TOXIC EFFECTS ON BACTERIAL METABOLISM OF THE REDOX DYE 5-CYANO-2,3-DITOLYL TETRAZOLIUM CHLORIDE

The monotetrazolium redox dye 5-cyano-2,3-ditolyl tetrazolium chloride (CTC) has been used as a vital stain of actively respiring bacteria f or several years, In this study, inhibitory effects on bacterial metab olism of this redox dye have been examined in a brackish water environ ment (Kiel Fjord, Germany) and a freshwater environment (Elbe River, G ermany), As the results from time series experiments (1 to 10 h) show, bacterial growth and respiration of the investigated natural communit ies were clearly reduced by CTC supply, Compared with untreated contro ls (100%), CTC-treated samples showed distinctly lower heterotrophic b acterial plate counts (0 to 24 and 11.8 to 23.7%, respectively), bacte rial production (0.9 to 14.1 and 1.1 to 9.6%, respectively), bacterial respiration (4.1 to 9.4 acid 6.8 to 43.8% for several concentrations of C-14-labeled glucose), and [C-14]glucose incorporation (0.2 to 4.2% ), Additionally, toxicity of CTC was demonstrated by luminescence in a Microtox bioassay, CTC concentrations of 0.1 and 5.0 mu M required on ly 15 min for decreases of approximately 50 and 100%, respectively, Th e suppression of CTC on several bacterial metabolic processes suggests that determination by the CTC technique underestimates the actual num ber of active cells distinctly, This conclusion is confirmed by the co mparison of generation times calculated on the basis of thymidine upta ke data and active bacterial counts determined by the CTC assay and mi croautoradiography. While unrealistic short generation times (0.5 to 5 h) resulted from the CTC assay, the generation times calculated accor ding to microautoradiography ranged within values (7 to 21 h) reported elsewhere for comparable aquatic environments, The inhibitory effect of CTC demonstrated in our experiments is an aspect with regard to the application of this tetrazolium dye for the estimation of active bact eria in natural aquatic environments which hitherto has not been consi dered.