Estimation of nitrogen removal rate in aqueous phase based on delta15N in microorganisms in solid phase.

Microbial nitrification and denitrification are important processes for removing nitrogenous compounds in aqueous systems. Nitrogen removal rate estimation is essential for controlling nitrogen removal processes and modeling the nitrogen cycle in ecosystems. The model described the relationship between ammonium removal rate (aqueous phase) and the nitrogen stable isotope ratio (delta15N) of microorganisms (solid phase) when a coupled nitrification-denitrification process occurs and assimilation and advections are maintained in a steady state. An oxidation ditch in a municipal wastewater treatment plant was evaluated for 3 years using the model. The ammonium removal rate was calculated from the data of delta15N of the activated sludge, it correlated significantly with the observed removal rate. The isotope fractionation factor (epsilon) was determined to be -5.5 per thousand by using a nonlinear method. The model and obtained factor value were applicable for standard activated-sludge processes performed in parallel in the oxidation ditch and a river watershed. The model may help illustrate nitrogen behavior in ecosystems.

Total RNA concentration as an index of microbial activity and oxygen supply in an oxidation ditch.

Total RNA and chromosomal DNA concentrations at a municipal wastewater treatment plant with an oxidation ditch (OD) were monitored for 1.5 years using commercial extraction kits for DNA and RNA. No parameters correlated with the chromosomal DNA concentration. The total RNA concentration exhibited better correlation than the solids retention time and the mixed liquor suspended solids with the removal rate of total organic carbon, and can be regarded as an index of microbial activity. The total RNA concentration varied with a cycle of one year and increased at lower water temperatures in this OD. When diffusion theory was taken into account, it was found that the oxygen dissolution rate increased at lower temperature, and a small change in the oxygen dissolution rate caused a large variation in microbial activity and also affected nitrification and denitrification. The information was insufficient to clarify the various reaction relationships, but total RNA concentration will likely be useful as an index of microbial activity in actual wastewater treatment reactors.

Exchange characteristics of monocarboxylic acids and monosulfonic acids onto anion-exchange resins.

In the previous paper (N. Kanazawa, K. Urano, N. Kokado, Y. Urushigawa, J. Colloid Interface Sci. 238 (2001) 196), the equilibria of propionic acid and benzoic acid adsorption onto three anion-exchange resins were investigated, and an equation was proposed that summed of the physical adsorption of the carboxylic acid molecule and the ion exchange of the dissociated carboxylate ion. The ion exchange equation, including a selectivity coefficient to chloride ion for each combination between carboxylate ions and anion-exchange resins, could be used in wide ranges of concentration and pH. In this research, ion-exchange equilibria using 16 anion-exchange resins and 9 organic acids including monocarboxylic and monosulfonic acids were investigated. It could be confirmed that the proposed equation applied to the ion exchange with these monoorganic acids. Characteristics of ion exchange between the organic anions and anion exchange were also studied by the selectivity coefficients.

Interference of contaminating DNA in the quantification of a toluene-induced tod gene in Pseudomonas putida.

