Detection of Phytophthora nicotianae and P. citrophthora in Citrus Roots and Soils by Nested PCR

The polymerase chain reaction (PCR) was used for the specific detection of Phytophthora nicotianae and P. citrophthora in citrus roots and soils. Primers were based on the nucleotide sequences of the internal transcribed space regions (ITS1 and ITS2) of 16 different species of Phytophthora. Two primer pairs, Pn5B–Pn6 and Pc2B–Pc7, were designed specifically to amplify DNA from P. nicotianae and P. citrophthora, respectively. Another primer pair (Ph2–ITS4) was designed to amplify DNA from many Phytophthora species. All primer pairs were assessed for specificity and absence of cross-reactivity, using DNA from 118 isolates of Phytophthora and 82 of other common soil fungi. In conventional PCR, with a 10-fold dilution series of template DNA, the limit of detection was of 1pgl^–1 DNA for all the primer pairs (Ph2–ITS4, Pn5B–Pn6, and Pc2B–Pc7). In nested PCR, with primers Ph2–ITS4 in the first round, the detection limit was of 1fgl^–1 for both the primer sets (Pn5B–Pn6 and Pc2B–Pc7). Simple, inexpensive and rapid procedures for direct extraction of DNA from soil and roots were developed. The method yielded DNA of a purity and quality suitable for PCR within 2–3h. DNA extracted from soil and roots was amplified by nested PCR utilizing primers Ph2–ITS4 in the first round. In the second round the primer pairs Pn5B–Pn6 and Pc2B–Pc7 were utilized to detect P. nicotianae and P. citrophthora, respectively. Comparison between the molecular method and pathogen isolation by means of a selective medium did not show any significant differences in sensitivity.     

Real-time detection of <Emphasis Type="Italic">Phytophthora nicotianae</Emphasis> and <Emphasis Type="Italic">P. citrophthora</Emphasis>in citrus roots and soil

Two primers, specific for Phytophthora nicotianae (Pn6) and P. citrophthora (Pc2B), were modified to obtain Scorpion primers for real-time identification and detection of both pathogens in citrus nursery soils and roots. Multiplex PCR with dual-labelled fluorogenic probes allowed concurrent identification of both species ofPhytophthora among 150 fungal isolates, including 14 species of Phytophthora. Using P. nicotianaespecific primers a delayed and lower fluorescence increase was also obtained from P. cactorumDNA. However, in separate real-time amplifications, the aspecific increase of fluorescence from P. cactorum was avoided by increasing the annealing temperature. In multiplex PCR, with a series of 10-fold DNA dilutions, the detection limit was 10 pg l^-1 for P. nicotianaeand 100 pg l^–1 for P. citrophthora, whereas in separate reaction DNA up to 1 pg l^-1 was detected for both pathogens.Simple and rapid procedures for direct DNA extraction from soil and roots were utilised to yield DNA whose purity and quality was suitable for PCR assays. By combining these protocols with a double amplification (nested Scorpion-PCR) using primers Ph2-ITS4 amplifying DNA from the main Phytophthora species (first round) and PnB5-Pn6 Scorpion and Pc2B Scorpion-Pc7 (second round), it was possible to achieve real-time detection of P. nicotianaeand P. citrophthora from roots and soil. The degree of sensitivity was similar to that of traditional detection methods based on the use of selective media. The analyses of artificially and naturally infested soil showed a high and significant correlation between the concentration of pathogen propagules and the real-time PCR cycle threshold.     

Toxigenic Fusarium Species of Liseola Section in Pre-harvest Maize Ear Rot, and Associated Mycotoxins in Slovakia

