Plasmid-associated biodegradation of an organophosphorus pesticide, Monocrotophos, by Pseudomonas mendocina

Pseudomonas mendocina isolated from soil degraded an insecticide, Monocrotophos (MCP), by 67% and harbored a 7.4 kb plasmid, designated as pMCP424. On the basis of curing and transformation experiments, MCP degradation by Pseudomonas mendocinawas plasmid-borne and transferable to other bacteria.     

The Potentiality of Free Gram-negative Bacteria for Removing Oil and Grease from Contaminated Industrial Effluents

Summary Eight bacterial species were isolated from vegetable oil and grease-contaminated industrial wastewater, only four of which were found to have the ability to degrade oil and grease in the contaminated wastewater. These isolates were identified according to morphological and biochemical profiles as, Pseudomonas sp. (L1), P. diminuta (L2), P. pseudoalcaligenes (L3), and Escherichia sp. (L5). The degradative capabilities of the identified bacterial isolates for Tween 20 (Tw20) were investigated under different pH levels (6.5, 7, 7.5, and 8), different temperatures (30 and 37 °C) and different concentrations of Tw20 (1, 1.5, and 2%). Results revealed differences in their optimum conditions for maximum degradation of vegetable oil. Bacterial isolates were tested individually or in combinations using synthetic aqueous medium supplemented with 1% palm oil, incubated at 30 °C, and agitated at 150 rev/min for 13 days. All the tested bacteria were able to degrade the palm oil completely and utilized the free fatty acids (FFA) as a carbon source. The combination M1 (Pseudomonas sp. and P. diminuta) produced the highest degradative activity, followed by M3 (Pseudomonas sp., P. diminuta and P. pseudoalcaligenes). Also M1 produced the highest activity in reducing COD (93%) and BOD₅ (100%).     

Growth of <Emphasis Type="Italic">Salinispora tropica</Emphasis> strains CNB440, CNB476, and NPS21184 in nonsaline,low-sodium media

Effective wastewater treatment using microbial fuel cells (MFCs) will require a better understanding of how operational parameters and solution chemistry affect treatment efficiency, but few studies have examined power generation using actual wastewaters. The efficiency of wastewater treatment of a beer brewery wastewater was examined here in terms of maximum power densities, Coulombic efficiencies (CEs), and chemical oxygen demand (COD) removal as a function of temperature and wastewater strength. Decreasing the temperature from 30°C to 20°C reduced the maximum power density from 205 mW/m² (5.1 W/m³, 0.76 A/m²; 30°C) to 170 mW/m² (20°C). COD removals (R _COD) and CEs decreased only slightly with temperature. The buffering capacity strongly affected reactor performance. The addition of a 50-mM phosphate buffer increased power output by 136% to 438 mW/m², and 200 mM buffer increased power by 158% to 528 mW/m². In the absence of salts (NaCl), maximum power output varied linearly with wastewater strength (84 to 2,240 mg COD/L) from 29 to 205 mW/m². When NaCl was added to increase conductivity, power output followed a Monod-like relationship with wastewater strength. The maximum power (P _max) increased in proportion to the solution conductivity, but the half-saturation constant was relatively unaffected and showed no correlation to solution conductivity. These results show that brewery wastewater can be effectively treated using MFCs, but that achievable power densities will depend on wastewater strength, solution conductivity, and buffering capacity.     

