For propagation of certified horticultural but also agricultural plant material, a phytosanitary virus screening is obligatory in order to avoid the spread of plant viruses regionally but also globally. To date, most high throughput standard virus screenings are still based on the immunological detection method ELISA because of its robustness, high throughput potential and very good cost-benefit ratio. However, in some cases molecular biological applications are compulsorily. For instance in the process of seed potato certification, low virus titres present in seed potatoes require a long virus enrichment phase for adequate virus detection by DAS-ELISA. This time consuming step can be avoided by a change to more sensitive molecular biological techniques. Most nucleic acid based detection protocols are accompanied by high costs for their performance. We introduce a TaqMan (R) qPCR based one step protocol, named direct reverse transcriptase quantitative PCR (DiRT-qPCR), for the detection of RNA encoded viruses without sophisticated nucleic acid purification at lower costs but with high throughput potential exemplarily on (+)ssRNA Potato leafroll virus (PLRV).

Polyphenols like anthocyanins, flavan-3-ols and other flavonoids have proven to be good for human health. Therefore, there is an interest in polyphenols rich fruits like plums. But the European plum cultivars show a large variability in the characteristics of the fruit like form, size, skin colour, sugar content, taste, among others. For that reason it can be expected that the polyphenolic pattern is different between them. This study aims to identify the differences in the polyphenolic pattern of 19 European plum (Prunus domestica L.) cultivars with different fruit skin colours. For that purpose, fruits were harvested during 3 consecutive years. The juice was extracted with methanol using solid phase extraction. The extracts were measured with inverse phase HPLC to identify anthocyanins, flavan-3-ols, flavonols, flavones and hydroxycinnamic acids. After analysing the extracts, two groups were identified in relation to the total flavonoid content. One group of cultivars has a higher amount of total flavonoids than the other. This difference is principally influenced by the amount of hydroxycinnamic acids in the fruit juice, mainly neochlorogenic acid. Flavan-3-ol content is higher in those cultivars with higher flavonoid content. These differences were independent from the skin colour, therefore from the presence of anthocyanin. Flavonol and flavone content are relatively constant in all cultivars. This suggests that the final amount of polyphenols is early regulated in the biosynthetic pathway.

Phytoplasmas are cell-wall-less bacteria that cause diseases in approximately 1,000 plant species. ‘Candidatus Phytoplasma pyri’, the causal agent of pear decline, induces various symptoms on its hosts, leading to weakening and dieback of the plants, reduced fruit size and yield, and, consequently, considerable financial losses in all pear-growing areas. Fighting this disease requires a reliable and inexpensive method for pathogen detection in propagation material as well as plant stocks in orchards and breeding facilities. Here, we present a field-suitable detection protocol for ‘Ca. P. pyri’ based on loop-mediated isothermal amplification (LAMP) targeting the phytoplasmal 16S ribosomal DNA sequence. The combination of a simplified sample preparation method based on sodium hydroxide and colorimetric visualization of LAMP results enables a laboratory-independent pathogen detection. The detection limit is comparable with analysis by polymerase chain reaction; however, the pear decline LAMP detection method is superior in terms of ease of use, cost, and time effectiveness for obtaining results.

Prunus domestica (European plum) and Prunus armeniaca (apricot) are related species which can be hybridized. However, there are only very few reports on true interspecific hybrids between P. domestica and P. armeniaca. These hybrids must not be confused with the so called pluots or plumcots which are hybrids between P. salicina (Japanese plum) and P. armeniaca. In 2005, a program for breeding interspecific hybrids between P. domestica and P. armeniaca was initiated at Technische Universitat Munchen. As female parent, a P. domestica clone with hypersensitivity resistance to Plum pox virus (PPV) was used. As male parent, apricot cultivars with different degrees of susceptibility to PPV were chosen. Several interspecific hybrids were obtained. Leaves, shoots and fruits of the seedlings are intermediate between European plum and apricot. The seedlings were tested for their resistance to PPV. Interspecific hybrids with strong resistance to PPV could be obtained. To the best of our knowledge, this is the first report on the generation of PPV resistant hybrids between European plum and apricot. The hybrids could help to better understand the genes which are involved in hypersensitivity resistance to PPV. Moreover, the interspecific hybrid can be used for transferring the PPV resistance to the apricot genome by repeatedly backcrossing them with apricot. The interspecific hybrids represent a new kind of fruit species as the outer and inner fruit traits are intermediate between the parent species.

