Combination of linkage and association mapping with genomic prediction to infer QTL regions associated with gray leaf spot and northern corn leaf blight resistance in tropical maize.

Among the diseases threatening maize production in Africa are gray leaf spot (GLS) caused by Cercospora zeina and northern corn leaf blight (NCLB) caused by Exserohilum turcicum. The two pathogens, which have high genetic diversity, reduce the photosynthesizing ability of susceptible genotypes and, hence, reduce the grain yield. To identify population-based quantitative trait loci (QTLs) for GLS and NCLB resistance, a biparental population of 230 lines derived from the tropical maize parents CML511 and CML546 and an association mapping panel of 239 tropical and sub-tropical inbred lines were phenotyped across multi-environments in western Kenya. Based on 1,264 high-quality polymorphic single-nucleotide polymorphisms (SNPs) in the biparental population, we identified 10 and 18 QTLs, which explained 64.2% and 64.9% of the total phenotypic variance for GLS and NCLB resistance, respectively. A major QTL for GLS, qGLS1_186 accounted for 15.2% of the phenotypic variance, while qNCLB3_50 explained the most phenotypic variance at 8.8% for NCLB resistance. Association mapping with 230,743 markers revealed 11 and 16 SNPs significantly associated with GLS and NCLB resistance, respectively. Several of the SNPs detected in the association panel were co-localized with QTLs identified in the biparental population, suggesting some consistent genomic regions across genetic backgrounds. These would be more relevant to use in field breeding to improve resistance to both diseases. Genomic prediction models trained on the biparental population data yielded average prediction accuracies of 0.66-0.75 for the disease traits when validated in the same population. Applying these prediction models to the association panel produced accuracies of 0.49 and 0.75 for GLS and NCLB, respectively. This research conducted in maize fields relevant to farmers in western Kenya has combined linkage and association mapping to identify new QTLs and confirm previous QTLs for GLS and NCLB resistance. Overall, our findings imply that genetic gain can be improved in maize breeding for resistance to multiple diseases including GLS and NCLB by using genomic selection.

Population genomic analyses suggest recent dispersal events of the pathogen Cercospora zeina into East and Southern African maize cropping systems.

A serious factor hampering global maize production is gray leaf spot disease. Cercospora zeina is one of the causative pathogens, but population genomics analysis of C. zeina is lacking. We conducted whole-genome Illumina sequencing of a representative set of 30 C. zeina isolates from Kenya and Uganda (East Africa) and Zambia, Zimbabwe, and South Africa (Southern Africa). Selection of the diverse set was based on microsatellite data from a larger collection of the pathogen. Pangenome analysis of the C. zeina isolates was done by (1) de novo assembly of the reads with SPAdes, (2) annotation with BRAKER, and (3) protein clustering with OrthoFinder. A published long-read assembly of C. zeina (CMW25467) from Zambia was included and annotated using the same pipeline. This analysis revealed 790 non-shared accessory and 10,677 shared core orthogroups (genes) between the 31 isolates. Accessory gene content was largely shared between isolates from all countries, with a few genes unique to populations from Southern Africa (32) or East Africa (6). There was a significantly higher proportion of effector genes in the accessory secretome (44%) compared to the core secretome (24%). PCA, ADMIXTURE, and phylogenetic analysis using a neighbor-net network indicated a population structure with a geographical subdivision between the East African isolates and the Southern African isolates, although gene flow was also evident. The small pangenome and partial population differentiation indicated recent dispersal of C. zeina into Africa, possibly from 2 regional founder populations, followed by recurrent gene flow owing to widespread maize production across sub-Saharan Africa.

Deep Learning Diagnostics of Gray Leaf Spot in Maize under Mixed Disease Field Conditions.