The occurrence of Fusarium species of Liseola section and related toxins was investigated for two years (1996 and 1998) on maize ear rot samples collected in the most important areas for maize growing in Slovakia. The species most frequently isolated was F. verticillioides, followed by F. proliferatum in 1996 and F. subglutinans in 1998. Most of the strains belonged to mating populations A, D, and E of the teleomorph Gibberella fujikuroi. Fusarium graminearum was also frequently recovered in both the years of investigations. Toxin analysis of maize ears showed that most of the samples (21 out of 22) were contaminated with at least one toxin. In particular, the concentration of fumonisin B₁, and fumonisin ₂ was up to 26.9 and 5.1gg^-1, respectively in 1996, and up to 12.1 and 6.3gg^-1, respectively in 1998. Beauvericin was detected only in one sample in 1996. Seven samples in 1996 were contaminated by fusaproliferin up to 8.2gg^-1, but just traces of the toxin were found in one sample in 1998. All 29 strains of F. verticillioides, two of three strains of F. proliferatum and none of eight F. subglutinans strains isolated from samples produced fumonisin B₁ in culture on whole maize kernels (0.1–5646 and 940–1200ugg^-1, respectively). Two strains of F. subglutinans and two of F. proliferatum produced beauvericin (up to 65 and 70gg^-1, respectively). Ten strains of F. verticillioides produced beauvericin: 9 strains produced a low amount (up to 3gg^-1), while only one of them produced a high level of toxin (375gg^-1). Fusaproliferin was produced by two F. proliferatum strains (220 and 370gg^-1), by seven F. subglutinans (20–1335gg^-1) and by three F. verticillioides (10–35gg^-1). This is the first report on fusaproliferin production by F. verticillioides, although at low level.     

Electron microscope observations of nuclear polyhedra fromMalacosoma neustria (Lepidoptera: Lasiocampidae

The nuclear polyhedral bodies fromMalacosoma neustria are enclosed within a membrane. The diameter of the nuclear polyhedra varies from 0.9 to 2.8 with an average of 1.8 . In the nuclear polyhedra the rod-like virus particles occur both singly and in bundles. The single virus rods are enclosed within two membranes, namely the intimate membrane and the developmental membrane. The virus rods which occur in bundles have an intimate membrane just like the single virus rods, whereas the developmental membrane encloses the whole bundle. The virus rods are closely packed by the intimate membrane and between the intimate and the developmental membrane is a space. The diameter of the virus rods without membranes, determined from sectioned polyhedra, is about 25 m and the length 250 m.     

Production of Beauvericin by Different Races of Fusarium Oxysporum F. sp. Melonis, The Fusarium Wilt Agent of Muskmelon

Fourty-four strains of Fusarium oxysporum were isolated from plants of melon with Fusarium wilt symptoms. Among these strains, thirty-nine were characterized for their pathogenicity on melon. Thirty-seven strains belonged to known races of F. oxysporum f. sp. melonis, while two strains were non-pathogenic. Four strains belonged to race 0, seven to race 1, four to race 2, and twenty-two to race 1,2. Beauvericin was produced by thirty-six strains in a range from 1 to 310gg^–1. Eight isolates of race 1,2 did not produce the toxin. In addition, of the two non-pathogenic strains, only one strain produced the toxin (290gg^–1). The production of enniatin A₁, enniatin B₁, and enniatin B was also investigated. Enniatin B was the only enniatin detected, being produced by eleven strains belonging to all the races, with a range of production from traces to 60gg^–1. Finally, melon fruits belonging to two different cultivars (Cantalupo and Amarillo) were artificially inoculated with one strain of F. oxysporum f. sp. melonis (ITEM 3464). Beauvericin was detected in the fruit tissues of both cultivars at a level of 11.2 and 73.8gg^–1, respectively. These data suggest that the production of both the toxins is not related to the pathogenicity of F. oxysporum f. sp. melonis, nor to the differential specificity of the races. The results confirm that beauvericin is a common metabolite of phytopathogenic Fusarium species.     

Epidemiology of Fusarium Infection and Deoxynivalenol Content in Winter Wheat in the Rhineland, Germany

Details of our long-term research programme concerning the epidemiology of Fusarium spp. and mycotoxin production are summarized. Evaluation of the occurrence of Fusarium spp., mainly on winter wheat (Triticum aestivum), was carried out by investigating Fusarium infection and mycotoxin contamination. Two to 15% of grains were infested during 1995–1998 at three climatologically differing localities of the Rhineland, Germany. Disease progress was accelerated by rainfall during the flowering season. The species most frequently isolated were Fusarium avenaceum, F. poae, F. culmorum and F. graminearum. The mean deoxynivalenol (DON) content varied from 19gkg^–1 (1995) to 310gkg^–1 (1998) and was not always correlated with disease severity. Organic farming systems showed lower rates of infection with ear blight and lower mycotoxin contamination than conventional farming systems.     

