Comparative analysis of mycorrhizal communities associated with Struthiopteris spicant (L.) Weiss across Europe and North America.

Introduction: Ferns constitute the second largest group of vascular plants. Previous studies have shown that the diversity and composition of fern communities are influenced by resource availability and water stress, among other factors. However, little is known about the influence of these environmental factors on their biotic interactions, especially regarding the relationship between mycorrhizal fungi and ferns. The present study compares the mycorrhizal communities associated with 36 populations of Struthiopteris spicant L. Weiss across Europe and North America. This species exhibits a great tolerance to variations in light, nutrient, and pH conditions, and it can survive with and without mycorrhizae. Methods: With the aim of determining which environmental factors impact the composition and abundance of the root-associated fungal communities in this species, we used an ITS-focused metabarcoding approach to identify the mycorrhizal fungi present and analyzed the influence of climatic and edaphic variables at global and regional scales. Results and discussion: We encountered striking differences in the relative abundance of arbuscular mycorrhizal fungi (AMF) between S. spicant populations at both spatial levels. We recorded a total of 902 fungal ASVs, but only 2- 4% of the total fungal diversity was observed in each individual, revealing that each fern had a unique fungal community. Light availability and the interactive action of pH and soil nitrogen concentration showed a positive influence on AMF relative abundance, explaining 89% of the variance. However, environmental factors could only explain 4- 8% of the variability in AMF community composition, indicating that it might be determined by stochastic processes. These results support the hypothesis that ferns may be more independent of mycorrhization than other plant groups and interact with fungi in a more opportunistic manner.

Facultative mycorrhization in a fern (Struthiopteris spicant L. Weiss) is bound to light intensity.

BACKGROUND: The establishment of mycorrhizal relationships between a fungus and a plant typically enhances nutrient and water uptake for the latter while securing a carbon source for the fungus. However, under a particular set of environmental conditions, such as low availability of light and abundant nutrients in the soil, the resources invested in the maintenance of the fungi surpass the benefits obtained by the host. In those cases, facultative mycorrhizal plants are capable of surviving without symbiosis. Facultative mycorrhization in ferns has been overlooked until now. The present study measured the response of Struthiopteris spicant L. Weiss, and its root-associated fungi to different levels of light and nutrient availability in terms of growth, mycorrhizal presence, and leaf nutrient content. This fern species exhibits a great tolerance to variable light, nutrient, and pH conditions, and it has been found with and without mycorrhizae. We conducted a greenhouse experiment with 80 specimens of S. spicant and three factors (Light, Phosphorus, and Nitrogen) resulting in eight treatments. RESULTS: We found a significant influence of the factor light on fungal community composition, plant biomass, and nutrient accumulation. Departing from a lack of colonization at the initial stage, plants showed a remarkable increment of more than 80% in the arbuscular mycorrhizal fungi (AMF) richness and abundance in their roots when grown under high light conditions, compared with the ones in low light. We also observed an upward trend of C:P and C:N ratios and the above- and belowground biomass production when AMF abundance increased. Furthermore, the compositional analysis of the whole fungal communities associated with S. spicant roots revealed clear differences among low-light and high-light treatments. CONCLUSIONS: This study is the first to investigate the importance of light and nutrient availability in determining fern-AMF relationships. We confirmed that Struthiopteris spicant is a facultative mycorrhizal plant. The composition and diversity of AMF found in the roots of this fern are strongly influenced by light and less by nutrient conditions. Our study shows that ferns respond very sensitively to changes in environmental factors, leading to shifts in the associated mycorrhizal communities.

A metabarcoding protocol targeting two DNA regions to analyze root-associated fungal communities in ferns and lycophytes.

Premise: Detailed studies of the fungi associated with lycophytes and ferns provide crucial insights into the early evolution of land plants. However, most investigations to date have assessed fern-fungus interactions based only on visual root inspection. In the present research, we establish and evaluate a metabarcoding protocol to analyze the fungal communities associated with fern and lycophyte roots. Methods: We used two primer pairs focused on the ITS rRNA region to screen the general fungal communities, and the 18S rRNA to target Glomeromycota fungi (i.e., arbuscular mycorrhizal fungi). To test these approaches, we collected and processed roots from 12 phylogenetically distant fern and lycophyte species. Results: We found marked compositional differences between the ITS and 18S data sets. While the ITS data set demonstrated the dominance of orders Glomerales (phylum Glomeromycota), Pleosporales, and Helotiales (both in phylum Ascomycota), the 18S data set revealed the greatest diversity of Glomeromycota. Non-metric multidimensional scaling (NMDS) ordination suggested an important geographical effect in sample similarities. Discussion: The ITS-based approach is a reliable and effective method to analyze the fungal communities associated with fern and lycophyte roots. The 18S approach is more appropriate for studies focused on the detailed screening of arbuscular mycorrhizal fungi.

Premisa: El estudio de los hongos asociados a licofitas y helechos proporciona información crucial sobre la evolución temprana de las plantas terrestres. Sin embargo, hasta el momento, la mayoría de las investigaciones ha evaluado las interacciones helecho­hongo basándose solamente en la observación directa de las raíces. En la presente investigación, establecemos y evaluamos un protocolo de metabarcoding enfocado en dos regiones de ADN para analizar las comunidades fúngicas asociadas a las raíces de helechos y licofitas. Métodos: Utilizamos dos pares de primer orientados hacia la región ITS ARNr, para la detección de las comunidades fúngicas generales, y la región 18S ARNr, para captar hongos pertenecientes al phylum Glomeromycota (i.e., hongos micorrícicos arbusculares). Para evaluar estos procedimientos, nosotros recolectamos y procesamos raíces de 12 especies de helechos y licofitas distantes desde el punto de vista filogenético. Resultados: Se observaron claras diferencias de composición entre los sets de datos ITS y 18S: mientras el primero demostró un predominio de los órdenes Glomerales (phylum Glomeromycota), Pleosporales y Helotiales (ambos en phylum Ascomycota), el set 18S reveló la mayor diversidad de hongos micorrizógenos arbusculares. Ninguno de los marcadores moleculares utilizados detectó miembros del phylum Mucoromycota en las muestras. El escalamiento multidimensional no métrico (NMDS) sugirió un papel importante de la región geográfica de origen en la determinación de las similitudes entre muestras. Discusión: El método basado en la región ITS es consistente, replicable y eficaz para analizar las comunidades fúngicas asociadas con raíces de helechos y licofitos. El enfoque 18S es más apropiado para estudios centrados en la detección de los hongos micorrizógenos arbusculares.