My dissertation research has focused on identifying host cellular factors required by the bunyavirus Rift Valley fever virus (RVFV), an RNA virus that causes disease in humans and animals, and the secreted Toxin B (TcdB) from Clostridium difficile, a bacterial pathogen that causes severe antibiotic-associated diarrheal disease. In 2015, the WHO named the ten emerging diseases most likely to cause severe outbreaks in the near future, and three are caused by bunyaviruses (including RVFV). Concern is likewise mounting about the increasing incidence, virulence, and antibiotic-resistance of C. difficile infection worldwide. A better understanding of the molecular details of the pathogenesis of these diseases is urgently needed in order to inform the development and application of therapeutic interventions. The data presented in this thesis summarize the results of two independent screening projects, each utilizing a strategy of forward genetic screening in a mutagenized human haploid cell library. Our RVFV screen identified a suite of enzymes involved in glycosaminoglycan biogenesis and transport, including several components of the cis-oligomeric Golgi (COG) complex. In addition, we identified the gene PTAR1, disruption of which led to RVFV resistance and reduced heparan sulfate surface levels. Biochemical and genetic approaches were utilized to show that both pathogenic and attenuated RVFV strains require GAGs for efficient infection in some cell types, with the block to infection being at the level of virion attachment. Our TcdB screen identified the Wiskott–Aldrich syndrome protein and SCAR homologue (WASH) complex as a host cellular factor supporting TcdB intoxication. Involvement of the WASH complex in TcdB entry was validated by pharmacologic inhibition of recycling endosomes and the use of mouse fibroblasts lacking a functional WASH complex due to genetic ablation of the core WASH1 gene. The host factors supporting TcdB internalization and transport are largely unknown, and our data help to elucidate the mechanism of intoxication of this important and poorly-characterized virulence factor.