BACKGROUND, AIMS AND SCOPE: Research and development has its own benefits and inconveniences. One of the inconveniences is the generation of enormous quantity of diverse toxic and hazardous wastes and its eventual contamination to soil and groundwater resources. Ethidium bromide (EtBr) is one of the commonly used substances in molecular biology experiments. It is highly mutagenic and moderately toxic substance used in DNA-staining during electrophoresis. Interest in phytoremediation as a method to solve chemical contamination has been growing rapidly in recent years. The technology has been utilized to clean up soil and groundwater from heavy metals and other toxic organic compounds in many countries like the United States, Russia, and most of European countries. Phytoremediation requires somewhat limited resources and very useful in treating wide variety of environmental contaminants. This study aimed to assess the potential of selected tropical plants as phytoremediators of EtBr. MATERIALS AND METHODS: This study used tomato (Solanum lycopersicum), mustard (Brassica alba), vetivergrass (Vetiveria zizanioedes), cogongrass (Imperata cylindrica), carabaograss (Paspalum conjugatum), and talahib (Saccharum spontaneum) to remove EtBr from laboratory wastes. The six tropical plants were planted in individual plastic bags containing soil and 10% EtBr-stained agarose gel. The plants were allowed to establish and grow in soil for 30 days. Ethidium bromide content of the test plants and the soil were analyzed before and after soil treatment. Ethidium bromide contents of the plants and soils were analyzed using an UV VIS spectrophotometer. RESULTS: Results showed a highly significant (p≤0.001) difference in the ability of the tropical plants to absorb EtBr from soils. Mustard registered the highest absorption of EtBr (1.4±0.12 μg kg⁻¹) followed by tomato and vetivergrass with average uptake of 1.0±0.23 and 0.7±0.17 μg kg⁻¹ EtBr, respectively. Cogongrass, talahib, and carabaograss had the least amount of EtBr absorbed (0.2±0.6 μg kg⁻¹). Ethidium bromide content of soil planted to mustard was reduced by 10.7%. This was followed by tomato with an average reduction of 8.1%. Only 5.6% reduction was obtained from soils planted to vetivergrass. Soils planted to cogongrass, talahib, and carabaograss had the least reduction of 1.52% from its initial EtBr content. DISCUSSION: In this study, mustard, tomato, and vetivergrass have shown their ability to absorb EtBr from contaminated soil keeping them from expanding their reach into the environment and preventing further contamination. Its downside, however, is that living creatures including humans, fish, and birds, must be prevented from eating the plants that utilized these substances. Nonetheless, it is still easier to isolate, cut down, and remove plants growing on the surface of the contaminated matrices, than to use strong acids and permanganates to chemically neutralize a dangerous process that can further contaminate the environment and pose additional risks to humans. Though this alternative method does not totally eliminate eventual environmental contamination, it is by far produces extremely insignificant amount of by-products compared with the existing processes and technologies. CONCLUSIONS: Mustard had the highest potential as phytoremediator of EtBr in soil. However, the absorption capabilities of the other test plants may also be considered in terms of period of maturity and productivity. RECOMMENDATIONS AND PERSPECTIVES: It is recommended that a more detailed and complete investigation of the phytoremediation properties of the different plants tested should be conducted in actual field experiments. Plants should be exposed until they reach maturity to establish their maximum response to the toxicity and mutagenecity of EtBr and their maximum absorbing capabilities. Different plant parts should be analyzed individually to determine the movement and translocation of EtBr from soil to the tissues of plants. Since this study has established that some plants can thrive and dwell in EtBr-treated soil, an increased amount of EtBr application should be explored in future studies. It is suggested therefore that a larger, more comprehensive exploration of phytoremediation application in the management of toxic and hazardous wastes emanating from biotechnology research activities should be considered especially on the use of vetivergrass, a very promising tropical perennial grass.

