Short communication: Rapid <i>in vitro</i> tests to determine the toxicity of raw wastewater and treated sewage effluents

Abstract

Wastewater consists of a complex mixture of substances. During wastewater treatment these harmful substances can be eliminated or degraded. However, persistent compounds released with the treated sewage effluents enter the environment and pose a risk to animal and human life. To determine the potential risks involved, screening tests are needed to monitor wastewater for potential toxic contaminants. The aim of this study was to validate and use screening tests to determine the toxicity of raw wastewater and treated sewage effluents from 3 sewage treatment plants in the Western Cape, South Africa. Raw wastewater and treated sewage effluents were screened for cytotoxicity using lactate dehydrogenase (LDH) release from cells as biomarker, for neurotoxicity using acetylcholinesterase (AChE) inhibition and for genotoxicity using the Save Our Soul (SOS) test. Results showed no cytotoxicity for both raw wastewater and treated sewage effluents from all sewage treatment plants. Raw wastewater from all sewage treatment plants contained AChE inhibitors and sewage treatment processes were not effective at eliminating these AChE inhibitors. Raw wastewater from all sewage treatment plants tested positive for genotoxicity. Treated sewage effluents from all three sewage treatment plants displayed no genotoxicity indicating effective removal of genotoxins by all three sewage treatment plants investigated.

The assessment of inflammatory activity and toxicity of treated sewage using RAW264.7 cells

“…None of the water samples tested resulted in significantly different WST‐1‐ or XTT‐based metabolic assay compared to controls, which implies that samples had no toxic effects to RAW264.7 cells. Lack of toxicity of wastewater or contaminated water has been reported in previous studies (Pool et al ., ; Hendricks and Pool ). This observation may be attributed to lower concentration of pollutants required to induce immunotoxicity as compared to that required for induction of cell death (Hymery et al .…”

In vitro acetylcholinesterase inhibitory activity and chemical composition of the dichloromethane extracts of the leaves and inflorescences of the tumbleweed, Trachyandra laxa (N.E.Br.) Oberm. var. rigida (Suesseng.) Roessl

“…The in vitro AChE inhibition assay was performed on the different extracts by measuring optical densities (ODs) at different time intervals to monitor the change in acetylcholine concentration, using a method adapted from the one used by Hendricks and Pool (Hendricks and Pool, 2012). The assay was performed in triplicate.…”

“…A dilution of each extract was prepared in phosphate buffer to obtain final concentrations of 0.1 mg/mL. The positive control consisted of 0.1 mM chlorpyrifos, an organophosphate insecticide that inhibits acetylcholinesterase, prepared in phosphate buffer (Hendricks and Pool, 2012;Rhee et al, 2013). The negative control, 10% DMSO, was also prepared in phosphate buffer, and serial dilutions of both the positive and negative controls were performed.…”

“…When compounds inhibit AChE enzymes and cause a disruption of neurotransmission, they are essentially acting as neurotoxins ( Colovi c et al, 2013). Therefore, in vitro AChE inhibition assays are commonly used for the determination of the presence of neurotoxins (Hendricks and Pool, 2012;Lionetto et al, 2013). AChE hydrolyses the substrate acetylcholine (ACh) at the membrane of mammalian neurons and, while predominantly associated with the neuromuscular junction and cholinergic synapses in the central nervous system (CNS), it is also found in the blood where it is associated with erythrocytes, lymphocytes, megakaryocytes, and platelets (Husain, 1994;Paulus et al, 1981;Zajicek, 1957).…”

“…For instance, the serine in the active site of AChE-E produces a covalent bond with an organophosphate pesticide in the same way as the AChE isoforms found in the nervous system (Dafferner et al, 2017). Consequently, one version of the AChE assay is performed by using AChE extracted from blood (Ellman et al, 1961;Hendricks and Pool, 2012). Although the AChE used in this case is mostly the erythrocyte isoform it will still provide an indication of neurotoxicity (Dafferner et al, 2017;Gupta et al, 2015).…”