Kerstin Flath

Multiple alleles for resistance and susceptibility modulate the defense response in the interaction of tetraploid potato (Solanum tuberosum) with Synchytrium endobioticum pathotypes 1, 2, 6 and 18

The obligate biotrophic, soil-borne fungus Synchytrium endobioticum causes wart disease of potato (Solanum tuberosum), which is a serious problem for crop production in countries with moderate climates. S. endobioticum induces hypertrophic cell divisions in plant host tissues leading to the formation of tumor-like structures. Potato wart is a quarantine disease and chemical control is not possible. From 38 S. endobioticum pathotypes occurring in Europe, pathotypes 1, 2, 6 and 18 are the most relevant. Genetic resistance to wart is available but only few current potato varieties are resistant to all four pathotypes. The phenotypic evaluation of wart resistance is laborious, time-consuming and sometimes ambiguous, which makes breeding for resistance difficult. Molecular markers diagnostic for genes for resistance to S. endobioticum pathotypes 1, 2, 6 and 18 would greatly facilitate the selection of new, resistant cultivars. Two tetraploid half-sib families (266 individuals) segregating for resistance to S. endobioticum pathotypes 1, 2, 6 and 18 were produced by crossing a resistant genotype with two different susceptible ones. The families were scored for five different wart resistance phenotypes. The distribution of mean resistance scores was quantitative in both families. Resistance to pathotypes 2, 6 and 18 was correlated and independent from resistance to pathotype 1. DNA pools were constructed from the most resistant and most susceptible individuals and screened with genome wide simple sequence repeat (SSR), inverted simple sequence region (ISSR) and randomly amplified polymorphic DNA (RAPD) markers. Bulked segregant analysis identified three SSR markers that were linked to wart resistance loci (Sen). Sen1-XI on chromosome XI conferred partial resistance to pathotype 1, Sen18-IX on chromosome IX to pathotype 18 and Sen2/6/18-I on chromosome I to pathotypes 2,6 and 18. Additional genotyping with 191 single nucleotide polymorphism (SNP) markers confirmed the localization of the Sen loci. Thirty-three SNP markers linked to the Sen loci permitted the dissection of Sen alleles that increased or decreased resistance to wart. The alleles were inherited from both the resistant and susceptible parents.

Managing potato wart: a review of present research status and future perspective

Key messageIdentification of resistance genes to potato wart disease caused bySynchytrium endobioticumis the key for developing diagnostic markers for breeding resistant cultivars. We present an overview on the current knowledge of this host-pathogen system and molecular advances while highlighting future research focus.AbstractPotato wart is a quarantined disease of cultivated potato (Solanum tuberosum L.) caused by the obligate biotrophic, soil-borne fungus Synchytrium endobioticum (Schilb.) Perc. Since its discovery by Schilberszky in 1896, the management of wart disease was enabled by research efforts focusing on understanding and classifying the causative agent, its mode of infection, pathogenesis, geographical distribution, detection and chemical control, on developing screening methods for host resistance and on genetic analyses, which led to the development of resistant cultivars. These early successes are currently challenged by new S. endobioticum pathotypes evolving and the increased risk of dissemination by potato tuber trade. New research efforts are therefore required to ensure continuation of effective and sustainable management of the potato wart disease. Advances in molecular biology and genomic tools offer potential for innovations. This review presents an overview on what we know about this complex host-pathogen interaction, highlights recent molecular work and embarks on an outlook towards future research directions.

