Cultural conditions were optimized so that disease reactions of plantlets and callus tissue from partially resistant and susceptible genotypes of Pisum sativum could be differentiated when inoculated with Fusarium solani f.sp. pisi. Disease was assessed using both semi‐quantitative (disease score/fungal diameter) and quantitative methods (ergosterol assay). After optimization of inoculum concentration and incubation period it was possible to differentiate the disease reactions of plantlets using disease scores and amount of ergosterol in the tissues. Incubation period, temperature and the auxin added to the media were important in allowing differentiation of callus tissue cultures. These differences were visible by measurement of the diameter of fungal growth on the callus and the amount of ergosterol in the tissues, but not with disease scores. The plantlet test was also applicable to the other foot rot pathogens Phoma medicaginis var. pinodella and Aphanomyces euteiches, allowing differentiation of the disease reactions of a partially resistant and susceptible genotype using disease scores. The ergosterol assay could be used to quantify infection caused by P. medicaginis var. pinodella, but this sterol was not detectable in mycelium of A. euteiches. In this fungus the presence of cholesterol was detected, which may be used to quantify infection.

A simplified method for the quantitative assessment of the fungal lipid ergosterol was used to assess the levels of infection in tissue cultures of oilseed rape (Brassica napus ssp. oleifera) inoculated with Leptosphaeria maculans. The growth of L. maculans in liquid culture throughout a 36‐day period correlated well (r = 0·92) with the amount of ergosterol extracted from the mycelium. There were significant differences (P < 0·05) in the amount of ergosterol extracted from infected thin cell layer (TCL) explants and callus tissue of two resistant and three susceptible cultivars of oilseed rape. Amounts of ergosterol extracted from resistant cultivars were < 100 (g and from susceptible > 100 (g. The mean amounts of ergosterol extracted from shoot cultures of two resistant and four susceptible cultivars were similar to those for TCL explants and callus tissue, although the values obtained were variable. This technique can be used in in vitro breeding programmes to accurately assess the resistance of tissue cultures of B. napus to L. maculans and could also have value in conventional breeding programmes.

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.

As a basis for devising an in vitro screening programme, culture conditions were optimized so that tissue cultures from two resistant cultivars of Brassica napus ssp. oleifera (Mikado, Bienvenu) and two susceptible cultivars (Lesira, Ceres) could be differentiated using a disease scoring scheme, when inoculated with Leptosphaeria maculans. Tissues inoculated included thin cell layer explants from soil‐grown plants and in vitro‐grown shoot cultures and callus tissue formed on such explants. The period of incubation and the incubation temperature were of importance in the development of differential disease reactions. Increasing temperature generally resulted in an increase in infection and too great an incubation period resulted in total overgrowth of the tissue. Increasing concentrations (1 × 10−6 M‐1 ×10−4 M) of the auxins 1‐naphthylacetic acid (NAA), 2,4‐dichlorophenoxyacetic acid (2,4‐D) and mdole‐3‐acetic acid (IAA) in the culture medium, resulted in a decrease in disease score of the thin cell layer (TCL) explants from soil‐grown plants. The cytokinins examined 6‐benzyl‐aminopurine (BAP) and 6‐4‐hydroxy‐3‐methyl‐2‐enylaminopurine (zeatin), reduced the extent of infection of the TCL explants when used in combination with the auxin NAA. Medium containing NAA at a concentration of 1 × 10−6 M in combination with BAP at a concentration of 1× 10−6 or 1 × 10−4 M allowed differentiation of the disease reactions of the resistant and susceptible cultivars, when the explants were incubated for 10 days at 20 °C after inoculation. Similar conditions of incubation and the addition of NAA (1 × 10−6 M) combined with BAP (1 × 10−6 M) to the medium also allowed the differentiation of the disease reactions on TCL explants from stems of in vitro shoot cultures of the cultivars Mikado and Lesira. Increasing concentrations of the auxin NAA and the cytokinin BAP resulted in a reduction in the mean disease score of the callus tissue produced on TCL explants from soil‐grown plants, and NAA (1 × 10−5 M) combined with BAP (1 × 10−6 or 1 × 10−5 M) allowed differentiation of resistance and susceptibility in callus tissues when incubated for 5 days at 20 °C. 2,4‐D did not allow differentiation of the cultivars. This was in contrast to the inoculation of callus tissue attached to TCL explants of in vitro shoot cultures, where combinations of 2,4‐D and BAP at concentrations of 1 × 10−6 M allowed differentiation of the resistant and susceptible cultivars. These findings provide a basis for designing selection protocols of value in both traditional as well as in vitro breeding programmes to select lines of oilseed rape with resistance/novel resistance to L. maculans.