Powdery Mildew Susceptibility Research Articles

Sugar Transporter HmSWEET8 Cooperates with HmSTP1 to Enhance Powdery Mildew Susceptibility in Heracleum moellendorffii Hance.

The powdery mildew caused by Eeysiphe heraclei is a serious concern in Heracleum moellendorffii Hance. Therefore, exploring the mechanisms underlying sugar efflux from host cells to the fungus during the plant-fungus interaction showed great significance. The study successfully cloned HmSWEET8 and HmSTP1 genes based on RNA-seq technology. The complementation assays in yeast EBY.VW4000 found HmSWEET8 and HmSTP1 transporting hexose. Over-expressing or silencing HmSWEET8 in H. moellendorffii leaves increased or decreased powdery mildew susceptibility by changing glucose concentration in infective sites. Meanwhile, over-expressing HmSTP1 in H. moellendorffii leaves also increased powdery mildew susceptibility by elevating the glucose content of infective areas. Additionally, HmSTP1 expression was up-regulated obviously in HmSWEET8 over-expressed plants and inhibited significantly in HmSWEET8 silenced plants. Co-expressing HmSWEET8 and HmSTP1 genes significantly increased powdery mildew susceptibility compared with over-expressed HmSWEET8 or HmSTP1 plants alone. The results demonstrated that HmSTP1 may assist with HmSWEET8 to promote E. heraclei infection. Consequently, the infection caused by E. heraclei resulted in the activation of HmSWEET8, leading to an increased transfer of glucose to the apoplasmic spaces at the sites of infection, then, HmSTP1 facilitated the transport of glucose into host cells, promoting powdery mildew infection.

Analysis of Expression, Mutation, and Alternative Splice Variants of Candidate Genes, MLO2 and MLO6A, Involved in Powdery Mildew Susceptibility in Mulberry (Morus spp.)

Analysis of Expression, Mutation, and Alternative Splice Variants of Candidate Genes, MLO2 and MLO6A, Involved in Powdery Mildew Susceptibility in Mulberry (Morus spp.)

Interplay of EXO70 and MLO proteins modulates trichome cell wall composition and susceptibility to powdery mildew.

Exocyst component of 70-kDa (EXO70) proteins are constituents of the exocyst complex implicated in vesicle tethering during exocytosis. MILDEW RESISTANCE LOCUS O (MLO) proteins are plant-specific calcium channels and some MLO isoforms enable fungal powdery mildew pathogenesis. We here detected an unexpected phenotypic overlap of Arabidopsis thaliana exo70H4 and mlo2 mlo6 mlo12 triple mutant plants regarding the biogenesis of leaf trichome secondary cell walls. Biochemical and Fourier transform infrared spectroscopic analyses corroborated deficiencies in the composition of trichome cell walls in these mutants. Transgenic lines expressing fluorophore-tagged EXO70H4 and MLO exhibited extensive colocalization of these proteins. Furthermore, mCherry-EXO70H4 mislocalized in trichomes of the mlo triple mutant and, vice versa, MLO6-GFP mislocalized in trichomes of the exo70H4 mutant. Expression of GFP-marked PMR4 callose synthase, a known cargo of EXO70H4-dependent exocytosis, revealed reduced cell wall delivery of GFP-PMR4 in trichomes of mlo triple mutant plants. In vivo protein-protein interaction assays in plant and yeast cells uncovered isoform-preferential interactions between EXO70.2 subfamily members and MLO proteins. Finally, exo70H4 and mlo6 mutants, when combined, showed synergistically enhanced resistance to powdery mildew attack. Taken together, our data point to an isoform-specific interplay of EXO70 and MLO proteins in the modulation of trichome cell wall biogenesis and powdery mildew susceptibility.

