Aggressiveness of Phytophthora medicaginis on chickpea: Phenotyping method determines isolate ranking and host genotype-by-isolate interactions

Phytophthora medicaginis causing Phytophthora root rot of chickpea (Cicer arietinum) is an important disease, with genetic resistance using C. arietinum × Cicer echinospermum crosses as the main disease management strategy. We evaluated pathogenic variation in P. medicaginis populations with the aim of improving phenotyping methods for disease resistance. We addressed the question of individual isolate aggressiveness across four different seedling-based phenotyping methods conducted in glasshouses and one field-based phenotyping method. Our results revealed that a seedling media surface inoculation method used on a susceptible C. arietinum variety and a moderately resistant C. arietinum × C. echinospermum backcross detected the greatest variability in aggressiveness among 37 P. medicaginis isolates. Evaluations of different components of resistance, using our different phenotyping methods, revealed that differential pathogen–isolate reactions occur with some phenotyping methods. We found support for our hypotheses that the level of aggressiveness of P. medicaginis isolates depends on the phenotyping method, and that phenotyping methods interact with both isolate and host genotype reactions. Our cup-based root inoculation method showed promise as a non-field-based phenotyping method, as it provided significant correlations with genotype–isolate rankings in the field experiment for a number of disease parameters

Functional genomics identifies a small secreted protein that plays a role during the biotrophic to necrotrophic shift in the root rot pathogen Phytophthora medicaginis

IntroductionHemibiotrophic Phytophthora are a group of agriculturally and ecologically important pathogenic oomycetes causing severe decline in plant growth and fitness. The lifestyle of these pathogens consists of an initial biotrophic phase followed by a switch to a necrotrophic phase in the latter stages of infection. Between these two phases is the biotrophic to necrotrophic switch (BNS) phase, the timing and controls of which are not well understood particularly in Phytophthora spp. where host resistance has a purely quantitative genetic basis.MethodsTo investigate this we sequenced and annotated the genome of Phytophthora medicaginis, causal agent of root rot and substantial yield losses to Fabaceae hosts. We analyzed the transcriptome of P. medicaginis across three phases of colonization of a susceptible chickpea host (Cicer arietinum) and performed co-regulatory analysis to identify putative small secreted protein (SSP) effectors that influence timing of the BNS in a quantitative pathosystem.ResultsThe genome of P. medicaginis is ~78 Mb, comparable to P. fragariae and P. rubi which also cause root rot. Despite this, it encodes the second smallest number of RxLR (arginine-any amino acid-leucine-arginine) containing proteins of currently sequenced Phytophthora species. Only quantitative resistance is known in chickpea to P. medicaginis, however, we found that many RxLR, Crinkler (CRN), and Nep1-like protein (NLP) proteins and carbohydrate active enzymes (CAZymes) were regulated during infection. Characterization of one of these, Phytmed_10271, which encodes an RxLR effector demonstrates that it plays a role in the timing of the BNS phase and root cell death.DiscussionThese findings provide an important framework and resource for understanding the role of pathogenicity factors in purely quantitative Phytophthora pathosystems and their implications to the timing of the BNS phase

An evaluation of Solanum nigrum and S. physalifolium biology and management strategies to reduce nightshade fruit contamination of process pea crops