Fungal remediation of Amarula distillery wastewater

Amarula Cream is an alcoholic beverage derived from the distillation of fermented marula fruit and to date there is no scientific data as to the characteristics of the distillery wastewater generated from its production. The wastewater was found to have a chemical oxygen demand (COD) of 27 g/l, a pH of 3.8, a high concentration of phenolic compounds (866 mg/l) and a high suspended solids content (10.5 g/l), all of which could adversely affect biological treatment. Full-strength wastewater was treated using shake-flask monocultures of four white rot fungi (Trametes pubescens MB 89, Ceriporiopsis subvermispora, Pycnoporus cinnabarinus or Phanerochaete chrysosporium) at pH 5.0 with no additional carbon or nitrogen supplements. Trametes pubescens performed the best with regards to degrading phenolic compounds, COD and colour, while P. cinnabarinus improved the pH to the greatest extent. Laccase synthesis was only detected in the T. pubescens and C. subvermispora cultures. Six wastewater concentrations (100, 80, 60, 40, 20 and 10%) were assessed at pH 4.5 to establish an optimum concentration for remediation and laccase production by T. pubescens. Similar COD removal efficiencies (71–77%) and phenolic removal efficiencies (87–92%) were achieved at all concentrations. The phenolic removal efficiencies improved by approximately 5% compared to the screening experiment at pH 5.0, indicating that the laccase was more efficient at pH 4.5. The pH became more basic as a result of treatment and the colour decreased for samples below 60% wastewater concentration. The maximum laccase activity (1063 ± 26 units/l) was obtained in the 80% wastewater concentration. This study has resulted in the first characterization of Amarula distillery wastewater and showed that it has a high phenolic compound concentration, COD and suspended solids content. It was possible to biologically treat the wastewater at full strength using a number of white-rot fungi just by raising the pH.     

A Novel High Biosorbent of Pb-resistant Bacterium Isolate for the Removal of Hazardous Lead from Alkaline Soil and Water: Biosorption Isotherms In Vivo and Bioremediation Strategy

A novel Pb-resistant bacterium was isolated from aged lead-contaminated alkaline soils, and was identified as Bacillus megaterium via the MIDI protocol. The biosorption isotherms and kinetics of Pb(II) associated with B. megaterium in vivo in the alkaline environment were investigated at the first time. All the batch experiments of biosorption demonstrate that the B. megaterium uptake of lead is pH-dependent, exothermic (ΔH° = ?5224.86 KJ mol^?1), spontaneous, and fits well with the Langmuir isotherm, resulting in different kinetics under different examination temperatures. The maximum biosorption capacity is 503.86 mg g^?1 at optimum conditions, which is much better in comparison to the biosorbent reported at the acidic condition in the literature. The Fourier-transform Infrared spectroscopic analysis of lead-loaded biomass confirms that the biosorption between B. megaterium and lead is the chemical adsorption in vivo. A site test indicates that B. megaterium really increases mobility and bioavailability of lead in Pb-contaminated alkaline soil in terms of chemical fractionation in vivo, which will potentially increase its uptake by hyperaccumulated plants in alkaline soils in arid or semi-arid areas of NW, China. Therefore, the novel isolate of B. megaterium with the highest adsorption capacity is a new promising biosorbent for the lead removal in alkaline water and soil.     

Decolorization of synthetic melanoidins-containing wastewater by a bacterial consortium

The presence of melanoidins in molasses wastewater leads to water pollution both due to its dark brown color and its COD contents. In this study, a bacterial consortium isolated from waterfall sediment was tested for its decolorization. The identification of culturable bacteria by 16S rDNA based approach showed that the consortium composed of Klebsiella oxytoca, Serratia mercescens, Citrobacter sp. and unknown bacterium. In the context of academic study, prevention on the difficulties of providing effluent as well as its variations in compositions, several synthetic media prepared with respect to color and COD contents based on analysis of molasses wastewater, i.e., Viandox sauce (13.5% v/v), caramel (30% w/v), beet molasses wastewater (41.5% v/v) and sugarcane molasses wastewater (20% v/v) were used for decolorization using consortium with color removal 9.5, 1.13, 8.02 and 17.5%, respectively, within 2 days. However, Viandox sauce was retained for further study. The effect of initial pH and Viandox concentration on decolorization and growth of bacterial consortium were further determined. The highest decolorization of 18.3% was achieved at pH 4 after 2 day of incubation. Experiments on fresh or used medium and used or fresh bacterial cells, led to conclusion that the limitation of decolorization was due to nutritional deficiency. The effect of aeration on decolorization was also carried out in 2 L laboratory-scale suspended cell bioreactor. The maximum decolorization was 19.3% with aeration at KLa = 2.5836 h⁻¹ (0.1 vvm).     

