Effect of effluent from a nitrogen fertilizer factory and a pulp mill on the distribution and abundance of Aeromonas hydrophila in Albemarle Sound, North Carolina.

The density of Aeromonas hydrophila, standard count bacteria, fecal coliform bacteria, and 18 physical and chemical parameters were measured simultaneously at six sites for 12 months in Albemarle Sound, N.C. One site was above and two sites were below the discharge plume of a Kraft pulping process paper mill. The fourth site was above and the remaining two sites were below the discharge point of a nitrogen fertilizer factory. The impact of the pulp mill on water quality was acute, whereas that of the nitrogen fertilizer factory was chronic and much more subtle. Diffusion chamber studies indicated that A. hydrophila survival is increased by pulp mill effluent and decreased by nitrogen fertilizer factory effluent. From correlation and regression analysis, A. hydrophila was found to be directly affected by phytoplankton density and, thus, indirectly by concentrations of phosphate, nitrate, and total organic carbon. These two point sources are suspect as indirect causes of red-sore disease epizootics, a disease of fish caused by A. hydrophila.     

Effects of different nitrogen sources on lignocellulose degradation and APPL production

Naphthalene utilizing bacteria were isolated from several sites above and below the discharge from a coking plant. The distribution of the bacteria was influenced by the effluent. Of these isolates, 11·4% obtained from the effluent discharge site, contained plasmids. No conjugal transfer of naphthalene utilizing ability was observed in over 1000 matings. Curing and transformation experiments demonstrated that one plasmid pNB33 (101 kb) was concerned with naphthalene catabolism.     

Investigation of the biotransformation of pentachlorophenol and pulp paper mill effluent decolorisation by the bacterial strains in a mixed culture

Mixed culture of two bacterial strains Bacillus sp. and Serratia marcescens showed potential pentachlorophenol (PCP) degradation and decolorisation of pulp paper mill effluent. The physico-chemical quality of pulp paper mill effluent has been analyzed after 168 h incubation period degraded by mixed culture. The study revealed that it has decreased high load of BOD, COD, TS, TDS, TSS, sulphate, phosphate, total nitrogen, total phenols, metals and different salts (i.e. chloride, sodium, nitrate, potassium) at 168 h incubation period. PCP degradation in pulp paper mill effluent was confirmed by HPLC analysis. Mixed culture was found to degrade PCP up to (94%) present in pulp paper mill effluent with 1% glucose and 0.5% peptone (w/v) at 30 ± 1 °C, pH 8.0 ± 0.2 at 120 rpm in 168 h incubation period. The simultaneous release of chloride ion up to 1200 mg/l at 168 h emphasized the bacterial dechlorination in the medium. The pulp paper mill effluent degradation was also supported by decline in pH, AOX (absorbable organic halides), color, D.O., BOD, COD and PCP. The analysis of pulp paper mill effluent degradation products by GC–MS analysis revealed the formation of low molecular weight compound like 2-chlorophenol (RT = 3.8 min) and tetrachlorohydroquinone (RT = 11.86 min) from PCP extracted degraded sample. Further, mixed culture may be used for bioremediation of PCP containing pulp paper mill waste in the environment.     

The effect of sewage effluent on denitrification in Grasmere (English Lake District)

SUMMARY. Denitrification processes in Grasmere were studied using several techniques. Sediments were analysed for nitrate reductase activity and the accumulation of nitrogenous gases in samples incubated in the laboratory. Changes in the concentrations of inorganic nitrogen species in the water column and the release of gas bubbles into gas traps suspended above the sediment were also measured. Sewage effluent entered the lake via the main inflow and faecal bacteria were used as tracers to map the path of the river (and effluent) across the lake. The main river flow was west of the island in the centre of the lake and across the smaller of the two main basins. A preliminary study indicated that this western basin was the site of most active denitrification and this was confirmed in the following year when a seasonal survey showed that deoxygenation, nitrate reduction and ammonium accumulation was more advanced at this site. In addition, this basin was the only area where consistent discharge of nitrogen and methane from the profundal zone was observed. The nitrate reductase assay demonstrated that the sediment was the major site of denitrification, and that activity in the water column was higher in the anoxic hypolimnion than in the euphotic zone. Gas evolution from incubated sediment samples was analysed by gas liquid chromatography (GLC) and indicated that maximum denitrifying activity occurred below the mud surface in oxidized sediments. There was reasonable agreement between the enzyme assay and the GLC technique but neither predicted events in the field satisfactorily. Viable, most probable number (MPN) counts of denitrifying bacteria had similar limitations. A bimodal seasonal pattern of nitrate reductase activity was observed at four sites during thermal stratification, but this could not be explained fully. The major site of denitrification was identified and it was calculated that the nitrogen gas discharged was equivalent to c. 54% of the nitrate removed from the water column.     

