Effects of host plant resistance and fungicide applications on Ascochyta blight symptomology and yield of chickpea (Cicer arietinum L.).

Ascochyta blight (AB), caused by the pathogen Ascochyta rabiei, is a major threat to chickpea production worldwide causing major yield losses and decreasing quality. Control of AB requires integrating pest management options including resistant cultivars and fungicide applications. To address this, fungicides with different modes of action were evaluated on three chickpea cultivars with differing levels of susceptibility to AB under irrigated and dryland conditions in 2015 to 2017. The fungicides were applied once or twice and compared to a no fungicide application control on AB score and yield. The mean grain yields across locations and years were 1753, 1283 and 981 kg/ha, with a corresponding AB mean score of 2.6, 3.2, and 3.3 on 0 to 7 scale (where 0 is no disease and 7 is completely dead) for the moderately resistant, moderately susceptible, and susceptible chickpea cultivars, respectively. Fungicide application was not enough to control disease throughout the season. The use of AB resistant cultivars had the most significant impact on minimizing the disease and maximizing yield, irrespective of year and location. This study supports previous research indicating that planting AB resistant chickpea cultivars is essential for disease control, regardless of the fungicides applied.

Effect of Row Spacing and Seed Rate on Ascochyta Blight Severity and Yield of Chickpea (Cicer arietinum L.) in Tunisia

Background: In Tunisia, chickpea (Cicer arietinum L.) is the second major food legume. The development of chickpea production is facing several biotic constraints. Ascochyta blight (ABL) caused by Ascochyta rabiei (Pass.) Labr. is the most devastating disease and can cause complete yield losses under favorable conditions. In absence of chickpea variety totally resistance to ABL, some methods should be used to control and reduce this disease effects and help for its management. Therefore, this work was undertaken to evaluate the effect of row spacing and seed rate on ABL severity, growth and yield of chickpea. Methods: A split-plot design with three replicates was adopted to carry out this study during 2018 and 2019 cropping seasons. ‘Beja1’ chickpea variety was sown at 40 and 60 cm row spacing and at three seed rates (80, 110 and 140 kg ha-1). ABL severity was assessed visually on a 0-9 scale and agro morphological traits were measured. Analysis of variance was used to analyze the data. Correlations between agronomic traits, row spacing, seed rate and ABL severity were investigated. Result: Results showed that most of the variation in disease severity was associated with seed rate (r=0.61). The highest ABL score severity was noted at 140 kg ha-1 rate. Over both years, wide row spacing and low seed rate reduced ABL severity. Chickpea sown under narrow row spacing (40 cm) produced higher grain yield (1014 and 1099.7 kg ha-1 for 2018 and 2019 cropping seasons, respectively). Grain yield was tending to decrease with increasing sowing rates but at a density higher than optimal, grain yields decrease. In this study, ABL disease severity reached a score of 3.7 and 4.3 in 2018 and 2019, respectively. These disease severities levels had little effect on yield.

The nuclear effector ArPEC25 from the necrotrophic fungus Ascochyta rabiei targets the chickpea transcription factor CaβLIM1a and negatively modulates lignin biosynthesis for host susceptibility

Fungal pathogens deploy a barrage of secreted effectors to subvert host immunity, often by evading, disrupting, or altering key components of transcription, defense signaling, and metabolic pathways. However, the underlying mechanisms of effectors and their host targets are largely unexplored in necrotrophic fungal pathogens. Here, we describe the effector protein ArPEC25, which is secreted by the necrotroph Ascochyta rabiei, the causal agent of Ascochyta blight disease in chickpea (Cicer arietinum), and is indispensable for virulence. After entering host cells, ArPEC25 localizes to the nucleus and targets the host LIM transcription factor CaβLIM1a. CaβLIM1a is a transcriptional regulator of CaPAL1, which encodes phenylalanine ammonia lyase, the regulatory, gatekeeping enzyme of the phenylpropanoid pathway. ArPEC25 inhibits the transactivation of Ca?LIM1a by interfering with its DNA binding ability. This results in negative regulation of the phenylpropanoid pathway and decreased levels of intermediates of lignin biosynthesis, thereby suppressing lignin production. Our findings illustrate the role of fungal effectors in enhancing virulence by targeting a key defense pathway that leads to the biosynthesis of various secondary metabolites and antifungal compounds. This study provides a template for the study of less explored necrotrophic effectors and their host target functions.

Current population structure and pathogenicity patterns of Ascochyta rabiei in Australia

Ascochyta blight disease, caused by the necrotrophic fungus Ascochyta rabiei, is a major biotic constraint to chickpea production in Australia and worldwide. Detailed knowledge of the structure of the pathogen population and its potential to adapt to our farming practices is key to informing optimal management of the disease. This includes understanding the molecular diversity among isolates and the frequency and distribution of the isolates that have adapted to overcome host resistance across agroecologically distinct regions. Thanks to continuous monitoring efforts over the past 6 years, a comprehensive collection of A. rabiei isolates was collated from the major Australian chickpea production regions. To determine the molecular structure of the entire population, representative isolates from each collection year and growing region have been genetically characterized using a DArTseq genotyping-by-sequencing approach. The genotyped isolates were further phenotyped to determine their pathogenicity levels against a differential set of chickpea cultivars and genotype-phenotype associations were inferred. Overall, the Australian A. rabiei population displayed a far lower genetic diversity (average Nei’s gene diversity of 0.047) than detected in other populations worldwide. This may be explained by the presence of a single mating-type in Australia, MAT1-2, limiting its reproduction to a clonal mode. Despite the low detected molecular diversity, clonal selection appears to have given rise to a subset of adapted isolates that are highly pathogenic on commonly employed resistance sources, and that are occurring at an increasing frequency. Among these, a cluster of genetically similar isolates was identified, with a higher proportion of highly aggressive isolates than in the general population. The discovery of distinct genetic clusters associated with high and low isolate pathogenicity forms the foundation for the development of a molecular pathotyping tool for the Australian A. rabiei population. Application of such a tool, along with continuous monitoring of the genetic structure of the population will provide crucial information for the screening of breeding material and integrated disease management packages.

