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Rebecca Ford - One of the best experts on this subject based on the ideXlab platform.
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Determination of the Key Resistance Gene Analogs Involved in Ascochyta Rabiei Recognition in Chickpea
Frontiers Media S.A., 2019Co-Authors: Ziwei Zhou, Prabhakaran Thanjavur Sambasivam, Ido Bar, Rebecca FordAbstract:Chickpea (Cicer arietinum L.) is an important cool season food legume, however, its production is severely constrained by the foliar disease Ascochyta blight caused by the fungus Ascochyta Rabiei (syn. Phoma Rabiei). Several disease management options have been developed to control the pathogen, including breeding for host plant resistance. However, the pathogen population is evolving to produce more aggressive isolates. For host resistance to be effective, the plant must quickly recognize the pathogen and instigate initial defense mechanisms, optimally at the point of contact. Given that the most resistant host genotypes display rapid pathogen recognition and response, the approach taken was to assess the type, speed and pattern of recognition via Resistance Gene Analog (RGA) transcription among resistant and susceptible cultivated chickpea varieties. RGAs are key factors in the recognition of plant pathogens and the signaling of inducible defenses. In this study, a suite of RGA loci were chosen for further investigation from both published literature and from newly mined homologous sequences within the National Center for Biotechnology Information (NCBI) database. Following their validation in the chickpea genome, 10 target RGAs were selected for differential expression analysis in response to A. Rabiei infection. This was performed in a set of four chickpea varieties including two resistant cultivars (ICC3996 and PBA Seamer), one moderately resistant cultivar (PBA HatTrick) and one susceptible cultivar (Kyabra). Gene expression at each RGA locus was assessed via qPCR at 2, 6, and 24 h after A. Rabiei inoculation with a previously characterized highly aggressive isolate. As a result, all loci were differentially transcribed in response to pathogen infection in at least one genotype and at least one time point after inoculation. Among these, the differential expression of four RGAs was significant and consistently increased in the most resistant genotype ICC3996 immediately following inoculation, when spore germination began and ahead of penetration into the plant’s epidermal tissues. Further in silico analyses indicated that the differentially transcribed RGAs function through ADP-binding within the pathogen recognition pathway. These represent clear targets for future functional validation and potential for selective resistance breeding for introgression into elite cultivars
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Table_1_Determination of the Key Resistance Gene Analogs Involved in Ascochyta Rabiei Recognition in Chickpea.DOCX
2019Co-Authors: Ziwei Zhou, Prabhakaran Thanjavur Sambasivam, Ido Bar, Rebecca FordAbstract:Chickpea (Cicer arietinum L.) is an important cool season food legume, however, its production is severely constrained by the foliar disease Ascochyta blight caused by the fungus Ascochyta Rabiei (syn. Phoma Rabiei). Several disease management options have been developed to control the pathogen, including breeding for host plant resistance. However, the pathogen population is evolving to produce more aggressive isolates. For host resistance to be effective, the plant must quickly recognize the pathogen and instigate initial defense mechanisms, optimally at the point of contact. Given that the most resistant host genotypes display rapid pathogen recognition and response, the approach taken was to assess the type, speed and pattern of recognition via Resistance Gene Analog (RGA) transcription among resistant and susceptible cultivated chickpea varieties. RGAs are key factors in the recognition of plant pathogens and the signaling of inducible defenses. In this study, a suite of RGA loci were chosen for further investigation from both published literature and from newly mined homologous sequences within the National Center for Biotechnology Information (NCBI) database. Following their validation in the chickpea genome, 10 target RGAs were selected for differential expression analysis in response to A. Rabiei infection. This was performed in a set of four chickpea varieties including two resistant cultivars (ICC3996 and PBA Seamer), one moderately resistant cultivar (PBA HatTrick) and one susceptible cultivar (Kyabra). Gene expression at each RGA locus was assessed via qPCR at 2, 6, and 24 h after A. Rabiei inoculation with a previously characterized highly aggressive isolate. As a result, all loci were differentially transcribed in response to pathogen infection in at least one genotype and at least one time point after inoculation. Among these, the differential expression of four RGAs was significant and consistently increased in the most resistant genotype ICC3996 immediately following inoculation, when spore germination began and ahead of penetration into the plant’s epidermal tissues. Further in silico analyses indicated that the differentially transcribed RGAs function through ADP-binding within the pathogen recognition pathway. These represent clear targets for future functional validation and potential for selective resistance breeding for introgression into elite cultivars.
