Indigenous Microbial Consortium Research Articles

Potential of dissimilatory nitrate reduction pathways in polycyclic aromatic hydrocarbon degradation

This study investigates the potential of an indigenous estuarine microbial consortium to degrade two polycyclic aromatic hydrocarbons (PAHs), naphthalene and fluoranthene, under nitrate-reducing conditions. Two physicochemically diverse sediment samples from the Lima Estuary (Portugal) were spiked individually with 25 mg L−1 of each PAH in laboratory designed microcosms. Sediments without PAHs and autoclaved sediments spiked with PAHs were run in parallel. Destructive sampling at the beginning and after 3, 6, 12, 30 and 63 weeks incubation was performed. Naphthalene and fluoranthene levels decreased over time with distinct degradation dynamics varying with sediment type. Next-generation sequencing (NGS) of 16 S rRNA gene amplicons revealed that the sediment type and incubation time were the main drivers influencing the microbial community structure rather than the impact of PAH amendments. Predicted microbial functional analyses revealed clear shifts and interrelationships between genes involved in anaerobic and aerobic degradation of PAHs and in the dissimilatory nitrate-reducing pathways (denitrification and dissimilatory nitrate reduction to ammonium - DNRA). These findings reinforced by clear biogeochemical denitrification signals (NO3− consumption, and NH4+ increased during the incubation period), suggest that naphthalene and fluoranthene degradation may be coupled with denitrification and DNRA metabolism. The results of this study contribute to the understanding of the dissimilatory nitrate-reducing pathways and help uncover their involvement in degradation of PAHs, which will be crucial for directing remediation strategies of PAH-contaminated anoxic sediments.

Bioremediation of Petroleum Hydrocarbon Contaminated Soil: Optimization Strategy Using Taguchi Design of Experimental (DOE) Methodology

One of the major environmental challenges of the oil industry is the permeation of petroleum and related products that cause intense soil pollution. Hydrocarbon oil polluted soils cannot be used for agricultural target because of hydrocarbon toxiscity. Bioremediation is a biotechnological technique which permits the treatment of oil polluted soil. In this work, design of experimental (DOE) method using Taguchi orthogonal array (OA) was performed to assess the influence of five factors (i.e., different levels of molasses concentration, whey concentration, urea concentration, use of indigenous microbial consortium as inoculum and Tween 80 as surfactant) on the removal efficiency of total petroleum hydrocarbon (TPH) from Iranian crude oil contaminated soils. The percent contribution of each factor was also calculated. The results showed that use of indigenous microbial and whey concentration made the highest (25.3%) and the lowest (13.7%) contribution, respectively. The total hydrocarbon content was reduced to 72.63% of its initial value at the optimal conditions at the end of the experiments. The study demonstrates that Taguchi method is appropriate to optimize the experiments for the efficiency of TPH removal (R 2 = 0.974).

Efficiency of consortium for in-situ bioremediation and CO2 evolution method of refines petroleum oil in microcosms study

An in-situ bioremediation study was conducted in a laboratory by using mixed microbial consortium. An indigenous microbial consortium was developed by assemble of two Pseudomonas spp. and two Aspergillus spp. which were isolated from various oil contaminated sites of India. The laboratory feasibility study was conducted in a 225 m 2 block. Six treatment options-Oil alone, Oil+Best remediater, Oil+Bacterial consortium, Oil+Fungal consortium, Oil+Mixed microbial consortium, Oil+Indigenous microflora. Out of five treatments, the mixed microbial consortium (Block 5) degraded 55.12% refine petroleum oil compare to degradation of bacterial (Block 3) and Fungal consortium (Block 4) (i.e, degradation rate were 19.88% and 18.07% correspondingly) after the end of treatment (60 days). Bioremediation ability of this consortium was confirmed by CO 2 evolution method. The result showed that 136.36 % CO 2 release after 12 days incubation. 16r DNA sequencing showed that two bacterial species were Pseudomonas aeruginosa and morph taxonomical examination of fungus were Aspergillus terrus (At) and Aspergillus flavus (Af).The ability of degradation of synthetic mixture of refine petroleum oils makes the consortium potentially useful for bioremediation and microbial enhanced oil recovery.