Resistance to powdery mildew in Spanish barley landraces is controlled by different sets of quantitative trait loci

Twenty-two landrace-derived inbred lines from the Spanish Barley Core Collection (SBCC) were found to display high levels of resistance to a panel of 27 isolates of the fungus Blumeria graminis that exhibit a wide variety of virulences. Among these lines, SBCC145 showed high overall resistance and a distinctive spectrum of resistance compared with the other lines. Against this background, the main goal of the present work was to investigate the genetic basis underlying such resistance using a doubled haploid population derived from a cross between SBCC145 and the elite spring cultivar Beatrix. The population was genotyped with the 1,536-SNP Illumina GoldenGate Oligonucleotide Pool Assay (Barley OPA-1 or BOPA1 for short), whereas phenotypic analysis was performed using two B. graminis isolates. A major quantitative trait locus (QTL) for resistance to both isolates was identified on the long arm of chromosome 6H (6HL) and accounted for ca. 60% of the phenotypic variance. Depending on the B. graminis isolate tested, three other minor QTLs were detected on chromosomes 2H and 7H, which explained less than 5% of the phenotypic variation each. In all cases, the alleles for resistance derived from the Spanish parent SBCC145. The position, the magnitude of the effect observed and the proportion of phenotypic variation accounted for by the QTL on 6HL suggest this is a newly identified locus for broad-based resistance to powdery mildew.

Genomic architecture of potato resistance to Synchytrium endobioticum disentangled using SSR markers and the 8.3k SolCAP SNP genotyping array

BackgroundThe soil borne, obligate biotrophic fungus Synchytrium endobioticum causes tumor-like tissue proliferation (wart) in potato tubers and thereby considerable crop damage. Chemical control is not effective and unfriendly to the environment. S. endobioticum is therefore a quarantined pathogen. The emergence of new pathotypes of the fungus aggravate this agricultural problem. The best control of wart disease is the cultivation of resistant varieties. Phenotypic screening for resistant cultivars is however time, labor and material intensive. Breeding for resistance would therefore greatly benefit from diagnostic DNA markers that can be applied early in the breeding cycle. The prerequisite for the development of diagnostic DNA markers is the genetic dissection of the factors that control resistance to S. endobioticum in various genetic backgrounds of potato.ResultsProgeny of a cross between a wart resistant and a susceptible tetraploid breeding clone was evaluated for resistance to S. endobioticum pathotypes 1, 2, 6 and 18 most relevant in Europe. The same progeny was genotyped with 195 microsatellite and 8303 single nucleotide polymorphism (SNP) markers. Linkage analysis identified the multi-allelic locus Sen1/RSe-XIa on potato chromosome XI as major factor for resistance to all four S. endobioticum pathotypes. Six additional, independent modifier loci had smaller effects on wart resistance. Combinations of markers linked to Sen1/RSe-XIa resistance alleles with one to two additional markers were sufficient for obtaining high levels of resistance to S. endobioticum pathotypes 1, 2, 6 and 18 in the analyzed genetic background.ConclusionsPotato resistance to S. endobioticum is oligogenic with one major and several minor resistance loci. It is composed of multiple alleles for resistance and susceptibility that originate from multiple sources. The genetics of resistance to S. endobioticum varies therefore between different genetic backgrounds. The DNA markers described in this paper are the starting point for pedigree based selection of cultivars with high levels of resistance to S. endobioticum pathotypes 1, 2, 6 and 18.

Resistance to powdery mildew in one Spanish barley landrace hardly resembles other previously identified wild barley resistances

Two major quantitative trait loci (QTLs) associated with resistance to powdery mildew (Blumeria graminis f. sp. hordei) were previously identified on chromosome 7H of the Spanish barley line SBCC097. The two QTLs seemed to share the same chromosomal position as the major genes mlt and Mlf, which were formerly described in Hordeum vulgare ssp. spontaneum-derived lines. In the present work, different lines that carry mlt (RS42-6*O), Mlf (RS137-28*E), or a combination of both (SI-4 and SI-6) were compared with SBCC097 to evaluate their relatedness at the phenotypic, cellular, and genetic levels. The resistance of the lines was characterised by inoculating them with a set of 27 isolates of B. graminis, which displayed a wide range of virulence. It was revealed that SBCC097 possessed a distinctive resistance spectrum. Microscopic assessment of the cytological development of the resistance response showed that SBCC097 clearly formed fewer well-established colonies and secondary hyphae than the other lines. This was confirmed by the infection type recorded after visual inspection. Genetic analyses of all five lines, based on markers flanking the QTLs derived from SBCC097, supported the macroscopic and microscopic data and pointed to the presence of a combination of novel genes or alleles in SBCC097, which may be included in the category of “intermediate-acting” genes, governing resistance mainly at the post-penetration stage.

