Genetic barriers often prevent exploiting diploid cotton germplasm for the improvement of tetraploid cotton. The objective of this study was to investigate mating schemes to achieve introgression of Gossypium sturtianum J.H. Willis and G. australe F. Muell diploid cottons into tetraploid G. hirsutum L. Gossypium hirsutum x G. sturtianum and G. hirsutum x G. australe hexaploids were backcrossed to G. hirsutum to produce BC1 pentaploids and BC2 and BC1S1 euploid and aneuploid plants. The use of G. hirsutum x G. australe pentaploids as male parent in backcrosses with G. hirsutum allowed the production of an important progeny of BC2 self fertile plants that were euploid or carried one chromosome of G. australe in addition to the 52 chromosomes of G. hirsutum. The only two BC2 fertile plants issued from G. hirsutum x G. sturtianum hybrids were monosomic addition materials. Both of them were obtained with the pentaploid as female parent in the backcross to cv. Stam F. These results confirm that cotton male gametes are more limited than female gametes in the number of supernumerary chromosomes they can carry. This provides a means of developing alien monosomic addition lines in the G. hirsutum background from all the diploid species of Gossypium whose chromosomes do not carry genes preventing their individual male transfer. The analysis of the monosomic addition plants produced from the G. hirsutum x G. australe hexaploid with simple sequence repeat (SSR) markers allowed us to distinguish seven lines carrying different single chromosomes of G. australe. These lines constitute valuable materials with which to carry out fundamental and applied genetic investigations.