Ability of Indigenous Microbial Consortium in the Process of Ammonia Oxidation of Livestock Waste

Ability of Indigenous Microbial Consortium in the Process of Ammonia Oxidation of Livestock Waste

Quality and Microbiological Evaluation of Phosphocompost Prepared from the Hydrothermal Pre-treated Pine Needles using Indigenous Microbial Consortium

Quality and Microbiological Evaluation of Phosphocompost Prepared from the Hydrothermal Pre-treated Pine Needles using Indigenous Microbial Consortium

Effects of Indigenous Microbial Consortium in Crude Oil Degradation: A Microcosm Experiment

Microbial biodegradation is a bio-treatment method for attenuating crude oil contaminated soils. The purpose of present study was to select and recognize indigenous bacteria and to investigate the capability of the bacterial consortium to remove crude oil in a planned microcosm. Capability of the bacterial growth in agar plates containing crude oil was used to select the isolated crude oil eating bacteria. Soil samples were collected from different contaminated sites nearby the exploitation unit of Karoon-3, Ahvaz, Iran, and incubated by individual and mixed bacterial consortium at 30°C for a 90 day period time. Two indigenous isolates were selected and designated as S10 and S31. According to 16S rRNA sequencing, S10 was identified as a new strain of Bacillus subtilis and S31 as Bacillus licheniformis. The results showed that the highest cell mass was in 10th day of incubation time with about 1.7×107 CFU/g, for bacterial consortium. The highest total petroleum hydrocarbons degradation (C%) in the first 10 days of incubation time raised up to about 77.5% for bacterial consortium, and also residual C:N ratio deleted about 73%. Results emphasized that the planned microcosm in laboratory with controlled conditions using consortia of the selected indigenous bacteria can make the crude oil biodegradation close to the optimal conditions and have a good potential for application in bioremediation of crude oil contaminated soils, as compared to the individual strains. This is the first report on planned microcosm and microbial consortium of Karoon soil contaminated with crude oil.

Identification of an Indigenous Atrazine Herbicide Tolerant Microbial Consortium in Beans (Phaseolus vulgaris L.) as a Potential Soil Bioremediator

The present article reports the isolation and identification of atrazine-tolerant strains of indigenous microorganisms recovered from three representative agricultural sites representing agronomic characteristics of the Tulancingo Valley, Central part of Mexico (disturbed and undisturbed). Biochemical and morphological tests were performed for microorganism’s identification and the minimum inhibitory concentration assay was followed to assess atrazine tolerance. Results showed the microorganism populations varied from 10-5 to 10-6 UFC g-1 of soil for bacteria and 104 - 105 conidia g-1 of soil for fungi. The bacterial genera isolated and identified were: Agrobacterium sp., Bacillus sp., Erwinia sp., Micrococcus sp., Pediococcus sp., Rhizobium sp., Serrantia sp. and Sphingomonas sp. Identified fungal genera were: Alternaria sp., Aspergillus sp., Mucor sp., Cladosporium sp., Penicillium sp., Fusarium sp. and Trichoderma sp. Tests for herbicide tolerance indicate the isolated microorganisms do not show inhibitory growth at 500 to 2,500 ppm of atrazine concentrations under laboratory conditions. The strains of the fungal genera and Rhizobium sp. showed greater tolerance rates to atrazine, based on their growth without inhibition in the presence of 5,000 to 10,000 ppm of the agrochemical. Results suggest the isolated microorganisms may be useful as a viable inoculum for bioremediation purposes in agricultural atrazine-contaminated soils.

