Ammonia Oxidizer Enrichment Sarah Douglas Shelbi Russell Ammonia

Ammonia Oxidizer Enrichment Sarah Douglas Shelbi Russell

Ammonia oxidation NH 3+ NH 2 OH Amo. ABC Bacteria & Archaea NO 2 HAO Bacteria Archaeal pathway unknown, hydroxylamine may be oxidized by the nitroxyl pathway (Walker et al. , 2010)

Phylogenetic diversity of ammonia oxidizers Bacteria β-Proteobacteria (Nitrosomonas & Nitrospira) γ-Proteobacteria (Nitrosococcus) Crenarchaeota

Sampling Monterey Bay Aquarium activated sand 19°C and 20°C Hopkins sand by aquarium outflow and coast

Enrichment procedure • Crenarchaeota medium inoculated with 50 m. L of 10 m. M filtered slurry water sample – NH 4 Cl – Cofactors: • 7 -vitamin, selenite/tungstate, and SL 12 trace metal solution – Bicarbonate carbon source – Phosphate buffer • Store in the dark • Sample every 2 days for 2 weeks

Spectrophotometric determination of substrate and product concentration Hopkins Aquarium Sand 250 Concentration (u. M) 200 150 Aisle Ammonia Aisle Nitrite Window Ammonia 100 Window Nitrite 50 0 0 2 4 6 8 10 Days 12 14 16 18 20 22

Dead Seal Beach Sand 250 Concentration (u. M) 200 150 Dead Seal Beach Sand (Aisle) Ammonia Dead Seal Beach Sand (Aisle) Nitrite Dead Seal Beach (Window) Ammonia Dead Seal Beach (Window) Nitrite 100 50 0 0 2 4 6 8 10 Days 12 14 16 18 20 22

Monterey Bay Aquarium Sand 250 Concentration (u. M) 200 21 degrees C (Aisle) Ammonia 150 21 degrees C (Aisle) Nitrite 21 degrees C (Window) Ammonia 21 degrees C (Window) Nitrite 18 degrees C (Aisle) Ammonia 100 18 degrees C (Aisle) Nitrite 18 degrees C (Window) Ammonia 18 degrees C (Window) Nitrite 50 0 0 2 4 6 8 10 Days 12 14 16 18 20 22

Monterey Bay Aquarium Bradley’s experiment Added 200 u. M Ammonia 200 180 KJ - Nitrite Concentration (u. M) 160 KJ - Ammonia 140 BS - Nitrite 120 BS - Ammonia 100 80 BS Add - Nitrite 60 BS Add Ammonia BS Trans Nitrite BS Trans Ammonia 40 20 0 0 2 4 6 8 10 12 14 Time (days) 15 17 19 21

PCR amplification of bacterial and archaeal amo. A AOA amo. A AOB amo. A Monterey Bay Aquarium sand 21° Monterey Bay Aquarium sand 18° Hopkins beach sand Hopkins aquarium sand Positive control

Conclusion • Bacterial ammonia oxidizers appear to be more prevalent • Archaea at competitive disadvantage in high ammonia environments? • Lack of correlation between ammonia depletion and nitrite generation: complex ecosystem?