Anja Miltner

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.

Biodegradation of ciprofloxacin in water and soil and its effects on the microbial communities

While antibiotics are frequently found in the environment, their biodegradability and ecotoxicological effects are not well understood. Ciprofloxacin inhibits active and growing microorganisms and therefore can represent an important risk for the environment, especially for soil microbial ecology and microbial ecosystem services. We investigated the biodegradation of (14)C-ciprofloxacin in water and soil following OECD tests (301B, 307) to compare its fate in both systems. Ciprofloxacin is recalcitrant to biodegradation and transformation in the aqueous system. However, some mineralisation was observed in soil. The lower bioavailability of ciprofloxacin seems to reduce the compounds toxicity against microorganisms and allows its biodegradation. Moreover, ciprofloxacin strongly inhibits the microbial activities in both systems. Higher inhibition was observed in water than in soil and although its antimicrobial potency is reduced by sorption and aging in soil, ciprofloxacin remains biologically active over time. Therefore sorption does not completely eliminate the effects of this compound.

Classification and Modelling of Nonextractable Residue (NER) Formation of Xenobiotics in Soil – A Synthesis

This review provides a comprehensive overview about nonextractable residue (NER) formation and attempts to classify the various types. Xenobiotic NER derived from parent pesticides (or other environmental contaminants) and primary metabolites sorbed or entrapped within the soil organic matter (Type I) or covalently bound (Type II) pose a considerably higher risk than those derived from productive biodegradation. However, biogenic nonextractable residues (bioNER) (Type III) resulting from conversion of carbon (or nitrogen) from the compounds into microbial biomass molecules do not pose any risk. Experimental approaches to clearly distinguish between the types are provided, and a model to prospectively estimate bioNER formation in soil is proposed.

Formation and Fate of Bound Residues from Microbial Biomass during 2,4-D Degradation in Soil

During organic contaminant degradation in soil, bound or nonextractable residues (NER) are formed. Part of these residues may be biogenic, because degrading microorganisms assimilate carbon derived from the pollutant and mineralized CO(2) to form cellular components for example, [fatty acids (FA) and amino acids (AA)], which are subsequently stabilized within soil organic matter (SOM). We investigated the formation and fate of FA and AA from biodegradation of (13)C(6)-2,4-D in soil and the incorporation of the (13)C-label into living biomass via (13)CO(2) fixation. After 64 days of incubation, (13)C-AA in SOM indicated that 44% of the initially applied (13)C(6)-2,4-D equivalents had been converted to microbial biomass and finally to biogenic residues. The intermediate maximum of (13)C-FA in SOM indicated a 20% conversion of (13)C(6)-2,4-D to biomass, but (13)C-FA decreased to 50% of that value whereas (13)C-AA in the SOM remained stable. We provide the first evidence that nearly all bound residues from 2,4-D are biogenic, containing natural microbial residues stabilized in SOM. Because of biogenic residue formation, the potential risk of bound residues from readily metabolized xenobiotics in soils is highly overestimated. Hence, the formation of biogenic residues must be considered in general when performing mass balances of pollutant biodegradation in soils.

FATE OF MICROBIAL RESIDUES DURING LITTER DECOMPOSITION AS AFFECTED BY MINERALS

Minerals may protect organic matter against microbial decay, either by direct chemical and physical interactions or by inhibitory effects on the soil microbial community. To clarify the effects of minerals on organic matter cycling by microorganisms, we used amino sugars as tracers for C and N in dead microbial cells after 0, 15, 29, 90, 239, and 498 days of incubation of beech leaf litter mixed with quartz sand, Fe oxide, Al hydroxide, or Mn oxide. Beech leaf litter without addition of any mineral phases was used as the control. The results show that amino sugar concentrations increased as litter decomposition proceeded. Decreasing ratios of glucosamine to muramic acid and of glucosamine to galactosamine indicated that bacterial products accumulated increasingly relative to fungal cells with increasing incubation time. As the presence of Mn oxide promoted losses of plant-derived C, there was a more pronounced selective accumulation of the microbial-derived amino sugar C than in the other treatments. Aluminium hydroxide and Fe oxide inhibited synthesis of bacterial amino sugars by a factor of two. This resulted in lower amino sugar C proportions compared with the other treatments. Consequently, the amino sugar C proportions were sensitive to both increasing amino sugar synthesis and C mineralization rates. In contrast, the amino sugar N proportions were not affected by any mineral additions. Thus, the mere presence of minerals did not affect the cycling of N through the amino sugar pool, but minerals altered the relative proportions of N sequestered within residues of bacteria and fungi.

Prediction of the Adaptability of Pseudomonas putida DOT-T1E to a Second Phase of a Solvent for Economically Sound Two-Phase Biotransformations

