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Plant compartments provide unique niches that lead to specific microbial associations. The microbiota colonizing the endophytic compartment (endorhizosphere) and the rhizosphere contribute to productivity, plant growth, phytoremediation and carbon sequestration. The main objective of this study was to investigate how fungal communities are enriched in different habitats outside and inside of grapevine roots. For this purpose, the spatial dynamics of the fungal communities associated with three soil-plant compartments (bulk soil, rhizosphere and endorhizosphere) were characterized by ITS high-throughput amplicon sequencing (HTAS). Fungal communities were largely affected in their diversity and composition by soil-plant compartments, whereas the spatial variation (i.e. across five vineyards) was low. The endorhizosphere compartment differed most from the other two, suggesting that the root tissues entail a barrier for fungal colonization. The results of functional prediction via FUNGuild suggested an increase in the relative abundances of potential plant pathogens, endophytes and arbuscular mycorrhiza, and a decrease in wood, dung and undefined saprotrophs from bulk soil towards the endorhizosphere. Roots of asymptomatic vines were a microbial niche that is inhabited by soilborne fungi associated with grapevine trunk diseases, which opens up new perspectives in the study of the endophytic role of these pathogens on grapevines. Results obtained in this study provide helpful information to better know how the host shapes its microbiome and the implications for vineyard productivity and management., David Gramaje was supported by the DOC-INIA program from theINIA, co-funded by the European Social Fund. María del Pilar Martínez-Diz was supported by the FPI-INIA program from the INIA.

The ecology of total fungal communities in grapevine is so far largely derived from studies on culture-dependent methods or cultivation-independent rDNA approaches. Sequencing the ribosomal RNA transcripts (rRNA) would rather reveal the functionally and metabolically active important taxa of the fungal community and provide insights into its activity in the wood. The present study investigated changes in the potentially active fungal communities of internal grapevine wood after Hot-Water Treatment (HWT) in planting material from Czech Republic and Spain at two different times during the propagation process and from two plant zones. We examined fungal communities using both traditional isolation into culture and high-throughput amplicon sequencing (HTAS) of the internal transcribed spacer 2 (ITS2) region in extracted total RNA. HTAS from metatranscriptomic RNA increased the resolution of the fungal community analysis and revealed a highly diverse mycobiota of grapevine wood compared to the traditional method. Fungal diversity differed between grapevine genotypes and showed a temporal variation over the vegetative period. Grapevine planting materials exhibited high fungal diversity after HWT, which demonstrates that the HWT process does not sterilize the internal wood of grapevine. HWT reduced the infection caused by fungal trunk disease pathogens but was not completely effective in eliminating their growth. This study provides important and practically useful insights into the dynamics of active fungal communities in hot-water treated plants, and represents the first study of active fungal communities on grapevine grafted plants by comparing traditional and next-generation sequencing methods., This work was supported by Projects No. LD14051 from the Ministry of Education, Youth and Sports, Czech Republic covered by COST Action FA1303, and No. CZ.02.1.01/0.0/0.0/16_025/0007314 “Multidisciplinary research to increase application potential of nanomaterials in agricultural practice” from EFRR (Grant provider: Ministerstvo školství, mládeže a tělovýchovy České republiky (MŠMT). Access to computing and storage facilities owned by parties and projects contributing to the National Grid Infrastructure MetaCentrum provided under the programme "Projects of Large Research, Development, and Innovations Infrastructures" (CESNET LM2015042), is greatly appreciated. David Gramaje was supported by the DOC-INIA program from the National Institute for Agronomic Research (INIA), co-funded by the European Social Fund. The authors are grateful for the technical assistance of Jesús Alfonso Crespo and Aitor Gavara Vidal.

Timing of pruning may affect wound susceptibility to wood-invading fungi, such as those associated with grapevine trunk diseases (GTDs). This study aimed to determine the effect of pruning time on natural fungal infection in six vineyards in Galicia, Spain, during two periods of three months each, from November to February and from February to May by ITS2 high-throughput amplicon sequencing. Fungal microbiome composition did not differ significantly with year of sampling. Pruned canes harboured a core community of fungal species, which appeared to be independent of the infection period. Accumulated rainfall over 8 and 11 weeks after pruning positively correlated with the total fungal microbiome. A seasonal effect on GTD fungal infection was detected for most genera, with a higher percent detected after pruning in February (winter) as compared with that of pruning in November (mid-autumn). Early pruning is recommended to reduce the infections caused by GTD fungi in Galicia., Funding was provided by Ministerstvo Školství, Mládeže a Tělovýchovy (CZ.02.1.01./0.0/0.0/16_017/0002334).