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.