Cloning and expression analysis of Mildew Locus O (MLO) genes related to powdery mildew resistance in Vitis pseudoreticulata, and functional characterisation of VpMLO13

ABSTRACT In many plants, Mildew Locus O (MLO) genes facilitate infection by pathogens, including Erysiphe necator, the causative agent of powdery mildew (PM) in grapevines. The function of VvMLOs in Vitis vinifera has previously been demonstrated, but the identity and function of VpMLOs in V. pseudoreticulata are not yet clear. Here, we cloned MLO genes that have been implicated in PM susceptibility from the highly PM-resistant V. pseudoreticulata accession ‘Baihe-35-1’ and the PM-susceptible V. vinifera ‘Carinena’. The open reading frames (ORFs) of MLO3, MLO6, and MLO8 differed between ‘Baihe-35-1’ and ‘Carinena’. After inoculation of the leaves with Erysiphe necator, six MLO genes exhibited two distinct expression patterns in ‘Baihe-35-1’ and ‘Carinena’. Consistent with the MLO proteins in other plants, we also found that VpMLO3, VpMLO6, VpMLO11, and VpMLO13 are all localised to the plasma membrane. To further support a role for VpMLOs in susceptibility to PM, we expressed VpMlo13 in the PM-resistant Arabidopsis thaliana MLO2/6/12 triple mutant, which showed that transgenic expression of VpMLO13 restored susceptibility to PM. In addition, a yeast two-hybrid (Y2H) assay showed that VpMLO13 interacted with calmodulin (VpCaM). These results will provide a basis for further investigating the PM resistance mechanisms in V. pseudoreticulata.

Enhancing powdery mildew resistance in soybean by targeted mutation of MLO genes using the CRISPR/Cas9 system

BackgroundPowdery mildew is a major disease that causes great losses in soybean yield and seed quality. Disease-resistant varieties, which are generated by reducing the impact of susceptibility genes through mutation in host plants, would be an effective approach to protect crops from this disease. The Mildew Locus O (MLO) genes are well-known susceptibility genes for powdery mildew in plant. In this study, we utilized the CRISPR/Cas9 system to induce targeted mutations in the soybean GmMLO genes to improve powdery mildew resistance.ResultsA dual-sgRNA CRISPR/Cas9 construct was designed and successfully transferred into the Vietnamese soybean cultivar DT26 through Agrobacterium tumefaciens-mediated transformation. Various mutant forms of the GmMLO genes including biallelic, chimeric and homozygous were found at the T0 generation. The inheritance and segregation of CRISPR/Cas9-induced mutations were confirmed and validated at the T1 and T2 generations. Out of six GmMLO genes in the soybean genome, we obtained the Gmmlo02/Gmmlo19/Gmmlo23 triple and Gmmlo02/Gmmlo19/Gmmlo20/Gmmlo23 quadruple knockout mutants at the T2 generation. When challenged with Erysiphe diffusa, a fungus that causes soybean powdery mildew, all mutant plants showed enhanced resistance to the pathogen, especially the quadruple mutant. The powdery mildew severity in the mutant soybeans was reduced by up to 36.4% compared to wild-type plants. In addition, no pleiotropic effect on soybean growth and development under net-house conditions was observed in the CRISPR/Cas9 mutants.ConclusionsOur results indicate the involvement of GmMLO02, GmMLO19, GmMLO20 and GmMLO23 genes in powdery mildew susceptibility in soybean. Further research should be conducted to investigate the roles of individual tested genes and the involvement of other GmMLO genes in this disease infection mechanism. Importantly, utilizing the CRISPR/Cas9 system successfully created the Gmmlo transgene-free homozygous mutant lines with enhanced resistance to powdery mildew, which could be potential materials for soybean breeding programs.

The First Genome-Wide Mildew Locus O Genes Characterization in the Lamiaceae Plant Family.

