Microbial diversity and metabolic inference of diclofenac removal in optimised batch heterotrophic-denitrifying conditions by means of factorial design.

Using the Response Surface Methodology (RSM) and Rotational Central Composite Design (RCCD), this study evaluated the removal of DCF under denitrifying conditions, with ethanol as cosubstrate, in batch reactors, being 1 L Erlenmeyer flasks (330 mL of reactional volume) containing Dofing medium and kept under agitation at 130 rpm and incubated at mesophilic temperature (30 °C). It considered the individual and multiple effects of the variables: nitrate (130 - 230 mg NO3- L-1), DCF (60-100 µg DCF L-1) and ethanol (130 - 230 mg EtOH L-1). The highest drug removal efficiency (17.5%) and total nitrate removal were obtained at 176.6 ± 4.3 mg NO3 -L-1, 76.8 ± 3.7 µg DCF L-1, and 180.0 ± 2.5 mg EtOH L-1. Under such conditions, the addition of ethanol and nitrate was significant for the additional removal of diclofenac (p > 0.05). The prevalence of Rhodanobacter, Haliangium and Terrimonas in the inoculum biomass (activated sludge systems) was identified through the 16S rRNA gene sequencing. The potential of these genera to remove nitrate and degrade diclofenac was inferred, and the main enzymes potentially involved in this process were α-methylacyl-CoA racemase, long-chain fatty acid-CoA ligase, catalases and pseudoperoxidases.

Influence of cosubstrate and hydraulic retention time on the removal of drugs and hygiene products in sanitary sewage in an anaerobic Expanded Granular Sludge Bed reactor.

Diclofenac (DCF), ibuprofen (IBU), propranolol (PRO), triclosan (TCS) and linear alkylbenzene sulfonate (LAS) can be recalcitrant in Wastewater Treatment Plants (WWTP). The removal of these compounds was investigated in scale-up (69 L) Expanded Granular Sludge Bed (EGSB) reactor, fed with sanitary sewage from the São Carlos-SP (Brazil) WWTP and 200 mg L-1 of ethanol. The EGSB was operated in three phases: (I) hydraulic retention time (HRT) of 36±4 h; (II) HRT of 20±2 h and (III) HRT of 20±2 h with ethanol. Phases I and II showed no significant difference in the removal of LAS (63 ± 11-65 ± 12 %), DCF (37 ± 18-35 ± 11 %), IBU (43 ± 18-44 ± 16 %) and PRO (46 ± 25-51 ± 23 %) for 13±2-15 ± 2 mg L-1, 106 ± 32-462 ± 294 µg L-1, 166 ± 55-462 ± 213 µg L-1 and 201 ± 113-250 ± 141 µg L-1 influent, respectively. Higher TCS removal was obtained in phase I (72 ± 17 % for 127 ± 120 µg L-1 influent) when compared to phase II (51 ± 13 % for 135 ± 119 µg L-1 influent). This was due to its greater adsorption (40 %) in the initial phase. Phase III had higher removal of DCF (42 ± 10 % for 107 ± 26 µg L-1 influent), IBU (50 ± 15 % for 164 ± 47 µg L-1 influent) and TCS (85 ± 15 % for 185 ± 148 µg L-1 influent) and lower removal of LAS (35 ± 14 % for 12 ± 3 mg L-1 influent) and PRO (-142 ± 177 % for 188 ± 88 µg L-1 influent). Bacteria similar to Syntrophobacter, Smithella, Macellibacteroides, Syntrophus, Blvii28_wastewater-sludge_group and Bacteroides were identified in phase I with relative abundance of 3.1 %-4.7 %. Syntrophobacter was more abundant (15.4 %) in phase II, while in phase III, it was Smithella (12.7 %) and Caldisericum (15.1 %). Regarding the Archaea Domain, Methanosaeta was more abundant in phases I (84 %) and II (67 %), while in phase III it was Methanobacterium (86 %).

Influence of metabolic cosubstrates on methanogenic potential and degradation of triclosan and propranolol in sanitary sewage.

