Bacterial antibiotic resistance (AR) has become a critical global health threat. AR is mainly driven by adaptive resistance mutations and the horizontal gene transfer of resistance genes, both of which are enhanced by genome recombination. We have previously discovered that genome recombination-mediated tRNA up-regulation is important for AR especially in the early stages. RecA is the most important genome recombination factor. Therefore, RecA inhibitors should be effective in reducing AR. In this study, we found that BRITE338733 (BR), a RecA inhibitor, can prevent ciprofloxacin (CIP) resistance in subculturing Escherichia coli strain BW25113 in the early stages (up to the 7^th generation). In the presence of BR, the tRNA was decreased, so the bacteria cannot evolve resistance via the tRNA up-regulation-mediated AR mechanism. The RecA expression level was also not increased when treated with BR. Transcriptome sequencing revealed that BR causes a down-regulation of ATPase activity and therefore suppresses the energy state of bacteria. Also, the effective concentrations of BR do not harm human cell viability, indicating its clinical safety. These findings demonstrate that BR effectively delays the emergence of spontaneous antibiotic resistance by targeting RecA-mediated pathways. Our findings shed light on a new strategy to counteract the clinical AR: applying BR with the antibiotics together at the beginning.