Introducing an effective antibacterial agent is very important in controlling pathogenic and antibiotic-resistant bacterial strains. Recent advances in nanotechnology, especially the ability to fabricate biological nanoparticles, have led to the development of a wide range of antimicrobial agents. Therefore, the aim of this study is to investigate the antibacterial properties of ZnO nanoparticles (NPs) in inhibiting resistant strains of Acinetobacter baumannii. Ten samples of Acinetobacter were collected from clinical centers of Khorramabad city in 2020 and were identified based on biochemical tests. After treatment of MDR strains with sub-minimum inhibitory concentration (MIC) of ZnO NPs, the expression level of efflux pump genes (AdeA, AdeC) was evaluated using real-time PCR. After applying the effect of ZnO NPs on AdeA gene, 80% of the samples showed reduced expression. 20% of the sample also had incremental expression. Also in AdeC gene, 40% of the samples showed reduced expression after the effect of ZnO NPs. 60% of the samples also had incremental expression. According to the results, the optimal inhibitory effect of biosynthesized ZnO NPs on the expression of AdeA and AdC genes, which play an important role in drug resistance of the Acinetobacter species, was evident. On the other hand, due to the increased expression in 60% of the samples for the AdeC gene, the possibility of mutations in the regulatory genes was raised. It seems that ZnO NPs can be used as a pharmaceutical candidate in the pharmaceutical industry in the future.