This study was designed to evaluate the cross-resistance of Salmonella Typhimurium ATCC 19585 (ST), ciprofloxacin-resistant S. Typhimurium (ST), and clinically isolated S. Typhimurium CCARM 8009 (ST) serially exposed to a half MIC of ceftriaxone (CEF), ciprofloxacin (CIP), polymyxin B (POL), or tetracycline (TET), followed by 1 MIC of CEF, CIP, POL, and TET . The viability changes of ST, ST, and ST exposed to different classes of antibiotics were measured to evaluate the development of cross-resistance . The relative fitness was also estimated to evaluate the reproductive success of ST, ST, and ST consecutively exposed to a half and 1 MIC of CEF, CIP, POL, or TET . The expression levels of efflux pump-related genes (acrA, acrB, mdsA, mdsB, mdtB, and tolC) were evaluated by using a quantitative RT-PCR assay . The MIC of CIP against ST was increased by 128-fold compared to ST. ST was highly resistant to POL (MIC = 8 μg/ml) and TET (MIC = 256 μg/ml). ST exposed to CIP [1/2] was susceptible to POL  than CEF , CIP , and TET , ST exposed to CIP [1/2] was susceptible to CEF  and POL , and ST exposed to CIP [1/2] was susceptible to POL . ST was cross-resistant to CEF [1/2] -CIP , CIP [1/2] -CIP , POL [1/2] -CIP , and TET [1/2] -CIP (Ochoa et al. , 2020) . The efflux pump-related genes (acrA, acrB, mdsA, mdsB, mdtB, and tolC) were overexpressed in ST, ST, and ST exposed to serial antibiotic treatments, which was related to the development of cross-resistance . The antibiotic susceptibility profiles of ST, ST, and ST exposed to serial antibiotic treatments are worth understanding the evolution of antibiotic resistance and developing effective therapeutic regimens against antibiotic-resistant bacteria.