To investigate the effects of subminimum inhibitory concentrations of cephalosporins on bacterial biofilm formation, the biofilm production of 52 Escherichia (E.) coli strains was examined following treatment with cephalosporin compounds at 1/4 minimum inhibitory concentrations (MICs). Ceftazidime (CAZ) inhibited biofilm formation in seven isolates, while cefoperazone (CFP) enhanced biofilm formation in 18 isolates. Biofilm formation of E. coli E42 was inhibited by CAZ and induced by CFP. Therefore, using reverse transcription‑polymerase chain reaction, the expression of the biofilm‑modulating genes of this isolate was investigated. To monitor the production of the autoinducer of quorum sensing in E. coli, autoinducer‑2 (AI‑2) production was detected by measuring the bioluminescence response of Vibrio harveyi BB170. Antisense oligonucleotides (AS‑ODNs) targeting S‑ribosylhomocysteine lyase (luxS) inhibited the expression of the luxS gene in E. coli. CAZ at 1/4 MIC reduced luxS mRNA levels and the production of AI‑2, whereas CFP at 1/4 MIC had the opposite effect. AS‑ODNs targeting luxS significantly decreased the aforementioned inhibitory effects of CAZ and the induction effects of CFP on E. coli biofilm formation. Therefore, biofilm formation by the E. coli clinical isolate E42 was evoked by CFP but attenuated by CAZ at sub‑MICs, via a luxS/AI‑2‑based quorum sensing system.