Impacts of process parameters optimization on the performance of the annular single chamber microbial fuel cell in wastewater treatment

Energy harvest from optimized annular single chamber microbial fuel cell (ASCMFC) with novel configuration, which treats chocolate industry wastewater, was investigated. In this study, optimization of operational parameters of the ASCMFC in terms of efficiency water‐soluble organic matter reduction and capability of electricity generation was evaluated. During the experiment, effluent from the anode compartment was examined through current and power density curves for variation in temperature and pH, chemical oxygen demand (COD), and turbidity removal, and substrate concentration. The performance analyzed at different temperature ranges such as 25, 30, 35, and 40°C, which showed 88% increase by uprising temperature from 25 to 35°C. The ASCMFC was used to produce electricity by adjusting pH between 5 and 9 at resistance of 100 Ω. Under the condition of pH 7 power density (16.75 W/m³) was highest, which means natural pH is preferred to maximize microbial activities. Wastewater concentration with COD of 700 and 1400 mg/L were investigated to determine its affection on current production. Reduction of current density was observed due to decrease in wastewater concentration. Significant reduction in COD and turbidity of effluent were 91 and 78%, respectively. The coulombic efficiency of 45.1% was achieved.     

Biological treatment of a pulp and paper industry effluent by Fomes lividus and Trametes versicolor

White rot fungi Fomes lividus and Trametes versicolor, isolated from the Western Ghats region of Tamil Nadu, India, were used to treat pulp and paper industry effluents on a laboratory scale and in a pilot scale. On the laboratory scale a maximum decolourization of 63.9% was achieved by T. versicolor on the fourth day. Inorganic chloride at a concentration of 765 mg/l, which corresponded to 227% of that in the untreated effluent, was liberated by F. lividus on the 10th day. The chemical oxygen demand (COD) was also reduced to 1984 mg/l (59.3%) by each of the two fungi. On the pilot scale, a maximum decolourization of 68% was obtained with the 6-day incubation by T. versicolor, inorganic chloride 475 mg/l (103%) was liberated on the seventh day by T. versicolor, and the COD was reduced to 1984 mg/l corresponding to 59.32% by F. lividus. These results suggested that F. lividus seems to be another candidate efficient for dechlorination of wastewater.     

Use of native aquatic macrophytes in the reduction of organic matter from dairy effluents

Considering the diversity and the unexplored potential of regional aquatic flora, this study aimed to identify and analyze the potential of native aquatic macrophytes to reduce the organic matter of dairy wastewater (DW) using experimental constructed wetlands. The dairy wastewater (DW) had an average chemical oxygen demand (COD) of 7414.63 mg/L and then was diluted to 3133.16 mg/L (D1) and to 2506.53 mg/L (D2). Total solids, COD, temperature, and pH analyses were performed, and the biochemical oxygen demand (BOD) was estimated from the COD values. The best performance in the reduction of the organic matter was observed for Polygonum sp. (87.5% COD and 79.6% BOD) and Eichhornia paniculata (90% COD and 83.7% BOD) at dilution D1, on the 8th day of the experiment. However, the highest total solids removal was observed for Polygonum sp. (32.2%), on the 4th day, at dilution D2. The total solid (TS) concentration has also increased starting from the 8th day of the experiment was observed which may have been due to the development of mosquito larvae and their mechanical removal by sieving, thus changing the steady state of the experimental systems. The macrophytes Polygonum sp. and E. paniculata were considered suitable for the reduction of organic matter of DW using constructed wetlands.     