Influence of nitrate and sulfate reduction in the bioelectrochemically assisted dechlorination of cis-DCE

This paper investigated the reductive dechlorination (RD) of cis-dichloroethylene (cis-DCE) (average influent 14.2±0.7μM) by a bioelectrochemical system (BES), in the presence of real contaminated groundwater containing high levels of nitrate and sulfate. The BES enhanced both the RD and competing reactions, such as nitrate and sulfate reductions, which occurred with neither an external organic carbon source nor any inoculum other than the indigenous microbial consortia in the real groundwater. In preliminary batch tests, RD and full nitrate removal occurred after a short lag phase, whereas sulfate reduction occurred slowly and alongside the RD. Under continuous flow conditions (hydraulic retention time, HRT, 1.4d), the competition of different electron acceptors was strongly affected by the cathodic potential in the range −550 to −750mV vs. standard hydrogen electrode (SHE). Nitrate reduction was driven to completion at all tested cathodic potentials, whereas sulfate reduction and the RD rate increased as the cathodic potential became more negative. At −750mV vs. SHE, strong methanogenesis was also observed and became the most important sink of electrons. The overall coulombic efficiency decreased while the potential became more negative. The RD contribution was always less than 1%. Hence, greater energy consumption was required to obtain higher RD rate and better conversion. Anodic oxidation was only observed at −750mV vs. SHE where almost 39% of residual vinyl chloride (VC) was oxidized and the sulfate was formed back from sulfide (further contributing to electric waste).

Effects of nitrate injection on microbial enhanced oil recovery and oilfield reservoir souring.

Column experiments were utilized to investigate the effects of nitrate injection on sulfate-reducing bacteria (SRB) inhibition and microbial enhanced oil recovery (MEOR). An indigenous microbial consortium collected from the produced water of a Brazilian offshore field was used as inoculum. The presence of 150 mg/L volatile fatty acids (VFA´s) in the injection water contributed to a high biological electron acceptors demand and the establishment of anaerobic sulfate-reducing conditions. Continuous injection of nitrate (up to 25 mg/L) for 90 days did not inhibit souring. Contrariwise, in nitrogen-limiting conditions, the addition of nitrate stimulated the proliferation of δ-Proteobacteria (including SRB) and the associated sulfide concentration. Denitrification-specific nirK or nirS genes were not detected. A sharp decrease in water interfacial tension (from 20.8 to 14.5 mN/m) observed concomitantly with nitrate consumption and increased oil recovery (4.3 % v/v) demonstrated the benefits of nitrate injection on MEOR. Overall, the results support the notion that the addition of nitrate, at this particular oil reservoir, can benefit MEOR by stimulating the proliferation of fortuitous biosurfactant-producing bacteria. Higher nitrate concentrations exceeding the stoichiometric volatile fatty acid (VFA) biodegradation demands and/or the use of alternative biogenic souring control strategies may be necessary to warrant effective SRB inhibition down gradient from the injection wells.

Large Scale Bioremediation of Petroleum Hydrocarbon Contaminated Waste at Various Installations of ONGC. India: Case Studies

In situ and ex situ bioremediation of oil contaminated effluent pits, sludge pits, oil spilled land and tank bottom, and effluent treatment plant (ETP) oily sludge was carried out at Ankleshwar, Mehsana, Assam and Cauvery Asset of Oil and Natural Gas Corporation Limited (ONGC), India. The types of contaminant were heavy paraffinic, asphaltic and light crude oil and emulsified oily sludge /contaminated soil. An indigenous microbial consortium was developed by assembling four species of bacteria, isolated from various oil contaminated sites of India, which could biodegrade different fractions of total petroleum hydrocarbon (TPH) of the oily waste to environment friendly end products. The said consortium was on a large scale field applied to the above oil installations and it successfully bioremediated 30,706 tonnes of different types of oily waste. In 65 case studies of different batch size of in situ and ex situ bioremediation processes, the initial TPH content varying from 69.20 to 662.70 g/kg of oily waste has been biodegraded to 5.30 – 16.90 g/kg of oily waste in a range of 2 to 33 months. Biodegradation rate varied in the range of 0.22 – 1.10 Kg TPH /day/m2 area due to the climatic condition of the treatment zone and the type of waste treated. The bioremediated soil was non-toxic and natural vegetation was found to be grown on the same ground. Successful eco-restoration of one large effluent pit of 26,000 m2 area was carried out by cultivation of local fish species after completion of bioremediation. Bioremediation technology has helped ONGC with the management of their hazardous oily wastes in an environment friendly manner. DOI: http://dx.doi.org/10.5755/j01.erem.68.2.5632