Metal biosorption in lignocellulosic biofuel biorefinery effluent: an initial step towards sustainability of water resources

Biosorption of metals by microorganisms is a promising technology to remove accumulated non-process elements in highly recycled biorefinery process water. Removal of these elements would enable greater water reuse and reduce the environmental impact of effluent discharge. A model lignocellulosic ethanol biorefinery wastewater was created based on pulp mill effluent. This generated a wastewater with an environmentally realistic high loading of dissolved natural organic matter (900?mg/l), a potentially important factor influencing metal biosorption. Analysis of feedstock and pulp mill effluent indicated that Mn and Zn are likely to be problematic in highly recycled lignocellulosic ethanol biorefinery process water. Therefore, the growth of several bacteria and fungi from existing collections, and some isolated from pulp mill effluent were tested in the model wastewater spiked with Mn and Zn (0.2?mM). Wastewater isolates grew the best in the wastewater. Metal uptake varied by species and was much greater for Zn than Mn. A bacterium, Novosphingobium nitrogenifigens Y88(T), removed the most metal per unit biomass, 35 and 17?mg?Mn/g. No other organism tested decreased the Mn concentration. A yeast, Candida tropicalis, produced the most biomass and removed the most total metal (38?% of Zn), while uptake per unit biomass was 24?mg?Zn/g. These results indicate that microorganisms can remove significant amounts of metals in wastewater with high concentrations of dissolved natural organic matter. Metal sorption by autochthonous microorganisms in an anaerobic bioreactor may be able to extend water reuse and therefore lower the water consumption of future biorefineries.     

Bioremediation of pulp and paper mill effluent by a novel fungal consortium isolated from polluted soil

Two basidiomycetous fungi (Merulius aureus syn. Phlebia sp. and an unidentified genus) and a deuteromycetous fungus (Fusarium sambucinum Fuckel MTCC 3788) were isolated from soils affected with effluents of a pulp and paper mill over several years. These isolates were immobilized on nylon mesh and the consortium was used for bioremediation of pulp and paper mill effluent in a continuously aerated bench-top bioreactor. The treatment resulted in the reduction of color, lignin and COD of the effluent in the order of 78.6%, 79.0% and 89.4% in 4 days. A major part of reductions in these parameters occurred within first 24h of the treatment, which was also characterized by a steep decline in the pH of the effluent. During this period, total dissolved solids, electrical conductivity and salinity of the effluent also registered marked decline. It is pertinent to note that this is the first report of bioremediation of pulp and paper mill effluent by an immobilized fungal consortium.     

Effect, distribution, and prevalence of Glugea stephani (Microspora) in winter flounder (Pleuronectes americanus) living near two pulp and paper mills in Newfoundland

A study was conducted to determine the effects, geographical distribution, and prevalence of a microsporan parasite, Glugea stephani, in winter flounder (Pleuronectes americanus) in Newfoundland. Fish were captured by SCUBA divers in several coastal areas, including 2 embayments where pulp and paper mill effluent was discharged, as well as a number of pristine sites. Fish health was assessed by comparing histological profiles, condition factors (K), organosomatic indices and blood values between infected and uninfected samples. Multifocal xenomas of G. stephani were observed in several organs of fish taken near contaminated sites, whereas infected samples captured at a pristine site harbored the cysts only in the wall of the digestive tract. Proliferative inflammation, granuloma formation, and focal necrosis were associated with the infection primarily in the liver and kidney. Condition factors and blood values were lower and ovarian development inhibited or delayed in infected flounder. The multifocal infection occurred only in flounder in 2 embayments in western Newfoundland where pulp and paper mill effluent was discharged. Prevalence varied seasonally, with a peak in autumn and a low in spring. It is likely that the multifocal infection was associated with immunodepression after exposure to the contaminant.     