Maize yields worldwide are limited by foliar diseases that could be fungal, oomycete, bacterial, or viral in origin. Correct disease identification is critical for farmers to apply the correct control measures, such as fungicide sprays. Deep learning has the potential for automated disease classification from images of leaf symptoms. We aimed to develop a classifier to identify gray leaf spot (GLS) disease of maize in field images where mixed diseases were present (18,656 images after augmentation). In this study, we compare deep learning models trained on mixed disease field images with and without background subtraction. Performance was compared with models trained on PlantVillage images with single diseases and uniform backgrounds. First, we developed a modified VGG16 network referred to as "GLS_net" to perform binary classification of GLS, which achieved a 73.4% accuracy. Second, we used MaskRCNN to dynamically segment leaves from backgrounds in combination with GLS_net to identify GLS, resulting in a 72.6% accuracy. Models trained on PlantVillage images were 94.1% accurate at GLS classification with the PlantVillage testing set but performed poorly with the field image dataset (55.1% accuracy). In contrast, the GLS_net model was 78% accurate on the PlantVillage testing set. We conclude that deep learning models trained with realistic mixed disease field data obtain superior degrees of generalizability and external validity when compared to models trained using idealized datasets.

Population genetic structure and migration patterns of the maize pathogenic fungus, Cercospora zeina in East and Southern Africa.

Cercospora zeina is a causal pathogen of gray leaf spot (GLS) disease of maize in Africa. This fungal pathogen exhibits a high genetic diversity in South Africa. However, little is known about the pathogen's population structure in the rest of Africa. In this study, we aimed to assess the diversity and gene flow of the pathogen between major maize producing countries in East and Southern Africa (Kenya, Uganda, Zambia, Zimbabwe, and South Africa). A total of 964 single-spore isolates were made from GLS lesions and confirmed as C.zeina using PCR diagnostics. The other causal agent of GLS, Cercospora zeae-maydis, was absent. Genotyping all the C.zeina isolates with 11 microsatellite markers and a mating-type gene diagnostic revealed (i) high genetic diversity with some population structure between the five African countries, (ii) cryptic sexual recombination, (iii) that South Africa and Kenya were the greatest donors of migrants, and (iv) that Zambia had a distinct population. We noted evidence of human-mediated long-distance dispersal, since four haplotypes from one South African site were also present at five sites in Kenya and Uganda. There was no evidence for a single-entry point of the pathogen into Africa. South Africa was the most probable origin of the populations in Kenya, Uganda, and Zimbabwe. Continuous annual maize production in the tropics (Kenya and Uganda) did not result in greater genetic diversity than a single maize season (Southern Africa). Our results will underpin future management of GLS in Africa through effective monitoring of virulent C.zeina strains.

De novo Assembly of Transcriptomes From a B73 Maize Line Introgressed With a QTL for Resistance to Gray Leaf Spot Disease Reveals a Candidate Allele of a Lectin Receptor-Like Kinase.

Gray leaf spot (GLS) disease in maize, caused by the fungus Cercospora zeina, is a threat to maize production globally. Understanding the molecular basis for quantitative resistance to GLS is therefore important for food security. We developed a de novo assembly pipeline to identify candidate maize resistance genes. Near-isogenic maize lines with and without a QTL for GLS resistance on chromosome 10 from inbred CML444 were produced in the inbred B73 background. The B73-QTL line showed a 20% reduction in GLS disease symptoms compared to B73 in the field (p = 0.01). B73-QTL leaf samples from this field experiment conducted under GLS disease pressure were RNA sequenced. The reads that did not map to the B73 or C. zeina genomes were expected to contain novel defense genes and were de novo assembled. A total of 141 protein-coding sequences with B73-like or plant annotations were identified from the B73-QTL plants exposed to C. zeina. To determine whether candidate gene expression was induced by C. zeina, the RNAseq reads from C. zeina-challenged and control leaves were mapped to a master assembly of all of the B73-QTL reads, and differential gene expression analysis was conducted. Combining results from both bioinformatics approaches led to the identification of a likely candidate gene, which was a novel allele of a lectin receptor-like kinase named L-RLK-CML that (i) was induced by C. zeina, (ii) was positioned in the QTL region, and (iii) had functional domains for pathogen perception and defense signal transduction. The 817AA L-RLK-CML protein had 53 amino acid differences from its 818AA counterpart in B73. A second "B73-like" allele of L-RLK was expressed at a low level in B73-QTL. Gene copy-specific RT-qPCR confirmed that the l-rlk-cml transcript was the major product induced four-fold by C. zeina. Several other expressed defense-related candidates were identified, including a wall-associated kinase, two glutathione s-transferases, a chitinase, a glucan beta-glucosidase, a plasmodesmata callose-binding protein, several other receptor-like kinases, and components of calcium signaling, vesicular trafficking, and ethylene biosynthesis. This work presents a bioinformatics protocol for gene discovery from de novo assembled transcriptomes and identifies candidate quantitative resistance genes.

