Rui Zhao’s research while affiliated with Massachusetts Institute of Technology and other places

Anammox bacteria inhabiting oxygen-deficient zones (ODZs) are a major functional group mediating fixed nitrogen loss in the global ocean. However, many basic questions regarding the diversity, broad metabolisms, origin, and adaptive mechanisms of ODZ anammox bacteria remain unaddressed. Here we report two novel metagenome-assembled genomes of anammox bacteria affiliated with the Scalindua genus, which represent most, if not all, of the anammox bacteria in the global ODZs. Metagenomic read-recruiting and comparison with historical data show that they are ubiquitously present in all three major ODZs. Beyond the core anammox metabolism, both organisms contain cyanase, and the more dominant one encodes a urease, indicating most ODZ anammox bacteria can utilize cyanate and urea in addition to ammonium. Molecular clock analysis suggests that the evolutionary radiation of these bacteria into ODZs occurred no earlier than 310 million years ago, ~1 billion years after the emergence of the earliest modern-type ODZs. Different strains of the ODZ Scalindua species are also found in benthic sediments, and the first ODZ Scalindua is likely derived from the benthos. Compared to benthic strains of the same clade, ODZ Scalindua uniquely encodes genes for urea utilization but has lost genes related to growth arrest, flagellum synthesis, and chemotaxis, presumably for adaptation to thrive in the global ODZ waters. Our findings expand the known metabolisms and evolutionary history of the bacteria controlling the global nitrogen budget.

Nitrite-oxidizing bacteria (NOB) are important nitrifiers whose activity regulates the availability of nitrite and dictates the magnitude of nitrogen loss in ecosystems. In oxic marine sediments, ammonia-oxidizing archaea (AOA) and NOB together catalyze the oxidation of ammonium to nitrate, but the abundance ratios of AOA to canonical NOB in some cores are significantly higher than the theoretical ratio range predicted from physiological traits of AOA and NOB characterized under realistic ocean conditions, indicating that some NOBs are yet to be discovered. Here we report a bacterial phylum Candidatus Nitrosediminicolota, members of which are more abundant than canonical NOBs and are widespread across global oligotrophic sediments. Ca. Nitrosediminicolota members have the functional potential to oxidize nitrite, in addition to other accessory functions such as urea hydrolysis and thiosulfate reduction. While one recovered species (Ca. Nitrosediminicola aerophilus) is generally confined within the oxic zone, another (Ca. Nitrosediminicola anaerotolerans) additionally appears in anoxic sediments. Counting Ca. Nitrosediminicolota as a nitrite-oxidizer helps to resolve the apparent abundance imbalance between AOA and NOB in oxic marine sediments, and thus its activity may exert controls on the nitrite budget.

