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Precursors to Modern Aerobic Ammonium Oxidation: Oxygenic and Electrogenic Anaerobic Pathways

Periodic Reporting for period 2 - PAERADOX (Precursors to Modern Aerobic Ammonium Oxidation: Oxygenic and Electrogenic Anaerobic Pathways)

Reporting period: 2020-08-01 to 2021-07-31

Earth has undergone dramatic changes since the start of life, and among them, the rise of oxygen by Cyanobacteria greatly influenced the evolution of life. The appearance of oxygen-dependent (aerobic) metabolisms shaped the modern biosphere, as oxygen breathing is the dominant metabolism on modern Earth. On the other hand, oxygen-independent (anaerobic) metabolisms are still diverse and even persistent in the genomes of the strictly oxygen-dependent microbes. Can the evolutionary history of microbial metabolisms on our planet be uncovered by research on such persistent oxygen-independent metabolism in modern aerobic microbes?
PARAEDOX aims to discover the novel anaerobic microbial metabolisms and the story of ancient microbial metabolisms on Earth. Research on the origins and evolution of microbial metabolisms can ultimately help us explain how and when life co-evolved with biogeochemical cycles and may help to evaluate the state of evolution on other planets. We trace unidentified ancient anaerobic microbial pathways associated with ammonia oxidation and reconstruct the evolutionary history of those metabolisms.
To discover novel anaerobic metabolisms, we have mined all microbial genomes in the tree of ammonia oxidizers for anticipated pathways that were selected as the model for tracing the evolutionary origins and history of oxygen-dependent metabolisms. In particular, Nitrosomonas and Nitrosococcus spp. were selected to assess the presence and activity of anaerobic extracellular electron transfer coupled to ammonia oxidation and nitrate reduction coupled to ammonia oxidation, respectively. Careful physiological assessments based on advanced incubations, isotopic tracking, meta-transcriptomics, and cytochrome-staining coupled to electron microscopy evidenced the activity of the anticipated anaerobic metabolisms for the first time. The presence of such metabolisms in strictly aerobic ammonia oxidizers was striking and raised the question of why and how did such metabolisms persist in aerobes? The molecular phylogenetics aligned with paleogeochemical evidence tackled this question and revealed that extracellular electron transfer proteins in beta-proteobacterial ammonia oxidizers were acquired by gene transfer from gamma-proteobacteria during oxygen scarcity. This first study explained how beta-proteobacterial ammonia oxidizers have been coped with oxygen stress and survived under oxygen deprivation. These findings shed new light on the evolution of microbial metabolisms. Moreover, a complementary metagenomic and metatranscriptomic study revealed the ecological significance of the extracellular electron transfer metabolism in oxygen minimum zones.To discover novel anaerobic metabolisms, we have mined all microbial genomes in the tree of ammonia oxidizers for anticipated pathways that were selected as the model for tracing the evolutionary origins and history of oxygen-dependent metabolisms. In particular, Nitrosomonas and Nitrosococcus spp. were selected to assess the presence and activity of anaerobic extracellular electron transfer coupled to ammonia oxidation and nitrate reduction coupled to ammonia oxidation, respectively. Careful physiological assessments based on advanced incubations, isotopic tracking, meta-transcriptomics, and cytochrome-staining coupled to electron microscopy evidenced the activity of the anticipated anaerobic metabolisms for the first time. The presence of such metabolisms in strictly aerobic ammonia oxidizers was striking and raised the question of why and how did such metabolisms persist in aerobes? The molecular phylogenetics aligned with paleogeochemical evidence tackled this question and revealed that extracellular electron transfer proteins in beta-proteobacterial ammonia oxidizers were acquired by gene transfer from gamma-proteobacteria during oxygen scarcity. This first study explained how beta-proteobacterial ammonia oxidizers have been coped with oxygen stress and survived under oxygen deprivation. These findings shed new light on the evolution of microbial metabolisms. Moreover, a complementary metagenomic and metatranscriptomic study revealed the ecological significance of the extracellular electron transfer metabolism in oxygen minimum zones.
In summary, the novelties of the PARAEDOX are as follows:
• Provides direct evidence for the metabolisms of extracellular electron transfer and nitrite reduction in aerobic ammonia oxidizers, as the previously unrecognized mechanisms in the global N cycle
• Identifies the origins and the evolution of ammonia-oxidizing bacteria and their extracellular electron transfer metabolism
• Identifies the ecological significance of extracellular electron transfer in oxygen minimum zones.
• Constructs a novel research program merging microbial physiology and microbial phylogenomics
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