Autotrophic ammonia-oxidising bacteria (AOB) are a crucial component of the microbial communities of nitrifying wastewater treatment systems. Nitrification is known to occur in reactors of different configuration, but whether AOB communities are different in reactors of different design is unknown. We compared the diversity and community structure of the betaproteobacterial AOB in two full-scale treatment reactors - a biological aerated filter (BAF) and a trickling filter - receiving the same wastewater. Polymerase chain reaction (PCR) of 16S ribosomal RNA (rRNA) gene fragments with AOB-selective primers was combined with denaturing gradient gel electrophoresis (DGGE) to allow comparative analysis of the dominant AOB populations. The phylogenetic affiliation of the dominant AOB was determined by cloning and sequencing PCR-amplified 16S rRNA gene fragments. DGGE profiles were compared using a probability-based similarity index (Raup and Crick). The use of a probability-based index of similarity allowed us to evaluate if the differences and similarities observed in AOB community structure in different samples were statistically significant or could be accounted for by chance matching of bands in DGGE profiles, which would suggest random colonisation of the reactors by different AOB. The community structure of AOB was different in different sections of each of the reactors and differences were also noted between the reactors. All AOB-like sequences identified, grouped within the genus Nitrosomonas. A greater diversity of AOB was detected in the trickling filters than in the BAF though all samples analysed appeared to be dominated by AOB most closely related to Nitrosococcus mobilis. Numerical analysis of DGGE profiles indicated that the AOB communities in depth profiles from the filter beds were selected in a non-random manner.

The diversity and community structure of the beta-proteobacterial ammonia oxidising bacteria (AOB) in a range of different lab-scale industrial wastewater treatment reactors were compared. Three of the reactors treat waste from mixed domestic and industrial sources whereas the other reactor treats waste solely of industrial origin. PCR with AOB selective primers was combined with denaturing gradient ge electrophoresis to allow comparative analysis of the dominant AOB populations and the phylogenetic affiliation of the dominant AOB was determined by cloning and sequencing or direct sequencing of bands excised from DGGE gels. Different AOB were found within and between different reactors. All AOB sequences identified were grouped within the genus Nitrosomonas. Within the lab-scale reactors there appeared to be selection for a low diversity of AOB and predominance of a single AOB population. Furthermore, the industrial input in both effluents apparently selected for salt tolerant AOB, most closely related to Nitrosococcus mobilis and Nitrosomonas halophila.

Direct toxicity assessment of wastewater is becoming necessary, and new legislation may render it compulsory for the water industry. At present such assessment is performed at a laboratory away from a site, at considerable cost, and results often come too late, after a toxic event has occurred and the toxin has been released into the environment. Some of the rapid toxicity tests available today require certain conditions to function properly, or their results do not always correlate with other methods. The objective of this study was to assess a portable device, the Baroxymeter, for its suitability as an instrument to test wastewater toxicity. The way the device works is based on monitoring respiration of a bacterial culture by pressure measurements and using respiration inhibition as a toxicity alert. It has been shown that it is possible to detect toxic substances such as 3,5-dichlorophenol and bronopol within 5 min from a 1-mL sample. The benefits and future applications of the Baroxymeter as a high-throughput, cost-effective alternative for toxicity screening are discussed in this article.

A sandwich hybridization assay for high-throughput, rapid, simple, and inexpensive quantification of specific microbial populations was evaluated. The assay is based on the hybridization of a target rRNA with differentially labeled capture and detector probes. Betaproteobacterial ammonia-oxidizing bacteria (AOB) were selected as the target group for the study, since they represent a phylogenetically coherent group of organisms that perform a well-defined geochemical function in natural and engineered environments. Reagent concentrations, probe combinations, and washing, blocking, and hybridization conditions were optimized to improve signal and reduce background. The detection limits for the optimized RNA assay were equivalent to approximately 10 3 to 10 4 and 10 4 to 10 5 bacterial cells, respectively, for E. coli rRNA and RNA extracted from activated sludge, by using probes targeting the majority of bacteria. Furthermore, the RNA assay had good specificity, permitted discrimination of rRNA sequences that differed by a 2-bp mismatch in the probe target region, and could distinguish the sizes of AOB populations in nitrifying and nonnitrifying wastewater treatment plants.Quantification of specific bacterial populations is key to understanding the mechanisms that underlie many biologically mediated processes, including nitrification. The use of rRNAbased cultivation-independent techniques may be one potentially useful means to do this. Numerous rRNA-based techniques are now commonly employed to quantify microbial communities in environmental samples. These include fluorescence in situ hybridization (FISH) (see, e.g., references 11, 12, 17, 27, 37, and 40), quantitative dot blot hybridization (see, e.g., references 14 and 34), PCR-based methods (e.g., real time PCR [19] and competitive PCR [15,33]), and microarray technology (see, e.g., references 1, 18, and 38).Each of these methods has particular advantages and disadvantages. For example, FISH allows direct identification and quantification of microorganisms in environmental samples and, because of its ability to measure individuals accurately, represents the "gold standard." However, FISH has a low sample throughput and is thus too slow for routine analysis. In contrast, PCR-based methods allow high sample throughput, which may be required for statistically valid spatial and temporal analysis. However, PCR-based methods are prone to biases (typically introduced during nucleic acid extraction and amplification) that may make quantification of natural bacterial populations difficult (see, e.g., references 20, 39, 39a, and 43). In addition, relatively pure preparations of DNA are required to avoid problems of inhibition of DNA polymerases. Quantitative dot blots and microarrays, on the other hand, often require the use of radioactive isotopes or expensive equipment and consumables.An alternative sensitive and specific method is a sandwich hybridization assay for the detection of rRNA, developed by Wicks and coworkers (46). The RNA assay is based on hybridization of...

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