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In many species sperm competition selects for large ejaculates while females eliminate or disable a high percentage of the spermatozoa in their storage organs in order to control paternity. Therefore, sperm viability is an excellent measure of the reproductive success of both males and females. Here we assess the viability of spermatozoa in males, freshly mated queens and old queens at the end of a colony cycle of the monandrous bumblebee Bombus terrestris. We found that the accessory testes of males contained a significantly lower percentage of dead sperm (i.e. higher average viability) than the spermathecae of both freshly mated and old queens. In each case, however, the percentage of dead spermatozoa was very small. No differences could be detected between sperm viability in freshly mated and old queens. To test for the possible incompatibility of the sperm and the environment provided by female spermathecae males and females either from different geographic regions (north and south of the Alps) or the same region were mated. We did not find any differences between matings of individuals from within or between regions. The mechanism that causes the lower sperm viability in females remains unknown.