Quantitative detection of _Potato virus Y_ in potato plants and aphids - Discussion of diverse applications in potato research

Every year potato growers worldwide complain about severe yield losses caused by _Potato virus Y_ (PVY). Therefore, PVY along with _Potato leafroll virus_ belongs to the most important potato viruses. There are three main strains of PVY: PVY^O^, PVY^N^ and PVY^C^. However, also recombinant forms exist such as PVY^N^Wilga and PVY^NTN^, both of which increase in importance due to their potential to displace the non-recombinant strains at a high percentage. They appear also in mixed infections. In recent years PCR and qPCR assays were developed to differentiate PVY isolates. In order to identify PVY isolates by PCR often large amplicons have to be generated which requires the input of expensive enzymes. On the other hand, qPCR assays until now do not allow the differentiation between PVY^N^Wilga and PVY^NTN^. 

For the discrimination between PVY^O^/PVY^N^Wilga and PVY^N^/PVY^NTN^ a qPCR assay was developed, which allows the differentiation and highly efficient quantification of both strains and recombinants, respectively. For this purpose dual-labeled hydrolysis probes tagged with different fluorophores were designed. The assay is suitable for many different applications, for example safety research on genetically modified (GM) potato plants. The goal of this research is to determine whether genetic modification causes changes in resistance to viruses. Two different GM cultivars were examined for signs of altered resistance to an infection with PVY in comparison to their near-isogenic lines and three reference cultivars. Reference cultivars are included to determine the baselines for resistance and thus to be able to decide if the changes could represent a biological risk. The plants to be investigated were mechanically inoculated with PVY^N^Wilga or PVY^NTN^ and analyzed by means of the developed assay after two weeks. The results of the experiment indicate that the differences in virus titer between the reference cultivars are higher than between the GM potatoes and their isogenic lines. Therefore, in our experiments the GM potato plants showed no alteration in PVY resistance to neither one of the tested strains.

Since _Myzus persicae_ is one of the most important vectors transmitting PVY, the developed assay will also be applied to the quantification of PVY particles in aphids. The displacement of PVY^O^ and PVY^N^ by PVY^N^Wilga and PVY^NTN^ may be due to a difference in efficiency of transmission by _M. persicae_. Therefore, the objective is to test whether more virus particles of the recombinant forms in comparison to the non-recombinant strains PVY^O^ and PVY^N^ bind in the stylets of _M. persicae_. 

A third possible application of the developed assay may be of interest in potato breeding. The exact quantification of PVY particles in plants allows the classification of resistance in potato plants. It is possible to estimate whether a resistance is extreme or not. Extreme resistance is characterized by the absence or presence of very low amounts of virus particles in plants several days after inoculation. When testing the plants for PVY infection by ELISA, often unspecific reactions occur which makes it difficult to differentiate between plants weakly infected and plants very weakly infected. An exact quantification of the PVY titer gives more certainty for the determination of the resistance type.

In conclusion, the developed assay is an efficient and low-cost method that allows the differentiation and quantification of PVY^O^/PVY^N^Wilga on the one hand and PVY^N^/PVY^NTN^ on the other hand with high throughput. The method can be utilized for a wide range of applications in potato research.
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Effect of N-fertilization, fungicide treatment, seed density and abiotic stress factors on the total ß-glucan content of six-rowed winter barley (Hordeum vulgare L.)

In Germany, the first six-rowed waxy winter barley cultivar (cv.) `Waxyma` was registered in 2008. Besides changes in starch composition, waxy barley is rich in ß-glucans offering new applications in the food industry as ß-glucans reduce the blood cholesterol level. To investigate the stability of the ß-glucan content, cv. `Waxyma`, three waxy breeding lines and the non-waxy cv. `Lomerit` were grown in field trials, shelter-, green house- and growth chamber experiments. Besides this, `Waxyma` was grown in field trials at varying nitrogen levels and different seed rates, both with and without fungicide treatment. Waxyma showed a significant increase of the ß-glucan content under optimal nitrogen fertilization and fungicide treatment, but the influence of the seed density was not significant. Under shelter and greenhouse conditions the influence of drought stress during grain filling was analyzed. The increase of the ß-glucan content under drought stress in the shelter and under rising temperatures in growth chambers was only significant for the non waxy cv. `Lomerit` while no influence of drought stress was  observed under green house conditions.  In summary the high ß-glucan content of cv. `Waxyma` seems to be relatively stable with respect to growing conditions making it a suitable raw material for human nutrition

Breeding for resistance to insect-transmitted viruses in barley – an emerging challenge due to global warming

