Sevcan Aydin

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.

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.

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.

Assessment of anaerobic bacterial diversity and its effects on anaerobic system stability and the occurrence of antibiotic resistance genes

This study evaluated the link between anaerobic bacterial diversity and, the biodegradation of antibiotic combinations and assessed how amending antibiotic combination and increasing concentration of antibiotics in a stepwise fashion influences the development of resistance genes in anaerobic reactors. The biodegradation, sorption and occurrence of the known antibiotic resistance genes (ARGs) of erythromycin and tetracycline were investigated using the processes of UV-HPLC and qPCR analysis respectively. Ion Torrent sequencing was used to detect microbial community changes in response to the addition of antibiotics. The overall results indicated that changes in the structure of a microbial community lead to changes in biodegradation capacity, sorption of antibiotics combinations and occurrence of ARGs. The enhanced biodegradation efficiency appeared to generate variations in the structure of the bacterial community. The results suggested that controlling the ultimate Gram-negative bacterial community, especially Acinetobacter-related populations, may promote the successful biodegradation of antibiotic combinations and reduce the occurrence of ARGs.

Combined effect of erythromycin, tetracycline and sulfamethoxazole on performance of anaerobic sequencing batch reactors.

The combined effects of erythromycin-tetracycline-sulfamethoxazole (ETS) and sulfamethoxazole-tetracycline (ST) antibiotics on the performance of anaerobic sequencing batch reactors were studied. A control reactor was fed with wastewater that was free of antibiotics, while two additional reactors were fed with ETS and ST. The way in which the ETS and ST mixtures impact COD removal, VFA production, antibiotic degradation, biogas production and composition were investigated. The effects of the ETS mixtures were different from the ST mixtures, erythromycin can have an antagonistic effect on sulfamethoxazole and tetracycline. The anaerobic pre-treatment of these antibiotics can represent a suitable alternative to the use of chemical treatments for concentrations at 10 mg/L of S and 1 mg/L of T; 2 mg/L of E, 2 mg/L of T and 20 mg/L of S for the ST and ETS reactors respectively, which corresponds to min 70% COD removal efficiency.

Inhibitory effects of antibiotic combinations on syntrophic bacteria, homoacetogens and methanogens.

Antibiotics have the potential to adversely affect the microbial community that is present in biological wastewater treatment processes. The antibiotics that exist in waste streams directly inhibit substrate degradation and also have an influence on the composition of the microbial community. The aim of this study was to evaluate the short-term inhibition impact that various antibiotic combinations had on the syntrophic bacteria, homoacetogenic and methanogenic activities of a microbial community that had been fed with propionate and butyrate as the sole carbon source and VFA mixture (acetate, propionate and butyrate). Acute tests were constructed using on a two way-factorial design, where one factor was the composition of antibiotic mixture and another was the concentration of antibiotics added. In addition, the inhibitory effect of antibiotics was evaluated by monitoring biogas production and the accumulation of individual volatile fatty acids. Specific methanogenic activity batch tests showed a significant (p<0.05) decrease in the maximum methane production rate in the presence of 1 mg L(-1) of antibiotics for the substrate in a VFA mixture and propionate; 1 mg L(-1) of ETS, 25 mg L(-1) of ET, 10 mg L(-1) of ST and ES combination for substrates butyrate. The addition of antibiotics to the batch tests affected the utilization of acetate, propionate and butyrate. This study indicated that antibiotic mixtures have an effect on homoacetogenic bacteria and methanogens, which may exert inhibitory effects on propionate and butyrate-oxidizing syntrophic bacteria, resulting in unfavorable effects on methanogenesis.

Evaluation of microbial population and functional genes during the bioremediation of petroleum-contaminated soil as an effective monitoring approach

