A newly improved Bacillus calidolactis tube diffusion test and two postscreening test systems-a receptor assay (Charm HVS; Charm Sciences, Inc., Maiden, MA) and a newly developed Bacillus cereus ATCC 11778 mycoides test system-were evaluated for the detection and identification of tetracycline residues using 973 samples of bulk milk taken at random in The Netherlands. All milk samples were assayed with the B. calidolactis tube and the receptor test. The milk samples testing as suspect or positive with one or both of the test systems were analyzed with HPLC (limit of detection, 10 ng/ml) and the recently developed B, cereus test system. The B. calidolactis tube diffusion test detected tetracycline residues >45 ng/ml in milk. With the B. cereus test plate, residues of oxytetracycline and tetracycline of >30 ng/ml milk were detected; for chlortetracycline and doxycycline, the detection limit was 10 ng/ml. Raw milk exhibiting inhibition diameters of <20 mm on the B. cereus test plate fulfilled the European Union criterion for maximum residue level for tetracyclines of <100 ng/ml (including their 4-epimer derivatives). The detection limits of the receptor assay depended on the type of milk used. The scintillation counts that were obtained for control samples of bulk milk were considerably lower than for the milks obtained from Charm Sciences, Inc. or processed using UHT pasteurization. One of 973 milk samples was suspect for tetracycline residues by means of the B. calidolactis tube test as well as by the receptor assay; 8 other samples were also considered to be positive using the receptor assay alone. The presence of tetracycline residues could not be proved for these 9 samples (residue concentration, <10 ng/ml) with HPLC. We concluded that the receptor assay was not reliable to detect tetracycline residues in raw milk at <150 ng/ml. The B, cereus test plate was determined to be an inexpensive, reliable alternative for the receptor assay for detection of tetracycline residues.