ABSTRACT The strain Pseudomonas putida DOT-T1E was tested for its ability to tolerate second phases of different alkanols for their use as solvents in two-liquid-phase biotransformations. Although 1-decanol showed an about 10-fold higher toxicity to the cells than 1-octanol, the cells were able to adapt completely to 1-decanol only and could not be adapted in order to grow stably in the presence of a second phase of 1-octanol. The main explanation for this observation can be seen in the higher water and membrane solubility of 1-octanol. The hydrophobicity (log P) of a substance correlates with a certain partitioning of that compound into the membrane. Combining the log P value with the water solubility, the maximum membrane concentration of a compound can be calculated. With this simple calculation, it is possible to predict the property of an organic chemical for its potential applicability as a solvent for two-liquid-phase biotransformations with solvent-tolerant P. putida strains. Only compounds that show a maximum membrane concentration of less than 400 mM, such as 1-decanol, seem to be tolerated by these bacterial strains when applied in supersaturating concentrations to the medium. Taking into consideration that a solvent for a two-liquid-phase system should possess partitioning properties for potential substrates and products of a fine chemical synthesis, it can be seen that 1-decanol is a suitable solvent for such biotransformation processes. This was also demonstrated in shake cultures, where increasing amounts of a second phase of 1-decanol led to bacteria tolerating higher concentrations of the model substrate 3-nitrotoluene. Transferring this example to a 5-liter-scale bioreactor with 10% (vol/vol) 1-decanol, the amount of 3-nitrotoluene tolerated by the cells is up to 200-fold higher than in pure aqueous medium. The system demonstrates the usefulness of two-phase biotransformations utilizing solvent-tolerant bacteria.

Non-phototrophic CO2 fixation by soil microorganisms

Although soils are generally known to be a net source of CO2 due to microbial respiration, CO2 fixation may also be an important process. The non-phototrophic fixation of CO2 was investigated in a tracer experiment with 14CO2 in order to obtain information about the extent and the mechanisms of this process. Soils were incubated for up to 91 days in the dark. In three independent incubation experiments, a significant transfer of radioactivity from 14CO2 to soil organic matter was observed. The process was related to microbial activity and could be enhanced by the addition of readily available substrates such as acetate. CO2 fixation exhibited biphasic kinetics and was linearly related to respiration during the first phase of incubation (about 20–40 days). The fixation amounted to 3–5% of the net respiration. After this phase, the CO2 fixation decreased to 1–2% of the respiration. The amount of carbon fixed by an agricultural soil corresponded to 0.05% of the organic carbon present in the soil at the beginning of the experiment, and virtually all of the fixed CO2 was converted to organic compounds. Many autotrophic and heterotrophic biochemical processes result in the fixation of CO2. However, the enhancement of the fixation by addition of readily available substrates and the linear correlation with respiration suggested that the process is mainly driven by aerobic heterotrophic microorganisms. We conclude that heterotrophic CO2 fixation represents a significant factor of microbial activity in soils.

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.

Terrestrial organic matter in surface sediments of the Baltic Sea, Northwest Europe, as determined by CuO oxidation

Abstract We studied the distribution and composition of terrestrial organic matter in sediments of the Baltic Sea (Northwest Europe). To this end, surface sediments from all basins of the Baltic Sea were analyzed for their lignin oxidation product yields and compositions after CuO oxidation. Lignin oxidation product yields depend on the concentration of organic carbon and range from 0.4 to 10.2 mg g−1 total organic carbon (TOC). On the basis of an average of 13 mg g−1 TOC in two river sediments, we estimate that the upper limit of terrestrial organic matter in Baltic Sea sediments is 30% of TOC. The contribution of terrestrial organic matter differed between the individual basins, depending on the distance from runoff discharge areas and on the area occupied by each submarine catchment. Lignin composition showed a relative decrease of angiosperm tissue from the Southwest to the Northeast, reflecting the shift from temperate to boreal vegetation type. The Gotland and the Bornholm Seas, which have no significant river input, were characterized by high relative contributions of nonwoody, strongly altered material. The source may either be a mixture of pollen and peat being eroded from geologically older strata at the seafloor or laterally advected material from the other basins. However, the pronounced compositional differences between the basins indicated that interbasin transport of terrestrial organic matter is less important than direct river input, although river signals can only be traced at a few places in the Baltic Sea.

Microbial degradation of the pharmaceutical ibuprofen and the herbicide 2,4-D in water and soil - use and limits of data obtained from aqueous systems for predicting their fate in soil.

The persistence of chemicals is a key parameter for their environmental risk assessment. Extrapolating their biodegradability potential in aqueous systems to soil systems would improve the environmental impact assessment. This study compares the fate of (14/13)C-labelled 2,4-D (2,4-dichlorophenoxyacetic acid) and ibuprofen in OECD tests 301 (ready biodegradability in aqueous systems) and 307 (soil). 85% of 2,4-D and 68% of ibuprofen were mineralised in aqueous systems, indicating ready biodegradability, but only 57% and 45% in soil. Parent compounds and metabolites decreased to <2% of the spiked amounts in both systems. In soil, 36% of 2,4-D and 30% of ibuprofen were bound in non-extractable residues (NER). NER formation in the abiotic controls was half as high as in the biotic treatments. However, mineralisation, biodegradation and abiotic residue formation are competing processes. Assuming the same extent of abiotic NER formation in abiotic and biotic systems may therefore overestimate the abiotic contribution in the biotic systems. Mineralisation was described by a logistic model for the aquatic systems and by a two-pool first order degradation model for the soil systems. This agrees with the different abundance of microorganisms in the two systems, but precludes direct comparison of the fitted parameters. Nevertheless, the maximum mineralisable amounts determined by the models were similar in both systems, although the maximum mineralisation rate was about 3.5 times higher in the aqueous systems than in the soil system for both compounds; these parameters may thus be extrapolated from aqueous to soil systems. However, the maximum mineralisable amount is calculated by extrapolation to infinite times and includes intermediately formed biomass derived from the labelled carbon. The amount of labelled carbon within microbial biomass residues is higher in the soil system, resulting in lower degradation rates. Further evaluation of these relationships requires comparison data on more chemicals and from different soils.