Nutrient enrichment of northern rivers in response to pulp mill and municipal discharges

To evaluate the effects of pulp mill and municipal sewageeffluents on the nitrogen (N) and phosphorus (P) status of northernCanadian rivers, the Northern River Basins Study required an integratedresearch and assessment program consisting of field observation andexperimentation. Analysis of monitoring data collected over 3–13 yshowed that on an annual basis, pulp mills contributed 22% of theP and 20% of the N load discharged from the Wapiti to the Smokyriver, and 6 to 16% of the P load and 4 to 10% of the Nload in the Athabasca River. Despite these low contributions, N and Pconcentrations were elevated below pulp mill discharges on all threerivers during the low discharge period of September to April. Insitu experiments conducted with nutrient diffusing substrata showedthat periphyton biomass was maintained at low levels by insufficient Pin the upper reaches of the Athabasca River and insufficient N+P inthe Wapiti River upstream of point-source discharges. In contrast,effluent loading from pulp mill and sewage inputs alleviated nutrientlimitation downstream of major discharges on both rivers. Experimentsconducted in artificial streams located beside the Athabasca Riverupstream of the first pulp mill showed that P addition increasedperiphyton biomass (expressed as chlorophyll a content) suchthat biomass increased sharply at low P concentrations (2–5µg/L soluble reactive P) and approached saturation at 35µg/L soluble reactive P. As a result of recommendations fromthis integrated program of river monitoring and experimentation,nutrient loading has been reduced to the most affected river(Wapiti-Smoky) and federal and provincial departments of environment arereviewing loading limits for other river reaches.     

Influence of Long Term Irrigation with Pulp and Paper Mill Effluent on the Bacterial Community Structure and Catabolic Function in Soil

Microbial communities play a vital role in maintaining soil health. A multiphasic approach to assess the effect of pulp and paper mill effluent on both the structure and function of microbial soil communities is taken. Bacterial communities from agricultural soils irrigated with pulp and paper mill effluent were compared to communities form soils irrigated with well water. Samples were taken from fields in the state of Uttarakhand, India, where pulp and paper mill effluent has been used for irrigation for over 25 years. Comparisons of bacterial community structure were conducted using sequencing of the 16S rRNA gene from both isolates and clone libraries attained from the soil. Community-level physiological profiling was used to characterize the functional diversity and catabolic profile of the bacterial communities. The multiphasic approach using both physiological and molecular techniques proved to be a powerful tool in evaluating the soil bacterial community population and population differences therein. A significant and consistent difference in the population structure and function was found for the bacterial communities from soil irrigated with effluent in comparison to fields irrigated with well water. The diversity index parameters indicated that the microbial community in pulp and paper mill effluent irrigated fields were more diverse in both structure and function. This suggests that the pulp and paper mill effluent is not having a negative effect on the soil microbial community, but in fact may have a positive influence. In terms of soil health, this finding supports the continued use of pulp and paper mill effluent for irrigation. This is however only one aspect of soil health which was evaluated. Further studies on soil resistance and robustness could be undertaken to holistically evaluate soil health in this situation.     

Levels of damage of Scots pine and Norway spruce caused by needle miners along a SO2 gradient

Needle damages caused by mining insects on Scots pine and Norway spruce were studied in the vicinity or a pulp mill. The abundance of needles mined by the pine bud moth Exoteleia dodecella (Lepidoptera: Gelechiidae) on Scots pine Pinus sylvestris , and the spruce needle miner Epinotia tedella (Lepidoptera: Tortricidae) on Norway spruce Picea abies had a significantly peaked response curve on logarithmic distance scale. For pine bud moth, the maximum expected population density was estimated to be at the distance of 1.35 km from the factory. The maximum expected population density for the spruce needle miner was at the distance of 1.53 km from the factory. However, for both species the curves were significantly different among transects. Both species had a peaked and significant response to sulphur level in needles as well. The maximum expected density in pine was at 1270 ppm, and in spruce at 1070 ppm sulphur concentration in pine needles. The results are consistent with earlier reports demonstrating that these mining insects frequently attack trees suffering from air pollution. The nonlinear response of both species to distance from the pulp mill suggests that E. dodecella on pine and E. tedella on spruce are rather indicators of the zone of intermediate air pollution than of strongly polluted or nearly unpolluted sites. This also agrees with the plant stress-insect performance hypothesis indicating that insect response varies with the magnitude of stress, and at very high stress levels a tree no longer provides the insects with relevant food.     