Pesticide relevance and their microbial degradation: a-state-of-art

The extensive use of pesticide causes imbalance in properties of soil, water and air environments due to having problem of natural degradation. Such chemicals create diverse environmental problem via biomagnifications. Currently, microbial degradation is one of the important techniques for amputation and degradation of pesticide from agricultural soils. Some studies have reported that the genetically modified microorganism has ability to degrade specific pesticide but problem is that they cannot introduce in the field because they cause some other environmental problems. Only combined microbial consortia of indigenous and naturally occurring microbes isolated from particular contaminated environment have ability to degrade pesticides at faster rate. The bioaugumentation processes like addition of necessary nutrients or organic matter are required to speed up the rate of degradation of a contaminant by the indigenous microbes. The use of indigenous microbial strains having plant growth activities is ecologically superior over the chemical methods. In this review, we have attempted to discuss the recent challenge of pesticide problem in soil environment and their biodegradation with the help of effective indigenous pesticides degrading microorganisms. Further, we highlighted and explored the molecular mechanism for the pesticide degradation in soil with effective indigenous microbial consortium. This review suggests that the use of pesticide degrading microbial consortia which is an eco-friendly technology may be suitable for the sustainable agriculture production.

Analysis of Bio-gas Production from Cow Dung by Indigenous Microbial Consortia

The study analyzed quantitatively and qualitatively the biogas produced from cowdung by indigenous microbial consortia. Four 20L bioreactors were used for the study. The bioreactors were constructed to imitate the fixed batch prototype. The study lasted for six months and it was carried out at the Microbiology Laboratory of Anambra State University, Anambra State, Nigeria. Slurry was prepared in bioreactors. The substrates in the bioreactors were water and manure (WM), rumen fluid and manure (RM-1 and RM-2), medium and manure (MM). The pH, the total solids (TS), volatile solids (VS) and total volatile fatty acid (VFA) characteristics of the substrate before and after digestion were determined using standard method. Quantification and qualitative analysis of biogas production was by liquid displacement and gas chromatography methods respectively. The microbial analysis of the substrate was carried out using spread plate method. The results of the TS, VS and VFA were 400 mg/l, 92mg/l and 16.7 mg/l respectively in the predigested samples and 92 mg/l, 17.4mg/l and 28.3mg/l respectively in the post digested samples. The quantity of biogas produced at fourth month was 60 ml, 128ml and 220ml from WM, MM and RM-1 respectively. The qualitative analysis showed that the prominent biogas produced was methane. The cultural morphology revealed Gram positive rods with creamy irregular edges. The average heterotrophic counts at the end of each month for a period of four months were 18.5x10cfu/ml, 21.1 x10cfu/ml and 26.7x10cfu/ml for WM, MM and RM respectively. The results of the research concluded that high quantity of biogas can be produced using cow dung. Approaches and technology for more efficient biogas producing consortia are proposed. Original Research Article British Biotechnology Journal, 4(5): 630-639, 2014 631

Airlift bioreactors for hydrocarbon water pollution remediation in a tourism development pole

Underwater sinkholes in Quintana Roo, Mexican Caribbean, are affected by the run-off of nearby highways promoting the presence of aromatic and polyaromatic hydrocarbons. Phenanthrene, naphthalene, and benzene derivatives were found as the most common hydrocarbon contaminants present in underwater sinkholes located in Cancún and Playa del Carmen, two well-developed tourism poles. This confirms the impact of urban activities related with the increased transportation of locals and tourists around these areas, in comparison with Holbox which is a recent touristic development where transportation of tourists and locals is scarce; there is no hydrocarbon detectable pollution. Additionally, the development of a new water cleaning approach by airlift bioreactors, using indigenous microbial consortium, shows promising results and may be used in future ex-situ remediation technique in tourism poles.

