Supporting transitions to sustainable, resilient agri-food systems is important to ensure stable food supply in the face of growing climate extremes. Agroecology, or diversified farming systems based on ecological principles, can contribute to such systems. Based on a qualitative case study of Nicaragua, a forerunner in agroecology, this paper unpacks an ongoing transition to agroecology, focusing on how the transition has been shaped by knowledge flows and intermediary actors. Using a niche development framework based on knowledge processes, we analyze the growth of the agroecological niche in Nicaragua over three phases of niche development. The findings show how knowledge processes’ emphases have shifted over time, as have functions enacted by intermediaries. Dedicated, diversified intermediaries have been key in creating momentum for agroecology, as have individual actors moving between niche and regime. Agency in niche development has come from both niche and regime actors. Finally, we find that Nicaragua’s transition to agroecology has been ambiguous: While the niche has succeeded in changing the mainstream selection environment to its favor in some arenas, transition dynamics lag in others. Drawing lessons from this ambiguity, we suggest entry points for broader systems change, such as market stimulation, value chain development, phase-out policies, and supportive policy in related arenas. We also point out possible actions for niche actors such as integration of financial and commercial actors into niches and creation of dedicated market-focused intermediaries. Our results provide evidence of an ongoing transition and action points for supporting niche development in (sustainable agri-food) transitions around the globe.