Abstract A double-blind, placebo-controlled phase II trial (e-Track 444563-014/ NCT00346892 ) was conducted in South Africa to evaluate the co-administration of RIX4414 (live-attenuated human G1P8 rotavirus vaccine) and oral poliovirus vaccine (OPV) administered simultaneously. Healthy infants ( n = 450) were randomized into three groups (RIX4414 + OPV, RIX4414 + IPV or Placebo + OPV) to receive two oral doses of RIX4414/placebo with OPV or IPV using two vaccination schedules (6–10 weeks and 10–14 weeks). Serum anti-rotavirus IgA antibodies (ELISA) and neutralizing antibodies (micro-neutralization assay) to poliovirus serotypes 1, 2 and 3 were measured. Co-administration of RIX4414 with OPV did not result in a decrease in the high sero-protection rates against poliovirus serotypes 1, 2 and 3 detected after the third OPV dose (98–100%). The anti-rotavirus IgA antibody sero-conversion rates were higher for the 10–14 weeks schedule (55–61%) compared to the 6–10 weeks schedule (36–43%). Solicited symptoms were reported at similar rates between RIX4414 and placebo groups and no serious adverse events related to RIX4414 were reported. This study provided evidence that RIX4414 can be co-administered with routine EPI immunizations including OPV and that two doses of RIX4414 were well tolerated and immunogenic in South African infants.