The shellfish culture industry in Atlantic Canada has been adversely affected by the presence of non-indigenous, invasive tunicates since the mid-1990’s. A Fisheries and Oceans Canada Aquatic Invasive Species (DFO-AIS) monitoring program documented the presence, establishment, and spread of five tunicate species at geo-referenced coastal monitoring stations in Nova Scotian waters from 2006-2009. Styela clava (Herdman, 1881) and Didemnum vexillum (Kott, 2002) were not found in Nova Scotia during the course of this study, despite their problematic presence in Prince Edward Island and the Gulf of Maine, respectively. Botryllus schlosseri (Pallas, 1766) was the most widely distributed species, found at more than 69% of sites monitored in all years. Ciona intestinalis (Linnaeus, 1767) was present at about half of the stations in all years, and while its populations were heaviest and most persistent in the Halifax – St. Margaret’s Bay, Shelburne – Port La Tour, and Canso – Isle Madame areas, there was evidence of spread on the eastern and Fundy shores, and in Cape Breton. Botrylloides violaceus (Oka, 1927), was the least common tunicate encountered, but its distribution increased from 19% of stations in 2006 to 50% of stations in 2009. Tunicates occupied a wide variety of hard substrates (natural and artificial) in waters with 13.0 to 33.2 salinity and at oxygen saturations of 32.5 to 124.8%.