Microbial dinitrogen (N2) fixation, the nitrogenase enzyme-catalysed reduction of N2 gas into biologically available ammonia, is the main source of new nitrogen (N) in the ocean. For more than 50 years, oceanic N2 fixation has mainly been attributed to the activity of the colonial cyanobacterium Trichodesmium1,2. Other smaller N2-fixing microorganisms (diazotrophs)—in particular the unicellular cyanobacteria group A (UCYN-A)—are, however, abundant enough to potentially contribute significantly to N2 fixation in the surface waters of the oceans3,4,5,6. Despite their abundance, the contribution of UCYN-A to oceanic N2 fixation has so far not been directly quantified. Here, we show that in one of the main areas of oceanic N2 fixation, the tropical North Atlantic7, the symbiotic cyanobacterium UCYN-A contributed to N2 fixation similarly to Trichodesmium. Two types of UCYN-A, UCYN-A1 and -A2, were observed to live in symbioses with specific eukaryotic algae. Single-cell analyses showed that both algae–UCYN-A symbioses actively fixed N2, contributing ∼20% to N2 fixation in the tropical North Atlantic, revealing their significance in this region. These symbioses had growth rates five to ten times higher than Trichodesmium, implying a rapid transfer of UCYN-A-fixed N into the food web that might significantly raise their actual contribution to N2 fixation. Our analysis of global 16S rRNA gene databases showed that UCYN-A occurs in surface waters from the Arctic to the Antarctic Circle and thus probably contributes to N2 fixation in a much larger oceanic area than previously thought. Based on their high rates of N2 fixation and cosmopolitan distribution, we hypothesize that UCYN-A plays a major, but currently overlooked role in the oceanic N cycle.