Evaluation of methods of screening strawberry cultivars for resistance to crown rot caused by Phytophthora cactorum

Four methods were evaluated in measuring resistance of strawberry cultivars to crown rot caused by Phytophthora cactorum. Meristem propagated plants grown in vitro were inoculated with mycelial discs. Four to five days after inoculation, it was possible to distinguish between cultivars with large differences in susceptibility to the disease. Ten days later, all plants were totally necrotic making it impossible to distinguish between cultivars. When detached leaves were inoculated by inoculating a plug of mycelium into the petiole, disease symptoms developed more slowly in resistant cultivars, but leaf age greatly affected the rate of symptom development. When plug plants (not cold stored) were lightly wounded in the rhizome with a scalpel and inoculated with either zoospores or mycelium, differences in disease development between cultivars were mainly as would be expected from previous information on susceptibility, but both age and size of plants influenced the rate of disease development. Unwounded, inoculated plants did not develop symptoms. When cold‐stored plug plants were either unwounded or lightly wounded with a scalpel in the rhizome and inoculated with zoospores, the relative rates of disease development consistently reflected the susceptibility to crown rot. At the time of final assessment, disease was much more severe in wounded plants, but the relative susceptibility of cultivars was not affected by the wounding.     

Systemic infection of black currant flowers by Botrytis cinerea and its possible involvement in premature abscission of fruits

Emasculated flowers of several black currant cultivars were pollinated and then inoculated with dry conidia of Botrytis cinerea in the field and glasshouse. The infection of pistils was examined by U.V. fluorescence microscopy and the incidence of premature flower abscission recorded. Conidia germinated in the stigmatic fluid in all cultivars and hyphae spread symptomlessly throughout the style to infect the pericarp and ovules. Of six cultivars inoculated in the field, cv. Ojebyn was the most, and cv. Ben More the least resistant to flower shedding. Natural infection of stigmas by B. cinerea was common in the field and a high proportion of apparently healthy non-inoculated flowers which abscissed were found to contain infected ovules. Fewer flowers abscissed if inoculations were made 6 days after pollination. Symptomless or latent infection of black currant flowers by B. cinerea may be a contributory cause of premature abscission of developing fruits, or ‘running-off’, recorded in these experiments.     

Rooted leaf cuttings as an aid in screening for resistance to Plasmodiophora brassicae

Rooted leaf cuttings were used to screen a range of genotypes and cultivars from Brassica spp. and Raphanus sativus for resistance to Plasmodiophora brassicae. The optimum conditions for obtaining equivalent disease incidence and disease severity on leaf cuttings and on seedlings were investigated. In the method used leaves were cut near the base of the petiole before the lamina was fully expanded. They were kept in a mist propagator for 6 to 18 days and then transferred to a clubroot test bench and individually inoculated with a suspension of resting spores. There was an optimum period for each cultivar between cutting and inoculation which gave the maximum incidence of infection: c. 15 days for Doon Major (swede); 10–15 days for Civasto (stubble turnip) and Giant Rape (forage rape); c. 22 days for Maris Kestrel (marrow stem kale) and c. 10 days for Slobolt (fodder radish). Gall size on rooted leaf cuttings gave a quantitative measure of disease severity equivalent to that on seedlings in tests using the European Clubroot Differential host set, the five above-mentioned cultivars and four swede cultivars. There was little callus production except on some B. campestris hosts and on cuttings with shoot tissue; microscopic examination was used to confirm the presence or absence of infection in galls. Hormone rooting powder did not increase the rate of root production in two cultivars of forage rape. Root development was less extensive in B. campestris than in B. oleracea or B. napus. Apparently normal shoots developed on 96% of leaf cuttings (inoculated and uninoculated) in which a fragment of axillary bud tissue was included. The results are discussed in relation to screening for resistance in whole plants and in tissue culture.     