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evidence and consequence of a highly adapted clonal haplotype within the australian Ascochyta Rabiei population
Frontiers in Plant Science, 2017Co-Authors: Yasir Mehmood, Celeste C. Linde, Audrey E Leo, Prabhakaran Thanjavur Sambasivam, Sukhjiwan Kaur, J A Davidson, Kristy Hobson, Kevin Moore, Jeremy C Brownlie, Rebecca FordAbstract:The Australian Ascochyta Rabiei (syn. Phoma Rabiei) population has low genotypic diversity with only one mating type detected to date, potentially precluding substantial evolution through recombination. However, a large diversity in aggressiveness exists. In an effort to better understand the risk from selective adaptation to currently used resistance sources and chemical control strategies, the population was examined in detail. For this, a total of 598 isolates were quasi-hierarchically sampled between 2013-2015 across all major Australian chickpea growing regions and commonly grown host genotypes. Although a large number of haplotypes were identified (66) through short sequence repeat (SSR) genotyping, overall low gene diversity (Hexp = 0.066) and genotypic diversity (D = 0.57) was detected. Almost 70% of the isolates assessed were of a single dominant haplotype (ARH01). Disease screening on a differential host set, including three commonly deployed resistance sources, revealed distinct aggressiveness among the isolates, with 17% of all isolates identified as highly aggressive. Almost 75% of these were of the ARH01 haplotype. A similar pattern was observed at the host level, with 46% of all isolates collected from the commonly grown host genotype Genesis090 (classified as “resistant” during the term of collection) identified as highly aggressive. Of these, 63% belonged to the ARH01 haplotype. In conclusion, the ARH01 haplotype represents a significant risk to the Australian chickpea industry, being not only widely adapted to the diverse agro-geographical environments of the Australian chickpea growing regions, but also containing a disproportionately large number of aggressive isolates, indicating fitness to survive and replicate on the best resistance sources in the Australian germplasm.
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Changes in Foliar Host Reaction to Ascochyta Rabiei With Plant Maturity
2016Co-Authors: Vicki L. Elliott, Paul W. J. Taylor, Rebecca FordAbstract:Ascochyta Rabiei is the most significant foliar disease of chickpea (Cicer arietinum) worldwide, and can cause both yield loss and reduced seed quality. Significant mean disease score differences were observed from the reactions among a collection of Ascochyta Rabiei isolates and chickpea hosts at both seedling and maturity growth stages (P<0.000). Although isolates ranked similarly within their range of pathogenicity at either growth stage, the ranking of host disease reaction was different. This suggests that genotype-specific expression of resistance genes to A. Rabiei may be related to growth stage. Hence, durable selection methods for resistance will require screening of isolates representative of the pathogenicity within the population on target genotypes at growth stages representative of the type of resistance being sought, for example seedling resistance or pod resistance
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Defence gene expression profiling to Ascochyta Rabiei aggressiveness in chickpea
Theoretical and Applied Genetics, 2016Co-Authors: Celeste C. Linde, Rebecca FordAbstract:Key message Significant differences in defence pathway-related gene expression were observed among chickpea cultivars following A. Rabiei infection. Differential gene expression is indicative of diverse resistances, a theoretical tool for selective breeding. Abstract A high number of Ascochyta Rabiei pathotypes infecting chickpea in Australia has severely hampered efforts towards breeding for sustained quantitative resistance in chickpea. Breeding for sustained resistance will be aided by detailed knowledge of defence responses to isolates with different aggressiveness. As an initial step, the conserved and differential expressions of a suit of previously characterised genes known to be involved in fungal defence mechanisms were assessed among resistant and susceptible host genotypes following inoculation with high or low aggressive A. Rabiei isolates. Using quantitative Real-Time PCR (qRT-PCR), 15 defence-related genes, normalised with two reference genes, were temporally differentially expressed ( P
Praveen Kumar Verma - One of the best experts on this subject based on the ideXlab platform.