Influence of farming practices on the population genetics of the maize pathogen Cercospora zeina in South Africa.

Gray leaf spot (GLS) is an important foliar disease of maize. This disease, caused by Cercospora zeina, is prevalent in both smallholder and commercial maize farms in South Africa. Notably, smallholder practices are geared towards conservation agriculture, planting diverse maize genotypes within a field and avoiding chemical control. This study examined the population genetic structure of 129 C. zeina isolates from three smallholder farm sites in KwaZulu-Natal in South Africa using 13 microsatellite markers. These were analysed, together with 239 isolates previously analysed from four commercial farms in the same province, to determine whether farming systems influence the genetic diversity of C.zeina. In addition, we wanted to determine whether the smallholder farming system harboured a greater diversity of C.zeina haplotypes due to lack of chemical spraying of these crops. Overall, farming systems exhibited partial, but significant, population differentiation, contributing 10% of the genetic variation observed. A 16% genetic variation conferred between KwaNxamalala (smallholder) and Cedara (commercial) areas that are in close proximity, confirmed this. Private alleles accounted for 29% of the 52 alleles observed in smallholder farms. Smallholder farms harboured a higher gene and genotypic diversity, with a clonal fraction of only 13% compared to 32% in commercial farms. Mating type ratios indicative of sexual recombination and lower linkage disequilibrium in most smallholder populations were consistent with higher levels of diversity. This study suggests that commercial farming practices, such as fungicides and monoculture crop planting, may result in a narrower genetic diversity of the pathogen that is then propagated by asexual reproduction. In contrast, management of GLS disease in smallholder farms should consider the greater diversity of pathogen genotypes, especially if future research shows that this equates to a greater diversity of pathogenicity alleles.

Application of Chloroplast Phylogenomics to Resolve Species Relationships Within the Plant Genus Amaranthus.

Amaranthus species are an emerging and promising nutritious traditional vegetable food source. Morphological plasticity and poorly resolved dendrograms have led to the need for well resolved species phylogenies. We hypothesized that whole chloroplast phylogenomics would result in more reliable differentiation between closely related amaranth species. The aims of the study were therefore: to construct a fully assembled, annotated chloroplast genome sequence of Amaranthus tricolor; to characterize Amaranthus accessions phylogenetically by comparing barcoding genes (matK, rbcL, ITS) with whole chloroplast sequencing; and to use whole chloroplast phylogenomics to resolve deeper phylogenetic relationships. We generated a complete A. tricolor chloroplast sequence of 150,027 bp. The three barcoding genes revealed poor inter- and intra-species resolution with low bootstrap support. Whole chloroplast phylogenomics of 59 Amaranthus accessions increased the number of parsimoniously informative sites from 92 to 481 compared to the barcoding genes, allowing improved separation of amaranth species. Our results support previous findings that two geographically independent domestication events of Amaranthus hybridus likely gave rise to several species within the Hybridus complex, namely Amaranthus dubius, Amaranthus quitensis, Amaranthus caudatus, Amaranthus cruentus and Amaranthus hypochondriacus. Poor resolution of species within the Hybridus complex supports the recent and ongoing domestication within the complex, and highlights the limitation of chloroplast data for resolving recent evolution. The weedy Amaranthus retroflexus and Amaranthus powellii was found to share a common ancestor with the Hybridus complex. Leafy amaranth, Amaranthus tricolor, Amaranthus blitum, Amaranthus viridis and Amaranthus graecizans formed a stable sister lineage to the aforementioned species across the phylogenetic trees. This study demonstrates the power of next-generation sequencing data and reference-based assemblies to resolve phylogenies, and also facilitated the identification of unknown Amaranthus accessions from a local genebank. The informative phylogeny of the Amaranthus genus will aid in selecting accessions for breeding advanced genotypes to satisfy global food demand.

