Qualifying the T-2 Toxin-Degrading Properties of Seven Microbes with Zebrafish Embryo Microinjection Method

Abstract

T-2 mycotoxin degradation and detoxification efficiency of seven bacterial strains were investigated with zebrafish microinjection method in three steps ((1) determination of mycotoxin toxicity baseline, (2) examination of bacterial metabolites toxicity, (3) identification of degradation products toxicity). Toxicity of T-2 was used as a baseline of toxic effects, bacterial metabolites of strains as control of bacterial toxicity and degradation products of toxin as control of biodegradation were injected into one-cell stage embryos in the same experiment. The results of in vivo tests were checked and supplemented with UHPLC-MS/MS measurement of T-2 concentration of samples. Results showed that the Rhodococcus erythropolis NI1 strain was the only one of the seven tested (R. gordoniae AK38, R. ruber N361, R. coprophilus N774, R. rhodochrous NI2, R. globerulus N58, Gordonia paraffinivorans NZS14), which was appropriated to criteria all aspects (bacterial and degradation metabolites of strains caused lower toxicity effects than T-2, and strains were able to degrade T-2 mycotoxin). Bacterial and degradation metabolites of the NI1 strain caused slight lethal and sublethal effects on zebrafish embryos at 72- and 120-h postinjection. Results demonstrated that the three-step zebrafish microinjection method is well-suited to the determination and classification of different bacterial strains by their mycotoxin degradation and detoxification efficiency.

Evaluation of the Multimycotoxin-Degrading Efficiency of Rhodococcus erythropolis NI1 Strain with the Three-Step Zebrafish Microinjection Method

“…Toxicological effects of Rhodococcus erythropolis NI1 bacterial metabolites, single and combined mycotoxins and their NI1 degradation products were examined with the same injected doses (0.22, 0.52, 1.77, and 4.17 nL). These injected volumes were selected so that the mortality values of toxins and mixtures were interpretable above and below the baseline in every dose [ 53 ]. The initial concentrations of AFB1, ZEN, and T-2 were 1 mg/L in individual solutions, and 1-1 mg/L in combinations (ratio was 1:1), where the initial concentration was in the range where interactions could be plotted (antagonism, additive, and synergism) for simultaneous exposure of mycotoxin mixtures.…”

“…An earlier study had described the concentration-dependent mortality of T-2, which aligned with dose-dependent lethality in this study [ 24 ]. Phenotypic lesions were the same except for the lack of hatching as characterized in previous studies [ 24 , 53 ].…”

“…Degrading enzymes of T-2 mycotoxins are not available. A biotest was not available to examine the effects of T-2 degradation products until the Csenki–Garai three-step method, which is an in vivo test on zebrafish embryos [ 53 , 56 ]. The advantages of this method are that mycotoxins and both types of metabolites (degradation products and bacterial metabolites) can be tested in a complex, synergistic and antagonistic, can be also detected, and the more efficient and safe strains can be selected.…”

“…Previously, the Csenki-Garai three-step method ((1) determination of mycotoxin toxicity baseline, (2) examination of bacterial metabolites toxicity, and (3) identification of degradation products toxicity) was developed, which is a microinjection-based technique in a zebrafish model system, for qualification of degradation and detoxification efficiency of bacteria, and suitable for indirect testing of toxin metabolites [ 53 , 56 ]. During the development of the method, it was demonstrated that the injection volumes alone do not cause mortality or other malformations in the treated embryos.…”

“…The results also showed that the effects observed in the treatments were derived only from the toxin and the normal and degradation metabolites of the microbe [ 56 ]. With the help of the Csenki-Garai three-step method, seven different bacterial strains and T-2 toxin-degrading properties were examined for classifying the strains [ 53 ]. The results confirmed that this microinjection technique may provide an opportunity for the selection of microbial strains that are able to degrade toxins and the identification of the most effective and environmentally safe microbes from the selected strains.…”