Detection of ciprofloxacin residues in cow milk: A novel and rapid optical β-galactosidase-based screening assay

Abstract

Ciprofloxacin (a member of the fluoroquinolone class) is one of the most widely used antibacterial agents for the treatment of bacterial infections in livestock. The improper use of such antibacterial agents could lead to the presence of residues in animal origin foods (including milk) and consequently harmful effects for health of consumers, together with the spread of antibiotic-resistant bacterial strains. Therefore, in order to support good farming practices and to ensure food safety, antimicrobial (in particular fluoroquinolones) residues surveillance through improved monitoring techniques is crucial. However, commercial available kits for the detection of fluoroquinolones residues in food samples are time-consuming and still fail to detect MRL concentrations for fluoroquinolones (e.g. 0.1mg/kg for the sum of enrofloxacin and ciprofloxacin residues). Here a novel and rapid assay for ciprofloxacin residual detection through optical microbiological screening in commercially pasteurized cow's milk samples is described. Escherichia coli ATCC 11303 cell proliferation was optically monitored by measuring endogenous β-gal activity that was determined through colorimetric assay in the presence of a chromogenic β-gal artificial substrate. Optical density of E. coli cell culture (linked to cell proliferation) was positively correlated with endogenous β-gal activity. As the presence of ciprofloxacin residues inhibits the E. coli cell proliferation in tested samples, β-gal levels decreased more in exposed samples than in control. The essential step of β-gal induction (usually obtained by IPTG) was obtained by exploiting the lactose present in the milk. Our findings show a detection of ciprofloxacin residues at 1 MRL concentration after 1h using ONPG as chromogenic β-gal artificial substrate and lactose as β-gal inducer. Compared to previously described methods, this assay proved to be a rapid, proficient and more eco-friendly (i.e. minimizing the use of additional reagents) system that could be potentially employed as screening method for detection ciprofloxacin residues in cow's milk.