Evaluation of a single plate microbiological growth inhibition assay as a screening test for the presence of antimicrobial agents in compound animal feedingstuffs at therapeutic and contaminating concentrations

Abstract

The Inhibitory Substance Test (IST), a microbiological growth inhibition test, is used for screening animal feedingstuffs for the presence of (contaminating) antimicrobial compounds. The effectiveness of the IST was established for 33 compounds that may be incorporated in feedingstuffs. Minimum detectable concentrations (MDCs) for standard solutions were established and compared with those obtained following solvent extraction of an antimicrobial-free compound feedingstuff spiked with each compound at 0-20 mg/kg. Of the 33 standard solutions examined, the test organism was not sensitive to 11 and the MDC for one was greater than its maximum inclusion rate in complete feedingstuffs. Following routine extraction (25% acetone-phosphate buffer) of feedingstuffs spiked with each of the 22 compounds to which the organism was sensitive, 10 were not detected, 15 were detectable at both minimum and maximum feed-inclusion rates and four were only detectable at their maximum feed-inclusion rates. Extraction with methanol (25%) had a deleterious effect with 12 compounds not detected, nine detectable at both minimum and maximum feed-inclusion rates and five detectable at their maximum feed-inclusion rates. Increasing acetone and methanol concentrations to 40 and 55% respectively resulted in larger inhibitory zones for antibiotic-free feedingstuff (25.3 + 2.43 mm vs 21.1 + 1.02 mm) compared with both 25% acetone (11.3 + 0.22 mm) and 25% methanol (11.2 + 0.22 mm), requiring the establishment of greater threshold zone diameters and negating any advantage in increasing the solvent concentration under these test conditions. It is concluded that the IST may be particularly useful for detection of a number of the zootechnical feed-additives recently banned in the EU, which, if used illegally, may be present at sufficiently high inclusion rates to facilitate detection. Further alteration of extraction conditions may improve the scope of the assay.

Incidence of Antibiotic Residues in Cultured African Catfish C. gariepinus in Selected Zones at Enugu, Nigeria

“…(1994) who said housing of un-medicated pigs in boxes where pigs had previously been treated orally with sulfamethazine resulted in residues in urine, kidney and diaphragm. Kaneene and Miller,[49] and Higgins et al [50], argued that high drug residues can also occur as a result of improper use of a licensed product or through the illegal use of an unlicensed substance or extra-label dosages and use. Residues can also occur in calves fed milk and/or colostrum from cows receiving antimicrobials as suggested by Guest and Paige, [51].…”

Maximum Residue Levels of Veterinary Medicines in Relation to Food Safety: European Community Legislation and Ethical Aspects

“…1) or through the illegal use 6 of an unlicensed substance (Papich et al, 1993;Kaneene and Miller, 1997;Higgins et al, 1999).…”

Collaborative study of a microbiological screening method (three-plate) for the banned antimicrobial growth promotors tylosin, virginiamycin, spiramycin, zinc bacitracin and avoparcin in animal feed

“…The available methods of analysis (adopted in the European Commission) at that time were based on microbiological inhibition tests (European Community 1972, 1981, 1984a, 1984b. These specific methods mainly focused on checking the content of one single compound at the registered additive level, and were therefore not suited as a broad screening method for all substances (Higgins et al 1999). Gafner (1999) described a multi-analyte method using TLC and bio-autography.…”