This study investigated the abundance and diversity of soil n-alkane and polycyclic aromatic hydrocarbon (PAH)-degrading bacterial communities. It also investigated the quantity of the functional genes, the occurrence of horizontal gene transfer (HGT) in the identified bacterial communities and the effect that such HGT can have on biostimulation process. Illumina sequencing was used to detect the microbial diversity of petroleum-polluted soil prior to the biostimulation process, and quantitative real-time PCR was used to determine changes in the bacterial community and functional genes (alkB, phnAc and nah) expressions throughout the biostimulation of petroleum-contaminated soil. The illumine results revealed that γ-proteobacteria, Chloroflexi, Firmicutes, and δ-proteobacteria were the most dominant bacterial phyla in the contaminated site, and that most of the strains were Gram-negative. The results of the gene expression results revealed that gram-negative bacteria and alkB are critical to successful bioremediation. Failure to maintain the stability of hydrocarbon-degrading bacteria and functional gene will reduce the extend to which alkanes and PAHs are degraded. According to the results of the study, the application of a C:N:P ratio of was 100:15:1 in the biodegradation experiment resulted in the highest rate at which petroleum hydrocarbons were biodegraded. The diversity of pollutant-degrading bacteria and the effective transfer of degrading genes among resident microorganisms are essential factors for the successful biostimulation of petroleum hydrocarbons. As such, screening these factors throughout the biostimulation process represents an effective monitoring approach by which the success of the biostimulation can be assessed.

Performance of anaerobic sequencing batch reactor in the treatment of pharmaceutical wastewater containing erythromycin and sulfamethoxazole mixture.

This study evaluates the joint effects of erythromycin-sulfamethoxazole (ES) combinations on anaerobic treatment efficiency and the potential for antibiotic degradation during anaerobic sequencing batch reactor operation. The experiments involved two identical anaerobic sequencing batch reactors. One reactor, as control unit, was fed with synthetic wastewater while the other reactor (ES) was fed with a synthetic substrate mixture including ES antibiotic combinations. The influence of ES antibiotic mixtures on chemical oxygen demand (COD) removal, volatile fatty acid production, antibiotic degradation, biogas production, and composition were investigated. The influent antibiotic concentration was gradually increased over 10 stages, until the metabolic collapse of the reactors, which occurred at 360 days for the ES reactor. The results suggest that substrate/COD utilization and biogas/methane generation affect performance of the anaerobic reactors at higher concentration. In addition, an average of 40% erythromycin and 37% sulfamethoxazole reduction was achieved in the ES reactor. These results indicated that these antibiotics were partly biodegradable in the anaerobic reactor system.

Enhanced biodegradation of antibiotic combinations via the sequential treatment of the sludge resulting from pharmaceutical wastewater treatment using white-rot fungi Trametes versicolor and Bjerkandera adusta

While anaerobic treatment is capable of treating pharmaceutical wastewater and removing antibiotics in liquid phases, solid phases may still contain significant amounts of antibiotics following this treatment. The main goal of this study was to evaluate the use of white-rot fungi to remove erythromycin, sulfamethoxazole, and tetracycline combinations from biosolids. The degradation potential of Trametes versicolor and Bjerkandera adusta was evaluated via the sequential treatment of anaerobic sludge. Polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) analyses were used to identify competition between the autochthonous microbial communities and white-rot fungi. Solid-phase treatment using white-rot fungi substantially reduced antibiotic concentrations and toxicity in sludge. According to PCR-DGGE results, there is an association between species of fungus and antibiotic type as a result of the different transformation pathways of fungal strains. Fungal post-treatment of sludge represents a promising method of removing antibiotic combinations, therefore holding a significant promise as an environmentally friendly means of degrading the antibiotics present in sludge.

Microbial monitoring of ammonia removal in a UASB reactor treating pre-digested chicken manure with anaerobic granular inoculum

Performance and microbial community dynamics in an upflow anaerobic sludge bed (UASB) reactor coupled with anaerobic ammonium oxidizing (Anammox) treating diluted chicken manure digestate (Total ammonia nitrogen; TAN=123±10mg/L) were investigated for a 120-d operating period in the presence of anaerobic granular inoculum. Maximum TAN removal efficiency reached to above 80% with as low as 20mg/L TAN concentrations in the effluent. Moreover, total COD (tCOD) with 807±215mg/L in the influent was removed by 60-80%. High-throughput sequencing revealed that Proteobacteria, Actinobacteria, and Firmicutes were dominant phyla followed by Euryarchaeota and Bacteroidetes. The relative abundance of Planctomycetes significantly increased from 4% to 8-9% during the late days of the operation with decreased tCOD concentration, which indicated a more optimum condition to favor ammonia removal through anammox route. There was also significant association between the hzsA gene and ammonia removal in the UASB reactor.