The diversity of cultured actinomycete bacteria was compared between near- and offshore marine sediments. Strains were tested for the effects of seawater on growth and analyzed for 16S rRNA gene sequence diversity. In total, 623 strains representing 6 families in the order Actinomycetales were cultured. These strains were binned into 16 to 63 operational taxonomic units (OTUs) over a range of 97 to 100 % sequence identity. The majority of the OTUs were closely related (> 98 % sequence identity) to strains previously reported from non-marine sources; indicating that most are not restricted to the sea. However, new OTUs averaged 96.6 % sequence identity with previously cultured strains and ca. one-third of the OTUs were marine-specific, suggesting that sediment communities include considerable actinomycete diversity that does not occur on land. Marine specificity did not increase at the off-shore sites, indicating high levels of terrestrial influence out to 125 km from shore. The requirement of seawater for growth was observed among < 6%. of the strains, while all members of 9 OTUs possessed this trait, revealing a high degree of marine adaptation among some lineages. Statistical analyses predicted greater OTU diversity at the off-shore sites and provided a rationale for expanded exploration of deep-sea samples. A change in community composition was observed, with the number of Micromonospora OTUs increasing in the off-shore samples. UniFrac (see http://bmf2.colorado.edu/unifrac) statistics support a difference in community composition between near- and off-shore locations. Overall, 123 of 176 strains had distinct 16S rRNA gene sequences, indicating a high level of actinomycete diversity in marine sediments.