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United States Department of Agriculture

Agricultural Research Service

Mycotoxin Research
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Overview

At NPRL we are concerned with mycotoxins, harmful chemicals produced by certain species of fungi. Under certain conditions, these fungi can infect and grow in various crops, including peanuts and corn, contaminating them with mycotoxins.

Moldy peanuts

Because mycotoxin contamination threatens the safety of food and feed, extensive monitoring must be done to ensure that mycotoxins are below levels set by the Food and Drug Administration. The most important mycotoxin that may contaminate peanuts and corn is aflatoxin. Not only is aflatoxin contamination a food safety concern, it is a major economic problem for the peanut and corn industries, which have to pay for the monitoring and take other expensive steps to be sure that products going into the edible market are essentially aflatoxin-free.

 Aflatoxin represented by:

 Aflatoxin displayed on TLC plate

 Aflatoxin displayed on HPLC chromatogram

 TLC

 HPLC

The NPRL has conducted research on aflatoxin contamination of peanuts for many years and has recently developed and patented technology for biological control of aflatoxin contamination. This technology has been licensed and commercialized by Circle One Global, Inc., under the trade name, afla-guard®. Since receiving EPA registration as a biopesticide in 2004, afla-guard has been used in commercial peanut production in the southeastern US where aflatoxin contamination is historically the highest. It is also being used in large-scale tests on corn under an EPA experimental use permit. If those trials prove to be safe and successful, it is expected that the label would be extended to include that crop as well.

Biocontrol agent under peanut plant

 

For more information contact Bruce Horn.

 


Biological Control of Aflatoxin

Aflatoxin contamination of peanuts is a food safety concern and major economic burden for the peanut industry. Aflatoxin contamination occurs when seeds are colonized by the mold, Aspergillus flavus, under late-season conditions of drought and high temperature.
 
A competitive strain of A. flavus that cannot make aflatoxin (nontoxigenic) is applied to crops at mid-season. When drought and high temperatures make them susceptible to infection by A. flavus later in the growing season, the applied strain out-competes the toxin-producing strains naturally present, and the result is reduced levels of aflatoxin.
 

The competitive fungus is bound to a small grain, which serves as (1) a carrier, to get the fungus to the field, and (2) a substrate, for the fungus to grow and multiply on. Hulled barley coated with non-toxigenic spores of A. flavus.
AflaGuard® is applied to peanut about 60 days after planting. The fungus begins to grow and produces a multitude of spores on the surface of the grain. Application of biocontrol
The spores spread into the soil so the nontoxigenic strain is in position to compete with the strains capable of producing aflatoxin. Sporulating barley next to peanut peg

This ARS technology has been licensed, commercialized, and is now marketed
by Syngenta Crop Protection under the trade name, AflaGuard®. In May 2004, AflaGuard® received EPA Section 3 registration as a biopesticide for use on peanuts. The label was amended to include corn in October, 2008.


» Peanut Test Results

Large-scale field studies were conducted in peanuts in 2004 to demonstrate efficacy prior to EPA registration. AflaGuard® was applied at 20 lb/acre to about 5000 acres of peanuts in Alabama and Georgia. Analyses of farmers’ stock peanuts from 582 loads showed that AflaGuard® reduced average aflatoxin by 85%, and consistent reductions were seen in all areas. Analyses of shelled stock peanuts from 2 warehouses resulted in no lots from treated areas being rejected because of aflatoxin compared with rejection rates of 16 and 48%, respectively,  for untreated peanuts.

 Commercial Use Results - Peanuts
Aflatoxin (ppb)

 Untreated

Treated 

Farmers' Stock

78.9

11.7 

Shelled Stock
Warehouse 1

36.2

0.9

Shelled Stock
Warehouse 2

7.2

2.2


» Corn Test Results

A two-year experimental use permit was granted by EPA to determine the efficacy of AflaGuard® in corn. About 3000 acres were treated in two areas of Texas with either 10 or 20 lb/ac in 2007 and 2008. AflaGuard® reduced aflatoxin contamination by averages of 85% and 88% in respective years. There was no significant difference in the reduction achieved by the different rates.

