Abstract: In order to explore the body surface structure and molecular biological classification of three species of the nematodes of the family Strongylidae parasitizing the digestive tract of donkeys, in this experiment, scanning electron microscope was used to observe and measure the specimens of Cyathostomum alveatum, Cyathostomum montgomeryi and Cylicodontophorus bicoronatus from the intestines of Equus asinu from Xinjiang region, China. At the same time, the ITS gene of three nematodes was amplified by PCR, and the homology analysis was carried out after the sequencing of positive products and the phylogenetic tree was constructed. The results showed that the outer leaf crown of Cyathostomum alveatum was slender, which amounted to about 21. The male copulatory bursa was short, with a finger-like appendage on the dorsal lip of the genital cone and a papillomatoid protrusion on the ventral lip; the copulatory spines were linear and hooked at the distal end. The outer leaf crown of Cyathostomum montgomeryi was slender, curled outward, with a total number of 21. The male copulatory bursa was long, with a pair of sac-like appendages on the dorsal lip of the genital cone, one mastoid protrusion on each appendage; the copulatory spines were linear and hooked at the distal end. There was only one thin annular groove at the junction of the oral collar and body of Cylicodontophorus bicoronatus, and the outer leaf crown amounted to about 30; the male reproductive cone was well developed, and the ventral lip was conical and had a mound-like process; on the dorsal lip, there were two symmetrical spiny appendages with thick intersecting spines. The nucleotide similarity between the sequenced ITS gene sequence and other Cylicodontophorus bicoronatus ITS gene sequences in NCBI was 99.88%-100%. The similarity with the ITS gene sequences of other species of nematodes in the subfamily Cyathostominae was 86.46%-96.45%. The similarities between the ITS gene sequences of Cyathostomum alveatum and Cyathostomum montgomeryi and the ITS gene sequences of other species of the nematodes in the subfamily Cyathostominae were 85.38%-96.42% and 85.38%-96.11%, respectively. Cyathostomum alveatum and Cyathostomum montgomeryi were clustered into one branch with Cyathostomum, Cylicostephanus, Coronocyclus nematodes, and Cylicodontophorus bicoronatus and Cylicodontophorus nematodes were clustered into one branch. The results suggested that the body surface structure of three Strongylidae nematodes was accurately observed by scanning electron microscopy, and the phylogenetic analysis results based on ITS gene were consistent with the morphological classification results.