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The abiotic conditions required for eradication of Polymyxa betae, the vector of Beet necrotic yellow vein virus in sugar beet, were investigated. Survival of resting spores of P. betae was determined under aerobic (30 min, 4 days and 21 days) and anaerobic (4 days) conditions under several temperature regimes in a water suspension and in leachate extracted from an aerobic compost heap. In water under aerobic conditions the lethal temperature was 60, 55 and 40°C for exposure times of 30 min, 4 days and 21 days, respectively. The effect of compost leachate and/or anaerobic conditions on survival of P. betae depended on temperature. After incubation for 4 days at 20°C, no significant effects of anaerobic conditions or leachate on the survival of P. betae were found. However, at 40°C for 4 days under anaerobic conditions, survival of P. betae was significantly lower than survival under aerobic conditions in water as well as in leachate. In leachate taken from an aerobic compost heap, aerobically incubated at 40°C for 4 days, survival of P. betae was significantly lower than survival in water at the same temperature. As anaerobic spots are prevalent in aerobic compost heaps, especially during the thermophilic phase, actual inactivation temperatures under composting conditions are likely to be lower than the temperatures we found for eradication in water under aerobic conditions.

Production of sugar beet, the most important source of sugar in Syria, has suffered from many problems in the past, especially from diseases. No previous surveys have been made in Syria for viral diseases and soil-borne fungi of sugar beet. In 1998, samples were collected from plants showing symptoms of virus infection (yellowing, wilting, necrosis and mosaic). Root samples (341) were collected from crops of autumn-sown seed from 115 localities in seven provinces, 173 root samples from spring-sown crops and 39 leaf samples were collected during both seasons. The root samples were tested for the presence of viruses by double antibody sandwich-enzyme-linked immunosorbent assay (ELISA) and triple antibody sandwich-ELISA, and for soil-borne fungi by red plate (Rose Bengal) dishes. We have shown for the first time the presence of Beet necrotic yellow vein virus, Beet soil-borne virus, Beet yellows virus and Beet mild yellowing virus in Syrian sugar beet fields in which Rhizoctonia sp. and Fusarium sp. were also widely distributed.

Polymyxa betae isolates were obtained by means of bait plants from a large number of soil samples collected in eastern Germany. Additional P. betae isolates were received from several institutions in western Germany and abroad. Isolates were grown on sugarbeet seedlings and tested for the presence of beet necrotic yellow vein virus (BNYVV) and beet soilborne virus (BSBV). BNYVV was only present in isolates from western Germany and abroad but absent in all isolates from eastern Germany., In contrast, BSBV was detected in more uniform geographic distribution in 14 out of 33 P. betae isolates tested. The virulence of P. betae isolates was estimated on the basis of the extent of resting spore formation in the root system of sugarbeet seedlings. Differences in virulence were found among virus-free as well as virus-carrying P. betae isolates. The mean value of virulence ratings was distinctly lower with BNYVV-carrying isolates and slightly lower with BSBV-carrying isolates as compared to virus-free isolates. Zusammenfassung Virusgehalt und Virulenz von Polymyxa betae Keskin-Isolaten aus verschiedenen Regionen Europas Polymyxa betae-Isolate wurden mit Hilfe von Fangpflanzen aus einer grosen Anzahl Bodenproben gewonnen, die in Ostdeutschland gesammelt worden waren. Weitere P. betae-Isolate wurden von mehreren Institutionen in Westdeutschland und dem Ausland erhalten. Die Isolate wurden an Zukerrubensamlingen kultiviert und auf ihren Gehalt an beet necrotic yellow vein virus (BNYW) und beet soilborne virus (BSBV) getestet, BNYVV war nur in Isolaten aus Westdeutschland und dem Ausland enthalten und fehlte in allen ostdeutschen Isolaten. Im Gegensatz dazu wurde BSBV geographisch gleichmasiger verteilt in 14 von 33 getesteten Isolaten nachgewiesen. Die Virulenz der P. betae-lsolate wurde anhand der Starke des Dauersporenbefalls im Wurzelsystem eingeschatzt. Virulenzunterschiede wurden sowohl innerhalb der virusfreien als auch innerhalb der virustragenden P. betae-lsolate gefunden. Der Mittelwert der Virulenz war bei den BNYVV-tragenden Isolaten deutlich geringer und bei den BSBV-tragenden Isolaten etwas geringer als bei den virusfreien Isoiaten.

