Bacteria that can oxidize both ammonia to nitrite and nitrite to nitrate seem to be better adapted to ammonia-limited environments than most cultured microbes that oxidize ammonia to nitrite only, contrary to expectations. See Letter p.269
The oxidation of ammonia to nitrite and of nitrite to nitrate, known as nitrification, is a central part of the nitrogen cycle, which is a fundamental biogeochemical process. Competition for limited amounts of ammonia is a major factor in shaping the complex communities of bacteria and archaea responsible for nitrification. It has been thought that ammonia-oxidizing archaea (AOA) outcompete ammonia-oxidizing bacteria (AOB) in environments that have low (submicromolar) concentrations of ammonia1,2. But on page 269, Kits et al.3 show that an unusual bacterium called Nitrospira inopinata, which can perform both steps of nitrification4,5,6, is better adapted to low-ammonia conditions than are most cultured AOA species.
Ammonia is formed during the breakdown of organic nitrogen, and is rare in most environments, but ammonia oxidizers are abundant in nature. At first, these observations seem difficult to reconcile. However, the apparent rarity of ammonia can be attributed to the ability of ammonia oxidizers that have high ammonia affinity to scoop up even tiny scraps of ammonia. For example, AOA that live in the ocean grow on almost undetectable amounts of ammonia by tuning their cellular machinery — including the key ammonia-oxidizing enzyme ammonia monooxygenase — to extremely low substrate concentrations (less than 10 nM)1,7,8. It has been thought that terrestrial AOA, like their marine relatives, have a high affinity for ammonia and so could outcompete AOBs under low ammonia concentrations2,9,10. But ammonia-affinity measurements for pure cultures of these archaea have been lacking.
Kits et al. isolated a pure culture of N. inopinata from the mixed culture in which it was previously identified4. They then determined the nitrification kinetics for this species and for four pure cultures of terrestrial AOA isolated from soils and hot springs (Fig. 1). The authors found that these cultured AOA had surprisingly poor ammonia affinities (about 0.4–6 μM; affinity is typically expressed as the substrate concentration at which the reaction rate is half the maximum rate), similar to those of cultured AOB. By contrast, N. inopinata exhibited a higher affinity for ammonia than most AOA and all other cultured AOB (about 60 nM).
Although N. inopinata is closely related to Nitrospira species that oxidize only nitrite, the researchers found that it had a poor affinity for nitrite (about 500 μM). This poor affinity probably prevents it from growing as a pure nitrite oxidizer in most environments, where nitrite concentrations are typically low. When N. inopinata grows in environments suited to its role as an ammonia oxidizer, nitrite can accumulate intracellularly, thus overcoming this apparent shortcoming.
Kits and colleagues next showed that N. inopinata has a higher growth yield per molecule of ammonia oxidized than cultured AOA or AOB. This confirms theoretical predictions11 positing that growth yield would be more efficient owing to the biochemical pathway involved in complete nitrification, which produces more energy-carrying ATP molecules than does a single oxidation step. Kits et al. suggest that the efficient carbon-fixation pathway of N. inopinata — by which carbon dioxide is assimilated into cellular biomass — could also contribute to the high yield.
Together, the authors' data change our view of the potential role of complete ammonia-oxidizers for nitrification. Rather than being rare organisms inhabiting peripheral habitats, it turns out that they are well adapted to the low ammonia concentrations that characterize most of the world's biosphere.
Both its superior ammonia affinity and growth yield could explain why N. inopinata — along with other complete ammonia-oxidizing (comammox) Nitrospira species that perform both steps of nitrification — is abundant in ammonia-depleted groundwaters, drinking-water treatment systems and some soils12. However, other ammonia-depleted environments such as forest soils and the ocean are dominated by AOA and AOB1,12,13. If comammox species have a higher affinity for ammonia than AOA and AOB, why would this be the case? It could be that the affinities and growth yields of ammonia oxidizers in these environments are actually higher than has been documented for their cultivated relatives. Alternatively, factors other than ammonia affinity and growth yield could determine the composition of ammonia-oxidizing communities.
Aerobic ammonia oxidation is an ancient trait, probably dating back more than 2 billion years. Both bacterial and archaeal ammonia oxidizers have therefore had plenty of time to evolve. 'Horizontal' transfer of genes encoding high-affinity ammonia monooxygenases between species could rapidly change the ammonia affinity of an ammonia oxidizer from poor to high, making it difficult to imagine that a single trait such as high ammonia affinity can lead to the dominance of any given group of ammonia oxidizers in an environment.
That horizontal gene transfer does occur has been shown by analyses4,14 of the genomes of comammox Nitrospira suggesting that the microbes' ammonia-oxidizing pathway was either transferred to or obtained from betaproteobacterial AOB (β-AOB). Cultured β-AOB have a lower ammonia affinity than N. inopinata, making this a puzzling observation at first sight. However, an uncultured β-AOB with a high ammonia affinity might exist, or the enzymes in each species might have evolved different affinities after horizontal gene transfer.
The apparent dominance of AOA over AOB in the ocean is probably the result of a combination of factors, in addition to high ammonia affinity. These might include: their ability to efficiently use organic nitrogen instead of ammonia; the use of copper as a major cofactor for catalytic processes other than ammonia oxidation, instead of the less-abundant iron used by bacteria; and the small size of AOA, which means that less ammonia is required to double their cellular biomass.
So far, only a few ecosystems have been screened for the abundance of comammox bacteria, but the current study demonstrates that these microbes are probably particularly common in low-ammonia environments. There is now an urgent need to determine their contribution to nitrification in other ecosystems. Their abundance in some ammonia-depleted environments is probably due not only to their high affinity for ammonia, but also to factors such as their ability to grow on substrates other than ammonia and their low energy requirements.
Such variables might have general roles in shaping communities of ammonia-oxidizing microbes. Identifying which of these are most important will require greater insight into the physiology of AOA, AOB and comammox bacteria, and a more profound understanding of the ecology and evolution of these intriguing organisms.
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