BACKGROUND, AIM AND SCOPE: The presence of heavy metals in wastewater is one of the main causes of water and soil pollution. The aim of the present study was to investigate the removal of Cd, Cu, Pb, Hg, Mn, Cr and Zn in urban effluent by a biological wastewater treatment, as well as to quantify the levels of As, Be, Cd, Cr, Cu, Fe, Hg, Mn, Ni, Pb, Sn, Tl, V and Zn in dewatering sludge from the Biological Wastewater Treatment Plant to Ribeirão Preto (RP-BWTP), Brazil. MATERIALS AND METHODS: Concentrations of Cd, Cr, Cu, Mn and Pb in wastewater and those of Ni in sludge were determined by atomic absorption spectrophotometry with graphite furnace atomization. Mercury concentrations in wastewater were measured by hydride generation atomic spectrophotometry, and Zn levels were determined by atomic absorption spectrophotometry using acetylene flame. In sludge, the levels of As, Be, Cd, Cr, Cu, Fe, Hg, Mn, Pb, Sn, Tl, V and Zn were determined by inductively coupled plasma-mass spectrometry. RESULTS: The percentages of removal efficiency (RE) were the following: Hg 61.5%, Cd 60.0%, Zn 44.9%, Cu 44.2%, PB 39.7%, Cr 16,5% and Mn 10.4%. In turn, the mean concentrations (mg/kg) of metals in dewatering sludge followed this increasing order: Tl (<0.03), Hg (0.31), Be (0.43), As (1.14), Cd (1.34), V (59.2), Pb (132.1), Sn (166.1), Cr (195.0), Mn (208.1), Ni (239.4), Cu (391.7), Zn (864.4) and Fe (20537). DISCUSSION: The relationship between metal levels in untreated wastewater, as well as the removal efficiency are in agreement with previous data from various investigators, It is important to note that metal removal efficiency is not only affected by metal ion species and concentration, but also by other conditions such as operating parameters, physical, chemical, and biological factors. CONCLUSIONS: Metal values recorded for treated wastewater and sludge were within the maximum permitted levels established by the Environmental Sanitation Company (CETESB), São Paulo, Brazil. RECOMMENDATIONS: There is an urgent need for the authorities who are responsible for legislation on sludge uses in agriculture of establishing safety levels for As, Be, Hg, Sn, Tl and V. PERSPECTIVES: According to the current metal levels, RP-BWTP sludge might be used for agriculture purposes. However, for an environmentally safe use of sewage sludge, further studies including systematic monitoring are recommended. Annual metal concentrations and predicted variations of those elements in the sludge should be monitored.

- DOI: http://dx.doi.org/10.1065/espr2006.04.299 Goal, Scope and Background This paper describes a statistical modelling approach, suggested as a policy tool in the Athens area for the assessment of the emissions reduction level required to meet the air quality standards for two criteria air pollutants, namely CO and NO2. Methods. More than ten years of hourly CO and NOx-NO2 concentration data measured by the monitoring network of the Hellenic Ministry for the Environment, Physical Planning and Public Works were analyzed and the original dataset has been reduced using a data evaluation procedure. Results and Discussion Seasonal pollutant concentration trends suggested that the reduction of CO and NOx concentrations observed in the beginning of the '90s is almost entirely attributed to the increase of the catalyst-equipped cars during this period. The numerical parameters of an empirical model relating EU standard exceedances with mean annual concentrations were defined and the model was validated using datasets from years that were not used for the estimation of these parameters. This model was used in conjunction with a roll-back equation as a policy tool for the assessment of the effect of different CO and NOx emissions reduction scenarios on air quality standard compliance for CO and NO2. Results predicted with this empirical modelling approach were assessed with monitored data averaged over a 3-year period, giving satisfactory results. Conclusion A methodology suggested for assessing the effects of different emissions reduction scenarios on air quality standard attainment was successfully applied for CO and NO2 in the Athens area. Recommendation and Perspective The proposed methodology can provide a useful tool for the evaluation of policies already in progress as well as the development of future policies for emissions reduction in urban areas with similar characteristics, aiming at air quality standard compliance on a timely manner. Such a methodology could be applied in other urban areas of Greece characterized by dense traffic, therefore assisting the development of national policies in relation to air pollutants for which standard exceedances occur.

