Microbial exposureisrelated to the health of passengers on commercial aircraft, but nostudies characterizedthe microbial compositionat the species level and identified their ecological determinants. We collected vacuum dust from floor and seatsurfaces in flight decks and cabins of 18 aircraft, and amplification-freeshotgunmetagenomicswas conducted to characterize the microbial composition. In total,7,437 microbial taxa were identified. The relative abundance for bacteria, eukaryote, viruses and archaea were 96.9%, 1.8%, 0.3% and 0.03%, respectively.The top bacterial species weremainly derived from outdoor air and human skin, included Sphigomonas, Corynebacterium, Micrococcus leutens, Variovorax paradoxus, Paracoccus dentrificans and Propionibacterium acnes.The abundance ofNIAID definedpathogens was low, accounted for only 0.23% of total microbes.Themicrobial species and functional composition were structured by theindoor surface type (R = 0.38, Adonis), followed by themanufacturer of the aircraft (R = 0.12) andflight duration (R = 0.07).Indoor surfacesaffected speciesderivedfrom different habitats; the abundance of dry skin anddesiccatedspecies were higheron textile surfaces, whereas the abundance of moist and oilyskin species washigher on leather surfaces.The growth rates for most microbes were stopped and almost stopped.