Triclosan (TCS) and propranolol (PRO) are emerging micropollutants that are difficult to remove in wastewater treatment plants. In this study, methanogenic potential (P) of anaerobic sludge submitted to TCS (3.6 ± 0.1 to 15.5 ± 0.1 mg L-1) and PRO (6.1 ± 0.1 to 55.9 ± 1.2 mg L-1) in sanitary sewage, was investigated in batch reactors. The use of cosubstrates (200 mg L-1 of organic matter) ethanol, methanol:ethanol and fumarate was evaluated for micropollutant degradation. Without cosubstrates, P values for 5.0 ± 0.1 mgTCS L-1, 15.5 ± 0.1 mgTCS L-1 and 55.0 ± 1.3 mgPRO L-1 were 50.53%, 98.24% and 17.66% lower in relation to Control assay (855 ± 5 µmolCH4) with sanitary sewage, without micropollutants and cosubstrates, respectively. The use of fumarate, ethanol and methanol:ethanol favored greater methane production, with P values of 2144 ± 45 µmolCH4, 2960 ± 185 µmolCH4 and 2239 ± 171 µmolCH4 for 5.1 ± 0.1 mgTCS L-1, respectively; and of 10,827 ± 185 µmolCH4, 10,946 ± 108 µmolCH4 and 10,809 ± 210 µmolCH4 for 55.0 ± 1.3 mgPRO L-1, respectively. Greater degradation of TCS (77.1 ± 0.1% for 5.1 ± 0.1 mg L-1) and PRO (24.1 ± 0.1% for 55.9 ± 1.2 mg L-1) was obtained with ethanol. However, with 28.5 ± 0.5 mg PRO L-1, greater degradation (88.4 ± 0.9%) was obtained without cosubstrates. With TCS, via sequencing of rRNA 16S gene, for Bacteria Domain, greater abundance of phylum Chloroflexi and of the genera Longilinea, Arcobacter, Mesotoga and Sulfuricurvum were identified. With PRO, the genus VadinBC27 was the most abundant. Methanosaeta was dominant in TCS with ethanol, while in PRO without cosubstrates, Methanobacterium and Methanosaeta were the most abundant. The use of metabolic cosubstrates is a favorable strategy to obtain greater methanogenic potential and degradation of TCS and PRO.

Methanogenic potential of diclofenac and ibuprofen in sanitary sewage using metabolic cosubstrates.

Diclofenac (DCF) and ibuprofen (IBU) are widely used anti-inflammatory drugs and are frequently detected in wastewater from Wastewater Treatment Plants and in aquatic environments. In this study, the methanogenic potential (P) of anaerobic sludge subjected to DCF (7.11 ± 0.02 to 44.41 ± 0.05 mg L-1) and IBU (6.11 ± 0.01 to 42.61 ± 0.05 mg L-1), in sanitary sewage, was investigated in batch reactors. Cosubstrates (200 mg L-1 of organic matter) in the form of ethanol, methanol:ethanol and fumarate were tested separately for the removal of drugs. In the DCF assays, P was 6943 ± 121 µmolCH4, 9379 ± 259 µmolCH4, 9897 ± 212 µmolCH4 and 11,530 ± 368 µmolCH4 for control, fumarate, methanol:ethanol and ethanol conditions, respectively. In the IBU assays, under the same conditions, P was 6145 ± 101 µmolCH4, 6947 ± 66 µmolCH4, 8141 ± 191 µmolCH4and 10,583 ± 512 µmolCH4, respectively. Without cosubstrates, drug removal was below 18% for 43.10 ± 0.01 mgDCF L-1 and 43.12 ± 0.03 mgIBU L-1, respectively. Higher P and removal of DCF (28.24 ± 1.10%) and IBU (18.72 ± 1.60%) with ethanol was observed for 43.20 ± 0.01 mgDCF L-1 and 43.42 ± 0.03 mgIBU L-1, respectively. This aspect was better evidenced with DCF due to its molecular structure, a condition that resulted in a higher diversity of bacterial populations. Through the 16S rRNA sequencing, bacteria genera capable of performing aromatic ring cleavage, ß-oxidation and oxidation of ethanol and fatty acids were identified. Higher relative abundance (>0.6%) was observed for Smithella, Sulfuricurvum and Synthophus for the Bacteria Domain and Methanosaeta (>79%) for the Archaea Domain. The use of ethanol favored greater mineralization of organic matter and greater methane production, which can directly assist in the metabolic pathways of microorganisms.

The effect of cations (Na+, Mg2+, and Ca2+) on the activity and structure of nitrifying and denitrifying bacterial communities.

