Chlorination of the secondary effluent from domestic wastewater treatment plants (WWTP) is suspicious of toxic effects due to the formation of trihalomethanes (THMs). In order to clarify this mechanism, samples were collected from the inlet and outlet of a WWTP, and a series of analyses were conducted on the relationship between THMs formation and the toxicity of the secondary effluent after chlorination. By three-dimensional fluorescence spectrometry it was clarified that the biological wastewater treatment process was accompanied with the formation of humic- and fulvic-like substances which were typical THMs precursors as shown by their high thihalomethane formation potential (THMFP) as well as high specific ultraviolet absorbance (SUVA). As a result of bioassay using a freshwater luminescent bacterium Vibrio-qinghaiensis sp.-Q67, a linear relationship was obtained between the concentration of THMs after chlorinating the effluent and its toxicity expressed as the inhibition of the luminescent light. The effective THMs concentration corresponding to a 50% inhibition was 318 mu g/L at a chlorine dose of 19.6 mg/L as Cl-2. THMs formation may be the direct reason for the increase of toxicity in the secondary effluent when chlorine was used as disinfectant. (C) 2013 Elsevier B.V. All rights reserved.