IMA Genome-F 8: Draft genome of Cercospora zeina, Fusarium pininemorale, Hawksworthiomyces lignivorus, Huntiella decipiens and Ophiostoma ips.

The genomes of Cercospora zeina, Fusarium pininemorale, Hawksworthiomyces lignivorus, Huntiella decipiens, and Ophiostoma ips are presented in this genome announcement. Three of these genomes are from plant pathogens and otherwise economically important fungal species. Fusarium pininemorale and H. decipiens are not known to cause significant disease but are closely related to species of economic importance. The genome sizes range from 25.99 Mb in the case of O. ips to 4.82 Mb for H. lignivorus. These genomes include the first reports of a genome from the genus Hawksworthiomyces. The availability of these genome data will allow the resolution of longstanding questions regarding the taxonomy of these species. In addition these genome sequences through comparative studies with closely related organisms will increase our understanding of how these species or close relatives cause disease.

RNA-Seq analysis of resistant and susceptible sub-tropical maize lines reveals a role for kauralexins in resistance to grey leaf spot disease, caused by Cercospora zeina.

BACKGROUND: Cercospora zeina is a foliar pathogen responsible for maize grey leaf spot in southern Africa that negatively impacts maize production. Plants use a variety of chemical and structural mechanisms to defend themselves against invading pathogens such as C. zeina, including the production of secondary metabolites with antimicrobial properties. In maize, a variety of biotic and abiotic stressors induce the accumulation of the terpenoid phytoalexins, zealexins and kauralexins. RESULTS: C. zeina-susceptible line displayed pervasive rectangular grey leaf spot lesions, running parallel with the leaf veins in contrast to C. zeina-resistant line that had restricted disease symptoms. Analysis of the transcriptome of both lines indicated that genes involved in primary and secondary metabolism were up-regualted, and although different pathways were prioritized in each line, production of terpenoid compounds were common to both. Targeted phytoalexin analysis revealed that C. zeina-inoculated leaves accumulated zealexins and kauralexins. The resistant line shows a propensity toward accumulation of the kauralexin B series metabolites in response to infection, which contrasts with the susceptible line that preferentially accumulates the kauralexin A series. Kauralexin accumulation was correlated to expression of the kauralexin biosynthetic gene, ZmAn2 and a candidate biosynthetic gene, ZmKSL2. We report the expression of a putative copalyl diphosphate synthase gene that is induced by C. zeina in the resistant line exclusively. DISCUSSION: This study shows that zealexins and kauralexins, and expression of their biosynthetic genes, are induced by C. zeina in both resistant and susceptible germplasm adapted to the southern African climate. The data presented here indicates that different forms of kauralexins accumulate in the resistant and susceptible maize lines in response to C. zeina, with the accumulation of kauralexin B compounds in a resistant maize line and kauralexin A compounds accumulating in the susceptible line.

Complementation of CTB7 in the Maize Pathogen Cercospora zeina Overcomes the Lack of In Vitro Cercosporin Production.

Gray leaf spot (GLS), caused by the sibling species Cercospora zeina or Cercospora zeae-maydis, is cited as one of the most important diseases threatening global maize production. C. zeina fails to produce cercosporin in vitro and, in most cases, causes large coalescing lesions during maize infection, a symptom generally absent from cercosporin-deficient mutants in other Cercospora spp. Here, we describe the C. zeina cercosporin toxin biosynthetic (CTB) gene cluster. The oxidoreductase gene CTB7 contained several insertions and deletions as compared with the C. zeae-maydis ortholog. We set out to determine whether complementing the defective CTB7 gene with the full-length gene from C. zeae-maydis could confer in vitro cercosporin production. C. zeina transformants containing C. zeae-maydis CTB7 were generated by Agrobacterium tumefaciens-mediated transformation and were evaluated for in vitro cercosporin production. When grown on nitrogen-limited medium in the light-conditions conducive to cercosporin production in other Cercospora spp.-one transformant accumulated a red pigment that was confirmed to be cercosporin by the KOH assay, thin-layer chromatography, and ultra performance liquid chromatography-quadrupole-time-of-flight mass spectrometry. Our results indicated that C. zeina has a defective CTB7, but all other necessary machinery required for synthesizing cercosporin-like molecules and, thus, C. zeina may produce a structural variant of cercosporin during maize infection.

