Many symbiotic marine invertebrates acquire free-living Symbiodinium from the environment. Abundance and diversity of free-living Symbiodinium could influence recovery from bleaching, resilience, and the long-term adaptation of host organisms. Although free-living Symbiodinium have been detected in the water column and substrates of coral reefs, their diversity and availability to the hosts are poorly understood. Tank experiments were conducted to test whether asymbiotic coral larvae of Acropora monticulosa acquired free-living Symbiodinium from the water column or sediment to become symbiotic. Treatments included filtered (0.22 μm) seawater (FSW), unfiltered seawater (SW), FSW and sediment, and SW and sediment. Our results showed that greater proportions of larvae in sediment-containing treatments acquired Symbiodinium earlier and had greater in hospite Symbiodinium densities when compared to seawater-only treatments. Additionally, clade A Symbiodinium was only recovered in the larvae from the sediment-containing treatments, whereas clades B and C were recovered from all treatments. Differences in distribution, abundance, replication and motility patterns of Symbiodinium, as well as larval behavior, may have contributed to the observed differences between uptake from the sediment and the water column. However, our results suggest that the sediment may represent an important source of free-living Symbiodinium available for uptake during primary acquisition by coral larvae.