Assessing Genotypic and Environmental Effects on Endophyte Communities of Fraxinus (Ash) Using Culture Dependent and Independent DNA Sequencing

Abstract

Fraxinus excelsior populations are in decline due to the ash dieback disease Hymenoscyphus fraxineus. It is important to understand genotypic and environmental effects on its fungal microbiome to develop disease management strategies. To do this, we used culture dependent and culture independent approaches to characterize endophyte material from contrasting ash provenances, environments, and tissues (leaves, roots, seeds). Endophytes were isolated and identified using nrITS, LSU, or tef DNA loci in the culture dependent assessments, which were mostly Ascomycota and assigned to 37 families. Few taxa were shared between roots and leaves. The culture independent approach used high throughput sequencing (HTS) of nrITS amplicons directly from plant DNA and detected 35 families. Large differences were found in OTU diversity and community composition estimated by the contrasting approaches and these data need to be combined for estimations of the core endophyte communities. Species richness and Shannon index values were highest for the leaf material and the French population. Few species were shared between seed and leaf tissue. PCoA and NMDS of the HTS data showed that seed and leaf microbiome communities were highly distinct and that there was a strong influence of Fraxinus species identity on their fungal community composition. The results will facilitate a better understanding of ash fungal ecology and are a step toward identifying microbial biocontrol systems to minimize the impact of the disease.

Fungal Communities in Re-Emerging Fraxinus excelsior Sites in Lithuania and Their Antagonistic Potential against Hymenoscyphus fraxineus

“…Most frequently isolated fungi from leaves were A. alternata , Aureobasidium pullulans , Apiospora montagnei , and G. avenacea . The fungal taxa detected in shoots and leaves of F. excelsior were similar to those reported in numerous other culture-based studies [ 11 , 12 , 13 , 14 ]. However, high-throughput sequencing of the shoot and leave samples collected in the present study revealed a very contrasting fungal community structure [ 15 ].…”

First Report of the Ash Sawfly, Tomostethus nigritus, Established on Fraxinus excelsior in the Republic of Ireland

“…A weakened tree can also become vulnerable to more severe threats, such as ash dieback disease caused by the fungus Hymenoscyphus fraxineus [22]. Ash dieback is already present on the island of Ireland and it is a major threat to the ash tree population [23]. The combination of ash dieback Adult sawflies have been seen clinging to clothing of cyclists and walkers on the Lagan towpath in Belfast and it is plausible that they could be carried on sheltered parts of cars and lorries in the same manner.…”

“…A weakened tree can also become vulnerable to more severe threats, such as ash dieback disease caused by the fungus Hymenoscyphus fraxineus [22]. Ash dieback is already present on the island of Ireland and it is a major threat to the ash tree population [23]. The combination of ash dieback with the outbreaks of ash sawflies could acerbate even further the already precarious situation of the ash tree population in Ireland.…”

Ash Dieback and Other Pests and Pathogens of Fraxinus on the Island of Ireland

“…A combination effect of sunken canker with H. fraxineus further aggravates ash dieback, accelerating the decline of the tree (Linaldeddu et al 2020). All of the three species here described have not been formally recorded in Ireland yet (FRDBI database 2021) except by Lahiri, Murphy and Hodkinson (2021).…”