Background <p>Climate change, globalization, and urbanization continue to reshape the ecology of vector-borne diseases, allowing mosquito-transmitted arboviruses to expand into previously non-endemic areas. Increasing insecticide resistance, ecological disruption, and declining efficacy of conventional vector control strategies have prompted an interest in microbe-based alternatives. Insect-specific viruses (ISVs), a group of viruses restricted to replicating in arthropods like mosquitoes, have emerged as potential modulators of mosquito vector competence. This systematic review assesses current in vivo evidence on the effects of ISVs on arboviral replication, infection, and transmission, for the purpose of considering their potential application as biocontrol agents for arboviruses.Please confirm if the author names are presented accurately and in the correct sequence (given name, middle name/initial, family name).</p> Methods <p>A systematic search strategy was applied to PubMed, Scopus, ScienceDirect, and Google Scholar to identify relevant studies. The selection process followed PRISMA guidelines and PICO-based inclusion criteria. RAYYAN AI was used to remove duplicates, for abstract and full text screening. Only in vivo studies analyzing interactions between ISVs and medically relevant arboviruses in mosquitoes were retained. Study quality was assessed using the Mixed Methods Appraisal Tool (MMAT), and data extraction followed JBI guidelines.</p> Results <p>A total of 3386 studies were found using the search strategy, of these 15 studies met the inclusion criteria. The effects of ISVs on co-infecting arboviruses were highly variable, depending on mosquito species, ISV strain, route of infection, and the phylogenetic relationship between the ISV and the arbovirus. Cell-fusing agent virus (CFAV), Nhumirim virus (NHUV), Palm Creek virus (PCV), Culex flavivirus (CxFV), Eilat virus (EILV), Espirito Santo virus (ESV), Yichang virus (YCV), Anopheles gambiae densovirus (AgDNV), and Aedes aegypti densovirus (AaeDV) showed suppression in either one or more of the parameters of vector competence used, such as replication, transmission, and infection rate, likely associated with mechanisms consistent with superinfection exclusion or competition for shared cellular resources. In contrast, Phasi Charoen-like virus (PCLV) and Humaita-Tubiacanga virus (HTV) enhanced flavivirus infection and transmission in <i>Aedes aegypti,</i> suggesting that some ISVs may facilitate, rather than inhibit, arbovirus spread. Additionally, in some studies where infection routes differed, neutral or inconsistent interactions were observed.</p> Conclusions <p>ISVs act as context-dependent modulators of arboviral dynamics in mosquitoes. While some indicate biological inhibitory potential, others enhance viral replication or have negligible effects, underlining the importance of meticulous viral selection and ecological risk assessment before considering ISVs as biocontrol candidates. Carefully designed experimental, mechanistic and semi-field evaluations are important for ISVs to be considered as biocontrol agents within integrated vector management programs.</p> Graphical Abstract <p>LE: This title is mismatch with jobsheet title. Please check if it is correct or not and proceed further.we have changed the picture title to match the manuscript title</p>