Infective behaviour and aphid-transmissibility of Italian isolates of potato virus Y in tobacco and peppers

When mechanically inoculated to susceptible tobacco (Nicotiana tabacum L.) cultivars, nine isolates of PVY from Umbria (Central Italy) and two from Southern Latium gave rise to rapid systemic infection which developed within 6–8 days after inoculation. Systemic spread of the same isolates was slower, or much slower, in infected pepper (Capsicum annuum L.) cultivars, 8–14 days for Southern Latium isolates and 20 - 35 days for Umbrian ones. Aphid (Myzus persicae)-moculation of pepper and tobacco plants with two of the Umbrian and one of the Southern Latium isolates confirmed the results from sap-transmission and showed that fewer inoculated pepper plants become infected, especially with Umbrian isolates. In agreement with the data on systemic spread, aphid-acquisition trials indicated that tobacco plants became efficient PVY sources for vectors 6–8 days after inoculation with either group of isolates. Peppers became efficient acquisition hosts 8–15 days after inoculation with Southern Latium isolates but not until 22–45 days after inoculation with Umbrian ones. Southern Latium isolates induced more severe symptoms in pepper cultivars than Umbrian isolates did. One of the Southern Latium isolates was able to systemically infect the resistant pepper cv. Yolo Y, which was never infected by the Umbrian isolates. The Umbrian isolates tested seem to be better adapted to tobacco than peppers, while Southern Latium ones are well adapted to both.     

Arbuscular mycorrhizal fungal-induced alteration to root architecture in strawberry and induced resistance to the root pathogenPhytophthora fragariae

Three strawberry cultivars Elsanta, Cambridge Favourite and Rhapsody were inoculated with eitherGlomus fasciculatum orGlomus etunicatum and their growth compared with non-inoculated plants. The roots of all inoculated plants were 55 to 70% colonised after 98 days. Increases in both root and shoot dry weights were measured. Root architecture was also determined and increases in branching were evident in AMF colonised root systems. The remaining plants were then inoculated with the root pathogenPhytophthora fragariae and allowed to grow for a further 58 days before harvest. In two of the cultivars, Cambridge Favourite and Elsanta, AMF reduced root necrosis by approximately 60 and 30% respectively. Only in the least susceptible cultivar, Rhapsody, was no reduction measured in AMF colonised plants. There were differences in the control conferred by the two arbuscular mycorrhizal fungi and this suggests there may be practical benefits of inoculation. Relationships between the presence of roots of different orders, on inoculation with the pathogen, and subsequent necrosis provided a mechanism for identifying root-architecture driven alteration to susceptibility. Root system necrosis was positively correlated with the proportion of the root system made up of higher order roots (3° to 4°) in non-colonised plants and negatively correlated in AMF colonised plants. These data suggest that root-architecture changes are not important per se but factors expressed concurrently may be.     

Induction of Susceptibility and Resistance of Wheat Leaves by Preinoculation with High and Low Virulence Races of Puccinia recondita f. sp. Tritici

