Isolation of microorganisms and substances inhibitory to aflatoxin production

Abstract

Aflatoxins are toxic and carcinogenic secondary metabolites produced by Aspergillus flavus and A. parasiticus. The contamination of crops, feeds, and foods with aflatoxins can have serious effects on the health of humans and animals. Although many studies have been done to develop aflatoxin-control strategies, most are limited in their effectiveness. As part of an effort to develop control procedures, we have devised simple and safe methods that are useful for identifying microorganisms that effectively inhibit aflatoxin production by fungi. These include the microtitre agar plate assay using norsolorinic acid-accumulating mutant fungi, the ultraviolet light photography method using an instant film, the tip culture method, a convenient RNA extraction method for reverse transcription-polymerase chain reaction (RT-PCR) analysis, and other methods. Results of a recent trial have shown that Achromobacter xylosoxidans significantly inhibited aflatoxin production by A. parsiticus, and that the main inhibitory substance produced by the bacterium was cyclo(L-leucyl-L-prolyl). This result confirms that the methods described herein are useful for identifying microorganisms that inhibit aflatoxin production by fungi and could contribute to the development of methods to reduce aflatoxin contamination in commodities.

Addition of Carbon to the Culture Medium Improves the Detection Efficiency of Aflatoxin Synthetic Fungi

“…Next, we investigated the application of this method to a liquid culture using a pipette tip culture method [34]. After incubation of the Aspergillus strains for seven days, thick colonies were observed on the surface of the potato dextrose (PD) medium (Figure 5).…”

“…A small-scale culture method using a micropipette tip [34] was conducted in this study. Culture medium (295 µL) was dispensed into a 1-mL tip, the end of which was pre-stuffed with glass wool and sealed using parafilm.…”

Predicted Roles of the Uncharacterized Clustered Genes in Aflatoxin Biosynthesis

“…It had been widely assumed that NAA is converted to NA spontaneously, by non-enzymatic O 2 oxidation [22,35,36,37,38,39]. Oxidation of bacterial anthrones is known to be catalyzed by an unusual low molecular weight (less that 20 kDa) monooxygenase that does not require a co-factor [40,41].…”

Improvement of the Culture Medium for the Dichlorvos-Ammonia (DV-AM) Method to Selectively Detect Aflatoxigenic Fungi in Soil

“…The aflatoxigenic fungus FUT-A1 from soil of a FUT flowerbed was identified to be A. nomius . Although the aflatoxigenic fungi have been isolated from agricultural fields in Okinawa area [36], Kyusyu area [46], Kanto area [47,48], and Ibaraki prefecture [34,48,49]. This is the first report that aflatoxigenic fungi are present in Hokuriku area (the Japan Sea side area with a heavy snow in winter), where aflatoxigenic fungi had not been expected meteorologically.…”