Plutella xylostella L. (Lepidoptera: Plutellidae), is an important insect pest of brassica crops. This pest is globally distributed and reported to cause annual losses of US $4-5 billion to the world economy. Continuous and intensive use of many classes of insecticides has led to resistance development in some populations of P. xylostella, particularly in the tropics and subtropics where brassica crops are grown year-round and insect pressure is intense. Long distance migration, its ability to establish in new habitats and elimination of local natural enemies from the agroecosystem due to the intense use of broad-spectrum insecticides are some of the reasons that have contributed to outbreaks of this devastating pest. The development and commercialization of insect-resistant genetically modified (IRGM) crops expressing insecticidal crystal (Cry) proteins from the bacterium Bacillus thuringiensis (Bt) have revolutionized insect pest management in two field crops, cotton and corn. In 2012, these crops were grown on more than 69 million hectares worldwide. Several studies have reported that Bt genes coding for insecticidal Cry toxins, when introduced in brassica crops, confer resistance to P. xylostella. Introduction of Bt brassica crops along with other pest management tactics including cultural, chemical and natural enemies could be a feasible solution to keep P. xylostella under control in the near and long term. However, assessing the environmental safety and compatibility of Bt crops with other control measures, particularly with biological control agents, is imperative. Entomopathogenic nematodes (EPN) in the family Steinernematidae and Heterorhabditidae are important biological control agents for many insect pests. Laboratory and field studies have reported the potential use of EPN for the control of P. xylostella. In the future, if Bt brassica crops are commercialized, EPN applied against P. xylostella may become exposed to Bt proteins directly through root exudates or indirectly by feeding on P. xylostella that developed on Bt crops. The main purpose of this research project was to assess the potential effects of Cry1Ac on Heterorhabditis bacteriophora Poinar (Rhabditida: Heterorhabditidae) under tri-trophic conditions. Using Cry1Ac-resistant P. xylostella larvae as hosts, we evaluated the potential impact of Cry1Ac-expressing broccoli on several fitness parameters of H. bacteriophora. Virulence, reproductive potential, time of emergence and preference of H. bacteriophora for the host (P. xylostella) were not significantly affected when Cry1Ac-resistant P. xylostella larvae were reared on leaves of Cry1Ac or non-Bt broccoli. Also the above-mentioned parameters of the subsequent generation of H. bacteriophora did not differ between nematodes obtained from P. xylostella reared on Cry1Ac broccoli compared to those obtained from P. xylostella reared on non-Bt broccoli.