Orhan Ince

Chronic impact of tetracycline on the biodegradation of an organic substrate mixture under anaerobic conditions

The study evaluates the chronic impact of the antibiotic tetracycline on the biodegradation of organic substrate under anaerobic conditions. The experiments involved an anaerobic sequencing batch reactor fed with a synthetic substrate mixture including glucose, starch and volatile fatty acids, and operated in a sequence of different phases with gradually increasing tetracycline doses of 1.65-8.5 mg/L, for more than five months. Tetracycline exerted a terminal/lethal effect at 8.5 mg/L on the microbial community under anaerobic conditions, which caused the inhibition of substrate/COD utilization and biogas generation and leading to a total collapse of the reactor. The microbial activity could not be recovered and re-started within a period of more than 10 days, even after stopping tetracycline dosing. At lower doses, substrate utilization was not affected but a reduction of 10-20% was observed in the biogas/methane generation, suggesting that substrate utilization of tetracycline to the biomass was limiting their bioavailability. During the experiments, tetracycline was partially removed either through biodegradation or conversion into its by-products. The adverse long-term impact was quite variable for fermenting heterotrophic and methanogenic fractions of the microbial community based on changes inflicted on the composition of remaining/residual organic substrate.

Control of organic loading rate using the specific methanogenic activity test during start-up of an anaerobic digestion system

The specific methanogenic activity test (SMA) was used to determine a suitable organic loading rate during the start-up phase of a crossflow ultrafiltration membrane anaerobic reactor system (CUMAR). The SMA test was immediately carried out after seeding the reactor with sludge taken from a municipal wastewater treatment plant digester in order to determine the most acceptable initial organic loading rate. The initial test results during the first week showed that the acetoclastic capacity of the reactor sludge, at an organic loading rate (OLR) of 1 kg COD/m3 · d, was found to be low which resulted in a very poor COD removal efficiency, i.e. 14%. Therefore the OLR was maintained at approximately 0.7 kg COD/m3 · d in the next two weeks of operation resulting in the COD removal efficiency of over 65% in the effluent. Based upon the SMA tests results, the OLRs were either increased or decreased in order to improve performance of the system and to achieve stable conditions which further resulted in an excellent organic matter removal, i.e. over 98% COD and almost 100% BOD removal at an OLR of over 1 kg COD/m3 · d. At this point the ratio of actual methane production (AMP) rate of the CUMAR system to potential methane production (PMP) rate determined from the SMA test was 0.45, which ensured that the CUMAR system could be loaded at higher organic loading rates for the remainder of the operating period.

Application of real-time PCR to determination of combined effect of antibiotics on Bacteria, Methanogenic Archaea, Archaea in anaerobic sequencing batch reactors.

This study evaluated the long-term effects of erythromycin-tetracycline-sulfamethoxazole (ETS) and sulfamethoxazole-tetracycline (ST) antibiotic combinations on the microbial community and examined the ways in which these antimicrobials impact the performance of anaerobic reactors. Quantitative real-time PCR was used to determine the effect that different antibiotic combinations had on the total and active Bacteria, Archae and Methanogenic Archae. Three primer sets that targeted metabolic genes encoding formylterahydrofolate synthetase, methyl-coenzyme M reductase and acetyl-coA synthetase were also used to determine the inhibition level on the mRNA expression of the homoacetogens, methanogens and specifically acetoclastic methanogens, respectively. These microorganisms play a vital role in the anaerobic degradation of organic waste and targeting these gene expressions offers operators or someone at a treatment plant the potential to control and the improve the anaerobic system. The results of the investigation revealed that acetogens have a competitive advantage over Archaea in the presence of ETS and ST combinations. Although the efficiency with which methane production takes place and the quantification of microbial populations in both the ETS and ST reactors decreased as antibiotic concentrations increased, the ETS batch reactor performed better than the ST batch reactor. According to the expression of genes results, the syntrophic interaction of acetogens and methanogens is critical to the performance of the ETS and ST reactors. Failure to maintain the stability of these microorganisms resulted in a decrease in the performance and stability of the anaerobic reactors.