Real-time Scorpion-PCR detection and quantification of <Emphasis Type="Italic">Erwinia amylovora</Emphasis> on pear leaves and flowers

A specific primer couple (E3–E4) amplifying a single DNA fragment of 111 bp from plasmid pEA29 was designed to identify, detect and quantify Erwinia amylovora by real-time Scorpion-PCR. Specificity of primers and probe was assessed both by means of BLAST analyses and by using genomic DNA from a large number of E. amylovora isolates and other bacteria. In Scorpion-PCR, the limit of detection was of 1 pg of total DNA and a high correlation (r = 0.999) was achieved between target DNA quantity and cycle threshold (Ct). Combining two sequential amplifications with conventional reported primers (PEANT1–PEANT2) and Scorpion primers (E3 Scorpion-E4) the detection limit was of 1 fg (nested Scorpion-PCR). Using serial dilution of the bacterial suspensions the limit of detection was 3.2 × 10⁴ CFU ml⁻¹ in Scorpion-PCR and 2.8 × 10² CFU ml⁻¹ in nested Scorpion-PCR. Real-time PCR combined with effective procedures for DNA extraction enabled the detection and the quantification of the epiphytic population of E. amylovora in the washings of flowers and leaves of artificially inoculated pear. A significant correlation (r = 0.92) was achieved between pathogen CFU on semi-selective media and the corresponding target DNA concentration evaluated by real-time PCR.     

Species-specific PCRs differentiate <Emphasis Type="Italic">Rosellinia necatrix</Emphasis> from <Emphasis Type="Italic">R. compacta</Emphasis> as the prevalent cause of white root rot in Japan

Rosellinia compacta, described recently, resembles R. necatrix and also causes white root rot. Here a species-specific PCR was developed for R. compacta, and the two R. necatrix-specific primer sets already available were validated in terms of species specificity. PCRs using the primer sets for R. necatrix amplified specific products exclusively from R. necatrix isolates. The R. compacta-specific primer set exclusively detected R. compacta, which appears to be a rare but widely distributed species. We conclude that R. necatrix is the major cause of the disease in Japan but that the involvement of R. compacta should be studied further.     

Effect of acetochlor treatment on Fusarium wilt and sugar content in melon seedlings

Pretreatment of soil with the herbicide acetochlor at 0.1–1g g^–1 significantly decreased incidence of wilt due toFusarium oxysporum f. sp.melonis in melon seedlings. Glucose, fructose and sucrose increased in leaves of inoculated and uninoculated melon plants following acetochlor treatment. The increase in sugar levels in stems and roots was less pronounced. Light intensity affected sugar content and disease incidence. The percentage of diseased plants was significantly higher in untreated plants grown under 165E m^–2 sec^–1 compared to plants grown under 300E m^–2 sec^–1. Lowering light intensity resulted in reduction of levels of total sugars on the third and sixth day after inoculation. Acetochlor had little or no effect on growth rate or sporulation of the pathogen in culture. The colonization rate of diseased plant stems by the pathogen was similar in herbicide-treated and untreated plants, thus excluding the possibility that disease reduction by the herbicide is related to direct fungitoxicity.Contribution from the Agricultural Research Organization. No. 1560-E, 1995 series.     

