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Several strategies have been in place in food animal production to reduce the unnecessary use of antimicrobial agents. Beyond the monitoring of their use, the evaluation of the effect of these strategies on the occurrence and types of antimicrobial resistance (AMR) associated genes is crucial to untangle the potential emergence and spread of AMR to humans through the food chain. In the present study, the occurrence of these genes was evaluated in commensal Escherichia coli isolated from broiler carcasses "produced without the use of antibiotics" in 3 antibiotic-free (AB-free) farms in Italy in 2019. Sequenced data were analyzed along with publicly available genomes of E. coli collected in Italy from the broiler food chain from previous years (2017-2018). The genetic relationships among all 93 genomes were assessed on de novo assemblies by in silico MLST and SNP calling. Moreover, the resistomes of all genomes were investigated. According to SNP calling, genomes were gathered in three clades. Clade A encompassed, among others, ST117, ST8070, and ST1011 genomes. ST10 belonged to clade B, whereas Clade C included ST58, ST297, ST1101, and ST23 among others. Regarding the occurrence of AMR genes, a statistically significant lower occurrence of these genes in the genomes of this study in comparison to the public genomes was observed considering the whole group of genes as well as genes specifically conferring resistance to aminoglycosides, β-lactams, phenicols, trimethoprim, and lincosamides. Moreover, significant reductions were observed by comparing the whole group of AMR associated mutations, as well as those specifically for fluoroquinolones and fosfomycin resistance. Although the identification of 3° generation cephalosporin resistance associated genes in AB-free E. coli is a concern, this study provides a first indication of the impact of a more prudent use of antimicrobial agents on the occurrence of AMR genes in Italian broiler production chain. More studies are needed in next years on a higher number of genomes to confirm this preliminary observation., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Escherichia coli is a normal inhabitant of the intestinal tract of chickens, but when an imbalance in the gut microbiota occurs, E. coli may overgrow and cause extraintestinal infections. The aims of this study were to assess the distribution and spread of E. coli isolates with specific phylogenetic groups and antimicrobial resistance characters among asymptomatic breeder flocks and their broiler progenies with early symptoms of colibacillosis. Broiler flocks were treated with lincospectin during the first week of life and sampled at one, 21 and 42 days. The majority of the 363 E. coli isolates belonged to phylogenetic group A (53.17%), followed by groups D (23.14%), B1 (19.28%) and B2 (4.41%). In broilers, group A was the most represented in birds of 21 and 42 days of age whereas group B1 was the most represented phylogroup in one-day old chicks. More than 90.00% of the isolates were resistant to one or more antimicrobials. Along the life-time of broilers, no differences were found on the occurrence of resistant isolates except for the number of E. coli with elevated MIC to spectinomycin, which increased significantly after the lincospectin treatment. According to XbaI-macrorestriction analysis, a high genetic diversity among E. coli isolates was underlined. Four antimicrobial resistant E. coli isolates of phylogroups A, B1 and D collected from breeders showed similar PFGE patterns to five isolates collected from the respective broiler progenies suggesting a potential spread of these isolates from breeders to broilers., (Copyright © 2015 Elsevier B.V. All rights reserved.)

The mutant prevention concentration (MPC) is a new concept meant to face the increased prevalence of antibiotic resistance by using antibiotic concentrations able to prevent the selection of single-step resistant mutants. In the present study, the MPCs of ciprofloxacin and enrofloxacin were evaluated against fully susceptible strains of Escherichia coli, Salmonella Typhimurium and Pseudomonas aeruginosa. Additionally, representative single-step mutants arising after exposure to sub-MPC antibiotic concentrations were investigated for molecular basis of their fluoroquinolone resistance phenotypes. MPC value was recorded when more than 10(10) CFU/mL were spread on Muëller Hinton Agar supplemented with different antibiotic concentrations (from 1X to 16X MIC value). MICs of original strains as well as single-step mutants were determined in presence or absence of the Efflux Pump Inhibitor Phe-Arg-beta-naphthylamide (PAbetaN). Moreover point mutations in the QRDR of the gyrA and parC genes were investigated by sequencing. The enrofloxacin MPC values were 4-16-fold higher than ciprofloxacin values. E. coli and S. Typhimurium representative single-step mutants showed reduced susceptibilities associated with point mutations in the QRDR of the gyrA gene or efflux pump system. P. aeruginosa mutants showed resistance phenotypes associated predominantly with efflux pump system activity. According to in vitro MPC data, ciprofloxacin showed a better efficacy than enrofloxacin, in preventing the selection of E. coli, S. Typhimurium and P. aeruginosa single-step mutants. However, in relation to AUC/MPC ratio, the MPC concept can be applied in vivo to ciprofloxacin and enrofloxacin for E. coli and S. Typhimurium but not for P. aeruginosa.

