATERIALS AND METHODS: Pneumoproteins and markers of inflammation and platelet activation were analysed in blood samples of 102 grain elevator and compound feed mill workers. Differences between exposed (n = 67) and controls (n = 35), and associations with exposure measurements and respiratory health were investigated by multiple linear regression analyses. RESULTS: Concentrations of CC-16 and IL-6 were higher in exposed workers compared with controls (p < 0.001 for both), whereas fibrinogen was lower (p = 0.005). Concentrations of CRP, TNF-α, sCD40L and sP-selectin were similar in both groups. Serum CC-16 was significantly higher in workers with farm childhood, regardless of exposure. The impact of farm childhood on CC-16 interacted with smoking. None of the biomarkers were associated with exposure measurements or any of the tested respiratory health parameters. CONCLUSION: Dust exposure induced inflammatory and anti-inflammatory reactions, but did not induce systemic inflammation and had no effect on platelet activation. No cause-effect relationship could be established in spite of relatively high exposure levels, particularly to endotoxin. Whether increased serum CC-16 is an early sign of lung damage or a reversible defense reaction remains unclear.

Exposure to Wood Dust, Microbial Components, and Terpenes in the Norwegian Sawmill Industry.

Sawmill workers are exposed to wood dust (a well-known carcinogen), microorganisms, endotoxins, resin acids (diterpenes), and vapours containing terpenes, which may cause skin irritation, allergy, and respiratory symptoms including asthma. The health effects of most of these exposures are poorly understood as most studies measure only wood dust. The present study assessed these exposures in the Norwegian sawmill industry, which processes predominantly spruce and pine. Personal exposures of wood dust, resin acids, endotoxin, fungal spores and fragments, mono-, and sesquiterpenes were measured in 10 departments in 11 saw and planer mills. The geometric mean (GM) and geometric standard deviation (GSD) thoracic exposures were: 0.09 mg m-3 dust (GSD 2.6), 3.0 endotoxin units (EU) m-3 (GSD 4.9), 0.4 × 105 fungal spores m-3 (GSD 4.2), 2 × 105 fungal fragments m-3 (GSD 3.2), and 1560 ng m-3 of resin acids (GSD 5.5). The GM (GSD) inhalable exposures were: 0.72 mg m-3 dust (2.6), 17 EU m-3 (4.3), 0.4 × 105 fungal spores m-3 (3.8), and 7508 ng m-3 (4.4) of resin acids. The overall correlation between the thoracic and inhalable exposure was strong for resin acid (rp = 0.84), but moderate for all other components (rp = 0.34-0.64). The GM (GSD) exposure to monoterpenes and sesquiterpenes were 1105 µg m-3 (7.8) and 40 µg m-3 (3.9), respectively. Although mean exposures were relatively low, the variance was large, with exposures regularly exceeding the recommended occupational exposure limits. The exposures to spores and endotoxins were relatively high in the dry timber departments, but exposures to microbial components and mono-and sesquiterpenes were generally highest in areas where green (undried) timber was handled. Dust and resin acid exposure were highest in the dry areas of the sawmills. Low to moderate correlation between components (rp ranging from 0.02 to 0.65) suggests that investigations of exposure-response associations for these components (both individually and combined) are feasible in future epidemiological studies.

Cross-shift study of exposure-response relationships between bioaerosol exposure and respiratory effects in the Norwegian grain and animal feed production industry.

OBJECTIVE: We have studied cross-shift respiratory responses of several individual bioaerosol components of the dust in the grain and feed industry in Norway. METHODS: Cross-shift changes in lung function and nasal congestion, as well as in respiratory and systemic symptoms of 56 exposed workers and 36 referents, were recorded on the same day as full-shift exposure to the inhalable aerosol fraction was assessed. Exposure-response associations were investigated by regression analysis. RESULTS: The workers were exposed on average to 1.0 mg/m(3) of grain dust, 440 EU/m(3) of endotoxin, 6 µg/m(3) of ß-1,3-glucans, 17×10(4)/m(3) of bacteria and 4×10(4)/m(3) of fungal spores during work. The exposure was associated with higher prevalence of self-reported eye and airway symptoms, which were related to the individual microbial components in a complex manner. Fatigue and nose symptoms were strongest associated with fungal spores, cough with or without phlegm was associated with grain dust and fungal spores equally strong and wheeze/tight chest/dyspnoea was strongest associated with grain dust. Bioaerosol exposure did not lead to cross-shift lung function decline, but several microbial components had influence on nose congestion. CONCLUSIONS: Exposure to fungal spores and dust showed stronger associations with respiratory symptoms and fatigue than endotoxin exposure. The associations with dust suggest that there are other components in dust than the ones studied that induce these effects.