Abstract. The sea surface microlayer (SML) represents the boundary layer at the air-sea interface. Microbial eukaryotes in the SML potentially influence air-sea gas exchange directly by taking up and producing gases, and indirectly by excreting and degrading organic matter, which may modify the viscoelastic properties of the SML. However, little is known about the controlling factors that influence microbial eukaryote community composition in the SML. We studied the composition of the microbial community, transparent exopolymer particles and polysaccharides in the SML during the PEACETIME cruise along a west-east transect in the Mediterranean Sea, covering the western basin, Tyrrhenian Sea and Ionian Sea. At the stations located in the Ionian Sea, fungi were found in high relative abundances determined by 18S sequencing efforts, making up a significant proportion of the sequences recovered. At the same time, bacterial and phytoplankton counts were decreasing from west to east, while transparent exopolymer particle (TEP) abundance and total carbohydrate (TCHO) concentrations remained the same between Mediterranean basins. Thus, the presence of substrates for fungi, such as Cladosporium known to take up phytoplankton-derived polysaccharides, in combination with decreased substrate competition by bacteria suggests that fungi could be thriving in the neuston of the Ionian Sea.