Use of <Emphasis Type="Italic">Pichia fermentans</Emphasis> and <Emphasis Type="Italic">Candida</Emphasis> sp. strains for the biological treatment of stored olive mill wastewater

Of 105 isolates screened for growth on plates containing olive mill wastewater (OMW), five were selected and identified as Pichia fermentans (Y1, Y4) and Candida sp. (Y2, Y11, and Y18). On the basis of their ability to use phenol at 716 mg l⁻¹, strains Y2 (15% reduction) and Y4 (18% reduction) were then used to detoxify stored OMW under various operational conditions. Yeast treatment of OMW increased the pH and, in the best conditions (aeration and no glucose addition), the COD decreased (47%) and phytotoxicity was also decreased (56%) probably due to the changes in the composition of phenolic compounds.     

Sporulation and regeneration efficiency of phosphobacteria (<Emphasis Type="Italic">Bacillus megaterium</Emphasis> var <Emphasis Type="Italic">phosphaticum</Emphasis>)

Sporulation in Bacillus megaterium var phosphaticum (PB — 1) was induced using modified nutrient media. This modified medium induced sporulation within 36 h. After spore induction the spores were kept under refrigerated (5°C) and room temperature (32°C) for five months and survival of spores was studied at 15 days intervals by plating them in nutrient agar medium. It was observed that there was not much variation in the storage temperature (5°C & 32°C). The spore cells of Bacillus megaterium var phosphaticum (PB — 1) were observed up to five months of storage under refrigerated (5°C) and room temperature (32°C). Regeneration of spore cells into vegetative cells was studied in tap water, rice gruel, nutrient broth, sterile lignite and sterile water at different concentrations of spore inoculum. The multiplication of sporulated Bacillus megaterium var phosphaticum culture was fast and reached its maximum (29.5 × 10⁸ cfu ml⁻¹) in nutrient broth containing 5 per cent inoculum level.     

Identification of a denitrifying bacterium and verification of its anaerobic ammonium oxidation ability

A strain D₃ of denitrifying bacterium was isolated from an anammox reactor, and identified as Pseudomonas mendocina based on the morphological and physiological assay, Vitek test, Biolog test, (G+C) mol% content, and 16S rDNA phylogenetic analysis. As a typical denitrifying bacterium, strain D₃ achieved the maximal nitrate reduction rate of 26.2 mg/(L·d) at the nitrate concentration of 88.5 mg N/L. The optimal pH and growth temperature were 7.84 and 34.9°C, respectively. Strain D₃ was able to oxidize ammonia under anaerobic condition. The maximum nitrate and ammonium utilization rates were 6.37 mg/(L·d) and 3.34 mg/(L·d), respectively, and the consumption ratio of ammonia to nitrate was 1:1.91. Electron microscopic observation revealed peculiar cell in clusions in strain D₃. Because of its relation to anammox activity, strain D₃ was presumed to be anammoxosome. The present investigation proved that denitrifying bacteria have the anammox ability, and the results have engorged the range of anammox populations.     

Bioleaching of a Complex Co-Ni-Cu Sulfide Flotation Concentrate by Bacillus megaterium QM B1551 at Neutral pH

Bioleaching of sulfide minerals at neutral pH has been rarely reported. In this study, a bacterium, Bacillus megaterium QM B1551, was isolated from Jinchuan sulfide tailings and used to leach a complex sulfide flotation concentrate for the extraction of Co²⁺, Ni²⁺ and Cu²⁺ at near neutral pH. A total of 38.2% Co, 44.7% Ni and 3.6% Cu were extracted from the sulfide concentrate in 5 days with an initial pH of 6. An enhanced Co²⁺, Ni²⁺ and Cu²⁺ extraction extent was achieved by first bioleaching the concentrate with Bacillus megaterium QM B1551 at 35°C and then followed by chemical leaching with 4 M sulfuric acid at 90°C. As a result, a total of 60.7% Co²⁺, 76.3% Ni²⁺ and 39.8% Cu²⁺ were extracted. On an industrial scale, the profits from the metal recovery by such a combined leaching procedure are optimum if considering the cost-benefit ratio.     

