Gokhan Turker

The way that antibiotic residues in manure follow is one of the greatest concerns due to its potential negative impacts on microbial communities, the release of metabolites and antibiotic resistant genes (ARGs) into the nature and the loss of energy recovery in anaerobic digestion (AD) systems. This study evaluated the link between different operating conditions, the biodegradation of oxytetracycline (OTC) and the formation of its metabolites and ARGs in anaerobic digesters treating cow manure. Microbial communities and ARGs were determined through the use of quantitative real-time PCR. The biodegradation of OTC and occurrence of metabolites were determined using UV-HPLC and LC/MS/MS respectively. The maximum quantity of resistance genes was also examined at the beginning of AD tests and concentration was in the order of: tetM >tetO. The numbers of ARGs were always higher at high volatile solids (VS) content and high mixing rate. The results of the investigation revealed that relationship between mixing rate and VS content plays a crucial role for elimination of ARGs, OTC and metabolites. This can be attributed to high abundance of microorganisms due to high VS content and their increased contact with elevated mixing rate. An increased interaction between microorganisms triggers the promotion of ARGs.

Management of manure containing veterinary antibiotics is a major concern in anaerobic treatment systems because of their possible adverse effects on microbial communities. Therefore, the aim of study was to investigate how oxytetracycline (OTC) influences bacteria and acetoclastic and hydrogenotrophic methanogens under varying operational conditions in OTC-medicated and non-medicated anaerobic cow manure digesters. Concentrations of OTC and its metabolites throughout the anaerobic digestion were determined using ultraviolet-high-performance liquid chromatography (UV-HPLC) and tandem liquid chromatography-mass spectrometry (LC/MS/MS), respectively. Fluorescent in situ hybridization, denaturing gradient gel electrophoresis, cloning, and sequencing analyses were used to monitor changes in microbial community structures. According to the results of analytical and molecular approaches, operating conditions highly influence active microbial community dynamics and associate with biogas production and elimination of OTC and its metabolites during anaerobic digestion of cow manure in the presence of an average initial concentration of 2.2 mg OTC/L. The impact of operating conditions has a drastic effect on acetoclastic methanogens than hydrogenotrophic methanogens and bacteria.

The effect of veterinary antibiotics in anaerobic digesters is a concern where methane production efficiency is highly dependent on microbial community structure. In this study, both anaerobic degradation of a common veterinary antibiotic, oxytetracycline (OTC), and its effects on an anaerobic digester microbial community were investigated. Qualitative and quantitative molecular tools were used to monitor changes in microbial community structure during a 60-day batch incubation period of cow manure with the addition of different concentrations of the antibiotic. Molecular data were interpreted by a further redundancy analysis as a multivariate statistics approach. At the end of the experiment, approximately 48, 33, and 17 % of the initially added 50, 100, and 200 mg l−1 of OTC was still present in the serum bottles which reduced the biogas production via accumulation of some of the volatile fatty acids (VFAs). Biogas production was highly correlated with Methanobacteriales and Methanosarcinales gene copy numbers, and those parameters were negatively affected with oxytetracycline and VFA concentrations.

This study aimed to determine the fate and effect of oxytetracycline (OTC) and its metabolites during thermophilic anaerobic digestion of cow manure. OTC-medicated and non-medicated digesters were operated at 55°C with different volatile solids (VS) concentrations (4% and 6%) and mixing rates (90 and 120rpm). OTC and its metabolites were measured by HPLC and LC/MS/MS, respectively. Microbial community dynamics were monitored by denaturing gradient gel electrophoresis (DGGE) and real-time PCR (qPCR). Approximately 2mg/L initial OTC concentration caused 10-30% inhibition on biogas production and higher inhibition was observed as mixing rate increased. DGGE results indicated that OTC caused a shift in bacterial community structure and several species became dominant with time. Archaeal community decreased throughout the digestion period. RNA based qPCR analyses showed that gene copy numbers of bacteria and Methanomicrobiales declined in all digesters whereas gene copy numbers of Methanobacteriales and Methanosarcinales increased in high mixing rate digesters.

Possible adverse effects of a commonly used veterinary antibiotic, oxytetracycline (OTC), on acidogenic phase of anaerobic digestion of cattle manure along with optimum operating conditions were investigated. A standard veterinary practice of 50 ml OTC solution (20 mg/kg cattle weight) was injected into the muscles of cattle and then manure samples were collected for 5 days following the injection. The 5-day samples were equally mixed and used throughout digestion experiments. Preliminary batch tests were conducted to obtain the optimum pH range and observe volatile fatty acids (VFAs) production. In this regard, different sets of batch digesters were operated at pH ranging from 5.2 +/- 0.1 to 5.8 +/- 0.1 at mesophilic conditions with total solids content of 6.0 +/- 0.2%. The pH of 5.5 +/- 0.1 was found to be the optimum value for acidification for both non-medicated and OTC-medicated conditions. Under predetermined conditions, maximum total VFA (VFAtot) of 830 +/- 3 mg (as acetic acid)/L was produced and maximum acidification rate was evaluated as 11% for OTC-medicated cattle manure, whereas they were 900 +/- 6 mg (as acetic acid)/L and 12% for non-medicated manure. Digestion studies were further continued in a semi-continuous mode at pH 5.5 +/- 0.1 and SRT/HRT of 5 days. VFAtot concentrations and maximum acidification rate increased up to 2181 +/- 19 mg (as acetic acid)/L and 29% for non-medicated cattle manure. For OTC-medicated cattle manure, lower acidification rate of 18% was observed.

The aim of this study was to investigate the effect of a common veterinary antibiotic in biogas plants. 20 mg/kg of oxytetracycline was intramuscularly injected into a cow and its concentration in manure, which was sampled daily during the following 20 days, was measured. A total of 20 % of the injected oxytetracycline was detected in manure. Collected manure samples on days 1, 2, 3, 5, 10, 15, and 20 were digested in triplicate serum bottles at 37 °C for 30 days. Control serum bottles produced 255 ± 13 mL biogas, whereas 50-60 % inhibitions were obtained for the serum bottles operated with samples collected for the 5 days after medication. Multivariate statistics used for the evaluation of FISH results showed that Methanomicrobiales were the main methanogenic group responsible for most of the biogas production. Numbers of active Bacteria and Methanomicrobiales were negatively correlated with the presence of oxytetracycline, whereas Methanosarcinales and Methanobacteriales were less affected.

The way that antibiotic residues in manure follow is one of the greatest concerns due to its potential negative
impacts on microbial communities, the release of metabolites and antibiotic resistant genes (ARGs) into the
nature and the loss of energy recovery in anaerobic digestion (AD) systems. This study evaluated the link between
different operating conditions, the biodegradation of oxytetracycline (OTC) and the formation of its
metabolites and ARGs in anaerobic digesters treating cow manure. Microbial communities and ARGs were determined
through the use of quantitative real-time PCR. The biodegradation of OTC and occurrence of metabolites
were determined using UV-HPLC and LC/MS/MS respectively. The maximum quantity of resistance genes
was also examined at the beginning of AD tests and concentration was in the order of: tetM>tetO. The numbers
of ARGs were always higher at high volatile solids (VS) content and high mixing rate. The results of the investigation
revealed that relationship between mixing rate and VS content plays a crucial role for elimination of
ARGs, OTC and metabolites. This can be attributed to high abundance of microorganisms due to high VS content
and their increased contact with elevated mixing rate. An increased interaction between microorganisms triggers
the promotion of ARGs.