Albrecht Serfling

Performance of the Biocontrol Fungus Piriformospora indica on Wheat Under Greenhouse and Field Conditions

ABSTRACT The endophyte Piriformospora indica colonizes roots of a range of host plants and increases biomass production and resistance to fungal pathogens and, thus has been considered a biocontrol fungus. However, the field performance of this fungus has not yet been tested in temperate climates. Therefore, we evaluated the performance of this fungus in different substrata under greenhouse and practical field conditions. Roots of winter wheat were colonized efficiently, and biomass was particularly increased on poor substrata. In greenhouse experiments, symptom severity of a typical leaf (Blumeria graminis f. sp. tritici), stem base (Pseudocercosporella herpotrichoides), and root (Fusarium culmorum) pathogen was reduced significantly. However, in field experiments, symptoms caused by the leaf pathogen did not differ in Piriformospora indica-colonized compared with control plants. In the field, Pseudocercosporella herpotrichoides disease severity was significantly reduced in plants colonized by the endophyte. Increased numbers of sheath layers and hydrogen peroxide concentrations after B. graminis attack were detected in Piriformospora indica-colonized plants, suggesting that root colonization causes induction of systemic resistance or priming of the host plant. Although the endophyte is not well suited for growth at Central European temperature conditions, it remains to be shown whether P. indica is more suitable for tropical or subtropical farming.

Diagnostic value of molecular markers for Lr genes and characterization of leaf rust resistance of German winter wheat cultivars with regard to the stability of vertical resistance

Breeding for resistance is an efficient strategy to manage wheat leaf rust caused by Puccinia triticina f. sp. tritici. However, a prerequisite for the directed use of Lr genes in breeding and the detection of new races virulent to these Lr genes is a detailed knowledge on Lr genes present in wheat cultivars. Therefore, respective molecular markers for 18 Lr genes were tested for specificity and used to determine Lr genes in 115 wheat cultivars. Results obtained were compared to available pedigree data. Using respective molecular markers, genes Lr1, Lr10, Lr26, Lr34 and Lr37 were detected, but data were not always in accordance with pedigree data. However, leaf rust scoring data of field trials confirmed the reliability of DNA markers. These reliable marker data facilitated the analyses of the development of virulent leaf rust races from 2002 to 2009 based on released cultivars. A sudden change from low infection rates to susceptibility was observed for Lr1, Lr3, Lr10, Lr13, Lr14, Lr16, Lr26 and Lr37 since 2006. Cultivars carrying several leaf rust resistance genes showed no significant shift to susceptibility except one cultivar which revealed an increasing infection rate at a low level. In summary, it turned out that pedigree data are often not reliable and a detection of Lr genes by diagnostic markers is fundamental to combine Lr genes in cultivars for a durable resistance against leaf rust, and to conduct reliable surveys based on released cultivars, instead of ‘Thatcher’ NILs.

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.

High-resolution mapping of the barley Ryd3 locus controlling tolerance to BYDV

Barley yellow dwarf disease (BYD) is transmitted by aphids and is caused by different strains of Barley yellow dwarf virus (BYDV) and Cereal yellow dwarf virus (CYDV). Economically it is one of the most important diseases of cereals worldwide. Besides chemical control of the vector, growing of tolerant/resistant cultivars is an effective way of protecting crops against BYD. The Ryd3 gene in barley (Hordeum vulgare L.) confers tolerance to BYDV-PAV and BYDV-MAV and the locus was previously mapped on the short arm of barley chromosome 6H near the centromere. We applied a strategy for high-resolution mapping and marker saturation at the Ryd3 locus by exploiting recent genomic tools available in barley. In a population of 3,210 F2 plants, 14 tightly linked markers were identified, including 10 that co-segregated with Ryd3. The centromeric region where Ryd3 is located suffers suppressed recombination or reduced recombination rate, suggesting potential problems in achieving (1) map-based cloning of Ryd3 and (2) marker selection of the resistance in breeding programmes without the introduction of undesirable traits via linkage drag.

