The chemical and biological characteristics of the surface microlayer were determined during a transect across the South Pacific Ocean in October-December 2004. Concentrations of particulate organic carbon (1.3 to 7.6-fold) and nitrogen (1.4 to 7-fold), and POC:PON ratios were consistently higher in the surface microlayer as compared to surface waters (5 m). The large variability in particulate organic matter enrichment was negatively correlated to wind speed. No enhanced concentrations of dissolved organic carbon were detectable in the surface microlayer as compared to 5 m, but chromophoric dissolved organic matter was markedly enriched (by 2 to 4-fold) at all sites. Based on pigment analysis and cell counts, no consistent enrichment of any of the major components of the autotrophic and heterotrophic microbial community was detectable. CE-SSCP fingerprints and CARD FISH revealed that the bacterial communities present in the surface microlayer had close similarity (>76%) to those in surface waters. By contrast, bacterial heterotrophic production (³H-leucine incorporation) was consistently lower in the surface microlayer than in surface waters. By applying CARD-FISH and microautoradiography, we observed that <i>Bacteroidetes</i> and <i>Gammaproteobacteria</i> dominated leucine uptake in the surface microlayer, while in surface waters <i>Bacteroidetes</i> and <i>Alphaproteobacteria</i> were the major groups accounting for leucine incorporation. Our results demonstrate that the microbial community in the surface microlayer closely resembles that of the surface waters of the open ocean. Even a short residence in the surface microlayer influences leucine incorporation by different bacterial groups, probably as a response to the differences in the physical and chemical nature of the two layers.