Systems genetics reveals a transcriptional network associated with susceptibility in the maize-grey leaf spot pathosystem.

We used a systems genetics approach to elucidate the molecular mechanisms of the responses of maize to grey leaf spot (GLS) disease caused by Cercospora zeina, a threat to maize production globally. Expression analysis of earleaf samples in a subtropical maize recombinant inbred line population (CML444 × SC Malawi) subjected in the field to C. zeina infection allowed detection of 20 206 expression quantitative trait loci (eQTLs). Four trans-eQTL hotspots coincided with GLS disease QTLs mapped in the same field experiment. Co-expression network analysis identified three expression modules correlated with GLS disease scores. The module (GY-s) most highly correlated with susceptibility (r = 0.71; 179 genes) was enriched for the glyoxylate pathway, lipid metabolism, diterpenoid biosynthesis and responses to pathogen molecules such as chitin. The GY-s module was enriched for genes with trans-eQTLs in hotspots on chromosomes 9 and 10, which also coincided with phenotypic QTLs for susceptibility to GLS. This transcriptional network has significant overlap with the GLS susceptibility response of maize line B73, and may reflect pathogen manipulation for nutrient acquisition and/or unsuccessful defence responses, such as kauralexin production by the diterpenoid biosynthesis pathway. The co-expression module that correlated best with resistance (TQ-r; 1498 genes) was enriched for genes with trans-eQTLs in hotspots coinciding with GLS resistance QTLs on chromosome 9. Jasmonate responses were implicated in resistance to GLS through co-expression of COI1 and enrichment of genes with the Gene Ontology term 'cullin-RING ubiquitin ligase complex' in the TQ-r module. Consistent with this, JAZ repressor expression was highly correlated with the severity of GLS disease in the GY-s susceptibility network.

Cercospora zeina from Maize in South Africa Exhibits High Genetic Diversity and Lack of Regional Population Differentiation.

South Africa is one of the leading maize-producing countries in sub-Saharan Africa. Since the 1980s, Cercospora zeina, a causal agent of gray leaf spot of maize, has become endemic in South Africa, and is responsible for substantial yield reductions. To assess genetic diversity and population structure of C. zeina in South Africa, 369 isolates were collected from commercial maize farms in three provinces (KwaZulu-Natal, Mpumalanga, and North West). These isolates were evaluated with 14 microsatellite markers and species-specific mating type markers that were designed from draft genome sequences of C. zeina isolates from Africa (CMW 25467) and the United States (USPA-4). Sixty alleles were identified across 14 loci, and gene diversity values within each province ranged from 0.18 to 0.35. High levels of gene flow were observed (Nm = 5.51), and in a few cases, identical multilocus haplotypes were found in different provinces. Overall, 242 unique multilocus haplotypes were identified with a low clonal fraction of 34%. No distinct population clusters were identified using STRUCTURE, principal coordinate analysis, or Weir's theta θ statistic. The lack of population differentiation was supported by analysis of molecular variance tests, which indicated that only 2% of the variation was attributed to variability between populations from each province. Mating type ratios of MAT1-1 and MAT1-2 idiomorphs from 335 isolates were not significantly different from a 1:1 ratio in all provinces, which provided evidence for sexual reproduction. The draft genome of C. zeina CMW 25467 exhibited a complete genomic copy of the MAT1-1 idiomorph as well as exonic fragments of MAT genes from both idiomorphs. The high level of gene diversity, shared haplotypes at different geographical locations within South Africa, and presence of both MAT idiomorphs at all sites indicates widespread dispersal of C. zeina between maize fields in the country as well as evidence for sexual recombination. The outcomes of this genome-enabled study are important for disease management since the high diversity has implications for dispersal of fungicide resistance should it emerge and the need for diversified resistance breeding.

