Inhibitory Effects of Eurotium cristatum on Growth and Aflatoxin B1 Biosynthesis in Aspergillus flavus

Abstract

Probiotic strain <i>Eurotium cristatum</i> was isolated from Chinese Fuzhuan brick-tea and tested for its <i>in vitro</i> activity against aflatoxigenic <i>Aspergillus flavus</i>. Results indicated that <i>E. cristatum</i> can inhibit the radial growth of <i>A. flavus.</i> Furthermore, this inhibition might be caused by <i>E. cristatum</i> secondary metabolites. The ability of culture filtrate of strain <i>E. cristatum</i> against growth and aflatoxin B₁ production by toxigenic <i>A. flavus</i> was evaluated <i>in vitro</i>. Meanwhile, the influence of filtrate on spore morphology of <i>A. flavus</i> was analyzed by scanning electron microscopy (SEM). Results demonstrated that both radial growth of <i>A. flavus</i> and aflatoxin B₁ production were significantly weakened following increases in the <i>E. cristatum</i> culture filtrate concentration. In addition, SEM showed that the culture filtrate seriously damaged hyphae morphology. Gas chromatography mass spectrometry (GC/MS) analysis of the <i>E. cristatum</i> culture supernatant revealed the presence of multiple antifungal compounds. Real-time quantitative polymerase chain reaction (RT-qPCR) analysis showed that the expression of aflatoxin biosynthesis-related genes (<i>aflD</i>, <i>aflQ</i>, and <i>aflS</i>) were down-regulated. Importantly, this latter occurrence resulted in a reduction of the AflS/AflR ratio. Interestingly, cell-free supernatants of <i>E. cristatum</i> facilitated the effective degradation of aflatoxin B₁. In addition, two degradation products of aflatoxin B₁ lacking the toxic and carcinogenic lactone ring were identified. A toxicity study on the HepG2 cells showed that the degradation compounds were less toxic when compared with AFB₁.