The volatile composition of four southern highbush blueberry cultivars ('Primadonna', 'Jewel', 'Snowchaser', and 'FL02-40') grown in two locations (Gainesville and Haines City, FL) and harvested multiple times was investigated. A total of 42 volatiles were identified, including 8 esters, 12 terpenoids, 11 aldehydes, 7 alcohols, and 4 ketones. Twelve of these volatiles are reported for the first time in highbush blueberries, with 10 being positively identified: (Z)-3-hexenal, (E,E)-2,4-hexadienal, (E,Z)-2,6-nonadienal, (E,E)-2,4-nonedienal, methyl 2-methylbutanoate, butyl acetate, 2-methylbutyl acetate, and geranyl acetate. The dominant volatiles were aldehydes followed by terpenoids and esters, with distinct varietal profiles. 'Primadonna' was characterized by a large amount of esters and C-6 aldehydes. In contrast, fewer than 4 esters were found in 'FL02-40' and 'Snowchaser', respectively, but they produced more terpenoids than 'Primadonna' and 'Jewel'. Location and/or harvest date affected the production of volatiles in 'Primadonna', but not so much in the other cultivars.