Flow-cytometric study of vital cellular functions in Escherichia coli during solar disinfection (SODIS).
The effectiveness of solar disinfection (SODIS), a low-cost household water treatment method for developing countries, was investigated with flow cytometry and viability stains for the enteric bacterium Escherichia coli. A better understanding of the process of injury or death of E. coli during SODIS could be gained by investigating six different cellular functions, namely: efflux pump activity (Syto 9 plus ethidium bromide), membrane potential [bis-(1,3-dibutylbarbituric acid)trimethine oxonol; DiBAC4(3)], membrane integrity (LIVE/DEAD BacLight), glucose uptake activity (2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]-2-deoxy-d-glucose; 2-NBDG), total ATP concentration (BacTiter-Glo) and culturability (pour-plate method). These variables were measured in E. coli K-12 MG1655 cells that were exposed to either sunlight or artificial UVA light. The inactivation pattern of cellular functions was very similar for both light sources. A UVA light dose (fluence) of <500 kJ m(-2) was enough to lower the proton motive force, such that efflux pump activity and ATP synthesis decreased significantly. The loss of membrane potential, glucose uptake activity and culturability of >80 % of the cells was observed at a fluence of approximately 1500 kJ m(-2), and the cytoplasmic membrane of bacterial cells became permeable at a fluence of >2500 kJ m(-2). Culturable counts of stressed bacteria after anaerobic incubation on sodium pyruvate-supplemented tryptic soy agar closely correlated with the loss of membrane potential. The results strongly suggest that cells exposed to >1500 kJ m(-2) solar UVA (corresponding to 530 W m(-2) global sunlight intensity for 6 h) were no longer able to repair the damage and recover. Our study confirms the lethal effect of SODIS with cultivation-independent methods and gives a detailed picture of the 'agony' of E. coli when it is stressed with sunlight.
Berney
Michael
M
Swiss Federal Institute of Aquatic Science and Technology (EAWAG), PO 611, CH-8600 Dübendorf, Switzerland.
Weilenmann
Hans-Ulrich
HU
Swiss Federal Institute of Aquatic Science and Technology (EAWAG), PO 611, CH-8600 Dübendorf, Switzerland.
Egli
Thomas
T
Swiss Federal Institute of Aquatic Science and Technology (EAWAG), PO 611, CH-8600 Dübendorf, Switzerland.
D001426:Bacterial Proteins / Q000378:metabolism; D004203:Disinfection / Q000379:methods*; D004926:Escherichia coli / Q000254:growth & development / Q000378:metabolism / Q000528:radiation effects*; D005434:Flow Cytometry / Q000379:methods*; D005947:Glucose / Q000378:metabolism; D008564:Membrane Potentials; D009930:Organic Chemicals / Q000378:metabolism; D013472:Sunlight; D014466:Ultraviolet Rays; D014871:Water Microbiology; D018508:Water Purification / Q000379:methods; D014881:Water Supply
D001426:Bacterial Proteins; D009930:Organic Chemicals; C103389:SYTO 9; D005947:Glucose