Spodoptera littoralis (Boisd.) (Lepidoptera: Noctuidae) is one of the most destructive pests of several vegetables and fruits worldwide. In spite of various control methods, this pest has still continued to cause significant damage. In this study, the culturable bacterial flora of S. littoralis was determined. New isolates from S. littoralis, as well as 12 different Bacillus isolates belong to 5 species that were previously isolated from different pests, were tested on S. littoralis larvae. In total, 9 bacteria were characterized based on their morphological, biochemical, physiological, and molecular characteristics. The bacterial flora of S. littoralis was determined as Flavobacterium sp. (SL1), Klebsiella pneumonia (SL2), Enterobacter sp. (SL3), Enterobacter sp. (SL4), Klebsiella sp. (SL5), Serratia marcescens (SL6), Pseudomonas aeruginosa (SL7), Acinetobacter baumannii (SL8), and Staphylococcus sp. (SL9). The insecticidal activity tests were performed on the third-instar larvae of S. littoralis. SL1 and SL5 from S. littoralis caused the highest mortalities with 67% and 77%, respectively. Among previously isolated Bacillus isolates, Bacillus thuringiensis subsp. kurstaki (MnD) and B. thuringiensis subsp. kurstaki (BnBt) were found to be the most effective, causing 100% mortality within 10 days after treatment. A concentration-response test was conducted with these isolates and it was found that the isolate MnD was more effective than BnBt. Therefore, further bioassay experiments were conducted with the isolate MnD and results were discussed with respect to the biocontrol potential of the bacterial isolates.