C. Büttner

Host Species-Dependent Population Structure of a Pollen-Borne Plant Virus, Cherry Leaf Roll Virus

ABSTRACT Cherry leaf roll virus (CLRV) belongs to the Nepovirus genus within the family Comoviridae. It has a host range which includes a number of wild tree and shrub species. The serological and molecular diversity of CLRV was assessed using a collection of isolates and samples recovered from woody and herbaceous host plants from different geographical origins. Molecular diversity was assessed by sequencing a short (375-bp) region of the 3′ noncoding region (NCR) of the genomic RNAs while serological diversity was assessed using a panel of seven monoclonal antibodies raised initially against a walnut isolate of CLRV. The genomic region analyzed was shown to exhibit a significant degree of molecular variability with an average pairwise divergence of 8.5% (nucleotide identity). Similarly, serological variability proved to be high, with no single monoclonal antibody being able to recognize all isolates analyzed. Serological and molecular phylogenetic reconstructions showed a strong correlation. Remarkably, the diversity of CLRV populations is to a large extent defined by the host plant from which the viral samples are originally obtained. There are relatively few reports of plant viruses for which the genetic diversity is structured by the host plant. In the case of CLRV, we hypothesize that this situation may reflect the exclusive mode of transmission in natural plant populations by pollen and by seeds. These modes of transmission are likely to impose barriers to host change by the virus, leading to rapid biological and genetic separation of CLRV variants coevolving with different plant host species.

Double-stranded RNA pattern and partial sequence data indicate plant virus infection associated with the ringspot disease of European mountain ash (Sorbus aucuparia L.)

Summary.Double-stranded RNA (dsRNA) has been extracted from tissue of European mountain ash trees (Sorbus aucuparia L.) showing typical ringspot and mottling symptoms on leaves and a gradual decay in general. A characteristic dsRNA pattern was found in leaf samples of symptomatic mountain ash trees from various stands in Germany. Bands of dsRNA molecules of approximately 7 kb, 2.3 kb, 1.5 kb, and 1.3 kb, respectively, were repeatedly detected. By random primed reverse transcription cDNA was synthesised from dsRNA and amplified by degenerate oligonucleotide primed PCR. After TA cloning, the cDNA clones obtained were screened with an enhanced-chemiluminescence-labelled dsRNA probe. Positive clones were further analysed by using them as hybridisation probes in Northern blots of total plant RNA and in Southern hybridisation with genomic DNA from Sorbus aucuparia leaves. From cDNA clones that were found to be specific for dsRNA in Northern analysis, primers were deduced for 5′-RACE analyses and further cloning. Finally, a cDNA fragment of 3,737 bp was obtained, which showed homology to viral proteins, particularly to the RNA-dependent RNA polymerase of members of the family Bunyaviridae, but without high similarity to a known genus. The dsRNA pattern and the sequence information strongly indicate a virus associated with the mountain ash ringspot disease. The putative virus remains still unidentified.

Aphidius colemani Vier. (Hymenoptera, Braconidae, Aphidiinae) detected in cereal fields in Germany

Studies conducted in the 2000 cropping season at two different localities, Flaeming and Magdeburger Boerde in Germany, have provided new information on cereal aphid (Sitobion avenae (F.), Metopolophium dirhodum (Walker), and Rhopalosiphum padi (L.)) parasitoids in winter wheat; their species composition, relative abundance, hosts, and location effects. The average aphid population density was higher at Magdeburger Boerde and lower at Flaeming. Among the aphid species, Sitobion avenae was more abundant at Flaeming and Metopolophium dirhodum at Magdeburger Boerde. In total, eight species of primary parasitoids were recorded: Aphidius colemani Viereck, Aphidius rhopalosiphi DeStefani Perez, Aphidius uzbekistanicus Luzhetzki, Aphidius ervi Haliday, Aphidius picipes (Nees), Ephedrus plagiator (Nees), Praon gallicumStarý, and Praon volucre (Haliday). The predominant parasitoid species were Aphidius colemani and Aphidius rhopalosiphi on Metopolophium dirhodum and Aphidius uzbekistanicus on Sitobion avenae. A low number of hyperparasitoids were also recorded. Aphidius colemani was recorded for the first time in the open winter wheat fields in Germany, although it has been used as a biocontrol agent in glasshouses in many European countries and overseas. An analysis of the aforementioned information shows that Aphidius colemani detected as a parasitoid of cereal aphids in Germany is likely a result of an accidental escape of parasitoids from a glasshouse, as well as their successful overwintering and establishment in the area. This study provides baseline information essential for assessing future changes in aphid parasitoid species guild and dynamics in cereal fields in Germany.

