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Omono River virus (OmRV), which infects Culex mosquitoes, belongs to the group of dsRNA toti-like viruses. Some members of toti-like viruses have a large negative impact on the fishing industry, which has increased the interest in understanding their propagation cycle. The structural elements of the OmRV such as the obstructed 5-fold pores and the surface crown proteins (CrPs) have been hypothesized to be of importance for viral particle replication as well as the ability of the virus to recognize and enter the host as part of the transmission cycle. In this study, a newly developed method based on fluorescence in situ hybridization (FISH) was used to observe the effect of OmRV pore mutants on viral particle genome transcription. Immobilized-metal affinity chromatography-based pull-down assays and Förster resonance energy transfer (FRET) assays were used to study the interactions between the CrPs and the viral capsid in vitro. The results show that perturbation of the obstructed pore function has a negative impact on viral propagation. One of the mutants, R925A, resulted in a reduction of viral nascent (+)ssRNA synthesis. The findings are to our knowledge the first indication that positively charged arginine residues in the pore structure might be crucial for intraparticle genome synthesis. Specific CrPs-capsid interaction could however not be detected using these in vitro methods, indicating the need for additional optimization to allow for detection of the weak and transient interactions between CrPs and capsid structure. In summary, these studies and a further development of quantitative methods will serve as starting point for and lead to a deeper understanding of the transmission cycle of the dsRNA toti-like virus family, including also the economically and societal important variants.