Mechanism of action of aflatoxin B1 in Bacillus megaterium

Bacillus megaterium cells from various growth phases were equally susceptible to the lethal effects of aflatoxin B1. Known surfactants (EDTA and Tween-80) accentuated the effects of aflatoxin B1. Viability and inulin uptake in aflatoxin B1-exposed cells decreased considerably. The effect was concentration dependent. A straight-line relationship observed in the death curve indicated a single target for aflatoxin B1 action in B. megaterium. Leakage of intracellular constituents in B. megaterium was also concentration dependent, and this can be related to the extent of cell membrane damage.     

Mechanism of action of aflatoxin B1 in Bacillus megaterium.

Bacillus megaterium cells from various growth phases were equally susceptible to the lethal effects of aflatoxin B1. Known surfactants (EDTA and Tween-80) accentuated the effects of aflatoxin B1. Viability and inulin uptake in aflatoxin B1-exposed cells decreased considerably. The effect was concentration dependent. A straight-line relationship observed in the death curve indicated a single target for aflatoxin B1 action in B. megaterium. Leakage of intracellular constituents in B. megaterium was also concentration dependent, and this can be related to the extent of cell membrane damage.     

Transformation of Bacillus megaterium by electroporation

Summary Plasmidic DNA was introduced into intact cells of Bacillus megaterium by electroporation. The procedure showed an efficiency of 10³ transformants g^–1 DNA.     

Conditions for induction of bacteriophage from lysogenic Bacillus megaterium with aflatoxin B1.

The present study was conducted to determine whether or not aflatoxin B1 was an effective inducing agent for lysogenic bacteria and to characterize some of the parameters involved in induction. A lysogenic strain of Bacillus megaterium (NRRL-B-3695) and an indicator strain of this species (NRRL-B-3694) were used. Cultures of the lysogenic strain were incubated for various periods of time in the presence of aflatoxin B1. Plaque-forming units as well as colony-forming units were then determined. Results of the present study indicated that bacteriophage lysogenizing B. megaterium could be induced with aflatoxin B1. The optimum concentration for induction was 25 micrograms of toxin per ml of early-log-phase culture. Evidence suggested that: (i) higher concentrations of aflatoxin B1 formed hydrophobic complexes which would not efficiently induce B. megaterium; (ii) the toxic effect of aflatoxin B1 severely limited the number of cells which could be induced prior to killing action of the toxin; and (iii) concentrations less than 25 micrograms of aflatoxin B1 per ml were not efficient inducers of bacteriophage production nor did they demonstrate the toxic effect observed at higher concentrations.     

Conditions for induction of bacteriophage from lysogenic Bacillus megaterium with aflatoxin B1.

The present study was conducted to determine whether or not aflatoxin B1 was an effective inducing agent for lysogenic bacteria and to characterize some of the parameters involved in induction. A lysogenic strain of Bacillus megaterium (NRRL-B-3695) and an indicator strain of this species (NRRL-B-3694) were used. Cultures of the lysogenic strain were incubated for various periods of time in the presence of aflatoxin B1. Plaque-forming units as well as colony-forming units were then determined. Results of the present study indicated that bacteriophage lysogenizing B. megaterium could be induced with aflatoxin B1. The optimum concentration for induction was 25 micrograms of toxin per ml of early-log-phase culture. Evidence suggested that: (i) higher concentrations of aflatoxin B1 formed hydrophobic complexes which would not efficiently induce B. megaterium; (ii) the toxic effect of aflatoxin B1 severely limited the number of cells which could be induced prior to killing action of the toxin; and (iii) concentrations less than 25 micrograms of aflatoxin B1 per ml were not efficient inducers of bacteriophage production nor did they demonstrate the toxic effect observed at higher concentrations.     

Energy-dependent uptake of urea by Bacillus megaterium

Evidence for the existence of an energy-dependent urea uptake system in Bacillus megaterium DSM 90 was obtained by studying the uptake of 14C-urea. In vivo urea uptake and in vitro urease activity differed significantly with respect to temperature- and pH-dependence, kinetic parameters and response towards metabolic inhibitors. Highest uptake activities were observed during exponential growth, and a rapid decrease in urea uptake occurred when cells entered the stationary growth phase and started to sporulate. Significant differences in the uptake rates were observed during growth with different nitrogen sources, suggesting that the formation of the system is under nitrogen control.     

Uptake of inorganic pyrophosphate by Bacillus megaterium

Cells of Bacillus megaterium take up inorganic pyrophosphate, employing a saturable carrier which is sensitive to sulfhydryl reagents, orthophosphate, and arsenate. Uptake is stimulated by proton ionophores, including CCCP and nigericin, indicating that proton cotransport can lead to an opposing gradient. Inhibitor sensitivity, as well a relatively high Km for inorganic pyrophosphate render it likely that uptake is mediated by an orthophosphate transport system.     

Uptake of aflatoxin B1 and T-2 toxin by two mycotoxin bioassay microorganisms: Kluyveromyces marxianus and Bacillus megaterium

Uptake of aflatoxin B1 (AFB1) and trichothecene T-2 toxin from growth medium by mycotoxin bioassay strains of Klutyveomyces marxianus and Bacillus megaterium was assessed by incubating, washing, and sonicating the cells, extracting samples with chloroform, and analysing the extracts by a combination of high-performance thin-layer chromatography (HPTLC) and fluorescence densitometry. Using cultures of K. marxianus, the entire AFB1 dose was recovered and no AFB1 metabolites were detected. Less than 1% of the AFB1 was recovered from the cells, and AFB1 did not inhibit growth. Methanol in the incubation medium had no significant effect on the levels of AFB1 associated with K. marxianus cells. The entire dose of T-2 toxin was also recovered from K. marxianus cultures, and no metabolites were detected; again, less than 1% of T-2 toxin was cell-associated, but growth was completely inhibited. AFB1 partially inhibited the growth of B. megaterium; approximately 12% of the dose could not be recovered, and no AFB1-related metabolites were detected. Methanol increased the levels of recoverable AFB1 associated with B. megaterium cells. In the case of T-2 toxin, around 8% of the dose was not recovered, and no metabolites were detected; growth of B. megaterium was stimulated. These results suggest irreversible binding of both toxins, or derivatives of them, to the cells of B. megaterium.