Background Wastewater treatment plant (WWTP) workers are exposed to biological hazards, including bacteria, fungi, and viruses with pathophysiological properties, posing a risk to workers’ health. Due to the complexity of WWTP bioaerosol composition, exposure assessment is challenging, and new methods are needed for a more holistic evaluation. Objectives (1) Characterize bacterial exposure in three Norwegian wastewater treatment plants (WWTPs), (2) Assess the bioaerosol’s and cultivated viruses’ ability to activate toll-like receptors in vitro, and (3) Elucidate adverse health effects in WWTP workers. Methods Inhalable bioaerosol fractions were collected with personal and stationary sampling, and endotoxin levels and 16S rRNA gene as a biomarker for bacteria were quantified. The inflammatory potential of the bioaerosol was assessed in vitro with HEK-Blue reporter cells, and we investigated the applicability of these cells for viral ligand detection by exposing them to cultivated Influenza A virus, Adenovirus F/40, E. coli phage MS2, and bovine Coronavirus. We used self-reported questionnaire data for an observational health assessment. Results TLR2 and TLR4 were activated in 85 and 91% of bioaerosol samples and activation correlated with airborne endotoxin levels. Airborne bacteria were present in all samples, with higher levels in personal (2.7 × 104 gc/m3) versus stationary samples (1.3 × 104 gc/m3). Endotoxin levels ranged from <LOD to 4800 EU/m3 in stationary, and <LOD to 920 EU/m3 in personal samples, with median levels of 54 and 47 EU/m3, respectively. Workers reported a higher prevalence of respiratory symptoms, skin dryness, diarrhea, and fever than unexposed controls. No bioaerosol sample activated TLR3 and 7, while moderate activation was observed by bovine Coronavirus. Significance Workers are continuously exposed to high levels of airborne bacteria levels of endotoxins exceeding 90 Endotoxin units/m3, which can induce a TLR2- or TLR4/NF-κB immune response. This exposure occurs throughout their workday, independent of workstations and workers’ tasks. Impact <UnorderedList Mark="Bullet"> <ItemContent> This study highlights the continuous exposure of WWTP workers to bioaerosols with pathophysiological properties, inducing TLR2- and TLR4-NF-κB immune responses. This effect occurs regardless of workstations and workers’ tasks, despite median endotoxin levels remaining below 90 EU/m³. The presence of airborne bacteria and bioaerosols with inflammatory potential may contribute to workers’ health symptoms, and our results highlight the importance of a holistic exposure assessment. We further demonstrate that HEK TLR reporter cell lines provide an innovative method to assess bioaerosol exposure, though they may underestimate exposure to viral-laden bioaerosols. </ItemContent> </UnorderedList>

TLR/NF-κB activation in vitro as a tool in the assessment of occupational bioaerosol exposure and health risks in wastewater treatment plants

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2026年4月20日

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Anna Jacobsen Lauvås,
Anne Straumfors,
Sarah Alsaedi,
Pål Graff,
Mette Myrmel,
Anani K. Afanou

摘要

Background

Wastewater treatment plant (WWTP) workers are exposed to biological hazards, including bacteria, fungi, and viruses with pathophysiological properties, posing a risk to workers’ health. Due to the complexity of WWTP bioaerosol composition, exposure assessment is challenging, and new methods are needed for a more holistic evaluation.

Objectives

(1) Characterize bacterial exposure in three Norwegian wastewater treatment plants (WWTPs), (2) Assess the bioaerosol’s and cultivated viruses’ ability to activate toll-like receptors in vitro, and (3) Elucidate adverse health effects in WWTP workers.

Methods

Inhalable bioaerosol fractions were collected with personal and stationary sampling, and endotoxin levels and 16S rRNA gene as a biomarker for bacteria were quantified. The inflammatory potential of the bioaerosol was assessed in vitro with HEK-Blue reporter cells, and we investigated the applicability of these cells for viral ligand detection by exposing them to cultivated Influenza A virus, Adenovirus F/40, E. coli phage MS2, and bovine Coronavirus. We used self-reported questionnaire data for an observational health assessment.

Results

TLR2 and TLR4 were activated in 85 and 91% of bioaerosol samples and activation correlated with airborne endotoxin levels. Airborne bacteria were present in all samples, with higher levels in personal (2.7 × 10⁴ gc/m³) versus stationary samples (1.3 × 10⁴ gc/m³). Endotoxin levels ranged from <LOD to 4800 EU/m³ in stationary, and <LOD to 920 EU/m³ in personal samples, with median levels of 54 and 47 EU/m³, respectively. Workers reported a higher prevalence of respiratory symptoms, skin dryness, diarrhea, and fever than unexposed controls. No bioaerosol sample activated TLR3 and 7, while moderate activation was observed by bovine Coronavirus.

Significance

Workers are continuously exposed to high levels of airborne bacteria levels of endotoxins exceeding 90 Endotoxin units/m³, which can induce a TLR2- or TLR4/NF-κB immune response. This exposure occurs throughout their workday, independent of workstations and workers’ tasks.

Impact

This study highlights the continuous exposure of WWTP workers to bioaerosols with pathophysiological properties, inducing TLR2- and TLR4-NF-κB immune responses. This effect occurs regardless of workstations and workers’ tasks, despite median endotoxin levels remaining below 90 EU/m³. The presence of airborne bacteria and bioaerosols with inflammatory potential may contribute to workers’ health symptoms, and our results highlight the importance of a holistic exposure assessment. We further demonstrate that HEK TLR reporter cell lines provide an innovative method to assess bioaerosol exposure, though they may underestimate exposure to viral-laden bioaerosols.