This paper deals with the ozonation and biological treatment for achieving the mineralisation of the non-biodegradable compound naphthalene-1,5-disulphonic acid (NADSA; C₁₀H₈O₆S₂), a basic chemical for dyes and building materials. This combination of ozonation and biological treatment is expected to minimise the ozone consumption compared to ozonation alone. To examine the influence of ozone on the biodegradability of NADSA, a lab-scale plant was run semi-continuously and batch tests were conducted for comparison. For total ozonation, 1.8 mol O₃/mol DOC have to be used. For optimum biodegradation, only 0.8 mol O₃/mol DOC are needed. The results show that by using a combination of ozonation and biological treatment the ozone consumption can be reduced by more than 50% compared to ozonation alone.

Some of the intermediate products are isolated and suggestions for their chemical structures are made. A proposal for the first steps of ozonation is also presented to illustrate a possible pathway of ozonation. All the major identifiable compounds are non-biodegradable. From these results, it is quite clear that further intermediate products are formed, which are biodegradable.

A study was undertaken to investigate the distribution of biosurfactant producing and crude oil degrading bacteria in the oil contaminated environment. This research revealed that hydrocarbon contaminated sites are the potent sources for oil degraders. Among 32 oil degrading bacteria isolated from ten different oil contaminated sites of gasoline and diesel fuel stations, 80% exhibited biosurfactant production. The quantity and emulsification activity of the biosurfactants varied. Pseudomonas sp. DS10-129 produced a maximum of 7.5 ± 0.4 g/l of biosurfactant with a corresponding reduction in surface tension from 68 mN/m to 29.4 ± 0.7 mN/m at 84 h incubation. The isolates Micrococcus sp. GS2-22, Bacillus sp. DS6-86, Corynebacterium sp. GS5-66, Flavobacterium sp. DS5-73, Pseudomonas sp. DS10-129, Pseudomonas sp. DS9-119 and Acinetobacter sp. DS5-74 emulsified xylene, benzene, n-hexane, Bombay High crude oil, kerosene, gasoline, diesel fuel and olive oil. The first five of the above isolates had the highest emulsification activity and crude oil degradation ability and were selected for the preparation of a mixed bacterial consortium, which was also an efficient biosurfactant producing oil emulsifying and degrading culture. During this study, biosurfactant production and emulsification activity were detected in Moraxella sp., Flavobacterium sp. and in a mixed bacterial consortium, which have not been reported before.

The promotory effects have been studied of the non-ionic surfactant, Pluronic F-68, on bud induction/shoot regeneration in epicotyl and cotyledon explants of Citrus depressa and on shoot regeneration from leaf segments of 4–6 week-old axenic nodal segment-derived in vitro plants of Passiflora mollissima, P. giberti and P. edulis var. flavicarpa. For epicotyls of C. depressa, supplementation of agar-solidified MS-based bud induction/shoot regeneration medium with 0.5% [w/v] Pluronic F-68 significantly (P < 0.05) increased mean fresh weight gain of cultures, percentage of explants giving shoots and number of shoots per explant. The same Pluronic concentration also enhanced the mean percentage of cotyledons exhibiting bud induction and the number of buds regenerated per cotyledon explant. Fresh weight gain was unaffected across the range of concentrations (0.001–0.5% w/v) of Pluronic F-68 evaluated for this latter explant source. For leaf explants from axenic shoot cultures of P. mollissima, supplementation of NN-based medium, containing 3 mg/l 6-benzyladenine and 2.0 mg/l kinetin with 0.001–0.5% [w/v] Pluronic F-68, significantly (P < 0.05) increased mean (± s.e.m.) biomass gain by a maximum of 2.7 ± 0.1 g fresh weight (g f.wt.) over the control. Similarly, for leaf explants of P. giberti, 0.001–0.5% [w/v] Pluronic F-68 in MS-based medium, containing 1.0 mg/l 6-BAP and 0.5 mg/l kinetin significantly (P < 0.05) increased mean percentage of explants undergoing shoot regeneration. For P. edulis leaf explants, mean f.wt. gain was also significantly (P < 0.05) higher with Pluronic F-68 at 0.001–0.5% [w/v].

The effect of an immuno-stimulative β-glucan from Schizophyllum commune, schizophyllan, on the enhancement of fish immunity was evaluated with carp and flounder. The oral administration of schizophyllan induced a reduction in cumulative mortality after a bacterial challenge. Whereas mortality in the control groups was 100% after seven days, it was only 60% after challenging carp (Cyprinus carpio) with 1 × 10⁶ cells of Aeromonas hydrophila and 70% for flounder (Paralichthys olivaceus) using 1 × 10⁶ cells of Edwardsiella tarda. The numbers of peripheral macrophages and neutrophiles, the phagocytic activities of leukocytes and the activity of serum lysozyme were increased in the fish being fed a schizophyllan-supplemented diet. These results support the findings that non-specific defence activities in fish such as the number of leukocytes, phagocytic activities and serum lysozyme activity could be enhanced by oral administration of schizophyllan. Moreover, the oral administration of schizophyllan can reduce mortality after bacterial infections depending on the size of fish and the concentrations of bacteria.

