eCALIBRATOR : a comparative tool to identify key genes and pathways for eucalyptus defense against biotic stressors

Many pests and pathogens threaten Eucalyptus plantations. The study of defense responses in this economically important wood and fiber crop enables the discovery of novel pathways and genes, which may be adopted to improve resistance. Various functional genomics experiments have been conducted in Eucalyptus-biotic stress interactions following the availability of the Eucalyptus grandis genome, however, comparisons between these studies were limited largely due to a lack of comparative tools. To this end, we developed eCALIBRATOR http://ecalibrator.bi.up.ac.za, a tool for the comparison of Eucalyptus biotic stress interaction. The tool, which is not limited to Eucalyptus, allows the comparison of various datasets, provides a visual output in the form of Venn diagrams and clustering and extraction of lists for gene ontology enrichment analyses. We also demonstrate the usefulness of the tool in revealing pathways and key gene targets to further functionally characterize. We identified 708 differentially expressed E. grandis genes in common among responses to the insect pest Leptocybe invasa, oomycete pathogen Phytophthora cinnamomi and fungus Chrysoporthe austroafricana. Within this set of genes, one of the Gene Ontology terms enriched was “response to organonitrogen compound,” with NITRATE TRANSPORTER 2.5 (NRT2.5) being a key gene, up-regulated under susceptible interactions and downregulated under resistant interactions. Although previous functional genetics studies in Arabidopsis thaliana support a role in nitrate acquisition and remobilization under long-term nitrate starvation, the importance of NRT2.5 in plant defense is unclear. The T-DNA mutants of AtNRT2.5 were more resistant to Pseudomonas syringae pv. tomato pv tomato DC3000 inoculation than the wild-type counterpart, supporting a direct role for NRT2.5 in plant defense. Future studies will focus on characterizing the Eucalyptus ortholog of NRT2.5

eCALIBRATOR : a comparative tool to identify key genes and pathways for Eucalyptus defense against biotic stressors

Many pests and pathogens threaten Eucalyptus plantations. The study of defense responses in this economically important wood and fiber crop enables the discovery of novel pathways and genes, which may be adopted to improve resistance. Various functional genomics experiments have been conducted in Eucalyptus-biotic stress interactions following the availability of the Eucalyptus grandis genome, however, comparisons between these studies were limited largely due to a lack of comparative tools. To this end, we developed eCALIBRATOR http://ecalibrator.bi.up.ac.za, a tool for the comparison of Eucalyptus biotic stress interaction. The tool, which is not limited to Eucalyptus, allows the comparison of various datasets, provides a visual output in the form of Venn diagrams and clustering and extraction of lists for gene ontology enrichment analyses. We also demonstrate the usefulness of the tool in revealing pathways and key gene targets to further functionally characterize. We identified 708 differentially expressed E. grandis genes in common among responses to the insect pest Leptocybe invasa, oomycete pathogen Phytophthora cinnamomi and fungus Chrysoporthe austroafricana. Within this set of genes, one of the Gene Ontology terms enriched was “response to organonitrogen compound,” with NITRATE TRANSPORTER 2.5 (NRT2.5) being a key gene, up-regulated under susceptible interactions and downregulated under resistant interactions. Although previous functional genetics studies in Arabidopsis thaliana support a role in nitrate acquisition and remobilization under longterm nitrate starvation, the importance of NRT2.5 in plant defense is unclear. The T-DNA mutants of AtNRT2.5 were more resistant to Pseudomonas syringae pv. tomato pv tomato DC3000 inoculation than the wild-type counterpart, supporting a direct role for NRT2.5 in plant defense. Future studies will focus on characterizing the Eucalyptus ortholog of NRT2.5.Supplementary Material: Figure S1 : Selection of atnrt2.5 T-DNA mutants. (A) Diagram of the AtNRT2.5 gene consisting of three exons and two introns with the positions of the T-DNA insertions in GK213H10 (AtNRT2.5-A) and GK046H04 (AtNRT2.5-B) in the second and first exons, respectively. (B,C) PCR detection of T-DNA in AtNRT2.5-A (B) and AtNRT2.5-B (C), respectively, using a combination of the T-DNA left border oligonucleotide and gene-specific oligonucleotides. No amplification was observed in AtNRT2.5-A and AtNRT2.5-B using gene-specific oligonucleotides spanning the T-DNA insertion sites.Table S1 : Summary of the mapping of RNA-seq libraries per Eucalyptus–pathogen interaction sample.Table S2 : Functionally enriched terms of the gene ontology biological processes category in common between the resistant and susceptible defense responses.Table S3 : Functionally enriched terms of the gene ontology biological processes category in the unique set of differentially expressed genes between the resistant and susceptible interactions.The Department of Science and Technology grant for Forest Genomics and Biotechnology, the South African National Research Foundation Grant for Y-rated researchers (UID105767) Incentive funding for rated researchers (UID95807), Technology and Human Resources for Industry Program (THRIP, Grant ID 96413) and the Technology Innovation Agency (TIA) Forest Molecular Genetics Cluster Program.http://www.frontiersin.org/Microbiologyam2020BiochemistryForestry and Agricultural Biotechnology Institute (FABI)GeneticsMicrobiology and Plant Patholog

