Effect of increasing salinity to adapted and non-adapted Anammox biofilms.

The Anammox process is an efficient low energy alternative for the elimination of nitrogen from wastewater. The process is already in use for side stream applications. However, some industrial wastewaters, e.g. from textile industry are highly saline. This may be a limit for the application of the Anammox process. The aim of this study was to evaluate the effects of different NaCl concentrations on the efficiency of adapted and non-adapted Anammox biofilms. The tested NaCl concentrations ranged from 0 to 50 g NaCl*L-1. Concentrations below 30 g NaCl*L-1 did not significantly result in different nitrogen removal rates between adapted and non-adapted bacteria. However, adapted bacteria were significantly more resilient to salt at higher concentrations (40 and 50 g NaCl*L-1). The IC50 for adapted and non-adapted Anammox bacteria were 19.99 and 20.30 g NaCl*L-1, respectively. Whereas adapted biomass depletes the nitrogen in ratios of NO2- / NH4+ around 1.20 indicating a mainly Anammox-driven consumption of the nitrogen, the ratio increases to 2.21 at 40 g NaCl*L-1 for non-adapted biomass. This indicates an increase of other processes like denitrification. At lower NaCL concentrations up to 10 g NaCl*L-1, a stimulating effect of NaCl to the Anammox process has been observed.

Autofluorescent characteristics of Candidatus Brocadia fulgida and the consequences for FISH and microscopic detection.

An enrichment culture of Candidatus Brocadia fulgida was identified by three independent methods: analysis of autofluorescence using different microscope filter blocks and a fluorescence spectrometer, fluorescence in situ hybridization (FISH) with anammox-specific probes and partial sequencing of the 16S rDNA, hydrazine synthase hzsA and hydrazine oxidoreductase hzo. The filter block BV-2A (400-440, 470 LP, Nikon) was suitable for preliminary detection of Ca. B. fulgida. An excitation-emission matrix revealed three pairs of excitation-emission maxima: 288-330 nm, 288-478 nm and 417-478 nm. Several autofluorescent cell clusters could not be stained with DAPI or by FISH, suggesting empty but intact cells (ghost cells) or inhibited permeability. Successful staining of autofluorescent cells with the FISH probes Ban162 and Bfu613, even at higher formamide concentrations, suggested insufficient specificity of Ban162. Under certain conditions, Ca. B. fulgida lost its autofluorescence, which reduced the reliability of autofluorescence for identification and detection. Non-fluorescent Ca. Brocadia cells could not be stained with Ban162, but with Bfu613 at higher formamide concentrations, suggesting a dependency between both parameters. The phylogenetic analysis showed only good taxonomical clustering of the 16S rDNA and hzsA. In conclusion, careful consideration of autofluorescent characteristics is recommended when analysing and presenting FISH observations of Ca. B. fulgida to avoid misinterpretations and misidentifications.

Highly efficient long-term storage of carrier-bound anammox biomass.

The anammox process is a potential alternative to the conventional nitrogen removal from wastewater. However, due to large generation times of anammox bacteria, the start-up of treatment reactors may be impeded. An efficient storage technique can handle this drawback and may be also suitable for seasonally operated treatment plants like in touristic areas. In the current study, several storage techniques were investigated with respect to its suitability for the preservation of the specific anammox activity after long-term storage. Storing conditions differed in terms of temperature, redox buffer and nutrient supplementation. The specific activity of immobilized anammox bacteria (Candidatus Kuenenia stuttgartiensis) was determined three times during a long-term preservation of 78 days and 106 days, respectively. The highest activity was ensured at a storing temperature of 4 °C, providing nitrate as redox buffer and a nutrient supplement every 23 days. Thus, 91.4% of the initial anammox activity could be preserved after a storage of 106 days. Superiority of the presented treatment condition was confirmed by a calculated nitrate-ammonium consumption rate close to the optimal ratio of 1.32. This technique provided an economical and simple method suitable for long-term storage of immobilized anammox biomass.