A novel isolated Terrabacter-like bacterium can mineralize 2-aminopyrimidine, the principal metabolite of microbial sulfadiazine degradation.

Recently we showed that during the degradation of sulfadiazine (SDZ) by Microbacterium lacus strain SDZm4 the principal metabolite 2-aminopyrimidine (2-AP) accumulated to the same molar amount in the culture as SDZ disappeared (Tappe et al. Appl Environ Microbiol 79:2572-2577, 2013). Although 2-AP is considered a recalcitrant agent, long-term lysimeter experiments with (14)C-pyrimidine labeled SDZ ([(14)C]pyrSDZ) provided indications for substantial degradation of the pyrimidine moiety of the SDZ molecule. Therefore, we aimed to enrich 2-AP degrading bacteria and isolated a pure culture of a Terrabacter-like bacterium, denoted strain 2APm3. When provided with (14)C-labeled SDZ, M. lacus strain SDZm4 degraded [(14)C]pyrSDZ to [(14)C]2-AP. Resting cells of 2APm3 at a concentration of 5 × 10(6) cells ml(-1) degraded 62 µM [(14)C]2-AP to below the detection limit (0.6 µM) within 5 days. Disappearance of 2-AP resulted in the production of at least two transformation products (M1 and M2) with M2 being identified as 2-amino-4-hydroxypyrimidine. After 36 days, the transformation products disappeared and 83 % of the applied [(14)C]2-AP radioactivity was trapped as (14)CO2. From this we conclude that a consortium of two species should be able to almost completely degrade SDZ in soils.

Degradation of sulfadiazine by Microbacterium lacus strain SDZm4, isolated from lysimeters previously manured with slurry from sulfadiazine-medicated pigs.

Sulfadiazine (SDZ)-degrading bacterial cultures were enriched from the topsoil layer of lysimeters that were formerly treated with manure from pigs medicated with (14)C-labeled SDZ. The loss of about 35% of the applied radioactivity after an incubation period of 3 years was attributed to CO2 release due to mineralization processes in the lysimeters. Microcosm experiments with moist soil and soil slurries originating from these lysimeters confirmed the presumed mineralization potential, and an SDZ-degrading bacterium was isolated. It was identified as Microbacterium lacus, denoted strain SDZm4. During degradation studies with M. lacus strain SDZm4 using pyrimidine-ring labeled SDZ, SDZ disappeared completely but no (14)CO2 was released during 10 days of incubation. The entire applied radioactivity (AR) remained in solution and could be assigned to 2-aminopyrimidine. In contrast, for parallel incubations but with phenyl ring-labeled SDZ, 56% of the AR was released as (14)CO2, 16% was linked to biomass, and 21% remained as dissolved, not yet identified (14)C. Thus, it was shown that M. lacus extensively mineralized and partly assimilated the phenyl moiety of the SDZ molecule while forming equimolar amounts of 2-aminopyrimidine. This partial degradation might be an important step in the complete mineralization of SDZ by soil microorganisms.

Growth-inhibitory effects of sulfonamides at different pH: dissimilar susceptibility patterns of a soil bacterium and a test bacterium used for antibiotic assays.

The ionic speciation of sulfonamides is pH-driven and this may be crucial for their bioavailability and sorption to soil constituents, as well as for their uptake into bacterial cells. The inhibition behaviour of a bacterial test strain (Pseudomonas aeruginosa; DSM 1117), which was grown in the presence of different concentrations of 8 sulfonamides at pH values from 5 to 8, could be predicted by models that take the speciation of sulfonamides in- and outside of bacterial cells into account. Assuming a pH of 7.5 inside the cells (pH homeostasis), the strongest inhibition was predicted for the lowest external pH and for sulfonamides with the lowest pK(a) values. Growth experiments with Ps. aeruginosa basically reflected this predicted behaviour. However, Pantoea agglomerans -- a bacterial strain isolated from arable soil -- behaved surprisingly different regarding its pH dependency: all sulfonamides showed the strongest effects at pH 7 to 8 instead of being most effective at lowest pH, although the pK(a) dependencies followed the same pattern. Experimental and modeling results could be brought into good agreement for P. agglomerans if the cell-internal pH was admitted to approximate the external pH instead of implying pH homeostasis for modeling calculations. Thus, besides the actual concentration of sulfonamides, the pH dependent mode of reaction of different bacteria to sulfonamides may additionally govern the population dynamics in soils.