β-Amylase fromBacillus megaterium

Bacillus megaterium strain B₆ producing extracellular β-amylase was isolated and grown in a medium supplemented with waste potato starch. It showed highest enzyme synthesis in the early stationary phase. The partially purified β-amylase had a temperature optimum at 60°C and a pH optimum at 6.9 and was not affected by Schardinger dextrins. These properties would allow its application in sugar industry.     

Identification of a denitrifying bacterium and verification of its anaerobic ammonium oxidation ability

A strain D3 of denitrifying bacterium was isolated from an anammox reactor,and identi-fied as Pseudomonas mendocina based on the morphological and physiological assay,Vitek test,Biolog test,(G C) mol% content,and 16S rDNA phylogenetic analysis.As a typical denitrifying bac-terium,strain D3 achieved the maximal nitrate reduction rate of 26.2 mg/(L·d) at the nitrate concen-tration of 88.5 mg N/L.The optimal pH and growth temperature were 7.84 and 34.9℃,respectively.Strain D3 was able to oxidize ammonia under anaerobic condition.The maximum nitrate and ammo-nium utilization rates were 6.37 mg/(L·d) and 3.34 mg/(L·d) ,respectively,and the consumption ratio of ammonia to nitrate was 1:1.91.Electron microscopic observation revealed peculiar cell inclusions in strain D3.Because of its relation to anammox activity,strain D3 was presumed to be anammoxosome.The present investigation proved that denitrifying bacteria have the anammox ability,and the results have engorged the range of anammox populations.     

Purification, characterization and application of a novel extracellular agarase from a marine Bacillus megaterium

A marine, gram positive, aerobic, spore forming, and non flagellated bacterium which degrades low melting point (LMP) — agarose was isolated from the west coast of India and identified as Bacillus megaterium based on its morphological, biochemical, and molecular characterization. This bacterium produced clear haloes or zone of clearance on agar containing plates which was a clear indication of its agarolytic property. The extracellular agarase thus obtained was purified 8.8 and 78 fold from the culture supernatant by ammonium sulfate precipitation and gel filtration, respectively. Molecular mass by gel filtration and SDS-PAGE gave values of 15 and 12 kDa, respectively. The optimum temperature and pH for maximum agarase activity were 40°C and 6.6. The activity of agarase was drastically reduced by addition of metal ions in the assay system. This agarase, gave a K _m and V _max value of 4 mg/mL and 2.75 μmol/min/mg. The isolation of protoplast from agarophyte like Gelidiella acerosa using indegenous agarase is reported for the first time.     

Phosphorus removal in pilot plant using biofilm filter process from farm wastewater

Various environmental conditions affecting total phosphorus removal from farm wastewater in a biofilm filter, process were investigated using loess balls andChromobacterium LEE-38 at a pilot plant. WhenChromobacterium LEE-38 was used, the removal efficiency of total phosphorous was approximately 10- or 5-fold higher than that ofAcinetobacter CHA-2-14 orAcinetobacter CHA-4-5, respectively. When a loess ball of 11–14 mm manufactured at a 960°C calcining temperature was used, the removal efficiency of total phosphorous was 90.0%. When 70% of the volume fraction was used, the maximum efficiency of total phosphorus removal was 93.1%. Notably, when the initial pH was in the range of 6.0 to 8.0, the maximum removal efficiency of total phosphorus was obtained after 30 days. When the operating temperature was in the range of 30 to 55°C, the maximum removal efficiencies of total phosphorus, 95.6 to 94.6%, were obtained. On the other hand, at operating temperatures below 20°C or above 40°C, the removal efficiency of total phosphorous decreased. Among the various processes, biofilm filter process A gave the highest removal efficiency of 96.4%. Pilot tests of total phosphorus removal using farm wastewater from the biofilm filter process A were carried out for 60 days under optimal condition. WhenAcinetobacters sp. Lee-11 was used, the average removal efficiency in thep-adsorption area was only 32.5%, and the removal efficiencies of chemical oxygen demand (COD) and biological oxygen demand (BOD) were 56.7 and 62.5%, respectively. On the other hand, whenChromobacterium LEE-38 was used, the average removal efficiency was 95.1%, and the removal efficiencies of COD and BOD were 91.3 and 93.2%, respectively. The first two authors contributed equally to this work.     