Quantitative studies on dispersal of Pseudocercosporella herpotrichoides spores from infected wheat straw by simulated rain

Splash dispersal ofPseudocercosporella herpotrichoides spores from infected wheat straw was investigated using simulated rainfall (rate 13.8l h^–1 m^–2, volume mean diameter 2.9 mm) and wind (2 m sec^–1) in a raintower/wind tunnel complex. Spores were deposited on the floor of the wind tunnel up to 1 m upwind and 2.5 m downwind from the centre of the straw and impacted on vertical surfaces at heights up to 36 cm above it. Fewer spores were collected with increasing distance from the straw and with increasing height. Most spore-carrying splash droplets were in the size range 400–600 m and very few were less than 200 m.Our results show that these spores are generally dispersed over short distance, which is consistent with field observations.Samenvatting Spetterverspreiding vanPseudocercosporella herpotrichoides van besmet tarwestro werd onderzocht met behulp van nagebootste regen (13,8 l uur^–1 m^–2, gemiddelde druppel diameter 2,9 mm) en wind (snelheid 2 m sec^–1) in een proefopstelling van een regentoren en een windtunnel. Sporen werden op de vloer van de windtunnel gedeponeerd tot 1 m afstand tegen de wind in, gerekend vanaf het centrum van het tarwestro, en met de wind mee tot 2,5 m afstand daarvan. Op verticale vlakken werden sporen op een hoogte van 0 tot 36 cm boven de vloer van de windtunnel opgevangen. Naarmate de afstand en de hoogte toenamen werden er minder sporen gevonden. De meeste spetters met sporen hadden een diameter van 400–600 m en slechts enkele waren kleiner dan 200 m.Onze resultaten tonen aan dat de sporen in het algemeen slechts over korte afstanden verspreid worden, hetgeen overeenkomt met veldwaarnemingen.     

PCR-based specific detection of <Emphasis Type="Italic">Ralstonia solanacearum</Emphasis> race 4 strains

Two primer sets were designed based on the sequence of polymorphic bands that were derived from repetitive sequence-based polymerase chain reaction (rep-PCR) fingerprinting and specifically detected in Ralstonia solanacearum race 4 strains (ginger, mioga, and curcuma isolates). One primer set (AKIF-AKIR) amplified a single band (165bp) from genomic DNA obtained from all mioga and curcuma and some ginger isolates; another set (21F-21R) amplified one band (125bp) from the other ginger isolates. These primer sets did not amplify the bands from genomic DNA of other R. solanacearum strains or of other related bacteria. PCR detection limit for the pathogen was 2 × 10²cfu.The nucleotide sequence data reported are available in the DDBJ/EMBL/GenBank databases under accession numbers AB118756 and AB118757     

Biological control of cucumber powdery mildew by Tilletiopsis minor

Samenvatting Onder optimale omstandigheden konT. minor de ontwikkeling van komkommermeeldauw (Sphaerotheca fuliginea) tegengaan.Spuiten met 2×10⁷ sporen ml^–17 dagen na inoculatie met komkommermeeldauw gaf een reductie van meeldauwontwikkeling van ongeveer 90%. Wanneer een tweede bespuiting met dezelfde concentratie sporen 3 dagen na de eerste werd toegepast bleven de planten vrij van meeldauw tot ze werden opgeruimd 3 weken later.Bij een R.L. lager dan 70% en een temperatuur boven 30 °C had geen van de behandelingen succes. T. minor bleek ongevoelig voor dimethirimol (Milcurb) bij een concentratie van 125 g ml^–1, terwijl er gemakkelijk een mutant kon worden verkregen, die resistent was tegen 100 g fenarimol ml^–1, bij gelijk blijvende groeikracht en pathogeniteit ten opzichte van komkommermeeldauw, waardoorT. minor ingepast kan worden in een schema voor geïntegreerde bestrijding.     

Drain of nutrients imposed when Aphis sambuci and a. fabae feed on the inflorescence of Yucca flaccida,and the amounts retained by the aphids