Field scale boscalid residues and dissipation half-life estimation in a sandy soil.

The aim of this study was to analyze the environmental fate of the fungicide boscalid in a sandy soil. Boscalid was applied in spring 2010/11 to a cropland site in western Germany. Three years after second application 65 undisturbed soil samples were taken. Boscalid was extracted using accelerated solvent extraction (ASE). Boscalid contents in the plough horizon ranged between 0.12 and 0.53 with a field mean of 0.20 ± 0.09 µg kg(-1). These contents were considerably lower compared to calculation using literature DT50 values, whereby a concentration of 16.89 µg kg(-1) was expected assuming a literature DT50 value of 345 days. Therefore, the measured field boscalid concentration only yields 1.2% of the expected value. To test whether the unknown extraction efficiency, losses from spray drift and interception can explain the mismatch between calculated and measured concentrations all these uncertainties were taken into account into calculations, but field concentrations and DT50 were still lower as expected. Leaching to deeper horizons was also studied but could not explain the discrepancy either. Moreover, a short-term incubation experiment using (14)C labelled boscalid revealed also shorter DT50 values of 297-337 compared to the 345 days taken from literature. However, this DT50 value is still considerably larger compared to the 104-224 days that were calculated based on the field experiment. Our results indicate that boscalid dissipation under field conditions is much faster at agricultural sites with sandy soil type as expected from laboratory incubation experiments.

Dissipation of bentazone, pyrimethanil and boscalid in biochar and digestate based soil mixtures for biopurification systems.

Biopurification systems, such as biofilters, are biotechnological tools to prevent point sources of pesticide pollution stemming from on-farm operations. For the purification processes pesticide sorption and mineralization and/or dissipation are essential and both largely depend on the type of filling materials and the pesticide in use. In this paper the mineralization and dissipation of three contrasting (14)C-labeled pesticides (bentazone, boscalid, and pyrimethanil) were investigated in laboratory incubation experiments using sandy soil, biochar produced from Pine woodchips, and/or digestate obtained from anaerobic digestion process using maize silage, chicken manure, beef and pig urine as feedstock. The results indicate that the addition of digestate increased pesticide mineralization, whereby the mineralization was not proportional to the digestate loads in the mixture, indicating a saturation effect in the turnover rate of pesticides. This effect was in correlation with the amount of water extractable DOC, obtained from the digestate based mixtures. Mixing biochar into the soil generally reduced total mineralization and led to larger sorption/sequestration of the pesticides, resulting in faster decrease of the extractable fraction. Also the addition of biochar to the soil/digestate mixtures reduced mineralization compared to the digestate alone mixture but mineralization rates were still higher as for the biochar/soil alone. In consequence, the addition of biochar to the soil generally decreased pesticide dissipation times and larger amounts of biochar led to high amounts of non-extractable residues of pesticide in the substrates. Among the mixtures tested, a mixture of digestate (5%) and biochar (5%) gave optimal results with respect to mineralization and simultaneous sorption for all three pesticides.

Diffuse atrazine pollution in German aquifers.

Until its prohibition in Germany in 1991, atrazine was the most frequently applied herbicide in maize cultivation. Moreover, it was used in orchards and vineyards and as a total herbicide on non-cultivated grounds (railways, factory grounds). Later on, atrazine was substituted mainly by terbutylazine. Terbutylazine and terbutryn are the only s-triazines presently permitted in Germany. Nevertheless, atrazine and its metabolite desethylatrazine are by far the most abundant herbicides detected in near surface groundwater. This might be due to wash-outs from the pools of atrazine and its metabolites from the soil into the groundwater or continuing illegal applications. Samples taken from maize fields in 1994 showed that 6.2% of 471 fields tested were treated with atrazine despite the prohibition of its use. Nevertheless, the overall trend is in fact a slow decrease in atrazine concentrations where it is detected in groundwater and, simultaneously often a slight increase in desethylatrazine concentrations. But this is not the case for all sampling points, and increasing concentrations in several aquifers are observed as well. Factors governing the adsorption, degradation, persistence and the possible transfer into the aquifer and the current situation concerning atrazine occurrence in German aquifers will be discussed.