Archaea belonging to the DPANN (Diapherotrites, Parvarchaeota, Aenigmarchaeota, Nanoarchaeota, and Nanohaloarchaeota) superphylum have been found in an expanding number of environments and perform a variety of biogeochemical roles, including contributing to carbon, sulfur, and nitrogen cycling. Generally characterized by ultrasmall cell sizes and reduced genomes, DPANN archaea may form mutualistic, commensal, or parasitic interactions with various archaeal and bacterial hosts, influencing the ecology and functioning of microbial communities. While DPANN archaea reportedly comprise a sizeable fraction of the archaeal community within marine oxygen-deficient zone (ODZ) water columns, little is known about their metabolic capabilities in these ecosystems. We report 33 novel metagenome-assembled genomes (MAGs) belonging to the DPANN phyla Nanoarchaeota, Pacearchaeota, Woesearchaeota, Undinarchaeota, Iainarchaeota, and SpSt-1190 from pelagic ODZs in the Eastern Tropical North Pacific and the Arabian Sea. We find these archaea to be permanent, stable residents of all three major ODZs only within anoxic depths, comprising up to 1% of the total microbial community and up to 25%–50% of archaea as estimated from read mapping to MAGs. ODZ DPANN appear to be capable of diverse metabolic functions, including fermentation, organic carbon scavenging, and the cycling of sulfur, hydrogen, and methane. Within a majority of ODZ DPANN, we identify a gene homologous to nitrous oxide reductase. Modeling analyses indicate the feasibility of a nitrous oxide reduction metabolism for host-attached symbionts, and the small genome sizes and reduced metabolic capabilities of most DPANN MAGs suggest host-associated lifestyles within ODZs. IMPORTANCE Archaea from the DPANN (Diapherotrites, Parvarchaeota, Aenigmarchaeota, Nanoarchaeota, and Nanohaloarchaeota) superphylum have diverse metabolic capabilities and participate in multiple biogeochemical cycles. While metagenomics and enrichments have revealed that many DPANN are characterized by ultrasmall genomes, few biosynthetic genes, and episymbiotic lifestyles, much remains unknown about their biology. We report 33 new DPANN metagenome-assembled genomes originating from the three global marine oxygen-deficient zones (ODZs), the first from these regions. We survey DPANN abundance and distribution within the ODZ water column, investigate their biosynthetic capabilities, and report potential roles in the cycling of organic carbon, methane, and nitrogen. We test the hypothesis that nitrous oxide reductases found within several ODZ DPANN genomes may enable ultrasmall episymbionts to serve as nitrous oxide consumers when attached to a host nitrous oxide producer. Our results indicate DPANN archaea as ubiquitous residents within the anoxic core of ODZs with the potential to produce or consume key compounds.

Anammox bacteria inhabiting oxygen deficient zones (ODZs) are a major functional group mediating fixed nitrogen loss and thus exerting a critical control on the nitrogen budget in the global ocean. However, the diversity, origin, and broad metabolisms of ODZ anammox bacteria remain unknown. Here we report two novel metagenome-assembled genomes of Scalindua , which represent most, if not all, of the anammox bacteria in the global ODZs. Beyond the core anammox metabolism, both organisms contain cyanase and the more dominant one encodes a urease, indicating ODZ anammox bacteria can utilize cyanate and urea in addition to ammonium. The first ODZ Scalindua likely derived from the benthos ∼200 million years ago. Compared to benthic strains of the same clade, ODZ Scalindua uniquely encode genes for urea utilization but lost genes related to growth arrest, flagellum synthesis, and chemotaxis, presumably for adaptation to the anoxic water column.

Archaea belonging to the DPANN superphylum have been found within an expanding number of environments and perform a variety of biogeochemical roles, including contributing to carbon, sulfur, and nitrogen cycling. Generally characterized by ultrasmall cell sizes and reduced genomes, DPANN archaea may form mutualistic, commensal, or parasitic interactions with various archaeal and bacterial hosts, influencing the ecology and functioning of microbial communities. While DPANN archaea reportedly comprise 15–26% of the archaeal community within marine oxygen deficient zone (ODZ) water columns, little is known about their metabolic capabilities in these ecosystems. We report 33 novel metagenome-assembled genomes belonging to DPANN phyla Nanoarchaeota, Pacearchaeota, Woesarchaeota, Undinarchaeota, Iainarchaeota, and SpSt-1190 from pelagic ODZs in the Eastern Tropical North Pacific and Arabian Sea. We find these archaea to be permanent, stable residents of all 3 major ODZs only within anoxic depths, comprising up to 1% of the total microbial community and up to 25–50% of archaea. ODZ DPANN appear capable of diverse metabolic functions, including fermentation, organic carbon scavenging, and the cycling of sulfur, hydrogen, and methane. Within a majority of ODZ DPANN, we identify a gene homologous to nitrous oxide reductase. Modeling analyses indicate the feasibility of a nitrous oxide reduction metabolism for host-attached symbionts, and the small genome sizes and reduced metabolic capabilities of most DPANN MAGs suggest host-associated lifestyles within ODZs. Importance Archaea from the DPANN superphylum have diverse metabolic capabilities and participate in multiple biogeochemical cycles. While metagenomics and enrichments have revealed that many DPANN are characterized by ultrasmall genomes, few biosynthetic genes, and episymbiotic lifestyles, much remains unknown about their biology. We report 33 new DPANN metagenome-assembled genomes originating from the 3 global marine oxygen deficient zones (ODZs), the first from these regions. We survey DPANN abundance and distribution within the ODZ water column, investigate their biosynthetic capabilities, and report potential roles in the cycling of organic carbon, methane, and nitrogen. We test the hypothesis that nitrous oxide reductases found within several ODZ DPANN genomes may enable ultrasmall episymbionts to serve as nitrous oxide consumers when attached to a host nitrous oxide producer. Our results indicate DPANN archaea as ubiquitous residents within the anoxic core of ODZs with the potential to produce or consume key compounds.