Virulence and diversity of Puccinia striiformis f. sp. tritici in Ethiopia

Stripe rust of wheat (Triticum spp.), caused by Puccinia striiformis f. sp. tritici, is an important disease in Ethiopia. To investigate the population structure of the pathogen, 107 single-pustule isolates were collected from four regions (northern, central, southern, and southeastern Ethiopia) and tested on 24 differential genotypes with known resistance genes. The isolates were classified into 39 pathotypes. All isolates were virulent on Yr7 and avirulent on Yr1, Yr5, Yr15, and YrSp. Virulence complexity of the isolates ranged from 7 to 16, with a mean of 11.72. No common pathotype was shared by these regional collections. Distribution of pathotypes within the regions was quite even, whereas the richness of populations varied considerably between 1.78 (the central region) and 2.96 (the southeastern region). The genotypic, gene, and genetic diversities within populations were characterized using the Simpson, Nei, and Kosman indices. Significant differences among the regional populations were detected for both gene and genetic diversity within populations, whereas the genotypic diversity was rather stable. The lowest and highest diversities of wheat stripe rust occurred in the southern and northern regions, respectively. Genetic variation in the population structure of the pathogen observed in different geographical areas might indicate that unique wheat cultivars should be developed and released for production on a regional basis.

Virulence phenotypes in powdery mildew (Blumeria graminis) populations and resistance genes in triticale (x Triticosecale)

Triticale (xTriticosecale) is a new host for powdery mildew (Blumeria graminis) being not infected by this pathogen in Germany before 2001. To evaluate population structure of the pathogen and race-specific resistances in the host, 694 isolates were collected in 12 states of Germany in the years 2007 to 2009 on triticale. They were tested by a newly developed initial differential set of 20 triticale cultivars. Corresponding virulences were found for all differentials except for cultivar Grenado. In total, 272 different virulence phenotypes (=pathotypes) were detected. The virulence complexity of the isolates ranged from 6 to 19 of 20 possible virulences with a mean of 15. In all years, a high level of diversity of the powdery mildew populations was observed with Simpson indices in the range 0.95 to 0.97. The distribution of the pathotypes was even across Germany with an Evenness index in the range 0.82 to 0.88. A set of 19 isolates with different virulence pathotypes and 10 cultivars were selected to be used to identify race-specific resistances of triticale cultivars and breeding lines. Some cultivars susceptible to most of the isolates in seedling stage were moderately resistant in adult-plant stage. The high diversity and complexity of the pathotypes found in German powdery mildew populations as well as an increasing acreage of only a few dominating triticale cultivars accelerate the adaptation of the pathogen to race-specific host resistances suggesting restricted durability only. More durable resistance might be achieved by combining new effective race-specific (qualitative) resistance genes with race-nonspecific (quantitative) resistances effective in the adult-plant stage that are already available.

Genomic and Transcriptomic Resources for Marker Development in Synchytrium endobioticum, an Elusive but Severe Potato Pathogen

