Multidrug-resistant (MDR) Escherichia coli is a major foodborne pathogen posing a critical threat to public health, particularly through the contamination of animal products. The increasing prevalence and virulence of MDR E. coli strains underscore the urgent need for alternative antimicrobial strategies. This study aimed to synthesize and characterize zinc oxide nanoparticles (ZnO-NPs) using Stevia rebaudiana as a sustainable capping and reducing agent, aligning with green chemistry principles. Of the 120 fish samples, 74.2% (89/120) were positive for E. coli contamination. Among the identified E. coli strains, 77.8% (119/153) were classified as MDR. Resistance profiling revealed 22 distinct patterns, and seven highly resistant and virulent strains were selected for further analyses. The eco-friendly auto-combustion synthesis of ZnO-NPs produced nanoparticles with semi-spherical to hexagonal shapes and an average size ranging from 12 to 25 nm. Scanning Electron Microscope–Energy Dispersive X-ray analysis (SEM-EDS) confirms that ZnO-NPs primarily consist of zinc (37.5%) and oxygen (19.9%), with carbon (42.6%) indicating the green synthesis process. ZnO-NPs demonstrated potent, dose-dependent antibacterial and antibiofilm activity against the selected MDR E. coli strains. Additionally, mechanistic studies revealed that ZnO-NPs disrupt bacterial cell membranes, alter cellular morphology, and interfere with DNA integrity. These findings highlight the potential of eco-friendly ZnO-NPs as a promising nanomaterial for enhancing food safety and addressing the growing challenge of MDR foodborne bacteria.

Background: The emergence of multiple antibiotic resistance (MAR) Escherichia coli (E. coli) and virulent non-O157 Shiga toxin-producing Escherichia coli (STEC) poses a growing concern to the meat industry. Non-O157 STEC strains including O26, O45, O103, O111, O121, and O145 have been implicated in the occurrence of bloody diarrhea and hemolytic uremic syndrome in humans. This research assessed prevalence, matrix-assisted laser desorption/ionization-time of flight mass-spectrometry (MALDI-TOF MS) protein mass-spectra profiles, multidrug-resistance traits, polymerase chain reaction detection of virulence, and antibiotic-resistance genes of E. coli isolated from beef carcasses and slaughterhouse environments. Methods: A total of 180 convenience sponge samples were collected from two different sources-specific parts of beef carcasses and surfaces of the processing environment at the slaughterhouse of Ha′il, Saudi Arabia between September and November 2020. MALDI BioTyper and phylotype-based identification methods accurately identified and classified the samples as belonging to the genus belonging to the Escherichia coli domain of bacteria (NCBI txid: 562). Results: Expected changes were seen in the mass peak spectrum defining nine closely related isolates and four unrelated E. coli isolates. Serological typing of E. coli revealed enterotoxigenic E. coli O166 (19.10%); enteropathogenic E. coli O146 (16.36%) and O44 (18.18%); enterohemorrhagic E. coli O111 (31.18%) and O26 (14.54%). Forty-five percent of examined E. coli were resistant to seven antimicrobials; 75% of 20 selected isolates were resistant to three or more antimicrobials. phoA and bla_TEM genes were detected in all selected E. coli isolates. Conclusion: This study confirmed the efficiency and validity of Matrix-assisted laser desorption/ionization time of flight mass-spectrometry in screening for multi-drug resistant E. coli isolated from slaughterhouse derived beef carcasses in Ha’il, Saudi Arabia. We contributed by revealing the distinction between related and non-related strains of E.coli in livestock. The findings in this study can inform improved policy development decision making and resource allocation related to livestock processing regarding antimicrobial use in food animals and rapid screening for effective multiple antibiotic resistance E. coli and virulent non-O157 STEC control in the slaughterhouses.

Listeria monocytogenes is one of the most severe foodborne pathogens found in several habitats. Therefore, this study aims to investigate the antilisterial activity of different essential oils (EOs) against multidrug-resistant (MDR) L. monocytogenes strains isolated from fresh chicken meat. Our results showed that the prevalence of L. monocytogenes in the examined samples was 48%. Seventy-eight isolates were identified as L. monocytogenes. Out of these, 64.1% were categorized as MDR and were categorized in 18 patterns with 50 MDR isolates. One isolate was selected randomly from each pattern to investigate their biofilm-forming ability, resistance, and virulence genes incidence. Out of 18 MDR isolates, 88.9% showed biofilm-forming ability. Moreover, the most prevalent resistance genes were ermB (72%), aadA (67%), penA (61%), and floR genes (61%). However, the most prevalent virulence genes were inlA (94.4%), prfA (88.9%), plcB (83.3%), and actaA (83.3%). The antilisterial activity of EOs showed that cinnamon bark oil (CBO) was the most effective antilisterial agent. CBO activity could be attributed to the bioactivity of cinnamaldehyde which effects cell viability by increasing the bacterial cell electrical conductivity, ion leakage, and salt tolerance capacity loss. Therefore, CBO could be an effective alternative natural agent for food safety applications.

The rise of multidrug-resistant (MDR) bacteria in food products poses a significant threat to public health, necessitating innovative and sustainable antimicrobial solutions. This study investigates the green synthesis of zinc oxide nanoparticles (ZnO-NPs) using Stevia rebaudiana extracts to evaluate their antibacterial and antibiofilm activities against MDR Staphylococcus aureus strains isolated from sold fish samples. The obtained results show that the contamination with S. aureus reached 54.2% in the tested fish samples (n = 120), underscoring the urgent need for effective interventions. ZnO-NPs were successfully synthesized and characterized using UV-visible spectroscopy, FT-IR, XRD, and TEM, confirming their formation with an average size of 15.7 nm and reflecting their suitability for antimicrobial and biological applications. ZnO-NPs exhibited potent antibacterial activity, with a maximum inhibition zone of 24.4 ± 0.4 mm at 20 μg/disk, MIC values of 6.25–25 μg/mL, and MBC values of 12.5–50 μg/mL. Additionally, biofilm formation was inhibited by up to 92.1% at 250 μg/mL. Our mechanistic study confirmed that ZnO-NPs damage bacterial membranes and DNA, leading to the intracellular leakage of cell components that lead to bacterial cell lysis. The use of S. rebaudiana in ZnO-NP synthesis aligns with green chemistry principles, offering an eco-friendly alternative to conventional antibiotics and enhancing the bioactivity of ZnO-NPs, and may address the growing issue of antimicrobial resistance, thereby contributing to improved food safety and public health protection.

A total of 100 rabbits weighed about 1000-1500 grams (free from any antibiotics by feeding the rabbits on a balanced ration free from antibiotics for three weeks) were divided into two groups ; control group (50) and test group (50) which were injected subcutaneously with oxytetracycline hydrochloride 20mg/kg body weight for five successive days. 5 rabbits from each group were slaughtered at zero time, 24hr, 48hr, 72hr, 96hr, 120hr, 144hr, 168hr (1week), 2weeks and 3weeks. Residue depletion of oxytetracycline was determined using high performance liquid chromatography (HPLC). The slaughter time strongly affects the concentration of oxytetracycline residues in animal meats, so that by increasing the slaughter time the oxytetracycline residues decreased. Boiling for 30 minutes caused a great degradation of oxytetracyline residues in rabbit meats but not complete destruction. Freezing at -20º c caused a lower degradation than that caused by boiling. So neither boiling nor freezing could be used as reliable methods to get rid of oxytetracycline residues in rabbit meats. The recovery rates for oxytetracycline in rabbit muscles, kidneys and livers were 102%, 92% and 86%, respectively, at a concentration of 20 µg/kg of spiked samples.