Nucella lapillus (L.) is a marine gastropod mollusc widely used as a bioindicator of TBT pollution in the North Atlantic coastlines. The species reproductive cycle and the male penis length seasonal/spatial variation were studied at a single site at Aveiro seashore (NW Portugal) between December 2005 and June 2007. The main objective of this work is to assess if the "Relative Penis Size Index" (RPSI)--an important imposex assessment index--varies seasonally and spatially in the same sampling site and how this can affect results obtained in imposex monitoring programmes. Animals able to reproduce were found every month but a seasonal pattern in N. lapillus reproductive cycle was evident. Female gametogenic maturation varied seasonally and a decrease in capsule gland volume and condition factor occurred in late summer/early autumn. The gametogenesis in males did not show a significant seasonal variation as in females but the condition factor, penis length, amount of sperm and prostate volume also diminished in late summer/early autumn. On the other hand, males that were close to egg capsules clusters had larger penises than those far away from clusters. The temporal and spatial male penis length variation introduces a bias on imposex assessment results when using RPSI and the magnitude of the error involved is evaluated for different TBT pollution levels scenarios. We consider that RPSI provides interesting and complementary information that should not be excluded from monitoring programmes, but temporal or spatial comparisons of imposex should be based on other more reliable imposex indices like the VDSI.