Microbial diversity of Alcyonium digitatum

Marine multi-cellular organisms are described as sources of many newly discovered bioactive compounds. Meanwhile, it has been demonstrated repeatedly for several natural products of reputed multicellular origin that they are, in fact, produced by endophytic unicellular organisms-such as microbial fungi or bacteria. Consequently, while studying compounds isolated from a living organism, it is essential to ensure that the sample integrity is not compromised. To test the diversity of the endobiome from Alcyonium digitatum, a cold water coral found along the Atlantic coasts of the northern hemisphere, we performed a culture dependent surveyed using a phylogenetic approach. A 1 cm(3) cube from the interior tissue of A. digitatum was excised under aseptic conditions, homogenized, spread onto agar-based growth medium plates and incubated in 22 A degrees C to promote microbial growth. Colonies were transferred to secondary medium plates, incubated, and after harvesting lysed using sterile water to release DNA. 16S and 23S rDNA regions were amplified using PCR, and sequenced for systematic evaluation using phylogenetic analysis. From this survey we identified a broad selection of bacteria, predominantly of the alpha-proteobacterial, bacteriodete, actinobacterial and firmicute lineages, demonstrating a significant biodiversity of the coral bacterial endobiome.