1. Barak Vasker ID 036966281 Marine Biotechnology Keuter et al. 2011

2. Introduction  Recirculation Aquaculture Systems (RAS) in which waste of aquaculture is processed. Water and “safe” nutrients re-used.  In the biofilter, dense biofilm will be created on carrier elements (“moving bed”), mostly comprising of: Ammonium Oxidizing Bacteria (AOB) & Nitrite Oxidizing Bacteria (NOB) working together as a pair.

3. Introduction  Nitrospira sp. Is a curved rod. Usually single celled, but also capable of colonization (biofilm). Non-obligatory autotrophs.  NH 4 +  NO 2 -  NO 3 (Requires aerobic atmosphere).  Dominant NOB, optimal activity and mechanism was not well understood, in part due to low energy yields of Ammonia oxidation and even lower yields on Nitrite oxidation (total work ~ 6 years (!)).

4. Introduction  Chemotaxonomic identification of Nitrospira sp. (Lipski 2001) via FAME (Fatty Acid Methyl Ester) Markers: 1)16:1 cis 7 2)16:1 cis 11

5. Results  Nitrospira cells were repeatedly diluted, enriched and transferred to fresh medium. This process repeated several times, until a new strain was isolated: “Ecomares 2.1”. (even pure, still secrets a tough polymer)  16S rRNA sequences clones were retrieved (1502 bp).  13 C-bicarbonate labeled fatty acids shows Nitrospira’s role.

6. Results  What about the 16S rRNA?  Ecomares 2.1 (isolated in Büsum, Germany) was found to match at most 99.1 % of the sequence of Nitrospira in Rehovot, Israel.  All strains showed are a part of cluster IV.

7. Results  Nitrospira colonies (circled) & nitrosomonas (marked “X”) colonies function as a pair, with each one enjoying the other’s produce. (mutualism?)

8. Discussion  High Nitrogenic load should have selected Nitrobacter (or other proteobacteria), consistent with recent strategy theories. How can Nitrospira dominate the RAS? Lab procedures/conditions. Theory / de-facto not always consistent.  Who is the most durable strain to high nitrite concentrations? Could this be enough to establish dominance? Pure Ecomares 2.1 still coated with thick, tough polymer.

9. Discussion  Selective Shear forces inside RAS? A stronger adherence of Nitrospira might be the final blow.  What is happening in other marine RAS? 16S rRNA sequences from other marine RAS suggest Nitrospira dominance.

10. Conclusions  Nitrospira-like bacteria dominates RAS biofilters, in contrast to what previously thought. Nitrospira characteristics critical for Marine aquaculture systems.  Nitrospira is supported by Nitrosomonas in an AOB/NOB biofilm.  Nitrospira Ecomares 2.1 is the most resisting strain to nitrite peaks in RAS. Therefore, it’s favorable in establishing dominance as the major Nitrite Oxidizing Bacteria (NOB) of the Biofilm.

11. Conclusions  Physiology is still under-investigated. Further research will reveal metabolic pathways, inhibatory effects & optimization of the process. We might be able to get different bacteria strains growing in different conditions.

12. Bibliography  Amann, R.I., Ludwig, W., and Schleifer, K.H. (1995) Phylogenetic identification and in situ detection of individual microbial cells without cultivation. Microbiol Rev 59: 143– 169.  Camargo, J.A., Alonso, A., and Salamanca, A. (2005) Nitrate toxicity to aquatic animals: a review with new data for freshwater invertebrates. Chemosphere 58: 1255–1267.  Daims, H., Nielsen, J.L., Nielsen, P.H., Schleifer, K.H., and Wagner, M. (2001) In situ characterization of Nitrospira-like nitrite-oxidizing bacteria active in wastewater treatment plants. Appl Environ Microbiol 67: 5273–5284  Knief, C., Altendorf, K., and Lipski, A. (2003) Linking autotrophic activity in environmental samples with specific bacterial taxa by detection of 13C-labelled fatty acids. Environ Microbiol 5: 1155–1167.

13. Thank you !