The contamination of process pea (Pisum sativum L.) crops by the immature fruit of black nightshade (Solanum nigrum L.) and hairy nightshade (S. physalifolium Rusby var. nitidibaccatum (Bitter.) Edmonds) causes income losses to pea farmers in Canterbury, New Zealand. This thesis investigates the questions of whether seed dormancy, germination requirements, plant growth, reproductive phenology, or fruit growth of either nightshade species reveal specific management practices that could reduce the contamination of process peas by the fruit of these two weeds. The seed dormancy status of these weeds indicated that both species are capable of germinating to high levels (> 90%) throughout the pea sowing season when tested at an optimum germination temperature of 20/30 °C (16/8 h). However, light was required at this temperature regime to obtain maximum germination of S. nigrum. The levels of germination in the dark at 20/30 °C and at 5/20 °C, and in light at 5/20 °C, and day to 50 % germination analyses indicated that this species cycled from nondormancy to conditional dormancy throughout the period of investigation (July to December 2002). For S. physalifolium, light was not a germination requirement, and dormancy inhibited germination at 5/20 °C early in the pea sowing season (July and August). However, by October, 100% of the population was non-dormant at this test temperature. Two field trials showed that dark cultivation did not reduce the germination of either species. Growth trials with S. nigrum and S. physalifolium indicated that S. physalifolium, in a non-competitive environment, accumulated dry matter at a faster rate than S. nigrum. However, when the two species were grown with peas there was no difference in dry matter accumulation. Investigation of the flowering phenology and fruit growth of both species showed that S. physalifolium flowered (509 °Cd, base temperature (Tb) 6 °C) approximately 120 °Cd prior to S. nigrum (633 °Cd). The fruit growth rate of S. nigrum (0.62 mm/d) was significantly faster than the growth rate of S. physalifolium (0.36 mm/d). Because of the earlier flowering of S. physalifolium it was estimated that for seedlings of both species emerging on the same date that S. physalifolium could produce a fruit with a maximum diameter of 3 mm ~ 60 °Cd before S. nigrum. Overlaps in flowering between peas and nightshade were examined in four pea cultivars, of varying time to maturity, sown on six dates. Solanum physalifolium had the potential to contaminate more pea crops than S. nigrum. In particular, late sown peas were more prone to nightshade contamination, especially late sowings using mid to long duration pea cultivars (777-839 °Cd, Tb 4.5 °C). This comparison was supported by factory data, which indicated that contamination of crops sown in October and November was more common than in crops sown in August and September. Also, cultivars sown in the later two months had an ~ 100 °Cd greater maturity value than cultivars sown in August and September. Nightshade flowering and pea maturity comparisons indicated that the use of the thermal time values for the flowering of S. nigrum and S. physalifolium can be used to calculate the necessary weed free period required from pea sowing in order to prevent the flowering of these species. The earlier flowering of S. physalifolium indicates that this species is more likely to contaminate pea crops than is S. nigrum. Therefore, extra attention may be required where this species is present in process pea crops. The prevention of the flowering of both species, by the maintenance of the appropriate weed free period following pea sowing or crop emergence, was identified as potentially, the most useful means of reducing nightshade contamination in peas

Chickpea shows genotype-specific nodulation responses across soil nitrogen environment and root disease resistance categories

Background: The ability of chickpea to obtain sufficient nitrogen via its symbiotic relationship with Mesorhizobium ciceri is of critical importance in supporting growth and grain production. A number of factors can affect this symbiotic relationship including abiotic conditions, plant genotype, and disruptions to host signalling/perception networks. In order to support improved nodule formation in chickpea, we investigated how plant genotype and soil nutrient availability affect chickpea nodule formation and nitrogen fixation. Further, using transcriptomic profiling, we sought to identify gene expression patterns that characterize highly nodulated genotypes. Results: A study involving six chickpea varieties demonstrated large genotype by soil nitrogen interaction effects on nodulation and further identified agronomic traits of genotypes (such as shoot weight) associated with high nodulation. We broadened our scope to consider 29 varieties and breeding lines to examine the relationship between soilborne disease resistance and the number of nodules developed and real-time nitrogen fixation. Results of this larger study supported the earlier genotype specific findings, however, disease resistance did not explain differences in nodulation across genotypes. Transcriptional profiling of six chickpea genotypes indicates that genes associated with signalling, N transport and cellular localization, as opposed to genes associated with the classical nodulation pathway, are more likely to predict whether a given genotype will exhibit high levels of nodule formation. Conclusions: This research identified a number of key abiotic and genetic factors affecting chickpea nodule development and nitrogen fixation. These findings indicate that an improved understanding of genotype-specific factors affecting chickpea nodule induction and function are key research areas necessary to improving the benefits of rhizobial symbiosis in chickpea

Chickpea shows genotype-specific nodulation responses across soil nitrogen environment and root disease resistance categories