Evaluation of a method for quantification of Pythium aphanidermatum in cucumber roots at different temperatures and inoculum densities

A method for the quantification of Pythium aphanidermatum density in cucumber roots based on an indirect enzyme-linked immunosorbent assay (Elisa) was established and tested. This approach was applied in three experiments under various environmental conditions. In addition, different inoculum densities were applied to vary the disease severity but also, to search for suitable inoculum densities in long-term epidemiological studies. Cucumber plants were grown in containers with aerated nutrient solution in a growth chamber at air temperatures of 15, 20, 25 and 30°C, and were inoculated with 0, 10, 103 and 105 oospores of P. aphanidermatum per litre nutrient solution. The pathogen density in the roots increased with inoculum density and temperature and resulted in growth reduction of the cucumber plants. Clearly, low temperatures delayed the development of the disease in the plant, while high temperatures combined with high inoculum densities led to sudden death of some plants. Therefore, inoculum densities not higher than 103 oospores per litre should be applied in long-term experiments. Independent of inoculum density and temperature, correlations were established between the mycelium density in the roots and the crop biomass, indicating that the indirect Elisa produces robust estimates of P. aphanidermatum density in cucumber roots under various conditions.ZusammenfassungEine Methode zur Bestimmung der Dichte von Pythium Aphanidermatum in Gurkenwurzeln wurde durch Anwendung eines indirekten enzyme-linked immunosorbent assay (ELISA) entwickelt und in drei Experimenten mit unterschiedlichen klimatischen Bedingungen getestet. Dabei wurde auch die Inokulumdichte variiert, um weitere Abstufungen der Erkrankung zu erhalten, aber auch, um geeignete Inokulumdichten für Langzeitstudien zur Epidemiologie herauszufinden. Gurkenpflanzen wurden in Gefäýen mit belüfteter Nährlösung in Klimakammern bei Lufttemperaturen von 15, 20, 25 und 30°C kultiviert und mit 0, 10, 103 and 105 Oosporen von P. aphanidermatum pro Liter Nährlösung inokuliert. Die Pathogendichte in der Wurzel stieg mit der Inokulumdichte und der Temperatur, wodurch sich das Wachstum der Pflanzen verringerte. Bei 20°C war die Krankheitsentwicklung deutlich verzögert, während 25 und 30°C in Kombination mit hoher Inokulumdichte zum vorzeitigen Absterben einiger Pflanzen führten. Deshalb sollten Inokulumdichten von mehr als 103 Oosporen je Liter in Langzeitexperimenten nicht verwendet werden. Über alle Inokulumdichten und Temperaturen wurden Korrelationen zwischen der Myzeldichte in der Wurzel und der Pflanzenbiomasse gefunden. Diese Korrelationen zeigen, dass der indirekte ELISA eine robuste Schätzung der Dichte von P. aphanidermatum in Gurkenwurzeln unter verschieden Bedingungen liefert.

Occurrence of fumonisins in asparagus (asparagus officinalis L.) and garlic (allium sativum L.) from Germany.

Fusarium proliferatum is able to produce fumonisins and is considered a pathogen of many economically important plants (e.g. corn, rice, asparagus) [1]. The occurrence of fumonisin FB1 inF. proliferatum infected asparagus spears from Germany was investigated using a liquid chromatography/electrospray ionization-mass spectrometry (LC-ESI-MS) method with isotopically labeled fumonisin FB1-d6 as internal standard. Asparagus samples were harvested in July 2000 and screened forFusarium species. AltogetherF. oxysporum, F. proliferatum and F. sambucinum were isolated from the spears. The samples infected with F.proliferatum were subsequently analyzed for fumonisins. FB1 was detected in 9 of the 10 samples in amounts ranging from 36.4 ng/g to 4513.7 ng/g (based on dry weight). Fumonisins FB2 and FB3 were found in six samples in lower concentrations. In asparagus spears of June 2002 we could findF. proliferatum in 6% of the samples, however no fumonisins were detectable.Furthermore the capability of producing FB1 by the fungus in garlic bulbs was investigated. Therefore garlic was cultured inF. proliferatum contaminated soil and the bulbs were screened for infection with F.proliferatum and for the occurrence of fumonisins by LC-MS. F.proliferatum was detectable in the garlic tissue and all samples contained FB1 (26.0 ng/g to 94.6 ng/g).This is the first report of the natural occurrence of FB1 in German asparagus spears and furthermore our findings suggest a potential for natural contamination of garlic bulbs with fumonisins. For detailed results and methods see Ref. [2].