Source bioaerosol concentration and rRNA gene-based identification of microorganisms aerosolized at a flood irrigation wastewater reuse site

Reuse of partially treated domestic wastewater for agricultural irrigation is a growing practice in arid regions throughout the world. A field sampling campaign to determine bioaerosol concentration, culturability, and identity at various wind speeds was conducted at a flooded wastewater irrigation site in Mexicali, Baja California, Mexico. Direct fluorescent microscopy measurements for total microorganisms, culture-based assays for heterotrophs and gram-negative enteric bacteria, and small-subunit rRNA gene-based cloning were used for microbial characterizations of aerosols and effluent wastewater samples. Bioaerosol results were divided into two wind speed regimens: (i) below 1.9 m/s, average speed 0.5 m/s, and (ii) above 1.9 m/s, average speed 4.5 m/s. Average air-borne concentration of total microorganisms, culturable heterotrophs, and gram-negative enteric bacteria were, respectively, 1.1, 4.2, and 6.2 orders of magnitude greater during the high-wind-speed regimen. Small-subunit rRNA gene clone libraries processed from samples from air and the irrigation effluent wastewater during a high-wind sampling event indicate that the majority of air clone sequences were more than 98% similar to clone sequences retrieved from the effluent wastewater sample. Overall results indicate that wind is a potential aerosolization mechanism of viable wastewater microorganisms at flood irrigation sites.     

Source Bioaerosol Concentration and rRNA Gene-Based Identification of Microorganisms Aerosolized at a Flood Irrigation Wastewater Reuse Site

Reuse of partially treated domestic wastewater for agricultural irrigation is a growing practice in arid regions throughout the world. A field sampling campaign to determine bioaerosol concentration, culturability, and identity at various wind speeds was conducted at a flooded wastewater irrigation site in Mexicali, Baja California, Mexico. Direct fluorescent microscopy measurements for total microorganisms, culture-based assays for heterotrophs and gram-negative enteric bacteria, and small-subunit rRNA gene-based cloning were used for microbial characterizations of aerosols and effluent wastewater samples. Bioaerosol results were divided into two wind speed regimens: (i) below 1.9 m/s, average speed 0.5 m/s, and (ii) above 1.9 m/s, average speed 4.5 m/s. Average air-borne concentration of total microorganisms, culturable heterotrophs, and gram-negative enteric bacteria were, respectively, 1.1, 4.2, and 6.2 orders of magnitude greater during the high-wind-speed regimen. Small-subunit rRNA gene clone libraries processed from samples from air and the irrigation effluent wastewater during a high-wind sampling event indicate that the majority of air clone sequences were more than 98% similar to clone sequences retrieved from the effluent wastewater sample. Overall results indicate that wind is a potential aerosolization mechanism of viable wastewater microorganisms at flood irrigation sites.     

Detection of Aeromonas hydrophila in a drinking-water distribution system: a field and pilot study

A 16-month study was conducted on the presence of Aeromonas hydrophila in drinking water in Indiana, U.S.A. Enumeration was conducted in source water, in various sites within a water treatment plant, and in the distribution system in both bulk water and biofilm, as well as in a simulated (annular reactors) drinking-water distribution system. Presumptive Aeromonas spp. counts on source waters regularly approached 10(3)-10(4) CFU/100 mL, during summer months and granular activated carbon - filtered water counts ranged from <1 to 490 CFU/100 mL. In source water, presumptive Aeromonas levels were related to water temperature. Aeromonas hydrophila was never detected in the treatment plant effluent or distributed bulk water, showing disinfectant efficiency on suspended bacteria; however, isolates of A. hydrophila were identified in 7.7% of the biofilm samples, indicating a potential for regrowth and contamination of drinking-water distribution systems.     

Nitrogen fertilizer factory effects on the amino acid and nitrogen content in the needles of Scots pine

The aim of the research was to evaluate the content of amino acids in the needles of Pinus sylvestris growing in the area affected by a nitrogen fertilizer factory and to compare them with other parameters of needles, trees, and sites. Three young-age stands of Scots pine were selected at a distance of 0.5 km, 5 km, and 17 km from the factory. Examination of the current-year needles in winter of the year 2000 revealed significant (p < 0.05) differences between the site at a 0.5-km distance from the factory and the site at a 17-km distance from the factory--with the site closest to the factory showing the highest concentrations of protein (119%), total arginine (166%), total other amino acids (depending on amino acid, the effect ranged between 119 and 149%), free arginine (771%), other free amino acids (glutamic acid, threonine, serine, lysine--depending on amino acid, the effect ranged between 162 and 234%), also the longest needles, widest diameter, largest surface area, and heaviest dry weight (respectively, 133, 110, 136, and 169%). The gradient of nitrogen concentration in the needles was assessed on the selected plots over the period of 1995-2000, with the highest concentration (depending on year, 119 to 153%) documented in the site located 0.5 km from the factory. Significant correlations were determined between the total amino acid contents (r = 0.448 -0.939, p < 0.05), some free amino acid (arginine, aspartic acid, glutamic acid, lysine, threonine, and serine) contents (r = 0.418 - 0.975, p < 0.05), and air pollutant concentration at the sites, the distance between the sites and the factory, and characteristics of the needles. No correlation was found between free or total arginine content and defoliation or retention of the needles. In conclusion, it was revealed that elevated mean monthly concentration of ammonia (26 microg m(-3)) near the nitrogen fertilizer factory caused changes in nitrogen metabolism, especially increasing (nearly eight times) concentration of free arginine in the needles of Scots pine.     