Isolates R1 and R2 of Puccinia recondita f. sp. tritici of low or high virulence on leaves of a series of cultivars of wheat behaved very similarly up to the formation of appressoria above stomata and sub-stomatal vesicles. Differences in growth of hyphae and in responses of host cells became clearly apparent only 24 h after infection; they then increased so that 10 days after inoculation high virulence isolates had produced freely sporulating sori (infection type 4) whereas low virulence isolates produced small colonies which did not erupt and sporulate and were associated with large numbers of dead host cells. The cultivar Timmo was selected for experiments on induced susceptibility and induced resistance. It was highly susceptible (infection type 4) to isolate Rl and to a mutant RlM, similar in pathogenicity on many cultivars but with uredospores different in colour from those of R1 and those of isolate R2 to which Timmo was highly resistant (infection type 0). This difference facilitated interpretation of results of experiments in which leaves were inoculated with more than one isolate. Inoculation with R1 (or with RUM which always behaved similarly) made leaves susceptible to R2 inoculated 4 days later. R2 sori were about 70 % the area of and about 70 % as frequent as Rl sori when inoculated on the same area on the same surface or on an area on the lower surface below a similar area on the upper surface inoculated with Rl. There also developed in the same area small, non-erumpent colonies associated with large numbers of dead host cells; these necrotic lesions were very similar to those produced in leaves inoculated only with R2. There were significantly fewer sori and more necrotic lesions when Rl was inoculated 1 or 2 days before R2. Totals of sori and necrotic lesions were about the same when the interval between inoculations was 1, 2 or 4 days; they were also similar with an interval of 3 days when numbers of sori were substantially less than after 4 days. Inoculation with Rl had some but much less effect in making tissues susceptible to R2 when again it was inoculated later but about 2 mm away. Although inoculation with R1 sufficiently in advance had the striking effect of making tissue almost as susceptible to normally low virulence R2 as it was to R1, colonies and sori of R1 which developed in association with those of R2 were substantially smaller than those which developed when R1 was inoculated alone. But there were similar differences in sizes of colonies and sori when R1 and RIM were inoculated together or separately. They probably depended on competition for nutrients. Possible mechanisms of induced susceptibility and induced resistance and implications for disease in the field are discussed briefly.     

Receptivity of various wheat cultivars to infection by VA-mycorrhizal fungi as influenced by inoculum potential and the relation of VAM-effectiveness to succinic dehydrogenase activity of the mycelium in the roots

Wheat cultivars grown in pots in a greenhouse were inoculated either once or twice with the vesicular-arbuscular mycorrhizal fungus (VAMF) Glomus mosseae. If inoculum was only added to the soil once (before planting) the cultivars showed differences in subsequent VAM development. If additional inoculum was added ten days after planting, VAM development was much increased and cultivars which remained without VAM after only one inoculum application now became mycorrhizal.Succinate dehydrogenase (SDH) activity decreased throughout the experiment, but this decrease was less rapid in mycelium in twice-inoculated plants.No close relationship between SDH-activity and plant growth (VAM effectiveness) was found.     

Screening of cassava varieties for resistance to root-knot nematodes

Twenty cassava cultivars were screened in micro-plot experiments in the green house of Institute of Agricultural Research and Training, Obafemi Awolowo University, Moor Plantation, Ibadan for root-knot nematode resistance. Four weeks after planting, each plant was inoculated with 5000 eggs of Meloidogyne incognita. There were three replications per cultivar. The plants were uprooted and assessed for resistance after 60 days of inoculation. Assessment of plant damaged was determined by gall index and reproductive factor. Based on host status rating, 3 cultivars were found to be resistant, 4 cultivars were tolerant, 10 cultivars ere susceptible while .3 cultivars were hypersusceptible. All the screened cultivars are resistant to cassava mosaic disease and other major pests of cassava like bacterial disease, anthracnose disease, cassava green mite and cassava mealy bug. They are all high yielding, suitable for food, industry and livestock feed.     

Mycotoxin production on different cultivars and lines of broad bean (Vicia faba L.) seeds in Egypt

One hundred different cultivars and lines of broad bean (Vicia faba L.) seed samples were inoculated with Aspergillus flavus Link (CMI 102135) to determine varietal differences which may support or resist aflatoxin production. Thin-layer chromatographic analysis of the chloroform extracts of the different seed samples revealed that 11 cultivars/lines were highly resistant to seed invasion and aflatoxin production while 9 cultivars/lines showed partial resistance. The remaining 80 samples were susceptible to the establishment of A. flavus and aflatoxin accumulation. All the resistant cultivars/lines seed samples were inoculated also with three local isolates of fungi namely; Stachybotrys chartarum (Ehrenb. ex Link) Hughes, Aspergillus ochraceus Wilhelm, and Fusarium oxysporum Schlecht. The resistant seed samples were also resistant for colonization with these fungi and mycotoxin formation.     

