The spread of antibiotic-resistant bacteria. Analysis of water sources and “ready to eat” salads

Antibiotic resistance is becoming a major concern for human health. Several studies have shown that antibiotic resistant bacteria can be found in the environment, where aquatic ecosystems act as a reservoir as well as food products, such as ready to eat foods, milk, and meat products. This project aims to assess the presence of antibiotic resistant bacteria in water sources in both Norway and Italy, and in ready to eat salads. Between March and June 2022, a total of 7 water sources were sampled. Five of them were sampled in Norway, of these four were sampled in Ås and 1 in Ullesvang municipalities. The 2 water samples from Italy were obtained in two different municipalities of the province of Vicenza (Marostica and Pojana Maggiore). In June, 3 samples of ready to eat salads were sampled in three different Norwegian supermarkets in Ås. From both water and salad samples, a total of 26 different bacteria were isolated (nwater = 20; nsalad = 6) using two different selective media, extended spectrum β-lactamase agar and carbapenem resistant Enterobacteriaceae agar and later identified by 16s rDNA sequencing. The bacteria were tested for the presence of 12 commonly known genes for antibiotic resistance through PCR. None of the samples showed the presence these genes. Sixteen samples (nwater = 14; nsalad = 2) were subjected to Minimum Inhibitory Concentration test for 7 different classes of antibiotic for a total of 9 antibiotics (ampicillin, amoxicillin, penicillin G, cefepime, cefotaxime, meropenem, ciprofloxacin, erythromycin, and tetracycline). Ten samples were found to be multi-drug resistant (nwater = 9; nsalad = 1), 4 of which were part of the Pseudomonas non aeruginosa group, 1 was part of the Acinetobacter group, and 5 were part of the non-specie related group. Four samples were subjected to full genome sequencing using Illumina Miseq. Results show the presence of a β-lactamase (OXA-334) but also novel genes (CpxR and MexB) that code for efflux pumps that can effectively remove various classes of antibiotics including carbapenems. Two samples also show the presence of several novel virulence factors, closely related to the ones found in Pseudomonas aeruginosa.