Perennial ryegrass is one of the most important food sources in animal production. However, several pests affect this crop, and one of the primary control strategies is the symbiotic relationships between ryegrass endophyte fungi. This fungus produces alkaloids that exhibit toxic activity against arthropods. Furthermore, the effect of fungi may extend to higher trophic levels, including predators (spiders and/or insects), decreasing their abundance and diversity. Given the importance of spiders and insects as predators, whether the symbiotic interaction between perennial ryegrass and endophyte fungus reduces the abundance and diversity of predators pose an important question. To address this question, natural enemies in perennial ryegrass were collected and analyzed over a year, and the percentage of endophyte fungus was evaluated by the presence of hyphae from two ryegrass cultivars, Jumbo (E-) and Alto AR1 (E+). We observed an 80% endophyte infection rate for (E+) and 0% for (E-). Moreover, 222 individual spiders corresponding to 10 families were identified in both perennial ryegrasses, including 209 individuals for (E-) and 13 for (E+). The most abundant spider family was Lycosidae, representing 71.17% of the total spiders. In addition, 65 insects were collected, corresponding to 6 families, with Carabidae being the most abundant. Furthermore, the Simpson index indicated the dominance of the family Lycosidae. Overall, spider and insect abundance and diversity were reduced in (E+), suggesting a negative effect of the endophyte on predator populations.