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Modulation of fungal virulence through CRZ1 regulated F-BAR-dependent actin remodeling and endocytosis in chickpea infecting phytopathogen Ascochyta Rabiei.
'Public Library of Science (PLoS)', 2021Co-Authors: Manisha Sinha, Kunal Singh, Ankita Shree, Kamal Kumar, Shreenivas Kumar Singh, Vimlesh Kumar, Praveen Kumar VermaAbstract: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
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Comparative Structural Modeling of Six Old Yellow Enzymes (OYEs) from the Necrotrophic Fungus Ascochyta Rabiei: Insight into Novel OYE Classes with Differences in Cofactor Binding, Organization of Active Site Residues and Stereopreferences
2016Co-Authors: Shadab Nizam, Rajesh Kumar Gazara, Kunal Singh, Hya Verma, Praveen Kumar VermaAbstract:Old Yellow Enzyme (OYE1) was the first flavin-dependent enzyme identified and characterized in detail by the entire range of physical techniques. Irrespective of this scrutiny, true physiological role of the enzyme remains a mystery. In a recent study, we systematically identified OYE proteins from various fungi and classified them into three classes viz. Class I, II and III. However, there is no information about the structural organization of Class III OYEs, eukaryotic Class II OYEs and Class I OYEs of filamentous fungi. Ascochyta Rabiei, a filamentous phytopathogen which causes Ascochyta blight (AB) in chickpea possesses six OYEs (ArOYE1-6) belonging to the three OYE classes. Here we carried out comparative homology modeling of six ArOYEs representing all the three classes to get an in depth idea of structural and functional aspects of fungal OYEs. The predicted 3D structures of A. Rabiei OYEs were refined and evaluated using various validation tools for their structural integrity. Analysis of FMN binding environment of Class III OYE revealed novel residues involved in interaction. The ligand para-hydroxybenzaldehyde (PHB) was docked into the active site of the enzymes and interacting residues were analyzed. We observed a unique active site organization of Class III OYE in comparison to Class I and II OYEs. Subsequently, analysis of stereopreference through structural features of ArOYEs was carried out, suggesting differences in R/S selectivity of these proteins. Therefore, our comparative modeling study provides insights into the FMN binding, active site organization and stereopreference of different classes of ArOYEs and indicates towards functional differences of these enzymes. This stud
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draft genome sequencing and secretome analysis of fungal phytopathogen Ascochyta Rabiei provides insight into the necrotrophic effector repertoire
Scientific Reports, 2016Co-Authors: Sandhya Verma, Rajesh Kumar Gazara, Shadab Nizam, Sabiha Parween, Debasis Chattopadhyay, Praveen Kumar VermaAbstract:Constant evolutionary pressure acting on pathogens refines their molecular strategies to attain successful pathogenesis. Recent studies have shown that pathogenicity mechanisms of necrotrophic fungi are far more intricate than earlier evaluated. However, only a few studies have explored necrotrophic fungal pathogens. Ascochyta Rabiei is a necrotrophic fungus that causes devastating blight disease of chickpea (Cicer arietinum). Here, we report a 34.6 megabase draft genome assembly of A. Rabiei. The genome assembly covered more than 99% of the gene space and 4,259 simple sequence repeats were identified in the assembly. A total of 10,596 high confidence protein-coding genes were predicted which includes a large and diverse inventory of secretory proteins, transporters and primary and secondary metabolism enzymes reflecting the necrotrophic lifestyle of A. Rabiei. A wide range of genes encoding carbohydrate-active enzymes capable for degradation of complex polysaccharides were also identified. Comprehensive analysis predicted a set of 758 secretory proteins including both classical and non-classical secreted proteins. Several of these predicted secretory proteins showed high cysteine content and numerous tandem repeats. Together, our analyses would broadly expand our knowledge and offer insights into the pathogenesis and necrotrophic lifestyle of fungal phytopathogens.