Escherichia coli can harbor a broad repertoire of virulence and antimicrobial resistance (AMR) genes, which can be exchanged across the human gastrointestinal microflora, thus posing a public health risk. In this study, 6 batches of artisanal soft cheese and a 6-month ripened fermented dried sausage were investigated to assess the occurrence, phylogeny, and genomic traits (AMR, virulence, and mobilome) of E. coli. 30 and 3 strains isolated from salami and cheese food chains, respectively, were confirmed as E. coli by whole genome sequencing. The accumulation of single nucleotide polymorphism differences within small clusters of strains encompassing batches or processing stages, combined with high serotype and phylogroup diversity, suggested the occurrence of different contamination phenomena among the facilities. A total of 8 isolates harbored plasmid-mediated resistance genes, including one cheese strain that carried an IncQ1 plasmid carrying AMR determinants to macrolides [mph(B)], sulfonamides (sul1, sul2), trimethoprim (dfrA1), and aminoglycosides [aph(3")-Ib and aph(6)-Id]. A pool of virulence-associated genes in the class of adhesion, colonization, iron uptake, and toxins, putative ColV-positive iron uptake systems sit, iro, or iuc (8 salami and 2 cheese), plasmid-encoded hemolysin operon hlyABCD (one salami), and potential atypical enteropathogenic E. coli (3 salami environment) were reported. Overall, our findings underscore the importance of routine surveillance of E. coli in the artisanal food chain to prevent the dissemination of AMR and virulence. [ABSTRACT FROM AUTHOR]

Simple Summary: Although the use of antimicrobials and the related selection of antimicrobial-resistant (AMR) pathogens are recognized worldwide, limited or no information is available on the effect of the antibiotic treatment on the genetic structure as well as the dissemination and persistence of plasmids carrying AMR genetic determinants. This is of particular interest for bacterial pathogens like Avian Pathogenic Escherica coli (APEC), which have the potential to persist and be transmitted from broilers to humans through the food chain as well as for their AMR genetic determinants, which have the potential to be mobilized and spread. With a genomic approach, results of the present study revealed that during the enrofloxacin treatment of broilers, initial strains of extraintestinal E. coli disappeared, being substituted by new clones. Plasmid-mediated fluoroquinolone resistance did not appear to disseminate or persist among observed genomes, confirming that QRDR mutations rather than plasmids are the main drivers of quinolone resistance. Interestingly, plasmids carrying other AMR genes than quinolone-determinant ones were disseminated and persistent since they were found in different clones both before and after the treatment. The persistence of plasmids without a direct antimicrobial selective pressure, if confirmed with further studies, might give insights on the so-called plasmid paradox. The aim of the present study was to investigate the genetic diversity and antimicrobial resistance (AMR) of E. coli during enrofloxacin therapy in broilers affected by colisepticemia. Three unrelated farms with ongoing colibacillosis outbreaks were sampled at day 1 before treatment and at days 5, 10 and 24 post-treatment. A total of 179 E. coli isolates were collected from extraintestinal organs and submitted to serotyping, PFGE and the minimum inhibitory concentration (MIC) against enrofloxacin. PFGE clusters shifted from 3–6 at D1 to 10–16 at D5, D10 and D24, suggesting an increased population diversity after the treatment. The majority of strains belonged to NT or O78 and to ST117 or ST23. PFGE results were confirmed with SNP calling: no persistent isolates were identified. An increase in resistance to fluoroquinolones in E. coli isolates was observed along the treatment. Resistome analyses revealed qnrB19 and qnrS1 genes along with mutations in the gyrA, parC and parE genes. Interestingly, despite a fluoroquinolone selective pressure, qnr-carrying plasmids did not persist. On the contrary, two conjugative AMR plasmid clusters (AB233 and AA474) harboring AMR genes other than qnr were persistent since they were identified in both D1 and D10 genomes in two farms. Further studies should be performed in order to confirm plasmid persistence not associated (in vivo) to antimicrobial selective pressure. [ABSTRACT FROM AUTHOR]