Localisation and movement of Plum pox virus in apricot stem tissues

Localisation and movement of Plum pox virus (PPV), sharka disease, in stem tissues of susceptible and resistant apricot (Prunus armeniaca L.) cultivars was studied. Two different assays were performed. In the first assay, apricot cultivars were grafted on to a non‐inoculated GF305 peach rootstock and, after two months, the sprouted apricot was inoculated by chip‐grafting. In the second assay, apricot cultivars were grafted on to a previously chip‐inoculated GF305 showing strong PPV symptoms. Localisation of virus was studied in apricot stem by immuno‐tissue printing and sharka symptoms in GF305 and apricot leaves were also observed. Virus was mainly localised in the xylem, and sometimes in the cortex and pith. Results revealed that, while all the cultivars allowed limited virus movement from the inoculation point, only the susceptible cultivars (Screara, Bebeco and Colomer) allowed long distance movement and even showed symptoms in leaves.     

In vitro evaluation of sugarcane resistance to gumming disease and of Xanthomonas campestris pv. vasculorum aggressiveness

Sugarcane plantlets were sectioned halfway between the base and the youngest ligule and then inoculated by soaking the wound in a suspension of Xanthomonas campestris pv. vasculorum. The infection caused rapid necrosis of the inoculated leaves, chlorosis of uninoculated leaves, or death of the inoculated plantlet. New tillers sometimes showed chlorosis or white streaks. The effects of the inoculum concentration, the cultivar, and the bacterial strain on symptom severity were determined. The ranking of cultivars depended on the inoculum concentration, and strains were found to differ with regard to aggressiveness. However, cultivars and strains were more effectively classified in greenhouse trials. The poor expression of leaf resistance appeared to limit the use of the in vitro test.     

Cytology of infection,development and expression of resistance to Plasmodiophora brassicae in canola

The timing and expression of resistance to four isolates of Plasmodiophora brassicae, collected from research sites where pathotypes 2, 3, 5 and 6 (Williams' system) had been dominant when characterised in 2006, were assessed in four new commercial cultivars of canola (Brassica napus) with resistance to clubroot. Each of the resistant cultivars was highly resistant to all four of the isolates, and there was no difference in their response to infection. Root hair infection occurred at high levels, but pathogen development occurred more slowly than in a susceptible cultivar (control). Secondary infection and development in cortical cells was severely inhibited in each of the resistant cultivars; only a few bi‐nucleated plasmodia were observed at 12 days after inoculation (DAI), and plasmodia were rarely observed at 18 and 24 DAI. In contrast, development in the susceptible cultivar had progressed to resting spores by 24 DAI. A dense ring of accumulated reactive oxygen species (ROS) was observed in the endodermis, pericycle and vascular cambium of non‐inoculated controls and inoculated plants of the resistant cultivars. However, the ROS ring disappeared rapidly in infected plants of the susceptible cultivar. Plasmodia invaded the stele of susceptible roots by preferentially colonising the xylem parenchyma cells. Expansion and enlargement of lignified xylem cells was observed by 35 DAI. The absence of any specific points of ROS accumulation or lignification of epidermal or cortical cells in the resistant cultivars indicates that a hypersensitive response is not the main mechanism of resistance in these lines. The uniform response of these resistant cultivars to the four isolates of P. brassicae indicates that the resistance in each cultivar may be conditioned by a gene(s) from a single source that confers broad resistance, because most sources of resistance to P. brassicae are pathotype specific.     

