Visual and PCR assessment of light leaf spot (Pyrenopeziza brassicae) on winter oilseed rape (Brassica napus) cultivars

Methods to assess light leaf spot ( Pyrenopeziza brassicae ) on winter oilseed rape cultivars were compared in laboratory, controlled-environment and field experiments. In controlled-environment experiments with seedling leaves inoculated at GS 1,4, the greatest differences in percentage area affected by P. brassicae sporulation were observed with inoculum concentrations of 4 × 10³ or 4 × 10⁴ spores mL⁻¹, rather than 4 × 10² or 4 × 10⁵ spores mL⁻¹, but older leaves had begun to senesce before assessment, particularly where they were severely affected by P. brassicae . In winter oilseed rape field experiments, a severe light leaf spot epidemic developed in 2002/03 (inoculated, September/October rainfall 127·2 mm) but not in 2003/04 (uninoculated, September/October rainfall 40·7 mm). In-plot assessments discriminated between cultivars best in February/March in 2003 and June in 2004, but sometimes failed to detect plots with many infected plants (e.g. March/April 2004). Ranking of cultivar resistance differed between seedling experiments done under controlled-environment conditions and field experiments. The sensitivity of detection of P. brassicae DNA extracted from culture was greater using the PCR primer pair PbITSF/PbITSR than using primers Pb1/Pb2. P. brassicae was detected by PCR (PbITS primers) in leaves from controlled-environment experiments immediately and up to 14 days after inoculation, and in leaves sampled from field experiments 2 months before detection by visual assessment.     

Effects of inoculum concentration, wetness duration and plant age on development of early blight (Alternaria solani) and on shedding of leaves in tomato plants

Effects of inoculum concentration, wetness duration and plant age on the development of tomato early blight were evaluated in relation to host susceptibility under controlled environmental conditions. The main effect of early blight was premature defoliation, which was linearly related to the percentage of leaf area showing symptoms. As ln(inoculum concentration, conidia mL⁻¹) increased from 6·2 to 11·5, the percentages of leaf area affected and of defoliation increased linearly. Four h of leaf wetness after inoculation were sufficient to initiate the disease on plants of hybrid Skala RZ but not on those of cv. Rio Rojo, for which at least 6 h leaf wetness were needed. As wetness duration increased up to 24 h, there was an increase in the percentage leaf area showing symptoms and in the percentage of defoliation, but thereafter there was no significant increase in either parameter. Tomato plants were susceptible to Alternaria solani at all growth stages, but susceptibility increased as plants matured. There were no significant differences in susceptibility between tomato cultivars and hybrids.     

Sporulation of Pyrenopeziza brassicae (light leaf spot) on oilseed rape (Brassica napus) leaves inoculated with ascospores or conidia at different temperatures and wetness durations

In controlled environment experiments, sporulation of Pyrenopeziza brassicae was observed on leaves of oilseed rape inoculated with ascospores or conidia at temperatures from 8 to 20°C at all leaf wetness durations from 6 to 72 h, except after 6 h leaf wetness duration at 8°C. The shortest times from inoculation to first observed sporulation ( l ₀), for both ascospore and conidial inoculum, were 11–12 days at 16°C after 48 h wetness duration. For both ascospore and conidial inoculum (48 h wetness duration), the number of conidia produced per cm² leaf area with sporulation was seven to eight times less at 20°C than at 8, 12 or 16°C. Values of Gompertz parameters c (maximum percentage leaf area with sporulation), r (maximum rate of increase in percentage leaf area with sporulation) and l ₃₇ (days from inoculation to 37% of maximum sporulation), estimated by fitting the equation to the observed data, were linearly related to values predicted by inserting temperature and wetness duration treatment values into existing equations. The observed data were fitted better by logistic equations than by Gompertz equations (which overestimated at low temperatures). For both ascospore and conidial inoculum, the latent period derived from the logistic equation (days from inoculation to 50% of maximum sporulation, l ₅₀) of P. brassicae was generally shortest at 16°C, and increased as temperature increased to 20°C or decreased to 8°C. Minimum numbers of spores needed to produce sporulation on leaves were ≈25 ascospores per leaf and ≈700 conidia per leaf, at 16°C after 48 h leaf wetness duration.     