Background, Aim and Scope Numerous herbicides and xenobiotic organic pollutants are detoxified in plants to glutathione conjugates. Following this enzyme catalyzed reaction, xenobiotic GS-conjugates are thought to be compartmentalized in the vacuole of plant cells. In the present study, evidence is presented for long range transport of these conjugates in plants, rather than storage in the vacuole. To our knowledge this is the first report about the unidirectional long range transport of xenobiotic conjugates in plants and the exudation of a glutathione conjugate from the root tips. This could mean that plants possess an excretion system for unwanted compounds which give them similar advantages as animals. Materials and Methods: Barley plants (Hordeum vulgare L. cv. Cherie) were grown in Petri dishes soaked with tap water in the greenhouse. - Fluorescence Microscopy. Monobromo- and Monochlorobimane, two model xenobiotics that are conjugated rapidly in plant cells with glutathione, hereby forming fluorescent metabolites, were used as markers for our experiments. Their transport in the root could be followed sensitively with very good temporal and spatial resolution. Roots of barley seedlings were cut under water and the end at which xenobiotics were applied was fixed in an aperture with a thin latex foil and transferred into a drop of water on a cover slide. The cover slide was fixed in a measuring chamber on the stage of an inverse fluorescence microscope (Zeiss Axiovert 100). - Spectrometric enzyme assay. Glutathione S-transferase (GST) activity was determined in the protein extracts following established methods. Aliquots of the enzyme extract were incubated with 1-chloro-2,4-dinitrobenzene (CDNB), or monochlorobimane. Controls lacking enzyme or GSH were measured. - Pitman chamber experiments. Ten days old barley plants or detached roots were inserted into special incubation chambers, either complete with tips or decapitated, as well as 10 days old barley plants without root tips. Compartment A was filled with a transport medium and GSH conjugate or L-cysteine conjugate. Compartments B and C contained sugar free media. Samples were taken from the root tip containing compartment C and the amount of conjugate transported was determined spectro-photometrically. Results: The transport in roots is unidirectional towards the root tips and leads to exsudation of the conjugates at rates between 20 and 200 nmol min-1. The microscopic studies have been complemented by transport studies in small root chambers and spectroscopic quantification of dinitrobenzene-conjugates. The latter experiments confirm the microscopic studies. Furthermore it was shown that glutathione conjugates are transported at higher rates than cysteine conjugates, despite of their higher molecular weights. This observation points to the existence of glutathione specific carriers and a specific role of glutathione in the root. Discussion: It can be assumed that long distance transport of glutathione conjugates within the plant proceeds like GSH or amino acid transport in both, phloem and xylem. The high velocity of this translocation of the GS-X is indicative of an active transport. For free glutathione, a rapid transport-system is essential because an accumulation of GSH in the root tip inhibits further uptake of sulfur. Taking into account that all described MRP transporters and also the GSH plasmalemma ATPases have side activities for glutathione derivatives and conjugates, co-transport of these xenobiotic metabolites seems credible. - On the other hand, when GS-B was applied to the root tips from the outside, no significant uptake was observed. Thus it can be concluded that only those conjugates can be transported in the xylem which are formed inside the root apex. Having left the root once, there seems to be no return into the root vessels, probably because of a lack of inward directed transporters. Conclusions: Plants seem to possess the capability to store glutathione conjugates in the vacuole, but under certain conditions, these metabolites might also undergo long range transport, predominantly into the plant root. The transport seems dependent on specific carriers and is unidirectional, this means that xenobiotic conjugates from the rhizosphere are not taken up again. The exudation of xenobiotic metabolites offers an opportunity to avoid the accumulation of such compounds in the plant. Recommendations and Perspectives: The role of glutathione and glutathione related metabolites in the rhizosphere has not been studied in any detail, and only scattered data are available on interactions between the plant root and rhizosphere bacteria that encounter such conjugates. The final fate of these compounds in the root zone has also not been addressed so far. It will be interesting to study effects of the exuded metabolites on the biology of rhizosphere bacteria and fungi.