Nitrite-oxidizing bacteria (NOB) are important nitrifiers whose activity regulates the availability of nitrite and links reduced ammonium and oxidized nitrate in ecosystems. In oxic marine sediments, ammonia-oxidizing archaea (AOA) and NOB together catalyze the oxidation of ammonium to nitrate, but the observed abundance ratios of AOA to canonical NOB are significantly higher than the theoretical ratio predicted from microbial physiology, indicating that many novel NOBs are yet to be discovered. Here we report a new bacterial phylum Candidatus Nitrosediminicolota, members of which are more abundant than canonical NOBs and are widespread across global oligotrophic sediments. Ca. Nitrosediminicolota members have the functional potential to oxidize nitrite, in addition to other accessory functions such as urea hydrolysis and thiosulfate reduction. While one recovered species ( Ca. Nitrosediminicola aerophilis) is generally confined within the oxic zone, another ( Ca. Nitrosediminicola anaerotolerans) can additionally thrive in anoxic sediments. Counting Ca. Nitrosediminicolota as a nitrite-oxidizer resolves the apparent abundance imbalance between AOA and NOB in oxic marine sediments, and thus its activity may exert a critical control on the nitrite budget.

Bacteria specialized in anaerobic ammonium oxidation (anammox) are widespread in many anoxic habitats and form an important functional guild in the global nitrogen cycle by consuming bio-available nitrogen for energy rather than biomass production. Due to their slow growth rates, cultivation-independent approaches have been used to decipher their diversity across environments. However, their full diversity has not been well recognized. Here, we report a new family of putative anammox bacteria, "Candidatus Subterrananammoxibiaceae," existing in the globally distributed terrestrial and marine subsurface (groundwater and sediments of estuary, deep-sea, and hadal trenches). We recovered a high-quality metagenome-assembled genome of this family, tentatively named "Candidatus Subterrananammoxibius californiae," from a California groundwater site. The "Ca. Subterrananammoxibius californiae" genome not only contains genes for all essential components of anammox metabolism (e.g., hydrazine synthase, hydrazine oxidoreductase, nitrite reductase, and nitrite oxidoreductase) but also has the capacity for urea hydrolysis. In an Arctic ridge sediment core where redox zonation is well resolved, "Ca. Subterrananammoxibiaceae" is confined within the nitrate-ammonium transition zone where the anammox rate maximum occurs, providing environmental proof of the anammox activity of this new family. Phylogenetic analysis of nitrite oxidoreductase suggests that a horizontal transfer facilitated the spreading of the nitrite oxidation capacity between anammox bacteria (in the Planctomycetota phylum) and nitrite-oxidizing bacteria from Nitrospirota and Nitrospinota. By recognizing this new anammox family, we propose that all lineages within the "Ca. Brocadiales" order have anammox capacity. IMPORTANCE Microorganisms called anammox bacteria are efficient in removing bioavailable nitrogen from many natural and human-made environments. They exist in almost every anoxic habitat where both ammonium and nitrate/nitrite are present. However, only a few anammox bacteria have been cultured in laboratory settings, and their full phylogenetic diversity has not been recognized. Here, we present a new bacterial family whose members are present across both the terrestrial and marine subsurface. By reconstructing a high-quality genome from the groundwater environment, we demonstrate that this family has all critical enzymes of anammox metabolism and, notably, also urea utilization. This bacterium family in marine sediments is also preferably present in the niche where the anammox process occurs. These findings suggest that this novel family, named "Candidatus Subterrananammoxibiaceae," is an overlooked group of anammox bacteria, which should have impacts on nitrogen cycling in a range of environments.