Berry infection and the development of bunch rot in grapes caused by Aspergillus carbonarius

The effects of different levels of inoculum of Aspergillus carbonarius and time of inoculation on berry infection and the development of aspergillus bunch rot on grapevines (cv. Sultana) were studied under field conditions. Inflorescences at full bloom were inoculated with aqueous spore suspensions of A. carbonarius containing 0 or 1 × 10⁶ spores mL⁻¹ in 2004/05 and 0, 1 × 10² or 1 × 10⁵ spores mL⁻¹ in 2005/06. In both years, the incidence of infection in inoculated berries was significantly higher than in uninoculated berries. Incidence of infection in berries from veraison until harvest was higher than at earlier stages of bunch development (berry set to berries that were still hard and green). Inoculation of bunches at veraison did not significantly increase A. carbonarius infection prior to harvest, at harvest, 6 days after harvest or when berries were over-ripe. Bunches inoculated at harvest did not significantly increase infection 6 days after harvest or when berries were over-ripe. Aspergillus carbonarius was isolated more frequently from the pedicel end (53·1%) than from the middle section (37·5%) and distal end (35·0%) of berries that were inoculated with 10⁵ spores mL⁻¹.     

Genetic variation in Danish populations of Erysiphe graminis f.sp. hordei: estimation of gene diversity and effective population size using RFLP data

Genetic variation of the barley powdery mildew fungus ( Erysiphe graminis f.sp. hordei ) was estimated in three Danish local populations. Genetic variation was estimated from the variation amongst clones of Egh , and was therefore an estimate of the maximum genetic variation in the local populations. The average gene diversity, Ĥ , was estimated as 0.84. The effective population size was estimated as: log₁₀ ( N^ _e ) = 0.64 − log₁₀(μ), or 4.4 × 10⁹, assuming a nucleotide mutation rate ( μ) of 10⁻⁹ per base per generation. There was no significant genetic differentiation between locations.     

Biological control of damping-off of sugar beet by Pseudomonas putida applied to seed pellets

Pseudomonas putida 40RNF applied to seed pellets reduced the occurrence of Pythium damping-off of sugar beet. A density of 6 × 10⁷ 40RNF per pellet reduced Pythium damping-off from 70 to 26% when seeds were sown in artificially infested soil (250 propagules Pythium ultimum per g dry soil). The efficacy of 40RNF was dependent on its density in the seed pellet (in the range 2 × 10⁴–6 × 10⁸ per pellet) and on the number of propagules of Pythium in soil. 40RNF declined to or stabilized at approximately 1 × 10⁶ per pellet 3 days after planting, and this was independent of the inoculum density. This indicated that the crucial steps resulting in damping-off of sugar beet caused by Pythium ultimum must occur within 3–4 days of sowing. 40RNF reduced pericarp colonization by P. ultimum by 43% 48 h after planting and caused a 68% decrease in the number of sporangia of P. ultimum in the surrounding soil (0.0–5.0 mm). P. putida 40RNF also reduced pre and post-emergence damping-off (from 69.5 to 37.5%) caused by indigenous populations of Pythium species in an infested soil and this was as effective as the fungicide hymexazol (69.5 to 40%).     

Production of conidia by Peronosclerospora sorghi on sorghum crops in Zimbabwe

Factors affecting the production of conidia of Peronosclerospora sorghi , causing sorghum downy mildew (SDM), were investigated during 1993 and 1994 in Zimbabwe. In the field conidia were detected on nights when the minimum temperature was in the range 10–19°C. On 73% of nights when conidia were detected rain had fallen within the previous 72 h and on 64% of nights wind speed was < 2.0 m s⁻¹. The time period over which conidia were detected was 2–9 h. Using incubated leaf material, conidia were produced in the temperature range 10–26°C. Local lesions and systemically infected leaf material produced 2.4–5.7 × 10³ conidia per cm². Under controlled conditions conidia were released from conidiophores for 2.5 h after maturation and were shown to be well adapted to wind dispersal, having a settling velocity of 1.5 × 10⁻⁴ m s⁻¹. Conditions that are suitable for conidia production occur in Zimbabwe and other semi-arid regions of southern Africa during the cropping season.     

