Abstract
Resistance to antimicrobials is a growing problem of worldwide concern. Plasmids are thought to be major drivers of antibiotic resistance spread. The present work reports a simple way to recover replicative plasmids conferring antibiotic resistance from the bacteria in cheese. Purified plasmid DNA from colonies grown in the presence of tetracycline and erythromycin was introduced into plasmid-free strains of Lactococcus lactis, Lactiplantibacillus plantarum and Lacticaseibacillus casei. Following antibiotic selection, the plasmids from resistant transformants were isolated, analyzed by restriction enzyme digestion, and sequenced. Seven patterns were obtained for the tetracycline-resistant colonies, five from L. lactis, and one each from the lactobacilli strains, as well as a single digestion profile for the erythromycin-resistant transformants obtained in L. lactis. Sequence analysis respectively identified tet(S) and ermB in the tetracycline- and erythromycin-resistance plasmids from L. lactis. No dedicated resistance genes were detected in plasmids conferring tetracycline resistance to L. casei and L. plantarum. The present results highlight the usefulness of the proposed methodology for isolating functional plasmids that confer antibiotic resistance to LAB species, widen our knowledge of antibiotic resistance in the bacteria that inhabit cheese, and emphasize the leading role of plasmids in the spread of resistance genes via the food chain.
Highlights
The wide use of antibiotics in human and veterinary medicine, agriculture and aquaculture has promoted the appearance and spread of resistance to antibiotics, compromising their therapeutic effectiveness [1]
The results provide knowledge on the prevalence and diversity of plasmids conferring tetracycline and erythromycin resistance to cheese bacteria, and provide preliminary insight into the role of plasmids in the spread of antibiotic resistance throughout dairy ecosystems
Counts of resistant aerobic mesophilic bacteria and lactic acid bacteria (LAB) were performed on Plate Count Milk (PCM) and de Man, Rogosa, and Sharpe (MRS) agar plates, respectively, both supplemented with tetracycline or erythromycin, from samples of Cabrales cheese at day 3 (3D; early manufacture) and day 60 (60D; end of ripening)
Summary
The wide use of antibiotics in human and veterinary medicine, agriculture and aquaculture has promoted the appearance and spread of resistance to antibiotics, compromising their therapeutic effectiveness [1]. Antibiotic resistance genes (ARG) can spread via horizontal transfer between bacteria that share the same habitat. The metabolic cost of plasmid replication and maintenance can place an energetic burden on bacterial cells [9], but plasmids can encode traits that confer advantages to the host. These are frequently associated with survival and adaptation to changeable environments, including the ability to metabolize different carbon and nitrogen sources, resistance and tolerance to heavy metals, disinfectants, antibiotics, and other environmental pollutants, and the ability to synthesize antimicrobial agents [10,11,12,13]. The presence of ARG-carrying plasmids, increases the risk of the transfer of resistance [15,16]; the study of such plasmids is crucial if the risk they pose to human health is to be understood, and if strategies to reduce the horizontal transfer of the genes they carry are to be developed [12]
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