Chitin: a potential new alternative nitrogen source for the tertiary,algal-based treatment of pulp and paper mill wastewater

Every day, pulp and paper mills in the USA discharge millions of liters of wastewater. Primary and secondary treatment of this wastewater often enriches it with phosphorus, resulting in uncontrolled eutrophication of receiving water bodies. A new method of tertiary wastewater treatment uses controlled growth of algae in a photobioreactor to sequester phosphorus into algal biomass, which is then harvested. This typically requires addition of a nitrogen fertilizer (nitrate, ammonium, or urea) to the water. We show on the laboratory scale that chitin can be used as an alternative source of nitrogen for the tertiary treatment of pulp mill wastewater using algae. We demonstrate that phosphorus can be efficiently removed from pulp wastewater using algae and chitin. Furthermore, phosphorus removal with chitin did not result in an increase in dissolved nitrogen in the wastewater because it is insoluble, unlike conventional nitrogen fertilizers. Despite its insolubility, it has recently been found that many diverse algae and cyanobacteria can use it as a source of nitrogen. Chitin has many advantages over conventional nitrogen fertilizers for use in wastewater treatment technologies. It is the second-most abundant natural polymer and is a waste product of the shellfish industry. Chitin is sustainable, inexpensive, and carbon neutral. Thus, chitin improves the sustainability and carbon footprints associated with water treatment, while the production of commercially attractive algal biomass helps to offset costs associated with the water treatment system itself.     

Coliform bacteria and nitrogen fixation in pulp and paper mill effluent treatment systems

The majority of pulp and paper mills now biotreat their combined effluents using activated sludge. On the assumption that their wood-based effluents have negligible fixed N, and that activated-sludge microorganisms will not fix significant N, these mills routinely spend large amounts adding ammonia or urea to their aeration tanks (bioreactors) to permit normal biomass growth. N(2) fixation in seven Eastern Canadian pulp and paper mill effluent treatment systems was analyzed using acetylene reduction assays, quantitative nitrogenase (nifH) gene probing, and bacterial isolations. In situ N(2) fixation was undetectable in all seven bioreactors but was present in six associated primary clarifiers. One primary clarifier was studied in greater detail. Approximately 50% of all culturable cells in the clarifier contained nifH, of which >90% were Klebsiella strains. All primary-clarifier coliform bacteria growing on MacConkey agar were identified as klebsiellas, and all those probed contained nifH. In contrast, analysis of 48 random coliform isolates from other mill water system locations showed that only 24 (50%) possessed the nifH gene, and only 13 (27%) showed inducible N(2)-fixing activity. Thus, all the pulp and paper mill primary clarifiers tested appeared to be sites of active N(2) fixation (0.87 to 4.90 mg of N liter(-1) day(-1)) and a microbial community strongly biased toward this activity. This may also explain why coliform bacteria, especially klebsiellas, are indigenous in pulp and paper mill water systems.     

Niche Differentiation of Ammonia-Oxidising Archaea (AOA) and Bacteria (AOB) in Response to Paper and Pulp Mill Effluent

Sediment organic loading has been shown to affect estuarine nitrification and denitrification, resulting in changes to sediment biogeochemistry and nutrient fluxes detrimental to estuarine health. This study examined the effects of organic loading on nutrient fluxes and microbial communities in sediments receiving effluent from a paper and pulp mill (PPM) by applying microcosm studies and molecular microbial ecology techniques. Three sites near the PPM outfall were compared to three control sites, one upstream and two downstream of the outfall. The control sites showed coupled nitrification–denitrification with minimal ammonia release from the sediment. In contrast, the impacted sites were characterised by nitrate uptake and substantial ammonia efflux from the sediments, consistent with a decoupling of nitrification and denitrification. Analysis of gene diversity demonstrated that the composition of nitrifier communities was not significantly different at the impacted sites compared to the control sites; however, analysis of gene abundance indicated that whilst there was no difference in total bacteria, total archaea or ammonia-oxidising archaea (AOA) abundance between the control and impacted sites, there was a significant reduction in ammonia-oxidising bacteria (AOB) at the impacted sites. The results of this study demonstrate an effect of organic loading on estuarine sediment biogeochemistry and highlight an apparent niche differentiation between AOA and AOB.     

