Background: Pseudomonas aeruginosa, an opportunistic pathogen associated with hospital infections is a potent biofilm forming bacteria. A new class of genes termed the chaperone/usher (cup) encoding fimbrial subunit, chaperone and outer membrane usher protein have been identified which facilitates biofilm formation in Pseudomonas aeruginosa independent of the Type IV pili. Since antibiotics at low concentrations can change biological response in bacteria; therefore in the present study the response of cupA1 and cupA5 genes was studied at a sub-inhibitory concentration of carbapenem to check if it influences biofilm formation. Methods and materials: Four clinical isolates of Pseudomonas aeruginosa harboring blaOXA-48&blaNDM associated with hospital infection were selected for the study. All the isolates had MIC above breakpoint against imipenem and meropenem. The isolates were cultured overnight in fresh Luria Bertani broth with and without 2 μg/mL carbapenem (imipenem, meropenem) stress respectively. Total RNA was isolated from the overnight cultures grown to log phase using RNeasy kit (Qiagen, India); cDNA was prepared using the Quanti Tect Reverse Transcription kit (Qiagen, India) and was then subjected to quantitative real-time PCR to estimate the relative expression of cupA1 and cupA5 genes in the study isolates using the ΔΔCt method. Results: The transcriptional analysis of cupA1 and cupA5 genes in the isolates revealed that the expression levels of the genes in all of the four isolates under normal condition i.e. without any antibiotic pressure were lower than the expression level in control strain Pseudomonas aeruginosa ATCC 278533 (RQ=1). Whereas, overexpression of the genes was observed in all the isolates when these were treated with 2 μg/mL imipenem and meropenem respectively. The expression of cupA5 gene increased multiple folds under meropenem stress in all the isolates. Conclusion: The results of the present study imply that carbapenem at a concentration below the lethal dose increases the expression of cupA1 and cupA5 genes which facilitates biofilm formation. Thus from the present study, it can be concluded that imipenem and meropenem at sub-inhibitory concentration favours the biofilm forming ability in clinical strains of P. aeruginosa establishing infection and thus complicating antibiotic therapies which may further lead to treatment failure.