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comparative structural modeling of six old yellow enzymes oyes from the necrotrophic fungus Ascochyta Rabiei insight into novel oye classes with differences in cofactor binding organization of active site residues and stereopreferences
PLOS ONE, 2014Co-Authors: Shadab Nizam, Sandhya Verma, Rajesh Kumar Gazara, Kunal Singh, Praveen Kumar VermaAbstract:Old Yellow Enzyme (OYE1) was the first flavin-dependent enzyme identified and characterized in detail by the entire range of physical techniques. Irrespective of this scrutiny, true physiological role of the enzyme remains a mystery. In a recent study, we systematically identified OYE proteins from various fungi and classified them into three classes viz. Class I, II and III. However, there is no information about the structural organization of Class III OYEs, eukaryotic Class II OYEs and Class I OYEs of filamentous fungi. Ascochyta Rabiei, a filamentous phytopathogen which causes Ascochyta blight (AB) in chickpea possesses six OYEs (ArOYE1-6) belonging to the three OYE classes. Here we carried out comparative homology modeling of six ArOYEs representing all the three classes to get an in depth idea of structural and functional aspects of fungal OYEs. The predicted 3D structures of A. Rabiei OYEs were refined and evaluated using various validation tools for their structural integrity. Analysis of FMN binding environment of Class III OYE revealed novel residues involved in interaction. The ligand para-hydroxybenzaldehyde (PHB) was docked into the active site of the enzymes and interacting residues were analyzed. We observed a unique active site organization of Class III OYE in comparison to Class I and II OYEs. Subsequently, analysis of stereopreference through structural features of ArOYEs was carried out, suggesting differences in R/S selectivity of these proteins. Therefore, our comparative modeling study provides insights into the FMN binding, active site organization and stereopreference of different classes of ArOYEs and indicates towards functional differences of these enzymes. This study provides the basis for future investigations towards the biochemical and functional characterization of these enigmatic enzymes.
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comparative transcriptome analysis of the necrotrophic fungus Ascochyta Rabiei during oxidative stress insight for fungal survival in the host plant
PLOS ONE, 2012Co-Authors: Kunal Singh, Shadab Nizam, Manisha Sinha, Praveen Kumar VermaAbstract:Localized cell death, known as the hypersensitive response (HR), is an important defense mechanism for neutralizing phytopathogens. The hallmark of the HR is an oxidative burst produced by the host plant. We aimed to identify genes of the necrotrophic chickpea blight fungus Ascochyta Rabiei that are involved in counteracting oxidative stress. A subtractive cDNA library was constructed after menadione treatment, which resulted in the isolation of 128 unigenes. A reverse northern blot was used to compare transcript profiles after H2O2, menadione and sodium nitroprusside treatments. A total of 70 unigenes were found to be upregulated by more than two-fold following menadione treatment at different time intervals. A large number of genes not previously associated with oxidative stress were identified, along with many stress-responsive genes. Differential expression patterns of several genes were validated by quantitative real-time PCR (qRT-PCR) and northern blotting. In planta qRT-PCR of several selected genes also showed differential expression patterns during infection and disease progression. These data shed light on the molecular responses of the phytopathogen A. Rabiei to overcome oxidative and nitrosative stresses and advance the understanding of necrotrophic fungal pathogen survival mechanisms.
Mauricio Carrillo Avila - One of the best experts on this subject based on the ideXlab platform.
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Permanent genetic resources added to Molecular Ecology Resources Database 1 October 2010-30 November 2010.