Powdery mildew (PM) is a widespread plant disease that causes significant economic losses in thousands crops of temperate climates, including Lamiaceae species. Multiple scientific studies describe a peculiar form of PM-resistance associated at the inactivation of specific members of the Mildew Locus O (MLO) gene family, referred to as mlo-resistance. The characterization of Lamiaceae MLO genes, at the genomic level, would be a first step toward their potential use in breeding programs. We carried out a genome-wide characterization of the MLO gene family in 11 Lamiaceae species, providing a manual curated catalog of 324 MLO proteins. Evolutionary history and phylogenetic relationships were studied through maximum likelihood analysis and motif patter reconstruction. Our approach highlighted seven different clades diversified starting from an ancestral MLO domain pattern organized in 18 highly conserved motifs. In addition, 74 Lamiaceae putative PM susceptibility genes, clustering in clade V, were identified. Finally, we performed a codon-based evolutionary analysis, revealing a general high level of purifying selection in the eleven Lamiaceae MLO gene families, and the occurrence of few regions under diversifying selection in candidate susceptibility factors. The results of this work may help to address further biological questions concerning MLOs involved in PM susceptibility. In follow-up studies, it could be investigated whether the silencing or loss-of-function mutations in one or more of these candidate genes may lead to PM resistance.

Identification of acyl-CoA-binding protein gene in Triticeae species reveals that TaACBP4A-1 and TaACBP4A-2 positively regulate powdery mildew resistance in wheat

Plant acyl-CoA-binding proteins (ACBPs), which contain the conserved ACB domain, participate in multiple biological processes, however, there are few reports on wheat ACBPs. In this study, the ACBP genes from nine different species were identified comprehensively. The expression patterns of TaACBP genes in multiple tissues and under various biotic stresses were determined by qRT-PCR. The function of selected TaACBP genes was studied by virus-induced gene silencing. A total of 67 ACBPs were identified from five monocotyledonous and four dicotyledonous species and divided into four classes. Tandem duplication analysis of the ACBPs suggested that tandem duplication events occurred in Triticum dicoccoides, but there was no tandem duplication event in wheat ACBP genes. Evolutionary analysis suggested that the TdACBPs may have experienced gene introgression during tetraploid evolution, while TaACBP gene loss events occurred during hexaploid wheat evolution. The expression pattern showed that all the TaACBP genes were expressed, and most of them were responsive to induction by Blumeria graminis f. sp. tritici or Fusarium graminearum. Silencing of TaACBP4A-1 and TaACBP4A-2 increased powdery mildew susceptibility in the common wheat BainongAK58. Furthermore, TaACBP4A-1, which belonged to class III, physically interacted with autophagy-related ubiquitin-like protein TaATG8g in yeast cells. This study provided a valuable reference for further investigations into the functional and molecular mechanisms of the ACBP gene family.

Comparative Analysis of Powdery Mildew Disease Resistance and Susceptibility in Brassica Coenospecies

Erysiphe cruciferarum, a causative agent of powdery mildew disease, has emerged as a serious threat in Brassica juncea and its closely related species. To date, no resistant cultivars have been identified in Brassica species against powdery mildew. Here, we used histopathological, biochemical, and molecular approaches to elucidate the powdery mildew disease progression and host responses in three Brassica cenospecies, namely B. juncea, Camelina sativa, and Sinapis alba. Based on the results of disease progression, S. alba was found to be extremely resistant to powdery mildew infection, whereas B. juncea and C. sativa were highly vulnerable. In addition, the disease spread rate to uninfected parts was comparatively higher in B. juncea and C. sativa. Histopathological results revealed more pathogen-induced cell death in B. juncea and C. sativa compared to S. alba. We also examined the role of antioxidant enzymes such as superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) in B. juncea, C. sativa, and S. alba after powdery mildew infection. Based on our findings, the enzyme activity of SOD, POD, and CAT was relatively higher in S. alba then that of B. juncea and C. sativa after powdery mildew infection. Furthermore, we evaluated the expression levels of salicylic acid (SA) signature genes, including pathogenesis-related protein viz., PR1, PR2, and PR5 in B. juncea, C. sativa, and S. alba after E. cruciferarum infection. Based on our findings, the expression levels of SA marker genes PR1, PR2, and PR5 increased in all three species after infection. However, the fold change was relatively higher in S. alba than in B. juncea and C. sativa. In future, further studies are required to identify the potential candidates in S. alba that are involved in powdery mildew disease resistance.