Abstract Background The ability of chickpea to obtain sufficient nitrogen via its symbiotic relationship with Mesorhizobium ciceri is of critical importance in supporting growth and grain production. A number of factors can affect this symbiotic relationship including abiotic conditions, plant genotype, and disruptions to host signalling/perception networks. In order to support improved nodule formation in chickpea, we investigated how plant genotype and soil nutrient availability affect chickpea nodule formation and nitrogen fixation. Further, using transcriptomic profiling, we sought to identify gene expression patterns that characterize highly nodulated genotypes. Results A study involving six chickpea varieties demonstrated large genotype by soil nitrogen interaction effects on nodulation and further identified agronomic traits of genotypes (such as shoot weight) associated with high nodulation. We broadened our scope to consider 29 varieties and breeding lines to examine the relationship between soilborne disease resistance and the number of nodules developed and real-time nitrogen fixation. Results of this larger study supported the earlier genotype specific findings, however, disease resistance did not explain differences in nodulation across genotypes. Transcriptional profiling of six chickpea genotypes indicates that genes associated with signalling, N transport and cellular localization, as opposed to genes associated with the classical nodulation pathway, are more likely to predict whether a given genotype will exhibit high levels of nodule formation. Conclusions This research identified a number of key abiotic and genetic factors affecting chickpea nodule development and nitrogen fixation. These findings indicate that an improved understanding of genotype-specific factors affecting chickpea nodule induction and function are key research areas necessary to improving the benefits of rhizobial symbiosis in chickpea. </jats:sec

Inoculum production of Phytophthora medicaginis can be used to screen for partial resistance in chickpea genotypes

Phytophthora root rot caused by Phytophthora medicaginis is an important disease of chickpeas (Cicer arietinum) in Australia with limited management options, increasing reliance on breeding for improved levels of genetic resistance. Resistance based on chickpea–Cicer echinospermum crosses is partial with a quantitative genetic basis provided by C. echinospermum and some disease tolerance traits originating from C. arietinum germplasm. Partial resistance is hypothesised to reduce pathogen proliferation, while tolerant germplasm may contribute some fitness traits, such as an ability to maintain yield despite pathogen proliferation. To test these hypotheses, we used P. medicaginis DNA concentrations in the soil as a parameter for pathogen proliferation and disease assessments on lines of two recombinant inbred populations of chickpea–C. echinospermum crosses to compare the reactions of selected recombinant inbred lines and parents. Our results showed reduced inoculum production in a C. echinospermum backcross parent relative to the C. arietinum variety Yorker. Recombinant inbred lines with consistently low levels of foliage symptoms had significantly lower levels of soil inoculum compared to lines with high levels of visible foliage symptoms. In a separate experiment, a set of superior recombinant inbred lines with consistently low levels of foliage symptoms was tested for soil inoculum reactions relative to control normalised yield loss. The in-crop P. medicaginis soil inoculum concentrations across genotypes were significantly and positively related to yield loss, indicating a partial resistance-tolerance spectrum. Disease incidence and the rankings for in-crop soil inoculum were correlated strongly to yield loss. These results indicate that soil inoculum reactions may be useful to identify genotypes with high levels of partial resistance.</jats:p

Selection for Phytophthora Root Rot Resistance in Chickpea Crosses Affects Yield Potential of Chickpea × <i>Cicer echinospermum</i> Backcross Derivatives

Phytophthora root rot (PRR) of chickpea (Cicer arietinum) caused by Phytophthora medicaginis is an important disease. Partial resistance to PRR is sourced from Cicer echinospermum. In this study, we evaluated if lines with low levels of PRR foliage symptoms in two contrasting recombinant inbred line (RIL) populations parented by chickpea cultivars (Yorker and Rupali) and 04067-81-2-1-1 (C. echinospermum, interspecific breeding line) had a significant drag on yield parameters. For the Yorker × 04067-81-2-1-1 population with the highest level of PRR resistance, in the absence of PRR, low foliage symptom RIL had significantly later flowering and podding, lower grain yields, and lighter seed and shorter plant phenotypes than high foliage symptom RIL. A quantitative trait locus analysis identified significant QTL for flowering, height, 100-seed weight, and yield, and there was a significantly higher frequency of alleles for the negative agronomic traits (i.e., drag) from the 04067-81-2-1-1 parent in low foliage symptom RIL than in high foliage symptom RIL. For the Rupali × 04067-81-2-1-1 population with lower levels of PRR resistance, in the absence of PRR, low foliage symptom RIL had significantly lighter seed and shorter plants than high foliage symptom RIL. Significant QTL were detected, the majority were for the timing of flowering and podding (n = 18), others were for plant height, yield, and 100-seed weight. For this second population, the frequency of alleles for the negative agronomic traits from the 04067-81-2-1-1 parent did not differ between low and high foliage symptom RIL. The 100 seed weight of RIL under moderate PRR disease pressure showed some promise as a yield component trait to identify phenotypes with both high levels of PRR resistance and grain yield potential for further seed number evaluations. We identified that large population sizes are required to enable selection among chickpea × C. echinospermum crosses for high levels of PRR resistance without a significant drag on yield

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.</jats:p

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&ndash;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