Quantitative phenotyping of grey leaf spot disease in maize using real-time PCR

Korsman, Jeanne; Meisel, Barbara; Kloppers, F. J.; Crampton, Bridget Genevieve; Berger, David Kenneth

doi:10.1007/s10658-011-9920-1

Cited by 30 publications

(31 citation statements)

References 29 publications

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“…Fungal material was quantified using an E. turcicum-specific cpr1 gene primer set and normalized relative to the amount of maize material estimated using the gst3 gene primer set (Langenhoven et al, under review). The method developed and validated by Langenhoven et al (under review) was based on the fungal quantification method developed for Cercospora zeina (Korsman et al, 2012). Quantities of E. turcicum and maize were extrapolated from standard curve graphs, and fungal quantification was determined as nanogram E. turcicum DNA per microgram maize DNA.…”

Section: Fungal Quantificationmentioning

confidence: 99%

Time-Course RNAseq Reveals Exserohilum turcicum Effectors and Pathogenicity Determinants

2020

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Exserohilum turcicum (sexual stage Setosphaeria turcica) is the hemibiotrophic causal agent of northern leaf blight of maize and sorghum. This study aimed to identify the genes involved in host colonization during the biotrophic and necrotrophic phases of infection. It also aimed to identify race-specific differences in gene expression. RNAseq of maize seedlings inoculated with a race 13N or 23N E. turcicum isolate was conducted before inoculation and at 2, 5, 7, and 13 days post-inoculation (dpi). Biological replicates were pooled per time point for each race and sequenced. A bioinformatics pipeline was used to identify candidate effectors, and expression was validated for selected candidates. Fungal biomass was positively correlated with the percentages of E. turcicum reads mapped, which were low at early time points (2-7 dpi) with a significant increase at 13 dpi, indicating a lifestyle switch from biotrophy to necrotrophy between 7 and 13 dpi. AVRHt1 is the putative E. turcicum effector recognized by the maize resistance gene Ht1. Consistent with this, AVRHt1 was expressed in planta by race 23N, but transcripts were absent in race 13N. In addition, specific transposable elements were expressed in 23N only. Genes encoding the virulence-associated peptidases leupeptin-inhibiting protein 1 and fungalysin were expressed in planta. Transcriptional profiles of genes involved in secondary metabolite synthesis or cell wall degradation revealed the importance of these genes during late stages of infection (13 dpi). A total of 346 expressed candidate effectors were identified, including Ecp6 and proteins similar to the secreted in xylem (SIX) effectors common to formae speciales of Fusarium oxysporum, SIX13 and SIX5. Expression profiling of Ecp6 and SIX13-like indicated a peak in expression at 5 and 7 dpi compared to 2 and 13 dpi. Sequencing of SIX13-like from diverse isolates of E. turcicum revealed host-specific polymorphisms that were mostly non-synonymous, resulting in two groups of SIX13-like proteins that corresponded to the maize or sorghum origin of each isolate. This study suggests putative mechanisms whereby E. turcicum causes disease. Identification of the candidate effector SIX13-like is consistent with the infection mode of E. turcicum through the xylem of susceptible hosts.

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Section: Fungal Quantificationmentioning

confidence: 99%

Time-Course RNAseq Reveals Exserohilum turcicum Effectors and Pathogenicity Determinants

2020

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“…RT-qPCR and kauralexin analysis were conducted on maize B73 plants grown in a glasshouse. At the V8-V10 stage the plants were leaf inoculated with C. zeina CMW25467 using methods described in Korsman et al (2012). Three biological replicate leaf samples from independent plants were sampled at 0 and 18 dpi (after GLS lesions had developed), and subjected to RNA extraction and kauralexin quantification.…”

Section: Rna-seq Rt-qpcr and Kauralexin Analysis Of Maize Inbred Linmentioning

confidence: 99%

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

et al. 2017

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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.

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“…First, DNA sequence information has facilitated the design of rapid species-specific PCR assays which can be used to screen many isolates from GLS lesions to identify the causal species of Cercospora Korsman et al 2012;Meisel et al 2009). Second, microsatellite markers could be designed from the genome sequence, overcoming the need for tedious and inefficient laboratory approaches for marker discovery (Santana et al 2009).…”

Section: Discussionmentioning

confidence: 99%

“…Further work on the importance of these related sibling species will be facilitated by DNA-based diagnostic methods. These include the histone H3 PCR which can be implemented directly from GLS lesions (Meisel et al 2009), and the cpr1 quantitative PCR assay, which can distinguish C. zeina from C. zeae-maydis based on the Tm of the amplicon (Korsman et al 2012). …”

Section: Discussionmentioning

confidence: 99%

Cercospora zeinafrom Maize in South Africa Exhibits High Genetic Diversity and Lack of Regional Population Differentiation

Müller

Barnes

Kunene³

et al. 2016

Phytopathology®

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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 (GLS) 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 fourteen microsatellite markers and species-specific mating type markers that were designed from draft genome sequences of C. zeina isolates from Africa (CMW 25467) and USA (USPA-4). Sixty alleles were identified across 14 loci, and gene diversity values within each

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Quantitative phenotyping of grey leaf spot disease in maize using real-time PCR

Cited by 30 publications

References 29 publications

Time-Course RNAseq Reveals Exserohilum turcicum Effectors and Pathogenicity Determinants

Time-Course RNAseq Reveals Exserohilum turcicum Effectors and Pathogenicity Determinants

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

Cercospora zeinafrom Maize in South Africa Exhibits High Genetic Diversity and Lack of Regional Population Differentiation

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