Effects of nodulation on resistance to alfalfa sickness among ten alfalfa cultivars

Summary Ten alfalfa cultivars were used to study the effects of nodulation and variations in resistance to alfalfa sickness. The alfalfa seedlings were planted in sick soil with three treatment,,i.e.: pasteurized sick soil inoculated with Rhizobium ‘Nitragin’ which served as the control, the inoculated non-pasteurized sick soil and the non-inoculated non-pasteurized sick soil. None of the alfalfa cultivars were immune from the sickness. Cultivar ‘Anik’ fromMedicago falcata was among the most resistant cultivars. Three Phytophthora root rot resistant cultivars including ‘Agate’, ‘Apollo’ and ‘Ramsey’ were not resistant to the disease. Alfalfa inoculated with Rhizobium showed greatly improved seedling growth. Correlation coefficients showed that those alfalfa cultivars more resistant to alfalfa sickness produced more dry weight. Dry weight increase due to nodulation (82%) had more than compensated for the loss of dry weight due to alfalfa sickness (33%). The present study suggested that the poor growth of alfalfa on sick soil was attributed to both the soil borne pathogens and the poor nodulation of alfalfa, probably due to the absence of effective Rhizobium in sick soil. Highly significant differences were also obtained among olfalfa cultivars for plant dry weight in the inoculation treatment. Selection for effective Rhizobium strains and for alfalfa genotypes which are resistant to alfalfa sickness and are high in nitrogen fixation rates could improve alfalfa yield in sick soil.     

Phosphorus allocation in P-efficient and inefficient barley cultivars as affected by mycorrhizal infection

A glasshouse experiment was undertaken to investigate the effect of mycorrhizal infection on the allocation of phosphorus (P) in agronomically P-efficient (i.e. high yields at low P supply) and inefficient barley (Hordeum vulgare L.) cultivars. Four barley cultivars differing in agronomic P-efficiency were inoculated or not inoculated with Glomus etunicatum. Cultivars did not differ in percentage of root length infected. The concentration of P in roots of the inefficient cultivars was higher than that of the efficient cultivars. However, because of changes in root to shoot dry weight ratio and below-ground productivity, mycorrhizal infection significantly reduced the percentage of total plant P in roots of the inefficient cultivars. The distribution of P between root and shoot of P-efficient cultivars was not affected by mycorrhizal infection. Root to shoot dry weight ratio of the P-efficient cultivars was lower than that of the inefficient cultivars, and the decrease in the ratio following infection was significant in inefficient but not in P-efficient cultivars. This study indicates that mycorrhizal infection alters the allocation of P in inefficient cultivars and effectively improves the efficiency of P utilization with respect to shoot growth.     

Resistance in rice to Rice yellow mottle virus and evidence of non-seed transmission

Nine rice cultivars were evaluated under screenhouse conditions for resistance to Rice yellow mottle virus (RYMV) and possible seed transmission. Completely randomised design with three replications was used. In Experiment 1, the seedlings were inoculated with the virus at two weeks after planting. In Experiment 2, the seeds collected from Experiment 1 were dried for four weeks before planting. For each genotype, the seeds from healthy plants were planted as a control. Disease incidence and severity (scales 1–9), yield and yield components were recorded. Statistical analyses included Area Under the Disease Progress Curve (AUDPC) and independent t test. The cultivars FARO 37, FARO 52 and Gigante were highly resistant, whereas WAB189-B38HB was resistant. Paddy yield was highest (3.6 g) in FARO 37. There were no symptoms of virus disease in all the plants originating from the seeds of RYMV-infected plants. The differences between the seeds from infected and healthy plants for all the measured traits were not significant (p > 0.05). The number of days to seedling emergence was uniform (5.5 days) in all the cultivars. Plant height, number of tillers per plant, number of days to heading and paddy yield from the seeds of virus-infected plants varied from 54.8 to 68.4 cm, 17 to 21, 85.3 to 96 days and 2.7 to 4 g, respectively. Conversely, a range of 54.9–68.7 cm, 17–22, 83–95 days and 2.8–4.1 g was found in the seeds of healthy plants. Selection and cultivation of high-yielding, resistant and healthy seeds would enhance food security.     

