Infections due to extended-spectrum β-lactamase- (ESBL-) producing Gram-negative bacteria have led to increased mortality, morbidity, and economic burden worldwide. These bacteria can colonize the healthy intestine of human beings and can disseminate in communities and hospital. This study aimed to investigate the prevalence of fecal carriage of ESBL-producing Escherichia coli and Klebsiella species among health science (HS) and non-health science (NHS) students. This descriptive cross-sectional study was conducted on 104 HS and 104 NHS students in which one stool sample from each student was collected and processed for bacterial culture and sensitivity testing according to standard bacteriological procedures. Each morphotype was identified and characterized phenotypically. The antimicrobial sensitivity profile of bacterial isolates was determined by the Kirby–Bauer disk diffusion technique. ESBL production was tested by combination disk method as recommended by the Clinical and Laboratory Standards Institute. Out of 208 stool samples, E. coli and Klebsiella spp. were recovered from 203 (86.8%) and 31 (13.2%) stool samples, respectively. Among those 234 isolates, 69 were positive for ESBL which included E. coli (n = 66, 95.7%) and Klebsiella spp. (n = 3, 4.3%). Fifty (42.4%) out of 118 isolates from HS students and 19 (16.4%) out of 116 from NHS students were colonized by ESBL-producers. Compared to non-ESBL producers, a higher number of ESBL-producing isolates were resistant to ciprofloxacin (14.5% vs. 1.8%, p