Abstract: The Spodoptera exigua multiple nucleopolyhedrovirus (SeMNPV) is a species-specific pathogen that can efficiently control this pest in sweet pepper crop systems of southern Spain. Studies on SeMNPV persistence in covertly infected adults revealed the presence of RNA viruses in insect colonies. These were identified as two iflaviruses (SeIV1 and SeIV2). Subsequent experiments indicated that SeMNPV and SeIV could be detected in both field and laboratory insect populations using PCR-based techniques. In this study, we evaluated the effect of SeIV1 co-infection on the insecticidal characteristics of SeMNPV. For this, bioassays were performed on second instar larvae by inoculating at the same time one of five SeMNPV inoculum concentrations and one of the following iflavirus treatments: i) SeIV1; ii) SeIV2; iii) SeIV1+SeIV2; and mock-infected control. Overall, iflavirus co-inoculation consistently reduced median lethal concentrations (LC50) for SeMNPV compared to larvae infected with SeMNPV alone. However, the speed of kill of SeMNPV was similar in the presence or absence of the iflaviruses. Adults’ survivors of a sublethal SeMNPV treatment were examined for covert infection and SeMNPV DNA was found to be present at a high prevalence in all treatments. In conclusion, simultaneous SeMNPV-SeIV co-infection resulted in improved pathogenicity of SeMNPV and did not alter the capacity of SeMNPV for vertical transmission.