Wastewaters generated in regions with water scarcity usually have high alkalinity, hardness, and elevated osmotic pressure (OP). Those characteristics should be considered when using biological systems for wastewater treatment along with the salinity heterogeneity. The interaction of different salts in mixed electrolyte solutions may cause inhibition, antagonism, synergism, and stimulation effects on microbial communities. Little is known about those effects on microbial activity and community structure of nitrifying and denitrifying bacteria. In this work, factorial design was used to evaluate the effects of NaCl, MgCl2 and CaCl2 on nitrifying and denitrifying communities. Antagonistic relationships between all salts were observed and they had greater magnitude on the nitrifying community. Stimulus and synernism were more evident on the nitrifying and denitrifying experiments, respectively. For this reason, the highest nitrification and denitrification specific rates were 1.1 × 10-1 mgN-NH4+ gSSV-1 min-1 for condition 01 and 6.5 × 10-2 mgN-NO3- gSSV-1 min-1 for control condition, respectively. The toxicity of the salts followed the order of NaCl > MgCl2 > CaCl2 and the antagonism between MgCl2 and NaCl was the most significant. PCR/DGGE analyses showed that Mg2+ may be the element that expresses the least influence in the differentiation of microbial structure even though it significantly affects the activity of the autotrophic microorganisms. The same behavior was observed for Ca2+ on denitrifying microorganism. In addition, microbial diversity and richness was not negatively affected by different salinities. Genetic sequencing suggested that the genus Aeromonas, Alishewanella, Azospirillum, Pseudoalteromonas, and Thioalkalivibrio were outstanding on ammonium and nitrate removal under saline conditions. The specific toxicity of each salt and the interactions among them are the major effects on microbial activity in biological wastewater treatments rather than the osmotic pressure caused by the final salinity.

Establishing simultaneous nitrification and denitrification under continuous aeration for the treatment of multi-electrolytes saline wastewater.

Simultaneous nitrification and denitrification (SND) was established under continuous aeration (6 mgO2 L-1) aiming at achieving a feasible and simple operational strategy for treating multi-electrolyte saline wastewaters. Two Structured Fixed-Bed Reactors (SFBR) were used to assess SND performance with (Saline Reactor, SR) and without (Control Reactor, CR) salinity interference. Salinity was gradually increased (from 1.7 to 9 atm) based on the composition of water supplied in arid regions of Brazil. At 1.7 atm, N-NH4+ oxidation and Total Nitrogen (TN) removal efficiencies of 95.9 ±â€¯2.8 and 65.76 ±â€¯7.5%, respectively, were obtained. At osmotic pressure (OP) of 9 atm, the system was severely affected by specific salt toxicity and OP. High chemical oxygen demand (COD) removal efficiency was achieved at all operational conditions (97.2 ±â€¯1.6 to 78.5 ±â€¯4.6%). Salinity did not affect microbial diversity, although it modified microbial structure. Halotolerant genera were identified (Prosthecobacter, Chlamydia, Microbacterium, and Paenibacillus).

Microbial Characterization of Methanogenic and Iron-reducing Consortium in Reactors with Polychlorinated Biphenyls.

Recent papers have confirmed current environmental pollution and the continuous release of polychlorinated biphenyls (PCBs) despite the prohibition of its manufacture worldwide. As the dehalogenating microorganisms are able to remove halogens from various analogous compounds, the characterization of PCB metabolisms can improve the degradation of similar compounds. Thus, this study extensively evaluated the microbial community developed in methanogenic and iron-reducing reactors. The horizontal-flow anaerobic reactor (HAIB) with real waste of Aroclor (1 mL L-1) was fed with mineral medium, ethanol, and sodium formate. Bacteria belonging to Thermotogaceae (Thermotogae), Geobacteraceae, Chloroflexi, Proteobacteria, and Firmicutes (Clostridium) were identified in the HAIB reactor. Bacteria belonging to the Chloroflexi, Firmicutes, and Geobacteraceae are associated with the degradation of hydrocarbons and could be related to the Aroclor waste in this paper. Furthermore, 5.26 × 1012 cells gTVS-1 of iron-reducing bacteria were quantified by the most probable number method in the HAIB reactor, suggesting that this group has an important role in aromatic degradation. Moreover, the evaluation of methanogenic and iron-reducing microorganisms in batch reactors with Aroclor 1260 was performed and the biomass growth was not affected by the addition of PCB. The methane production reached 0.38 µmol CH4 gTVS-1 and the iron reduction attained 90% in batch reactors. Through microbial analyses from HAIB and batch reactors, lower diversity was evidenced in the presence of PCB. This paper indicates the relevant role of iron-reducing organisms and Chloroflexi, Geobacteraceae, and Firmicutes group in PCB metabolism.