Six published fungal specific primer sets (NS1/NS2, SSU-0817/SSU11-96, SSU-0817/SSU-1536, EF4/EF3, EF4/fung5 and FR1/FF390) were examined for their applicability to the analysis of soil fungal communities using bioinformatic tools as well as real PCR systems. Virtual primer matching for EF4/EF3 and EF4/fung5 revealed good matching with zygomycetous, ascomycetous and basidiomycetous 18S rDNA database entries. Whereas primer EF4/EF3 had no cross matches in the rDNA databases for plant and invertebrate, primer EF4/fung5 gave one signal with the corresponding database. Similar results were obtained for the primer set SSU-0817/SSU-1536. Two matches with plant rDNAs and 22 or 12 matches with the invertebrate database could be identified for the primer sets SSU-0817/SSU-1196 and FR1/FF390, respectively. Primer pair NS1/NS2 showed only a 70% match with fungal 18S rDNA sequences, but a 75% to 90% match with non-fungal sequences. Alignments of 2000 eukaryotic sequences using “ARB” confirmed that PCR fragments obtained by the primer sets EF4/EF3, EF4/fung5, SSU-0817/SSU-1536 and FR1/FF390 were supposed to include hypervariable regions (V4, V7, V9), whereas the others included regions which were more phylogenetically conserved. Practical PCR approaches affirmed fungal specificity as predicted by virtual primer matching for EF4/EF3, EF4/fung5 and FR1/FF390. However FR1/FF390 amplified only 60% of the fungal samples under investigation. All other primer sets amplified fungal as well as non-fungal samples.

The β-glucuronidase (GUS) gene is a widely used reporter gene in transgenic research. This study shows that although histochemical localisation of GUS activity may be very specific, differences in incubation conditions and tissue status can lead to artificial localisations that are independent of gene activity. The objective of the current studies was to evaluate the factors that affect the in-situ localisation of β-glucuronidase using transgenic tobacco plants as model tissues. The aspects considered include tissue size as well as and the addition of surfactants, vacuum infiltration and chemical fixatives. Transgenic tobacco plants exhibited variable staining patterns dependent on the size of tissue assayed and the treatments that affected the infiltration of substrate. A gradient of blue staining was observed in larger tissue pieces (10 mm²), where staining in central areas was light blue in contrast to edges, which stained deep indigo. More intense staining was associated with peripheral cell layers and regions adjacent to leaf veins. Thinner tissue strips incubated under similar conditions exhibited intense and even X-Gluc staining. Addition of Triton X-100 (1%) surfactant and vacuum infiltration (2 min) produced considerably quicker and more uniform staining (intense and consistent indigo blue colour) of the examined tissue after a 4 to 6-h incubation. Chemical fixation of tissues before GUS assay resulted in quantitative and histochemical differences in enzyme activity that were dependent on the fixative type and duration. Quantitative measurements using the MUG fluorometric assay showed that Histochoice™ provided the highest retention of GUS activity, maintaining more than 80 and 50% of the activity after fixation for 15 and 30 min, respectively. Activity in decreasing order was obtained with paraformaldehyde, glutaraldehyde, ethanol and FAA. GUS activity was affected not only by the type of fixative, but also by the duration of fixation with longer fixation producing lower GUS activity. From the experiments performed it can be concluded that those treatments that enhance substrate penetration, i.e., the addition of surfactant and vacuum infiltration, improve the consistency and speed of X-Gluc staining.

The bactericidal effect of atmospheric gas plasma (AGP) on Escherichia coli K12 cells in 0.9% [w/v] sodium chloride was investigated under normal atmospheric pressure. A plasma-generating unit was supplied with working gas (air) at 25 °C and a relative humidity (RH) of 0, 14, 32, 43, 50, 60 or 70%. Gas plasma was generated using radio frequency discharge (20 kHz) under atmospheric pressure, and blown onto the surface of an E. coli cell suspension. Seven log-cycles of cells were completely inactivated within 15 minutes of the start of AGP treatments using working gas at 32, 43, 50 or 60% RH. AGP at 14% RH inactivated only 2 log-cycles of cells, but no inactivation was observed when air at 0% or 70% RH was used. The inactivation curves were biphasic and the rate constants for both stages were closely related to the RH of the working gas. The rate of cell inactivation was at its maximum at an RH of 43%. The bactericidal effect of the AGP treatment was not a result of ozone generation, however, the moisture content of the working gas was a significant factor in ensuring that E. coli K12 inactivation occurred. The interpretation of these data was that chemical species generated from the water molecules in the working gas were bactericidal in their effects.