BACKGROUND AND AIM: In non-eutrophicated freshwaters, humic substances (HS) pose chemical stresses on aquatic organisms and, hence, separating sensitive from less sensitive or even tolerant species. One of the stresses, identified so far, is the reduction of photosynthetic oxygen production and reduction in growth in freshwater macrophytes and algae. In a previous paper, it has been shown that even closely related coccal green algae responded differently upon identical stress by HS, which is consistent with the hypothesis above. Due to their much simpler cellular ultrastructure, cyanobacteria are supposed to be more sensitive to HS exposure than eukaryote should be. One coccal green algal species (Desmodesmus communis) and one cyanobacterium (Chroococcus minutus) were exposed to four natural organic matter (NOM) isolates. One NOM has been isolated from a brown water lake (Schwarzer See) in Brandenburg State; three were obtained from a comprehensive Scandinavian NOM research project and originated from Norway (Birkenes), Finland (Hietajärvi), and Sweden (Svartberget). METHODS: Cultures of D. communis and C. minutus were obtained from the Culture Collection of Algae, Göttingen, and maintained in a common medium. The cultures were non-axenic. The algae and cyanobacteria were exposed under identical conditions to environmentally realistic NOM concentrations. Cell numbers were counted microscopically in Neugebauer cuvettes in triplicates. To avoid limitation by nutrient depletion, the experiments were terminated after 14 to 15 days. Until culture day 12, no growth limitation has been observed in the controls. RESULTS: All NOM isolates modulated the growth of the algae and cyanobacteria. During the early culture days, there was a slight growth promotion with the coccal green alga and to a much lesser degree with the cyanobacterium. Yet, the major effect were significant reductions in cell yield in both primary producer cultures. C. minutus was much more affected than D. communis. This applies particularly to the three tested Scandinavian NOM isolates, which were effective at concentrations even below 1 mg L-1 DOC. DISCUSSION: The growth promoting effect may be due to an increase in bioavailability of some trace nutrients in the presence of NOM, the release of some growth promoting substances by microbial or photochemical processing of the NOM, and/or a hormetic effect. The growth reducing effect can be explained as a herbicide-like mode of action that affects the photosystem II most prevalent by blocking the electron transport chain, absorption of electrons, or production of an internal oxidative stress after processing the bioconcentrated HS. Furthermore, it may be postulated that also phototoxicity of these HS in the algal cells contributes to the overall toxicity; however, experimental evidence is lacking so far. CONCLUSIONS: Upon exposure to HS, cyanobacteria appear to be much more sensitive than coccal green algae and respond in growth reduction. This high sensitivity of cyanobacteria to HS may explain phytoplankton patterns in the field. Eutrophic, humic-rich lakes do not support the cyanobacterial blooms characteristic of eutrophic, but humic-poor lakes. In the humic-rich systems, raphidophytes or, less frequent, specific coccal greens are more common. Obviously, cyanobacteria appear to be unable to make advantage of their accessory pigments (phycocyanin) to exploit the reddish light prevailing in humic-rich lakes. RECOMMENDATION: At present, no effective structure can be figured out which may be responsible for the adverse effect on the cyanobacterial species. It is reserved to future research whether or not HS may be applied more specifically (for instance, with elevated moieties of the effective structures) as a natural geochemical to combat cyanobacterial blooms.

