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ABSTRACT Vitamin B 1 (thiamin) is a cofactor for critical enzymatic processes and is scarce in surface oceans. Several eukaryotic marine algal species thought to rely on exogenous thiamin are now known to grow equally well on the precursor 4-amino-5-hydroxymethyl-2-methylpyrimidine (HMP), including the haptophyte Emiliania huxleyi . Because the thiamin biosynthetic capacities of the diverse and ecologically important haptophyte lineage are otherwise unknown, we investigated the pathway in transcriptomes and two genomes from 30 species representing six taxonomic orders. HMP synthase is missing in data from all studied taxa, but the pathway is otherwise complete, with some enzymatic variations. Experiments on axenic species from three orders demonstrated that equivalent growth rates were supported by 1 µM HMP or thiamin amendment. Cellular thiamin quotas were quantified in the oceanic phytoplankter E. huxleyi using the thiochrome assay. E. huxleyi exhibited luxury storage in standard algal medium [(1.16 ± 0.18) × 10 −6  pmol thiamin cell −1 ], whereas quotas in cultures grown under more environmentally relevant thiamin and HMP supplies [(2.22 ± 0.07) × 10 −7 or (1.58 ± 0.14) × 10 −7  pmol thiamin cell −1 , respectively] were significantly lower than luxury values and prior estimates. HMP and its salvage-related analog 4-amino-5-aminomethyl-2-methylpyrimidinemore »(AmMP) supported higher growth than thiamin under environmentally relevant supply levels. These compounds also sustained growth of the stramenopile alga Pelagomonas calceolata . Together with identification of a salvage protein subfamily (TENA_E) in multiple phytoplankton, the results indicate that salvaged AmMP and exogenously acquired HMP are used by several groups for thiamin production. Our studies highlight the potential importance of thiamin pathway intermediates and their analogs in shaping phytoplankton community structure. IMPORTANCE The concept that vitamin B 1 (thiamin) availability in seawater controls the productivity and structure of eukaryotic phytoplankton communities has been discussed for half a century. We examined B 1 biosynthesis and salvage pathways in diverse phytoplankton species. These comparative genomic analyses as well as experiments show that phytoplankton thought to require exogenous B 1 not only utilize intermediate compounds to meet this need but also exhibit stronger growth on these compounds than on thiamin. Furthermore, oceanic phytoplankton have lower cellular thiamin quotas than previously reported, and salvage of intermediate compounds is likely a key mechanism for meeting B 1 requirements under environmentally relevant scenarios. Thus, several lines of evidence now suggest that availability of specific precursor molecules could be more important in structuring phytoplankton communities than the vitamin itself. This understanding of preferential compound utilization and thiamin quotas will improve biogeochemical model parameterization and highlights interaction networks among ocean microbes.« less