For the production of 1 g dry weight of aphids about 10 g dry weight (55 ml) of sieve tube sap ofYucca flaccida is required, provided all N taken up by the aphids is retained by them. The production capacity of an averageYucca leaf was calculated at 4 mg dry weight of aphids per day or 30 g per cm² leaf area per day. Compared with N only about 30% of the P, K, Na, Mg, and Ca ingested by the aphids is retained by them, apparently in the same mutual ratios as these elements occur in the sieve tube fluid.Samenvatting Bij een aangenomen 100% benutting van de door luizen uit zeefvaten opgenomen N, is voor de ontwikkeling van 1 g drooggewicht vanA. fabae enA. sambuci ca. 10 g droog zeefvatensap nodig (55 ml). De produktiecapaciteit van eenYucca blad van gemiddelde grootte werd berekend op 4 mg droge luis per dag, overeenkomend met ca. 30 g per cm² bladoppervlak.Vergeleken met die van de N is de benutting van de door de luizen opgenomen hoeveelheden P, K, Na, Mg, en Ca slechts ongeveer 30%; dit gebeurt in ongeveer dezelfde onderlinge verhoudingen als waarin deze elementen in het zeefvatensap aanwezig zijn (Tabel 1).     

Effect of benomyl on soil fungi associated with rye. 1. Effect on the incidence of sharp eyespot caused by Rhizoctonia cerealis

Effects of benomyl on incidence of pathogens affecting the culm base of rye were studied in field trials and growth chamber experiments. Spraying of the crop with the fungicide at a high dosage (2.4 kg.ha^–1) resulted in a tenfold increase of sharp eyespot caused byRhizoctonia cerealis and reduced foot rot symptoms caused by fusaria by 50%. In a field trial at a low dosage (0.24 kg.ha^–1) a slight increase of sharp eyespot was observed. In one year, probably because of wet conditions during the infection period, sharp eyespot did not occur in either benomyl-treated or untreated plots, but eyespot caused byPseudocercosporella herpotrichoides was abundant. Its occurrence was reduced from 74% affected culm bases in untreated plots to 8% and 1% in plots that received 0.24 and 2.4 kg.ha^–1 of the fungicide, respectively.In growth chambers seedlings were grown in two sandy soils inoculated withR. cerealis. The soil was kept dry at about 35% of the moisture holding capacity. In plots with benomyl (1 mg.kg^–1; moisture content 11% of fresh weight), fewer seedlings emerged than in plots without the fungicide. This result was highly significant (P<0.01) for one soil but not for the other. The number of seedlings that remained free of disease symptoms was higher (P<0.01) in untreated than in fungicide-treated plots of both soils.Isolates of pathogens obtained from diseased culms were tested for their sensitivity to benomyl. Growth of all of them includingR. cerealis was inhibited, although not always completely suppressed, at 10 g.ml^–1 on potato-dextrose agar. ED₅₀ values of most isolates ofR. cerealis were between 2.2 and 3.1 g.ml^–1. The fungus was slightly but consistently less sensitive thanF. culmorum. Mycelial growth ofF. nivale was appreciably more sensitive than that of the otherFusarium spp. from cereals.P. herpotrichoides andF. nivale were the most sensitive pathogens tested with ED₅₀ values of <1 g.ml^–1. Accordingly,F. nivale was absent on culms from treated plots. In a growth chamber experiment, seedlings were protected from infection by supplying the fungicide (1 mg.kg^–1) to previously inoculated soil.In a laboratory assay the effect of benomyl on microbial antagonism toR. cerealis was estimated for rhizosphere soil. Enhanced incidence of sharp eyespot in treated crops was associated with adverese effects of the fungicide on microbial antagonism. There is presumptive evidence thatR. cerealis is suppressed by bacteria after wet periods during the vegetation period of the crop and by fungi after dry periods. Only fungal antagonism, which may be less effective, is affected by benomyl. The response to benomyl of the microflora in different soils varied. Reasons for this inconsistency are suggested.Samenvatting In veldproeven en in een klimaatkamer werd de invloed van benomyl op het optreden van voetziekten in rogge onderzocht. In veldjes die bespoten waren met een hoge dosis van het fungicide (in totaal 2.4 kg.ha^–1) bleken tienmaal zoveel halmen met scherpe oogvlekken, veroorzaakt doorRhizoctonia cerealis, voor te komen dan in onbespoten veldjes. Daarentegen was voetrot veroorzaakt doorFusarium-soorten met 50% verminderd. In een volgende veldproef, waarbij een voor de praktijk geadviseerde dosis (0.24 kg.ha^–1) was toegepast, werd een lichte toename van scherpe oogvlekken waargenomen.In een ander jaar trad scherpe oogvlekkenziekte in het geheel niet op, ook niet in met benomyl behandelde veldjes. De vochtige omstandigheden tijdens de infectieperiode zijn daarvan waarschijnlijk de oorzaak. Daarentegen werd de oogvlekkenziekte, welke doorPseudocercosporella herpotrichoides werd veroorzaakt, veel aangetroffen. In de onbehandelde veldjes waren 74% van de halmen aangetast tegen 8 en 1% in de veldjes die met het fungicide waren behandeld in doseringen van 0.24 en 2.4 kg.ha^–1.De invloed van het fungicide op de aantasting van kiemplanten werd in klimaatkamerproeven onderzocht. Daartoe werden twee zandgronden metR. cerealis geënt. De grond werd droog gehouden (op 35% van het waterhoudend vermogen). In grond met fungicide (1 mg.kg^–1) was de opkomst minder dan in grond zonder fungicide. Dit was zeer significant (P<0.01) voor één van de beide zandgronden, maar niet voor de andere. Het aantal gezonde kiemplanten was in beide gevallen duidelijk hoger (P<0.01) voor de onbehandelde grond.De isolaten van ziekteverwekkers uit aangetaste halmen werden op hun gevoeligheid voor het fungicide getoetst. Op aardappel-glucoseagar werden alle isolaten in hun groei geremd bij een benomyl-concentratie van 10 g.ml^–1.R. cerealis was iets minder gevoelig danF. culmorum. Voor het overgrote deel van de isolaten vanR. cerealis lag de ED₅₀ waarde tussen 2,2 en 3,1 g.ml^–1. De myceliumgroei vanF. nivale werd meer geremd dan die van de andereFusarium-soorten.P. herpotrichoides enF. nivale waren met een ED₅₀ waarde van <1 g.m.^–1 de gevoeligste pathogenen die uit de halmvoeten werden geïsoleerd. Dat de populatie vanF. nivale in benomylhoudende grond wordt onderdrukt, blijkt uit (1) het feit dat de schimmel niet voorkwam op halmen uit behandelde veldjes en (2) de bescherming tegen infectie van kiemplanten als aan de besmette grond fungicide (1 mg.kg^–1) was toegevoegd.In laboratoriumproeven werd de invloed van benomyl op het microbiële antagonisme in rhizosfeergrond tegenR. cerealis bepaald. Een toename in het optreden van scherpe oogvlekkenziekte in behandelde gewassen bleek gepaard te gaan met een remming van het antagonisme tegen de ziekteverwekker. Er zijn sterke aanwijzingen datR. cerealis na vochtige perioden tijdens de vegetatieperiode door bacteriën wordt onderdrukt en na droge perioden door schimmels. Het antagonisme van de laatste groep lijkt minder effectief te zijn en alleen dit antagonisme wordt door benomyl verlaagd. Tenslotte wordt een mogelijke oorzaak aangegeven voor de ongelijke respons op het fungicide van het microbieel antagonisme in verschillende gronden.     

