The Streptomyces genus comprises a large and diverse group of bacteria, many of which are commercially exploited for the production of antibiotics and lytic enzymes. The thermophilic species are less studied than the predominant mesophilic species. However, the first ones are a potential source of thermostable bioactive products and enzymes with novel properties. In this study, two selected thermophilic streptomycetes were identified and their chitinolytic activities were evaluated. The identification of these two isolates was performed by microscopic morphology, partial 16S rDNA sequences, and its phylogenetic analysis. To study the chitinolytic activities of these isolates, the effects of colloidal chitin (CC) and fungal cell walls (FCW) on the chitinase activities and chitinase and chitosanase isoform patterns were determined. Additionally, in vitro confrontations against chitinolytic phytopathogenic fungi were performed at 45 °C and 65 °C. Both isolates (AC4 y AC7) were identified as members of the streptomycete thermophilic clade. The highest chitinolytic activities were observed in the combinations 0.1% FCW/0.1% CC and 0.1% FCW/0.3% CC with maximum values of 0.7 U/μg and 0.45 U/μg, respectively for the AC4 strain, and with values of 0.48 U/μg in both treatments for the AC7 strain. The electrophoretic profiles of chitinase and chitosanase activity showed not only differences in bands intensity, but also few new bands were observed. Both isolates inhibited the growth of Rhizoctonia solani and Fusarium oxysporum. The present study shows that thermophilic streptomycetes have potential bioactivities that might be exploited in horticulture.