The Road to Resistance in Forest Trees

In recent years, forests have been exposed to an unprecedented rise in pests and pathogens. This, coupled with the added challenge of climate change, renders forest plantation stock vulnerable to attack and severely limits productivity. Genotypes resistant to such biotic challenges are desired in plantation forestry to reduce losses. Conventional breeding has been a main avenue to obtain resistant genotypes. More recently, genetic engineering has become a viable approach to develop resistance against pests and pathogens in forest trees. Tree genomic resources have contributed to advancements in both these approaches. Genome-wide association studies and genomic selection in tree populations have accelerated breeding tools while integration of various levels of omics information facilitates the selection of candidate genes for genetic engineering. Furthermore, tree associations with non-pathogenic endophytic and subterranean microbes play a critical role in plant health and may be engineered in forest trees to improve resistance in the future. We look at recent studies in forest trees describing defense mechanisms using such approaches and propose the way forward to developing superior genotypes with enhanced resistance against biotic stress

The road to resistance in forest trees

In recent years, forests have been exposed to an unprecedented rise in pests and pathogens. This, coupled with the added challenge of climate change, renders forest plantation stock vulnerable to attack and severely limits productivity. Genotypes resistant to such biotic challenges are desired in plantation forestry to reduce losses. Conventional breeding has been a main avenue to obtain resistant genotypes. More recently, genetic engineering has become a viable approach to develop resistance against pests and pathogens in forest trees. Tree genomic resources have contributed to advancements in both these approaches. Genome-wide association studies and genomic selection in tree populations have accelerated breeding tools while integration of various levels of omics information facilitates the selection of candidate genes for genetic engineering. Furthermore, tree associations with non-pathogenic endophytic and subterranean microbes play a critical role in plant health and may be engineered in forest trees to improve resistance in the future. We look at recent studies in forest trees describing defense mechanisms using such approaches and propose the way forward to developing superior genotypes with enhanced resistance against biotic stress.The Department of Science and Technology grant for Forest Genomics and Biotechnology, the South African National Research Foundation Grant for Y-rated researchers (UID105767) Incentive funding for rated researchers (UID95807) and Technology and Human Resources for Industry Programme (THRIP, Grant ID 96413)as well as the Australian Research Council (DP160102684).http://www.frontiersin.org/Plant_Scienceam2020Forestry and Agricultural Biotechnology Institute (FABI)GeneticsMicrobiology and Plant Patholog

Chickpea Roots Undergoing Colonisation by Phytophthora medicaginis Exhibit Opposing Jasmonic Acid and Salicylic Acid Accumulation and Signalling Profiles to Leaf Hemibiotrophic Models

Hemibiotrophic pathogens cause significant losses within agriculture, threatening the sustainability of food systems globally. These microbes colonise plant tissues in three phases: a biotrophic phase followed by a biotrophic-to-necrotrophic switch phase and ending with necrotrophy. Each of these phases is characterized by both common and discrete host transcriptional responses. Plant hormones play an important role in these phases, with foliar models showing that salicylic acid accumulates during the biotrophic phase and jasmonic acid/ethylene responses occur during the necrotrophic phase. The appropriateness of this model to plant roots has been challenged in recent years. The need to understand root responses to hemibiotrophic pathogens of agronomic importance necessitates further research. In this study, using the root hemibiotroph Phytophthora medicaginis, we define the duration of each phase of pathogenesis in Cicer arietinum (chickpea) roots. Using transcriptional profiling, we demonstrate that susceptible chickpea roots display some similarities in response to disease progression as previously documented in leaf plant–pathogen hemibiotrophic interactions. However, our transcriptomic results also show that chickpea roots do not conform to the phytohormone responses typically found in leaf colonisation by hemibiotrophs. We found that quantified levels of salicylic acid concentrations in root tissues decreased significantly during biotrophy while jasmonic acid concentrations were significantly induced. This study demonstrated that a wider spectrum of plant species should be investigated in the future to understand the physiological changes in plants during colonisation by soil-borne hemibiotrophic pathogens before we can better manage these economically important microbes

Abscisic acid supports colonization of Eucalyptus grandis roots by the mutualistic ectomycorrhizal fungus Pisolithus microcarpus

The pathways regulated in ectomycorrhizal (EcM) plant hosts during the establishment of symbiosis are not as well understood when compared to the functional stages of this mutualistic interaction. Our study used the EcM host Eucalyptus grandis to elucidate symbiosis-regulated pathways across the three phases of this interaction. Using a combination of RNA sequencing and metabolomics we studied both stage-specific and core responses of E. grandis during colonization by Pisolithus microcarpus. Using exogenous manipulation of the abscisic acid (ABA), we studied the role of this pathway during symbiosis establishment. Despite the mutualistic nature of this symbiosis, a large number of disease signalling TIRNBS-LRR genes were induced. The transcriptional regulation in E. grandis was found to be dynamic across colonization with a small core of genes consistently regulated at all stages. Genes associated to the carotenoid/ABA pathway were found within this core and ABA concentrations increased during fungal integration into the root. Supplementation of ABA led to improved accommodation of P. microcarpus into E. grandis roots. The carotenoid pathway is a core response of an EcM host to its symbiont and highlights the need to understand the role of the stress hormone ABA in controlling host–EcM fungal interactions