The development of Polymyxa betae within the roots of a sugar beet variety susceptible to Rhizomania was observed in hydroponic culture over a period of 10 days. Light microscope studies showed that at an average temperature of 20 °C the life cycle of the fungus, containing beet necrotic yellow vein virus (BNYW), was completed within 10 days. A change from the multiplication phase to the survival phase of P. betae became evident. At the beginning of the life cycle the fungus produced mainly zoospores whereas at a later stage plasmodia developed into resting spores. Zoospore density in the nutrient solution reached a maximum between the 5th and 7th day after inoculation and then declined to the initial concentration. The number of zoospores attached to the root surface increased progressively at 48 h intervals, correlated with a parallel increase in BNYW-content of the roots. Light- and fluorescence microscopy revealed that zoospores of P. betae often attach near the point of release and do not move very long distances. In addition it became evident that zoospores may attach to the thallus wall inside the zoosporangium that they have developed in.

BNYVV concentration in the tap roots of sugar beet varieties grown in rhizomania-infested fields During plant development, the BNYVV concentration in several commercially available rhizomania-tolerant sugar beet varieties and one susceptible variety was examined as an index of the intensity of infection. The root weight, sugar content and sugar yield of the same varieties in fields naturally infested with rhizomania were also measured. Significant negative correlations were found between the average virus concentration in the tap root and yield parameters in infested fields. These were largely independent of the growth stage of beet plants used for virological investigations. However, the negative correlations between virus concentration and yield were not significant if rhizomania-tolerant varieties only were compared. The possibility that virus concentration might be used as a criterion for selection in addition to yield performance is discussed. This may lead to selection that is targeted more directly at rhizomania resistance and thereby accelerate breeding work. Zusammenfassung An mehreren, bereits kommerziell verfugbaren rizomaniatoleranten Zuckerriibensorten und einer anfalligen Vergleichssorte wurde der BNYW-Gehalt wahrend der Entwicklung der Pflanzen als Maβ fur die Befallsstarke der Pflanzen untersucht. Gegenubergestellt wurde die Leistung (Ertrag, Zuckergehalt, bereinigter Zuckerertrag) dieser Sorten beim Anbau im Feld unter naturlichen Befalls-bedingungen. Zwischen dem durchschnittlichen Virusgehalt in der Hauptwurzel (Rube) und den pflanzen-baulich relevanten Leistungsmerkmalen unter Befall bestand eine enge negative Korrelation, und zwar offensichtlich weitgehend unabhangig vom Entwicklungsstadium der fur die virologische Untersu-chung verwendeten Rubenpflanzen. Wurden nur die Sorten mit Rizomaniatoleranz in die statistischen Betrachtungen einbezogen, war die negative Beziehung zwischen Virusgehalt und Leistungsmerk-malen jedoch nicht signifikant. Es wird diskutiert, inwieweit die Berucksichtigung des Virusgehaltes als zusatzliches Selek-tionskriterium (neben Leistungsmerkmalen) zu einer gezielteren Selektion in Richtung Rizomaniaresi-stenz und damit auch zu einer Beschleunigung der Zuchtungsarbeit fuhren kann.

