The development of a drug able to distinguish between tumor and host cells has been long sought, but the solid tumor microenvironment (TME) confounds many current therapeutics. Solid tumors present several challenges for oncotherapeutics, primarily, (1) aberrant vascularization, resulting in hypoxia, necrosis, abnormally high pH, and (2) tumor immune suppression. Oncolytic microbes are drawn to this microenvironment by an innate ability to selectively penetrate, colonize, and eradicate solid tumors as well as reactivate tumor associated immune components. To consider oncolytic bacteria deployment into this microenvironment, Chapter 1 dives into the background of oncolytic microbes. A discussion of the oncolytic bacterial field state, identifying Clostridium novyi¸ as a promising species, and details genetic engineering techniques to develop customized bacteria. Despite the promise of C.novyi in preclinical/clinical trials when administered intratumorally, the genetic and biochemical uniqueness of C.novyi necessitated the development of new methodologies to facilitate more widespread acceptance. Chapter 2 reports the development of methods that facilitate experimental work and therapeutic translation of C.novyi, including the ability to work with this obligate micro-anaerobe aerobically on the benchtop. While methods development is a necessary step in the clinical translation of C.novyi so too is choosing the correct model of the TME within which to test a potential anti-cancer therapy. While the typical solid TME includes both phenotypic and genotypic heterogeneity, the methods used to model this disease state often do not reflect this complexity. This simplistic approach may have contributed to stagnant five-year survival rates over the past four decades. Nevertheless, simplistic models are a necessary first step in clinical translation. Chapter 3 explores the impact of cancer cell lines co-cultured with C. novyi to establish the efficacy of this oncolytic bacteria in a monolayer culture. Chapter 4 extends this analysis adding not only a level of complexity by using an in vivo model, but also using CRISPR/Cas9 to modify the genome of C.novyi to encode a tumor targeting peptide, RGD, for expression within the spore coat. The combination of these studies indicates that C. novyi is uniquely poised to accomplish the long sought after selective tumor localization via intravenous delivery.