English

In contaminated soils, the efficiency of natural attenuation or engineered bioremediation largely depends on the biodegradation capacities of the total microflorae. In the present study, the biodegradation capacities of various bacteria towards petroleum-hydrocarbons were determined under laboratory conditions. The purpose of the study was to isolate and characterize petroleum-degrading bacteria from contaminated soil obtained from a refinery in Arzew, Algeria. A collection of 15 bacterial isolates were obtained by enrichment cultivation from oil-contaminated soil and an indigenous microbial consortium was developed by assembling four species of bacteria which could degrade different fractions of the petroleum hydrocarbons. 16S rRNA gene analysis was used to identify members of the consortium and oil biodegra- dability was analyzed by Thin Layer Chromatography (TLC) with Flame Ionization Detection (FID) that performs quantitative compositional analysis of oil samples. The Iatroscan TLC/FID system measured the relative percentages of the four major fractions of petroleum i.e. satu- rates aromatics, resins and asphaltenes. Results indicated that the constructed consortium which comprised the genera Pseudomonas, She- wanella, Enterobacter and Serratia used the hydrocarbons as sole sources of carbon where biodegradation was defined by an initial rapid decrease in the saturate and aromatic fractions from 56.44% and 34.72% to 51.77% and 27.77% respectively, coinciding with an increase in the asphaltene fraction. The resin content remained relatively constant throughout the project. Alkane hydroxylase genes (alkB) were positively amplified in the Pseudomonas isolate by the polymerase chain reaction (PCR) method using degenerate primers. This functional gene was used as a marker to assess the catabolic potential of the bacteria for alkane pollutant biodegra- dation. The selected bacterial consortium looks promising for its application in bioremediation technologies.

Growth kinetics of an indigenous mixed microbial consortium during methylene chloride degradation in a batch reactor

Biodegradation of methylene chloride by a mixed microbial culture, isolated from a common sewage treatment plant, was investigated in a batch system. Batch experiments were performed at room temperature (27 °C) and pH value of 7. The methylene chloride concentration in growth media varied from 25 mgl−1 to 250mgl−1. A maximum observed degradation was 1 mgl−1h−1 at 100 mgl−1 of methylene chloride. The culture followed substrate inhibition kinetics and specific growth rate were fitted to different substrate inhibition models (Haldane, Aiba and Edwards models) by MATLAB 7.1@. Among all models, Haldane was found to better fit with root mean square of 0.947. The biokinetic constants estimated using these models show good potential of the mixed microbial culture in methylene Chloride degradation. Escherichia coli and Staphylococcus aureus are predominant microbes present in the mixed culture.

Biosorption of heavy metals in a photo-rotating biological contactor—a batch process study

Metal removal potential of indigenous mining microorganisms from acid mine drainage (AMD) has been well recognised in situ at mine sites. However, their removal capacity requires to be investigated for AMD treatment. In the reported study, the capacity of an indigenous AMD microbial consortium dominated with Klebsormidium sp., immobilised in a photo-rotating biological contactor (PRBC), was investigated for removing various elements from a multi-ion synthetic AMD. The synthetic AMD was composed of major (Cu, Mn, Mg, Zn, Ca, Na, Ni) and trace elements (Fe, Al, Cr, Co, Se, Ag, Mo) at initial concentrations of 2 to 100 mg/L and 0.005 to 1 mg/L, respectively. The PRBC was operated for two 7-day batch periods under pH conditions of 3 and 5. The maximum removal was observed after 3 and 6 days at pH 3 and 5, respectively. Daily water analysis data demonstrated the ability of the algal-microbial biofilm to remove an overall average of 25-40 % of the major elements at pH 3 in the order of Na > Cu > Ca > Mg > Mn > Ni > Zn, whereas a higher removal (35-50 %) was observed at pH 5 in the order of Cu > Mn > Mg > Ca > Ni > Zn > Na. The removal efficiency of the system for trace elements varied extensively between 3 and 80 % at the both pH conditions. The batch data results demonstrated the ability for indigenous AMD algal-microbial biofilm for removing a variety of elements from AMD in a PRBC. The work presents the potential for further development and scale-up to use PBRC inoculated with AMD microorganisms at mine sites for first or secondary AMD treatment.

