Insight Into the Microbial Co-occurrence and Diversity of 73 Grapevine (Vitis vinifera) Crown Galls Collected Across the Northern Hemisphere

Abstract

Crown gall (CG) is a globally distributed and economically important disease of grapevine and other important crop plants. The causal agent of CG is Agrobacterium or Allorhizobium strains that harbor a tumor-inducing plasmid (pTi). The microbial community within the CG tumor has not been widely elucidated and it is not known if certain members of this microbial community promote or inhibit CG. This study investigated the microbiotas of grapevine CG tumor tissues from seven infected vineyards located in Hungary, Japan, Tunisia, and the United States. Heavy co-amplification of grapevine chloroplast and mitochondrial ribosomal RNA genes was observed with the widely used Illumina V3–V4 16S rRNA gene primers, requiring the design of a new reverse primer to enrich for bacterial 16S rRNA from CG tumors. The operational taxonomic unit (OTU) clustering approach is not suitable for CG microbiota analysis as it collapsed several ecologically distinct Agrobacterium species into a single OTU due to low interspecies genetic divergence. The CG microbial community assemblages were significantly different across sampling sites (ANOSIM global R = 0.63, p- value = 0.001) with evidence of site-specific differentially abundant ASVs. The presence of Allorhizobium vitis in the CG microbiota is almost always accompanied by Xanthomonas and Novosphingobium , the latter may promote the spread of pTi plasmid by way of acyl-homoserine lactone signal production, whereas the former may take advantage of the presence of substrates associated with plant cell wall growth and repair. The technical and biological insights gained from this study will contribute to the understanding of complex interaction between the grapevine and its microbial community and may facilitate better management of CG disease in the future.

Origin of Pathogens of Grapevine Crown Gall Disease in Hokkaido in Japan as Characterized by Molecular Epidemiology of Allorhizobium vitis Strains

“…The genetic diversity of A. vitis (Ti) isolated in some countries has previously been reported [22,36,37]. The results from cluster analysis based on repetitive sequence-based (rep)-PCR and inter-simple sequence-repeat (ISSR)-PCR data concurrently showed a potential genomic diversity that separates the Virginia strains from the Japanese strains using a total of 12 strains [22].…”

“…However, results from MLSA showed some Virginia strains and Japanese strains formed the same cluster (strain LCCH15 belonging to group A, DCCS15B to group B, ACME15 and HNVR15 to group E) (Figure 1, Table 1). Kuzmanović et al [36] reported that genetic varieties of A. vitis (Ti) of 29 strains isolated in European countries and the USA were analyzed by random amplified polymorphic DNA (RAPD) PCR, and sequence analysis of housekeeping genes of dnaK, gyrB, and recA. RAPD divided them into 17 groups, but a phylogenetic tree based on recA gene divided them into four clades [36].…”

“…Kuzmanović et al [36] reported that genetic varieties of A. vitis (Ti) of 29 strains isolated in European countries and the USA were analyzed by random amplified polymorphic DNA (RAPD) PCR, and sequence analysis of housekeeping genes of dnaK, gyrB, and recA. RAPD divided them into 17 groups, but a phylogenetic tree based on recA gene divided them into four clades [36]. It seems that PCR-based analysis tends to divide into more groups than partial sequence analysis.…”

Influence of Tourism Disturbance on Soil Microbial Community Structure in Dawei Mountain National Forest Park

“…We employed the following software for data processing: Fastp software (for quality control of the original sequencing sequence), FLASH software (for sequence splicing), UPARSE software (97% similarity) (for performing OTU clustering and chimera removal) [29,30], and RDP classifier software (using a 70% threshold to compare the fungus and bacteria database to annotate the species information for each sequence). Then, Mothur software was used to analyze and evaluate the OTU similarity level of 97% (0.97) and Student's t-test was used to calculate the difference in the α diversity index between samples.…”

Phenotypic and Molecular-Phylogenetic Analyses Reveal Distinct Features of Crown Gall-Associated Xanthomonas Strains

“…The nutrient-rich environment of crown gall tissue is a sink of amino acids and sugars or organic acids that can also be used as nutrients for the growth of several bacterial groups other than the pathogenic agrobacteria. Hence, gall tissue is a suitable environment to be colonized by epiphytic and endophytic populations of bacterial species ( 3 ).…”