Hydrogen has emerged as a clean and sustainable carrier of energy, capable of searching for solutions for the ongoing world energy and environmental concerns. The production of hydrogen by microorganisms is seen as a renewable and environment-friendly approach, exploiting the inherent metabolic capabilities of microorganisms. This chapter covers the respective routes of microbial hydrogen production, including fermentative hydrogen generation (both dark fermentation and photofermentation) and biophotolysis (direct and indirect), state-of-the-art hybrid systems exploiting more than one approach. Certain promising microorganisms include purple non-sulfur bacteria, cyanobacteria, green algae, clostridia, and enterobacteria, with contributions of utmost importance toward hydrogen production; different advanced gene modification methods are highlighted, for instance, CRISPR-Cas9, TALENs, ZFNs, RNA interference, and in synthetic biology approaches for optimally tuning microbial production of hydrogen. Metabolic engineering strategies that can be employed to improve hydrogen yields are discussed and include redirection of carbon flow, enhancement of hydrogenase activity, suppression of competing pathways, and enhancement of the tolerance to environmental stresses. Applications of biohydrogen in renewable energy and environmental sustainability cover such aspects as an environmental-friendly fuel, waste-to-energy conversion, and carbon capture and utilization. Having made significant progress in this area, some challenges yet outstanding include scalability, economic feasibility, genetic stability, regulation, and compatibility with existing infrastructures. Going forward, potential avenues include exploring novel microorganisms and pathways, as well as developing integrated systems for large-scale biohydrogen production. The chapter brings the latest knowledge regarding microbial hydrogen production in view of the combination offered toward a sustainable energy future.