Matthias Kaestner

Identification of bacterial micropredators distinctively active in a soil microbial food web

ABSTRACT The understanding of microbial interactions and trophic networks is a prerequisite for the elucidation of the turnover and transformation of organic materials in soils. To elucidate the incorporation of biomass carbon into a soil microbial food web, we added 13C-labeled Escherichia coli biomass to an agricultural soil and identified those indigenous microbes that were specifically active in its mineralization and carbon sequestration. rRNA stable isotope probing (SIP) revealed that uncultivated relatives of distinct groups of gliding bacterial micropredators (Lysobacter spp., Myxococcales, and the Bacteroidetes) lead carbon sequestration and mineralization from the added biomass. In addition, fungal populations within the Microascaceae were shown to respond to the added biomass after only 1 h of incubation and were thus surprisingly reactive to degradable labile carbon. This RNA-SIP study identifies indigenous microbes specifically active in the transformation of a nondefined complex carbon source, bacterial biomass, directly in a soil ecosystem.

Microbial cell-envelope fragments and the formation of soil organic matter: a case study from a glacier forefield

Genesis of soil organic matter (SOM) during pedogenesis is still a matter of controversy in soil science. Recently, it was hypothesized that microbial cell-envelope fragments contribute significantly to SOM formation. We tested the relevance of this process during pedogenesis by evaluating the development of SOM along a chronosequence of a glacier forefield (Damma glacier). Samples of increasing soil age collected along the forefield were analyzed for C and N contents, phospholipid and total fatty acids (PLFA and tFA), water contact angle, micro-hydrophobicity and surface coverage by microbial cell-envelope residues. The surface coverage was visualized and quantified by analysis of representative, equally-scaled scanning electron micrographs (SEM). Increasing SOM contents were accompanied by increasing coverage and overall abundance of microbial cell-envelope fragments as evaluated on the basis of scanning electron microscopy; this is also reflected in the amounts of tFA and PLFA, the trend of C/N ratios, and the increasing hydrophobicity and water contact angles of the soil samples. Using SEM and the image analysis approach, we can provide a process-based description of the development of SOM in the newly developing ecosystem of the glacier forefield. The majority of small-sized SOM visible with scanning electron microscopy appears to consist of bacterial cell envelope fragments that remain stable after cell death, such that their shape does not change with soil age. Our results show the importance of microbial processing of SOM, and highlight the existence of microbial necromass as a significant part of the fine-particulate SOM even in later stages of soil development.

Bioremediation of benzene-, MTBE- and ammonia-contaminated groundwater with pilot-scale constructed wetlands.

In this pilot-scale constructed wetland (CW) study for treating groundwater contaminated with benzene, MTBE, and ammonia-N, the performance of two types of CWs (a wetland with gravel matrix and a plant root mat) was investigated. Hypothesized stimulative effects of filter material additives (charcoal, iron(III)) on pollutant removal were also tested. Increased contaminant loss was found during summer; the best treatment performance was achieved by the plant root mat. Concentration decrease in the planted gravel filter/plant root mat, respectively, amounted to 81/99% for benzene, 17/82% for MTBE, and 54/41% for ammonia-N at calculated inflow loads of 525/603 mg/m(2)/d, 97/112 mg/m(2)/d, and 1167/1342 mg/m(2)/d for benzene, MTBE, and ammonia-N. Filter additives did not improve contaminant depletion, although sorption processes were observed and elevated iron(II) formation indicated iron reduction. Bacterial and stable isotope analysis provided evidence for microbial benzene degradation in the CW, emphasizing the promising potential of this treatment technique.

Performance evaluation of different horizontal subsurface flow wetland types by characterization of flow behavior, mass removal and depth-dependent contaminant load.

For several pilot-scale constructed wetlands (CWs: a planted and unplanted gravel filter) and a hydroponic plant root mat (operating at two water levels), used for treating groundwater contaminated with BTEX, the fuel additive MTBE and ammonium, the hydrodynamic behavior was evaluated by means of temporal moment analysis of outlet tracer breakthrough curves (BTCs): hydraulic indices were related to contaminant mass removal. Detailed investigation of flow within the model gravel CWs allowed estimation of local flow rates and contaminant loads within the CWs. Best hydraulics were observed for the planted gravel filter (number of continuously stirred tank reactors N = 11.3, dispersion number = 0.04, Péclet number = 23). The hydroponic plant root mat revealed lower N and pronounced dispersion tendencies, whereby an elevated water table considerably impaired flow characteristics and treatment efficiencies. Highest mass removals were achieved by the plant root mat at low level: 98% (544 mg m⁻² d⁻¹), 78% (54 mg m⁻² d⁻¹) and 74% (893 mg m⁻² d⁻¹) for benzene, MTBE and ammonium-nitrogen, respectively. Within the CWs the flow behavior was depth-dependent, with the planting and the position of the outlet tube being key factors resulting in elevated flow rate and contaminant flux immediately below the densely rooted porous media zone in the planted CW, and fast bottom flow in the unplanted reference.

