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Evaluating acute toxicity in enriched nitrifying cultures: Lessons learned
Carla Lopez1, Charles W Knapp1
1Centre for Water, Environment, Sustainability and Public Health, Department of Civil and Environmental Engineering, University of Strathclyde, Glasgow G1 1XJ, UK.
Journal of Microbiological Methods
|November 19, 2021
Summary
Experimental design critically impacts toxicological batch assays for nitrifying microorganisms. Minor changes in culture preparation, like excessive centrifugation or improper aeration, can skew results and affect toxicity endpoint determination.
Area of Science:
- Environmental Microbiology
- Ecotoxicology
- Wastewater Treatment
Background:
- Toxicological batch assays are vital for assessing contaminant effects on microorganisms, particularly in wastewater treatment plants (WWTPs).
- Nitrifying populations are sensitive indicators, but experimental design criteria for these assays are often overlooked.
- Deviations in culture preparation and conditions can significantly impact assay outcomes and the reliability of toxicological endpoints.
Purpose of the Study:
- To investigate how variations in experimental design affect nitrification activity in batch assays.
- To identify critical factors in culture preparation and assay conditions that influence toxicological results.
- To provide practical guidance for optimizing nitrifying batch assays and ensuring data accuracy.
Main Methods:
- Evaluated the impact of media and biomass preparation, including washing and centrifugation, on nitrification activity.
- Assessed the effects of aeration levels and nitrite build-up on assay performance.
- Tested the influence of organic solvents (DMSO, acetone) and culture concentration on nitrification inhibition.
- Compared the effects of different nutrient media compositions on nitrification capacity and toxicity responses.
Main Results:
- Extensive centrifugation destabilized nitrification activity, while proper media and biomass preparation are crucial for baseline conditions.
- Optimized aeration is necessary to prevent nitrite build-up and ammonia sparging, with DMSO and acetone suitable up to 0.03% (v/v).
- Dilute cultures showed more significant inhibition by allylthiourea (ATU), but biomass effects minimally impacted EC50 values.
- Switching mineral media reduced nitrification capacity; using original cultivation media is recommended.
Conclusions:
- Factors such as centrifugation, aeration, and media choice substantially impact nitrifying inoculum performance in acute bioassays.
- These overlooked experimental variables can significantly affect the determination of toxicological endpoints for ammonia-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria (NOB).
- The findings offer practical insights for improving the quality control and reliability of toxicological testing methodologies in environmental microbiology.
