Of all the recognized public health threats, antimicrobial resistance (AMR) is among the most serious. Antimicrobial resistance may have a higher prevalence in low- and middle-income countries, where antimicrobial misuse is compounded by inadequate infection control measures. In this study, we examined AMR trends in bacterial infections in a reference laboratory in Kathmandu, Nepal, from 2021 to 2024. We collected the clinical records of infected patients, including demographic details, pathogen profiles, and antibiograms, from the electronic laboratory information system and analyzed them using SPSS version 17.0 (IBM Corp., Armonk, NY). Among 3,720 patients, 703 (19.7%; median age: 40 years; 395 [54.0%] female) had infections primarily caused by bacteria (91.7%; 723/731), followed by yeast (3.8%; 28/731). Urinary tract infections (55.9%; 290/703) were the most common. The predominant pathogen was Escherichia coli (41.3%; 290/703). Resistance rates in Enterobacterales, nonfermenters, and Gram-positive cocci were 31.0%, 43.4%, and 13.5%, respectively, for aminoglycosides; 49.7%, 42.8%, and 40.2%, respectively, for fluoroquinolones; 64.9%, 59.0%, and 65.6%, respectively, for cephalosporins; 47.3%, 77.4%, and 27.2%, respectively, for trimethoprim–sulfamethoxazole; and 83.0%, 100.0%, and 85.0%, respectively, for penicillins. Extensively drug-resistant (XDR; 19.1%; 134/703) and multidrug-resistant (29.2%; 205/703) bacteria exhibited >90% resistance to ciprofloxacin, erythromycin, and beta-lactams. Over time (2022–2024), resistance increased for aminoglycosides (27.7–32.3%), fluoroquinolones (43.5–54.1%), and penicillin with beta-lactamase inhibitors (PwBLIs; 35.9–58.7%) but decreased for cephalosporins (67.5–62.8%), carbapenems (23.3–18.7%), penicillins (85.2–82.1%), trimethoprim–sulfamethoxazole (59.6–42.2%), and nitrofurantoin (39.0–3.7%). A low rate of bacterial infections was detected among hospital visitors. Urinary tract infections were the most prevalent type of infection. Multidrug-resistant bacteria predominated over XDR bacteria. Resistance to beta-lactams declined over time; however, resistance to aminoglycosides, fluoroquinolones, and PwBLIs increased.