Nineteen crosses were carried out in vitro between seven and nine isolates of Phytophthora infestans originating from potato and tomato, respectively. Oospores were produced abundantly in all but two crosses, but oospore germination was generally low (a few per cent) and depended on the combination of parental isolates. The highest fertility in F1 progeny was observed when at least one parental isolate originated from tomato, and the lowest in crosses of isolates from potato; half the crosses were not fertile. Forty‐three F1 and 51 F2 single‐oospore progeny of a selected cross, along with the 16 field isolates, were analysed phenotypically and with molecular markers. Phenotypic characterization included mating type; sensitivity to the phenylamide fungicide metalaxyl‐M; specific virulence on potato R‐gene differentials; and aggressiveness (infected leaf area) on potato and tomato leaf discs (host preference). Isolates and progeny were also assessed for mitochondrial DNA haplotype with RFLP–PCR (restriction fragment length polymorphism–polymerase chain reaction), and characterized with AFLP (amplified fragment length polymorphism) and SSR (simple sequence repeats, microsatellites). Parental isolates were sensitive and resistant to metalaxyl‐M, whereas all F1 were intermediate phenotypes. In the F2 progeny, the majority of isolates (43 of 51) were intermediate in sensitivity and four each were sensitive and resistant to metalaxyl‐M, respectively. In both F1 and F2 progeny, four isolates emerged through selfing. The A1 : A2 ratio was 25 : 18 in F1 and 24 : 21 (plus six self‐fertiles) among the F2 progeny. Many F1 progeny isolates were highly aggressive on both hosts, but 15 and 23% of isolates preferred tomato and potato, respectively. Among F2 progeny, few highly aggressive isolates were recorded and a quarter of isolates lost pathogenicity almost completely. Isolates preferring tomato increased, and those preferring potato significantly decreased in the F2 progeny. Inheritance of mitochondrial haplotype in F1 progeny was uniparental and mostly (25 of 27) from one parent only. Six and four different SSR genotypes were identified in F1 and F2 progeny, respectively, of which two were identical to the parents. The two microsatellite loci, 4B and 4G, segregated in the ratios 15 : 22 : 2 and 22 : 17 in F1 and 24 : 17 and 26 : 15 in F2 progeny, respectively, while the majority of AFLP markers segregated in either a 1 : 0, 1 : 1, 3 : 1 or 1 : 2: 1 ratio. There was no obvious association between AFLP and SSR genotypes, nor between genotypic and phenotypic traits.