Background: Entomopathogenic nematodes (EPNs) of the families Steinernematidae and Heterorhabditidae that are symbiotically associated with Xenorhabdus and Photorhabdus bacteria are one of the effective biological control agents of insect pests. Native isolates can probably be more efficacious to control insect pests than exotic ones due to their adaptability to indigenous environmental conditions. Results: In this study, Steinernema feltiae isolate FUM221 was recovered from soil samples collected from the Fandoghloo pasture, Ardabil province, Iran. Morphological investigations of the first and second-generation adults, infective juveniles, and molecular characterizations were given based on ITS and 18S rDNA genes. Besides, molecular analysis based on the 16S rRNA region and phenetic data recognized Xenorhabdus bovienii as its symbiont bacterium. The scanning electron microscopy (SEM) images verified the identification of this isolate.Conclusion: The molecular characterization using two loci and phylogenetic analyses provided more evidence for the classification of this steinernematid and its difference of the same species from other countries. Moreover, molecular and phenetic characterizations of its symbiotic bacterium were provided with low variations compared to other isolates. Herein, the comprehensive taxonomic data of this steinernematid from Iran is presented.