© 2015 The Authors. Marine sanctuaries are areas where the extraction of biota is not permitted. Although most marine sanctuaries have a positive influence on biotic communities, not all sanctuaries are meeting their conservation objectives. Amidst possible explanations (e.g., size, age and isolation), insufficient enforcement is often speculated to be a key driver of marine sanctuary underperformance. Despite this, there are few studies directly linking quantitative enforcement data to changes in biotic communities within marine sanctuaries. Here, we used an asymmetrical-BACI experimental design from 2006-2012 to test whether new enforcement initiatives enhanced abundances of target fishes and threatened species in an existing large sub-tropical marine sanctuary relative to areas open to fishing. Implementation of the new enforcement initiatives in 2010 was associated with a 201% increase in annual fine rate and a significant increase in target fish and elasmobranch abundance, as well as sightings of a critically-endangered shark, in the marine sanctuary relative to areas open to fishing. Overall, these results demonstrate that strengthening enforcement can have a rapid positive influence on target fish and perhaps threatened species in a subtropical marine sanctuary. From this, we contend that increased enforcement guided by risk-based compliance planning and operations may be a useful first step for improving underperforming marine sanctuaries.