The global carbon cycle is affected by biological processes in the oceans, which export carbon from surface waters in form of organic matter and store it at depth; a process called the 'biological carbon pump'. Most of the exported organic carbon is processed by the water column biota, which ultimately converts it into CO2 via respiration (remineralization). Variations in the resulting decrease in organic flux with depth 9 can, according to models, lead to changes in atmospheric CO 2 of up to 200 ppm 3 , indicating a strong coupling between biological activity in the ocean interior and oceanic storage of CO 2 .A key constraint in the analysis of carbon fluxes in the twilight zone is that, at steady state, the attenuation of particulate organic carbon (POC) flux with depth should be balanced by community metabolism. Published estimates of POC flux attenuation with depth are, however, up to 2 orders of magnitude lower than corresponding estimates of heterotrophic metabolism [4][5][6][7] . This discrepancy indicates that either estimates of POC flux and/or community metabolism are unreliable, or that additional, unaccounted for, sources of organic carbon to the twilight zone exist 8 .We compiled a comprehensive carbon budget of the twilight zone based on an based on the ratio between DOC concentrations and apparent oxygen utilization 15 , and on DOC gradients coupled to turbulent diffusivity measured from previous work at the study site 16 (Methods; Extended Data Fig. 2). DOC was estimated to supply 17% of total export in agreement with previous estimates of 9-20% across the North Atlantic basin 17 . Organic matter input via lateral advection was assumed to be negligible based on analyses of back-trajectories (derived from satellite-derived near-surface velocities over 3 months) of the water masses arriving at the PAP site during the study period, which suggested that the water had not passed over the continental slope (Extended Data Fig. 1b). The final source of DOC, excretion at depth by active flux, was estimated using net samples of zooplankton biomass and allometric equations 6,18 , giving a supply of 3 mg C m -2 d -1 . Defecation and mortality at depth present further sources of organic carbon to the twilight zone, but these were excluded from the budget due to large uncertainties associated with their estimation. Finally, chemolithoautotrophy has been suggested to be a significant source of organic matter in the deep ocean 19 , but without strong evidence that this poorly understood process could provide a major contribution at our study site, we chose to exclude it from our carbon budget.The remineralization of organic carbon by zooplankton and prokaryotes was estimated from zooplankton biomass and prokaryotic activity. It is crucial to note that in a steady state system, such as we assume this to be, organic carbon is lost from the system only by export or by remineralization. We focus entirely on community respiration as a measure of remineralization, a fundamental advance over previous methods to derive...

SummaryEffects of hydrostatic pressure on pure cultures of prokaryotes have been studied extensively but impacts at the community level in the ocean are less well defined. Here we consider hydrostatic pressure effects on natural communities containing both unadapted (piezosensitive) prokaryotes originating from surface water and adapted (including piezophilic) prokaryotes from the deep sea. Results from experiments mimicking pressure changes experienced by particle-associated prokaryotes during their descent through the water column show that rates of degradation of organic matter (OM) by surface-originating microorganisms decrease with sinking. Analysis of a much larger data set shows that, under stratified conditions, deep-sea communities adapt to in situ conditions of high pressure, low temperature and low OM. Measurements made using decompressed samples and atmospheric pressure thus underestimate in situ activity. Exceptions leading to overestimates can be attributed to deep mixing events, large influxes of surface particles, or provision of excessive OM during experimentation. The sediment-water interface, where sinking particles accumulate, will be populated by a mixture of piezosensitive, piezotolerant and piezophilic prokaryotes, with piezophilic activity prevailing deeper within sediment. A schematic representation of how pressure shapes prokaryotic communities in the ocean is provided, allowing a reasonably accurate interpretation of the available activity measurements.