Seasonal and evolutionary patterns of planktonic Hikarchaeia reveal ecological niches of marine rare biosphere constituents

Members of Marine Group IV, now termed Hikarchaeia, hold an important transitional evolutionary position among Archaea but remain uncultivated, and their ecology remains uncharacterized. Over a 5-year period, we performed monthly profile sampling in the North Atlantic Subtropical Gyre and sequenced V4 16S rRNA gene amplicons, while also collecting additional size-fractionated and metagenomic samples. Hikarchaeal relative abundance annually peaked at ~1% of prokaryotic amplicons. Their relative abundances were significantly higher after deep winter mixing when thermal stratification isolated exported dissolved organic matter in the twilight zone (TZ), the boundary region between the epipelagic and mesopelagic. A diminutive 1.17 Mbp complete hikarchaeal genome was recovered by sequencing and co-assembling diel TZ metagenomes from 200 m during summer, where their relative abundances were highest. Phylogenomic reconstruction incorporating our new metagenome-assembled genome (MAG) and public MAGs established five clades. Clades I and III were in subtropical and tropical environments, with vertical partitioning: Clade I dominated the TZ, and Clade III detection initiated in the lower mesopelagic and extended into the bathypelagic. Clade V is polar-associated. Data sparsity makes Clade II and IV distributions unclear. All Hikarchaeia recovered thus far appear to be free-living aerobes capable of heterotrophy. Unlike MGI, MGII, and MGIII Archaea, and most marine pelagic bacteria, the Hikarchaeia encode bacterioruberin biosynthesis, except deep Clade III. Hikarchaeia also have nitrile hydratases alongside enzymes for carbon monoxide oxidation and menaquinone biosynthesis, suggesting previously unrecognized roles in nitrogen and carbon cycling. Collectively, our studies reveal niche partitioning of evolutionarily distinct uncultivated Hikarchaeia clades and patterning within the rare biosphere.IMPORTANCEThe ocean and its vertical water-column structure are changing with planetary warming; yet, there are many enigmatic microbes, especially those that inhabit the deep ocean. Here, we illuminate a group of uncultivated Archaea that were discovered over two decades ago but still lack clear distributions and habitat preferences. With the first complete hikarchaeal genome now in hand, we identified distinct lineages within the Hikarchaeia that occupy different marine niches. Strong reoccurring seasonal patterns were observed that closely followed the annual cycle of temperature-driven ocean stratification. Given rapid natural and anthropogenic perturbations at hand, establishing baselines for these Archaea is critical for improving our understanding and modeling of marine microbiomes and their biogeochemical functions.