Survival,Growth and Condition of Freshwater Mussels: Effects of Municipal Wastewater Effluent

Freshwater mussels (Family Unionidae) are among the most imperiled group of organisms in the world, with nearly 65% of North American species considered endangered. Anthropogenic disturbances, including altered flow regimes, habitat alteration, and pollution, are the major driver of this group''s decline. We investigated the effects of tertiary treated municipal wastewater effluent on survivorship, growth, and condition of freshwater mussels in experimental cages in a small Central Texas stream. We tested the effluent effects by measuring basic physical parameters of native three ridge mussels (Amblema plicata) and of non-native Asian clams (Corbicula fluminea), before and after 72-day exposure at four sites above and below a municipal wastewater treatment plant outfall. Survivorship and growth of the non-native Asian clams and growth and condition indices of the native three ridge mussels were significantly higher at the reference site above the outfall than in downstream sites. We attribute this reduction in fitness below the outfall to elevated nutrient and heavy metal concentrations, and the potential presence of other untested-for compounds commonly found in municipal effluent. These results, along with an absence of native mussels below the discharge, indicate a significant negative impact of wastewater effluent on both native and non-native mussels in the stream.     

Seasonal occurrence of black flies (Diptera: Simuliidae) in a desert stream receiving trout farm effluent

The distribution and abundance of black flies (Diptera: Simuliidae) in a small desert stream were influenced by environmental changes caused by recharge of water supply storage basins and an aquaculture operation. Simulium virgatum was the most abundant benthic insect collected in Whitewater Canyon (Riverside County, CA) after April; however, it was never found in trout farm effluent where the ammonium‐nitrogen concentration was > 0.25 mg/liter. S. virgatum densities downstream of the input of water from the Colorado River aqueduct were lower than at other sampling sites in the Whitewater River. Simulium tescorum, an especially anthropophilic black fly, was most abundant during February and March, was not collected from late spring through early autumn, and was found only in the highly enriched, less variable flow of trout farm effluent. The mean concentrations of ammonium nitrogen and nitrate nitrogen in the trout farm effluent nearly 1 km from the fish holding ponds were ten and two times, respectively, the ambient levels in the Whitewater River upstream of the effluent discharge point. A combination of factors probably contributed to the presence of S. tescorum in the trout farm effluent including homogenization of the flow regime, enrichment of larval resources, and the development of riparian vegetation that provided oviposition and attachment sites.     

Cloning and identification of cellulase genes from uncultured microorganisms in pulp sediments from paper mill effluent]

The metagenomic DNA of pulp sediments from paper mill effluent was extracted and purified. The 16S rDNA was amplified using the purified metagenomic DNA as template and a 16S rDNA library was prepared. Sequence analysis of 16S rDNA clones showed that diverse of uncultured bacteria inhabit in this environment, which can be classified into 4 clusters as Spirochaetes, Proteobacteria, Bacteroidetes and Firmicutes. A metagenomic library containing 10000 clones was constructed into cosmid vector, and the capacity of inserted DNA of which was 3.53 x 10(8) bp. Functional screening of the library resulted in isolation of two independent clones expressing endoglucanase activity, three independent clones expressing exoglucanase activity and two independent clones expressing beta-glucosidase activity. One clone expressing strongest enzyme activity from each activity category was chosen to be further analyzed. Three novel cellulase genes designated as umcel5L, umcel5M and umbgl3D were identified by subcloning, sequencing and expression. The umcel5L encodes an endoglucanase belonging to glycosyl hydrolase family 5, which is most related to an endoglucanase from Bradyrhizobium japonicum at 43% identity and 59% similarity. The umcel5M encodes a cellodextrinase belonging to glycosyl hydrolase family 5, which is most similar to a cellodextrinase from Fibrobacter succinogenes at 48% identity and 69% similarity. The umbgl3D encodes a putative beta-glucosidase belonging to glycosyl hydrolase family 3, which shares highest homology with a beta-glucosidase from Thermotoga maritima at 46% identity and 61% similarity. It is the first time to reveal the bacterial diversity of pulp sediments from paper mill effluent and clone novel cellulase genes from the bacteria by culture-independent method.