Genome-wide characterization and expression analysis of the MLO gene family sheds light on powdery mildew resistance in Lagenaria siceraria

MLO (mildew locus O) genes play a vital role in plant disease defense system, especially powdery mildew (PM). Lagenaria siceraria is a distinct Cucurbitaceae crop, and PM is one of the most serious diseases threatening crop production and quality. Although MLOs have been exploited in many Cucurbitaceae species, genome-wide mining of MLO gene family in bottle gourd has not been surveyed yet. Here we identified 16 MLO genes in our recently assembled L. siceraria genome. A total of 343 unique MLO protein sequences from 20 species were characterized and compared to deduce a generally high level of purifying selection and the occurrence of regions related to candidate susceptibility factors in the evolutional divergence. LsMLOs were clustered in six clades containing seven conserved transmembrane domains and 10 clade-specific motifs along with deletion and variation. Three genes (LsMLO3, LsMLO6, and LsMLO13) in clade V showed high sequence identity with orthologues involved in PM susceptibility. The expression pattern of LsMLOs was tissue-specific but not cultivar-specific. Furthermore, it was indicated by qRT-PCR and RNA-seq that LsMLO3 and LsMLO13 were highly upregulated in response to PM stress. Subsequent sequence analysis revealed the structural deletion of LsMLO13 and a single nonsynonymous substitution of LsMLO3 in the PM-resistant genotype. Taken all together, it is speculated that LsMLO13 is likely a major PM susceptibility factor. The results of this study provide new insights into MLO family genes in bottle gourd and find a potential candidate S gene for PM tolerance breeding.

Evaluation of Malus Germplasm Identifies Genetic Sources of Powdery Mildew and Frogeye Leaf Spot Resistance for Apple Breeding.

Apple is an important fruit crop of temperate regions. The narrow genetic base of commercially cultivated apples has resulted in its vulnerability to a large number of fungal, bacterial, and viral pathogens. Apple breeders are always seeking new sources of resistance within the cross-compatible Malus species that can be deployed into elite genetic backgrounds. We have evaluated resistance to two major fungal diseases of apples: powdery mildew and frogeye leaf spot, using a germplasm collection of 174 Malus accessions to identify novel sources of genetic resistance. In 2020 and 2021, we evaluated these accessions for the incidence and severity of powdery mildew and frogeye leaf spot diseases at Cornell AgriTech, Geneva, New York, in a partially managed orchard. The severity and incidence of powdery mildew and frogeye leaf spot, as well as weather parameters were recorded in June, July, and August. Total incidence of powdery mildew and frogeye leaf spot infections increased from 33 to 38%, and 56 to 97% in 2020 and 2021, respectively. Our analysis showed that relative humidity and precipitation correlate with powdery mildew and frogeye leaf spot susceptibility. The predictor variables with highest impact to the variability of powdery mildew were accessions and relative humidity in May. A total of 65 Malus accessions were found to be resistant to powdery mildew, and only one accession showed moderate resistance to frogeye leaf spot. Several of these accessions belong to Malus hybrid species and domesticated apples and can therefore be potential sources of novel resistance alleles for apple breeding.

Volatile-mediated signalling in barley induces metabolic reprogramming and resistance against the biotrophic fungus Blumeria hordei.