Effect of wind on the dispersal of oospores of Peronosclerospora sorghi from sorghum

The effect of wind on the dispersal of oospores of Peronosclerospora sorghi , cause of sorghum downy mildew (SDM) is described. The oospores are produced within the leaves of aging, systemically infected sorghum plants. These leaves typically undergo shredding, releasing oospores into the air. Oospores are produced in large numbers (6.1 × 10³ cm⁻² of systemically infected leaf) and an estimate of the settling velocity of single oospores (0.0437 m s⁻¹) of P. sorghi indicated their suitability for wind dispersal. In wind tunnel studies wind speeds as low as 2 m s⁻¹ dispersed up to 665 oospores per m³ air from a group of leaves previously exposed to wind and displaying symptoms of leaf shredding. The number of oospores dispersed increased exponentially with increasing wind speed. At 6 m s⁻¹, up to 12 890 oospores per m³ air were dispersed. Gusts increased oospore dispersal. A constant wind speed of 3 m s⁻¹ dispersed a mean of 416 oospores per m³. When gusts were applied the mean was 15 592 oospores per m³. In field experiments in Zimbabwe, oospores were sampled downwind from infected plants in the field and at a height of 3.8 m above ground level immediately downwind of an infected crop. These data indicate that wind could play a major role in the dispersal of oospores from infected plants in areas where SDM infects sorghum, perhaps dispersing oospores over long distances.     

Antifungal vapour-phase activity of allyl-isothiocyanate against Penicillium expansum on pears

The fungitoxicity of allyl-isothiocyanate (AITC) vapour against Penicillium expansum , the causal agent of blue mould on pears (cvs Conference and Kaiser), was evaluated. The best control of blue mould was obtained by exposing fruits for 24 h in a 5 mg L⁻¹ AITC-enriched atmosphere, the extent of control depending on the inoculum density. Lesion diameter was inversely related to AITC concentration. In treated fruits the percentage of infected wounds increased with conidial concentration, with fewer than 20% affected at 1 × 10³ conidia mL⁻¹ to almost 80% at 1 × 10⁶ conidia mL⁻¹. In comparison, >98% of wounds were infected in untreated fruits irrespective of conidial concentration. AITC treatments were effective up to 24 h after inoculation for Conference and 48 h for Kaiser. AITC treatments also controlled a thiabendazole-resistant strain of P. expansum , reducing the incidence of blue mould by 90% in both cultivars. The use of AITC produced from pure sinigrin or from Brassica juncea defatted meal may be an economically viable alternative to synthetic fungicides against P. expansum .     

Rust resistance in Salix to Melampsora larici-epitea

A total of 174 Salix (willow) clones belonging to 57 species and 14 interspecific hybrids were inoculated with seven pathotypes of Melampsora larici-epitea using the leaf disc method. Infection types were scored based on the uredinial pustule area data and the inoculum density. A close correlation ( R ² = 0·82) was found between the average pustule area and the average number of spores produced. Most of the willows were also assessed in the field for rust in 1999. Most willow clones belonging to the species native to western Europe were infected by the rust. In inoculation experiments, uredinia developed on 46 S. viminalis clones, out of a total of 47. In the field, all the S. viminalis clones were infected by rust. Within the subgenus Vetrix , eight out of the 17 willow species that originated from North and South America produced rust pustules in inoculation experiments. Of these, S. pellita was most susceptible. Salix irrorata and S. lasiolepsis var. bracelinae produced well developed pustules after inoculation but no rust infections were detected in the field. In both leaf disc tests and field assessments, no rust infections were found on S. candida , S. cordata , S. drummondiana , S. eriocephala , S. hookeriana , S. houghtonii , S. humilis , S. rigida var. mackenziana and S. syrticola . Of 12 species of subgenus Vetrix native to northeast Asia and Japan, only S. kochiana was susceptible both in inoculation tests and in the field. Salix rossica produced no symptoms in leaf disc tests but showed low levels of infection in the field. The maximum infection type scores in leaf disc tests were highly significantly correlated with field disease severity scores (Spearman rank correlation coefficient was 0·76, P  < 1 × 10⁻¹⁰).     

