K. Killham

Nitrification in coniferous forest soils.

Net nitrification rates tend to be low or negligible in the forest floor of many coniferous forests of North-East Scotland. The most likely process controls are substrate availability, pH, allelopathy, water potential, nutrient status and temperature. These are discussed in relation to field and laboratory studies of net and potential rates of nitrification.Fungi make up by far the largest part of the nitrifier community in the coniferous forest floor. Very little is known about the distribution and activity of autotrophs in these systems, although it is certain that in vitro evidence suggesting autotrophs cannot nitrify at pH levels characteristic of coniferous forest soils is unrealistic.Because of the metabolic diversity of nitrifying fungi, a variety of organic and inorganic nitrification pathways may exist in coniferous forests. The possible involvement of free radicles in fungal nitrification in coniferous forest soils is also suggested.A complete understanding of nitrification in coniferous forest soils can only result from field characterisation of N flux such as through the use of 15N. This must be combined with ecophysiological characterisation of the organisms involved in order that the complexity of nitrification in coniferous forest soils can be resolved.

A review of rhizosphere carbon flow modelling

Rhizosphere processes play a key role in nutrient cycling in terrestrial ecosystems. Plant rhizodeposits supply low-molecular weight carbon substrates to the soil microbial community, resulting in elevated levels of activity surrounding the root. Mechanistic compartmental models that aim to model carbon flux through the rhizosphere have been reviewed and areas of future research necessary to better calibrate model parameters have been identified. Incorporating the effect of variation in bacterial biomass physiology on carbon flux presents a considerable challenge to experimentalists and modellers alike due to the difficulties associated with differentiating dead from dormant cells. A number of molecular techniques that may help to distinguish between metabolic states of bacterial cells are presented. The calibration of growth, death and maintenance parameters in rhizosphere models is also discussed. A simple model of rhizosphere carbon flow has been constructed and a sensitivity analysis was carried out on the model to highlight which parameters were most influential when simulating carbon flux. It was observed that the parameters that most heavily influenced long-term carbon compartmentalisation in the rhizosphere were exudation rate and biomass yield. It was concluded that future efforts to simulate carbon flow in the rhizosphere should aim to increase ecological realism in model structure.

Loss of exudates from the roots of perennial ryegrass inoculated with a range of micro-organisms

To determine the effect of microbial metabolites on the release of root exudates from perennial ryegrass, seedlings were pulse labelled with [14C]-CO2 in the presence of a range of soil micro-organisms. Microbial inoculants were spatially separated from roots by Millipore membranes so that root infection did not occur. Using this technique, only microbial metabolites affected root exudation. The effect of microbial metabolites on carbon assimilation and distribution and root exudation was determined for 15 microbial species. Assimilation of a pulse label varied by over 3.5 fold, dependent on inoculant. Distribution of the label between roots and shoots also varied with inoculant, but the carbon pool that was most sensitive to inoculation was root exudation. In the absence of a microbial inoculant only 1% of assimilated label was exuded. Inoculation of the microcosms always caused an increase in exudation but the percentage exuded varied greatly, within the range of 3–34%.

Assessment of bioavailability of heavy metals using lux modified constructs of Pseudomonas fluorescens

The bioluminescence response of a genetically modified (lux‐marked) bacterium to potentially toxic elements (PTEs) was monitored using an in vitro assay. Washed cells of Pseudomonas fiuorescens were added to solutions containing various concentrations of metal salts. Bioluminescence, involving either plasmid or chromosomally encoded lux genes, declined as the metal concentration increased. The plasmid marked construct was significantly more sensitive to all metals except Cr. The order of metal sensitivity was found to be Cu = Zn > Cd > Cr > Ni for the chromosomally marked construct and Cu = Zn > Cd > Ni > Cr for the plasmid marked construct. The very sensitive response of lux‐marked terrestrial bacteria to PTEs identified the potential for a rapid and flexible ecotoxicity assay for assessing the pollution of soil or fresh water environments.

Effect of elevated atmospheric CO2 concentration on C-partitioning and rhizosphere C-flow for three plant species

Abstract The effects of elevated atmospheric CO 2 concentration on the partitioning of dry matter and recent assimilate was investigated for three plant species (rye grass, wheat and Bermuda grass). This was evaluated in plant-soil microcosm systems maintained at specific growth conditions, under two CO 2 regimes (450 and 720 μmol mol −1 ). The distribution of recent assimilate between plant, microbial and soil pools was determined by 14 CO 2 pulse chase, for each plant species at both CO 2 concentrations. Growth of rye grass and wheat (both C 3 ) was ca. doubled at the higher CO 2 concentration. Dry matter partitioning was also significantly affected, with an increased root-to-shoot ratio for wheat (0.72–1.03), and a decreased root-to-shoot ratio for rye grass (0.68-0.47) at elevated CO 2 . For Bermuda grass (C 4 ), growth and partitioning of dry matter and 14 C were not affected by CO 2 concentration. 14 C-allocation to the rhizospheres of rye-grass and wheat was found to be increased by 62 and 19%, respectively, at the higher CO 2 concentration. The partitioning of 14 C within the rhizospheres of the two C 3 species was also found to be affected by CO 2 concentration. At the higher CO 2 concentration, proportionately less 14 C was present in the microbial fraction, relative to that in the soil. This indicates altered microbial utilisation of root-released compounds at the higher CO 2 concentration, which may be a consequence of altered quantity or quality of rhizodeposits derived from recent assimilate.

