Results Observed and estimated richness of the Archaea community in the activated sludge A 16S rRNA gene clone library was constructed from a sample

of activated sludge collected at the aeration tank of the Rya WWTP at a time of normal operating conditions. There were no atypical process parameter values or extreme events prior to sample collection. However, the F/M-ratio was higher at the time of the clone library sample collection (May 2007) compared with the times when samples were collected for FISH (December 2007) and T-RFLP analyses (May 2003 – August 2004) (Table 1). Cloning and sequencing generated 82 archaeal 16S rRNA gene sequences of lengths between 756 and 862 bases. Based on DNA check details similarity the sequences were assigned to operational taxonomic units (OTUs).

The sequences were assigned to OTUs corresponding to 25 species of 10 genera, 7 PX-478 manufacturer families/classes and 6 different phyla. The Archaea community richness was estimated to be at least 43 species of 19 different genera. Thus, the clone library covered at most 58% of the species and 53% of the genera present in the activated sludge. Accumulation curves (Figure  1) also illustrate that the clone library does not fully cover the Archaea community. Table 1 Comparison of WWTP parameters at the different sample collection times a Parameterb, c, d May 03 – Aug Captisol ic50 04e May 07 f Dec 07g Comment Temp b 15 ± 3 15 ± 1 11 ± 1 **   SRT b 3 ± 1 3 ± 0 2 ± 0   F/M b 0.008 ± 0.002 0.014 ± 0.004 ** 0.008 ± 0.002 Max value in May 2007 COD b 1058 ± 240 999 ± 194 1068 ±97±   NO23-N b 48 ± 8 46 ± 9 42 ± 22 Min and max values in Dec 07 SSVI c 80 ± 15 54 79   Effluent NSS c 23 ± 17 26 31   a The three periods were compared using the Kruskal-Wallis test. A statistically significant difference, p(same) < 0.05, is marked with asterisks (**). b Average values (± standard deviation) from all sample dates and the six days preceding

the sample dates. c Data only from sample dates, not including the six preceding days. d The parameters are water temperature (Temp, °C), solids retention time (SRT, days), food to mass ratio (F/M, g/kg*s), COD going into the Metalloexopeptidase activated sludge tanks (COD, g/s), nitrite/nitrate levels going in to the activated sludge tanks (NO23-N, g/s), standardized sludge volume index (SSVI, ml/g) and effluent non-settleable solids (Effluent NSS, mg/l). e Samples collected during this period were used for T-RFLP analysis. f A sample collected during this period was used for T-RFLP and clone library analysis. g A sample collected during this period was used for FISH analysis. Figure 1 Accumulation curves of archaeal 16S rRNA gene sequences. 82 archaeal 16S rRNA gene sequences were assigned to OTUs based on similarity thresholds representing the division in phylum (80%), family/class (90%), genus (95%) and species (98.7%) levels [23, 24].