The coat proteins of tymo-, tombus-, como-, nepo-, tobamo-, potex-, carla- and potyviruses were subjected to sodium dodecylsulphate (SDS)-polyacrylamide gel electrophoresis, electro-blotted onto nitrocellular membranes and reacted with homologous and heterologous antisera to intact plant viruses. Immune complexes were detected after reaction with alkaline phosphatase-labelled goat anti-rabbit antibodies. SDS coat proteins of comoviruses reacted only with antisera which were homologous for their intact particles. In all other groups, however, a rather broad range of serological relationships was detected. With a number of viruses, serological relations to other members of the same group were demonstrated for the first time. SDS coat proteins of maclura mosaic virus, a possible potyvirus, and of beet necrotic yellow vein virus, a possible tobamovirus, did not react with antisera to definitive members of the respective groups. Several unexpected reactivities of SDS coat proteins of viruses from one group with antisera to viruses in other groups were, however, also observed. This indicated that electro-blot immunoassay, like other serological methods, cannot be used reliably to assign a virus to a certain group. The SDS coat proteins of several tombusviruses were able to mimic serological reactions by a direct binding of enzymelabelled antibodies which occurred also without serum treatment. Zusammenfassung Electro-blot Immunoassay—eine Methode zum Studium serologischer Verwandtschaften zwischen Pflanzenviren? Nach Elektrophorese in SDS-Polyacrylamidgelen wurden die Hullproteine („SDS”-proteine) von verschiedenen Tymo-, Tombus-, Como-, Nepo-, Tobamo-, Potex-, Carla- und Potyviren elektrophoretisch auf Nitrozellulosemembranen ubertragen und mit Seren behandelt, die gegen intakte Viren hergestellt worden waren. Immunkomplexe wurden durch Ziegen-Anti-Kaninchen-Antikorper nachgewiesen, die mit alkalischer Phosphatase markiert waren. Die „SDS”-proccine der Comoviren reagierten nur mit den fur die intakten Viren homologen Antiseren. In allen anderen Gruppen wurden daruber hinaus auch Reaktionen mit Seren gegen andere Viren der gleichen Gruppe beobachtet. Eine Reihe von entfernten Verwandtschaften zwischen Gliedern einer Gruppe wurden zum ersten Mal festgestellt. Die „SDS”-proteine von Maclura mosaic virus, einem muglichen Potyvirus, und beet necrotic yellow vein virus, einem muglichen Tobamovirus, reagierten nicht mit Seren gegen definitive Viren der entsprechenden Gruppen. Unerwarteterweise reagierten die „SDS”-proteine von mehreren Viren auch mit Seren gegen Viren in anderen Gruppen. Electro-blot Immunoassay kann daher, ebenso wie andere serologische Methoden, nicht zuverlassig zur Bestimmung der Gruppenzugehorigkeit eines Virus benutzt werden. Die „SDS”-proteine einiger Tombusviren tauschten serologische Reaktivitaten durch eine direkte Bindung von enzymmarkierten Antikorpern vor, die auch ohne Serumbehandlung beobachtet wurde.

The best results in attempts to purify beet necrotic yellow vein virus were obtained with a slightly modified Triton × 100 method originally described by Hullet al. (1976) for cauliflower mosaic virus. Immunosorbent electronmicroscopy (ISEM), the double antibody sandwich form of enzyme-linked immunosorbent assay (ELISA) and electro-blot immunoassay (EBIA) yielded similar result when sugar beets were tested for the presence of virus. ISEM and EBIA ofer the advantage that in addition to serology a second parameter, i.e. either particle morphology or coat protein molecular weight, is used to identify the virus. Specific and possibly nonspecific reactions are thus more readily distinguished and antisera can be used which may still contain some antibodies to normal plant constituents. However, with these methods only a limited number of samples can be studied at a time, therefore ELISA is presently the method of choice for large scale routine testing. Zusammenfassung Beet necrotic yellow vein virus: Reindarstellung, Herstellung von Antiseren Und Nachweis Mit Hilfe von ELISA, Immunosorbent electronmicroscoy und Electro-Blot Immunoassay Bei Versuchen Zur Reindarstellung des beet necrotic yellow vein virus wurden die besten Ergebnisse mit einer leicht modifiziertem Triton × 100 Methode erzielt, die ursprung fur das cauliflower mosaic virus von Hullet al. (1976) entwickelt worden war. Immunosorbent electronmicroscopy (ISEM), double antibody-sandwid, ELISA und electro-blot immunoassay (EBIA) ergaben ahnliche Resultate bei der Unteuchung von Zuckerrubenwurzelproben. ISEM und EBIA haben den Vorteil, das neber der scrolognchen Reaktivitat noch ein zweites Merkmal zum Nachweis des Virus eingeserz wird, bei ISEM die Partikelmorphologie und bei EBIA das Molekulargewicht des Hullprodtim badurda kannen spezifische und eventuell unspezifische Reaktionen besser unterschreden werden und es konnen Antiseten verwendet werden, die noch Antikorper gegen normale Pflauzenbestandteile enthalten. Da mit diesen Methoden jedoch nur eine begrenzte Anzabl von Probcagleichzeitig untersucht werden kann, ist fur Massenuntersuchungen ELISA die Methode der Wahl.