Molecular ecology resources, 2011Co-Authors: Cecilia Agostini, P. A. Agudelo, P. A. Barber, Paolo M. Bisol, C. Brouat, Treena I. Burgess, Isabelle Calves, Mauricio Carrillo AvilaAbstract:This article documents the addition of 277 microsatellite marker loci to the Molecular Ecology Resources Database. Loci were developed for the following species: Ascochyta Rabiei, Cambarellus chapalanus, Chionodraco hamatus, Coptis omeiensis, Cynoscion nebulosus, Daphnia magna, Gerbillus nigeriae, Isurus oxyrinchus, Lates calcarifer, Metacarcinus magister, Oplegnathus fasciatus, Pachycondyla verenae, Phaethon lepturus, Pimelodus grosskopfii, Rotylenchulus reniformis, Scomberomorus niphonius, Sepia esculenta, Terapon jarbua, Teratosphaeria cryptica and Thunnus obesus. These loci were cross-tested on the following species: Austropotamobius italicus, Cambarellus montezumae, Cambarellus puer, Cambarellus shufeldtii, Cambarellus texanus, Chionodraco myersi, Chionodraco rastrospinosus, Coptis chinensis, Coptis chinensis var. brevisepala, Coptis deltoidea, Coptis teeta, Orconectes virilis, Pacifastacus leniusculus, Pimelodus bochii, Procambarus clarkii, Pseudopimelodus bufonius, Rhamdia quelen, Sepia andreana, Sepiella maindroni, Thunnus alalunga, Thunnus albacares, Thunnus maccoyii, Thunnus orientalis, Thunnus thynnus and Thunnus tonggol.
Shadab Nizam - One of the best experts on this subject based on the ideXlab platform.
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Comparative Structural Modeling of Six Old Yellow Enzymes (OYEs) from the Necrotrophic Fungus Ascochyta Rabiei: Insight into Novel OYE Classes with Differences in Cofactor Binding, Organization of Active Site Residues and Stereopreferences
2016Co-Authors: Shadab Nizam, Rajesh Kumar Gazara, Kunal Singh, Hya Verma, Praveen Kumar VermaAbstract:Old Yellow Enzyme (OYE1) was the first flavin-dependent enzyme identified and characterized in detail by the entire range of physical techniques. Irrespective of this scrutiny, true physiological role of the enzyme remains a mystery. In a recent study, we systematically identified OYE proteins from various fungi and classified them into three classes viz. Class I, II and III. However, there is no information about the structural organization of Class III OYEs, eukaryotic Class II OYEs and Class I OYEs of filamentous fungi. Ascochyta Rabiei, a filamentous phytopathogen which causes Ascochyta blight (AB) in chickpea possesses six OYEs (ArOYE1-6) belonging to the three OYE classes. Here we carried out comparative homology modeling of six ArOYEs representing all the three classes to get an in depth idea of structural and functional aspects of fungal OYEs. The predicted 3D structures of A. Rabiei OYEs were refined and evaluated using various validation tools for their structural integrity. Analysis of FMN binding environment of Class III OYE revealed novel residues involved in interaction. The ligand para-hydroxybenzaldehyde (PHB) was docked into the active site of the enzymes and interacting residues were analyzed. We observed a unique active site organization of Class III OYE in comparison to Class I and II OYEs. Subsequently, analysis of stereopreference through structural features of ArOYEs was carried out, suggesting differences in R/S selectivity of these proteins. Therefore, our comparative modeling study provides insights into the FMN binding, active site organization and stereopreference of different classes of ArOYEs and indicates towards functional differences of these enzymes. This stud
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draft genome sequencing and secretome analysis of fungal phytopathogen Ascochyta Rabiei provides insight into the necrotrophic effector repertoire
Scientific Reports, 2016Co-Authors: Sandhya Verma, Rajesh Kumar Gazara, Shadab Nizam, Sabiha Parween, Debasis Chattopadhyay, Praveen Kumar VermaAbstract:Constant evolutionary pressure acting on pathogens refines their molecular strategies to attain successful pathogenesis. Recent studies have shown that pathogenicity mechanisms of necrotrophic fungi are far more intricate than earlier evaluated. However, only a few studies have explored necrotrophic fungal pathogens. Ascochyta Rabiei is a necrotrophic fungus that causes devastating blight disease of chickpea (Cicer arietinum). Here, we report a 34.6 megabase draft genome assembly of A. Rabiei. The genome assembly covered more than 99% of the gene space and 4,259 simple sequence repeats were identified in the assembly. A total of 10,596 high confidence protein-coding genes were predicted which includes a large and diverse inventory of secretory proteins, transporters and primary and secondary metabolism enzymes reflecting the necrotrophic lifestyle of A. Rabiei. A wide range of genes encoding carbohydrate-active enzymes capable for degradation of complex polysaccharides were also identified. Comprehensive analysis predicted a set of 758 secretory proteins including both classical and non-classical secreted proteins. Several of these predicted secretory proteins showed high cysteine content and numerous tandem repeats. Together, our analyses would broadly expand our knowledge and offer insights into the pathogenesis and necrotrophic lifestyle of fungal phytopathogens.
