Wolbachia is a maternally transmitted bacterium that manipulates arthropod and nematode 28 biology in myriad ways. The Wolbachia strain colonizing Drosophila melanogaster creates sperm-29 egg incompatibilities and protects its host against RNA viruses, making it a promising tool for 30 vector control. Despite successful trials using Wolbachia-transfected mosquitoes for Dengue 31 control, knowledge of how Wolbachia and viruses jointly affect insect biology remains limited.32 Using the Drosophila model, transcriptomics and gene expression network analyses revealed 33 pathways with altered expression and splicing due to Wolbachia colonization and virus infection.34 Included are metabolic pathways previously unknown to be important for Wolbachia-host 35 interactions. Additionally, Wolbachia-colonized flies exhibit a dampened transcriptomic response 36 to virus infection, consistent with early blocking of virus replication. Finally, using Drosophila 37 genetics, we show Wolbachia and expression of nucleotide metabolism genes have interactive 38 effects on virus replication. Understanding the mechanisms of pathogen blocking will contribute 39 to the effective development of Wolbachia-mediated vector control programs.40 41 KEYWORDS 42 Pathogen blocking, symbiosis, metabolism, Drosophila melanogaster, arbovirus, Sindbis virus 43 44 45 46 47 48 49 50 51 52 Wolbachia is an alphaproteobacterium that establishes intracellular infections within arthropod 54 and nematode hosts. Wolbachia is famous for inducing reproductive manipulations of arthropods 55 in order to facilitate maternal transmission and spread throughout a population. In many cases, 56 this reproductive manipulation is linked to the ability to protect the same host from secondary 57 infections with pathogens, especially RNA viruses (Lindsey et al., 2018a). The Wolbachia strain 58 infecting Drosophila melanogaster (wMel) both induces sperm-egg incompatibilities (known as 59 cytoplasmic incompatibility, or CI), and blocks pathogens (Teixeira et al., 2008). These 60 phenotypes have made the wMel Wolbachia strain highly desirable for use in vector control 61 programs. Indeed, Aedes aegypti mosquitoes transfected with the wMel Wolbachia strain form 62