DIRECT ENUMERATION OF INJURED ESCHERICHIA-COLI-CELLS HARVESTED ONTO MEMBRANE FILTERS

Four methods testing either biosynthesis capacity (Direct Viable Count s), respiratory activity (CTC and XTT reduction assays), or membrane i ntegrity (LIVE/DEAD(R) BacLight(TM) VIABILITY kit) were performed to e numerate E. coli injured cells previously harvested onto 0.2-mu m blac k membranes. Some of them remained viable but were no longer culturabl e when exposed to hyperosmotic or oxidative stresses. The damaged cell ular functions were dependent on the stress applied, and the results p rovided by these methods (except the XTT reduction method which is not sensitive enough), gave information on cell behaviour to one stress, and on the different cellular targets of this stress. The osmotic shoc k in artificial seawater led to moderate plasma membrane damage (0.8 l og reduction in counts determined with the LIVE/DEAD(R) BacLight(TM) V IABILITY kit after 5 days in seawater), and to similar reductions of r espiration and elongation capacities (2.7 log reduction of CTC counts, and 2.6 log reduction of DVC counts, respectively). After a 30-min ox idative stress induced by peracetic acid (8 mg l(-1)), the plasma memb ranes remained intact (no reduction of LIVE/DEAD(R) BacLight(TM) VIABI LITY counts), and respiration was less impaired than elongation capaci ty (2.1 log reduction of CTC counts and 3.1 log reduction of DVC count s, respectively). (C) 1997 Elsevier Science B.V.