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comparative structural modeling of six old yellow enzymes oyes from the necrotrophic fungus Ascochyta Rabiei insight into novel oye classes with differences in cofactor binding organization of active site residues and stereopreferences
PLOS ONE, 2014Co-Authors: Shadab Nizam, Sandhya Verma, Rajesh Kumar Gazara, Kunal Singh, Praveen Kumar VermaAbstract:Old Yellow Enzyme (OYE1) was the first flavin-dependent enzyme identified and characterized in detail by the entire range of physical techniques. Irrespective of this scrutiny, true physiological role of the enzyme remains a mystery. In a recent study, we systematically identified OYE proteins from various fungi and classified them into three classes viz. Class I, II and III. However, there is no information about the structural organization of Class III OYEs, eukaryotic Class II OYEs and Class I OYEs of filamentous fungi. Ascochyta Rabiei, a filamentous phytopathogen which causes Ascochyta blight (AB) in chickpea possesses six OYEs (ArOYE1-6) belonging to the three OYE classes. Here we carried out comparative homology modeling of six ArOYEs representing all the three classes to get an in depth idea of structural and functional aspects of fungal OYEs. The predicted 3D structures of A. Rabiei OYEs were refined and evaluated using various validation tools for their structural integrity. Analysis of FMN binding environment of Class III OYE revealed novel residues involved in interaction. The ligand para-hydroxybenzaldehyde (PHB) was docked into the active site of the enzymes and interacting residues were analyzed. We observed a unique active site organization of Class III OYE in comparison to Class I and II OYEs. Subsequently, analysis of stereopreference through structural features of ArOYEs was carried out, suggesting differences in R/S selectivity of these proteins. Therefore, our comparative modeling study provides insights into the FMN binding, active site organization and stereopreference of different classes of ArOYEs and indicates towards functional differences of these enzymes. This study provides the basis for future investigations towards the biochemical and functional characterization of these enigmatic enzymes.
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comparative transcriptome analysis of the necrotrophic fungus Ascochyta Rabiei during oxidative stress insight for fungal survival in the host plant
PLOS ONE, 2012Co-Authors: Kunal Singh, Shadab Nizam, Manisha Sinha, Praveen Kumar VermaAbstract:Localized cell death, known as the hypersensitive response (HR), is an important defense mechanism for neutralizing phytopathogens. The hallmark of the HR is an oxidative burst produced by the host plant. We aimed to identify genes of the necrotrophic chickpea blight fungus Ascochyta Rabiei that are involved in counteracting oxidative stress. A subtractive cDNA library was constructed after menadione treatment, which resulted in the isolation of 128 unigenes. A reverse northern blot was used to compare transcript profiles after H2O2, menadione and sodium nitroprusside treatments. A total of 70 unigenes were found to be upregulated by more than two-fold following menadione treatment at different time intervals. A large number of genes not previously associated with oxidative stress were identified, along with many stress-responsive genes. Differential expression patterns of several genes were validated by quantitative real-time PCR (qRT-PCR) and northern blotting. In planta qRT-PCR of several selected genes also showed differential expression patterns during infection and disease progression. These data shed light on the molecular responses of the phytopathogen A. Rabiei to overcome oxidative and nitrosative stresses and advance the understanding of necrotrophic fungal pathogen survival mechanisms.
