Drinking water distribution systems (DWDSs) represent an ideal environment for biofilm formation, which can harbor pathogenic and antimicrobial-resistant bacteria. This study aimed to assess longitudinally the microbial community composition and antimicrobial resistance (AMR), as determined by 16S rRNA NGS and qPCR, respectively, in drinking water (DW) and biofilm from DWDSs, as well as faeces, of free-range organic broiler farms. The role of DWDSs in AMR gene (ARG) dissemination within the farm environment and transmission to animals, was also assessed. DW and biofilm microbial communities differed from those of faecal samples. Moreover, potentially pathogenic and opportunistic bacteria (e.g., Staphylococcaceae) were identified in water and biofilms. High prevalence and abundance of ARGs conferring resistance to carbapenems (i.e., bla_NDM), 3rd and 4th generation cephalosporins (i.e., bla_CMY-2), (fluoro)quinolones (i.e., qnrS), and polymyxins (i.e., mcr-3 and mcr-5) were detected in DW, biofilm, and faecal samples, which is of concern for both animal and human health. Although other factors (e.g., feed, pests, and wildlife) may contribute to the dissemination of AMR in free-range organic poultry farms, this study indicates that DWDSs can also play a role.

Escherichia coli able to produce extended spectrum β-lactamases (ESBLs) and plasmid-mediated AmpC β-lactamases (pAmpCs) represents a serious threat to public health, since these genes confer resistance to critically important antimicrobials (i.e., third generation cephalosporins) and can be transferred to non-resistant bacteria via plasmids. E. coli are known to be able to form a biofilm, which represents a favorable environment for the exchange of resistance determinants. Here, we assessed the ability of 102 ESBL/pAmpC-producing E. coli isolated from the broiler production pyramid to form a biofilm and to identify genetic factors involved in biofilm formation. All but one of the ESBL/pAmpC-producing E. coli were able to form a biofilm, and this represents a great concern to public health. E. coli belonging to phylogroups D, E, and F, as well as strains harboring the bla_CTX-M-type gene, seem to be associated with an increased biofilm capability (p < 0.05). Furthermore, virulence genes involved in adherence and invasion (i.e., csgBAC, csgDEFG, matABCDEF, and sfaX) seem to enhance biofilm formation in E. coli. Efforts should be made to reduce the presence of ESBL/pAmpC- and biofilm-producing E. coli in the broiler production pyramid and, therefore, the risk of dissemination of resistant bacteria and genes.

Aflatoxin B1 (AFB1) induces lipid peroxidation and mortality in bovine foetal hepatocyte-derived cells (BFH12), with underlying transcriptional perturbations associated mainly with cancer, cellular damage, inflammation, bioactivation, and detoxification pathways. In this cell line, curcumin and resveratrol have proven to be effective in mitigating AFB1-induced toxicity. In this paper, we preliminarily assessed the potential anti-AFB1 activity of a natural polyphenol, quercetin (QUE), in BFH12 cells. To this end, we primarily measured QUE cytotoxicity using a WST-1 reagent. Then, we pre-treated the cells with QUE and exposed them to AFB1. The protective role of QUE was evaluated by measuring cytotoxicity, transcriptional changes (RNA-sequencing), lipid peroxidation (malondialdehyde production), and targeted post-transcriptional modifications (NQO1 and CYP3A enzymatic activity). The results demonstrated that QUE, like curcumin and resveratrol, reduced AFB1-induced cytotoxicity and lipid peroxidation and caused larger transcriptional variations than AFB1 alone. Most of the differentially expressed genes were involved in lipid homeostasis, inflammatory and immune processes, and carcinogenesis. As for enzymatic activities, QUE significantly reverted CYP3A variations induced by AFB1, but not those of NQO1. This study provides new knowledge about key molecular mechanisms involved in QUE-mediated protection against AFB1 toxicity and encourages in vivo studies to assess QUE’s bioavailability and beneficial effects on aflatoxicosis.

Among veterinary antibiotics, flumequine (FLU) is still widely used in aquaculture due to its efficacy and cost-effectiveness. Although it was synthesized more than 50 years ago, a complete toxicological framework of possible side effects on non-target species is still far from being achieved. The aim of this research was to investigate the FLU molecular mechanisms in Daphnia magna, a planktonic crustacean recognized as a model species for ecotoxicological studies. Two different FLU concentrations (2.0 mg L⁻¹ and 0.2 mg L⁻¹) were assayed in general accordance with OECD Guideline 211, with some proper adaptations. Exposure to FLU (2.0 mg L⁻¹) caused alteration of phenotypic traits, with a significant reduction in survival rate, body growth, and reproduction. The lower concentration (0.2 mg L⁻¹) did not affect phenotypic traits but modulated gene expression, an effect which was even more evident under the higher exposure level. Indeed, in daphnids exposed to 2.0 mg L⁻¹ FLU, several genes related with growth, development, structural components, and antioxidant response were significantly modulated. To the best of our knowledge, this is the first work showing the impact of FLU on the transcriptome of D. magna.

Background: Short food supply chains are commonly perceived as more sustainable and safer alternatives to conventional production systems, often linked to organic, free-range livestock practices. Materials and methods: This study investigates, for the first time, the distribution of antimicrobial resistance genes (ARGs) and characterizes the microbial communities’ composition, using 16S rRNA sequencing and real-time PCR, respectively. Eleven fecal, 76 slaughterhouse surface, 11 cecal, and 11 carcass samples, from 11 poultry farms belonging to the same short food chain, were analyzed in the study. Results: While cleaning and disinfection procedures appeared to reduce the bacterial load on slaughterhouse surfaces, diverse and potentially resistant bacteria, including genera such as Staphylococcus and Streptococcus, persisted both before and after slaughter. ARGs conferring resistance to high-priority critically important antimicrobials (HPCIAs), such as fluoroquinolones and third-generation cephalosporins, were frequently detected on carcasses, with qnrS (76.15%, 95%CI 68.02-84.28%) and bla_CMY2 (57.8%, 95%CI 48.38-67.22%) being the most prevalent. The slaughtering process emerged as a critical step for ARG dissemination via intestinal bacteria, such as genus Lactobacillus. Additionally, the detection of mcr genes and bla_NDM on carcasses but not in the bird gut samples suggests possible anthropogenic contamination. Discussion: These findings highlight that the evisceration process, slaughterhouse environment, and personnel are all contributing factors in ARG spread and underscore the need for enhanced hygiene protocols and reduced gut ARG carriage in domestic birds to mitigate the risk for the consumer.

The dispersion of antibiotics in livestock farming represents a health concern worldwide, contributing to the spread of antimicrobial-resistant bacteria through animals, the environment, and humans. Phenolic compounds could be alternatives to antibiotics, once drawbacks such as their low water solubility, bioavailability, and reduced stability are overcome. Although nano- or micro-sized formulations could counter these shortcomings, they do not represent cost-effective options. In this study, three phenolic compounds, obtained from wood-processing manufacturers, were characterized, revealing suitable features such as their antioxidant activity, size, and chemical and colloidal stability for in-field applications. The minimum inhibitory concentration (MIC) of these colloidal suspensions was measured against six bacterial strains isolated from livestock. These particles showed different inhibition behaviors: Colloidal chestnut was effective against one of the most threatening antibiotic-resistant pathogens, i.e., S. aureus, but ineffective toward E. coli. Instead, colloidal pine showed a weak effect on S. aureus but specificity toward E. coli. The present proof-of-concept points at colloidal polyphenols as valuable alternatives for antimicrobial substitutes in the livestock context.