The vast majority of terrestrial plants exchange nutrients with fungal partners forming different mycorrhizal types. The minority of plants considered as non-mycorrhizal, however, are not necessarily free of any fungi, but are frequently colonized by elusive fungal endophytes, such as dark septate endophytes (DSE) or fine root endophytes (FRE). While a functional role of FRE in improvement of nutrient gain was recently elucidated, the function of DSE is still in discussion and was here addressed for 36 plant species belonging to the families Equisetaceae, Cypereaceae and Caryophyllaceae.

Molecular and microscopic staining approaches were conducted to verify the presence of DSE in the investigated species. Stable isotope natural abundances of the elements carbon, nitrogen, hydrogen and oxygen and total nitrogen concentrations were analyzed for the respective species of the target plant families and accompanying mycorrhizal and non-mycorrhizal (Brassicaceae) plant species.

Staining approaches confirmed the presence of DSE in all investigated species within the families Equisetaceae, Cyperaceae and Caryophyllaceae. A co-colonization with Paris-type arbuscular mycorrhiza (AM) was occasionally found by staining and molecular approaches in species of the Equisetaceae. Species of the Equisetaceae, Cyperaceae and Caryophyllaceae were significantly ¹⁵N-enriched in comparison to accompanying plants. In addition, a significant ¹³C and ²H enrichment and increased total nitrogen concentrations were found for representatives of the Equisetaceae.

The ¹⁵N-enrichment found here for representatives of Equisetaceae, Cyperaceae and Caryophyllaceae provides evidence for a functional role of the ubiquitous DSE fungi. DSE fungi obviously provide access to ¹⁵N-enriched soil organic compounds probably in exchange for organic carbon compounds from plant photosynthesis. As indicated by additional ¹³C- and ²H-enrichments, representatives of the Equisetaceae apparently gain simultaneously organic carbon compounds from their AM fungi of the Paris-morphotype. Thus, species of the Equisetaceae have to be considered as partially, or in case of the achlorophyllous fertile Equisetum arvense, as fully mycoheterotrophic at least in some stages of their life cycle.

So far mostly underappreciated fungi classified as DSE are suggested to occupy an ecologically relevant role similar to mycorrhizae and the occurrence of simultaneous functions of DSE and AM fungi in Equisetaceae is proposed.