Fate and seasonal change of Escherichia coli resistant to different antibiotic classes at each stage of conventional activated sludge process.

This study investigated the impact of each treatment stage of the activated sludge process on the fate of antibiotic resistant bacteria (ARB) in wastewater treatment plants (WWTPs). Wastewater and sludge samples were collected monthly at each stage of a commercial-scale WWTP. After 20-25 strains of indicator Escherichia coli were isolated from each sample on Chromocult Coliform Agar, antibiotic resistance of the isolates to amoxicillin (AMX), ciprofloxacin (CIP), norfloxacin (NFX), kanamycin (KM), sulfamethoxazole/trimethoprim (ST) and tetracycline (TC) were tested with the Kirby-Bauer disk diffusion method. As a result, activated sludge in the aeration tank and return sludge had higher abundance of antibiotic resistant E. coli than influent wastewater and secondary treatment effluent. AMX resistant E. coli was enriched in return sludge at the secondary clarifier. Higher temperature was also likely to cause an increase of AMX resistant E. coli in sludge. The antibiotic resistance profile of E. coli in secondary treatment effluent was more dependent on activated sludge than influent wastewater. These results suggested that activated sludge in WWTP possibly serves as a reservoir of ARB, and that behavior of ARB in WWTP differs by antibiotic classes.

Toxicity assessment of typical polycyclic aromatic hydrocarbons to Daphnia magna and Hyalella azteca in water-only and sediment-water exposure systems.

While the equilibrium partitioning (EqP) method has been demonstrated to effectively predict the adverse effects of nonionic organic chemicals in sediment on benthic organisms by sediment toxicity tests, only a limited number of studies have been performed both in water-only and whole-sediment toxicity tests using the same species and verified the validity of EqP-based toxicity assessment. To further examine the validity of the EqP method for application in a wide range of hydrophobicity, we conducted sorption/desorption experiments and both water-only and sediment toxicity tests using a popular aquatic crustacean species, Daphnia magna (48 h), and benthic species Hyalella azteca (96 h) for six typical polycyclic aromatic hydrocarbons (PAHs) with three to five rings and an amine derivative: anthracene, phenanthrene, fluoranthene, pyrene, benzo[a]pyrene, dibenzo[a,h]anthracene, and 1-aminopyrene. The linear sorption coefficient was determined and ranged from 2.7 × 102 (phenanthrene) and 1.2 × 104 L/kg (benzo[a]pyrene) highly depending on the hydrophobicity while the aqueous concentrations were stable after 24 h in the desorption test. As result of acute toxicity tests in the water-only exposure system, anthracene and dibenz[a,h]anthracene were found to be nontoxic to both species, while median effect/lethal concentrations (EC50/LC50) were determined as ranging from 0.66 (benzo[a]pyrene) to 330 µg/L (phenanthrene), and from 11 (1-aminopyrene) to 180 µg/L (phenanthrene) for D. magna and H. azteca, respectively. Among these compounds, three PAHs with three, four, and five rings each, and 1-aminopyrene were subjected to sediment-water toxicity tests. In the sediment-water tests, the LC50 of phenanthrene and pyrene was three to six times higher than that of the water-only tests for H. azteca while the EC50 was 1.1 to 2.0 times higher for D. magna. In contrast, the EC50/LC50 of benzo[a]pyrene (BaP) in the sediment-water toxicity test was more than 5 times higher than that in the water-only test for both H. azteca and D. magna. The EC50/LC50 values of 1-aminopyrene were similar in both the sediment-water and the water-only toxicity tests, ranging narrowly from 21 to 28 µg/L and 8.8 to 11 µg/L for D. magna and H. azteca, respectively. The EC50/LC50 based on the body residue (ER50/LR50) was investigated for two of the representative PAHs, pyrene, and BaP. The ER50/LR50 of pyrene in both species was 2.3 and 11 times higher in the water-only toxicity test for D. magna and H. azteca, respectively, while those of BaP in the sediment-water toxicity test were not calculated for the sediment-water toxicity tests, and the highest body concentration in the sediment-water tests was lower than the ER50/LR50 in the water-only toxicity test. Although the experimental results were comparable with the predicted sediment toxicity values based on the EqP method for the selected PAHs in this study, there is a risk of phenanthrene and pyrene being slightly underestimated (1.4-1.9 fold for phenanthrene and 3.7-6.1 fold for pyrene) by the EqP method for H. azteca. These results reaffirm that the bioavailability of poorly water-soluble chemicals is important for sediment toxicity and that the exposure pathway should be further investigated to avoid under- and overestimation via the EqP method.

Application of microalgae hydrolysate as a fermentation medium for microbial production of 2-pyrone 4,6-dicarboxylic acid.

