Intensive application of antibiotics in livestock husbandry can lead to an increasing occurrence of antibiotic-resistant bacteria in manure. Beside the application as biofertilizers, manure is a major input material for biogas plants. The aim of our study was to monitor the presence of antibiotic resistant and potentially pathogenic bacteria in input (mixed manure) and output samples of German biogas plants.
Two biogas plants were investigated in an annual cycle with respect to the presence of Extended Spectrum Beta-lactamases (ESBL)-producing E. coli, Vancomycin resistant Enterococci (VRE) and Methicillin resistant Staphylococci (MRS) using selective pre-enrichment procedures combined with subsequent cultivation approaches on CHROMagar media and resistance-genes-screenings. ESBL-producing E. coli and VRE were cultured from input and output-, MRS only from input material. ESBL-producing E. coli carried mainly blaCTX-M and blaTEM genes and belonged to several ST types (MLST analysis). VRE were most closely related to E. gallinarum- E. casseliflavus, E. viikkiensis and E. lemanii and were shown to carry vanA or vanB genes; MRS carried mecA genes and were most closely related to S. haemolyticus, S. lentus, and S. sciuri.
In addition, the microbial communities of input and output samples of 15 German biogas plants were analyzed once using molecular 16S rRNA gene based approaches. Community fingerprinting by denaturing gradient gel electrophoreses (DGGE) showed diverse community pattern in input, but more similar patterns in output samples. Pyrotaq sequencing showed a relatively low abundance of E. coli, Enterococci, and Staphylococci in input and output samples and a high abundance of uncultured bacteria in output samples. In conclusion, antibiotic resistant bacteria and high amounts of uncultured bacteria with so far unknown function that originate from livestock husbandry are released into the environment in case of application of biogas plant digestates as biofertilizers.