Subterranean clover mottle sobemovirus: its host range, resistance in subterranean clover and transmission through seed and by grazing animals

Subterranean clover mottle sobemovirus (SCMV) was transmitted by manual inoculation of sap to 27 cultivars belonging to three sub-species of subterranean clover. The virus readily infected systemically all inoculated plants of five susceptible cultivars of ssp. subterraneum. Ten others showed partial resistance as not all infected plants developed systemic infection; cold winter conditions further delayed or prevented systemic movement in four of them. Two cultivars of spp. brachycalycinum and four of spp. yanninicum failed to develop systemic infection following inoculation and were considered highly resistant. Resistance to SCMV in three of the spp. yanninicum was further confirmed by the failure to establish detectable primary infections in most of the inoculated leaves. Moreover, when the four ssp. yanninicum cultivars were graft-inoculated with SCMV, systemic infection eventually developed in them but the virus concentration was low. SCMV was also transmitted by manual inoculation of sap to a further 23 species of Trifolium, Medicago or Pisum. Three species were non-hosts, five were infected only in inoculated leaves and 18 others developed systemic infection in some or all plants. SCMV reached very high concentrations and was stable in subterranean clover sap. It was transmitted experimentally between subterranean clover plants by brushing infected leaves against healthy ones and in swards was readily transmitted by the trampling and grazing of sheep, but only poorly by mowing. Seed transmission of SCMV to seedlings of five cultivars of subterranean clover was low (0–0.12%). SCMV was not transmitted by Myzus persicae.     

The Effect of Temperature on Symptom Expression and Colonization in Resistant and Susceptible Carnation Cultivars Infected with Fusarium oxysporum f. sp. dianthi

Six commercial carnation cultivars were inoculated with Fusarium oxysporum f. sp. dianthi race 2, and grown under three different temperature regimes. Colonization by the pathogen and development of wilt symptoms were assessed at intervals up to 40 days. No symptoms and very little colonization were seen in any of the cultivars at 14–15°C. At a temperature of 22°C, the cultivars were clearly differentiated into three groups: those with resistance, partial resistance or susceptibility to the pathogen depending on the severity of symptoms and the extent of fungal colonization. Symptom severity was associated with the extent of colonization. This differentiation was not seen at 26°C, when all cultivars except the most resistant, cv.‘Carrier 929′, rapidly became diseased and died by 23 days after inoculation. ‘Carrier 929’ also showed some wilt symptoms at this temperature and was colonized throughout the height of the stem after 40 days. The pathogen caused disease at 26°C by a combination of vascular wilting and stem base and root rotting. Fungal colonization was assayed at 22°C by the dilution plate/homogenization method and by estimation of fungal chitin in a highly resistant (‘Carrier 929′) and in a highly susceptible (‘Red Baron’) cultivar. Both methods of assay gave similar results. In ‘Red Baron’, colonization increased slowly up to 20 days after inoculation then progressed rapidly, closely following the development of severe wilt symptoms. In ‘Carrier 929’, colonization remained very low. The low level of fungal biomass in ‘Carrier 929’ compared with ‘Red Baron’ indicated that the former cultivar showed true resistance as opposed to tolerance to the disease.     

A glasshouse screening method to assess resistance in onions to damage by onion fly (Delia antiqua) larvae

Factors influencing the resistance of onion (Allium cepa) cultivars to onion fly (Delia antiqua) damage were investigated. These included different environments, methods of plant raising and egg inoculation, and plant size. A glasshouse compartment maintained at 14 to 37 °C was as satisfactory for assessing the resistance as a controlled environment. The basic test unit comprised 10 onion seedlings raised in a 400 × 50 × 50 mm deep plastic trough and inoculated with 50 onion fly eggs from a laboratory culture. Resistance was assessed by counting wilted plants every two or three days for 21 days after egg inoculation. The reciprocal of number of days for each plant to wilt (wilting rate) was an appropriate variate for statistical analysis, adjusted for the length of the longest leaf at the time of inoculation. The method enables cultivars or breeding lines of onion species to be reliably screened for resistance to larvae of this pest in a glasshouse so that only the most promising need be tested in the field.     