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Comparative Transcriptome Analysis of the Necrotrophic Fungus Ascochyta Rabiei during Oxidative Stress: Insight for Fungal Survival in the Host Plant
2012Co-Authors: Kunal Singh, Shadab Nizam, Manisha Sinha, Praveen K. VermaAbstract:Localized cell death, known as the hypersensitive response (HR), is an important defense mechanism for neutralizing phytopathogens. The hallmark of the HR is an oxidative burst produced by the host plant. We aimed to identify genes of the necrotrophic chickpea blight fungus Ascochyta Rabiei that are involved in counteracting oxidative stress. A subtractive cDNA library was constructed after menadione treatment, which resulted in the isolation of 128 unigenes. A reverse northern blot was used to compare transcript profiles after H 2O 2, menadione and sodium nitroprusside treatments. A total of 70 unigenes were found to be upregulated by more than two-fold following menadione treatment at different time intervals. A large number of genes not previously associated with oxidative stress were identified, along with many stress-responsive genes. Differential expression patterns of several genes were validated by quantitative real-time PCR (qRT-PCR) and northern blotting. In planta qRT-PCR of several selected genes also showed differential expression patterns during infection and disease progression. These data shed light on the molecular responses of the phytopathogen A. Rabiei to overcom
Tobin L Peever - One of the best experts on this subject based on the ideXlab platform.
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production of the antibiotic secondary metabolite solanapyrone a by the fungal plant pathogen Ascochyta Rabiei during fruiting body formation in saprobic growth
Environmental Microbiology, 2017Co-Authors: Wonyong Kim, Tobin L Peever, Jeongjin Park, Frank M Dugan, David R Gang, George J Vandemark, Weidong ChenAbstract:Fungi are noted producers of a diverse array of secondary metabolites, many of which are of pharmacological importance. However, the biological roles of the vast majority of these molecules during the fungal life cycle in nature remain elusive. Solanapyrones are polyketide-derived secondary metabolites produced by diverse fungal species including the plant pathogen Ascochyta Rabiei. This molecule was originally thought to function as a phytotoxin facilitating pathogenesis of A. Rabiei. Chemical profiling and gene expression studies showed that solanapyrone A was specifically produced during saprobic, but not parasitic growth of A. Rabiei. Expression of the gene encoding the final enzymatic step in solanapyrone biosynthesis was specifically associated with development of the asexual fruiting bodies of the fungus on certain substrates. In confrontation assays with saprobic fungi that were commonly found in chickpea debris in fields, A. Rabiei effectively suppressed the growth of all competing fungi, such as Alternaria, Epicoccum and Ulocladium species. Solanapyrone A was directly detected in the inhibitory zone using a MALDI-imaging mass spectrometry, and the purified compound showed significant antifungal activities against the potential saprobic competitors. These results suggest that solanapyrone A plays an important role for competition and presumably the survival of the fungus.
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identification and function of a polyketide synthase gene responsible for 1 8 dihydroxynaphthalene melanin pigment biosynthesis in Ascochyta Rabiei
Current Genetics, 2010Co-Authors: Hajime Akamatsu, Martin I Chilvers, Jane E Stewart, Tobin L PeeverAbstract:Ascochyta Rabiei produces and accumulates one of the well-known fungal polyketides, 1,8-dihydroxynaphthalene-melanin pigment (DHN-melanin), in asexual and sexual fruiting bodies. Degenerate PCR primers were used to isolate an ArPKS1 of A. Rabiei encoding a polypeptide with high similarity to polyketide synthase (PKS) involved in biosynthesis of DHN-melanin in other ascomycetous fungi. Site-directed mutagenesis of ArPKS1 in A. Rabiei generated melanin-deficient pycnidial mutants but caused no significant reduction of pathogenicity to chickpea. Pycnidiospores in ArPKS1-mutant pycnidia showed higher sensitivity to UV light exposure compared to pycnidiospores in melanized pycnidia of the wild-type progenitor isolate. Integration of an orthologous PKS1 gene from Bipolaris oryzae into the genome of the mutants complemented the dysfunctional ArPKS1 gene. This study demonstrated that A. Rabiei uses a DHN-melanin pathway for pigmentation of pycnidia and this molecule may protect pycnidiospores from UV irradiation.