Performance Evaluation Using a Three Compartment Mass Balance for the Removal of Volatile Organic Compounds in Pilot Scale Constructed Wetlands

To perform a general assessment of treatment efficiency, a mass balance study was undertaken for two types of constructed wetlands (CWs), planted gravel filters and plant root mat systems, for treating VOC (benzene; MTBE) polluted groundwater under field conditions. Contaminant fate was investigated in the respective water, plant, and atmosphere compartments by determining water and atmospheric contaminant loads and calculating contaminant plant uptake, thereby allowing for an extended efficiency assessment of CWs. Highest total VOC removal was achieved during summer, being pronounced for benzene compared to MTBE. According to the experimental results and the calculations generated by the balancing model, degradation in the rhizosphere and plant uptake accounted for the main benzene removal processes, of 76% and 13% for the gravel bed CW and 83% and 11% for the root mat system. Volatilization flux of benzene and MTBE was low (<5%) for the gravel bed CW, while in the root mat system direct contact of aqueous and gaseous phases favored total MTBE volatilization (24%). With this applied approach, we present detailed contaminant mass balances that allow for conclusive quantitative estimation of contaminant elimination and distribution processes (e.g., total, surface, and phytovolatilization, plant uptake, rhizodegradation) in CWs under field conditions.

Response of ammonium removal to growth and transpiration of Juncus effusus during the treatment of artificial sewage in laboratory-scale wetlands.

The correlation between nitrogen removal and the role of the plants in the rhizosphere of constructed wetlands are the subject of continuous discussion, but knowledge is still insufficient. Since the influence of plant growth and physiological activity on ammonium removal has not been well characterized in constructed wetlands so far, this aspect is investigated in more detail in model wetlands under defined laboratory conditions using Juncus effusus for treating an artificial sewage. Growth and physiological activity, such as plant transpiration, have been found to correlate with both the efficiency of ammonium removal within the rhizosphere of J. effusus and the methane formation. The uptake of ammonium by growing plant stocks is within in a range of 45.5%, but under conditions of plant growth stagnation, a further nearly complete removal of the ammonium load points to the likely existence of additional nitrogen removal processes. In this way, a linear correlation between the ammonium concentration inside the rhizosphere and the transpiration of the plant stocks implies that an influence of plant physiological activity on the efficiency of N-removal exists. Furthermore, a linear correlation between methane concentration and plant transpiration has been estimated. The findings indicate a fast response of redox processes to plant activities. Accordingly, not only the influence of plant transpiration activity on the plant-internal convective gas transport, the radial oxygen loss by the plant roots and the efficiency of nitrification within the rhizosphere, but also the nitrogen gas released by phytovolatilization are discussed. The results achieved by using an unplanted control system are different in principle and characterized by a low efficiency of ammonium removal and a high methane enrichment of up to a maximum of 72.7% saturation.

Physiological responses of Juncus effusus (Rush) to chromium and relevance for wastewater treatment in constructed wetlands.

Constructed wetlands are increasingly applied for industrial wastewater treatment. However, current knowledge of the stress responses of helophytes to selected toxicants such as dichromate is limited. The goal of the experiments presented here was to characterize the physiological response of Juncus effusus to different concentrations of dichromate dependent upon the growth and constitution of the plants. The growth parameters, shoot length, and dry weight already were strongly affected at low dichromate concentrations of approximately 34 μM. Concentrations of 340 μM impaired chlorophyll fluorescence and a decrease in chlorophyll a started at concentrations higher than 170 μM dichromate. The concentrations of chlorophyll b and carotenoids also were influenced negatively. Thus, the reduction of the pigment content started before any obvious influence on the chlorophyll fluorescence. The highest concentration of dichromate, which caused no permanent inhibition of growth and photosynthesis, was found to be 17 μM K2Cr2O7. In principle, J. effusus is suitable for constructed wetlands to treat chromium-containing wastewater. Because the stress resistance of J. effusus is limited, the maximum concentration of dichromate in the treated wastewater should not exceed 34 μM. The growth parameters, shoot length, and dry weight were sensitive to much lower dichromate concentrations and did react more quickly than the biochemistry-related parameters chlorophyll fluorescence and pigment concentration. Therefore, the example of Juncus effusus shows that the use of only biochemical parameters to define concentration limits for the treatment of dichromate-containing wastewater can lead to incorrect conclusions and result in disturbed long-term operation of the system.

The impact of chemical pollution on the resilience of soils under multiple stresses: A conceptual framework for future research.

