Anshuman A. Khardenavis

Simultaneous nitrification and denitrification by diverse Diaphorobacter sp.

Eight bacterial isolates closely related to Diaphorobacter sp. were isolated from activated biomass surviving on wastewater laden with dyes and nitro-substituted chemicals and were identified by 16S rDNA sequence analysis. The isolates showed sequence similarity of 99–100% to other Diaphorobacter strains such as ZY 2006b, F2, NA5, PCA039, D. nitroreducens KSP4, and KSP3 and 98–99% sequence homology to D. nitroreducens NA10B (type strain JCM 11421). Neighbor-joining tree revealed that all the eight strains formed tight cluster together and also showed close clustering with other Diaphorobacter strains. Isolates demonstrated the ability to perform simultaneous nitrification and denitrification under aerobic conditions. Strains HPC 805, 815, 821, and 856 gave highest chemical oxygen demand removal (85–93%) and ammonia removal (92–96%), which correlated well with higher growth rates of the cultures. Simultaneously, complete removal of nitrate supplied in the medium in presence of ammonium and acetate (electron donor) was observed in addition to aerobic nitrite release from ammonium. Thus, the above strains showed ability to perform partial nitrification followed by further aerobic removal of common intermediate nitrite, which indicated their potential application in treatment systems for treatment of high-nitrogen-containing wastewaters.

Processing of poultry feathers by alkaline keratin hydrolyzing enzyme from Serratia sp. HPC 1383

The present study describes the production and characterization of a feather hydrolyzing enzyme by Serratia sp. HPC 1383 isolated from tannery sludge, which was identified by the ability to form clear zones around colonies on milk agar plates. The proteolytic activity was expressed in terms of the micromoles of tyrosine released from substrate casein per ml per min (U/mL min). Induction of the inoculum with protein was essential to stimulate higher activity of the enzyme, with 0.03% feathermeal in the inoculum resulting in increased enzyme activity (45U/mL) that further increased to 90U/mL when 3d old inoculum was used. The highest enzyme activity, 130U/mL, was observed in the presence of 0.2% yeast extract. The optimum assay temperature and pH for the enzyme were found to be 60 degrees C and 10.0, respectively. The enzyme had a half-life of 10min at 60 degrees C, which improved slightly to 18min in presence of 1mM Ca(2+). Inhibition of the enzyme by phenylmethyl sulfonyl fluoride (PMSF) indicated that the enzyme was a serine protease. The enzyme was also partially inhibited (39%) by the reducing agent beta-mercaptoethanol and by divalent metal ions such as Zn(2+) (41% inhibition). However, Ca(2+) and Fe(2+) resulted in increases in enzyme activity of 15% and 26%, respectively. The kinetic constants of the keratinase were found to be 3.84 microM (K(m)) and 108.7 microM/mLmin (V(max)). These results suggest that this extracellular keratinase may be a useful alternative and eco-friendly route for handling the abundant amount of waste feathers or for applications in other industrial processes.

Shifts in microbial community in response to dissolved oxygen levels in activated sludge

This study evaluates the degradative efficiency of activated biomass collected from a Common Effluent Treatment Plant (CETP) under three different dissolved oxygen (DO) levels, 1, 2 and 4mgl(-1). The change in bacterial diversity with reference to DO levels was also analyzed. Results demonstrate that degradative efficiency was the highest, when the reactor was maintained at 4mgl(-1) DO, but amplicon library analysis showed a greater diversity of bacteria in the reactor maintained at 2mgl(-1) DO. Bacteria belonging to the order Desulfuromonadales, Entomoplasmatales, Pasteurellales, Thermales and Chloroflexales have only been detected in this reactor. Ammonia and nitrate levels in all three reactors indicated efficient nitrification process. Results of this study offer new insights into understanding the performance of activated biomass vis-à-vis microbial diversity and degradative efficiency with reference to DO. This information would be useful in improving the efficiency of any wastewater treatment plant.

Biomethanation of vegetable market waste in an anaerobic baffled reactor: Effect of effluent recirculation and carbon mass balance analysis.

