Comparison of various assays used for detection of beta-lactam antibiotics in poultry meat

In this study, microbiological tests for the detection of beta-lactam antibiotics in meat and meat products were evaluated. The traditional FPT (four plate test, containing Bacillus subtilis and Kocuria rhizophila), BsDA (Bacillus stearothermophilus disc assay) and a newly developed microbiological test, Premi®Test (containing Bacillus stearothermophilus) were included in the study. The limit of detection (LOD) of the Premi®Test was compared with the LOD of the traditional methods. The detection limits of the tests were determined by using beta-lactam antibiotic standards dissolved in meat juice, as well as meat tissue obtained from laying hens after experimental administration of amoxicillin. Positive samples, based on inhibition of growth of the organism in the test, were confirmed by high performance liquid chromatography (HPLC). Growth inhibition in the traditional tests is visible as a clear zone on the plate, whereas for Premi®Test, this is based on the absence of a colour change of the test. The LODs of antibiotics tested were as follows: Penicillin G (PENG) 5 µg kg^-1, amoxicillin (AMOX) 10 µg kg^-1, ampicillin (AMP) 25 µg kg^-1, oxacillin (OXA) 30 µg kg^-1, and cloxacillin (CLOX) 30 µg kg^-1 on the plate with Bacillus stearothermophilus. Beta-lactam antibiotics can be detected also on one plate seeded with Kocuria rhizophila, although the LODs are higher: PENG 10 µg kg^-1, AMOX 25 µg kg^-1, AMP 30 µg kg^-1, OXA 50 µg kg^-1, and CLOX 50 µg kg^-1. Premi®Test was performed according to the Standard Operating Procedure intended for detection of beta-lactam antibiotics in poultry tissues with following LODs: PENG 4 µg kg^-1, AMOX 5 µg kg^-1, AMP 5 µg kg^-1, OXA 40 µg kg^-1, CLOX 50 µg kg^-1. All tests are able to detect beta-lactam antibiotics such as penicillin G, ampicillin, amoxicillin, oxacillin and cloxacillin below the maximum residue level (MRL). However, the detection limits of the Premi®Test for PENG, AMOX and AMP were below the limits of BsDA and the plate containing Kocuria rhizophila.