Gastrointestinal parasitism in horses can affect all categories with evident clinical signs. Control is made with the regular use of chemical products, which has cause resistance to all classes of drugs [1, 2]. New alternative control strategies are needed and biological control with the fungi Duddingtonia flagrans is an appropriate option. The fungi pass through the intestinal tract of the horse acting on the parasite free-living stages. The chlamydospores need to grow in particular environmental conditions for the hyphae to be able to predate the eggs and larvae [3]. The objective of this study was to evaluate the activity of D. flagrans against nematode parasites in naturally infected horses. The study was performed at the Sao Jose da Serra Stud in Pinhais, Brazil. Twenty Thoroughbred animals were divided based on their faecal egg count (EPG) values in 3 treatment groups and one control (n=5). Chlamydospores of a Brazilian isolate of the D. flagrans were orally administered together with grinded-corn at G1: 250.000, G2: 500.000, and G3: 1.000.000 per kilogram of live weight. The animals were treated daily during 5 days. Faecal samples were collected during the 5 days of treatment and 5 days after treatment for EPG counts, which was performed with a 1:25 concentration, individually, as well as the coproculture. Recovered larvae from coproculture were counted on triplicates from each sample, calculating the average and standard deviation. Statistical analysis was performed using Tukey test with GraphPad Prims 5 at 5% significance (P<0,05). The average EPG counts from all groups had a large (53%) daily variation during the experiment and did not show any statistical difference between groups (data not shown). Cyathostomum sp. was the predominant (above 94%) genus present at the faecal cultures in all groups. The fungi started to have an effect on the L3 stages 4 days after initiating the treatment reducing more than 97% (P<0.01) compared to the Control group. The predatory effect of the hyphae persisted for 4 days after the treatment was interrupted significantly (P<0.01) reducing larvae number (Figure 1). Duddingtonia flagrans showed a significant activity against infecting free-living larvae of Cyathostomins of horses.