Abstract: This study presents the data regarding species diversity of spider mites (Acari: Tetranychidae) which were collected over four growing seasons at 80 locations in 20 various protected natural areas of Serbia. A total of 21 tetranychid species were found: Bryobia angustisetis, B. graminum, B. kissophila, B. longisetis, B. praetiosa, B. rubrioculus, Tetranycopsis horridus, Amphitetranychus viennensis, Eotetranychus aceri, E. carpini, E. coryli, E. fraxini, E. pruni, E. rubiphilus, E. tiliarium, Neotetranychus rubi, Panonychus citri, P. ulmi, Schizotetranychus garmani, Tetranychus turkestani and T. urticae. Spider mites were found on 59 host plant species belonging to 21 plant families. Seven tetranychid species (B. praetiosa, E. aceri, E. carpini, E. fraxini, E. pruni, E. rubiphilus and P. citri), 37 host plant species and six plant families were new records for Serbia. The records of E. aceri, E. fraxini and E. rubiphilus were the first in the Balkans. Twelve plant species found in this survey were identified for the first time as spider mite hosts.