Actual biomass of microalgae was tested as a fermentation substrate for microbial production of 2-pyrone 4,6-dicarboxylic acid (PDC). Acid-hydrolyzed green microalgae Chlorella emersonii (algae hydrolysate) was diluted to adjust the glucose concentration to 2 g/L and supplemented with the nutrients of Luria-Bertani (LB) medium (tryptone 10 g/L and yeast extract 5 g/L). When the algae hydrolysate was used as a fermentation source for recombinant Escherichia coli producing PDC, 0.43 g/L PDC was produced with a yield of 20.1% (mol PDC/mol glucose), whereas 0.19 g/L PDC was produced with a yield of 8.6% when LB medium supplemented with glucose was used. To evaluate the potential of algae hydrolysate alone as a fermentation medium for E. coli growth and PDC production, the nutrients of LB medium were reduced from the algae hydrolysate medium. Interestingly, 0.17 g/L PDC was produced even without additional nutrient, which was comparable to the case using pure glucose medium with nutrients of LB medium. When using a high concentration of hydrolysate without additional nutrients, 1.22 g/L PDC was produced after a 24-h cultivation with the yield of 16.1%. Overall, C. emersonii has high potential as cost-effective fermentation substrate for the microbial production of PDC.

Effects of membrane orientation on fouling characteristics of forward osmosis membrane in concentration of microalgae culture.

Application of forward osmosis (FO) membrane to microalgae cultivation processes enables concentration of microalgae and nutrients with low energy consumption. To understand fouling characteristics of FO membrane in concentration of microalgae culture, we studied flux decline, flux recovery by cleaning, and foulants characteristics, in different membrane orientation of active-layer-facing-feed-solution (AL-FS) and active-layer-facing-draw-solution (AL-DS) modes. Batch concentration of Chlorella vulgaris was conducted with a cellulose-triacetate FO membrane. Rapid flux decline and lower flux recovery was observed in AL-DS mode because of inner-membrane fouling including internal pore clogging, adsorption and internal concentration polarization in the support layer. A proportion of polysaccharides in extracellular polymeric substances to soluble microbial products were larger in chemical cleaning effluent than physical one in AL-DS mode, although those were not significantly different in AL-FS mode. Excitation-emission matrix analysis revealed that proteins and humic-like substances were also possible irreversible foulants both in AL-DS and AL-FS modes.

Saccharification and ethanol fermentation from cholinium ionic liquid-pretreated bagasse with a different number of post-pretreatment washings.

Choline acetate (ChOAc), a cholinium ionic liquid (IL), was compared with 1-ethyl-3-methylimidazolium acetate (EmimOAc) with regard to biomass pretreatment, inhibition on cellulase and yeast, residuals in pretreated biomass, and saccharification and fermentation of pretreated biomass. Irrespective of ChOAc and EmimOAc, cellulose and hemicellulose saccharification of the IL-pretreated bagasse were over 90% and 60%, respectively. Median effective concentrations (EC50) based on cellulase activity were 32 wt% and 16 wt% for ChOAc and EmimOAc, respectively. The EC50 based on yeast growth were 3.1 wt% and 0.3 wt% for ChOAc and EmimOAc respectively. The residuals in IL-pretreated bagasse were 10% and 23% for ChOAc and EmimOAc, respectively, when washed 2 times after pretreatment. Ethanol yield on a bagasse basis were 60% and 24% for ChOAc and EmimOAc, respectively, in the saccharification and fermentation of IL-pretreated bagasse when washed 2 times. ChOAc-pretreated bagasse could be saccharified and fermented with fewer wash times than EmimOAc-pretreated bagasse.

Time-resolved DNA stable isotope probing links Desulfobacterales- and Coriobacteriaceae-related bacteria to anaerobic degradation of benzene under methanogenic conditions.

To identify the microorganisms involved in benzene degradation, DNA-stable isotope probing (SIP) with 13C-benzene was applied to a methanogenic benzene-degrading enrichment culture. Pyrosequencing of ribosomal RNA (rRNA) gene sequences revealed that the community structure was highly complex in spite of a 3-year incubation only with benzene. The culture degraded 98% of approximately 1 mM 13C-benzene and mineralized 72% of that within 63 d. The terminal restriction fragment length polymorphism (T-RFLP) profiles of the buoyant density fractions revealed the incorporation of 13C into two phylotypes after 64 d. These two phylotypes were determined to be Desulfobacterales- and Coriobacteriaceae-related bacteria by cloning and sequencing of the 16S rRNA gene in the 13C-labeled DNA abundant fraction. Comparative pyrosequencing analysis of the buoyant density fractions of 12C- and 13C-labeled samples indicated the incorporation of 13C into three bacterial and one archaeal OTUs related to Desulfobacterales, Coriobacteriales, Rhodocyclaceae, and Methanosarcinales. The first two OTUs included the bacteria detected by T-RFLP-cloning-sequencing analysis. Furthermore, time-resolved SIP analysis confirmed that the activity of all these microbes appeared at the earliest stage of degradation. In this methanogenic culture, Desulfobacterales- and Coriobacteriaceae-related bacteria were most likely to be the major benzene degraders.