To determine the extent of interference of co-extracted DNA contamination in the quantification of the tod gene transcript, two different concentrations of RNA (high, 500 ng/microl; low, 250 ng/microl) from a toluene-induced culture of Pseudomonas putida were treated with different amounts of DNase (2, 4, 6 and 8 U) and incubated for 30 and 60 min. The highly sensitive and reproducible TaqMan system was used to quantify the transcript of the tod gene, the tod gene in contaminating DNA and the 16S rRNA gene in DNase-treated RNA samples. For the high RNA concentration, the shorter incubation time (30 min) lowered the level of contaminating DNA as evidenced by the presence of 2.5 x 10(6) copies of the tod gene before treatment to 1.4 x 10(5) copies/microl (8 U), whereas, irrespective of the DNase units used, the longer incubation time (60 min) considerably lowered the level of DNA contamination (2.5 x 10(6) to 6.5 x 10(2) copies of the tod gene/microl). However, for the low RNA concentration, DNase treatment was found to be equally effective in lowering the level of contaminating DNA (10(6) to 10(2) copies of the tod gene/mu), irrespective of the incubation time and the amount of DNase used. Although the results of gel electrophoresis of conventional PCR amplification of the low RNA concentration revealed the absence of the target gene in contaminating DNA, the results of the TaqMan PCR indicated that a very low amount of contaminating DNA (less than 10(3) copies of the tod gene/mul) was still present in RNA samples, even after the DNase treatment. The number of copies of the tod gene transcript in RNA samples did not show any marked variation because of the DNase treatment. However, the proportion of contaminating DNA in RNA samples considerably decreased due to the treatment (0.01 to 0.000001). Furthermore, these results suggested that the extent of the removal of contaminating DNA from RNA samples depends on the concentration of RNA, the amount of DNase used and the incubation time. It is also suggested that the copies of the catabolic genes in contaminating DNA have to be quantified along with the target genes in RNA samples to have a more accurate quantification of the target genes for better understanding of their roles in many microbial processes.

Parallel characterization of anaerobic toluene- and ethylbenzene-degrading microbial consortia by PCR-denaturing gradient gel electrophoresis, RNA-DNA membrane hybridization, and DNA microarray technology.

A mesophilic toluene-degrading consortium (TDC) and an ethylbenzene-degrading consortium (EDC) were established under sulfate-reducing conditions. These consortia were first characterized by denaturing gradient gel electrophoresis (DGGE) fingerprinting of PCR-amplified 16S rRNA gene fragments, followed by sequencing. The sequences of the major bands (T-1 and E-2) belonging to TDC and EDC, respectively, were affiliated with the family Desulfobacteriaceae. Another major band from EDC (E-1) was related to an uncultured non-sulfate-reducing soil bacterium. Oligonucleotide probes specific for the 16S rRNAs of target organisms corresponding to T-1, E-1, and E-2 were designed, and hybridization conditions were optimized for two analytical formats, membrane and DNA microarray hybridization. Both formats were used to characterize the TDC and EDC, and the results of both were consistent with DGGE analysis. In order to assess the utility of the microarray format for analysis of environmental samples, oil-contaminated sediments from the coast of Kuwait were analyzed. The DNA microarray successfully detected bacterial nucleic acids from these samples, but probes targeting specific groups of sulfate-reducing bacteria did not give positive signals. The results of this study demonstrate the limitations and the potential utility of DNA microarrays for microbial community analysis.

Adsorption Equilibrium Equation of Carboxylic Acids on Anion-Exchange Resins in Water.

The adsorption of propionic acid and benzoic acid on anion-exchange resins was analyzed, and an adsorption equilibrium equation of carboxylic acids was proposed. The adsorption of carboxylic acids on the anion-exchange resins was considered to be the sum of the physical adsorption of the molecule and the ion-exchange adsorption of the ion, which were independent of each other. For the physical adsorption of carboxylic acids, it was conformed to the Freundlich equation. For the ion-exchange adsorption of carboxylate ions, the equilibrium equation corresponded well with the experimental results for wide ranges of concentration and pH. The equation contains a selectivity coefficient S(A)(Cl) for the chloride ion versus the carboxylate ion, which was considered essentially a constant. The influent of the bicarbonate ion from carbon dioxide in air could also be expressed by the additional equilibrium equation with the selectivity coefficient S(HCO(3))(Cl) for the chloride ion versus the bicarbonate ion. Consequently, an adsorption equilibrium equation can estimate the equilibrium adsorption amounts. Even the effect of a coexisting bicarbonate ion is inconsequential when the parameters of the Freundlich isotherm equation and the selectivity coefficients of the carboxylate ion and the bicarbonate ion in each resin are determined in advance. Copyright 2001 Academic Press.