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The recent development of culture-independent methods has revolutionized the study of complex microbial communities such as those present in activated sludge treatment systems. DNA probes that hybridize to genes coding for key enzymes that catalyze microbial processes have been widely used. Can such probes be used to quantify target genes and thus quantify the potential of a microbial community to carry out a reaction of interest? / Optimal conditions for DNA extraction, probe validation, hybridization, and activity measurements were determined for the pulp and paper treatment system environment under study. Using gene probes for key denitrification genes (nirS, nirK), the correlation between denitrifiers and denitrification activity in an enrichment culture and activated sludge samples was tested. The same correlation between nitrogen fixation and nitrogen-fixing bacteria in primary clarifiers was assessed using a probe for the gene encoding a component of the nitrogenase enzyme (nifH). This work was successful in establishing the correlation between gene numbers and their corresponding enzymatic activity and thus supports the quantitative hybridization approach for the monitoring of microbial communities. (Abstract shortened by UMI.)

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Studies were undertaken to determine the effect of Kraft mill effluent (KME) on a representative species of the aquatic environment. By transplanting a population of Pacific oysters (Crassostrea gigas) to the Port Mellon area (the site of a Kraft mill), the effect of varying concentrations (on a distance from the mill basis) of the pulp mill waste was monitored in terms of changes in shell dimension, body mass (meat) weight, visual observation of the oysters' physiological state and oyster mortality. A seasonal hydrographic survey was conducted at three regions within the study area to monitor changes in water quality imposed by the effluent. In the areas of effluent imposition (oyster stations 1, 2 and 3) the oysters decreased or showed little gain in shell dimension. The body mass of those oysters nearest the mill outfall began to decline (on a weight basis) shortly after placement. At Station 2 and 3 the deterioration in body mass due to changes in water quality began a short time after the decline at Station 1. Changes in the physiological state of the oyster expressed as a darkening of the gills and mantle edge and variations in body mass texture, can be correlated to an oyster's distance from the mill for each collection timei A mortality rate was calculated at each station for all collection times. The mortality rates at Station 1 (100% in 12 months), Station 2 (50% in 20 months), and Station 3 (20% in 24 months) were extensive and proportional to effluent levels. Several of the changes in water quality (increased temperature, reduced salinities, low oxygen contents, variable pH, dissolved and particulated organic matter and chemical additions) imposed by the effluent were individually tested as the major cause of oyster deterioration. Firstly, the critical oxygen tension (the P02 where V02 declines below the routine rate) was determined as 40 mmHg. Correlating this to the range of 02 levels at each station during a tidal cycle, the oxygen demand of the effluent was not considered as a major cause of oyster mortality. A range of filtered, neutralized (pH 7.0 at 22°C) and aerated percentage KME/volume (0-50%) test solutions were monitored in terms of their effect on the percentage time of shell closure. In these experiments percentages above 20 greatly increased the time of shell closure. The effect of shell closure on oysters was tested by continual (up to 28 days) periods of air exposure. In these experiments the P02, zC02 and pH of the pallial fluid was monitored (from time 0 to 28 days) to determine if anaerobic metabolism was undertaken and if it was, the time span of anaerobic life in juvenile oysters. Anaerobic metabolism was concluded to maintain life in juvenile oysters for 22 days. These results were consistent with the hypothesis that KME is deleterious to oyster populations. At high concentrations of effluent the duration of shell closure is extensive such that an anaerobic death results. At lower concentrations the effluent imposed changes in water quality are responsible for the gradual decline in oyster well-being. / Science, Faculty of / Zoology, Department of / Graduate

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Disposal of effluent from pulp and paper industry is one of the major problems faced by entrepreneur in view of increasing environmental standards day by day. In addition to this, industry loses economic value by disposing the effluent or selling it for a low price to other industries. Therefore, to address this problem, in the present study, 2 pulp mill effluents were selected to recover the economic value namely Hemi caustic stream and brown stock filtrate. As far as the recovery of value of hemi caustic stream is concerned, freeze concentration technique was used to recover water in its pure form and membrane separation was used to separate hemi cellulose from effluent so that permeate can be used as a pure source of caustic elsewhere. In addition to this, hemi caustic stream was subjected to acid hydrolysis to convert hemi cellulose into sugars. These sugars can be used to produce bioethanol. As far as the recovery of values of brown stock filtrate is concerned, it was proposed to recycle brown stock filtrate as a source of washing water for brown stock washers in the mill. However, continuous recycling of brown stock filtrate into the process causes building up of extractives in the recycle stream which in turn might deposit on the pulp and affects the quality of the pulp. Therefore, it was decided to separate extractives from the brown stock filtrate before recycle it into the mill. Dissolved Air flotation technique was used to achieve the above mentioned objective. An attempt was made to develop an improved and most reliable version of existing extractives measurement method to quantify the performance of Dissolved air flotation technique.

Phylogenetic hybridization and phenotypic fingerprinting were applied to the analysis of bacterial communities in wastewater treatment systems. These approaches were aimed at (i) developing monitoring tools able to foresee operational problems, and (ii) providing the rationale to optimize the operation of bioreactors. The work presented is intended to first describe the community found in two reactors treating pulp and paper mill effluent, and second evaluate the possibilities of these techniques with respect to the development of new monitoring tools. / Phylogenetic membrane hybridization showed that the bacterial communities were dominated by Alpha and Beta Proteobacteria, a structure probably linked to the low F:M ratio. Other important factors determining the community structure were the proportion of COD in the high molecular weight fraction, the sludge age, phosphate addition, and the concentration of specific compounds (alcohols, phenols, volatile fatty acids) in the influent. The community structure partly determined the sludge characteristics demonstrating its potential value in the assessment of reactor performance. The results obtained by phylogenetic membrane hybridization suggest that the probes used in a monitoring tool would not need to be targeted to the species level to provide relevant information. However, they also suggest that the technique is more sensitive to changes in population density as opposed to changes in bacterial metabolism. / Phenotypic fingerprinting measured a smaller difference between the communities of the two reactors studied than what was measured by phylogenetic membrane hybridization. However, differences in heterotrophic activities observed between the two communities were linked to differences in influent composition.

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