Plants have evolved diverse secondary metabolites to counteract biotic stress. Volatile organic compounds (VOCs) are released upon herbivore attack or pathogen infection. Recent studies suggest that VOCs can act as signalling molecules in plant defence and induce resistance in distant organs and neighbouring plants. However, knowledge is lacking on the function of VOCs in biotrophic fungal infection on cereal plants. We analysed VOCs emitted by 13 ± 1-day-old barley plants (Hordeum vulgare L.) after mechanical wounding using passive absorbers and TD-GC/MS. We investigated the effect of pure VOC and complex VOC mixtures released from wounded plants on the barley-powdery mildew interaction by pre-exposure in a dynamic headspace connected to a powdery mildew susceptibility assay. Untargeted metabolomics and lipidomics were applied to investigate metabolic changes in sender and receiver barley plants. Green leaf volatiles (GLVs) dominated the volatile profile of wounded barley plants, with (Z)-3-hexenyl acetate (Z3HAC) as the most abundant compound. Barley volatiles emitted after mechanical wounding enhanced resistance in receiver plants towards fungal infection. We found volatile-mediated modifications of the plant-pathogen interaction in a concentration-dependent manner. Pre-exposure with physiologically relevant concentrations of Z3HAC resulted in induced resistance, suggesting that this GLV is a key player in barley anti-pathogen defence. The complex VOC mixture released from wounded barley and Z3HAC induced e.g. accumulation of chlorophyll, linolenic acid and linolenate-conjugated lipids, as well as defence-related secondary metabolites, such as hordatines in receiving plants. Barley VOCs hence induce a complex physiological response and disease resistance in receiver plants.

Temperature Influences on Powdery Mildew Susceptibility and Development in the Hop Cultivar Cascade.

The hop cultivar 'Cascade' possesses partial resistance to powdery mildew (Podosphaera macularis) that can be overcome by recently emerged, virulent isolates of the fungus. Given that hop is a long-lived perennial and that brewers still demand Cascade, there is a need to better understand factors that influence the development of powdery mildew on this cultivar. Growth chamber experiments were conducted to quantify the effect of constant, transient, and fluctuating temperature on Cascade before, concurrent to, and after inoculation as contrasted with another powdery mildew-susceptible cultivar, 'Symphony'. Exposure of plants to supraoptimal temperature (26 and 32°C) before inoculation led to more rapid onset of ontogenic resistance in intermediately aged leaves in Cascade as compared with Symphony. Cascade was overall less susceptible to powdery mildew when exposed to constant temperature ranging from 18 to 32°C directly after inoculation. However, cultivar also interacted with temperature such that proportionately fewer and smaller colonies developed on Cascade than Symphony at supraoptimal yet permissive temperatures for disease. When plants were inoculated and then exposed to high temperature, colonies became progressively more tolerant to temperatures of 26 to 30°C with increasing time from inoculation to exposure, as moderated by cultivar, the specific temperature, and their interaction. Subjecting plants to simulated diurnal temperature regimes at the time of inoculation or 24 h later indicated Cascade and Symphony responded proportionately similarly on days predicted to be marginally unfavorable or marginally favorable for powdery mildew, although Cascade was quantitatively less susceptible than Symphony. In sum, this research indicates that Cascade is overall less susceptible to powdery mildew than Symphony, and supraoptimal temperature before, concurrent to, or after infection may interact differentially to moderate disease risk in Cascade. Therefore, cultivar-specific risk assessments for powdery mildew appear warranted.

Unmasking Mildew Resistance Locus O.

Loss of Mildew Resistance Locus O (MLO) in barley confers durable resistance to powdery mildew fungi, which has led to its wide deployment in agriculture. Although MLO is a susceptibility factor, it has become nearly synonymous with powdery mildew resistance. However, MLO has been recently implicated in colonization by arbuscular mycorrhizal fungi and a fungal endophyte, confirming its importance for biotrophic interactions and in promoting symbiosis. Other MLO proteins are involved in essential sensory processes, particularly fertilization and thigmotropism. We propose external stimulus perception as a common theme in these interactions and consider a unified biochemical role, potentially relating to reactive oxygen species (ROS) and calcium regulation, for MLOs across tissues and processes.

Genome-Wide Characterization of the MLO Gene Family in Cannabis sativa Reveals Two Genes as Strong Candidates for Powdery Mildew Susceptibility.

