Optimization of Yeast Bioassay for Trichothecene Mycotoxins

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Abstract:

An improved disc diffusion type bioassay was developed for T-2 toxin (T-2), HT-2 toxin (HT-2), diacetoxyscirpenol (DAS), deoxynivalenol (DON), nivalenol (NIV), neosolaniol (NSL), fusarenon-X (FUS-X), trichothecin (TTN), roridin A (RDA) and verrucarin A (VCA) using the yeast, Kluyveromyces marxianus. Factors such as type of medium, agar volume per plate, pre-incubation time and temperature, incubation temperature, inoculum size and pH had variable, and in some cases a dramatic effect on the sensitivity of the assay. The effect of pH of the assay medium was particularly pronounced. The highest sensitivity was obtained when 6 ml per plate of a tryptic-soy-agar (TSA) medium (pH 7.5) containing 105 CFU inoculum per ml was incubated at 38°C for 18 h. All of the trichothecenes (TNN) were able to inhibit the growth of the yeast with the detection limit being 0.005, 0.01, 0.02, 0.02, 0.1, 0.5, 10, 10, 10 and 50 μg/ disc for VCA, RDA, T-2, TTN, DAS, HT-2, DON, NSL, FUS-X and NIV, respectively. The detection limits for corn and wheat spiked with T-2 were 0.1 and 0.2 μg/g, respectively. Non-TNN mycotoxins that did not inhibit yeast at a concentration of 200 μg/disc were aflatoxin B1 (AFB1), ochratoxin A (OA), citrinin (CT), penicillic acid (PA), cyclopiazonic acid (CPA), penitrem-A (PTA) and zearalenone (ZEE). These results indicate that disc diffusion assay using K. marxianus under optimized conditions provides a sensitive method for the detection of low concentrations of several TNN.

Keywords: BIOASSAY; MYCOTOXINS; OPTIMIZATION; TRICHOTHECENES; YEAST

Document Type: Research Article

Affiliations: 1: Department of Animal Science, University of Manitoba, Winnipeg, Manitoba, Canada R3T 2N2 2: Radiation Application Research Branch, Whiteshell Laboratories, AECL Research, Pinawa, Manitoba, Canada R0E 1L0

Publication date: June 1, 1994

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