Biomonitoring of Indoor Air Fungal or Chemical Toxins with Caenorhabditis elegans nematodes

Abstract

Bad indoor air quality due to toxins and other impurities can have a negative impact on human well-being, working capacity and health. Therefore, reliable methods to monitor the health risks associated with exposure to hazardous indoor air agents are needed. Here, we have used transgenic Caenorhabditis elegans nematode strains carrying stress-responsive fluorescent reporters and evaluated their ability to sense fungal or chemical toxins, especially those that are present in moisture-damaged buildings. Liquid-based or airborne exposure of nematodes to mycotoxins, chemical agents or damaged building materials reproducibly resulted in time- and dose-dependent fluorescent responses, which could be quantitated by either microscopy or spectrometry. Thus, the C. elegans nematodes present an easy, ethically acceptable and comprehensive in vivo model system to monitor the response of multicellular organisms to indoor air toxicity.

Keywords: C. elegans; biomonitoring; chemicals; fungi; microbes; toxins.

Figure 1

Stress-responsive reporter strains sense fungal toxicity. Fluorescent responses of the sod-4::GFP strain to ethanol or water suspensions of Stachybotrys sp. (a/467) (A) or to ethanol suspensions of indicated Stachybotrys sp. strains (B), as analysed by spectrometry.

Figure 2

(A) Fluorescent responses of the cyp-34A9::GFP or sod-4::GFP strains to ethanol suspensions of Chaetomium sp. (a/459) or Stachybotrys sp. (a/467) prepared at 50 °C or 100 °C. (B) Fluorescent responses of the cyp-34A9::GFP strain to ethanol suspensions of several species of fungi: Trichoderma sp. (a/465), Paecilomyces variotii (a/462), Aspergillus ochraceus (Asp25), Aspergillus niger (a/464), Aspergillus fumigatus (a/466) and Stachybotrys sp. (a/468).

Figure 3

The C. elegans responses are dependent on mycotoxin dosage. (A) Effects of chaetoglobosin-containing Penicillium expansum (RcP61) exudates on fluorescent responses of the sod-4::GFP strain, as analysed by microscopy. Shown above are representative fluorescent images of individual nematodes. (B) Effects of chaetomin-containing samples (OT7) on fluorescent responses of the cyp-34A9::GFP strain, as analysed by spectrometry. Student’s t-test was used to analyse the statistical significance of the data. Significant differences (p < 0.05) as compared to control samples were marked with asterisks; ns refers to no significance. Error bars represent standard deviations.

Figure 4

Chemical toxins cause fluorescent responses in C. elegans, but also reduce their motility. (A) Time-dependent effects of glyoxal (0.2%) and methylglyoxal (0.04%) on fluorescent responses of the cyp-34A9::GFP strain, as analysed by spectrometry. Shown on the right are representative fluorescent images of nematode populations. (B) Dose-dependent effects of methylglyoxal on the motility of the sod-4::GFP strain, as analysed by video recordings. Student’s t-test was used to analyse the statistical significance of the data. Significant differences (p < 0.05) as compared to control samples were marked with asterisks. Error bars represent standard deviations. Shown on the right are representative brightfield images of nematode populations.

Figure 5

Representative images of the dose-dependent effects of DDDAC and Genapol X-080 on the fluorescent responses and motility of sod-4::GFP and cyp-34A9::GFP strains.

Figure 6

Airborne exposure to fungal cultures also causes fluorescent responses in C. elegans. Effects of Stachybotrys sp. exposure on fluorescent responses of the cyp-34A9::GFP strain, as analysed by microscopy (A) or spectrometry (B). (C) Relative effects of Wallemia sp. exposure on fluorescent responses of the sod-4::GFP strain, as analysed by microscopy. Student’s t-test was used to analyse the statistical significance of the data. Significant differences (p < 0.05) as compared to control samples were marked with asterisks; ns refers to no significance. Error bars represent standard deviations. Shown below are representative images of nematode populations. In those on the left, fluorescent images have been combined with brightfield images to highlight the localization of the fluorescence.

Figure 7

Moisture-damaged materials emit toxic volatile compounds. (A) Representative images of the fluorescent responses of the cyp-34A9::GFP strain to airborne exposure to control or moisture-damaged carpets, as analysed by microscopy, and the quantitation of them on the right. In chemotactic assays, the aversive effects of 2-ethyl-1-hexanol (B) or 1-octen-3-ol (C) on C. elegans were analysed using indicated dilutions in ethanol. Student’s t-test was used to analyse the statistical significance of the data. Significant differences (p < 0.05) as compared to control samples were marked with asterisks. Error bars represent standard deviations.

Figure 8

Experimental design. (A) Using shared fungal or chemical samples, the in vivo responses of the C. elegans nematodes were tested in a blinded fashion, and the results were compared to those previously obtained from in vitro PK-15 or BHK-21 cell proliferation assays and ex vivo sperm motility assays. (B) Our C. elegans bioassay is based on inducible green fluorescence produced upon liquid-based or airborne exposure to oxidative stress (with the sod-4 promoter) or xenobiotic stress (with the cyp-34A9 promoter).