AbstractThe number of marine environments known to harbor dinitrogen (N2)‐fixing (diazotrophic) microorganisms is increasing, prompting a reassessment of the biogeography of marine diazotrophs and N2fixation rates (NFRs). Here, we investigate the diversity, abundance, and activity of diazotrophic microorganisms in the North Pacific Subtropical Gyre (NPSG), a diazotrophic habitat, and the North Pacific Transition Zone (NPTZ), a region characterized by strong physical, chemical, and biological gradients. Samples were collected on two springtime meridional cruises during 2016 and 2017, spanning from 23.5°N to 41.4°N along 158°W. We observed an abrupt decrease in diazotrophic abundances near the southern edge of the NPTZ, which coincided with a salinity front and with a ∼10‐fold increase inSynechococcusabundance, but without a concomitant change in phosphate or nitrate concentrations. In NPSG waters south of this diazotrophic boundary,nifHgenes and NFRs were consistently detected and diazotrophic communities were dominated by UCYN‐A, an uncultivated, symbiotic cyanobacterium (2.8 × 103to 1.0 × 106nifHgene copies L−1). There was a significant positive relationship between quantitative polymerase chain reaction‐derived UCYN‐AnifHgene abundances and community NFRs in the NPSG, suggesting a large contribution of UCYN‐A to community NFRs. In the NPTZ waters to the north, NFRs were low or undetected andnifHgenes were rare, with the few detected sequences represented by UCYN‐A and noncyanobacterial diazotrophs. The patterns we observed in UCYN‐A abundance in the context of local biogeochemistry suggest that the environmental controls of this organism may differ from those of cultivated marine cyanobacterial diazotrophs.