Interaction of a bioherbicide and glyphosate for controlling hemp sesbania in glyphosate-resistant soybean

The bioherbicidal fungus, Colletotrichum truncatum (Schwein.) Andrus & Moore, was tested at different inoculum concentrations alone and in combination with, prior to or following treatment with different rates of glyphosate ( N -[phosphonomethyl]glycine) (Roundup Ultra) for the control of hemp sesbania ( Sesbania exaltata [Raf.] Rydb. ex A.W. Hill) in Roundup Ready soybean field plots. Colletotrichum truncatum and glyphosate were applied in all pair-wise combinations of 0, 1.25, 2.5, 5.0, and 10.0 × 10⁶ spores mL⁻¹ (i.e. 3.125, 6.25, 12.5, and 25 × 10¹¹ spores ha⁻¹), and 0.15, 0.30, 0.60, and 1.2 kg ha⁻¹, respectively. Weed control and disease incidence were enhanced at the two lowest fungal and herbicidal rates when the fungal spores were applied after glyphosate treatment. The application of the fungus in combination with or prior to glyphosate application at 0.30 kg ha⁻¹ resulted in reduced disease incidence and weed control regardless of the inoculum's concentration. At the highest glyphosate rates, the weeds were controlled by the herbicide alone. These results suggest that it might be possible to utilize additive or synergistic herbicide and pathogen interactions to enhance hemp sesbania control.     

Factors influencing infection of mechanical wounds by Fusarium circinatum on Monterey pines (Pinus radiata)

Pitch canker, caused by the fungus Fusarium circinatum , is a disease affecting many pine tree species. In California, Pinus radiata (Monterey pine) is the principal pine host affected by pitch canker. This investigation into factors affecting infection frequency by F. circinatum of P. radiata examined the influence of: (i) wound size; (ii) relative humidity; (iii) time of inoculation; (iv) inoculum density; and (v) wound age. Wounded branches sustained significantly more infections when large-diameter (1·6 mm) rather than small-diameter (0·5 mm) wounds were made. Infection frequencies tended to be higher at 100% RH than at ambient humidity, although these differences were not statistically significant. Infection frequencies were significantly higher on branches inoculated after 17·00 h than on branches inoculated before noon. Infection frequencies were significantly higher for wounded branches spray-inoculated with 5 × 10⁷ rather than 1 × 10⁷ spores mL⁻¹. Infection frequencies of pruning wounds declined as wounds aged.     

PCR-based specific and sensitive detection of Pectobacterium carotovorum ssp. carotovorum by primers generated from a URP-PCR fingerprinting-derived polymorphic band

A 24-mer primer pair was generated by sequencing a URP-PCR fingerprinting-derived polymorphic band that is uniquely shared in Pectobacterium carotovorum ssp . carotovorum strains (Pcc). The primer set (EXPCCF/EXPCCR) amplified a single band of expected size (0·55 kb) from genomic DNA obtained from 29 Pcc strains and three Pectobacterium carotovorum ssp. wasabiae (Pcw) strains, but not from other P. carotovorum subspecies atrosepticum , betavasculorum or odoriferum , or from other Erwinia spp. or bacterial genera. The Rsa I digestion profile of the amplified bands divided Pcc strains into five groups with a unique profile from Pcw strains. First-round PCR detected between 5 × 10² and 1 × 10³ colony forming units (CFU) mL⁻¹ and detection sensitivity was increased to as few as 2–4 CFU mL⁻¹ after second-round (nested) PCR. This PCR protocol was used directly to detect Pcc strains in infected plant tissues.     