Cannabis sativa is increasingly being grown around the world for medicinal, industrial, and recreational purposes. As in all cultivated plants, cannabis is exposed to a wide range of pathogens, including powdery mildew (PM). This fungal disease stresses cannabis plants and reduces flower bud quality, resulting in significant economic losses for licensed producers. The Mildew Locus O (MLO) gene family encodes plant-specific proteins distributed among conserved clades, of which clades IV and V are known to be involved in susceptibility to PM in monocots and dicots, respectively. In several studies, the inactivation of those genes resulted in durable resistance to the disease. In this study, we identified and characterized the MLO gene family members in five different cannabis genomes. Fifteen Cannabis sativa MLO (CsMLO) genes were manually curated in cannabis, with numbers varying between 14, 17, 19, 18, and 18 for CBDRx, Jamaican Lion female, Jamaican Lion male, Purple Kush, and Finola, respectively (when considering paralogs and incomplete genes). Further analysis of the CsMLO genes and their deduced protein sequences revealed that many characteristics of the gene family, such as the presence of seven transmembrane domains, the MLO functional domain, and particular amino acid positions, were present and well conserved. Phylogenetic analysis of the MLO protein sequences from all five cannabis genomes and other plant species indicated seven distinct clades (I through VII), as reported in other crops. Expression analysis revealed that the CsMLOs from clade V, CsMLO1 and CsMLO4, were significantly upregulated following Golovinomyces ambrosiae infection, providing preliminary evidence that they could be involved in PM susceptibility. Finally, the examination of variation within CsMLO1 and CsMLO4 in 32 cannabis cultivars revealed several amino acid changes, which could affect their function. Altogether, cannabis MLO genes were identified and characterized, among which candidates potentially involved in PM susceptibility were noted. The results of this study will lay the foundation for further investigations, such as the functional characterization of clade V MLOs as well as the potential impact of the amino acid changes reported. Those will be useful for breeding purposes in order to develop resistant cultivars.

Evolution of the MLO gene families in octoploid strawberry (Fragaria \xd7ananassa) and progenitor diploid species identified potential genes for strawberry powdery mildew resistance

Powdery mildew (PM) caused by Podosphaera aphanis is a major fungal disease of cultivated strawberry. Mildew Resistance Locus O (MLO) is a gene family described for having conserved seven-transmembrane domains. Induced loss-of-function in specific MLO genes can confer durable and broad resistance against PM pathogens. However, the genomic structure and potential role of MLO genes for PM resistance have not been characterized yet in the octoploid cultivated strawberry. In the present study, MLO gene families were characterized in four diploid progenitor species (Fragaria vesca, F. iinumae, F. viridis, and F. nipponica) and octoploid cultivated (Fragaria ×ananassa) strawberry, and potential sources of MLO-mediated susceptibility were identified. Twenty MLO sequences were identified in F. vesca and 68 identified in F. ×ananassa. Phylogenetic analysis divided diploid and octoploid strawberry MLO genes into eight different clades, in which three FveMLO (MLO10, MLO17, and MLO20) and their twelve orthologs of FaMLO were grouped together with functionally characterized MLO genes conferring PM susceptibility. Copy number variations revealed differences in MLO composition among homoeologous chromosomes, supporting the distinct origin of each subgenome during the evolution of octoploid strawberry. Dissecting genomic sequence and structural variations in candidate FaMLO genes revealed their potential role associated with genetic controls and functionality in strawberry against PM pathogen. Furthermore, the gene expression profiling and RNAi silencing of putative FaMLO genes in response to the pathogen indicate the function in PM resistance. These results are a critical first step in understanding the function of strawberry MLO genes and will facilitate further genetic studies of PM resistance in cultivated strawberry.

Evaluation of powdery mildew resistance of a diverse set of grape cultivars and testing the association between powdery mildew resistance and PR gene expression