Influence of mixed biocide GCSC-BtA on the pupae and adult stages of Apanteles plutellae Kurd. (Hym., Braconidae) and its host. Plutella xylostella (L.) (Lep., Plutellidae)

This paper deals with the influence of the mixed biocide GCSC-BtA on the pupal and adult stages of Apanteles plutellae Kurd. (Hym., Braconidae) and its host, Plutella xylostella (L.) (Lep., Plutellidae). The results show that mortalities of the pupae of P. xylostella in the direct-dip bioassay were 84.67%, that of the adults in the residue bioassay at 1.2500mg/ml concentration of GCSC-BtA were 78.00% which were significantly higher than the mortality values for the pupae with 54.62% and adults with 48.13% of A. plutellae. In contrast, cypermethrin showed extremely high toxicity to the pupae with 94.58% and adults with 86.00% mortality values of A. plutellae as compared to the low mortality values of 42.14% for the pupae and 32.11% for the adults of P. xylostella, with the same concentrations and bioassay methods. The LC₅₀ values of GCSC-BtA were 0.3402, 0.5516 and 1.2405, 1.9480mg/ml for the pupae and adults of P. xylostella and A. plutellae, respectively, while the LC₅₀ values for cypermethrin were 1.5652, 2.3471 and 0.1096, 0.1152mg/ml, respectively. GCSC-BtA was found more toxic to the pupae and adults of P. xylostella and safer to the pupae and adults of A. plutellae than cypermethrin. The possibilty of using GCSC-BtA against P. xylostella under partial control by A. plutellae in vegetable fields is discussed.     