Spargelstangenuntersuchungen zur Haupterntezeit auf Infektionen mitFusarium spp. und Kontaminationen mit Fumonisin B1

Asparagus spears collected from a total of six commercial plantings in Austria during the main harvest periods in May and June of 2003 and 2004 were examined for endophytic colonization byFusarium spp., particularlyF. proliferatum. Potentially toxigenic fungi such asF. proliferatum were isolated and identified by morphological characteristics using light microscopy. Fumonisin B1 inF. proliferatum-infected asparagus spears was detected with IAS-HPLC-FLD or HPLC-MS/MS. The identity of endophytic fungi colonizing of a total of 816 individual spears was determined. The incidence of infection byF. proliferatum and otherFusarium spp. was highly dependent on location and sampling date. The dominantFusarium species among the endophytic microflora wasF. oxysporum. Other frequently isolated species includedF. proliferatum, F. sambucinum, F. culmorum, F. avenaceum andF. equiseti. The incidence ofF. proliferatum-infected asparagus spears was less than 10% at four of the six sampling locations. At the two remaining locations, 20–47% of the spears examined were infected withF. proliferatum. Further exploration of FB1 generation in asparagus is required because the low levels of FB1 (10–50 (μg/kg) detected in harvested spears in 2003 and 2004 cannot be explained by the results of this study.

Cell-to-cell movement of plant viruses through plasmodesmata: a review.

Plasmodesmata are cytoplasmic bridges in plants through which intercellular communication occurs. This involves the transport of ions, photoassimilates, growth hormones as well as protein‐nucleic acid complexes. Although these molecules are rather small (< 1 kDa) plant viruses succeed in using these intercellular highways to transport their genome. These viruses alter the plasmodesmata in some way to allow the transport of such large molecules. This review deals with how plant viruses manage this with the help of movement proteins and the cytoskeleton.

Untersuchungen zur pathogenität und mykotoxinbildung vonFusarium sambucinum, dem erreger der trockenfäule an kartoffeln

AbstractInvestigation into virulence and mycotoxin formation of the dry rot causing pathogen Fusarium sambucinum on potatos 11 strains ofFusarium sambucinum were isolated from tubers with dry rot symptoms from three different depots in the Land Brandenburg and Saxony-Anhalt. All isolates produced diacetoxyscripenol in artificially infected potato tubers. Additionally, two isolates produced T-2 and HT-2 toxins as well. The virulence and mycotoxin formation of the isolates was dependent on the potato varieties ‘Sieglinde’ and ‘Berber’ used in the experiment. The amount of diacetoxyscripenol in diseased tissue was positively correlated with the virulence of theF. sambucinum isolate and the susceptibility of the potato variety as well.ZusammenfassungMehr als 11Fusarium sambucinum-Isolate wurden aus trockenfaulen Kartoffelknollen isoliert, die zum Zeitpunkt der Probenahme im Januar und Februar 2004 aus verschieden Lagern in Brandenburg und Sachen-Anhalt stammten. Alle Isolate produzierten im faulen Gewebe Diacetoxyscripenol. Zwei Isolate produzierten zusatzlich T2- und HT-2-Toxin. Es traten zwischen den beiden verwendeten Kartoffel-Sorten ‘Sieglinde’ und ‘Berber’ Unterschiede hinsichtlich der Virulenz und der Mykotoxinproduktion auf. Es konnte eine enge Korrelation zwischen der Fäulnisausprägung und der Diacetoxyscripenol-Konzentration nachgewiesen werden.

Virus-diseased Ulmus laevis in Eastern Germany

En olmos situados en un parque cercano a Postdam, se ha observado la presencia de sintomas foliares similares a los producidos por virus. Mediante bioensayos y pruebas serologicas se descarto la presencia de infecciones originados por el virus del enrollamiento de la hoja del cerezo (CLRV), el virus del moteado del olmo (EMV), el virus del mosaico de Arabis (ArMV) y el virus del anillamiento del tabaco (TRSV), todos ellos bien conocidos por afectar a los olmos. Repetidamente se aislo, en olmos enfermos, particulas flexibles de aproximadamente 750 mm de longitud similares a las de Potyvirus y Carlavirus. Las particulas fueron transmisibles a diversas especies de Chenopodium, un indicador herbaceo. Segun una prueba ELISA y un ensayo RT-PCR en que se usaron, respectivamente, un anticuerpo policlonal especifico de genero de Potyvirus de amplio espectro, y cebadores especificos de la familia, el virus no es miembro de la familia Potyviridae. Tampoco se ha encontrado, en estudios mediante microscopia electronica, inclusiones del tipo potyvirus en las celulas de las hojas de plantas indicadoras infectadas. En la actualidad se estan realizando nuevas caracterizaciones moleculares de estos aislamientos viricos.