Powdery mildew caused by the fungal pathogen Erysiphe necator is a global disease that reduces yield and quality in grapes (Vitis vinifera L.). Most grape cultivars are susceptible to this pathogen and thus the selection of powdery mildew resistant cultivars is a major objective of grape breeding programs. In this study, we evaluated powdery mildew resistance of a diverse set of 28 commonly cultivated local and international V. Vinifera cultivars after artificial inoculations with Erysiphe necator. Of these, 7 cultivars were found to be resistant while 10 and 11, were found to be moderately resistant and susceptible to powdery mildew, respectively. We also analyzed pathogen-responsive expression profiles of two defense genes, VvPR1 and VvGLP3 encoding a pathogenesis-related protein 1 homolog and a Germin-Like protein, respectively, in the same cultivars by qRT-PCR to determine whether defense gene expression is correlated with powdery mildew resistance. There was no significant correlation between powdery mildew ratings and induction patterns of these two defense genes. However, a significant correlation was found between VvPR1 and VvGLP3 expression only in cultivars that showed increased powdery mildew resistance. In addition, the grape cultivars we analyzed differed by berry ripening times, were shown to influence disease resistance. However, no significant correlation was found between powdery mildew ratings and the ripening times. Although potential mechanisms of powdery mildew resistance or susceptibility remain unclear, powdery mildew resistant grape cultivars identified here will be useful for growers and breeders interested in using these lines as parents in grape breeding programs.

The MLO1 powdery mildew susceptibility gene in Lathyrus species: The power of high-density linkage maps in comparative mapping and synteny analysis.

Powdery mildews are major diseases for a range of crops. The loss of function of specific Mildew Locus O (MLO) genes has long been associated with pre-haustorial plant resistance to powdery mildew and has proven to be durable in several species. Erysiphe pisi is the major causal agent of powdery mildew in pea (Pisum sativum L.) and in the closely related Lathyrus sativus L. and Lathyrus cicera L. PsMLO1 has been extensively studied in pea. However, no MLO gene family members have been isolated and characterized in Lathyrus species so far. In this study, MLO1 genes were isolated and characterized in L. sativus and L. cicera genotypes with varied levels of partial resistance against powdery mildew. Phylogenetic analyses confirmed that Lathyrus MLO1 belongs to Clade V, like all dicot MLO proteins associated with powdery mildew susceptibility. A L. sativus recombinant inbred line population (RIL) was genotyped by sequencing to develop a high-density L. sativus genetic linkage map. DNA sequence polymorphisms between the analyzed genotypes allowed the location of MLO1 in the newly developed L. sativus RIL genetic linkage map. Subsequent comparative mapping between L. sativus and L. cicera genetic maps and P. sativum, Lens culinaris Medik., and Medicago truncatula Gaertn. reference genomes revealed important aspects of the conservation of the MLO1 locus position and of the overall chromosomal rearrangements occurring during legume evolution, with relevance to legume disease resistance breeding programs.

Analysis of allelic variants of RhMLO genes in rose and functional studies on susceptibility to powdery mildew related to clade V homologs

Key messageRose has 19 MLO genes. Of these, RhMLO1 and RhMLO2 were shown to be required for powdery mildew infection, which suggests their potential as susceptibility targets towards disease resistance.Powdery mildew, caused by Podosphaera pannosa, is one of the most serious and widespread fungal diseases for roses, especially in greenhouse-grown cut roses. It has been shown that certain MLO genes are involved in powdery mildew susceptibility and that loss of function in these genes in various crops leads to broad-spectrum, long-lasting resistance against this fungal disease. For this reason, these MLO genes are called susceptibility genes. We carried out a genome-wide identification of the MLO gene family in the Rosa chinensis genome, and screened for allelic variants among 22 accessions from seven different Rosa species using re-sequencing and transcriptome data. We identified 19 MLO genes in rose, of which four are candidate genes for functional homologs in clade V, which is the clade containing all dicot MLO susceptibility genes. We detected a total of 198 different allelic variants in the set of Rosa species and accessions, corresponding to 5–15 different alleles for each of the genes. Some diploid Rosa species shared alleles with tetraploid rose cultivars, consistent with the notion that diploid species have contributed to the formation of tetraploid roses. Among the four RhMLO genes in clade V, we demonstrated using expression study, virus-induced gene silencing as well as transient RNAi silencing that two of them, RhMLO1 and RhMLO2, are required for infection by P. pannosa and suggest their potential as susceptibility targets for powdery mildew resistance breeding in rose.