Infection of Tomicus piniperda (Col., Scolytidae) with Canningia tomici sp. n. (Microsporidia, Unikaryonidae)

Canningia tomici sp. n. (Microsporidia, Unikaryonidae) infects the midgut epithelium, the gut muscules, Malpighian tubules, connective tissues, adipose tissues and the gonads of the pine shoot beetle, Tomicus piniperda (L.) (Coleoptera, Scolytidae). The infection is present in populations of Tomicus piniperda in Europe and in the United States. Uninucleate oval single spores occur in two sizes: 2.8±0.4× 1.4±0.4m and 3.8±0.3×2.0±0.2m. The polar filament of this microsporidium is fixed subapically in a flat anchoring disc. The thick posterior lamellae of the binary polaroplast are asymmetric due to the lateral fixation of the polar filament.     

Acquired resistance to benomyl and some other systemic fungicides in a strain of Botrytis cinerea in cyclamen

A benomyl-resistant strain (R) ofBotrytis cinerea was isolated from cyclamen that had been sprayed with relatively high doses of Benlate two weeks before. In vitro mycelial growth of this strain was less inhibited on PDA containing 1000 g/ml benomyl (Benlate, 50% W.P.) than that of another, wild isolate ofB. cinerea from cyclamen on PDA with 0.5 g/ml of the fungicide.The R-strain was also resistant to methyl-thiophanate, furidazol and to a lesser extent to thiabendazole. Mycelial growth of 5 other isolates was much more inhibited by benomyl than by thiabendazole.Resistance was retained for at least 20 weeks after repeated subculturing on fungicide-free agar.Samenvatting In een kwekerij, waar bespuiting met benomyl (Benlate, 50% W.P.) drie maal was toegepast ter bestrijding vanBotrytisrot in cyclamen, bleek de laatste bespuiting niet meer effectief. Integendeel, de ziekte breidde zich sneller uit dan onder normale omstandigheden het geval is. Uit bloemstelen van de aangetaste planten werd eenB. cinerea-stam (R) geïsoleerd, die zeer resistent bleek tegen benomyl. In vitro werd de groei van deze stam op aardappel-glucose-agar met 1000 g/ml benomyl (Benlate 50% W.P.) minder geremd dan die van een willekeurigB. cinerea-isolaat van cyclamen op het medium met 0.5 g/ml van het fungicide (Tabel 1, Fig. 1).De R-stam bleek eveneens resistent tegen methyl-thiophanaat, furidazol en in mindere mate tegen thiabendazol (Tabel 2).De myceliumgroei van vijf isolaten vanB. cinerea, verkregen van verschillende waardplanten, bleek in tegenstelling tot die van de R-stam juist sterker geremd te worden door benomyl dan door thiabendazol (Tabel 3).De R-stam bleef gedurende tenminste 20 weken resistent na regelmatig overenten op voedingsbodems zonder het fungicide.     

Resistance to benomyl and some chemically related compounds in strains of Penicillium species