GOALS, SCOPE AND BACKGROUND: Anaerobic digestion of organic household waste can lead to an increase in dioxin-like content, as determined by dioxin-specific bioassays. This may be a result of bioactivation of Ah receptor (AhR) agonists into more potent congeners. Work towards identifying the contributing compound groups is important in order to understand the mechanisms and to assess the relevance behind this increase in dioxin-like toxicity, since the residue can be used as a soil fertilising agent. The aim with the present work was to identify compound groups with AhR agonistic properties that caused the previously reported increase in dioxin-like activity after anaerobic biodegradation METHODS: Firstly, chemical fractionation combined with dioxin bioassay testing was used to find bioactive classes of compounds. Secondly, batch digestion experiments with an externally added polychlorinated biphenyl (PCB) mixture (Clophen A50) and with decabrominated diphenyl ether (decaBDE), respectively, were studied as a possible process for transformation of precursors into more potent, dioxin-like compounds. Mesophilic (37ºC) and thermophilic (55ºC) anaerobic digestion were studied. Two different dioxin-specific bioassays were used to analyse AhR agonists in the biodegraded material, the CELCAD and the DR-CALUX. RESULTS AND DISCUSSION: AhR agonist activity was detected in both di- and polyaromatic fractions of digestate extracts, which indicated that a diverse mixture of compounds contributed to the bioassay responses. No quantifiable activities were induced by the monoaromatic fractions. Further fractionation based on planarity revealed higher concentrations of AhR agonists than what was detected after the first fractionation, probably due to non-additive biological interactions of compounds in the extract that were removed in the second fractionation. These results showed significant activity in the non-planar diaromatic fractions and in the co-planar fractions of both diaromates and polyaromates. In the batch experiment with externally added PCB, an increase in dioxin-like activity was seen after 21 days of digestion at mesophilic conditions. After completed digestion, the content of AhR agonists was equal to the start concentration. PCB analysis with GC-MS indicated that dehalogenation of PCBs occurred in the digestors. The batch experiment with decaBDE showed no significant changes in TEQ-concentrations over time. CONCLUSIONS: The results show that the previously reported increase of AhR agonists during mesophilic anaerobic digestion is probably due to an accumulation of several different groups of AhR agonists, both diaromatic and polyaromatic, and both co-planar and non-planar. Batch experiments with externally added PCBs and decaBDE, respectively, did not result in any accumulation of AhR agonist activity after completed digestion, even though chemical analysis indicate a dechlorination of PCBs. Complex, unfractionated extracts were difficult to test using the bioassay approach. Removal of AhR antagonists or otherwise interacting compounds during fractionation may yield bio-TEQ values that are much higher than in the original extract. RECOMMENDATIONS AND PERSPECTIVE: Our results indicate that the environmental risk that AhR agonists may pose concerning large-scale anaerobic digestion of organic household waste probably depends on the efficiency of the digester and the sludge residence time. In order to obtain reliable results with the bioassays, an extensive cleanup and fractionation procedure is necessary. Without clean up and fractionation, there is a risk for false negatives and misleading conclusions. DR-CALUX and CELCAD were both suitable for these kinds of studies, provided that suitable fractionation methods are used.

GOAL, SCOPE AND BACKGROUND: Aquaculture activities are well known to be the major contributor to the increasing level of organic waste and toxic compounds in the aquaculture industry. Along with the development of intensive aquaculture in China, concerns are evoked about the possible effects of everincreasing aquaculture waste both on productivity inside the aquaculture system and on the ambient aquatic ecosystem. Therefore, it is apparent that appropriate waste treatment processes are needed for sustaining aquaculture development. This review aims at identifying the current status of aquaculture and aquaculture waste production in China. MAIN FEATURES: China is the world’s largest fishery nation in terms of total seafood production volume, a position it has maintained continuously since 1990. Freshwater aquaculture is a major part of the Chinese fishery industry. Marine aquaculture in China consists of both land-based and offshore aquaculture, with the latter mostly operated in shallow seas, mud flats and protected bays. The environmental impacts of aquaculture are also striking. RESULTS: Case studies on pollution hot spots caused by aquaculture have been introduced. The quality and quantity of waste from aquaculture depends mainly on culture system characteristics and the choice of species, but also on feed quality and management. Wastewater without treatment, if continuously discharged into the aquatic environment, could result in remarkable elevation of the total organic matter contents and cause considerable economy lost. Waste treatments can be mainly classified into three categories: physical, chemical and biological methods. DISCUSSION: The environmental impacts of different aquaculture species are not the same. New waste treatments are introduced as references for the potential development of the waste treatment system in China. The most appropriate waste treatment system for each site should be selected according to the sites’ conditions and financial status as well as by weighing the advantages and disadvantages of each system. Strategies and perspectives for sustainable aquaculture development are proposed, with the emphasis on environmental protection. CONCLUSIONS: Negative effects of waste from aquaculture to aquatic environment are increasingly recognized, though they were just a small proportion to land-based pollutants. Properly planned use of aquaculture waste alleviates water pollution problems and not only conserves valuable water resources but also takes advantage of the nutrients contained in effluent. It is highly demanding to develop sustainable aquaculture which keeps stocking density and pollution loadings under environmental capacity. RECOMMENDATIONS AND PERSPECTIVES: The traditional procedures for aquaculture waste treatment, mainly based on physical and chemical means, should be overcome by more site-specific approaches, taking into account the characteristics and resistibility of the aquatic environment. Further research needs to improve or optimize the current methods of wastewater treatment and reuse. Proposed new treatment technology should evaluate their feasibility at a larger scale for practical application.

