Background/Objectives: The widespread use of antibiotics in animal husbandry has led to an increase in antimicrobial-resistant Escherichia coli, particularly strains producing extended-spectrum β-lactamases (ESBL) and AmpC β-lactamases. This study aimed to isolate and characterize such strains from fecal samples of broiler chickens (n = 71) and slaughtered turkeys (n = 31) in northwestern Romania. Methods: Antimicrobial susceptibility testing and PCR were used to evaluate phenotypic resistance patterns and detect the presence of resistance genes (AmpC, blaZ, and blaTEM). Results: The results showed that 55% of turkey and 61% of broiler isolates were presumptive ESBL/AmpC producers. Among all isolates, 50% were classified as extensively drug-resistant (XDR), 44% as multidrug-resistant (MDR), and only 6% were fully susceptible. Gene detection revealed an overall prevalence of 44.2% for AmpC, 72.7% for blaZ, and 58.1% for blaTEM, yielding a total penetrance of 51.09%. The diagnostic odds ratio (DOR) values, ranging from 0.67 to 81, suggest the efficacy of the antibiotic susceptibility testing method used in detecting the presence of these resistance genes. Conclusion: Overall, these findings highlight a significant burden of antimicrobial-resistant, poultry-associated E. coli strains, warranting stricter antimicrobial stewardship.

Background/Objectives: Multidrug-resistant (MDR) Escherichia coli is a critical One Health challenge, with rising resistance in both humans and animals. The present study aimed to compare antimicrobial resistance (AMR) profiles of E. coli isolates from hospitalized patients and food-producing animals in Satu Mare, a county located in northwestern Romania. Methods: Between 2022–2023, 701 samples were collected, leading to 571 non-duplicate E. coli isolates (420 human, 151 animal). Human strains were recovered from 21 hospital departments and originated from feces, urine, blood, sputum, ear secretions, cerebrospinal fluid, purulent wound secretions, and puncture fluids. Animal isolates were obtained from ceca collected at local slaughterhouses serving farms in north-west Romania, including samples from turkeys, broilers, and pigs. Antimicrobial susceptibility testing was performed against eight antimicrobials (amikacin, ampicillin, cefotaxime, ceftazidime, cefepime, ciprofloxacin, gentamicin, sulfamethoxazole/trimethoprim) using standardized methods. Resistance classification followed international definitions of MDR. Statistical associations between host species and resistance were assessed with chi-square tests. Results: Resistance levels were consistently higher in E. coli strains isolated from animals compared with those from humans (p < 0.05). Among human isolates, resistance to ampicillin (41.9%), ciprofloxacin (41.4%), and sulfamethoxazole/trimethoprim (45.7%) approached, but did not exceed 50%. In contrast, E. coli strains recovered from animals showed markedly higher resistance, exceeding 50% for ampicillin (78.8%), ciprofloxacin (65.6%), and cefotaxime (55.0%). Amikacin retained full activity against all animal isolates, whereas 2.8% of human strains were resistant. Overall, multidrug resistance (MDR) was observed in 70.0% of E. coli isolates from humans and 79.7% from animals, with the highest resistance burden in pig-derived isolates. Conclusions: The study underscores the veterinary sector as a key contributor to the maintenance and spread of MDR E. coli. Even in clinically healthy animals, resistance levels exceeded those observed in human isolates. These findings emphasize the need for coordinated One Health monitoring and stricter antimicrobial use policies in livestock to reduce transmission risks across human and animal populations.