Evaluation of Saccharomyces cerevisiae strains as probiotic agent with aflatoxin B1 adsorption ability for use in poultry feedstuffs.

In this study the aflatoxin B1 (AFB1) removal capacity, the tolerance to salivary and gastrointestinal conditions, autoaggregation and coaggregation with pathogenic bacteria of Saccharomyces cerevisiae strains isolated from broiler feces, were evaluated. Only four of twelve isolated strains were identified as Saccharomyces cerevisiae using molecular techniques. The results obtained in AFB1 binding studies indicated that the amount of AFB1 removed was both strain and mycotoxin-concentration dependent. Therefore, a theoretical model was applied in order to select the most efficient strain to remove AFB1 in a wide range of mycotoxin concentration. The results indicated that S. cerevisiae 08 and S. cerevisiae 01 strains were the most efficient microorganisms in the mycotoxin removal. Viability on simulated salivary and gastrointestinal conditions was investigated and S. cerevisiae 08 strain showed the best results, achieving 98% of total survival whereas S. cerevisiae 01 reached only 75%. Autoaggregation and coaggregation assays showed S. cerevisiae 08 as the most appropriate strain, mainly because it was the unique strain able to coaggregate with the four bacterial pathogens assayed. Consequently, S. cerevisiae 08 is the best candidate for future in vivo studies useful to prevent aflatoxicosis. Further quantitative in vitro and in vivo studies are required to evaluate the real impact of yeast-binding activity on the bioavailability of AFB1 in poultry. However, this study could be useful in selecting efficient strains in terms of AFB1 binding and provide an important contribution to research into microorganisms with potential probiotic effects on the host.

Analysis of fumonisin B1 removal by microorganisms in co-occurrence with aflatoxin B1 and the nature of the binding process.

The objectives of this investigation were to evaluate the ability of Saccharomyces cerevisiae CECT 1891 and Lactobacillus acidophilus 24 to remove fumonisin B(1) (FB(1)) from liquid medium; to determine the nature of the mechanism involved in FB(1)-microorganism interaction and to analyze whether the presence of aflatoxin B(1) (AFB(1)) interferes with the removal of FB(1) and vice versa. The results obtained indicated that: (i) both microorganisms were able to remove FB(1) from liquid medium; (ii) the removal was a fast and reversible process; (iii) cell viability was not necessary; (iv) the amount of FB(1) removed was both toxin- and microorganism concentration-dependent; (v) the process did not involve chemical modification of FB(1) molecules; and (vi) cell wall structural integrity of the microorganisms was required for FB(1) removal. Consequently, we propose that the mechanism involved in the removal of FB(1) is a physical adsorption (physisorption) of the toxin molecule to cell wall components of the microorganisms. It is highly probable that FB(1) and AFB(1) co-occur in contaminated foods, since the fungal genera Aspergillus and Fusarium frequently occur simultaneously. Therefore, we analyzed whether the presence of AFB(1) interferes with the removal of FB(1) by the microorganisms previously evaluated, and vice versa. Studies of co-occurrence of both mycotoxins clearly showed that they did not compete for binding sites on the microorganism cell wall and the presence of one toxin did not modify the efficiency of the organism in the removal of the other mycotoxin. These findings may be useful for optimization of mycotoxin binding and provide an important contribution to research on microorganisms with ability to remove these secondary metabolites.

Pseudomonas putida A ATCC 12633 oxidizes trimethylamine aerobically via two different pathways.

The present study examined the aerobic metabolism of trimethylamine in Pseudomonas putida A ATCC 12633 grown on tetradecyltrimethylammonium bromide or trimethylamine. In both conditions, the trimethylamine was used as a nitrogen source and also accumulated in the cell, slowing the bacterial growth. Decreased bacterial growth was counteracted by the addition of AlCl(3). Cell-free extracts prepared from cells grown aerobically on tetradecyltrimethylammonium bromide exhibited trimethylamine monooxygenase activity that produced trimethylamine N-oxide and trimethylamine N-oxide demethylase activity that produced dimethylamine. Cell-free extracts from cells grown on trimethylamine exhibited trimethylamine dehydrogenase activity that produced dimethylamine, which was oxidized to methanal and methylamine by dimethylamine dehydrogenase. These results show that this bacterial strain uses two enzymes to initiate the oxidation of trimethylamine in aerobic conditions. The apparent K(m) for trimethylamine was 0.7 mM for trimethylamine monooxygenase and 4.0 mM for trimethylamine dehydrogenase, but both enzymes maintain similar catalytic efficiency (0.5 and 0.4, respectively). Trimethylamine dehydrogenase was inhibited by trimethylamine from 1 mM. Therefore, the accumulation of trimethylamine inside Pseudomonas putida A ATCC 12633 grown on tetradecyltrimethylammonium bromide or trimethylamine may be due to the low catalytic efficiency of trimethylamine monooxygenase and trimethylamine dehydrogenase.

Physical adsorption of aflatoxin B1 by lactic acid bacteria and Saccharomyces cerevisiae: a theoretical model.

The ability of lactic acid bacteria (LAB) and Saccharomyces cerevisiae to remove aflatoxin B1 (AFB1) from liquid medium was tested. The experimental results indicated that (i) AFB1 binding to microorganisms was a rapid process (no more than 1 min); (ii) this binding involved the formation of a reversible complex between the toxin and microorganism surface, without chemical modification of the toxin; (iii) the amount of AFB1 removed was both toxin- and bacteria concentration-dependent; and (iv) quantitatively similar results were obtained with viable and nonviable (heat-treated) bacteria. According to these details, a physical adsorption model is proposed for the binding of AFB1 to LAB and S. cerevisiae, considering that the binding (adsorption) and release (desorption) of AFB1 to and from the site on the surface of the microorganism took place (AFB1 + S <--> S - AFB1). The model permits the estimation of two parameters: the number of binding sites per microorganism (M) and the reaction equilibrium constant (K(eq)) involved, both of which are useful for estimating the adsorption efficiency (M x K(eq)) of a particular microorganism. Application of the model to experimental data suggests that different microorganisms have similar K(eq) values and that the differences in toxin removal efficiency are mainly due to differences in M values. The most important application of the proposed model is the capacity to select the most efficient microorganism to remove AFB1. Furthermore, it allows us to know if a modification of the adsorption efficiency obtained by physical, chemical, or genetic treatments on the microorganism is a consequence of changes in M, K(eq), or both.