Plum maturity is usually assessed by destructive methods in the laboratory. The determination of harvest date and quality factors of European plum is poorly studied but needs to be as accurate as possible. Thus, for precision agriculture or continuous following of plum maturation, more rapid and non-destructive methods are needed. Therefore, in addition to measurements of fruit color, a new optical method was recently proposed. It is based on the estimation of the screening of chlorophyll fluorescence by flavonols and anthocyanins accumulating in the fruit skin. This allows to indirectly quantifying these phenolics in the skin of intact fruit. Here, we present the first results obtained with the commercial device Multiplex on two European plum Prunus domestica L. cultivars 'Haganta' and 'Hoh4517' during the 2010 season. A good correlation between fruit development and non-destructive detection of these pigments by this device was found. The relation between the Multiplex anthocyanin index and fruit development may give a useful tool for following plum fruit maturation and, thus, for a more rapid and precise determination of harvest date.

ETHYLENE action inhibition by 1-methylcyclopropene (1-MCP) extends postharvest life of many climacteric fruits. This work evaluates the effect of 1-MCP (0.625 ppm) treatment on the physiological behavior and quality during postharvest life of ‘Katinka’ and ‘Haganta’ European plum (

The behavior of 6- to 10-year-old trees of two hypersensitive plum cultivars ('Jojo', 'Hoh 4517') grafted onto a PPV sensitive rootstock under natural PPV inoculation conditions was observed. Some of the rootstocks got infected with PPV. In case of 'Jojo', no effect on the phenotype of the crown part could be observed. In case of 'Hoh 4517', single young shoots showed necrosis on the young shoots. These lesions facilitated secondary infections with Pseudomonas bacteria which then led to the death of whole branches. In order to avoid these damages it is highly recommended to use rootstocks with hypersensitivity resistance for all scion cultivars with hypersensitivity resistance to PPV.

A breeding program with the aim of obtaining Plum pox virus (PPV) resistant rootstocks for stone fruit cultivars was established at Technische Universitat Munchen. Genetic crosses between European plum (P. domestica) genotypes with hypersensitivity resistance to PPV were used as one parent and either P. spinosa or P. cerasifera as the other parent resulting in either `Dospina' or `Docera' rootstock candidates, respectively. The seedlings were screened for hypersensitivity resistance. The PPV resistant clones were screened for easy vegetative propagation by green cuttings, hardwood cuttings and in vitro propagation. The selected seedlings were retested extensively for PPV resistance with several European plum cultivars differing in their susceptibility to PPV. PPV infected budsticks of these cultivars were grafted onto the rootstock candidates either in spring or in summer. In many cases, the infected buds were rejected by the rootstock before bud break. Otherwise the young shoot which started to grow from the infected bud or budwood was rejected by the rootstock within several weeks after bud break. Due to this self-eliminating effect of PPV infected buds or scions grafted onto the newly developed PPV resistant rootstocks, no PPV infected tree developed to a marketable size. First results obtained from pomological tests indicate that trees grafted onto `Docera 6' show similar vegetative and generative properties as those grafted onto `St. Julien A' rootstock. National and international tests for the pomological evaluation of the rootstock are underway. In a first step, the new hypersensitive rootstocks will be used for newly bred hypersensitive plum cultivars. In this way, neither the rootstock nor the scion cultivar can get infected with PPV under natural inoculation conditions. This gives the highest level of security to damages caused by PPV which is available in stone fruits species at the moment. In a second step, their use for PPV sensitive European plum cultivars as well as for apricot, Japanese plum and peach is under evaluation.

The screening of genotypes derived from the Weihenstephan plum breeding program for hypersensitivity resistance against the Plum pox virus is based on the monitoring of the hypersensitive reaction. Biological screening is supported by the Blue LAMP protocol for the detection of PPV because of its straightforward procedure and reliability. The specificity of the LAMP technology also allows for PPV strain differentiation which is important to validate the stability of hypersensitivity against all known subgroups of PPV during the selection process of resistant genotypes. The seven primer sets designed for this purpose were specific to each one of the seven PPV strains. Unspecific amplification observed in initial experiments was inhibited by adjusting the trehalose concentration.

