Abstract-Using the pollution-induced community tolerance (PICT) concept, this article presents a rapid method for detecting toxic effects of contaminated soil on soil microbial communities and for elucidating the causal relationship between exposure history and toxic effects in exposed microbial communities. The method is based on the use of multiwell plates (Biolog) for producing concentration versus effect relationships for (maximally) 95 different microbial metabolic activities. For this, artificial exposure in the multiwell plate is established to the contaminant supposedly causing toxic effects in the field. The method was tested in a gradient of zinc-contaminated field plots by studying community tolerance to zinc. For most substrates the metabolic activities showed an increased community tolerance for zinc with increased zinc concentration in the field. Consequently, PICT has evolved after 2.5 years of zinc exposure under field conditions. The perspectives for future use of the PICT concept in combination with the multiwell technique for demonstrating ecotoxicological effects at contaminated sites is exemplified by the comparison of Dutch soil quality criteria for zinc with the collected data. This comparison showed that the evolution of microbial-PICT for zinc has occurred at environmental concentrations near the Dutch intervention value, a quality criterion linked to remediation urgency.
In earlier studies, the pathway of toluene and m-and p-xylene degradation (TOL pathway) in Pseudomonas putida(pWW0) was found to be subject to catabolite repression when the strain was grown at the maximal rate on glucose or succinate in the presence of an inducer. This report describes catabolite repression of the TOL pathway by succinate in chemostat cultures run at a low dilution rate (D ؍ 0.05 h ؊1 ) under different conditions of inorganic-nutrient limitation. The activity of benzylalcohol dehydrogenase (BADH) in cell extracts was used as a measure of the expression of the TOL upper pathway. When cells were grown in the presence of 10 to 15 mM succinate under conditions of phosphate or sulfate limitation, the BADH activity in response to the nonmetabolizable inducer o-xylene was less than 2% of that of cells grown under conditions of succinate limitation. Less repression was found under conditions of ammonium or oxygen limitation (2 to 10% and 20 to 35%, respectively, of the BADH levels under succinate limitation). The BADH expression levels determined under the different growth conditions appeared to correlate well with the mRNA transcript levels from the upper pathway promoter (Pu), which indicates that repression was due to a blockage at the transcriptional level. The meta-cleavage pathway was found to be less susceptible to catabolite repression. The results obtained suggest that the occurrence of catabolite repression is related to a high-energy status of the cells rather than to a high growth rate or directly to the presence of growth-saturating concentrations of a primary carbon and energy source.
Using the pollution-induced community tolerance (PICT) concept, this article presents a rapid method for detecting toxic effects of contaminated soil on soil microbial communities and for elucidating the causal relationship between exposure history and toxic effects in exposed microbial communities. The method is based on the use of multiwell plates (Biolog) for producing concentration versus effect relationships for (maximally) 95 different microbial metabolic activities. For this, artificial exposure in the multiwell plate is established to the contaminant supposedly causing toxic effects in the field. The method was tested in a gradient of zinc-contaminated field plots by studying community tolerance to zinc. For most substrates the metabolic activities showed an increased community tolerance for zinc with increased zinc concentration in the field. Consequently, PICT has evolved after 2.5 years of zinc exposure under field conditions. The perspectives for future use of the PICT concept in combination with the multiwell technique for demonstrating ecotoxicological effects at contaminated sites is exemplified by the comparison of Dutch soil quality criteria for zinc with the collected data. This comparison showed that the evolution of microbial-PICT for zinc has occurred at environmental concentrations near the Dutch intervention value, a quality criterion linked to remediation urgency.
Previous studies have shown that expression of the toluene and m-and p-xylene degradation pathway in Pseudomonas putids ( p W 0 ) is subject to catabolite repression by succinate. We report here that the expression level of the upper part of this so-called TOL pathway in cells grown in chemostat culture is strongly influenced by nutrient limitation when m-xylene is the sole carbon and energy source. The benzylalcohol dehydrogenase (BADH) levels in cells that are growth-limited by anabolic processes [sulphate (S)-, phosphate (P)-or nitrogen (N)-limiting conditions] were 3-12O/0 of those in cells growing under oxygen limitation (when catabolism limits growth). BADH levels under S-, P-and N-limitation were further decreased (three-to fivefold) when succinate was supplied in addition to m-xylene. Levels of the meta-cleavage pathway enzyme catechoI2,3=dioxygenase were less affected by the growth conditions but the general pattern was similar. Dilution rate also influenced the expression of the TOL pathway: BADH levels gradually decreased with increasing dilution rates, from 1250 mU (mg protein)'l at D = 095 h' l under m-xylene limitation to 290 mU (mg protein)" a t D = 058 h'l (non-limited growth). BADH levels were shown to be proportional to the specific affinity of whole cells for m-xylene. It may, therefore, be expected that natural degradation rates are adversely affected by anabolic nutrient limitations, especially at relatively low concentrations of the xenobiotic compound.
Styrene degradation in Pseudomonas putida CA-3 has previously been shown to be subject to catabolite repression in batch culture. We report here on the catabolite-repressing effects of succinate and glutamate and the effects of a limiting inorganic-nutrient concentration on the styrene degradation pathway of P. putida CA-3 in a chemostat culture at low growth rates (0.05 h ؊1). Oxidation of styrene and the presence of styrene oxide isomerase and phenylacetaldehyde dehydrogenase activities were used as a measure of the expression of the styrene degradation pathway. Both glutamate and succinate failed to repress the styrene degradation ability under growth conditions of carbon and energy limitation. Lower levels of enzyme activities of the styrene degradation pathway were seen in cells grown on styrene or phenylacetic acid (PAA) under conditions of both ammonia and sulfate limitation than were seen under carbon and energy limitation. Cells grown on PAA under continuous culture oxidize styrene and styrene oxide and possess styrene oxide isomerase and NAD ؉-dependent phenylacetaldehyde dehydrogenase activities. Catabolite repression of styrene metabolism was observed in cells grown on styrene or PAA in the presence of growth-saturating (nonlimiting) concentrations of succinate or glutamate under sulfate limitation.
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