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Genomic in situ hybridization (GISH) was used for a chromosomal composition study of the later generations of interspecific hybrids between A. cepa L. and A. fistulosum L., which are relatively resistant to downy mildew (peronosporosis). GISH revealed that F2 hybrids, which did not produce seeds, were triploids (2n = 3x = 24) with 24 chromosomes and possessed in their compliments 16 chromosomes of A. fistulosum L. and eight chromosomes of A. cepa L. or eight chromosomes of A. fistulosum L. and 16 chromosomes of A. cepa L. The advanced F5 hybrid, which produced few seeds, was amphidiploid with 32 chromosomes. BC1F5 hybrid was triploid with eight chromosomes of A. fistulosum L. and 16 chromosomes of A. cepa L., which did not produce seeds. BC2 (BC1F5) plant was amphidiploid that possessed 4 recombinant chromosomes and produced few seeds. GISH results point to 2n-gametes formation in macro- and microsporogenesis of the hybrids. The mechanism of 2n-gametes formation and the possibility of apomixes events in the backcrossing progeny are discussed.