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historical and contemporary multilocus population structure of Ascochyta Rabiei teleomorph didymella Rabiei in the pacific northwest of the united states
Molecular Ecology, 2004Co-Authors: Tobin L Peever, W. J. Kaiser, S S Salimath, F J MuehlbauerAbstract:The historical and contemporary population genetic structure of the chickpea Ascochyta blight pathogen, Ascochyta Rabiei (teleomorph: Didymella Rabiei), was determined in the US Pacific Northwest (PNW) using 17 putative AFLP loci, four genetically characterized, sequence-tagged microsatellite loci (STMS) and the mating type locus (MAT). A single multilocus genotype of A. Rabiei (MAT1-1) was detected in 1983, which represented the first recorded appearance of Ascochyta blight of chickpea in the PNW. During the following year many additional alleles, including the other mating type allele (MAT1-2), were detected. By 1987, all alleles currently found in the PNW had been introduced. Highly significant genetic differentiation was detected among contemporary subpopulations from different hosts and geographical locations indicating restricted gene flow and/or genetic drift occurring within and among subpopulations and possible selection by host cultivar. Two distinct populations were inferred with high posterior probability which correlated to host of origin and date of sample using Bayesian model-based population structure analyses of multilocus genotypes. Allele frequencies, genotype distributions and population assignment probabilities were significantly different between the historical and contemporary samples of isolates and between isolates sampled from a resistance screening nursery and those sampled from commercial chickpea fields. A random mating model could not be rejected in any subpopulation, indicating the importance of the sexual stage of the fungus both as a source of primary inoculum for Ascochyta blight epidemics and potentially adaptive genotypic diversity.
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cloning and characterization of the mating type mat locus from Ascochyta Rabiei teleomorph didymella Rabiei and a mat phylogeny of legume associated Ascochyta spp
Fungal Genetics and Biology, 2003Co-Authors: M P Barve, F J Muehlbauer, S S Salimath, Tsutomu Arie, Tobin L PeeverAbstract:Degenerate primers designed to correspond to conserved regions of the high mobility group (HMG) protein encoded by the MAT1-2 gene of Cochliobolus heterostrophus, Cochliobolus sativus, and Alternaria alternata were used to amplify the portion of the sequence corresponding to the HMG box motif from Ascochyta Rabiei (teleomorph: Didymella Rabiei). A combination of TAIL and inverse PCR extended the MAT1-2 sequence in both directions, then primers designed to MAT1-2 flanking DNA were used to amplify the entire MAT1-1 idiomorph. MAT1-1 and MAT1-2 idiomorphs were 2294 and 2693 bp in length, respectively, and each contained a single putative open reading frame (ORF) and intron similar to MAT loci of other loculoascomycete fungi. MAT genes were expressed at high levels in rich medium. MAT-specific PCR primers were designed for use in a multiplex PCR assay and MAT-specific PCR amplicons correlated perfectly to mating phenotype of 35 ascospore progeny from a cross of MAT1-1 by MAT1-2 isolates and to the mating phenotype of field-collected isolates from diverse geographic locations. MAT-specific PCR was used to rapidly determine the mating type of isolates of A. Rabiei sampled from chickpea fields in the US Pacific Northwest. Mating type ratios were not significantly different from 1:1 among isolates sampled from two commercial chickpea fields consistent with the hypothesis that these A. Rabiei populations were randomly mating. The mating type ratio among isolates sampled from an experimental chickpea field where asexual reproduction was enforced differed significantly from 1:1. A phylogeny estimated among legume-associated Ascochyta spp. and related loculoascocmycete fungi using sequence data from the nuclear ribosomal internal transcribed spacer (ITS) demonstrated the monophyly of Ascochyta/Didymella spp. associated with legumes but was insufficiently variable to differentiate isolates associated with different legume hosts. In contrast, sequences of the HMG region of MAT1-2 were substantially more variable, revealing seven well-supported clades that correlated to host of isolation. A. Rabiei on chickpea is phylogenetically distant from other legume-associated Ascochyta spp. and the specific status of A. Rabiei, A. lentis, A. pisi, and A. fabae was confirmed by the HMG phylogeny
