Background: Trichostrongylus is an intestinal parasite that is highly prevalent in humans and livestock worldwide. There is limited information about the prevalence and epidemiology of Trichostrongylus species among the infected livestock in Mazandaran Province, northern Iran. This study aimed to identify Trichostrongylus spp. among small ruminants using morphometric and molecular methods. Materials and Methods: Small intestinal organs of sheep and goats, slaughtered in Mazandaran Province, were examined for infectivity with Trichostrongylus parasites. Primary species identification was conducted based on the morphological characterization of the male worms. The internal transcribed spacer (ITS) II regions of the ribosomal DNA of the worm tissues were amplified using the polymerase chain reaction (PCR) assay and then the product was subjected to sequencing. Subsequently, the PCR products of the ITS II region were subjected to digestion by HinfI and DraI restriction enzymes using the PCR-restriction fragment length polymorphism (RFLP). Results: Of 180 samples, 98 (54. 44%) were confirmed positive for Trichostrongylus based on the conventional PCR. The digestion of the PCR products with HinfI and DraI facilitated the identification of three Trichostrongylus species, namely Trichostrongylus colubriformis (35%, 90. 81%), Trichostrongylus axei (4%, 4. 08%), and Trichostrongylus vitrinus (5%, 5. 1%). Both morphometric and RFLP techniques resulted in the differentiation of the three Trichostrongylus species. Conclusion: The present study was the 1st attempt in the last 30 years for the identification of Trichostrongylus species in small ruminants in Mazandaran Province. The findings of this study can be helpful for epidemiological and ecological studies, the establishment of effective control programs, and the management of gastrointestinal parasites in Mazandaran Province.