Resistance of (Aegilops tauschii × Secale cereale) × Triticosecale Hybrids to leaf rust (Puccinia triticina) determined on the macroscopic and microscopic level

Leaf rust caused by Puccinia triticina Eriks belongs to the most important fungal pathogens of wheat (Triticum aestivum L.) and triticale (× Triticosecale). Effective resistance to leaf rust is both, cost-effective and environmentally safe. Many wild Aegilops species carry unknown resistances against fungal diseases and are characterized by a high genetic variability. The main goal of this work was to examine the resistance of (Aegilops tauschii × Secale cereale) × Triticosecale hybrids to leaf rust in inoculation tests with different races of P. triticina. Hybrid plants were selected for the presence of 2D chromosome/s in the triticale background using fluorescence and genomic in situ hybridization. The presence of leaf rust resistance genes was confirmed with closely linked molecular markers, i.e., Xgdm35 and Xgwm296. 14 genotypes of BC2F4 – BC2F6 hybrid plants with the monosomic addition of chromosome 2D (M2DA) were analyzed together with nine control lines. Resistance was determined at the macroscopic and microscopic level at the seedling and adult plant stage (flag leaf). In general, results revealed limited resistance of hybrid plants at the seedling stage, followed by an increase of the resistance level at later stages of plant development. This indicates that respective hybrid plants may exhibit APR resistance conferred by Lr22a introgressed from Ae. tauschii. On the basis of the macroscopic and microscopic analysis, this kind of resistance turned out to be additive and race-specific. We selected four monosomic 2D addition triticale genotypes highly resistant to P. triticina infection at the two main stages of plant development. From the selected genotypes, we obtained 26 doubled haploid lines among which two lines with doubled additional chromosomes 2D of Ae. tauschii can be used for further breeding to increase leaf rust resistance of cultivated triticale.

Azol Fungizid Resistenz in agrarischen Ökosystemen: Risikobewertung von Fungizid-Applikationsstrategien (AWECOS)

Die Daten der Vergleichsbetriebe stellen den Status quo im Pflanzenschutz in Deutschland dar. Der Behandlungsindex in Mohre und Frischkohl betrug im Durchschnitt der Jahre (2007-2014) bei 6,2 bzw. 9,2. Mohren wurden vorrangig mit Herbiziden und Frischkohl mit Insektiziden behandelt. Unter dem Aspekt des Pflanzenschutzes und zur Reduktion der Behandlungsintensitat mit Insektiziden kann Abbildung 1: Behandlungsindex der Herbizide (Herbizid-BI) in Abhangigkeit der Vorfrucht in den Vergleichsbetrieben fur Mohre, 2007-2014, in Deutschland, ungleiche Buchstaben symbolisieren signifikante Unterschiede (p<0,05), nBlattfrucht=58, nGemuse=28, nHalmfruchte=115 208 72. ALVA-Tagung, Seminarhotel Wesenufer, 2017 der Fruhkohlanbau empfohlen werden. Im Kohl- und Mohrenanbau konnen Halmvorfruchte Schadlingsbefall vorbeugen. Die Mittelaufwandmengen wurden vorrangig bei Herbizidanwendungen reduziert In beiden Kulturen wurden Abweichungen vom notwendigen Mas verzeichnet. Die Pflanzenschutzberatung sowie die regelmasige Durchfuhrung von Bonituren konnen Pflanzenschutzmittelanwendungen dem notwendigen Mas naher bringen und die Umsetzung des IPS in der Praxis verbessern. Fur die Ausschopfung nichtchemischer Masnahmen sind Praxisdemonstrationen und der intensive Austausch zwischen Wissenschaft und Praxis notig, genauso wie Forschungsforderung zur Entwicklung neuer und Optimierung bestehender integrierter Verfahren und neuer Wirkstoffe. Vorbildhaft wird der IPS von den Demonstrationsbetrieben u. a. in den Kulturen Mohre und Weiskohl, seit 2014 umgesetzt. Die Betriebe werden individuell und intensiv betraten, um Pflanzenschutzmittelanwendungen auf das notwendige Mas zu beschranken. Hierfur werden Bonituren unter Nutzung von Schadschwellen, Warndiensthinweisen und Monitoringverfahren durchgefuhrt.

