Overview - ISO 13829:2000 (umu-test) for water quality genotoxicity
ISO 13829:2000 specifies a standardized laboratory procedure to determine the genotoxicity of water and wastewater using the umu-test. The assay detects induction of the bacterial SOS repair system via the umuC-gene (fused to lacZ), so genotoxic activity is measured as increased β‑galactosidase activity. The test uses the genetically modified bacterium Salmonella typhimurium TA1535/pSK1002 and can be run with or without metabolic activation (S9 fraction). Warning: the method involves GMOs and regulated reagents; work should be done by qualified labs under applicable licensing.
Key topics and technical requirements
- Test principle: Exposure of TA1535/pSK1002 to sample dilutions in microplates; after incubation (4 h), measure umuC induction via β‑galactosidase (lacZ) activity with ONPG substrate.
- Controls and calibration:
- Negative controls (blank, untreated bacteria) and solvent controls (DMSO).
- Positive controls (e.g., 4‑NQO, 2‑AA) to validate assay sensitivity; tests run both with and without S9 metabolic activation.
- Test organism & culture:
- Use Salmonella typhimurium TA1535/pSK1002 (ampicillin‑resistant plasmid).
- Stock cultures preserved at ≤ −80 °C in DMSO or glycerol; prepare overnight and pre‑cultures under specified conditions.
- Reagents & media:
- TGA culture medium, B‑buffer, ONPG, S9 fraction and cofactors (NADP, glucose‑6‑phosphate).
- Note safety: DMSO can form mutagenic products; protect photo‑sensitive reagents (2‑AA).
- Apparatus:
- 96‑well flat‑bottom microplates, microplate incubator/shaker, microplate photometer, pipettes, centrifuge and autoclave.
- Sample handling:
- Test fresh or store at 4 °C (analyse within 48 h) or ≤ −18 °C when necessary (ISO 5667‑16 guidance).
- Adjust sample pH to 7.0 ± 0.2 before incubation; avoid concentrating extracts or solvent extraction that may remove genotoxins.
Practical applications and users
- Who uses it:
- Environmental testing laboratories, water utilities, wastewater treatment facilities, regulatory bodies, and research labs assessing genotoxic contaminants.
- Use cases:
- Screening drinking water, surface water, and effluents for genotoxic potential.
- Routine monitoring, incident investigations, and pre‑treatment or post‑treatment assessments.
- Complementary to chemical analyses to detect unknown genotoxic agents.
- Benefits:
- Sensitive, microplate‑based assay that detects a broad range of DNA‑damaging agents and can incorporate metabolic activation to mimic in vivo transformation.
Related standards
- ISO 5667‑16 - Water quality - Sampling - Guidance on biotesting of samples (normative reference in ISO 13829)
Keywords: ISO 13829:2000, umu-test, genotoxicity testing, water quality, Salmonella typhimurium TA1535/pSK1002, S9 fraction, umuC, lacZ, β‑galactosidase, environmental monitoring.