Growth and photosynthesis of four invasive aquatic plant species in Europe

Crassula helmsii , Hydrocotyle ranunculoides , Ludwigia grandiflora and Myriophyllum aquaticum are four well known invasive aquatic plants in European waters. In this study, plant growth at different nutrient availabilities, regeneration capacity and photosynthesis were investigated. Results show high relative growth rates (RGR) of the species of up to 0.132 ± 0.008 g g⁻¹ dry weight (dw) day⁻¹ ( H. ranunculoides ) and a significant increase in RGR with increasing nutrient availability. All species show a high regeneration capacity and the ability to form new shoots from single nodes, even though it differs between the species. Ludwigia grandiflora and M. aquaticum also show regeneration from single leaves. Species differed in maximal amounts, and in temperature and light optima of net assimilation rates: H. ranunculoides leaves reach maximum photosynthetic rates of up to 3500 μmol CO₂ × h⁻¹ g⁻¹ dw, L. grandiflora (leaves) up to 2200 μmol CO₂ × h⁻¹ g⁻¹ dw, M. aquaticum (shoots) 400   μmol CO₂ × h⁻¹ g⁻¹ dw and C. helmsii (shoots) up to 200 μmol CO₂ × h⁻¹ g⁻¹ dw. Hydrocotyle ranunculoides , L. grandiflora and M. aquaticum preferred high light intensity and high temperatures, whilst C. helmsii was negatively affected by intense sunlight. Summarising, it can be assumed that at least H. ranunculoides , L. grandiflora and M. aquaticum can grow well under current and likely future central European climate conditions.     

Controlled environment studies on the infection of Emex australis by Phomopsis emicis

The effect of temperature, dew period, inoculum concentration and host maturity on the growth and development of Emex australis after inoculation with Phomopsis emicis was studied in controlled environments. Disease was assessed by recording changes in the length of stems, number of fully expanded leaves, dry weights, number of fruits and disease severity index. Disease was as severe at 18°C as it was at 28°C. There was a trend of increased disease with longer dew period. Inoculum concentrations of 1·10⁶ and 1·10⁷ conidia per ml resulted in a significant net reduction in stem growth compared with lower concentrations. Progressively higher disease ratings occurred as the inoculum concentration increased to 1·10⁷ conidia per ml. There were no significant effects of inoculation with P. emicis on either seedlings, rosettes or flowering rosettes, but 10-week-old plants were highly susceptible to infection and stem collapse. The implications of these results for the potential development of P. emicis as a mycoherbicide are discussed.     

Effects of bean rust (Uromyces appendiculatus) epidemics on host dynamics of common bean (Phaseolus vulgaris)

Epidemics of bean rust ( Uromyces appendiculatus ) and their effects on host dynamics of common bean ( Phaseolus vulgaris ) were studied in three controlled greenhouse experiments, with and without fungicide sprays, on two susceptible bean cultivars, Dufrix and Duplika. Bean plants were artificially inoculated with a suspension of 10⁵ U. appendiculatus urediniospores mL⁻¹ water and temporal disease progress, as well as host growth dynamics (leaf size and defoliation), were monitored on a leaflet basis in comparison with non-inoculated plants, which were sprayed with deionized water. Progress curves of bean rust, expressed as the proportion of leaf area occupied by pustules (uredinia), or as the proportion occupied by total lesion area (= halo areas + pustule area), were well described by logistic functions with maximum disease levels clearly lower than 1. Bean rust epidemics substantially affected host growth by reducing the total leaf area formed by 17·4–35·6% and 35·3–46·2% compared with healthy plants for cvs Dufrix and Duplika, respectively. Fungicide sprays mitigated the negative effect of bean rust, leading to a gain in leaf area of 17–21% compared with unsprayed plants in both cultivars in two experiments, while in another experiment, disease control had no effect in Dufrix, but a clear effect in Duplika. In addition to the growth depression, bean rust also led to pronounced losses of leaf area as a result of reduced leaf size (leaf shrivelling) and accelerated defoliation.     

Infection of onion leaves by Alternaria porri and Stemphylium vesicarium and disease development in controlled environments

Infection of onion by Alternaria porri and Stemphylium vesicarium was investigated under a range of controlled temperatures (4–25°C) and leaf wetness periods (0–24 h). Conidia of A. porri and S. vesicarium germinated within 2 h when incubated at 4°C. Terminal and intercalary appressoria were produced at similar frequencies at or above 10°C. The maximum number of appressoria was produced after 24 h at 25°C. Penetration of leaves by both pathogens was via the epidermis and stomata, but the frequency of stomatal penetration exceeded that of epidermal penetration. There was a strong correlation ( R ² > 90%) between appressorium formation and total penetrations at all temperatures. Infection of onion leaves occurred after 16 h of leaf wetness at 15°C and 8 h of leaf wetness at 10–25°C, and infection increased with increasing leaf wetness duration to 24 h at all temperatures. Interruption of a single or double leaf wetness period by a dry period of 4–24 h had little effect on lesion numbers. Conidia of A. porri and S. vesicarium separately or in mixtures caused similar numbers of lesions. Alternaria porri and S. vesicarium are both potentially important pathogens in winter-grown Allium crops and purple leaf blotch symptoms were considered to be a complex caused by both pathogens.     

