Combating bacterial infections by antibiotic treatment is one of the greatest achievements in medicine. However, once administered antibiotic compounds are often not metabolized completely in humans and animals and are thus excreted, eventually ending up in sewage sludge or manure. As both are used as fertilizers on agricultural lands they represent a route for antibiotics into the soil environment where they may persist and affect levels of antibiotic
resistance in soil microbial communities over time. In this work the level of tolerance to the antibiotic sulfadiazine (SDZ) was studied in a number of soils applying the pollution-induced community tolerance (PICT)-approach. As SDZ is amphoteric and thus exist on either neutral, anionic or cationic form soil pH is likely to influence the toxicity and bioavailability of SDZ to soil bacteria.
In manuscript I the aim was to set a baseline, a PICT-threshold, of a non-contaminated soil environment at various pH of which to compare other soils. Soil samples representing a broad range of natural pH were collected from the pH gradient at the Hoosfield acid strip, part of the long-term field experiment at the Rothamstead Research Station (UK) and exposed to increasing SDZ-concentrations. Surprisingly, induced community tolerance was not detected either by [3H]-leucine incorporation or by quantification of two genes conferring resistance to sulfonamides. In contrast did SDZ-amendments lead to detection of PICT in soil samples from a long-term field experiment designed to test effects on soil quality of a range of different fertilizers in agriculture.
In manuscript II extracted bacteria from soil samples representing a broad range of natural soil pH values were tested for their toxicity response to SDZ when amended at different assay pH. Toxicity clearly increased with decreasing pH and SDZ inhibited growth at concentrations as low as 10 μg L-1 which is in the range found in manured soils.
In manuscript III experiments were conducted to test long-term effects of different fertilizers on the ability of SDZ to induce PICT-responses and to investigate the fate and transport of SDZ at the interphase between dewatered SDZ-amended sewage sludge and soil. SDZ was not mineralized within sludge aggregates and travelled more than 10 mm into the surrounding soil. The strongest PICT response was observed in soils fertilized with organic fertilizers or inorganic NPK fertilizer, whereas only a very weak PICT response was found for unfertilized soil. In conclusion, I show that fertilized soils are more conducive for PICT development, and therefore presumably also for selection of antibiotic resistance in individual bacteria, than unfertilized soil and that SDZ-loaded sludge can serve as a significant source of bioavailable SDZ to the surrounding soil. The sludge-soil interphase may therefore constitute a hotspot for proliferation of antibiotic resistance.
Collectively, results presented in this thesis indicate that selection for SDZ tolerant bacteria may take place at environmentally relevant concentrations at low pH especially in fertilized soils.