Comparative Analysis of Transcriptomes of Ophiostoma novo-ulmi ssp. americana Colonizing Resistant or Sensitive Genotypes of American Elm

Abstract

The Ascomycete Ophiostoma novo-ulmi threatens elm populations worldwide. The molecular mechanisms underlying its pathogenicity and virulence are still largely uncharacterized. As part of a collaborative study of the O. novo-ulmi-elm interactome, we analyzed the O. novo-ulmi ssp. americana transcriptomes obtained by deep sequencing of messenger RNAs recovered from Ulmus americana saplings from one resistant (Valley Forge, VF) and one susceptible (S) elm genotypes at 0 and 96 h post-inoculation (hpi). Transcripts were identified for 6424 of the 8640 protein-coding genes annotated in the O. novo-ulmi nuclear genome. A total of 1439 genes expressed in planta had orthologs in the PHI-base curated database of genes involved in host-pathogen interactions, whereas 472 genes were considered differentially expressed (DEG) in S elms (370 genes) and VF elms (102 genes) at 96 hpi. Gene ontology (GO) terms for processes and activities associated with transport and transmembrane transport accounted for half (27/55) of GO terms that were significantly enriched in fungal genes upregulated in S elms, whereas the 22 GO terms enriched in genes overexpressed in VF elms included nine GO terms associated with metabolism, catabolism and transport of carbohydrates. Weighted gene co-expression network analysis identified three modules that were significantly associated with higher gene expression in S elms. The three modules accounted for 727 genes expressed in planta and included 103 DEGs upregulated in S elms. Knockdown- and knockout mutants were obtained for eight O. novo-ulmi genes. Although mutants remained virulent towards U. americana saplings, we identified a large repertoire of additional candidate O. novo-ulmi pathogenicity genes for functional validation by loss-of-function approaches.

Independent Evolution Has Led to Distinct Genomic Signatures in Dutch Elm Disease-Causing Fungi and Other Vascular Wilts-Causing Fungal Pathogens

“…O. novo-ulmi has 15 members in family OG0000010, 8 members in each family OG0000011 and OG0000017, and only one current member of family OG0000093. We checked the transcript expression using previously published experimental in vitro and in planta studies [ 65 , 66 , 67 ] and one unpublished results study (T. de Oliveira, unpublished). One member of family OG0000010 and two members of family OG0000017 had a high or medium expression level in vitro in both yeast and mycelium growth phases ( Table S5 ).…”

“…The sequencing and annotation of genomes of O. ulmi [ 73 ] and O. novo-ulmi [ 38 , 74 ] opened the way for transcriptomic approaches to complement previous strategies used for unraveling the molecular bases of various biological traits in the DED fungi. Thus, the genes and pathways associated with yeast-mycelium dimorphism in vitro [ 65 , 66 ] and pathogenicity in planta [ 67 ] were identified and are now being validated by loss-of-function assays with targeted deletion mutants [ 75 ]. The wilt-causing fungi have so far been recorded in orders Hypocreales, Microascales, Glomerellales, and Ophiostomatales within the class Sordariomycetes.…”

“…Our previous experimental studies [ 65 , 66 , 67 ] and unpublished results allowed us to examine some of the gene families unique or enriched in wilt species that we found in this study, by looking at their expression in Ophiostoma novo-ulmi and other DED species after in vitro or in planta infections. A few of them proved to be good candidates for causing virulence in elms, by showing elevated expression in planta after infection and represent potential candidates for future loss-of-function studies.…”

“…The following supporting information can be downloaded at: , Figure S1: Gene counts for the most extreme expansions and contractions of gene classes in seven wilt groups; Figure S2: Presence/absence of orthologs of 10 Verticillium dahliae genes identified by [ 27 ] as unique to Fusarium and Verticillium wilt species; Table S1: List of species and strains used in this study; Table S2: Results of phylogenetic logistic regression predicting wilt ( n = 20) vs. non-wilt pathogens ( n = 19) based on the abundance of different gene families/groups; Table S3: Gene counts for gene families/groups in pathogenic species from four orders; Table S4: Inferred ancestral gene counts, fold changes and within-species raw gene counts for gene families/groups in seven wilt groups; Table S5: Number of transcripts in vitro and in planta for genes present in wilt-causing Ophiostoma novo-ulmi (Onu) and Ophiostoma spp. but absent/rare in other wilt-causing species [ 29 , 65 , 66 , 67 , 74 ]; Table S6: Gene homologs shared by different pairs of wilt groups.…”

Unraveling the transcriptional features and gene expression networks of pathogenic and saprotrophic Ophiostoma species during the infection of Ulmus americana

“…Pioneering works on elm- O. novo-ulmi interactions ( 36 , 37 ) allowed the recovery of transcripts for less than 200 fungal genes and were therefore not very informative regarding the biology of the pathogen. Nigg et al ( 38 ) presented the first comprehensive analysis of transcriptomes of O. novo-ulmi subsp. americana following inoculation of one susceptible and one resistant (Valley Forge) clone of Ulmus americana .…”