BACKGROUND, AIM AND SCOPE: Discussions on management of whales and whaling are factually monopolized by the International Whaling Commission (IWC), resulting in a limitation of information flow to outside communities. With an aim to improve the situation, this article briefly reviews whaling and dolphin/porpoise fisheries in Japan, which is recognized to be the world largest cetacean exploitation.MAIN FEATURES: The Japanese government grants an annual take of 22,647 cetaceans of 15 species for scientific whaling and various kinds of active dolphin/porpoise fisheries by the nationals. Further, over 100 baleen whales and numerous small cetaceans are taken in passive net fisheries. They are used mostly for human consumption and some for aquarium display.Results. Sustainability of the take is not evident and some populations have shown a historical decline. The Japanese program of scientific whaling has been reviewed by IWC and its Scientific Committee (SC), although they have arrived at no consensus. DISCUSSION: The current scientific whaling program invites arguments from the view points of science as well as concerning the ethics of scientists, economy, and interpretation of the International Convention for Regulation of Whaling (ICRW) of 1946. The scientific whaling and other Japanese cetacean fisheries are benefited from nationalistic public attitude, and ambiguity and weakness of the ICRW. CONCLUSIONS: Japanese cetacean harvest will continue supported by domestic demand for whale products as long as the proceeds can sustain the operation, even with criticisms from outside communities. RECOMMENDATIONS AND PERSPECTIVE: . For safe management of small cetaceans exploited by Japan, studies are urgent on the population structure, abundance and validity of catch statistics. The results should be open to scientific communities.

GOAL, SCOPE AND BACKGROUND: The aim of this work is to show the ability of several fungal species, isolated from arsenic polluted soils, to biosorb and volatilize arsenic from a liquid medium under laboratory conditions. Mechanisms of biosorption and biovolatilization play an important role in the biogeochemical cycle of arsenic in the environment. The quantification of production of volatile arsenicals is discussed in this article. METHODS: Heat-resistant filamentous fungi Neosartorya fischeri, Talaromyces wortmannii, T. flavus, Eupenicillium cinnamopurpureum, originally isolated from sediments highly contaminated with arsenic (more than 1403 mg.l-1 of arsenic), and the non-heat-resistant fungus Aspergillus niger were cultivated in 40 mL liquid Sabouraud medium (SAB) enriched by 0.05, 0.25, 1.0 or 2.5 mg of inorganic arsenic (H3AsO4). After 30-day and 90-day cultivation under laboratory conditions, the total arsenic content was determined in mycelium and SAB medium using the HG AAS analytical method. Production of volatile arsenic derivates by the Neosartorya fischeri strain was also determined directly by hourly sorption using the sorbent Anasorb CSC (USA). RESULTS: Filamentous fungi volatilized 0.025–0.321 mg of arsenic from the cultivation system, on average, depending on arsenic concentrations and fungal species. The loss of arsenic was calculated indirectly by determining the sum of arsenic content in the mycelium and culture medium. The amount of arsenic captured on sorption material was 35.7 ng of arsenic (22nd day of cultivation) and 56.4 ng of arsenic (29th day of cultivation) after one hour's sorption. Biosorption of arsenic by two types of fungal biomass was also discussed, and the biosorption capacity for arsenic of pelletized and compact biomass of Neosartorya fischeri was on average 0.388 mg and 0.783 mg of arsenic, respectively. DISCUSSION: The biosorption and amount of volatilized arsenic for each fungal species was evaluated and the effect of initial pH on the biovolatilization of arsenic was discussed. CONCLUSIONS: The most effective biovolatilization of arsenic was observed in the heat-resistant Neosartorya fischeri strain, while biotransformation of arsenic into volatile derivates was approximately two times lower for the non-heat-resistant Aspergillus niger strain. Biovolatilization of arsenic by Talaromyces wortmannii, T. flavus, Eupenicillium cinnamopurpureum was negligible. Results from biosorption experiments indicate that nearly all of an uptaken arsenic by Neosartorya fischeri was transformed into volatile derivates. RECOMMENDATIONS AND PERSPECTIVE: Biovolatilization and biosorption have a great potential for bioremediation of contaminated localities. However, results showed that not all fungal species are effective in the removal of arsenic. Thus, more work in this research area is needed.