In the present study, the occurrence of Listeria monocytogenes, Staphylococcus aureus, Salmonella spp. and Escherichia coli VTEC was investigated in two batches of artisanal Italian salami tested in winter and summer. Moreover, enumerations of total bacterial count, lactic acid bacteria and Enterobacteriaceae were performed as well as monitoring of water activity and pH. Samples were taken from raw materials, production process environment, semi-finished product and finished products. The results revealed an overall increase of total bacterial count and lactic acid bacteria during the ripening period, along with a decrease of Enterobacteriaceae, pH and water activity. No significant difference was observed between the two batches. The enterobacterial load appeared to decrease during the maturation period mainly due to a decrease in pH and water activity below the limits that allow the growth of these bacteria. E. coli VTEC, Salmonella spp. or L. monocytogenes were not detected in both winter and summer batches. However, Klebsiella pneumoniae was detected in both summer and winter products. Except for one isolate, no biological hazards were detected in the finished salami, proving the efficacy of the ripening period in controlling the occurrence of microbiological hazard in ripened salami. Further studies are required to assess the virulence potential of the Klebsiella pneumoniae isolates. [ABSTRACT FROM AUTHOR]

Cold plasma is an emerging non-thermal processing technology that could be used for large scale leaf decontamination as an alternative to chlorine washing. In this study the effect of an atmospheric cold plasma apparatus (air DBD, 15 kV) on the safety, antioxidant activity and quality of radicchio (red chicory, Cichorium intybus L.) was investigated after 15 and 30 min of treatment (in afterglow at 70 mm from the discharge, at 22 °C and 60% of RH) and during storage. Escherichia coli O157:H7 inoculated on radicchio leaves was significantly reduced after 15 min cold plasma treatment (−1.35 log MPN/cm 2 ). However, a 30 min plasma treatment was necessary to achieve a significant reduction of Listeria monocytogenes counts (−2.2 log CFU/cm 2 ). Immediately after cold plasma treatment, no significant effects emerged in terms of antioxidant activity assessed by the ABTS and ORAC assay and external appearance of the radicchio leaves. Significant changes between treated and untreated radicchio leaves are quality defects based on the cold plasma treatment. Atmospheric cold plasma appears to be a promising processing technology for the decontamination of leafy vegetables although some criticalities, that emerged during storage, need to be considered in future studies. [ABSTRACT FROM AUTHOR]

The antimicrobial efficacy of dielectric barrier discharge atmospheric gas plasma (DBD) was tested against Listeria monocytogenes and shigatoxin-producing Escherichia coli serogroups O157 and O26. Challenge tests were carried out with samples of cut celery and radicchio leaves inoculated with a mix of five strains of L. monocytogenes or the two strains of E. coli immersed in deionised water. The treatment efficacy was also assessed considering only the contaminated deionised water. For deionised inoculated water alone, a treatment time-dependent strong effect was observed and a pathogens reduction higher than 6 Log CFU/mL was obtained after 40 min of treatment. With the vegetables presence in the liquid medium, the efficacy appeared reduced and related to the treatment time, microorganism, substrate and storage duration (reduction up to 2.5 and 3.7 Log CFU/cm 2 for L. monocytogenes and E. coli , respectively). No significant changes were observed on celery visual attributes, soluble solids content and textural parameters. A significant decrease of the chroma colour parameter during storage was noted in treated radicchio samples respect to control ones. [ABSTRACT FROM AUTHOR]