**3. Conclusions**

For mycotoxin binding abilities, broths containing 0.2 µg/mL AFB1 or ST were inoculated with the *Lactobacillus* test strains. Before screening the strains for binding capacities, optimisation of the experiment parameters was carried out. Mycotoxin binding was detectable from a germ count of 10<sup>7</sup> cells/mL at 0.2 µg/mL mycotoxin concentration in MRS broth, so for the screening, a cell concentration of 10<sup>8</sup> cells/mL was chosen. The incubation time of the cells with the mycotoxins was studied from 10 min to 48 h. It was found that 2 days of co-incubation was not required for mycotoxin binding, after 10 min of incubation nearly the same binding values were obtained for the majority of the tested strains, though some anomalies could be observed as for *L. pentosus* TV3 shorter incubation time, while in the case of *L. plantarum* TS23 longer incubation time was slightly more efficient. Based on our experiments, it can be said that neither AFB1 nor ST affected the growth of bacterial strains at the studied concentration.

One hundred and five strains were tested for AFB1 binding; the highest capacities were obtained for *L. pentosus* TV3 with 11.5% and *L. plantarum* AT26, AT3, and AT1 with 8–9%. Interestingly, in the case of ST with a very similar structure, the degree of binding was more than 20%. ST binding ability was examined in 39 *Lactobacillus* strains. *L. plantarum* TV1, AT1, AT3, AT5, *L. paracasei* MA8, and *L. pentosus* TV3 proved to be the strains with the best adsorption abilities. The results found in the literature on the mycotoxin binding abilities of lactobacilli are diverse due to the different methodologies used.

Toxin binding of lactobacilli was measured in the MRS medium, the optimal medium for LAB. The highest mycotoxin binding values found in the literature for lactobacilli were measured in vitro in PBS buffer, 87% for AFB1 by *L. acidophilus* [31], 96% binding was found by Liew and co-workers by *L. casei* Shirota at AFB1 concentration of 2 µg/mL [32], nevertheless, Hernandez-Mendoza et al. showed that the percentage of AFB1 bound by the same species was approximately 30% at AFB1 concentration of 4.6 µg/mL after 4 h of incubation at 37 ◦C [33]. These latter findings underline that even in the same medium the same *Lactobacillus* species might present very different mycotoxin binding abilities in different experiments. Though the most results for AFB1 binding in the literature is measures in PBS buffer, however, MRS medium represents better the possible environment for LAB to be used for mycotoxin binding purposes. Thus, the results of our AFB1 binding assay could not be directly compared to values in the literature.

The same location of AFB1 and ST binding is assumed by our result that the most efficient mycotoxin binding species were representatives of *L. plantarum* and *L. pentosus* species for both mycotoxins (Figures 6 and 8). This is consistent with literature data for AFB1 binding, where these strains are among the most effective within the genus *Lactobacillus* [34].

In our studies, we consistently found that the ST binding potential of *Lactobacillus* strains was approximately twice that of AFB1 binding. This phenomenon may be due to the higher ST affinity of binding-critical cell wall polysaccharide fragments, but this may be explained by the nature of ST in aqueous media: ST in aqueous media may form a unique type of aggregate [35].

An interesting result of our studies is that we also found a large difference in AFB1 and ST binding potential between *Lactobacillus* strains belonging to a given species. This may be explained by the strain-specific, different polysaccharide composition of the WPS fraction of cell surface polysaccharides, as peptidoglucan has too conservative a structure to account for differences between strains [16].

Our work is the first report on microbial ST binding. The investigated LAB type strains had different ST and AFB1 binding abilities. These data, especially the altered binding potential of the *Lactobacillus* strains belonging to the same species, would be very useful in the future for investigating the molecular mechanism of bacterial mycotoxin adsorption and developing aflatoxin bio-binders.