GOALS, SCOPE AND BACKGROUND: It has been observed that hydrocarbon treated wastewaters still contain high COD and a number of intermediates. This suggests that the required catabolic gene pool for further degradation might be absent in the system or, that its titer value is not significant enough. By providing the desired catabolic potential, the overall efficiency of the treatment system can be improved. This study aims to demonstrate this concept by bioaugmentation of a lab-scale reactor treating refinery wastewater with a consortium having the capacity to complement the alkB genotype to the available microbial population. METHODS: Two reactors were set up using activated biomass collected from a refinery treatment plant and operated at a continuous mode for a period of 8 weeks. The feed to both reactors was kept constant. Crude oil was spiked regularly. One reactor was bioaugmented with a consortium previously described for crude oil spill remediation. The efficiency of the bioaugmented reactor was demonstrated by reduced COD. The changes in the microbial population over a period of time were analyzed by RAPD. Catabolic activity of the biomass in both reactors was monitored by PCR. The presence of the catabolic loci was confirmed by Southern Hybridization. RESULTS AND DISCUSSION: 52.2% removal of COD was observed in the bioaugmented reactor while only 15.1% reduction of COD was observed in the reactor without bioaugmentation. The change in microbial population can be seen from the 4th week, which also corresponds to improved catabolic activity. The presence of the bedA locus was seen in all samples, which indicates the presence of aromatic degraders, but the appearance of the alkB locus, from the 6th week onwards, which was observed only in the samples from the bioaugmented reactor. The results suggest that the gene pool of the bioaugmented reactor has catabolic loci that can degrade accumulated intermediates, thus improving the efficiency of the system. CONCLUSIONS: In this study, improvement of efficiency of bioremediation was demonstrated by addition of catabolic loci that are responsible for degradation. Bioaugmentation was carried out in biomass that was collected from an ETP (effluent treatment plant) treating hydrocarbon containing wastewater to study the strategies for improvement of the treatment system. Biostimulation, only marginally improved the efficiency, when compared to bioaugmentation. The improved efficiency was demonstrated by COD removal. The presence of the alkB locus suggests the importance of a catabolic gene pool that acts on accumulated intermediates. It is well documented that straight chain aliphatics and intermediates of aromatic compounds after ring cleavage, accumulate in refinery wastewater systems, thereby hindering further degradation of the wastewater. Supplementation of a catabolic gene pool that treats the lower pathway compounds and alkanes will improve the overall efficiency. In this study, results suggest that the alkB locus can also be used to monitor the degradative mode of the activated biomass. RECOMMENDATIONS AND PERSPECTIVE: . Pollution from petroleum and petroleum products around the globe are known to have grave consequences on the environment. Bioremediation, using activated sludge, is one option for the treatment of such wastes. Effluent treatment plants are usually unable to completely degrade the wastewater being treated in the biological unit (the aerator chambers). The efficiency of degradation can be improved by biostimulation and bioaugmentation. This study demonstrates the improved efficiency of a treatment system for wastewater containing hydrocarbons by bioaugmentation of a consortium that supports degradation. Further experiments on a pilot scale are recommended to assess the use of bioaugmentation on a large scale. The use of molecular tools, like DNA probes for alkB, to monitor the system also needs to be explored.