TwoPenicillium species, vizP. brevicompactum andP. corymbiferum, were isolated from senescent petioles of cyclamen and from bulbs of lilies, respectively, both samples treated previously with benomyl. The isolates turned out to be very resistant to this fungicide when grown on malt agar, supplied with the fungicide; at a concentration of 2000 g/ml they were less inhibited than randomly chosen isolates of the same species on agar with 1 g/ml.The strains retained their resistance at the same level for at least 3 months after repeated subculturing on fungicide-free agar.Resistance to benomyl coincided with resistance to methyl-thiophanate and, to a lesser extent, also to thiabendazole and furidazol.Samenvatting Uit afstervende bladstelen van cyclamen en uit schubben van leliebollen, welke eerder met benomyl waren behandeld, konden respectievelijkPenicillium brevicompactum enPenicillium corymbiferum worden geïsoleerd. De isolaten blekenin vitro zeer resistent tegen het fungicide. De myceliumgroei van deze isolaten werd op moutagar met 2000 g/ml benomyl minder geremd dan die van willekeurige isolaten van dezelfde soorten op agar met 1 g/ml (Fig. 1).De isolaten bleven gedurende tenminste 3 maanden resistent na regelmatig overenten op voedingsbodems zonder het fungicide.De resistente stammen van de beidePenicillium-soorten bleken eveneens resistent tegen methyl-thiophanaat en in mindere mate ook tegen thiabendazol en furidazol (Tabel 1). De volgorde van de groeiremmende werking van deze fungiciden was voor de willekeurig gekozen (gevoelige) isolaten: benomyl>thiabendazol>methyl-thiophanaat >furidazol. Voor de resistente stammen was deze: thiabendazol en furidazol >benomyl>methyl-thiophanaat. In het feit dat een dergelijke verandering in volgorde van remmend effect ook voorBotrytis cinerea geldt, ligt een aanwijzing, dat de wijze waarop de resistentie werkt, voor deze schimmels gelijk is.     

Metabolism of phaseollin by different races of Colletotrichum lindemuthianum

The ability of four races of the bean pathogenColletotrichum lindemuthianum to metabolize the phytoalexin phaseollin in shake cultures was compared. Apart from some differences in the rate of conversion, all races metabolized the phytoalexin in the same way. Phaseollin was first converted to 6a-hydroxyphaseollin, and this product was further metabolized to 6a, 7-dihydroxyphaseollin. No metabolites of the latter compound could be detected.6a, 7-Dihydroxyphaseollin was as inhibitory as phaseollin to race 11, but was only slightly inhibitory to races 1, 2 and 1.Samenvatting Een vergelijkend onderzoek werd verricht naar het vermogen van vier fysiologische rassen van het bonepathogeenColletotrichum lindemuthianum om het fytoalexine phaseolline om te zetten.In schudculturen waaraan 10 g phaseolline/ml was toegevoegd, werd dit door alle fysio's op gelijke wijze omgezet, hoewel met verschillende snelheid (Fig. 1). Steeds werd phaseolline eerst omgezet tot 6a-hydroxyphaseolline, en dit produkt vervolgens tot een verbinding die geïdentificeerd kon worden als 6a, 7-dihydroxyphaseolline. Hierna konden geen verdere produkten worden aangetoond.6a, 7-Dihydroxyphaseolline was even fungitoxisch als phaseolline voor fysio 11, maar was slechts weinig fungitoxisch voor de fysio's 1, 2 en 1 (Tabel 1).De verschillen in omzettingssnelheid van phaseolline en in gevoeligheid voor phaseolline ee zijn omzettingsprodukten die tussen de fysio's gevonden zijn, zijn onvoldoende om de fysiospecifieke interacties tussen de boon en de verschillende fysio's vanC. lindemuthianum te verklaren.     

Scab Response and Moniliformin Accumulation in Kernels of Oat Genotypes Inoculated with Fusarium avenaceum in Poland

Inoculation experiments with 14 genotypes of oats (10 cultivars and 4 lines) were performed during 1996, 1997 and 1998 in Sitaniec, South-Eastern Poland. Panicles of oats were inoculated with a conidial suspension of Fusarium avenaceum, which caused a reduction in yield by 33% and in 1000 kernel weight (TKW) by 21%. During the period between inoculation and harvest, F. avenaceum was able to accumulate moniliformin (MON) in kernels at an average level of 0.13mgkg^–1 (gg^–1). The highest reduction of yield components caused by the F. avenaceum inoculation was found for cv. Santor, followed by lines CHD 1171, STH 2795 and cvs: Kwant and Farys, while cvs Slawko, Dukat, Borys and Komes exhibited the highest resistance to the disease in terms of TKW and yield reductions after inoculation.