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Background: Studies on biting behaviours and infectivity status of insect vectors are pre-requisites in understanding the epidemiology of the vector- borne diseases and planning effective control measures. A longitudinal study was carried out to investigate the transmission index of Simulium damnosum complex species along Osun River, South Western Nigeria. Adult flies were collected on human attractants from 07:00 to 18:00 hours for two consecutive days from February 2008 to June 2009 at three communities: Osun Eleja, Osun Ogbere and Osun Budepo. The infectivity rate was determined by dissection and Polymerase Chain Reaction amplification (PCR) of 0-150 genes of Onchocerca parasite using the pool screening technique. Results: The results indicated that the majority of the flies collected at the three sampling points were nulliparous as they accounted for 53.90%, 57.86% and 59.58% of the flies dissected at Osun Budepo, Osun Ogbere and Osun Eleja, respectively. The parous rate was higher during the dry season than the wet season but the difference was not statistically significant (p < 0.05). The biting activity of the parous flies showed two peaks at Osun Budepo and three peaks at Osun Eleja and Osun Ogbere. Of the 1,472 flies dissected and 1,235 flies screened by molecular method, none was infected with Onchocerca parasite at the three sampling points however the annual biting rates at the three communities were higher than 1,000 considered as tolerable value for a person living in an onchocerciasis zone by Word Health Organization. Conclusion: The study has provided the baseline data for further study on onchocerciasis transmission dynamics and the need to intercept man- simuliid vector contact at the study area. [ABSTRACT FROM AUTHOR]