Improved Tolerance of Three Saudi Pearl Millet Cultivars (<i>Pennisetum spicatum</i>) to Salt Stress by Mycorrhiza

Seeds of three Saudi pearl millet cultivars (Pennisetum spicatum) from three regions (Madinah, Khulais and Jaizan) were inoculated with arbuscular mycorrhizal fungus Glomus mosseae obtained from the Agriculture Research Center of Giza, Egypt to enhance their salt tolerance. Five different NaCl concentrations (0, 30, 60, 90, and 120 mM) were used for treating cultivars with and without mycorrhiza. Growth rates, chlorophyll content, chlorophyll fluorescence (Fv/Fm), proline content and gas exchange were measured to determine the effect of salinity on these cultivars. The results indicated that compared to cultivars without mycorrhiza, all cultivars with mycorrhiza had enhanced growth and physiological parameters including shoot and root length, area and number of leaves, fresh and dry weights of shoots and roots, chlorophyll contents and gas exchanges at 0 and 30 mM of salinity. In addition, the measurements of the different growth rates showed higher growth performance of the cultivars from Madinah and Khulais than the cultivar from Jaizan. However, all cultivars with and without mycorrhiza showed significant reductions in growth rates, chlorophyll contents and gas exchanges at a salinity of 60 mM than those grown at 0 and 30 mM. Moreover, the values of Fv/Fm were significantly reduced in all cultivars with and without mycorrhiza grown at 60 mM than in those grown at 0 mM and 30 mM. Proline contents indicated a progressive increase with the elevation of NaCl concentration stress. The proline contents in the mycorrhiza-inoculated cultivars were significantly higher than those in the non-inoculated cultivars. On the other hand, all cultivars with and without mycorrhiza underwent senescence within four weeks of growth at salinity concentrations of 90 mM and 120 mM. Therefore, relatively low salinity must be maintained to achieve high growth rates and gas exchanges of these inoculated cultivars.     

A Comparison of the Disease Reactions of Stems and Detached Leaves of Soil and in vitro Grown Plants and Regenerants of Oilseed Rape to Leptosphaeria maculans and Protocols for Selection for Novel Disease Resistance

Protocols for selecting plant tissues of winter oilseed rape (Brassica napus subsp. oleifera) with resistance to Leptosphaeria maculans by either stem or leaf inoculation of both soil and in vitro grown plant material are described. The stem inoculation procedure gave good correlation (r = 0. 92) between the 50 day stem disease scores of eight out of nine cultivars of soil grown winter oilseed rape inoculated with isolate 41A4 of L. maculans and the N. A. B. esistance ratings or resistance data from field trials. The exception was the cultivar Liradonna. Inoculation of stems of five cultivars with isolates 41A4, 433 and 478 indicated a range of isolate virulence 478 > 41A4 > 433. This was the inverse of that observed in leaf inoculations. Application of the stem inoculation procedure to in vitro shoot cultures allowed differentiation of resistant and susceptible cultivars, including the cultivar Liradonna, after 20 days incubation at 20°C. The protocol was also applicable to plantlets regenerated from thin cell layer explants grown in vitro. Inoculations with isolate 433 allowed the differentiation of resistant, intermediately resistant and susceptible leaf material of soil grown plants, when leaf discs from young leaves were incubated on water agar supplemented with BAP (1 × 10^?5 M) at 25°C for 10 days. Intermediately resistant leaves were resistant after 10 days and susceptible after 15 days of incubation. Leaves of shoot cultures grown in vitro were more susceptible than the corresponding soil grown material. However, inoculation of old leaves with isolate 41A4 (an isolate of less virulence on leaves than 433) distinguished the cultivars after 15 days of incubation. These protocols allow the accurate assessment of resistance to L. maculans at the stem or leaf level and are of use in traditional as well as in vitro selection programmes.