Horse gastrointestinal myiasis caused by larvae of Gasterophilus spp. (Diptera, Oestridae) flies has a worldwide distribution and, where present, it is primarily caused by larvae of Gasterophilus intestinalis and Gasterophilus nasalis. Other species, i.e., Gasterophilus inermis, Gasterophilus pecorum and Gasterophilus haemorrhoidalis, present in different or in the same regions of the gastrointestinal tract, were only occasionally reported in very limited areas of eastern European Countries and in central Italy. With the aim to contribute data on the species composition of Gasterophilus and on the seasonal variation of the infection pattern in southern Italy, 152 native horses were necropsied from January to December 2003 and Gasterophilus larvae were collected from the stomach, the small intestine and the rectum of each of them. On the whole, 125 (82.2%) horses were infected by larvae of Gasterophilus spp. and 214 second stage larvae (L2) and 13,342 third stage larvae (L3) collected. Five species of Gasterophilus were identified with the following prevalence: G. intestinalis=95.2%, G. nasalis=44.8%, G. inermis=15.2%, G. pecorum=2.6% and G. haemorrhoidalis=0.8%. L3 were retrieved throughout the observation period with the highest mean burdens from January to August 2003 while the lowest mean was registered from September to November 2003. L2 were collected in February-March 2003 and from September to December 2003. The majority of the animals (n=66, 43.4%) were infected with a single Gasterophilus species, however, 45 animals (29.6%) harboured 2 species, 12 animals (7.9%) 3 species and 2 animals (1.3%) 4 species. The trend of abundance in the L3 of G. intestinalis and G. nasalis, the most represented species, was highly concordant (r=0.5, p<0.001). A retrospective comparison of our results and of other data from four seasons of observation (1983-1986) in central Italy showed that the number of G. inermis, G. pecorum and G. haemorrhoidalis have been decreasing relative to G. intestinalis and G. nasalis which may suggest tendency toward the extinction of the three former species of Gasterophilus.