By consuming ammonium and nitrite, anammox bacteria form an important functional guild in nitrogen cycling in many environments, including marine sediments. However, their distribution and impact on the important substrate nitrite has not been well characterized. Here we combined biogeochemical, microbiological, and genomic approaches to study anammox bacteria and other nitrogen cycling groups in two sediment cores retrieved from the Arctic Mid-Ocean Ridge (AMOR). We observed nitrite accumulation in these cores, a phenomenon also recorded at 28 other marine sediment sites and in analogous aquatic environments. The nitrite maximum coincides with reduced abundance of anammox bacteria. Anammox bacterial abundances were at least one order of magnitude higher than those of nitrite reducers and the anammox abundance maxima were detected in the layers above and below the nitrite maximum. Nitrite accumulation in the two AMOR cores co-occurs with a niche partitioning between two anammox bacterial families ( Candidatus Bathyanammoxibiaceae and Candidatus Scalinduaceae), likely dependent on ammonium availability. Through reconstructing and comparing the dominant anammox genomes ( Ca . Bathyanammoxibius amoris and Ca . Scalindua sediminis), we revealed that Ca . B. amoris has fewer high-affinity ammonium transporters than Ca . S. sediminis and lacks the capacity to access alternative substrates and/or energy sources such as urea and cyanate. These features may restrict Ca . Bathyanammoxibiaceae to conditions of higher ammonium concentrations. These findings improve our understanding about nitrogen cycling in marine sediments by revealing coincident nitrite accumulation and niche partitioning of anammox bacteria.

Ultrasmall-celled “ Ca. Patescibacteria” have been estimated to account for one-quarter of the total microbial diversity on Earth, the parasitic lifestyle of which may exert a profound control on the overall microbial population size of the local ecosystems. However, their diversity and metabolic functions in marine sediments, one of the largest yet understudied ecosystems on Earth, remain virtually uncharacterized.

By consuming ammonium and nitrite, anammox bacteria form an important functional guild in nitrogen cycling in many environments including marine sediments. Recent studies have shown that anammox bacteria can consume most of the upwardly diffusing ammonium from deep anoxic sediments; however, their impact on the other important substrate nitrite has not been well characterized. Here we show niche partitioning of two anammox families emerges in a 2.4 m long mostly anoxic sediment core retrieved from the Nordic Seas. We document high abundances of anammox bacteria in most investigated sediment layers, with two distinct anammox abundance maxima in two nitrite consumption zones. Between the two anammox abundance maxima, nitrite accumulates as observed in other marine sediment sites and aquatic environments, indicating anammox bacteria play a fundamental role in modulating the nitrite distribution. Anammox bacteria in the upper nitrite consumption zone are dominated by the Candidatus Bathyanammoxibiaceae family, while Ca. Scalinduaceae family dominate at the lower zone. A high-quality representative Ca. Bathyanammoxibiaceae genome is recovered, which, comparing to Ca. Scalindua sediminis, the representative of Scalinduaceae in marine sediments, has fewer high affinity ammonium transporters and lacks the capacity to access alternative substrates or energy sources such as urea and cyanate. These features may restrict Ca. Bathyanammoxibiaceae to conditions of higher ammonium concentrations or fluxes, and therefore drive the observed niche partitioning. These findings improve our understanding about nitrogen cycling in marine sediments by revealing the association between nitrite accumulation and niche partitioning of anammox bacteria.