Identification of candidate genes for the prehaustorial resistance of Triticum monococcum against Puccinia triticina

The project is dealing with the improvement of a universal pneumatic seeder which is suitable for sowing a wide range of different seeds in order to decrease particulate emissions of seed dressings. Aim of the project is to identify leakages in the pneumatic system in order to develop opportunities for the subsequent improvement of existing equipment to fulfill the high demands of user and environmental protection. Pneumatic seeders are characterized by a central, funnelshaped hopper. The batch feeder being an airlock and dosing feeder is located in the outlet of the seeder. The metered seed is fed into the air stream and transported to the sowing distributor by a conveying air stream.Due to climatic change, phenology traits are becoming increasingly important in grapevine breeding, since a premature flowering and ripening time could be observed for grapevine in the last decades. However, knowledge about these traits is still limited as they are genetically very complex and highly influenced by environmental factors. The analysis of the genetic basis of flowering time therefore will enable the development of tightly linked molecular markers useful for markerassisted selection of especially late flowering breeding lines.One recently occurred invasive insect pest has caught the attention for investigating biological control mechanisms and systems: The spotted wing drosophila (SWD, Drosophila suzukii Matsumura) is endemic in East China and Japan but has been introduced to the western hemisphere about 10 years ago and has been found in Europe since 2009. Nowadays, it has emerged to one of the most harmful pests to commercially grown fruit plants like stone fruits and nearly all kind of berries while it prefers ripe and overripe fruits. Our intention is to investigate the possible usage of natural antagonists for biological control. Therefore, we examine the natural load of parasites and pathogens (i.e. fungi, bacteria, viruses, microsporidia and protista) in fruit flies, isolate them and re-infect lab populations of D. suzukii for investigating the antagonistic potential. Furthermore, we will integrate the fruit pest codling moth (Cydia pomonella), which is an ongoing problem in apple orchards also because the pest develops resistance against commercially available insecticides. The long-term aim is to establish a stable system for pathogen detection that can be used for rapid identification of microbial antagonists in natural populations.Two main problems have to be considered in the development of control agents against the Fire Blight pathogen Erwinia amylovora: First, the ability of exponential growth leads to high cell densities in a short amount of time. Second, the most critical phase of Fire Blight infection occurs during blossoming, when the pathogen is transported to open flowers by various insects. To prevent infection, it is important to avoid invasion inside the plants tissue by interfering with growth of E. amylovora cells.The entomopathogenic fungus Isaria fumosorosea, formerly known as Paecilomyces fumosoroseus, has got a relatively wide host range. Within the scope of the EU project BIOCOMES investigations were done to validate the use of I. fumosorosea as a BCA against several pest insects. Under laboratory conditions it could be shown that I. fumosorosea seems to be a suitable BCA against Bemisia tabaci (silverleaf whitefly) and Spodoptera exigua (beet armyworm).Since the last two decades a number of fingerprinting methods have been developed to analyze microbial communities and their dynamics, including Terminal Restriction Fragment Length Polymorphism (T-RFLP), Length Heterogeneity-Polymerase Chain Reaction (LHPCR) and Automated Ribosomal Intergenic Spacer Analysis (ARISA). Because the latter provides a quick and cheap way together with high accuracy, we have chosen this