Soils are faced with man-made chemical stress factors, such as the input of organic or metal-containing pesticides, in combination with non-chemical stressors like soil compaction and natural disturbance like drought. Although multiple stress factors are typically co-occurring in soil ecosystems, research in soil sciences on this aspect is limited and focuses mostly on single structural or functional endpoints. A mechanistic understanding of the reaction of soils to multiple stressors is currently lacking. Based on a review of resilience theory, we introduce a new concept for research on the ability of polluted soil (xenobiotics or other chemical pollutants as one stressor) to resist further natural or anthropogenic stress and to retain its functions and structure. There is strong indication that pollution as a primary stressor will change the system reaction of soil, i.e., its resilience, stability and resistance. It can be expected that pollution affects the physiological adaption of organisms and the functional redundancy of the soil to further stress. We hypothesize that the recovery of organisms and chemical-physical properties after impact of a follow-up stressor is faster in polluted soil than in non-polluted soil, i.e., polluted soil has a higher dynamical stability (dynamical stability=1/recovery time), whereas resilience of the contaminated soil is lower compared to that of not or less contaminated soil. Thus, a polluted soil might be more prone to change into another system regime after occurrence of further stress. We highlight this issue by compiling the literature exemplarily for the effects of Cu contamination and compaction on soil functions and structure. We propose to intensify research on effects of combined stresses involving a multidisciplinary team of experts and provide suggestions for corresponding experiments. Our concept offers thus a framework for system level analysis of soils paving the way to enhance ecological theory.

Characterisation of microbial activity in the framework of natural attenuation without groundwater monitoring wells?: a new Direct-Push probe

At many contaminated field sites in Europe, monitored natural attenuation is a feasible site remediation option. Natural attenuation includes several processes but only the microbial degradation leads to real contaminant removal and very few methods are accepted by the authorities providing real evidence of microbial contaminant degradation activity. One of those methods is the recently developed in situ microcosm approach (BACTRAP®). These in situ microcosms consist of perforated stainless steel cages or PTFE tubes filled with an activated carbon matrix that is amended with 13C-labelled contaminants; the microcosms are then exposed within groundwater monitoring wells. Based on this approach, natural attenuation was accepted by authorities as a site remediation option for the BTEX-polluted site Zeitz in Germany. Currently, the in situ microcosms are restricted to the use inside groundwater monitoring wells at the level of the aquifer. The (classical) system therefore is only applicable on field sites with a network of monitoring wells, and only microbial activity inside the monitoring wells at the level of the aquifer can be assessed. In order to overcome these limitations, a new Direct-Push BACTRAP probe was developed on the basis of the Geoprobe® equipment. With respect to the mechanical boundary conditions of the DP technique, these new probes were constructed in a rugged and segmented manner and are adaptable to various sampling concepts. With this new probe, the approach can be extended to field sites without existing monitoring wells, and microbial activity was demonstrated to be measureable even under very dry conditions inside the vadose zone above the aquifer. In a field test, classical and Direct-Push BACTRAPs were applied in the BTEX-contaminated aquifer at the ModelPROBE reference site Zeitz (Germany). Both types of BACTRAPs were incubated in the centre and at the fringe of the BTEX plume. Analysis of phospholipid fatty acid (PLFA) patterns showed that the bacterial communities on DP-BACTRAPs were more similar to the soil than those found on classical BACTRAPs. During microbial degradation of the 13C-labelled substrate on the carrier material of the microcosms, the label was only slightly incorporated into bacterial biomass, as determined by PLFA analysis. This provides clear indication for decreased in situ natural attenuation potential in comparison to earlier sampling campaigns, which is presumably caused by a large-scale source remediation measure in the meantime. In conclusion, Direct-Push-based BACTRAPs offer a promising way to monitor natural attenuation or remediation success at field sites which are currently inaccessible by the technique due to the lack of monitoring wells or due to a main contamination present within the vadose zone.

Adaptation of a constructed wetland to simultaneous treatment of monochlorobenzene and perchloroethene.

Mixed groundwater contaminations by chlorinated volatile organic compounds (VOC) cause environmental hazards if contaminated groundwater discharges into surface waters and river floodplains. Constructed wetlands (CW) or engineered natural wetlands provide a promising technology for the protection of sensitive water bodies. We adapted a constructed wetland able to treat monochlorobenzene (MCB) contaminated groundwater to a mixture of MCB and tetrachloroethene (PCE), representing low and high chlorinated model VOC. Simultaneous treatment of both compounds was efficient after an adaptation time of 2½ years. Removal of MCB was temporarily impaired by PCE addition, but after adaptation a MCB concentration decrease of up to 64% (55.3 μmol L−1) was observed. Oxygen availability in the rhizosphere was relatively low, leading to sub-optimal MCB elimination but providing also appropriate conditions for PCE dechlorination. PCE and metabolites concentration patterns indicated a very slow system adaptation. However, under steady state conditions complete removal of PCE inflow concentrations of 10–15 μmol L−1 was achieved with negligible concentrations of chlorinated metabolites in the outflow. Recovery of total dechlorination metabolite loads corresponding to 100%, and ethene loads corresponding to 30% of the PCE inflow load provided evidence for complete reductive dechlorination, corroborated by the detection of Dehalococcoides sp.