Intra- and interspecific experimental pollinations were made to determine the mating systems and the interspecific crossing potential in individuals from 24 populations of five Pleurothallis species. Pleurothallis johannensis, P. ochreata and P. teres show weak or partial self-incompatibility while P. adamantinensis and P, fabiobarrosii are more strictly self-incompatible. We found no differences in Fruit set between intra- and interpopulation intraspecific crosses, and there was no correlation between Fruit set and genetic variability or inbreeding in these species. All species are interfertile and showed no correlation between genetic similarity and crossing potential. We scored fruits for seed viability and observed a strong inbreeding depression in all populations: however, there was no difference in seed viability among intrapopulation, interpopulation intraspecific and interspecific cross-pollinations. These species are pollinated by flies with a behavioural pattern that facilitates self-pollination. Self-incompatibility and inbreeding depression are apparently important in the maintenance of the unusually high levels of genetic variability found in these species. As generally observed in other orchid species, barriers to hybridization between these Pleurothallis species are at the level of pollination. (C) 2001 Annals of Botany Company.8818999