Plasmid and chromosomally encoded luminescence marker systems for detection of Pseudomonas fluorescens in soil

Luminescent strains of Pseudomonas fluorescens 10586 were constructed in which luciferase production was constitutive by introduction of Vibrio fischeri luxABE genes on the chromosome and on a multicopy plasmid. Light production in liquid batch culture was directly proportional to biomass concentration during exponential growth and enabled detection by luminometry of 1.7 × 103 and 8.9 × 104 cells/ml for the plasmid and chromosomally marked strains, respectively. Luminescent colonies of both strains were detectable by eye, enabling viable cell enumeration on solid media against a background of non‐luminescent strains. Following inoculation into sterile and non‐sterile soil lower levels of detection were increased but detection of 8.1–59 × 103and 2.2–30 × 103 cells per g of soil was possible for plasmid and chromosomally marked strains. Maximum specific growth rate in liquid culture was unaffected by introduction of lux marker genes on the chromosome, but was reduced in the plasmid marked strain. The chromosomally encoded marker was stable in both liquid culture and in soil, but the plasmid was unstable during continuous subculturing in liquid medium and during growth in soil. The chromosomally encoded luminescence‐marker system therefore provides a convenient, non‐extractive technique for quantification of genetically modified soil microbial inocula.

Survival of E. coli O157:H7 in organic wastes destined for land application

Aim:  To determine the persistence of Escherichia coli O157 in contrasting organic wastes spread to land and to assess the potential environmental risk associated with the disposal of these wastes to land.

Characterisation of the dynamics of C-partitioning within Lolium perenne and to the rhizosphere microbial biomass using 14C pulse chase

The dynamics of C partitioning with Lolium perenne and its associated rhizosphere was investigated in plant-soil microcosms using 14C pulse-chase labelling. The 14CO2 pulse was introduced into the shoot chamber and the plants allowed to assimilate the label for a fixed period. The microcosm design facilitated independent monitoring of shoot and root/soil respiration during the chase period. Partitioning between above- and below-ground pools was determined between 30 min and 168 h after the pulse, and the distribution was found to vary with the length of the chase period. Initially (30 min after the pulse), the 14C was predominantly (99%) in the shoot biomass and declined thereafter. The results indicate that translocation of recent photoassimilate is rapid, with 14C detected below ground within 30 min of pulse application. The translocation rate of 14C below ground was maximal (6.2% h-1) between 30 min and 3 h after the pulse, with greatest incorporation into the microbial biomass detected at 3 h. After 3 h, the microbial biomass 14C pool accounted for 74% of the total 14C rhizosphere pool. By 24 h, approximately 30% of 14C assimilate had been translocated below ground; thereafter 14C translocation was greatly reduced. Partitioning of recent assimilate changed with increasing CO2 concentration. The proportion of 14C translocated below ground almost doubled from 17.76% at the ambient atmospheric CO2 concentration (450 ppm) to 33.73% at 750 ppm CO2 concentration. More specifically, these changes occurred in the root biomass and the total rhizosphere pools, with two- and threefold 14C increases at an elevated CO2 concentration compared to ambient, respectively. The pulselabelling strategy developed in this study provided sufficient sensitivity to determine perturbations in C dynamics in L. perenne, in particular rhizosphere C pools, in response to an elevated atmospheric CO2 concentration.

Predicting bioremediation of hydrocarbons: Laboratory to field scale

There are strong drivers to increasingly adopt bioremediation as an effective technique for risk reduction of hydrocarbon impacted soils. Researchers often rely solely on chemical data to assess bioremediation efficiently, without making use of the numerous biological techniques for assessing microbial performance. Where used, laboratory experiments must be effectively extrapolated to the field scale. The aim of this research was to test laboratory derived data and move to the field scale. In this research, the remediation of over thirty hydrocarbon sites was studied in the laboratory using a range of analytical techniques. At elevated concentrations, the rate of degradation was best described by respiration and the total hydrocarbon concentration in soil. The number of bacterial degraders and heterotrophs as well as quantification of the bioavailable fraction allowed an estimation of how bioremediation would progress. The response of microbial biosensors proved a useful predictor of bioremediation in the absence of other microbial data. Field-scale trials on average took three times as long to reach the same endpoint as the laboratory trial. It is essential that practitioners justify the nature and frequency of sampling when managing remediation projects and estimations can be made using laboratory derived data. The value of bioremediation will be realised when those that practice the technology can offer transparent lines of evidence to explain their decisions.

A novel method of quantifying root exudation in the presence of soil microflora

A microcosm is described in which root exudation may be estimated in the presence of microorganisms. Ryegrass seedlings are grown in microcosms in which roots were spatially separated from a microbial inoculant by a Millipore membrane. Seedlings grown in the microcosms were labelled with [14C]-CO2, and the fate of the label within the plant and rhizosphere was determined. Inoculation of the microcosms with Cladosporium resinae increased net fixation of the [14C] label compared to plants grown under sterile conditions. Inoculation also increased root exudation. The use of the microcosm was illustrated and its applications discussed.