CaCO₃ optical detection with fluorescent in situ hybridization: A new method to identify and quantify calcifying microorganisms from the oceans

Open oceanic calcification is mainly driven by unicellular organisms and in particular by eukaryotes such as coccolithophores and foraminifers. Open ocean microcalcifiers, like most planktonic protists, are characterized by extremely fast generation times and occasional sexual reproduction. Populations can alternate between diploid and haploid stages, which often build different kinds of cell covers. In the most important pelagic calcifiers, the coccolithophores, the diploid and haploid stages, which can self-replicate and grow independently, display radically different morphologies with different modes of calcification or even with the absence of calcification in at least one life cycle stage. Although life cycle strategies seem likely to fundamentally influence the where and when of open ocean calcification, this issue has yet to be seriously addressed in the natural environment. Here, we introduce a new morphogenetic method, "combined CaCO₃ optical detection with fluorescent in situ hybridization," or COD-FISH, which is based on a combination of TSA-FISH and polarized optical microscopy. This technique allows simultaneous assessment of the taxonomic and life cycle status of single coccolithophore cells collected from the ocean. We demonstrate the application of COD-FISH using both laboratory culture and field samples and discuss its potential value for assessing the ecology, biodiversity, population structure, and life cycles of coccolithophores and other open ocean unicellular calcifiers.