method to investigate the fungal communities on grapevine, wood, leaves and berries.Saoussen Ben Tiba, Andreas Larem, Eva Fritsch, Karin Undorf-Spahn, Asma Laarif, Sami Fattouch, Johannes A. Jehle 1 Julius Kuhn-Institut, Institute for Biological Control, Darmstadt, Germany 2 Regional Center for Agriculture and Biological Agriculture, Chott Meriem, Tunisia 3 National Institute for applied technological Science Tunis (INSAT), Carthage University, Tunisia Email of corresponding author: saoussen.ben.tiba@jki.bund.deOne promising approach to a more environmentally friendly viticulture is growing fungus resistant cultivars with the novel cultivation method of the minimal pruning of trellis trained grapevines (MPTS). This practice reduces the tremendous amounts of fungicides needed to protect traditional cultivars and is expected to increase biodiversity compared to vineyards with traditional trellis trained grapevines (TS).Meiosis as the specialized cell division of sexual reproduction plays a crucial role in the exchange and reorganization of genetic material between two individuals by dividing the chromosome set in half and forming gametes. Even though in the last years major findings in the field of meiosis have been achieved, especially in plants, some key questions remain concealed. For a proper meiosis the initiation of double strand breaks (DSBs) during early prophase I is essential. Without DSBs no physical connection can occur between homologous chromosomes and recombination, pairing, and crossing over are excluded. So far in all analyzed eukaryotes SPO11, a meiosis specific transesterase, is the key enzyme inducing DSBs. But other than in animals and fungi where a single SPO11 is sufficient, plants need at least two different SPO11, referred to as SPO11-1 and SPO11-2, for proper meiosis. In Arabidopsis thaliana both have crucial functions and are essential in a functional form for the induction of meiotic DSBs as single knock out mutants are leading to near sterility by random chromosome distribution. Despite the same function of the homologs SPO111 and -2, the identity between both proteins is quite low. Homology of the orthologous SPO11 from different organisms is much higher. By exchanging SPO11-1 and -2 in Arabidopsis by their orthologs from various organisms we could demonstrate a species specific function of each SPO11, as a functional complementation of sterility could only be achieved with SPO11 from closely related species from the Brassicaceae. By exchanging non conserved regions between SPO11-1 and -2 of Arabidopsis we additionally could show a sequence specific function for each SPO11, as a functional rescue could not be achieved with all chosen regions. Interestingly, we could reveal a specific pattern of aberrant spliced isoforms for each SPO11 which are also sequence as well as species specific. By producing antibodies against AthSPO11-1 and -2 we were able to analyze for the first time the binding of SPO11-2 onto the DNA and perform co-immunolocalization studies with SPO11-1 and -2.European Stone Fruit Yellows (ESFY) is one of the most serious diseases in European fruit production. Infected Prunus cultivars yield poorly and lead to high economic losses. ESFY is caused by a specialized bacterium located in the phloem tissue of Prunus ssp., the Phytoplasma ‘Candidatus Phytoplasma prunorum’. It is spread by the phloemfeeding plum psyllid (Cacopsylla pruni) which acquires the bacterium by feeding on infected plants and is able to transmit it to healthy plants.Leaf rust caused by Puccinia triticina can cause yield losses up to 60 % and is the most common rust disease of wheat in the world. Vertical leaf rust resistance genes (Lr-genes) have been introduced in cultivars. Many of these resistances are broken down by virulent pathotypes. Horizontal resistances which are independent from races of a pathogen are known but show a quantitative characteristic which is carried by a few cultivars.

