Microorganisms are key to balancing marine ecosystems and have complex interactions at the ocean–atmosphere interface, affecting global climate and human health. This research investigated the diversity of cultivable bacteria and fungi in marine bioaerosols in the North Tropical Atlantic Ocean. Using the technique of spontaneous sedimentation in selective culture media, samples were collected during oceanographic expeditions. After isolation and purification, microbial strains were identified by phenotypic and genetic analyses. Fungi isolated included Acrophialophora, Aspergillus, Chrysosporium, Cladosporium, Fonsecaea, Mucor, Rhodotorula, Schizophyllum, Stemphylium, Candida, Curvularia, Cystobasidium, Exophiala, Neotestudina, Penicillium, Pestalotiopsis, and Preussia. The bacterial isolates belonged to the Bacillota, Pseudomonadota, Enterobacteriaceae family, Bacillus genus, and Serratia liquefaciens groups. About 40% of bacteria and 42% of fungi were identified as potential human pathogens, suggesting a relationship between human actions and the microbiota present in bioaerosols on the high seas. Sea surface temperature (SST) and wind speed influenced microorganisms. More studies and analyses in different scenarios should be conducted considering environmental and climate variables in order to deepen knowledge and generate information on the subject, so that standards can be established, and quality parameters determined.