Bioluminescence-based assay used for toxicity monitoring

A new ATP bioluminescence-based method is proposed in order to examine the effect of toxic shock on microbial communities in urban wastewater treatment plant (WWTP). As the efficiency of WWTP must be improved, the occurrence and toxic effect of hazardous substances have to be evaluated (Directive 2455/2001/EC). Among bioluminescence assay with firefly luciferase is commonly used for intracellular ATP monitoring. A multienzyme systems catalysed by three enzymes (adenylate kinase, pyruvate kinase and firefly luciferase) for adenine nucleotides (AN = ATP+ ADP+ AMP) estimation is used for monitoring metabolic changes in activated sludge. The AN method provides better sensitivity than ATP quantification alone. The pool of intracellular and extracellular AN is constant during growth and the rate of intracellular AN is correlated with the microbial growth. In the present investigation, the extraction of total adenylate assay is optimized (chemical and mechanical extraction) and applied for monitoring the effect of the toxic shock (eg. Naphthalene) on indigenous biomass activity (intracellular and extracellular AN rates). In the same time, the removal of naphthalene is addressed Laboratory-scale reactors (4 L) are inoculated with urban activated sludge is under continuous-flow aeration. The reactor, containing non acclimated sludge enriched with naphthalene. The other reactor containing sterilised activated sludge is used as abiotic and volatilisation control system. In all this batch cultures, ATP and AN (intracellular and extracellular) rates and naphthalene concentrations are controlled. One of the objectives of this research is to determine a relation between the ATP/AN ratio and the biodegradation of naphthalene. Evidence is given which demonstrates that the ATP/AN parameter is a possible alternative for monitoring very rapid metabolic changes in complex microbial community such as activated sludge.