Belowground microbiota analysis indicates that Fusarium spp. exacerbate grapevine trunk disease

Abstract

Background: Grapevine trunk diseases (GTDs) are disease complexes that are major threats to viticulture in most grapevine growing regions. The microbiomes colonizing plant belowground components form complex associations with plants, play important roles in promoting plant productivity and health in natural environments, and may be related to GTD development. To investigate associations between belowground fungal communities and GTD symptomatic or asymptomatic grapevines, fungal communities associated with three soil-plant compartments (bulk soils, rhizospheres, and roots) were characterized by ITS high-throughput amplicon sequencing across two years.

Results: The fungal community diversity and composition differs according to the soil-plant compartment type (PERMANOVA, p < 0.001, 12.04% of variation explained) and sampling year (PERMANOVA, p < 0.001, 8.83%), whereas GTD symptomatology exhibited a weaker, but still significant association (PERMANOVA, p < 0.001, 1.29%). The effects of the latter were particularly prominent in root and rhizosphere community comparisons. Many GTD-associated pathogens were detected, but their relative abundances were not correlated (or were negatively correlated) to symptomatology. Fusarium spp., were enriched in symptomatic roots and rhizospheres compared to asymptomatic counterparts, suggesting that their abundances were positively correlated with symptomatic vines. Inoculation tests revealed that Fusarium isolates, similar to Dactylonectria macrodidyma, a pathogen associated with black foot disease, caused dark brown necrotic spots on stems in addition to root rot, which blackened lateral roots. Disease indices were higher with co-inoculation than single inoculation with a Fusarium isolate or D. macrodidyma, suggesting that Fusarium spp. can exacerbate disease severity when inoculated with other known GTD-associated pathogens.

Conclusions: The belowground fungal microbiota of grapevines varied from soil-plant compartments, the years and whether showed GTD symptoms. The GTDs symptoms were related to the enrichment of Fusarium spp. rather than the relative abundances of GTD pathogens. These results demonstrate the effects of fungal microbiota of roots and rhizospheres on GTDs, while providing new insights into opportunistic pathogenesis of GTDs and potential control practices.

Keywords: Belowground; Fungi; Fusarium spp.; Grapevine trunk diseases; Microbiota.

Fig. 1

Differences in relative abundances of fungi in the belowground components of grapevines between 2019 and 2020. A and B The relative abundances of the top 10 most abundant fungal taxa at the family and genus levels, respectively. The less abundant taxa are referred to as “others”. C shows genera with significant differences in relative abundance between 2019 and 2020. t test, p < 0.05

Fig. 2

Principal coordinate analysis (PCoA) based on Bray–Curtis dissimilarity metrics. A–C The distances in the fungal communities of roots, rhizospheres, and bulk soils, respectively, between the sampling years of 2019 and 2020. The distances of different soil–plant compartments in fungal communities collected in 2019 and 2020 are shown in (D) and (E), respectively. The PERMANOVA test was used for statistical analysis

Fig. 3

Principal coordinate analysis (PCoA) based on Bray–Curtis dissimilarity metrics showing the distance in the fungal communities of asymptomatic and symptomatic roots collected in 2019 (A), asymptomatic and symptomatic rhizospheres collected in 2019 (B), asymptomatic and symptomatic bulk soils collected in 2019 (C), asymptomatic and symptomatic roots collected in 2020 (D), asymptomatic and symptomatic rhizospheres collected in 2020 (E), and asymptomatic and symptomatic bulk soils collected in 2020 (F). The PERMANOVA test was used for statistical analysis

Fig. 4

Linear discriminant analysis effect size (LEfSe) analysis of fungal taxa enrichment among treatments. Fungal biomarker enrichment among AsPE (the rhizoplane and endophyte fungi for samples collected from asymptomatic grapevines) and SPE (the rhizoplane and endophyte fungi for samples collected from symptomatic grapevines) with LDA values > 3 are shown in (A), and asymptomatic and symptomatic rhizosphere soils with LDA values > 4 are shown in (B). p, phylum; c, class; o, order; f, family; g, genus; s, species. LDA, linear discriminant analysis

Fig. 5

Relative abundances of grapevine trunk disease (GTD)-associated fungi detected in this study based on ITS amplicon sequencing at the species (A) and genus (B) levels

Fig. 6

Relative abundance differences of grapevine trunk disease (GTD)-associated fungi at the species (A) and genus (B) levels between comparable groups are indicated in the upper right side of the figure. Grey indicates no significant difference in the relative abundance of a certain species or genus between the groups being compared, while green and yellow indicate significant differences. Green indicates that the relative abundance of the former group is significantly higher than that of the latter one, while yellow indicates the opposite. For example, when AsPE.19 (the rhizoplane and endophyte fungi for samples collected from asymptomatic grapevines in 2019) was compared with AsPE.20 (the rhizoplane and endophyte fungi for samples collected from asymptomatic grapevines in 2020) (AsPE.19 vs. AsPE.20), the relative abundance of Dactylonectria torresensis in the former group was significantly lower than that in the latter group (indicated in yellow). The relative abundance of Cadophora luteo-olivacea was significantly higher in the former group than in the latter one (indicated in green). Statistical differences are indicated by q-values < 0.05 based on Wilcoxon tests

Fig. 7

Grapevine stem symptoms after inoculation with Fusarium and Dactylonectria macrodidyma isolates. A Control, B Fusarium sp. JZB3110090; C Fusarium sp. JZB3110089; D Fusarium sp. JZB3110102; E Fusarium sp. JZB3110103; F Fusarium sp. JZB3110180; G D. macrodidyma JZB3310008 associated with Black Foot Disease; H Fusarium sp. JZB3110090 + D. macrodidyma JZB3310008; I Fusarium sp. JZB3110089 + D. macrodidyma JZB3310008; J Fusarium sp. JZB3110102 + D. macrodidyma JZB3310008, K Fusarium sp. JZB3110103 + D. macrodidyma JZB3310008, L Fusarium sp. JZB3110180 + D. macrodidyma JZB3310008. The red arrow shows dark brown necrotic spots