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

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.

Genome-wide mapping of histone H3 lysine 4 trimethylation in Eucalyptus grandis developing xylem.

BACKGROUND: Histone modifications play an integral role in plant development, but have been poorly studied in woody plants. Investigating chromatin organization in wood-forming tissue and its role in regulating gene expression allows us to understand the mechanisms underlying cellular differentiation during xylogenesis (wood formation) and identify novel functional regions in plant genomes. However, woody tissue poses unique challenges for using high-throughput chromatin immunoprecipitation (ChIP) techniques for studying genome-wide histone modifications in vivo. We investigated the role of the modified histone H3K4me3 (trimethylated lysine 4 of histone H3) in gene expression during the early stages of wood formation using ChIP-seq in Eucalyptus grandis, a woody biomass model. RESULTS: Plant chromatin fixation and isolation protocols were optimized for developing xylem tissue collected from field-grown E. grandis trees. A "nano-ChIP-seq" procedure was employed for ChIP DNA amplification. Over 9 million H3K4me3 ChIP-seq and 18 million control paired-end reads were mapped to the E. grandis reference genome for peak-calling using Model-based Analysis of ChIP-Seq. The 12,177 significant H3K4me3 peaks identified covered ~1.5% of the genome and overlapped some 9,623 protein-coding genes and 38 noncoding RNAs. H3K4me3 library coverage, peaking ~600 - 700 bp downstream of the transcription start site, was highly correlated with gene expression levels measured with RNA-seq. Overall, H3K4me3-enriched genes tended to be less tissue-specific than unenriched genes and were overrepresented for general cellular metabolism and development gene ontology terms. Relative expression of H3K4me3-enriched genes in developing secondary xylem was higher than unenriched genes, however, and highly expressed secondary cell wall-related genes were enriched for H3K4me3 as validated using ChIP-qPCR. CONCLUSIONS: In this first genome-wide analysis of a modified histone in a woody tissue, we optimized a ChIP-seq procedure suitable for field-collected samples. In developing E. grandis xylem, H3K4me3 enrichment is an indicator of active transcription, consistent with its known role in sustaining pre-initiation complex formation in yeast. The H3K4me3 ChIP-seq data from this study paves the way to understanding the chromatin landscape and epigenomic architecture of xylogenesis in plants, and complements RNA-seq evidence of gene expression for the future improvement of the E. grandis genome annotation.

Investigating the molecular underpinnings underlying morphology and changes in carbon partitioning during tension wood formation in Eucalyptus.

Tension wood has distinct physical and chemical properties, including altered fibre properties, cell wall composition and ultrastructure. It serves as a good system for investigating the genetic regulation of secondary cell wall biosynthesis and wood formation. The reference genome sequence for Eucalyptus grandis allows investigation of the global transcriptional reprogramming that accompanies tension wood formation in this global wood fibre crop. We report the first comprehensive analysis of physicochemical wood property changes in tension wood of Eucalyptus measured in a hybrid (E. grandis × Eucalyptus urophylla) clone, as well as genome-wide gene expression changes in xylem tissues 3 wk post-induction using RNA sequencing. We found that Eucalyptus tension wood in field-grown trees is characterized by an increase in cellulose, a reduction in lignin, xylose and mannose, and a marked increase in galactose. Gene expression profiling in tension wood-forming tissue showed corresponding down-regulation of monolignol biosynthetic genes, and differential expression of several carbohydrate active enzymes. We conclude that alterations of cell wall traits induced by tension wood formation in Eucalyptus are a consequence of a combination of down-regulation of lignin biosynthesis and hemicellulose remodelling, rather than the often proposed up-regulation of the cellulose biosynthetic pathway.

Mapping QTL conferring resistance in maize to gray leaf spot disease caused by Cercospora zeina.

