Fungal root symbionts of the seagrass Posidonia oceanica in the central Adriatic Sea revealed by microscopy, culturing and 454-pyrosequencing

A significant part of the hitherto unexplored fungal diversity is hidden in the marine environment. At the same time, plant tissues host endophytic communities often dominated by yet undescribed fungal lineages. Here we focused on the Mediterranean endemic seagrass Posidonia oceanica and screened its root mycobionts at 8 localities in the Croatian central Adriatic Sea using (stereo-) microscopy, culturing from surface-sterilized root segments and 454-pyrosequencing. Our microscopic observations revealed that roots from all investigated localities possessed the typical dark septate endophytic association recently reported in this seagrass in the northwestern Mediterranean Sea. Both culturing and pyrosequencing detected very narrow fungal communities lacking typical terrestrial root endophytes. Similarly to the NW Mediterranean, these were dominated by an undescribed slow-growing mycobiont from the Pleosporales (1 operational taxonomic unit [OTU]: ca. 92% of 430 total isolates, and 3 molecular OTUs [MOTUs]: ca. 91% of 382 fungal sequences) and also comprised 2 yet undescribed mycobionts from the Lulworthiales (2 OTUs: ca. 8% of isolates, and 2 MOTUs: ca. 1.8% of sequences). Such a narrow, single-species dominated root mycobiont spectrum is unusual for photoautotrophic vascular plants and indicates a close symbiotic relationship between the dominating pleosporalean mycobiont and the dominant Mediterranean seagrass. Additionally, the apparent lack of typical terrestrial root endophytic fungi as well as their probable substitution by the pleosporalean mycobiont so far not known from other hosts or ecosystems implies relatively long specific coevolution of both marine organisms.