Synchytrium endobioticum is an obligate biotrophic fungus that causes wart diseases in potato. Like other species of the class Chytridiomycetes, it does not form mycelia and its zoospores are small, approximately 3 μm in diameter, which complicates the detection of early stages of infection. Furthermore, potato wart disease is difficult to control because belowground organs are infected and resting spores of the fungus are extremely durable. Thus, S. endobioticum is classified as a quarantine organism. More than 40 S. endobioticum pathotypes have been reported, of which pathotypes 1(D1), 2(G1), 6(O1), 8(F1), and 18(T1) are the most important in Germany. No molecular methods for the differentiation of pathotypes are available to date. In this work, we sequenced both genomic DNA and cDNA of the German pathotype 18(T1) from infected potato tissue and generated 5,422 expressed sequence tags (EST) and 423 genomic contigs. Comparative sequencing of 33 genes, single-stranded confirmation polymorphism (SSCP) analysis with polymerase chain reaction fragments of 27 additional genes, as well as the analysis of 41 simple sequence repeat (SSR) loci revealed extremely low levels of variation among five German pathotypes. From these markers, one sequence-characterized amplified region marker and five SSR markers revealed polymorphisms among the German pathotypes and an extended set of 11 additional European isolates. Pathotypes 8(F1) and 18(T1) displayed discrete polymorphisms which allow their differentiation from other pathotypes. Overall, using the information of the six markers, the 16 isolates could be differentiated into three distinct genotype groups. In addition to the presented markers, the new collection of EST from genus Synchytrium might serve in the future for molecular taxonomic studies as well as for analyses of the host-pathogen interactions in this difficult pathosystem. [Formula: see text] Copyright

Analyzing Genetic Diversity for Virulence and Resistance Phenotypes in Populations of Stem Rust (Puccinia graminis f. sp. secalis) and Winter Rye (Secale cereale)

Stem rust (Puccinia graminis f. sp. secalis) leads to considerable yield losses in rye-growing areas with continental climate, from Eastern Germany to Siberia. For implementing resistance breeding, it is of utmost importance to (i) analyze the diversity of stem rust populations in terms of pathotypes (= virulence combinations) and (ii) identify resistance sources in winter rye populations. We analyzed 323 single-uredinial isolates mainly collected from German rye-growing areas across 3 years for their avirulence/virulence on 15 rye inbred differentials. Out of these, 226 pathotypes were detected and only 56 pathotypes occurred more than once. This high diversity was confirmed by a Simpson index of 1.0, a high Shannon index (5.27), and an evenness index of 0.97. In parallel, we investigated stem rust resistance among and within 121 heterogeneous rye populations originating mainly from Russia, Poland, Austria, and the United States across 3 to 15 environments (location-year combinations). While German rye populations had an average stem rust severity of 49.7%, 23 nonadapted populations were significantly (P < 0.01) more resistant with a stem rust severity ranging from 3 to 40%. Out of these, two modern Russian breeding populations and two old Austrian landraces were the best harboring 32 to 70% fully resistant plants across 8 to 10 environments. These populations with the lowest disease severity in adult-plant stage in the field also displayed resistance in leaf segment tests. In conclusion, stem rust populations are highly diverse and the majority of resistances in rye populations seems to be race specific.

Euphresco Sendo: An international laboratory comparison study of molecular tests for Synchytrium endobioticum detection and identification

An international test performance study (TPS) was organised to generate validation data for three molecular Synchytrium endobioticum tests: van den Boogert et al. (European Journal of Plant Pathology 113, 47–57, 2005), and van Gent-Pelzer et al. (European Journal of Plant Pathology, 126, 129-133, 2010) for the detection of S. endobioticum, and the pathotype 1(D1) identification test described by Bonants et al. (European Journal of Plant Pathology, 143, 495-506, 2015). Two TPS rounds were organised focussing on different test matrices, i.e. round 1: warted potato tissue, and round 2: resting spore suspensions. When using the tests for detection and identification of S. endobioticum in warted potato tissue, no significant differences were observed for diagnostic sensitivity, diagnostic specificity, overall accuracy, analytical sensitivity and robustness. When using the tests for detection and identification of S. endobioticum in resting spore suspensions, the van den Boogert and van Gent-Pelzer tests significantly outperform the Bonants test for diagnostic sensitivity and diagnostic specificity. For overall accuracy and analytical sensitivity, the van Gent-Pelzer significantly outperforms the van den Boogert and Bonants tests and is regarded as the test of choice when identifying S. endobioticum from resting spores. Tests regarded fit for purpose for routine testing of wart material and resting spore suspensions are proposed for the update of EPPO standard PM7/28(1) Synchytrium endobioticum.