Microbial diversity of a full-scale UASB reactor applied to poultry slaughterhouse wastewater treatment: integration of 16S rRNA gene amplicon and shotgun metagenomic sequencing.

The 16S rRNA gene amplicon and whole-genome shotgun metagenomic (WGSM) sequencing approaches were used to investigate wide-spectrum profiles of microbial composition and metabolic diversity from a full-scale UASB reactor applied to poultry slaughterhouse wastewater treatment. The data were generated by using MiSeq 2 × 250 bp and HiSeq 2 × 150 bp Illumina sequencing platforms for 16S amplicon and WGSM sequencing, respectively. Each approach revealed a distinct microbial community profile, with Pseudomonas and Psychrobacter as predominant genus for the WGSM dataset and Clostridium and Methanosaeta for the 16S rRNA gene amplicon dataset. The virome characterization revealed the presence of two viral families with Bacteria and Archaea as host, Myoviridae, and Siphoviridae. A wide functional diversity was found with predominance of genes involved in the metabolism of acetone, butanol, and ethanol synthesis; and one-carbon metabolism (e.g., methanogenesis). Genes related to the acetotrophic methanogenesis pathways were more abundant than methylotrophic and hydrogenotrophic, corroborating the taxonomic results that showed the prevalence of the acetotrophic genus Methanosaeta. Moreover, the dataset indicated a variety of metabolic genes involved in sulfur, nitrogen, iron, and phosphorus cycles, with many genera able to act in all cycles. BLAST analysis against Antibiotic Resistance Genes Database (ARDB) revealed that microbial community contained 43 different types of antibiotic resistance genes, some of them were associated with growth chicken promotion (e.g., bacitracin, tetracycline, and polymyxin).

Biodegradation of linear alkylbenzene sulfonate in commercial laundry wastewater by an anaerobic fluidized bed reactor.

The biodegradation of linear alkylbenzene sulfonate (LAS) from commercial laundry wastewater was evaluated in an anaerobic fluidized bed reactor (FBR) fed with synthetic substrate (598 mg L(-1) to 723 mg L(-1) of organic matter) supplemented with 9.5±3.1 mg L(-1) to 27.9±9.6 mg L(-1) of LAS. The average chemical oxygen demand (COD) removal efficiency was 89% and the biodegradation of LAS was 57% during the 489 days of anaerobic FBR. Higher levels of volatile fatty acids (VFA) were observed in the effluent at the stage with the best LAS removal performance. Increasing the surfactant concentration did not increase the VFA production in the effluent. The predominant VFAs after the addition of LAS were as follows: isovaleric acid and valeric acid, followed by propionic acid, caproic acid and formic acid. The similarities of 64% and 45% to Archaea and Bacteria domains were observed in the samples taken in the operating period of anaerobic FBR fed with 23.6±10 mg L(-1) and 27.9±10 mg L(-1) of LAS. During the operation stages in the reactor, Gemmatimonas, Desulfobulbus and Zoogloea were determined as the most abundant genera related to surfactant degradation using 454-Pyrosequencing.

Optimization of linear alkylbenzene sulfonate (LAS) degradation in UASB reactors by varying bioavailability of LAS, hydraulic retention time and specific organic load rate.

Degradation of linear alkylbenzene sulfonate (LAS) in UASB reactors was optimized by varying the bioavailability of LAS based on the concentration of biomass in the system (1.3-16 g TS/L), the hydraulic retention time (HRT), which was operated at 6, 35 or 80 h, and the concentration of co-substrates as specific organic loading rates (SOLR) ranging from 0.03-0.18 g COD/g TVS.d. The highest degradation rate of LAS (76%) was related to the lowest SOLR (0.03 g COD/g TVS.d). Variation of the HRT between 6 and 80 h resulted in degradation rates of LAS ranging from 18% to 55%. Variation in the bioavailability of LAS resulted in discrete changes in the degradation rates (ranging from 37-53%). According to the DGGE profiles, the archaeal communities exhibited greater changes than the bacterial communities, especially in biomass samples that were obtained from the phase separator. The parameters that exhibited more influence on LAS degradation were the SOLR followed by the HRT.