title: Microbial screening for quinolones residues in cow milk by bio-optical method.
abstract: The use of antibiotics on lactating cows should be monitored for the possible risk of milk contamination with residues. Accordingly, Maximum Residue Levels (MRLs) are established by the European Commission to guarantee consumers safety. As pointed out by Dec 2002/657/EC, screening is the first step in the strategy for antibiotic residue control, thus playing a key role in the whole control procedure. However, current routine screening methods applied in milk chain still fail to detect residues of quinolones at concentrations of interest. This paper reports the findings of a new bio-optical method for the screening of quinolones residues in bovine milk, based on E. coli ATCC 11303 growth inhibition. The effect of blank and spiked cow milk samples (aliquots equivalents to 0.8%, v/v) is evaluated in Mueller Hinton Broth (MHb) and MHb enriched with MgSO4 2% (MHb-Mg) inoculated with the test strain at the concentration of 10(4)CFU/mL. The presence of quinolones inhibits the cellular growth in MHb, while this effect is neutralized in MHb-Mg allowing both detection and presumptive identification of quinolones. Growth of the test strain is monitored at 37 °C in a Bioscreen C automated system, and Optical Density (OD) at 600 nm is recorded every 10 min after shaking for 10s. Growth curves (OD vs. time) of E. coli ATCC 11303 are assessed in milk samples, with and without quinolones, and their differences in terms of ΔOD (ΔOD600nm=ODMHb-Mg-ODMHb) are calculated. The presence of quinolones is detected by the cellular growth inhibition (OD vs time, none increase in the value OD) and presumptively identified through the increase of the slope of ΔOD600nm curve (ΔOD vs. time), after about 3h of incubation. The detection limit for ciprofloxacin and enrofloxacin is at the level of MRL, for marbofloxacin is at 2-fold the MRL whereas for danofloxacin is at 4-fold the MRL. Although the sensitivity of the method could be further improved and the procedure automated, it is a promising step forward to integrate screening assays into the control process and, in particular, to fill in the gap for quinolones; moreover, these technological developments contribute to the One Health perspective through the monitoring of safe and correct use of veterinary antibiotics.
authors.affiliation: Department of Veterinary Public Health and Food Safety, Istituto Superiore di Sanità, Viale Regina Elena 299, 00161 Roma, Italy.
authors.forename: Brunella
authors.initials: B
authors.lastname: Appicciafuoco
authors.affiliation: Institute of Nanostructured Materials, Consiglio Nazionale delle Ricerche, c/o Department of Chemistry, 'Sapienza' University of Rome, Piazzale Aldo Moro 5, 00186 Roma, Italy. Electronic address: roberto.dragone@uniroma1.it.
authors.forename: Roberto
authors.initials: R
authors.lastname: Dragone
authors.affiliation: External Relations Office, Istituto Superiore di Sanità, Viale Regina Elena 299, 00161 Roma, Italy.
authors.forename: Chiara
authors.initials: C
authors.lastname: Frazzoli
authors.affiliation: National Reference Centre for Bovine Milk Quality, Istituto Zooprofilattico Sperimentale della Lombardia e dell'Emilia Romagna, Via Bianchi 9, 25124 Brescia, Italy.
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Bolzoni
authors.affiliation: Department of Veterinary Public Health and Food Safety, Istituto Superiore di Sanità, Viale Regina Elena 299, 00161 Roma, Italy.
authors.forename: Alberto
authors.initials: A
authors.lastname: Mantovani
authors.affiliation: Department of Veterinary Public Health and Food Safety, Istituto Superiore di Sanità, Viale Regina Elena 299, 00161 Roma, Italy.
authors.forename: Anna Maria
authors.initials: AM
authors.lastname: Ferrini
mesh_terms: D000818:Animals; D000900:Anti-Bacterial Agents / Q000032:analysis*; D001681:Biological Assay / Q000379:methods; D002417:Cattle; D004350:Drug Residues / Q000032:analysis*; D004926:Escherichia coli / Q000187:drug effects; D057230:Limit of Detection; D008892:Milk / Q000737:chemistry*; D015363:Quinolones / Q000032:analysis*; D013997:Time Factors
chemical_list: D000900:Anti-Bacterial Agents; D015363:Quinolones
keywords: Farm animals; Food safety; Milk; One Health; Quinolones; Veterinary drugs