Fruit size, flesh firmness, color of the fruit skin and the flesh, adherence of the fruit flesh to the stone, caverns within the fruit flesh, content of sugars, organic acids and aroma components are important quality traits in European plum. In order to improve the fruit quality of European plum the main influencing factors on fruit quality have to be known. Fruits of different European plum cultivars and breeding clones originating from the Hohenheim and Weihenstephan plum breeding program were harvested and analyzed concerning 20 different fruit characters amongst them the soluble solid content, the organic acid content, fruit mass, fruit length and width. The influence of the locations, the harvesting time and the crop load on fruit quality was compared. Both picking time, crop load and the genetic determination were shown to be important influencing factors on fruit quality. In many cultivars, fruits harvested from PPV infected trees were of lower fruit quality than fruits grown on healthy trees. Thus, breeding can be regarded to hold the key position in enhancing fruit quality in Prunus domestica. However, crop load and harvesting time have to be optimized as well in order to exhaust the full genetic potential of a respective genotype.

So far, the polymerase chain reaction (PCR) is the most widely used method for the amplification of nucleic acids in vitro, especially for pathogen detection because of its high sensitivity. However, numerous isothermal amplification methods were developed in the recent years to avoid the need for a thermal cycler. Meanwhile the most applied approach is loop-mediated isothermal amplification (LAMP). The great advantage of LAMP is the enormous rate of amplification paired with a very high specificity. This study presents a simplified procedure for Plum pox virus (PPV) detection. A crude plant extract was applied to a modified one-step RT-LAMP protocol of Varga and James (2006). Gel electrophoresis was circumvented by a colour change of the reaction mix upon nucleic acid amplification. This procedure takes only two and a half hour from sampling to result and requires minimal technical equipment. The risk of cross contamination is minimized since amplification and visualization take place in a single tube. The Blue LAMP provides a fast and reliable detection of PPV both for single samples and for large scale surveys.

Until now there have been many investigations according the inheritance of the hypersensitivity resistance against the Plum pox virus (PPV) in European plum (Prunus domestica L.). For establishing an effective breeding program for PPV resistance it is important to know more about the inheritance and the genetic determination of the hypersensitivity resistance. There have been investigations concerning the inheritance of the hypersensitive resistance by screening seedlings of selected crossing combinations. These seedlings resulted from crossings between two hypersensitive parents, between hypersensitive and sensitive parents and between two sensitive parents. Moreover quantitatively resistant genotypes were crossed with hypersensitive resistant genotypes. One can show that the inheritance depends on the degree of hypersensitivity resistance in the parent genotypes. The higher the degree of hypersensitivity the higher is the amount of hypersensitive progenies. Furthermore it strongly depends on the selection of the crossing partners. In combinations with two hypersensitive partners, the percentage of hypersensitive seedlings is higher, than in combinations of hypersensitive genotypes with sensitive ones. The amount of hypersensitive progenies in crossing combination of hypersensitive genotypes with a quantitative resistant one is lower than in combination of hypersensitive genotypes with a sensitive one. In order to clarify the origin of the hypersensitivity resistance, progenies of crossing combinations from Ortenauer x Stanley, Ortenauer x Ortenauer and Stanley x Stanley have been tested. This refers to Jojo, the first completely hypersensitive cultivar, which is a descendant of a crossing between Ortenauer and Stanley. The results show that the donor of the hypersensitivity resistance is the cultivar Ortenauer.

The dihydrochalcone phloridzin (phloretin 2′-O-glucoside) is the most abundant phenolic compound in apple trees (Malus × domestica) and was also discussed to have an influence on the pathogen defence by shifting the dihydrochalcone profile from the glucosides to the more active aglycones. The final step in the biosynthesis of phloridzin is the glycosylation of phloretin at position 2′. Three cDNA clones from apple encoding glycosyltransferases are available which are able to catalyze the reaction in vitro. We investigated the possible role of glycosyltransferase UGT71A15 in phloridzin biosynthesis. The recombinant enzyme showed broad substrate acceptance but highest activities were observed with flavonols. Specific activities and the kinetic data indicated that phloretin is not the preferred native substrate of the UGT71A15. However, an increase of the molar ratio phloridzin:phloretin was found in transgenic lines, indicating a physiological relevance of UGT71A15 in planta, although a decrease of the total amount of dihydrochalcones in the majority of the samples was found. Unexpectedly, the increase of the phloridzin:phloretin ratio was not reflected by an increase of the total glucosyltransferase activities. In contrast, the majority of transgenic plants showed a reduced glucosylating activity with both phloretin and quercetin as a substrate, but the observed activity changes in a given sample were not similar for the two substrates. An increased susceptibility of M. robusta against the fire blight causing bacterium E. amylovora as a result of UGT71A15 overexpression could not be observed. Overexpression of UGT71A15 in transgenic apple trees also did not lead to morphological changes.