Impact of Sublethal Levels of Environmental Pollutants Found in Sewage Sludge on a Novel Caenorhabditis elegans Model Biosensor

Abstract

A transgenic strain of the model nematode Caenorhabditis elegans in which bioluminescence reports on relative, whole-organism ATP levels was used to test an environmentally-relevant mixture of pollutants extracted from processed sewage sludge. Changes in bioluminescence, following exposure to sewage sludge extract, were used to assess relative ATP levels and overall metabolic health. Reproductive function and longevity were also monitored. A short (up to 8 h) sublethal exposure of L4 larval stage worms to sewage sludge extract had a concentration-dependent, detrimental effect on energy status, with bioluminescence decreasing to 50–60% of the solvent control (1% DMSO). Following longer exposure (22–24 h), the energy status of the nematodes showed recovery as assessed by bioluminescence. Continuous exposure to sewage sludge extract from the L4 stage resulted in a shorter median lifespan relative to that of solvent or medium control animals, but only in the presence of 400–600 µM 5-fluoro-2′-deoxyuridine (FUdR), which was incorporated to inhibit reproduction. This indicated that FUdR increased lifespan, and that the effect was counteracted by SSE. Exposure to sewage sludge extract from the L1 stage led to slower growth and a delayed onset of egg laying. When L1 exposed nematodes reached the reproductive stage, no effect on egg laying rate or egg number in the uterus was observed. DMSO itself (1%) had a significant inhibitory effect on growth and development of C. elegans exposed from the L1 stage and on reproduction when exposed from the L4 stage. Results demonstrate subtle adverse effects on C. elegans of a complex mixture of environmental pollutants that are present, individually, in very low concentrations and indicate that our biosensor of energy status is a novel, sensitive, rapid, quantitative, whole-organism test system which is suitable for high throughput risk assessment of complex pollutant mixtures.

A Screenable <em>In Vivo</em> Assay for Mitochondrial Modulators Using Transgenic Bioluminescent <em>Caenorhabditis elegans</em>

“…3 One of the compounds resulted in a substantial increase in GFP fluorescence consistent with the inhibitor increasing the levels of GFP-tagged firefly luciferase by binding to the luciferase enzyme's active site, and rendering it more stable. 15 In some instances (not in this particular case) this can lead to increased levels of bioluminescence when the inhibitor is displaced by excess substrate.…”

“…A number of luciferase inhibitors 15 are known and have been entered into PubChem. Therefore in the first instance, it is good practice to consult information on luciferase inhibitors held by PubChem; followed by testing of the compound effects in in vitro assays with the purified enzyme 3 and/or confirmation of effects on mitochondrial function by additional means. 16,17 GFP fluorescence measurements enable the detection of firefly luciferase levels (and the potential detection of some luciferase enzyme inhibitors as discussed above) making a strong case for measuring this endpoint alongside bioluminescence where possible.…”

“…The rationale is based on the transgenic expression of firefly luciferase in the nematode C. elegans [1][2][3] via the plasmid pSLGCV ( 1 https://www.addgene.org/49862). The luciferase enzyme is fused to the green fluorescent protein GFP and is expressed constitutively and ubiquitously in the cytoplasm (the luciferase peroxisome targeting signal was removed).…”

“…1 The bioluminescence strains generated were designated PE254(feIs4) and PE255(feIs5) 1 (see materials list) and can be used interchangeably. 3 A number of studies have utilized these sensor strains to report on cellular ATP levels in vivo following exposure to various xenobiotic chemicals such as sodium azide 1 , cadmium 2 , sewage sludge extract 3 , 5'-fluoro-2-deoxyuridine 6 , and a tobacco-specific nitrosamine 7 . The strains have also been useful to monitor effects of exposure to ultraviolet C radiation 8 and effects of disrupting the mitochondrial respiratory chain function.…”

Pollution by metals and toxicity assessment using Caenorhabditis elegans in sediments from the Magdalena River, Colombia

“…The later has been used as a model to assess toxicity of soils (Baderna et al, 2014;Harmon and Wyatt, 2008;H€ oss et al, 2009;Huguier et al, 2013), water (Ju et al, 2014), wastewater (Wang et al, 2008(Wang et al, , 2010, particulate matter (Sun et al, 2015;Zhao et al, 2014), sewage sludge (McLaggan et al, 2012), and river sediments (H€ oss et al, 2010;Tuikka et al, 2011;Wolfram et al, 2012). The transparency of its body, short life span, ability to self-fertilize and ease of being cultivated are advantages that make it ideal for use as a model in toxicology.…”