Vor dem Hintergrund längerer Perioden mit höheren Temperaturen im Herbst und im Winter ist künftig von einer zunehmenden Bedeutung insektenübertragener Viren auszugehen, wie es die vorliegenden Untersuchungen zum Auftreten von Barley yellow dwarf virus und Wheat dwarf virus in Mitteldeutschland belegen. Dabei zeigte sich u. a. eine deutliche Beziehung zwischen der Anzahl Infektionstage im Herbst und dem BYDV-Auftreten in der Wintergerste im darauffolgenden Frühjahr. In Klimakammeruntersuchungen zum Einfluss der Temperatur auf die BYDV-Übertragung wurden 10°C als Temperaturgrenze ermittelt, bis zu der eine Virusübertragung durch Rhopalosiphum padi erfolgt. Die ersten Ergebnisse zur Pyramidisierung von Ryd2, Ryd3 und eines QTL aus der Sorte ‘Post’, der auf Chromosom 2H lokalisiert ist, mittels molekularer Marker und DH-Linien zeigen, dass Linien mit 3 bzw. 2 Resistenz-/Toleranzgenen eine deutliche geringere Symptomausprägung und einen verringerten Virustiter aufweisen. Dabei hat der QTL auf Chromosom 2H im Vergleich zu Ryd3 und Ryd2 einen deutlich geringeren Effekt. Die genetische Basis der WDV-Toleranz ist sehr eng. Lediglich die Sorte ‘Post’ wies in den Infektionsversuchen im Freiland ein ausreichendes Toleranzniveau auf. Untersuchungen an Populationen doppelhaploider Linien lassen auf eine polygene Vererbung dieser Toleranz schließen.Due to global warming longer periods of higher temperature in autumn and winter are expected which may result in an increasing importance of insect-transmitted viruses. Investigations carried out in Saxony-Anhalt from 1998 to 2008 on the incidence of Barley yellow dwarf virus (BYDV) and Wheat dwarf virus (WDV) revealed a clear relation between the number of infection days in autumn and the BYDV-attack in winter barley fields in the following spring. In additional experiments carried out in growth chambers under controlled conditions it turned out that 10°C is the minimum temperature for an efficient transmission of BYDV by Rhopalosiphum padi. In order to enhance the level of resistance to BYDV, Ryd2, Ryd3 and a QTL derived from the cultivar ‘Post’ located on chromosome 2HL were combined using DH-lines and molecular markers. Concerning symptom expression and virus extinction first results indicate a reduction in those lines combining especially Ryd2 and Ryd3. Concerning WDV extensive screening programmes were conducted, but tolerance was only detected in cv. ‘Post’. First results of genetic analysis using DH-lines give hint that this tolerance is inherited in a quantitative manner

The control of aphids as insect pests - From chemical crop protection to plant resistance

Blattläuse sind aufgrund ihres massenhaften Auftretens, verbunden mit direkten Saugschäden und ihrer Fähigkeit, Viren zu übertragen, wichtige Schadinsekten. Entgegen dem belegten Rückgang der Insektenvielfalt in der Agrarlandschaft, sind Blattläuse von diesem Trend offenbar weniger betroffen und zeigen in Saugfallen konstante oder sogar steigende Zahlen. Lange Zeit konnten Blattläuse mit hoher Effizienz durch den Einsatz von chemischen Insektiziden bekämpft werden. Verschiedene Gründe (gesellschaftspolitische Akzeptanz sowie die Ausbreitung von Insektizidresistenzen) führen jedoch zu einer Einschränkung des Einsatzes chemischer Insektizide und damit zu der Notwendigkeit, Werkzeuge für alternative Strategien zu entwickeln. Das Konzept des Integrierten Pflanzenschutzes beinhaltet verschiedene alternative Ansätze zur Schädlingsbekämpfung, z. B. Pflanzen­resistenzen, für die es bereits mehrere Anwendungs­beispiele im Feld gibt. Darüber hinaus werden die Möglichkeiten des Blattlausmonitorings sowie Beispiele für den Einsatz neuer Züchtungsmethoden (Genome Editing) und neuer dsRNA-basierter Insektizide diskutiert.Aphids are important insect pests due to their mass occurrence combined with direct feeding damage and their abil­ity to transmit viruses. Contrary to the documented decline in insect diversity, aphids are apparently less affected by this trend, showing constant or even increasing numbers. For a long time, aphids could be controlled with a high efficiency by the use of chemical insecticides. However, various reasons (socio-political acceptance as well as spread of resistance) lead to a restriction of the use of conventional chemical insecticides and thus to the need to develop tools for alternative strategies. The concept of Integrated Pest Management provides various alternative approaches for pest control, e.g., plant resistance and several examples for prototypic field applications exist. In addition, the possibilities of aphid monitoring as well as examples of the use of new breeding methods (genome editing) and new dsRNA-based insecticides are discussed

Effect of the Soil pH on the Alkaloid Content of Lupinus angustifolius

Field studies were conducted in growing seasons 2004, 2005, and 2010 to investigate the effect of different soil pH values on the alkaloid content in seeds of Lupinus angustifolius. Two-year experiments with eleven cultivars were carried out in acid soils with an average of pH=5.8 (Mecklenburg-Western Pomerania) and on calcareous soils with an average pH of 7.1 (Bavaria), respectively. In addition, in 2010, eight cultivars were grown in field experiments in soils with pH values varying between pH=5.3 and pH=6.7. In all experiments conducted on soils with a higher pH (pH=6.7 and pH=7.1), a significantly lower alkaloid content was detected in all Lupinus angustifolius cultivars than on soils with a lower pH (pH=5.3 and pH=5.8). Results clearly show that the alkaloid content is significantly influenced by the soil pH but genotypic differences regarding the reaction to different pH values in the soil were observed

Effects of growing system and season on the alkaloid content and yield of different sweet <i>L. angustifolius</i> genotypes

Nine varieties and two breeding lines of sweet Lupinus angustifolius were cultivated under organic and conventional conditions in Northern Germany in growing seasons 2010, 2011 and 2012. The alkaloid content was significantly influenced by the growing system and year and also by genotype. The variety `Vitabor` and the breeding line Bo 083521AR revealed a  very low alkaloid content in all years and cropping systems, while  `Sanabor`, `Borlu` and `Boregine` had a higher content. In the years 2010-2012 significantly lower alkaloid contents (475 µg/g) were found under organic conditions than under conventional conditions (615 µg/g). The mean alkaloid level of all varieties and breeding lines in organic farming was highest in 2010 (640 µg/g) and lowest in 2012 (364 µg/g), depending on temperature during the seed filling period. Investigations on the yield of L. angustifolius revealed significant effects of the genotype, the year and the growing system