Isolation of pathogens from clinical cultures and their resistance patterns may be altered by antecedent antibiotic treatment. The objective of this study was to assess the influence of treatment with ceftriaxone versus that with ampicillin-sulbactam on recovery and superinfections with 10 nosocomial pathogens. The study was designed as a historical cohort study, using a propensity score to adjust for confounding by indication and multivariate survival analyses to adjust for other confounding. Two thousand four hundred forty-five patients were treated with ampicillin-sulbactam, and 1,308 were treated with ceftriaxone. The study analyzed two outcomes: (i) recovery of pathogens from clinical cultures and (ii) microbiologically documented infections. Data were obtained from administrative, pharmacy, clinical, and laboratory databases and by chart extraction. Following treatment, new isolation of at least 1 of the 10 target pathogens occurred for 244 patients. After adjustment, more infections occurred in the ampicillin-sulbactam group (hazard ratio [HR], 1.55; P = 0.009). This was observed with all gram-negative rods combined (HR, 3.6; P < 0.001) and with each genus of the family Enterobacteriaceae. No differences in isolation of gram-positive bacteria were evident (P = 0.33). Microbiologically documented superinfections occurred in 172 patients and were less frequent in the ceftriaxone group (3.8% versus 5%; HR, 1.6; P = 0.015). All the Escherichia coli and Klebsiella spp. isolates were susceptible to ceftriaxone, but half were resistant to ampicillin-sulbactam. The prevalence of oxacillin resistance among Staphylococcus aureus isolates was higher in the ceftriaxone group (63% versus 31%; odds ratio, 3.8; P = 0.08). Differences in the rates of superinfections and the likely causative organisms following treatment with ceftriaxone or ampicillin-sulbactam were evident. This may guide clinicians in empirical choices of antibiotics to treat superinfection.