CaProDH2-mediated modulation of proline metabolism confers tolerance to Ascochyta in chickpea under drought

Drought and leaf blight caused by the fungus Ascochyta rabiei often co-occur in chickpea (Cicer arietinum)-producing areas. While the responses of chickpea to either drought or A. rabiei infection have been extensively studied, their combined effect on plant defense mechanisms is unknown. Fine modulation of stress-induced signaling pathways under combined stress is an important stress adaptation mechanism that warrants a better understanding. Here we show that drought facilitates resistance against A. rabiei infection in chickpea. The analysis of proline levels and gene expression profiling of its biosynthetic pathway under combined drought and A. rabiei infection revealed the gene encoding proline dehydrogenase (CaProDH2) as a strong candidate conferring resistance to A. rabiei infection. Transcript levels of CaProDH2, pyrroline-5-carboxylate (P5C) quantification, and measurement of mitochondrial reactive oxygen species (ROS) production showed that fine modulation of the proline-P5C cycle determines the observed resistance. In addition, CaProDH2-silenced plants lost basal resistance to A. rabiei infection induced by drought, while overexpression of the gene conferred higher resistance to the fungus. We suggest that the drought-induced accumulation of proline in the cytosol helps maintain cell turgor and raises mitochondrial P5C contents by a CaProDH2-mediated step, which results in ROS production that boosts plant defense responses and confers resistance to A. rabiei infection. Our findings indicate that manipulating the proline-P5C pathway may be a possible strategy for improving stress tolerance in plants suffering from combined drought and A. rabiei infection.

The use of Real-Time PCR Assay for Screening Chickpea Genotypes Resistant to Ascochyta Rabiei

Ascochyta blight, caused by Ascochyta rabiei is a devastating disease of chickpea worldwide. Breeding for host resistance is an efficient means to reduce the damage by this pathogen. This study evaluated the utility of Real-time polymerase chain reaction (PCR) assay for screening chickpea genotypes for resistant to blight disease. Eight days after inoculation, the resistance level of 84 chickpea genotypes was quantified by Real-time PCR technique using a standard curve constructed by amplifying different known amounts of target DNA and compared with disease scores based on visual assessment. A significant variation was statistically found among chickpea genotypes with respect to disease reactions. The quantity of target DNA in infected samples varied from 0.004–83.37 ng. The results demonstrated a strong relationship between visual scoring of disease severity and quantification of the target DNA in chickpea genotypes. Tüb-35, Tüb-47, Tüb-26, Tüb-82, Tüb-65 and Tüb-69 genotypes were found highly resistant to Ascochyta blight based on the results of both assays utilized for screening chickpea genotypes for resistance. The real-time PCR assay could be used for quantifying disease progression in plant tissues and screening chickpea genotypes as a potential alternative to visual scoring in plant resistance breeding programs.

Modulation of fungal virulence through CRZ1 regulated F-BAR-dependent actin remodeling and endocytosis in chickpea infecting phytopathogen Ascochyta rabiei

Polarized hyphal growth of filamentous pathogenic fungi is an essential event for host penetration and colonization. The long-range early endosomal trafficking during hyphal growth is crucial for nutrient uptake, sensing of host-specific cues, and regulation of effector production. Bin1/Amphiphysin/Rvs167 (BAR) domain-containing proteins mediate fundamental cellular processes, including membrane remodeling and endocytosis. Here, we identified a F-BAR domain protein (ArF-BAR) in the necrotrophic fungus Ascochyta rabiei and demonstrate its involvement in endosome-dependent fungal virulence on the host plant Cicer arietinum. We show that ArF-BAR regulates endocytosis at the hyphal tip, localizes to the early endosomes, and is involved in actin dynamics. Functional studies involving gene knockout and complementation experiments reveal that ArF-BAR is necessary for virulence. The loss-of-function of ArF-BAR gene results in delayed formation of apical septum in fungal cells near growing hyphal tip that is crucial for host penetration, and impaired secretion of a candidate effector having secretory signal peptide for translocation across the endoplasmic reticulum membrane. The mRNA transcripts of ArF-BAR were induced in response to oxidative stress and infection. We also show that ArF-BAR is able to tubulate synthetic liposomes, suggesting the functional role of F-BAR domain in membrane tubule formation in vivo. Further, our studies identified a stress-induced transcription factor, ArCRZ1 (Calcineurin-responsive zinc finger 1), as key transcriptional regulator of ArF-BAR expression. We propose a model in which ArCRZ1 functions upstream of ArF-BAR to regulate A. rabiei virulence through a mechanism that involves endocytosis, effector secretion, and actin cytoskeleton regulation.