Co-metabolic degradation of tribenuron methyl, a sulfonylurea herbicide, by Pseudomonas sp. strain NyZ42

A co-metabolic degradation of tribenuron methyl bacterial strain NyZ42 was isolated from polluted agricultural soil and classified as genus Pseudomonas by its 16S rRNA gene sequencing. The degradation efficiency of tribenuron methyl was about 80% of the originally supplemented 200 mg l−1 tribenuron methyl in liquid minimal medium within four days, when either glucose or succinate was used as a supplemental carbon source. Three intermediates formed during the degradation of tribenuron methyl mediated by strain NyZ42 were captured by LC-MS, and two alternative pathways were proposed for the microbial mediated tribenuron methyl degradation, via either cleavage of the sulfonylurea bridge or saponification of alkyl-group. Furthermore, inoculation of strain NyZ42 enhanced the degradation of tribenuron methyl in the sterilized soil samples, although the biodegradation/co-metabolism ability of NyZ42 was not obvious in the nonsterilized soil samples when compared with the indigenous microbial consortium under current laboratory conditions.

Removal of Crude Oil by Microbial Consortium Isolated from Oil-Spilled Area in the Korean Western Coast

The feasibility of using an indigenous microbial consortium for the removal of crude oil from an oil-spilled coastal area was explored with the ultimate aim of applying for bioremediation. Initially, we obtained the microbial consortium TK-2 that catalyzed the dispersion as well as the degradation of crude oil in supplemented sea water. GC and GC-MS were used to evaluate the removal patterns of crude oil during the incubation. The effective removal of crude oil by TK-2 occurred, and above 95% of all aliphatic and aromatic compounds detected in this work was removed within 30 days of incubation. Two predominant crude oil-grown isolates derived from TK-2 revealed gram-negative, rod-shaped cells. Both BIOLOG system and 16S rRNA sequencing were conducted to identify the strains, which were assigned to Arthrobacter sp. HK-2 and Pseudoalteromonas sp. HK-3, and registered in GenBank as [FJ477042] and [FJ477041].

Bioremediation of Dyestuff Compounds using Indigenous Microorganism in a Bioreactor

Dyes are synthesized in a reactor, filtered, dried, blended with other color additives to produce the final product. The liquid effluent resulting from the manufacture of colored dye contains toxic organic residues and inorganic compounds. The present treatment technology involves: neutralization, flocculation, coagulation-settling, carbon adsorption, detoxification of organics by oxidation and biological treatments. In spite of the present treatment, the discharge effluent contains the color residual toxic compounds. In the present research study the dye effluent discharged at the waste disposal site has been characterized for physico-chemical and microbial characterization. The microorganisms adapted at contaminated sites have been isolated and cultured to use as a biomass for bioremediation of identified toxic organic compounds present in dye effluent. GC-MS has identified -octadecene; 5-Eicosene, (E)-; Di (2-ethylhexyl) phthalate; 1-nonadecene; Butyl-2-ethylhexyl phthalate; 5-Chloro-1,3-benzodioxole; 2-Oxazolidinone, 3, -dimethyl-5- phthalate; 1-Phenyl-5-methylnona-1, 3, 4-triene; and 2,2-Dimethyl-N-(3-methylpenyl) propanamide dye compounds in discharge effluents. The bioremediation of dye stuff- discharged effluent was carried out in a laboratory bioreactor using indigenous microbial consortium under controlled environmental conditions. The biodegradation studied at an interval of every hour for a day and thereafter each day for a period of 21 days, till the complete biodegradation takes place. Biodegradation has been studied using GC-MS; the intermediate compounds were found to be biodegraded into pthalic acid – a non toxic compound. The indigenous microbial consortium developed was found effective and efficient to biodegrade dye compounds into environmental friendly compounds.

Assessment of a process to degrade metal working fluids using Pseudomonas stutzeri CECT 930 and indigenous microbial consortia

The development of a novel biological process to treat metal working fluids (MWFs)-containing effluents at bioreactor scale was pursued in this work. The bacteria Pseudomonas stutzeri CECT 930 was investigated for the first time as an alternative agent for MWF degradation. An adequate medium design and mixing and aeration system, as well as an appropriate microorganism proved to be crucial for reaching high levels of degradation by P. stutzeri and by an indigenous consortium (about 70% and 50% of reduction in total petroleum hydrocarbon content in less than 2 wk, respectively). Additionally, as there is no information in literature trying to kinetically characterize an MWF-polluted effluent degradation process, all the experimental data were fitted to logistic and Luedeking and Piret models, that allowed to elucidate the growth-associated character of the biodegradation process.