Role of casein on induction and enhancement of production of a bacterial milk clotting protease from an indigenously isolated Bacillus subtilis

Aims: To isolate and enhance the yield of a bacterial milk clotting protease (MCP) through process optimization and scale up. Materials and Results: Bacillus subtilis was isolated as MCP producer with good milk clotting activity (MCA) per proteolytic activity (PA) index. The enzyme production was inducible with casein and enhanced with fructose and ammonium nitrate resulting in 571·43 U ml^?1 of enzyme. Conclusions: Medium containing 4% fructose, 0·75% casein, 0·3% NH₄NO₃ and 10 mmol l^–1 CaCl₂, pH 6·0, inoculated with 4% (v/v) inoculum, incubated at 37°C, 200 rev min^?1 for 72 h gave maximum production. A 6·67‐fold increase in MCP yield with very high MCA per PA index was observed after final optimization indicating similarity to rennets. Significance and Impact of the Study: Mostly fungal MCPs have been reported. The MCA and MCA per PA index of this bacterium is comparable to that of many fungal reports and better than quite a few bacterial MCPs. Thus, this enzyme by B. subtilis has good probability of successful use in cheese production.     

Optimization, production, and partial characterization of an alkalophilic amylase produced by sponge associated marine bacterium Halobacterium salinarum MMD047

An endosymbiont Halobacterium salinarum MMD047, which could produce high yields of amylase, was isolated from marine sponge Fasciospongia cavernosa, collected from the peninsular coast of India. Maximum production of enzyme was obtained in minimal medium supplemented with 1% sucrose. The enzyme was found to be produced constitutively even in the absence of starch. The optimum temperature and pH for the enzyme production was 40°C and 8.0, respectively. The enzyme exhibited maximum activity in pH range of 6∼10 with an optimum pH of 9.0. The enzyme was stable at 40°C and the enzyme activity decreased dramatically above 50°C. Based on the present findings, the enzyme was characterized as relatively heat sensitive and alkalophilic amylase which can be developed for extensive industrial applications.     

Low pH (6, 5 and 4) sulfate reduction during the acidification of sucrose under thermophilic (55 °C) conditions

The effect of a low pH (6, 5 and 4) and different COD/SO₄²⁻ ratios (9 and 3.5) on thermophilic (55 °C) sulfate reduction and acidification of sucrose was investigated using three upflow anaerobic sludge bed reactors fed with sucrose at an organic loading rate of 3.5 gCOD (l_reactor d)⁻¹. The three reactors showed nearly 100% acidification of sucrose for all pH values and COD/SO₄²⁻ ratios investigated. Sulfate reduction was complete at pH 6 and a COD/SO₄²⁻ ratio of 9. At pH 5, sulfate reduction efficiencies were 80–95% for both COD/SO₄²⁻ ratios (9 and 3.5). At pH 4, sulfate reduction efficiencies further dropped to 55–65% at a COD/SO₄²⁻ ratio of 9 and 30–40% at a COD/SO₄²⁻ ratio of 3.5. The pH decrease from 6 to 5 or 4 caused a shift in the acidification products from mainly acetate to butyrate, as well as a higher production of ethanol, especially at pH 4. At pH 4, propionate and methane were not formed and hydrogen concentrations in the biogas reached 50%, equivalent to a hydrogen yield of 1.3 mol H₂ (mol glucose)⁻¹. This study shows that sulfate reduction is possible in the acidification phase of anaerobic wastewater treatment at pH values as low as 6 till 4 and that the pH strongly affects both the acidification pathways and the sulfate reduction efficiencies.