Identification of Novel Loci and Candidate Genes for Resistance to Powdery Mildew in a Resequenced Cucumber Germplasm.

Powdery mildew (PM) is one of the most serious diseases in cucumber and causes huge yield loss. Multiple quantitative trait loci (QTLs) for PM resistance have been reported in previous studies using a limited number of cucumber accessions. In this study, a cucumber core germplasm (CG) consisting of 94 resequenced lines was evaluated for PM resistance in four trials across three years (2013, 2014, and 2016). These trials were performed on adult plants in the field with natural infection. Using genome-wide association study (GWAS), 13 loci (pmG1.1, pmG1.2, pmG2.1, pmG2.2, pmG3.1, pmG4.1, pmG4.2, pmG5.1, pmG5.2, pmG5.3, pmG5.4, pmG6.1, and pmG6.2) associated with PM resistance were detected on all chromosomes except for Chr.7. Among these loci, ten were mapped to chromosomal intervals where QTLs had been reported in previous studies, while, three (pmG2.1, pmG3.1, and pmG4.1) were novel. The loci of pmG2.1, pmG5.2, pmG5.3 showed stronger signal in four trials. Based on the annotation of homologous genes in Arabidopsis and pairwise LD correlation analysis, candidate genes located in the QTL intervals were predicted. SNPs in these candidate genes were analyzed between haplotypes of highly resistant (HR) and susceptible (HS) CG lines, which were defined based on combing disease index data of all trials. Furthermore, candidate genes (Csa5G622830 and CsGy5G015660) reported in previous studies for PM resistance and cucumber orthologues of several PM susceptibility (S) genes (PMR5, PMR-6, and MLO) that are colocalized with certain QTLs, were analyzed for their potential contribution to the QTL effect on both PM and DM in the CG population. This study shows that the CG germplasm is a very valuable resource carrying known and novel QTLs for both PM and DM resistance, which can be exploited in cucumber breeding.

Genome-wide identification and analysis of the MLO gene family for candidate powdery mildew susceptibility factors in Momordica charantia

Powdery mildew (PM) disease is one of the most serious threats to Momordica charantia. Specific mildew resistance locus O (MLO) genes associated with PM susceptibility have been explored in some plant species for mlo-based PM resistance. However, MLO has not been surveyed globally for candidate susceptibility factors in M. charantia. In this study, 15 MLO genes, designated McMLO1∼15, were identified in the recently reported M. charantia genome. Phylogenetic analysis clustered McMLOs into five clades. In clade V, McMLO3, McMLO4, McMLO6, and McMLO11 co-clustered with the known PM susceptibility (PMS) factors of dicotyledonous plants, making them putative PMS factors in M. charantia (McPMS). Three of the four McPMS candidates, McMLO4, McMLO6, and McMLO11, showed much higher sequence identity with PMS-related orthologues than with paralogues. Structure modelling and alignment revealed significant structural similarity between McMLO11 and McMLO4 and the functionally characterized PMS CsMLO8, CmMLO2, and CsMLO11. High structural similarity was also found between McMLO11 and McMLO6 but not between McMLO3 and any other MLO homologs. RNA-seq indicated that the four McPMS candidates were all specifically expressed at the highest level in leaves compared with other tissues. Furthermore, among the expressed McMLOs in leaves, McMLO6 was expressed at the lowest level, McMLO11, McMLO3 and McMLO4 were expressed at the first, second and fourth highest levels, respectively. During PM infection, McMLO11 was moderately induced but highly expressed at the highest level. McMLO4 and McMLO6 were significantly upregulated, but there was no significant differential expression of McMLO3. Consistent with the different expression profiles of the four McPMSs, different types and amounts of cis-acting regulatory elements were found in their promoters. Taken together, we speculate that in M. charantia McMLO4 is a minor PMS factor, and that McMLO11 is the major PMS factor and is functionally redundant with McMLO6. McMLO3 seems to be a unique PMS factor with a different functional mechanism. The results of the study provide valuable information for further study of mlo-based PM resistance improvement in M. charantia.