The role of trace metal micronutrients in the growth and niche separation of marine ammonia-oxidising microorganisms
Keywords: iron
trace metals
microbial ecology
marine microorganisms
oceanography
archaea
marine biogeochemistry
copper

Nitrogen is essential to life on Earth. In the oceans, the availability and chemical speciation of nitrogen constrains how effectively CO2 is removed from the atmosphere by phytoplankton and delivered to the deep ocean –– with implications for global climate. Nitrogen availability and speciation are controlled by marine microorganisms, with ammonia-oxidising archaea and bacteria (AOA and AOB) mediating the first step in the oxidative nitrogen cycle. A scarcity of trace metals (such as iron (Fe) and copper (Cu)) has been repeatedly shown to limit other components of microbial nitrogen cycling in oceans. Yet, the role of trace metals in AOA and AOB growth remains largely underexplored. The research presented here brings together microbial physiology, metallomics, proteomics and comparative genomics to ask whether there is physiological precedent for iron and copper availability to govern the growth, distribution and niche separation of AOA and AOB in the oceans. We found that growth rate of the marine AOA isolate, Nitrosopumilus maritimus, in culture is determined by Fe availability. N. maritimus displays an unchelated iron (Fe´) half-saturation constant 1–2 orders of magnitude greater for growth than numerous marine phytoplankton and heterotrophic bacterial species –– driven by a reduced uptake affinity for Fe´. When growth is limited by Fe´, proteomic analysis revealed that N. maritimus primarily minimises Fe demand by reducing Fe-dense ferredoxins which putatively interact with Complex I to generate the reduced co-factor NAD(P)H –– potentially functionally substituting these ferredoxins for Fe-free alternatives. In doing so, Fe-binding proteins which play a more central role in electron transfer and ammonia oxidation can be preferentially retained, allowing N. maritimus to continue in supporting key energy-yielding pathways under Fe limitation. Marine AOA and AOB isolates were found to display contrasting requirements and uptake strategies for Fe and Cu at the genetic, proteomic, and physiological level. We found that the AOB isolate, Nitrosococcus oceani displays a greater affinity for Fe´ but requires greater cupric Cu (Cu2+) for growth compared with the marine AOA isolate, N. maritimus. We predict that this may allow AOB to inhabit shallower, euphotic waters where NH4+ supply is high, but competition for Fe is rife. In contrast to AOB, AOA isolates have a greater affinity and toxicity threshold for Cu2+ providing an additional explanation to the greater success of AOA in the marine environment where Cu2+ availability can be highly variable. These data are used to develop the testable hypothesis, with support from comparative genomics and biogeochemical datasets, that Cu2+ widely limits AOA and AOB growth in the oceans, particularly in the Pacific Ocean where Cu2+ concentrations are often below that needed by AOA and AOB isolates for growth. Further, Fe and Cu ecophysiologies are hypothesised to contribute to the observed depth distribution of AOA and AOB in the oceans –– with contrasting trophic strategies for Fe and Cu acquisition potentially allowing AOA and AOB to occupy distinct trace metal niches in the oceans. This may explain the chemical niche that allows AOB to persist in the oceans, despite being uncompetitive for NH4+ compared with AOA. The research presented here develops an improved understanding of the physicochemical factors that govern ammonia oxidation in the marine environment –– with far-reaching implications for the current, past and future biogeochemical nitrogen and carbon cycles.