Current status of biological control of paddy weeds in Vietnam

Rice is a staple food in Vietnam and accounts for > 7.7 × 10⁶ cultivated ha, which provide 35.5 × 10⁶ t of rice, of which 4.2 × 10⁶ t were exported in 2004. The enlargement of the cropping area and the enhancement of rice yield have rapidly increased the amount of agrochemicals, including herbicides, in crop production in Vietnam. From 1990–2003, the percentage of herbicides in total pesticides has increased ≈ 10-fold to 30.2%. In addition, the improper use of herbicides caused environmental hazards, unsafe agricultural products, and human health problems. Biological management integrated with traditional weed control techniques might help to reduce the dependence on synthetic herbicides and build eco-friendly, sustainable agricultural production in Vietnam. This paper reviews the efforts in establishing a strategy for biological management of weeds that was conducted in recent years by Vietnamese weed scientists. This has included cropping system management, water and soil management, integrated pest management, and utilization of plant allelopathy as major components of the strategy. Many plants with strong allelopathic potential can be a source for biological weed suppression and soil fertility improvement. The utilization of allelopathic properties in rice might also help to provide new rice cultivars with weed-suppressing characteristics.     

Phytotoxic activity of root absorbed glyphosate in corn seedlings (Zea mays L.)

Growth chamber experiments were conducted in order to study the absorption, translocation and activity of glyphosate when applied to roots with aqueous solution avoiding any glyphosate–substrate interaction. Corn seedlings at the first leaf stage were set up in individual graduated cylinders containing different solutions of ¹⁴C-glyphosate (0–30 mg ae kg⁻¹). After 26 h of root exposure, plants were transferred to fresh nutrient solution and grown for the next 5 days. After harvest, plants were separated into seed, root, mesocotyle, coleoptile, cotyledon, first leaf and all new leaves (apex), and quantified ¹⁴C radioactivity contained in each part. Glyphosate uptake was only 11% of the theoretical mass flow into the plant. The amount of glyphosate translocated from roots was positively correlated with plant uptake ( P  < 0.01). Total plant fresh weight presented a logistic response to glyphosate amounts, including a growth stimulant effect (hormesis), when plants absorbed less than 0.6 µg. The treated plants presented a normal pattern of glyphosate allocation, with the apex the principal sink, accumulating more than 38% of mobilized glyphosate. When corn plants absorbed more than 0.6 µg they showed a decrease in growth. The relatively high glyphosate quantities allocated in the new leaves showed the relevance of the symplastic pathway in the translocation process for root absorbed glyphosate.     

The incorporation of pathogen spores into rain-splash droplets: a modelling approach

Simple, theoretical, physical principles and existing experimental data were used to derive an analytical model to describe the incorporation of plant pathogen spores into splash droplets. Data were obtained from experiments on splash dispersal of spores of Pseudocercosporella herpotrichoides (cereal eyespot), Pyrenopeziza brassicae (oilseed rape light leaf spot) and Septoria nodorum (wheat glume blotch). In these experiments, incident drops of diameter 4–5 mm were allowed to fall onto spore suspensions 0.5 mm deep with 1.2 × 10⁵ to 6.5 × 10⁵ spores/mL. The analytical model was constructed as the product of three functions of droplet diameter which described, respectively, the frequency distribution of droplet diameters, the proportion of droplets carrying spores and the mean number of spores in spore-carrying droplets in each diameter category. The frequency distribution of droplet sizes was described by a log-normal distribution, the proportion of droplets carrying spores was described by an exponential function and the adimensional spore concentration in spore-carrying droplets was described by a power law. The cumulative proportions of spores in droplets in diameter categories of increasing diameter were calculated to compare observed and fitted data.