Impact of a vesicular arbuscular mycorrhiza symbiosis on biotic and abiotic stress tolerance of wheat

The main role of MADS-box transcription factors in plant developmental processes has been well described in the model plant Arabidopsis thaliana. However, little is known about their function in crops of important agricultural and commercial value. Our study aims to investigate their role in two agronomical relevant Rosaceae crops: apple (Malus x domestica Borkh.) and strawberry (Fragaria vesca). Expression studies using qPCR and RNA seq have identified two apple Dormancy Associated MADS-box (DAM) genes. They group with the StMADS11 clade, and were named MdDAM1 and MdDAM2, the last one discovered ex novo. Real time expression studies in dormant buds collected during the chilling period and chromatin immunoprecipitation (ChIP) analyses confirmed that the genes are downregulated by exposure to cold and MdDAM1 is epigenetically repressed, as it has been demonstrated for Arabidopsis FLC and peach DAM genes. In parallel we worked on strawberry MADS-box genes of known function involved in flower development. We chose three MADS-box genes that are homologs of Arabidopsis PISTILLATA and AGAMOUS to perform gene expression and functional analysis using a RNA interference approach to obtain post-transcriptional gene silencing. The positive transgenic lines of each transformation were evaluated at the molecular and phenotypic level. Single gene mutants does not show altered flower phenotype, suggesting a different mechanism of flower development in strawberry, probably due to the peculiar flower structure.The quarantine pest pine wood nematode (PWN) Bursaphelenchus xylophilus, native in North America, can be found in wood and root parts of infected conifer trees with focus on Pinus species. At the beginning of the 20 century this species was introduced into Japan supposedly with timber imports where it induced pine wilt expression on susceptible native pine species. Further outbreaks outside Europe are reported from China, Korea, Taiwan and Mexico. In 1999 PWN was found in Portugal, which today is declared as complete infested, since followed by first single tree outbreaks in Spain. The European Union (EU) member states have to undertake measures to prevent further spread of this nematode according to the emergency measures of the EU Commission.The box tree pyralid Cydalima perspectalis (Walker 1859) is an invasive alien moth from East Asia which occurs in Central Europe since 2007. It is an insect pest on plants of the genus Buxus, causing serious damage. Because of the rapid spread in Germany and nearby countries like Switzerland, one part of this study was testing different ways of eco-friendly regulation with commercially available beneficials and biological control agents.1 Julius Kühn-Institute, Federal Research Centre for Cultivated Plants (JKI), Institute for Epidemiology and Pathogen Diagnostics, Braunschweig, Germany. 2 Geisenheim Research Center, Section Phytomedicine, Geisenheim, Germany. 3 Área de Genética, Facultad de Ciencias, Universidad de Málaga, Instituto de Hortofruticultura Subtropical y Mediterránea La Mayora (IHSM-UMA-CSIC), Málaga (Spain). 4 Suez Canal University, Faculty of Agriculture, Ismailia, Egypt. Email of corresponding author: namis.eltlbany@jki.bund.deExcessive use of mineral nitrogen (N) fertilizer is a common practice in rice-wheat rotations in southeastern China. At the same time N use efficiencies (NUEs) in these rice-based cropping system are very low (Peng et al., 2006). The consequences are high N losses from arable land to water bodies (surfaceand groundwater) and to the atmosphere (Richter and Roelcke, 2000). To investigate the scope and scale of reductions in mineral N fertilizer inputs, demonstration field experiments on farmers’ field sites were conducted for three consecutive winter wheat-summer rice double crop rotations in two counties of Jiangsu Province from 2008 to 2011. Results from the first two years are presented here. NUEsPolycyclic aromatic hydrocarbons (PAHs) are important environmental pollutants which often persist in soil. The biochemistry of microbe-driven degradation of PAHs is well-studied though less is known about abiotic soil environmental factors influencing the microbe-phenanthrene interaction. Since minerals and charcoal are major soil components we matured different microbial communities in artificial soils based only on variation in the mineral content (montmorillonite, illite, ferrihydrite) and presence of charcoal for 2 years after addition of Luvisol microorganisms and sterile manure. Established microbial communities in soils were exposed to PAHs’ model compound phenanthrene (2 mg/g) to study microbial functionality depending on soil composition. Furthermore, litter which was previously shown to enhance microbial activity was added to the soils (1 wt%). Both treatments with corresponding controls were further incubated for 63 days with sampling on day 0, 7, 21 and 63. A natural Luvisol soil was treated similarly for control. Fingerprints by denaturing gradient gel electrophoresis (DGGE) based on 16S rRNA or ITS gene fragments, respectively, amplified from extracted total community DNA were generated. The presence of phenanthrene-degradative genes was screened by PCR-Southern Blot detection and soil treatments were subjected to a chemical phenanthrene analysis. DGGE fingerprints revealed that the addition of phenanthrene and litter caused a shift in microbial community composition. Differences in bacterial response to phenanthrene were seen depending on litter addition and soil composition. The incubation time was determined as additional influencing factor. Furthermore, a dominance of a specific genotype for phenanthrene degradation is assumed based on PCR-Southern Blot analysis.1 Julius Kühn-Institute Federal Research Centre for Cultivated Plants (JKI), Institute for Epidemiology and Pathogen Diagnostics, Messeweg 11-12, D-38104 Braunschweig 2 Institute of Crop Science and Resource Conservation, Soil Science and Ecology, University of Bonn, Nussallee 13, D-53115 Bonn 3 Institute of Bioand Geosciences 3, Agrosphere, Forschungszentrum Jülich GmbH, D-52425 Jülich Email of corresponding author: sven.jechalke@jki.bund.de