The escalation of antimicrobial resistance (AMR) in uropathogens presents an urgent clinical and public health challenge for contemporary healthcare networks. In the present study, an evaluation of 3434 urine samples revealed an overall culture positivity rate of 24.9% (n = 857), a metric that aligns seamlessly with recent baseline surveillance trends reported across the Indian subcontinent, where positivity rates typically range between 20% and 30%. Notable differences in culture positivity were observed based on patient care setting, with inpatient department samples demonstrating significantly higher yields (70.1%) than outpatient department samples (29.9%). As previously documented, this discrepancy is heavily driven by clinical factors specific to hospital environments, such as severe underlying illness, prolonged hospitalization, indwelling urinary catheterization, and repetitive exposure to broad-spectrum empirical antimicrobial therapy. The GNB predominated, accounting for 77.4% of all culture-positive isolates, whereas GPC accounted for the remaining 22.6%. This distribution confirmed that GNB remain the primary therapeutic target in urological infectious pathologies, a finding consistently validated by international and regional epidemiological data.¹²

To ensure high diagnostic accuracy, organism identification and AST were performed using a BD Phoenix™ Automated System in accordance with the CLSI guidelines. Automated platforms offer critical advantages, including rapid species identification, highly standardized phenotypic susceptibility metrics, reduced observer variation, and significantly improved detection of complex multidrug-resistant (MDR) phenotypes. Such methodological frameworks have become benchmarks for modern AMR surveillance networks.^23,24

A significant finding of this study was the high prevalence of ESBL-producing Enterobacterales, which comprised 492 isolates annually and were heavily dominated by E. coli (63.4%, n = 312) and Klebsiella spp. (24.4%,
n = 120). This dense cluster of resistance severely compromises the clinical use of third-generation cephalosporins. Furthermore, it is frequently linked to cross-resistance against other critical drug classes, such as fluoroquinolones and aminoglycosides, owing to the co-selection of resistance determinants on shared mobile genetic elements.^25,26 Aththanayaka et al.¹⁶ previously characterized this therapeutic challenge, documenting similarly high resistance to cephalosporins and fluoroquinolones among ESBL-producing Enterobacterales causing UTIs.

Evaluation of specific therapeutic options for these ESBL-producing isolates revealed that oral nitrofurantoin retained a high susceptibility rate of 72.0% (354/492). This key finding indicates its continued utility as a reliable, targeted oral agent for uncomplicated lower UTIs and provides clinicians with an effective strategy to avoid the indiscriminate use of intravenous carbapenems. This observation is directly supported by prospective regional studies in which nitrofurantoin demonstrated sustained, robust in vitro activity against ESBL-producing E. coli urinary isolates.^27,28 Similarly, amikacin demonstrated comparatively satisfactory activity in the ESBL cohort in our study, maintaining a susceptibility rate of 68.1% (335/492). This profile matches the genomic and phenotypic surveillance data, showing that aminoglycosides are significantly more effective against multidrug-resistant uropathogenic E. coli than cephalosporins and fluoroquinolones.^29,30 In contrast, empirical monotherapy with conventional agents has become unviable, as evidenced by the suboptimal susceptibility rates to ciprofloxacin (22.0%) and ceftriaxone (5.1%).

However, the actual clinical emergency was underscored by the emergence of 171 carbapenem-resistant isolates. This cohort was dominated by E. coli (47.4%, n = 81) and Klebsiella spp. (28.1%, n = 48), whereas P. aeruginosa (14.0%, n = 24) emerged as a notable non-ESBL threat. As carbapenems are traditionally classified as reserve drugs for severe MDR Gram-negative infections, this expanding resistance network reflects a severe healthcare-associated burden in tertiary care environments, a trajectory previously forecasted by national and regional surveillance reports.^10,11

However, the susceptibility profiles of these carbapenem-resistant isolates are limited. Susceptibility to ceftriaxone plummeted to an absolute 0.0% (0/171), and ciprofloxacin tracking dropped to 9.9% (17/171), demonstrating an extensive, near-complete co-resistance phenotype. This severe therapeutic limitation mirrors contemporary findings reporting alarmingly high concurrent resistance to baseline beta-lactams and fluoroquinolones among carbapenem-resistant E. coli and Klebsiella pneumoniae urinary isolates.^31-33

In our study cohort, analysis of resistance mechanisms among urinary isolates demonstrated pronounced variations in beta-lactamase production between species. Among Escherichia coli isolates, 63.4% were identified as ESBL producers, whereas 47.4% exhibited carbapenemase production. The high rate of ESBL-producing E. coli aligns with established global and regional patterns of uropathogenic resistance, where plasmid-mediated genes rapidly disseminate within healthcare settings.^{16, 26,27,29} However, the significant co-burden of carbapenemase production in nearly half of the E. coli isolates (47.4%) represents a troubling shift toward carbapenem-resistance in routine urinary strains, severely limiting the utility of standard empirical beta-lactam regimens.^11,19,30

In contrast, Klebsiella species in our cohort exhibited a more balanced distribution, with 24.4% producing ESBLs and 28.1% demonstrating carbapenemase activity. While the overall ESBL burden was lower in Klebsiella than in E. coli, the comparable rate of carbapenemase positivity underscores the high capacity of Klebsiella spp. to acquire and express broad-spectrum carbapenemase genes under clinical selection pressure.^17,23,33 This species-specific divergence highlights the distinct evolutionary trajectories of uropathogens under local antibiotic selection and reinforces the imperative for routine epidemiological surveillance and targeted stewardship.^6,24,31

In this highly resistant landscape, colistin retained the highest overall susceptibility of 81.9% (140/171), making it the definitive drug of last resort for treating infections caused by carbapenemase-producing strains. This observation matches the data showing that colistin retains pivotal activity against carbapenemase-producing GNB, despite widespread resistance to all other baseline drug classes.^34-36 Nevertheless, the high dependency on polymyxins raises serious clinical concerns. Not only does it expose patients to significant nephrotoxic and neurotoxic risks, but it also increases the selective pressure driving plasmid-mediated colistin resistance (mcr genes), an emerging threat that has already been documented globally.³⁴

From a molecular epidemiology perspective, the high resistance rates and extensive multiclass co-resistance profiles observed in this study likely reflect intense selective antibiotic pressure within the hospital ecosystem. This pressure drives the horizontal dissemination of dominant resistance genes, such as the metallo-beta-lactamases bla_NDM, K. pneumoniae carbapenemases, and CTX-M-type extended-spectrum enzymes, among Enterobacterales sharing common anatomical niches. Recent molecular epidemiological studies by Xia et al. clearly demonstrated the widespread distribution of these mobile carbapenemase genes among urinary Enterobacterales, highlighting the rapid spread of these multidrug-resistance determinants across both hospital and community interfaces.³⁶

The highly restricted antimicrobial susceptibility profiles established in this study emphasize that the empirical prescriptions for nosocomial uropathogens are no longer sustainable. There is an urgent need to implement continuous institutional antibiogram surveillance, transition to culture-guided therapeutic pathways, rational antimicrobial prescription, and strengthen active antimicrobial stewardship programs to limit the spread of these multidrug-resistant uropathogens.

The present study has certain limitations. Because this was a retrospective laboratory-based study, detailed clinical information, including prior antibiotic exposure, catheterization status, comorbidities, and patient outcomes, could not be assessed. The molecular characterization of ESBL and carbapenemase genes has not yet been performed. Additionally, this study was conducted at a single tertiary care center, which may limit the generalizability of the findings to other healthcare settings.