Photorhabdus luminescens is an entomopathogenic rod-shaped bacterium infected with insect nematodes of the Heterorhabditidae family. It kills insects through the secretion of high molecular weight toxin complexes. In this study, Plutella xylostella larvae were orally administered P. luminescens for bioassay. After incubation in Luria-Bertani (LB) medium for a sufficiently long period, the mortality rates of P. xylostella observed after diluting the fermentation broth 50 times and diluting the supernatant 5 times were 18.89% and 91.11%, respectively. Retentates measuring more than 70 kDa showed 88% mortality after ultrafiltration (UF) membrane treatment. Thus, the supernatant of P. luminescens had insecticidal activity, and the main insecticidal toxin complexes had a molecular weight exceeding 70 kDa. The L-9 (3(4)) Taguchi orthogonal experimental optimized medium mode-predicted insecticidal activity levels were 84% and 119% in the 50-fold diluted fermentation broth and 5-fold diluted supernatant, respectively. Moreover, the insecticidal activity was improved to 92.2% in the 100-fold diluted fermentation broth and to 97.8% in the 10-fold diluted supernatant in the experiments. All combinations tested showed clear indications of lethality, including swelling, vesicle formation, cytoplasm vacuolization, and brush border membrane lysis. Thus, these results promote the use of P. luminescens 0805-P2R as a potent biopesticide to effectively control P. xylostella.