Microbiological study of two-stage anaerobic digestion during start-up

Abstract The changes in the microbial population of a two-stage anaerobic digestion system in terms of number and composition were studied during the start-up phase. The numbers of methanogens and non-methanogens slightly decreased in the upflow filter, whereas in the pre-acidification stage the number of acidogens was reasonably constant. Variations in the morphology of the methanogens occurred along the filter, for example the numbers of Methanosarcina and filamentous bacteria species decreased considerably in the drain, possibly becoming attached to the biofilm along the reactor, whereas in the pre-acidification stage only a few Methanosarcina species were identified. Slight variations within the bacterial populations occurred in terms of composition and numbers in the filter. The number of viable methanogens varied in the filter effluent depending on the OLR changes but this did not adversely affect the COD removal efficiency of the upflow filter. Among the autofluorescent methanogenic population, Methanococcus, Methanobacterium, Methanobrevibacteria , and Methanosarcina species were tentatively identified in the filter.

Development of antibiotic resistance genes in microbial communities during long-term operation of anaerobic reactors in the treatment of pharmaceutical wastewater.

Biological treatment processes offer the ideal conditions in which a high diversity of microorganisms can grow and develop. The wastewater produced during these processes is contaminated with antibiotics and, as such, they provide the ideal setting for the acquisition and proliferation of antibiotic resistance genes (ARGs). This research investigated the occurrence and variation in the ARGs found during the one-year operation of the anaerobic sequencing batch reactors (SBRs) used to treat pharmaceutical wastewater that contained combinations of sulfamethoxazole-tetracycline-erythromycin (STE) and sulfamethoxazole-tetracycline (ST). The existence of eighteen ARGs encoding resistance to sulfamethoxazole (sul1, sul2, sul3), erythromycin (ermA, ermF, ermB, msrA, ereA), tetracycline (tetA, tetB, tetC, tetD, tetE, tetM, tetS, tetQ, tetW, tetX) and class Ι integron gene (intΙ 1) in the STE and ST reactors was investigated by quantitative real-time PCR. Due to the limited availability of primers to detect ARGs, Illumina sequencing was also performed on the sludge and effluent of the STE and ST reactors. Although there was good reactor performance in the SBRs, which corresponds to min 80% COD removal efficiency, tetA, tetB, sul1, sul2 and ermB genes were among those ARGs detected in the effluent from STE and ST reactors. A comparison of the ARGs acquired from the STE and ST reactors revealed that the effluent from the STE reactor had a higher number of ARGs than that from the ST reactor; this could be due to the synergistic effects of erythromycin. According to the expression of genes results, microorganisms achieve tetracycline and erythromycin resistance through a combination of three mechanisms: efflux pumping protein, modification of the antibiotic target and modifying enzymes. There was also a significant association between the presence of the class 1 integron and sulfamethoxazole resistance genes.

Inert COD production in a membrane anaerobic reactor treating brewery wastewater

Abstract In this study, inert COD production in a crossflow ultrafiltration membrane anaerobic reactor (CUMAR) system treating brewery wastewater, was investigated. The system was operated at a wide range of influent chemical oxygen demand (COD) concentrations and solid retention times (SRT) in order to determine residual non-biodegradable COD produced by biomass in the anaerobic reactor. Previous studies showed that the wastewater had practically no inert soluble COD. The inert soluble COD in the effluent would, therefore, have been produced in the anaerobic reactor by the biomass. Significant increases in the effluent COD concentration were observed during the operation of the CUMAR system at SRT lower than 100 days. It was also calculated from the results that the soluble microbial products measured in terms of COD were found to be an average of 2.2% of the influent COD. These microbial products were not removed from the effluent even with extended operation times. Throughout the operation, a maximum of 28.5 kg COD/m 3 day was applied, at which point COD removal efficiencies of no lower than 97% in the anaerobic reactor and 99% in the membrane permeate were achieved during the steady-state operation of the CUMAR system.

