Dual RNA-Sequencing of Eucalyptus nitens during Phytophthora cinnamomi Challenge Reveals Pathogen and Host Factors Influencing Compatibility

Abstract

Damage caused by Phytophthora cinnamomi Rands remains an important concern on forest tree species. The pathogen causes root and collar rot, stem cankers, and dieback of various economically important Eucalyptus spp. In South Africa, susceptible cold tolerant Eucalyptus plantations have been affected by various Phytophthora spp. with P. cinnamomi considered one of the most virulent. The molecular basis of this compatible interaction is poorly understood. In this study, susceptible Eucalyptus nitens plants were stem inoculated with P. cinnamomi and tissue was harvested five days post inoculation. Dual RNA-sequencing, a technique which allows the concurrent detection of both pathogen and host transcripts during infection, was performed. Approximately 1% of the reads mapped to the draft genome of P. cinnamomi while 78% of the reads mapped to the Eucalyptus grandis genome. The highest expressed P. cinnamomi gene in planta was a putative crinkler effector (CRN1). Phylogenetic analysis indicated the high similarity of this P. cinnamomi CRN1 to that of Phytophthora infestans. Some CRN effectors are known to target host nuclei to suppress defense. In the host, over 1400 genes were significantly differentially expressed in comparison to mock inoculated trees, including suites of pathogenesis related (PR) genes. In particular, a PR-9 peroxidase gene with a high similarity to a Carica papaya PR-9 ortholog previously shown to be suppressed upon infection by Phytophthora palmivora was down-regulated two-fold. This PR-9 gene may represent a cross-species effector target during P. cinnamomi infection. This study identified pathogenicity factors, potential manipulation targets, and attempted host defense mechanisms activated by E. nitens that contributed to the susceptible outcome of the interaction.

Phytophthora cinnamomi

“…If the N-terminal truncation of four additional proteins is caused by assembly errors, these proteins may also be secreted, giving a total of 29 putative P. cinnamomi CRN effectors. A P. cinnamomi CRN homologue, shown in the current genome assembly to be truncated at the N-terminus, is the most highly expressed P. cinnamomi gene 5 days after inoculation of Eucalyptus nitens (Meyer et al, 2016). Thirteen of the 42 P. cinnamomi CRN homologues have a nuclear localization signal (NLS) in the C-terminal region (Table S3).…”

“…These unannotated data have been assembled into about 4000 contigs. At the time of writing, there were two published analyses of P. cinnamomi transcriptomes available (Meyer et al, 2016;Reitmann et al, 2016). The first reports data from Eucalyptus nitens, 5 days after inoculation with P. cinnamomi; the second reports data from a library of cysts and germinated cysts.…”

Time-resolved dual root-microbe transcriptomics reveals early induced Nicotiana benthamiana genes and conserved infection-promoting Phytophthora palmivora effectors

“…Finally, the N. benthamiana response to P. palmivora also includes upregulation of genes encoding late embryogenesis abundant (LEA) proteins as well as heat shock proteins. LEA proteins have been associated with the drought response [93,94] and upregulation of genes associated with water deprivation upon Phytophthora infection has been previously reported [77]. Conversely, downregulated genes were mostly associated with photosynthesis, cellulose biosynthesis and cell division.…”

“…So far, dual transcriptomics has only been used with limited time resolution and sequencing depth in plant-pathogenic oomycete studies [76,77]. Our study encompasses the full range of P. palmivora sequential lifestyle transitions occurring in N. benthamiana root, allowing reconstruction of comprehensive transcriptional landscape in both interacting organisms.…”

Las técnicas de secuenciación masiva en el estudio de la diversidad biológica

“…Esta aproximación se ha llevado a cabo fundamentalmente para la detección de hongos pató-genos en plantas (Hayden et al, 2013;Meyer et al, 2016). Usualmente, el RNA-seq dual se beneficia de la mayor facilidad de contar con un genoma de referencia para el patógeno en cuestión, de modo que se pueden separar los genes del huésped y el hospedante para así poder investigar los cambios en los patrones de expresión de diversos genes y relacionarlos con la virulencia del ataque.…”