From Population to Individual: Bulk and Single-Cell Analysis Reveal Fe Bioavailability, Limitation Thresholds, and Biological Responses for Marine Diatom Chaetoceros debilis.

In marine ecosystems, Fe fundamentally regulates algal productivity and drives biogeochemical cycling in the hydrosphere, yet its environmental controls and cellular responses remain insufficiently resolved, particularly at the single-cell level. Herein, we combined bulk and single-cell analysis to systematically investigate Fe bioaccumulation process in the marine diatom Chaetoceros debilis under varying Fe species, initial pH, and concentration conditions. Fe bioavailability followed the order Fe-ethylenediaminetetraacetate > Fe-deferoxamine B > Ferrihydrite and decreased with declining initial pH, highlighting the combined control of chemical speciation and ocean acidification. A critical bioavailable Fe limitation threshold of ∼10 nM was identified, below which algal growth, pigmentation, and cell size were significantly suppressed. Under Fe limitation, cells exhibited compensatory accumulation of other metals (e.g., Zn, Mn, Cu, and Ni), indicating metallome remodeling as an adaptive strategy. Notably, single-cell analysis revealed pronounced heterogeneity in Fe accumulation within populations, with heterogeneity positively related to bioavailable Fe levels. These findings demonstrate that algal Fe accumulation is governed by coupled environmental and physiological processes and that population-level averages mask critical cell-to-cell variability and their adaptive strategies under environmental stress. This work provides mechanistic insights into how environmental chemistry regulates metal bioavailability and biological responses, offering a broadly applicable framework for understanding the environmental behavior and ecological effects of trace metals and other contaminants in aquatic ecosystems.