In the present study, feasibility of biomethanation of vegetable market waste in a 4-chambered anaerobic baffled reactor (ABR) was investigated at 30d hydraulic retention time and organic loading rate of 0.5gVS/L/d for one year. Indicators of process stability viz., butyrate/acetate and propionate/acetate ratios were consistent with phase separation in the different chambers, which remained unaltered even during recirculation of effluent. Chemical oxygen demand (COD) and volatile solids (VS) removal efficiencies were observed to be consistently high (above 90%). Corresponding biogas and methane yields of 0.7-0.8L/g VS added/d and 0.42-52L/g VS added/d respectively were among the highest reported in case of AD of vegetable waste in an ABR. Process efficiency of the ABR for vegetable waste methanation, which is indicated by carbon recovery factor showed that, nearly 96.7% of the input carbon considered for mass balance was accounted for in the product.

Diverse Metabolic Capacities of Fungi for Bioremediation

Bioremediation refers to cost-effective and environment-friendly method for converting the toxic, recalcitrant pollutants into environmentally benign products through the action of various biological treatments. Fungi play a major role in bioremediation owing to their robust morphology and diverse metabolic capacity. The review focuses on different fungal groups from a variety of habitats with their role in bioremediation of different toxic and recalcitrant compounds; persistent organic pollutants, textile dyes, effluents from textile, bleached kraft pulp, leather tanning industries, petroleum, polyaromatic hydrocarbons, pharmaceuticals and personal care products, and pesticides. Bioremediation of toxic organics by fungi is the most sustainable and green route for cleanup of contaminated sites and we discuss the multiple modes employed by fungi for detoxification of different toxic and recalcitrant compounds including prominent fungal enzymes viz., catalases, laccases, peroxidases and cyrochrome P450 monooxygeneses. We have also discussed the recent advances in enzyme engineering and genomics and research being carried out to trace the less understood bioremediation pathways.

Identification and monitoring of nitrification and denitrification genes in Klebsiella pneumoniae EGD-HP19-C for its ability to perform heterotrophic nitrification and aerobic denitrification

Microbes capable of performing heterotrophic nitrification and aerobic denitrification simultaneously have application in nitrogen level management in effluent treatment plants. Klebsiella pneumoniae EGD-HP19-C is a metabolically versatile bacterium capable of utilising NH3–N, NO2–N and NO3–N as sole sources of nitrogen. The annotation was done for the genes involved in N-assimilation and N-dissimilation pathways from the draft genome sequences of this bacterium (NCBI GenBank accession no. AUTW02000000.1). The sequence data also suggested possible existence of plasmid associated with this bacterium. Multiple gene sequence alignments of glutamine synthetase (gln), hydroxylamine reductase (har), nitrite reductase (nir), nitric oxide reductase (nor), assimilatory nitrate reductase (nas) and respiratory nitrate reductase (nar) genes from EGD-HP19-C genome were performed to compare sequence identities with that of closely related bacterial species. The metabolic pathways were mapped using KAAS and 3D structures for representative enzyme sub-units were also elucidated. The study suggested that the organism, though it has incomplete nitrification and denitrification pathways still removes the inorganic nitrogen content from the system via ammonification reaction.

Activated sludge is a potential source for production of biodegradable plastics from wastewater

Increased utilization of synthetic plastics caused severe environmental pollution due to their non-biodegradable nature. In the search for environmentally friendly materials to substitute for conventional plastics, different biodegradable plastics have been developed by microbial fermentations. However, limitations of these materials still exist due to high cost. This study aims at minimization of cost for the production of biodegradable plastics P(3HB) and minimization of environmental pollution. The waste biological sludge generated at wastewater treatment plants is used for the production of P(3HB) and wastewater is used as carbon source. Activated sludge was induced by controlling the carbon : nitrogen ratio to accumulate storage polymer. Initially polymer accumulation was studied by using different carbon and nitrogen sources. Maximum accumulation of polymer was observed with carbon source acetic acid and diammonium hydrogen phosphate (DAHP) as nitrogen source. Further studies were carried out to optimize the carbon : nitrogen ratios using acetic acid and DAHP. A maximum of 65.84% (w/w) P(3HB) production was obtained at C/N ratio of 50 within 96 hours of incubation.