Comparison of porous and non-porous media in upflow anaerobic filters when treating dairy wastewater

Abstract The performance of porous and non-porous media in two mesophilic anaerobic upflow filters treating a wastewater from a milk bottling factory was investigated. The results indicated that at higher OLRs the porous media had a much better performance. The reactor with non-porous packing showed instability above an OLR of 4 kg COD/m3 ·d whereas the reactor with the porous packing was still stable at OLRs of up to 21 kg COD/m3 ·d. The behaviour of the media and biomass wash-out were observed at high upflow velocities (UV) and results demonstrated that porous media was not as susceptible to shear stresses as non-porous media. At the end of the investigation, attachment studies were carried out and the results showed a heavy biomass attachment in the porous media, whereas mainly unattached biomass was retained in the voids of the non-porous media.

Inhibition of Volatile Fatty Acid Production in Granular Sludge from a UASB Reactor

Abstract Inhibition of volatile fatty acids (VFA), namely acetate, butyrate, and propionate, on the activity of acetoclastic methanogens within a full-scale upflow anaerobic sludge blanket (UASB) reactor was investigated using specific methanogenic activity (SMA) test. SMA tests were carried out at acetate concentrations in a range of 1000–25,000 mg l−1, butyrate concentrations in a range of 3000–25,000 mg l−1 and propionate concentrations between 500–10,000 mg l−1. Maximum potential methane production (PMP) rates were obtained as 389 ml CH4 gTVS−1 .d−1 at 3000 mg l−1 acetate concentration, 432 ml CH4gTVS−1.d−1 at butyrate concentration of 5000 mg l−1, and 162 mlCH4gTVS−1.d−1 at 1000 mg l−1 propionate concentration. App. 50% and 100% inhibition occurred at acetate concentrations of 13,000 mg l−1 and 25,000 mg l−1, butyrate concentrations of 15,000 mg l−1 and 25,000 mg l−1, and propionate concentrations of 3500 mg l−1 and 5000 mg l−1, respectively.

Biogeographical distribution and diversity of bacterial and archaeal communities within highly polluted anoxic marine sediments from the marmara sea.

Physicochemical and microbiological characterization of anoxic sediments taken from seven highly polluted sites of the Marmara Sea was carried out. The 16S rRNA based microbial community structure analyses were performed using domain-specific PCR followed by denaturant gradient gel electrophoresis (DGGE) and sequencing of characteristic bands. The results showed that the microbial communities in these sediments were diverse and evenly distributed. Relating the prokaryotic and geochemical variables through statistical tools revealed that the microbial diversity in the sediments significantly related to depth, and S, Mn and Fe content of the sediments. Fermentative bacteria, denitrifying bacteria and hydrogenotrophic methanogens were dominant whereas sulfate reducing bacteria were absent in the DGGE patterns. This unusual microbial community structure implied that the newly discovered anaerobic methane oxidation coupled to denitrification process may occur in these subseafloor environments.

Use of PCR-DGGE based molecular methods to assessment of microbial diversity during anaerobic treatment of antibiotic combinations

As it is currently often not know how anaerobic bioreactors, e.g. for biogas production, react if the substrate is contaminated by toxic compounds like antibiotics. This study evaluated how anaerobic sequencing batch reactors were affected by amendments of different antibiotics and stepwise increasing concentrations. The compositions of microbial community were determined in the seed sludge using 16S rRNA gene clone libraries and PCR-DGGE analyses were used for the detection of microbial community changes upon antibiotics additions. According to PCR-DGGE results, the syntrophic interaction of acetogens and methanogens is critical to the performance of the reactors. Failure to maintain the stability of these microorganisms resulted in a decrease in the performance and stability of the anaerobic reactors. Assessment of DGGE data is also useful for suggesting the potential to control ultimate microbial community structure, especially derived from Gram-negative bacteria, through bioaugmentation to successful for antibiotic biodegradation.