A fluorescence-based assay for measuring the viable cell concentration of mixed microbial communities in soil.

Microbial cell concentration is a particularly important bioindicator of soil health and a yardstick for determining biological quotients which are likely to gain in ecological significance if they are calculated in relation to the viable, rather than total, microbial density. A dual-staining technique with fluorescent dyes was used for the spectrofluorimetric quantitative determination of the concentration of viable microbial cells present in three different soil types. This is a novel and substantially modified application of the dual-staining procedure implemented in the LIVE/DEAD BacLight viability kit which has never been successfully applied to the quantification of naturally occurring soil microbial communities. Indigenous microbial cell concentrations were quantified using an internal standard, i.e. spiking environmental samples with suspensions containing different concentrations of live E. coli cells, and external calibration, by comparing fluorescence emission by indigenous bacteria and known concentrations of E. coli in nutrient saline. Two types of environmental samples were tested: bacterial preparations obtained by density gradient centrifugation and soil suspensions. In both cases, prior dilution of the sample was necessary to minimise fluorescence quenching by soil particulate matter. Spectrofluorimetric measurements of indigenous cell concentration in bacterial preparations were in close agreement with those found using epifluorescence microscopy. Limits of detection of 5x10(6) for the soil bacterial preparations and 8x10(7) for the soil suspensions were estimated. Deviations observed when soil suspensions are dealt with are likely due to the selection of a unique bacterial strain for standardisation and calibration. Thorough testing of a variety of reference bacteria and fungi is suggested to determine a more accurate average fluorescence enhancement per microbial cell or mass unit.

2,4-Dichlorophenoxyacetic acid (2,4-D) sorption and degradation dynamics in three agricultural soils.

The fate and transport of 2,4-dichlorophenoxyacetic acid (2,4-D) in the subsurface is affected by a complex, time-dependent interplay between sorption and mineralization processes. 2,4-D is biodegradable in soils, while adsorption/desorption is influenced by both soil organic matter content and soil pH. In order to assess the dynamic interactions between sorption and mineralization, 2,4-D mineralization experiments were carried using three different soils (clay, loam and sand) assuming different contact times. Mineralization appeared to be the main process limiting 2,4-D availability, with each soil containing its own 2,4-D decomposers. For the clay and the loamy soils, 45 and 48% of the applied dose were mineralized after 10 days. By comparison, mineralization in the sandy soil proceeded initially much slower because of longer lag times. While 2,4-D residues immediately after application were readily available (>93% was extractable), the herbicide was present in a mostly unavailable state (<2% extractable) in all three soils after incubation for 60 days. We found that the total amount of bound residue decreased between 30 and 60 incubation days. Bioaccumulation may have led to reversible immobilization, with some residues later becoming more readily available again to extraction and/or mineralization.

Effect of metals on the adsorption and extractability of 14C-phenanthrene in soils.

The fate of polycyclic aromatic hydrocarbons (PAHs) in contaminated soils may be affected by several environmental factors including the presence of co-contaminants. This study was conducted in order to assess the effect of metals on (i) the adsorption of 14C-phenanthrene in soils and (ii) its extractability and ability to form non-extractable residues. The first objective was accomplished using batch adsorption experiments with an uncontaminated agricultural soil spiked with the metals Cd, Cu, Pb, and Zn. Adsorption of phenanthrene was significantly higher after the addition of the metals (Kf = 21.48 vs. 8.55) and the desorption less readily reversible when compared to the unspiked soil. The extractability of phenanthrene was assessed with incubation (4 months, laboratory conditions) and microlysimeter experiments (6 months, natural climatic conditions) on three soils spiked with metals. All the soils were labelled with 14C-phenanthrene. The amount of extractable phenanthrene residues was significantly higher when the metals had been added to the soils. Nevertheless, the quantity of non-extractable residues was non-significantly different between the spiked and unspiked soils. The mechanism leading to increased adsorption and extractability of phenanthrene in the presence of metals is still unknown. In perspective, it would be interesting to assess the bioavailability of PAHs in the presence of metals in further experiments.