Complementary deoxyribonucleic acids (cDNAs) to the four ribonucleic acid (RNA) species of isolate YU2 of beet necrotic yellow vein virus (BNYVV) were prepared by reverse transcription. The duplexes of these cDNAs were incorporated in the plasmid pBR322 and cloned into E. coli strain 5K. The insert sizes of the cDNA clones ranged from 280 to 2100 base pairs. In Southern blot hybridizations not all cDNAs specific for a particular RNA of BNYVV cross-reacted. This suggests that they are complementaryto different regions of the respective RNA. In dot blot hybridization experiments sixteen out of eighteen cDNA clones detected all thirteen BNYVV isolates which were obtained from sugar beets originating from Yugoslavia, Italy, Switzerland, France and various regions in Germany and which were maintained in Chenopodium quinoa. Two cDNA clones specific for RNA-3 failed to detect one German isolate. Pronounced differences between the thirteen virus isolates were seen when the electrophoretic migration of their RNAs was compared in Northern blot hybridizations. RNA-1 and RNA-2 of all isolates had the same sizes of 7100 and 5200 bases, respectively. The size, number and relative concentration of the smaller RNAs, however, varied greatly with different, isolates.

Attempts were made to introduce three different isolates of beet necrotic yellow vein virus (BNYW) into the roots of sugarbeets by means of mechanical inoculation. In Chenopodium quinoa these isolates had produced different patterns of viral RNA. The RNA pattern of isolate P was similar to that found in sugarbeets infected naturally by means of the vector Polymyxa betae. RNA 3 and 4 were partially deleted in isolate Y and apparently absent in isolate R. About 20% of the sugarbeet roots inoculated either with isolate P or Y became infected systemically, but with isolate R no systemic infections were recorded. In tap and side roots of plants infected with isolate P the virus concentration remained high for at least ten months and the tap roots of these plants showed a very poor growth. In tap and side roots of plants infected with isolate Y the virus concentration gradually dropped until it reached the limit of detectability. The tap roots of these plants became about five times as big as those infected with isolate P. Our results indicate that BNYW with full length RNA 3 and 4 can cause heavy damage to sugarbeets also in the absence of Polymyxa betae and that RNA 3 and/or 4 — in addition to a recently suggested involvement in virus transmission by P. betae — apparently also facilitate the persistent spread of the virus in sugarbeet roots. Zusammenfassung Mechanische Inokulation von Zuckerrubenwurzeln mit Rizomaniavirus-Isolaten unterschiedlicher RNA-Zusammensetzung Es wurde versucht, durch mechanische Inokulation Isolate des Rhizomaniavirus (BNYVV) unterschiedlicher RNA-Zusammensetzung in Zuckerruben-Keimlingswurzeln einzubringen. Das Isolat P hatte in Chenopodium quinoa ein RNA-Muster gezeigt, das von dem in naturlich infizierten Ruben nicht zu unterscheiden war, beim Isolat Y waren RNA 3 und 4 in verkurzter Form aufgetreten, beim Isolat R fehlten sie vollstandig. Systemische Wurzelinfektionen wurden 7 Wochen p.i. bei ca. 20% der entweder mit Isolat P oder Y inokulierten Pflanzen festgestellt, aber nicht bei den mit Isolat R inokulierten. Die BNYW-ELISA-Werte blieben in den Wurzeln der mit Isolat P beimpften Pflanzen wahrend der gesamten einjahrigen Versuchsdauer hoch, in den Wurzeln der mit Isolat Y beimpften Pflanzen wurden sie dagegen immer niedriger, etwa 5 Monaten p.i. war das Virus in der Mehrzahl dieser Pflanzen nicht mehr nachweisbar. Die Rubenkorper dieser Pflanzen wurden etwa funf Mal groser als die mit Isolat P infizierten. Daraus ergibt sich, das BNYW mit intakter RNA 3 und 4 auch in Abwesenheit von Polymyxa betae zu einer schweren Schadigung der Ruben fuhren kann. Unsere Ergebnisse deuten auserdem darauf hin, das intakte RNA 3 und/oder 4 die Ausbreitung des Virus in der Rube erleichtern und nicht nur, wie kurzlich vermutet wurde, seine Ubertragung durch P. betae.