The dissemination of resistant pathogens through food supply chains poses a significant public health risk, spanning from farm to fork. This study analyzed the distribution of Shiga toxin-producing Escherichia coli (STEC) across various sources within the animal-based food supply chain. A total of 500 samples were collected from livestock, poultry, the environment, fisheries, and dairy. Standard microbiological procedures were employed to isolate and identify E. coli isolates, which were further confirmed using MALDI-TOF and virulence-associated genes (VAGs) such as stx1, stx2, ompT, hylF, iutA, fimH, and iss. The phenotypic resistance patterns of the isolates were determined using the disc diffusion method, followed by molecular identification of antibiotic resistance genes (ARGs) through PCR. STEC were subjected to PCR-based O typing using specific primers for different O types. Overall, 154 (30.5%) samples were confirmed as E. coli, of which 77 (50%) were multidrug-resistant (MDR) E. coli. Among these, 52 (67.53%) isolates exhibited an array of VAGs, and 21 (40.38%) were confirmed as STEC based on the presence of stx1 and stx2. Additionally, 12 out of 52 (23.07%) isolates were identified as non-O157 STEC co-harbouring mcr-1 and bla_NDM-1. O26 STEC was found to be the most prevalent among the non-O157 types. The results suggest that the detection of STEC in food supply chains may lead to serious health consequences, particularly in developing countries with limited healthcare resources.

Background and Aim: The emergence of multidrug-resistant (MDR) Proteus mirabilis in food-producing animals and their associated environments is a growing public health concern. The indiscriminate use of antimicrobials in animal husbandry exacerbates resistance development, posing significant threats to food safety and sustainability. This study investigates the distribution, antibiotic resistance patterns, and virulence-associated genes (VAGs) of P. mirabilis isolated from poultry, livestock, fish, and their environments in Pakistan under a One Health perspective. Materials and Methods: A total of 225 samples were collected from poultry (n = 100), livestock (n = 75), and aquatic sources (n = 50) from March 2023 to September 2024. Standard microbiological methods were employed for the isolation and identification of P. mirabilis. Polymerase chain reaction (PCR)-based detection of antibiotic resistance genes and VAGs was performed using specific primers. Antibiotic susceptibility was assessed through the disk diffusion method following Clinical and Laboratory Standards Institute 2022 guidelines. Statistical analyses, including analysis of variance and correlation models, were applied to assess the relationships between variables. Results: P. mirabilis was detected in 28.44% (64/225) of the total samples, with the highest occurrence observed in poultry (38%), followed by livestock (22.67%) and aquatic sources (18%). Resistance to ampicillin (100%), chloramphenicol (82%), cefepime (75%), and ciprofloxacin (75%) was widespread. PCR analysis revealed a high occurrence of extended-spectrum beta-lactamase-producing P. mirabilis carrying blaCTX-M (49%), blaOXA (54%), and blaTEM (25.67%) genes. In addition, VAGs such as zapA (39.53%), ucaA (34.88%), and hpmA (32.55%) were frequently identified. The presence of MDR P. mirabilis in fish and related environments (18%) is alarming, highlighting potential zoonotic and foodborne transmission risks. Conclusion: The study underscores the widespread distribution of MDR P. mirabilis in animal-based food sources, raising significant concerns regarding food safety and antimicrobial resistance. The findings reinforce the need for stringent monitoring and regulatory policies to mitigate MDR bacterial dissemination across the food supply chain. Future research should employ metagenomic approaches for comprehensive surveillance and risk assessment. Keywords: animal-derived foods, antimicrobial resistance, multidrug-resistant Proteus mirabilis, One Health, virulence genes.

The indiscriminate use of antibiotics in food-producing animals serves as a major catalyst for the emergence of antibiotic-resistant infections. This study aimed to assess the genetic diversity and antibiotic resistance of Enterobacterales in animal-derived foods. A total of 905 animal-derived food samples, including meat, dairy, poultry, fish, and environmental sources, were collected from various locations in Pakistan. Isolates were confirmed through selective subculturing, morphological, biochemical, and MALDI-TOF analysis, followed by antibiotic susceptibility testing. Subsequently, PCR-based detection of antibiotic resistance genes and virulence-associated genes. Overall, a total of 263 (29.06%) Enterobacterales were identified, as follows: 58.55% (154/263) E. coli, 6.84% (18/263) K. pneumoniae, 21.29% (56/263) P. mirabilis, and 13.30% (35/263) Salmonella spp. Isolates showed a varying resistance pattern against different studied antibiotics, e.g., beta-lactams and inhibitors, ciprofloxacin, and tetracycline, while colistin and tigecycline remained most effective. All the isolates displayed an array of antibiotic resistance and virulence-associated genes. Particularly significant (<0.05) co-existence of bla_NDM and mcr-1 was observed among the Enterobacterales isolated from various animal-derived foods. This study underscores the need to monitor Enterobacterales in animal-derived foods, especially in developing countries, to curb the spread of resistant pathogens and ensure effective food safety measures.

High resistance to antimicrobials is associated with biofilm formation responsible for infectious microbes to withstand severe conditions. Therefore, new alternatives are necessary as biofilm inhibitors to control infections. In this study, the antimicrobial and antibiofilm activities of Fagonia indica extracts were evaluated against MDR clinical isolates. The extract exhibited its antibiofilm effect by altering adherence and disintegration of bacterial cell wall. Fagonia indica has antibacterial effect as minimum inhibitory concentration (MIC) values ranging from 125 to 500 µg mL-1 and minimum bactericidal concentration (MBC) value was 500-3000 µg mL-1 against multidrug resistant (MDR) clinical isolates. The extract exhibited its antibiofilm effect by altering adherence and disintegration of bacterial cell wall. Fagonia indica had antibacterial effect as minimum inhibitory concentration (MIC) values ranging from 125 to 500 µg mL-1 and minimum bactericidal concentration (MBC) value was 500-3000 µg mL-1 against MDR isolates. The maximum inhibitory effects of Fagonia indica chloroform extract on biofilm formation was observed on Staphylococcus aureus (71.84%) followed by Klebsiella pneumoniae (70.83%) after 48 hrs showing that inhibition is also time dependent. Our results about bacterial cell protein leakage indicated that MDR isolates treated with chloroform extract of Fagonia indica showed maximum protein leakage of K. pneumoniae (59.14 µg mL-1) followed by S. aureus (56.7 µg mL-1). Cell attachment assays indicated that chloroform extract resulted in a 43.5-53.5% inhibition of cell adherence to a polystyrene surface. Our results revealed that extracts of Fagonia indica significantly inhibited biofilm formation among MDR clinical isolates, therefore, could be applied as antimicrobial agents and cost effective biofilm inhibitor against these MDR isolates.

Shiga toxin-producing Escherichia coli (STEC) contamination and proliferation in aquaculture and aquatic systems is worrisome for global food safety, as well as veterinary and public health. Traditionally linked with terrestrial ruminant reservoirs, aquaculture matrices, including farmed finfish, shellfish, culture water, and benthic organisms, are increasingly acknowledged as potential conduits for the dissemination of STEC. Herein, the review documented data concerning the prevalence, genomic composition, and ecological dynamics of STEC across various aquaculture environments across different regions of the globe. As a result, the persistence of virulence-associated genes (VAGs) and antibiotic-resistant genes (ARGs) in STEC within the aquaculture supply chain presents a considerable risk to global food security and public health, highlighting the urgent necessity for comprehensive “One Health” surveillance frameworks aimed at alleviating aquatic biosecurity challenges.