Microbial community dynamics over large spatial and environmental gradients in a subtropical ocean basin

Microbes are fundamental to ocean ecosystem function, yet they remain understudied across broad spatial and environmental scales in dynamic regions like the Gulf of America/Gulf of Mexico (GOM). We employed DNA metabarcoding to characterize prokaryotes (16S V4-V5) and protists (18S V9) across 51 stations, spanning 16 inshore-offshore transects and three depths. Cluster analysis revealed three clusters corresponding to depth zones that integrated vertical and horizontal sampling: photic zone (inshore near surface-bottom and offshore surface), deep chlorophyll maximum (offshore), and aphotic zone (offshore near bottom). We applied group-specific generalized additive models (GAMs) to log-transformed abundance data of major taxa in the photic zone, identifying key environmental factors that explained 42%-82% of the variation in abundance. SAR11 and SAR86 were positively associated with temperature and dissolved inorganic carbon, while cyanobacterial genera (<i>Prochlorococcus</i> and <i>Synechococcus</i>) were differently impacted by nutrients, salinity, and pH in ways that often followed their expected ecological niches. Representatives of protist parasites (Syndiniales) and grazers (Sagenista) showed group-specific nonlinear associations with salinity, oxygen, nutrients, and temperature. Using GAMs, we expanded the spatial resolution of DNA sampling and predicted surface log abundances at 84 cruise sites lacking amplicon data. Indicator analysis was performed with sequence-level data, revealing several protists that were indicative of more acidic waters and the absence of any significant prokaryote indicators. Our results provide the first basin-scale survey of microbes in the GOM and highlight the need for coordinated omics and environmental sampling to improve predictions of microbial responses to changing conditions.IMPORTANCEMarine microbes are key indicators of environmental change and play central roles in ocean food webs and biogeochemical cycles. Yet, how natural microbial communities respond to shifting environmental conditions remains unclear, particularly in the Gulf of Mexico (GOM), a region shaped by dynamic physical and chemical gradients. Here, we conducted a novel basin-scale DNA metabarcoding survey of prokaryotes and protists in the GOM. We used generalized additive models and indicator analysis to reveal environmental drivers of microbial abundance, from broader taxonomic groups to unique sequences. Our results show group-specific associations with environmental factors such as temperature, nutrients, salinity, and carbonate chemistry parameters and identify several protist taxa associated with distinct ocean conditions. These findings provide a foundation for microbial monitoring in the GOM and shed light on the importance of integrating <i>in situ</i> biological, physical, and chemical data across spatial gradients to inform accurate ecosystem and biogeochemical models.