 Commercial Use Results - Corn
Aflatoxin (ppb)

 Treatment

2007 

2008 

 Control

 3.4

 12.4

 10 lb/ac

 0.4

  0.7

 20 lb/ac

 0.6

  2.1

Analyses for the type of A. flavus colonizing corn revealed that both the 10 and 20 lb/ac treatments displaced > 99% of the toxigenic strains of the fungus.  This is strong confirmation that AflaGuard® established the nontoxigenic A. flavus in the environment and that biological control through competitive exclusion is an effective aflatoxin prevention strategy.

 Incidence (%) of Toxigenic
A. flavus in Corn
 

 Treatment

2007 

2008 

 Control

 26.3

 21.0

 10 lb/ac

   0.2

   0.4

 20 lb/ac

   0.4

   1.3

For more information contact Bruce Horn or Marshall Lamb.


Fungal Invasion of Peanuts


Peanuts fruit underground and are in direct contact with soil populations of Aspergillus flavusand Aspergillus parasiticus, the two fungi responsible for producing aflatoxin in peanuts.

 Peanut plant with soil removed to show pods.

 Aspergillus flavus

Aspergillus parasiticus

 Aspergillus flavus

 Aspergillus parasiticus

A laboratory procedure has been developed in which viable peanut seeds are wounded and inoculated with soil directly from the field.  This research has shown that despite low spore densities in soil, aflatoxin-producing fungi preferentially invade peanuts over other soil-inhabiting fungi.  Invasion is greatly influenced by seed moisture and soil temperature.

Aspergillus infected kernel

Aspergillus infected kernel

Seeds colonized by Aspergillusspecies

 Chart showing the effect of soil population density on peanut infection

Effect of soil population density on peanut infection

This procedure is currently being used to assay different nontoxigenic A. flavusstrains for their ability to reduce aflatoxin in biological control.

Ability of eight nontoxigenic Aspergillus flavusstrains to inhibit aflatoxin production by specific aflatoxin-producing strains using the peanut seed assay
 

   Nontoxigenic strains
Aflatoxin production by strain F35  
Significant reduction of aflatoxin: Strain A > B > C > D > E > F 
No significant reduction in aflatoxin: Strain G, H
   
Aflatoxin production by strain F14  
Significant reduction of aflatoxin: Strain G > C > E > A > D > F> H > B
   
Aflatoxin production by strain F86  
Significant reduction of aflatoxin: Strain H > A > C
No significant reduction in aflatoxin: Strain B, E, D, G, F
   
Aflatoxin production by strain F44  
Significant reduction of aflatoxin: Strain C > A > D > G > E > F
No significant reduction in aflatoxin: Strain B, H
 
 F= Biological control strain in current use

For more information contact Bruce Horn.


Fungal Population Biology


A thorough understanding of the ecology and population biology of aflatoxin-producing fungi is a prerequisite for devising new strategies for controlling aflatoxin in peanuts.  We have examined populations at three spatial scales.

Single peanut field in Georgia:  Population structure is complex and aflatoxin production by fungi is extremely variable.

Peanut field
Chart of aflatoxin production by different genetic groups

Aflatoxin production by different genetic groups

Transect through the southern United States:  Peanut-growing regions having severe aflatoxin outbreaks are dominated by toxigenic strains of Aspergillus flavus.

Picture showing peanut-growing regions having severe aflatoxin outbreaks and dominated by toxigenic strains of Aspergillus flavus (1997).

Worldwide populations where peanuts are grown:  On-going study in collaboration with North Carolina State University shows large differences in population structure and genetic composition.

Worldwide populations where peanuts are grown
(Map courtesy of CIA - The World Factbook)

 

For more information contact Bruce Horn.

 


Last Modified: 1/6/2010