Mining of hemicellulose and lignin degrading genes from differentially enriched methane producing microbial community

Study creates a scenario for enrichment and selection of ligno-hemicellulose degrading genotypes with anaerobic bioreactor as a model using rice straw, vegetable waste and food waste as substrates. Relative discrimination analysis showed that the hydrolytic pathways and associated microbial communities for ligno-hemicellulose degradation were dominatingly colonized with rice straw as substrate. The dominating bacteria were Caldicellulosiruptor, Fervidobacterium, Cytophaga, Ruminococcus, Thermotoga associated with hemicellulose degradation and Burkholderia, Pandorea, Sphingomonas, Spirochaeta, Pseudomonas for lignocellulose hydrolysis. This was further supported by the abundance of anaerobic aromatic compound degrading genes along with genes for xylanase and xylosidase in rice straw enriched community. The metagenome analysis data was validated by evaluation of the biochemical methane potential for these substrates. Food waste being most amenable substrate yielded 1410mL of biogas/gVS added whereas, biogas yield of 1160mL/gVS and 1080mL/gVS was observed in presence of vegetable waste and rice straw respectively.

Management of various organic fractions of municipal solid waste via recourse to VFA and biogas generation.

A hybrid anaerobic solid–liquid system was used for anaerobic digestion of organic fraction of municipal solid waste (OFMSW) consisting of mixed food + fruit waste and vegetable waste. Hydrolysis and acidogenesis potential of the above wastes were evaluated with the aim of producing value-added products in the form of volatile fatty acids (VFAs) and biogas recovery. Efficient hydrolysis and acidogenesis of mixed food + fruit waste was observed at a hydraulic retention time (HRT) of 1–3 d with a five-fold increase in soluble chemical oxygen demand (SCOD) followed by VFA production consisting of 50–75% acetic acid. Longer time was required for hydrolysis of vegetable waste with optimum hydrolysis and SCOD generation at 9 d HRT followed by VFA synthesis consisting of 45% acetic acid. Higher inoculum:substrate ratios resulted in improved hydrolysis and acidogenesis rates for vegetable waste in shorter time of 6 d with higher VFA production and increase in acetic acid content to 70%. When acidogenic leachate was fed into methanogenic reactors, detectable biogas production was observed after 25 d with 37–53% SCOD removal from leachate from mixed food + fruit waste and methane production of 0.066–0.1 L g−1 SCOD removed and methane content of 38%. Though biogas yield from acidogenic leachate from vegetable waste was lower, nearly 94% volatile solids (VS) removal was observed in the reactors thereby providing methane yield of 0.13–0.21 L g−1 VS consumed. Thus, the study provided a method for generation of value-added products from an otherwise misplaced resource in the form of OFMSW.

Utilization of molasses spentwash for production of bioplastics by waste activated sludge

Present study describes the treatment of molasses spentwash and its use as a potential low cost substrate for production of biopolymer polyhydroxybutyrate (PHB) by waste activated sludge. Fluorescence microscopy revealed the presence of PHB granules in sludge biomass which was further confirmed by fourier transform-infra-red spectroscopy (FT-IR) and (13)C nuclear magnetic resonance (NMR). The processing of molasses spentwash was carried out for attaining different ratios of carbon and nitrogen (C:N). Highest chemical oxygen demand (COD) removal and PHB accumulation of 60% and 31% respectively was achieved with raw molasses spentwash containing inorganic nitrogen (C:N ratio=28) followed by COD removal of 52% and PHB accumulation of 28% for filtered molasses containing inorganic nitrogen (C:N ratio=29). PHB production yield (Y(p/s)) was highest (0.184 g g(-1) COD consumed) for deproteinized spentwash supplemented with nitrogen. In contrast, the substrate consumption and product formation were higher in case of raw spentwash. Though COD removal was lowest from deproteinized spentwash, evaluation of kinetic parameters suggested higher rates of conversion of available carbon to biomass and PHB. Thus the process provided dual benefit of conversion of two wastes viz. waste activated sludge and molasses spentwash into value-added product-PHB.