BACKGROUND: Gray leaf spot (GLS) is a globally important foliar disease of maize. Cercospora zeina, one of the two fungal species that cause the disease, is prevalent in southern Africa, China, Brazil and the eastern corn belt of the USA. Identification of QTL for GLS resistance in subtropical germplasm is important to support breeding programmes in developing countries where C. zeina limits production of this staple food crop. RESULTS: A maize RIL population (F7:S6) from a cross between CML444 and SC Malawi was field-tested under GLS disease pressure at five field sites over three seasons in KwaZulu-Natal, South Africa. Thirty QTL identified from eleven field trials (environments) were consolidated to seven QTL for GLS resistance based on their expression in at least two environments and location in the same core maize bins. Four GLS resistance alleles were derived from the more resistant parent CML444 (bin 1.10, 4.08, 9.04/9.05, 10.06/10.07), whereas the remainder were from SC Malawi (bin 6.06/6.07, 7.02/7.03, 9.06). QTLs in bin 4.08 and bin 6.06/6.07 were also detected as joint QTLs, each explained more than 11% of the phenotypic variation, and were identified in four and seven environments, respectively. Common markers were used to allocate GLS QTL from eleven previous studies to bins on the IBM2005 map, and GLS QTL "hotspots" were noted. Bin 4.08 and 7.02/7.03 GLS QTL from this study overlapped with hotspots, whereas the bin 6.06/6.07 and bin 9.06 QTLs appeared to be unique. QTL for flowering time (bin 1.07, 4.09) in this population did not correspond to QTL for GLS resistance. CONCLUSIONS: QTL mapping of a RIL population from the subtropical maize parents CML444 and SC Malawi identified seven QTL for resistance to gray leaf spot disease caused by C. zeina. These QTL together with QTL from eleven studies were allocated to bins on the IBM2005 map to provide a basis for comparison. Hotspots of GLS QTL were identified on chromosomes one, two, four, five and seven, with QTL in the current study overlapping with two of these. Two QTL from this study did not overlap with previously reported QTL.

The identification and differential expression of Eucalyptus grandis pathogenesis-related genes in response to salicylic acid and methyl jasmonate.

Two important role players in plant defence response are the phytohormones salicylic acid (SA) and jasmonic acid (JA); both of which have been well described in model species such as Arabidopsis thaliana. Several pathogenesis related (PR) genes have previously been used as indicators of the onset of SA and JA signaling in Arabidopsis. This information is lacking in tree genera such as Eucalyptus. The aim of this study was to characterize the transcriptional response of PR genes (EgrPR2, EgrPR3, EgrPR4, EgrPR5, and EgrLOX) identified in Eucalyptus grandis to SA and methyl jasmonate (MeJA) treatment as well as to qualify them as diagnostic for the two signaling pathways. Using the genome sequence of E. grandis, we identified candidate Eucalyptus orthologs EgrPR2, EgrPR3, EgrPR4, EgrPR5, and EgrLOX based on a co-phylogenetic approach. The expression of these genes was investigated after various doses of SA and MeJA (a derivative of JA) treatment as well as at various time points. The transcript levels of EgrPR2 were decreased in response to high concentrations of MeJA whereas the expression of EgrPR3 and EgrLOX declined as the concentrations of SA treatment increased, suggesting an antagonistic relationship between SA and MeJA. Our results support EgrPR2 as potentially diagnostic for SA and EgrPR3, EgrPR4, and EgrLOX as indicators of MeJA signaling. To further validate the diagnostic potential of the PR genes we challenged E. grandis clones with the fungal necrotrophic pathogen Chrysoporthe austroafricana. The tolerant clone showed high induction of EgrPR2 and decreased transcript abundance of EgrPR4. Pre-treatment of the susceptible genotype with 5 mM SA resulted in lesion lengths comparable to the tolerant genotype after artificial inoculation with C. austroafricana. Thus expression profiling of EgrPR2 and EgrPR4 genes could serve as a useful diagnostic approach to determine which of the two signaling pathways are activated against various pathogens in Eucalyptus.