Multiple responses of gram-negative bacteria to organic solvents.

Abstract

Since the Industrial Revolution, the production and use of chemicals has increased immensely. As a consequence, all kinds of wastes are produced, which are released into the environment. Many products such as herbicides or insecticides will be released into the environment to control weeds and pests; other products such as organic solvents or fuels will reach the biosphere as a result of production or storage losses, accidents and solvent evaporation. Nowadays, there is a growing awareness concerning the possible toxic or even carcinogenic effects of chemicals. Although the release of many of them is restricted by legislation, a number of pollutants have already reached the biosphere and need to be eliminated. The use of biological treatments for the removal of toxic chemicals seems to be promising (Ramos et al., 1994). However, chemical toxicity can hamper the application of microorganisms in the removal of pollutants from waste streams and dump sites. This is a serious problem when dealing with microbial bioremediation in reactors, biofilters and soils. The main function of the cell membrane of microorganisms is to form a permeability barrier, regulating the passage of solutes between the cell and the external environment (Nikaido, 1999). The barrier properties of the cytoplasmic membrane are of special importance for the energy transduction of the cell (Sikkema et al., 1995). The major damage caused by organic solvents on the cell membrane is the impairment of vital functions, e.g. loss of ions, metabolites, lipids and proteins, the dissipation of the pH gradient and electrical potential or the inhibition of membrane protein functions. This is often followed, in turn, by cell lysis and death (de Smet et al., 1978; Sikkema et al., 1995). The logarithm of the partitioning coefficient of a solvent in a defined octanol–water mixture (log Pow) is commonly used as a measure of the lipophilicity of a solvent (Rekker and de Kort, 1979). Aromatic solvents with a log Pow below 4.0, e.g. benzene (log Pow 2.0), styrene (log Pow 3.6), xylene (log Pow 3.2) and toluene (log Pow 2.5), accumulate in the cytoplasmic membrane of bacteria, causing disorganization of the cell membrane structure and impairment of the above-cited vital membrane functions (Sikkema et al., 1992; 1994). Nevertheless, several Pseudomonas species have been isolated that are able to grow on rich and minimal medium in the presence of high concentrations of toxic organic solvents, such as toluene, styrene and p-xylene (Inoue and Horikoshi, 1989; Cruden et al., 1992; Weber et al., 1994; Ramos et al., 1995). Recently, it has been shown that P. putida Idaho and P. putida DOT-T1 are not only resistant to toluene in a two-phase system, but can even use toluene at these high concentrations as a carbon and energy source (Cruden et al., 1992; Ramos et al., 1995). Regarding tolerance to aromatic hydrocarbons, a number of elements have been suggested as being involved in the response to these toxic chemicals: (i) metabolism of the toxic hydrocarbons, which can contribute to their transformation into non-toxic compounds; (ii) rigidification of the cell membrane via alteration in the composition of phospholipids; and (iii) efflux of the toxic compound in an energy-dependent process. Although the metabolism of the toxic chemicals can help to reduce their toxicity, two lines of evidence suggest that it is of minor importance. (i) A number of microorganisms tolerant to different organic solvents cannot metabolize the toxic compound, e.g. Escherichia coli strains tolerant to 1% (v/v) hexane do not use (or biotransform) this compound at all (Aono et al., 1991). Furthermore, a number of Pseudomonas strains tolerant to supersaturating concentrations of toluene did not use this compound as a carbon source (Inoue and Horikoshi, 1989). (ii) Pseudomonas putida DOT-T1E is a toluene-tolerant strain that degrades and uses this chemical via the toluene–dioxygenase Environmental Microbiology (1999) 1(3), 191–198