Advances in the Development of Host Resistance in Corn to Aflatoxin Contamination by Aspergillus flavus

Aflatoxins are toxic, highly carcinogenic secondary metabolites of Aspergillus flavus and A. parasiticus, which when produced during fungal infection of a susceptible crop in the field or after harvest contaminate food and feed and threaten human and animal health. Although there are several management strategies that may reduce aflatoxin contamination of corn, the preeminent strategy for elimination of aflatoxin is to develop preharvest host resistance to aflatoxin accumulation. This strategy has gained even greater prominence due to recent discoveries of natural resistance in corn that can be exploited in plant-breeding strategies. The ability to identify resistant corn genotypes has been enhanced by the development of a laboratory kernel-screening assay and by a strain of A. flavus genetically engineered to produce β-glucuronidase, an enzyme whose activity can be monitored to assess the degree of fungal infection in kernels. Investigations of resistant corn genotypes have associated kernel pericarp wax characteristics with resistance, identified kernel proteins associated with resistance to and inhibition of fungal growth or aflatoxin biosynthesis, and identified chromosome regions associated with resistance to Aspergillus ear rot and aflatoxin production. Such research advances could lead, in the near future, to commercially available, agronomically acceptable corn lines with multiple preharvest resistances to aflatoxin contamination.

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Evaluation of resistance to aflatoxin contamination in kernels of maize genotypes using a GFP-expressing Aspergillus flavus strain

World Mycotoxin Journal ◽

10.3920/wmj2012.1497 ◽

2013 ◽

Vol 6 (2) ◽

pp. 151-158 ◽

Cited By ~ 12

Author(s):

K. Rajasekaran ◽

C.M. Sickler ◽

R.L. Brown ◽

J.W. Cary ◽

D. Bhatnagar

Keyword(s):

Fungal Infection ◽

Aspergillus Flavus ◽

Fluorescent Protein ◽

Aflatoxin Contamination ◽

Clear Correlation ◽

Aleurone Layer ◽

Screening Assay ◽

Maize Germplasm ◽

Antifungal Proteins ◽

Green Fluorescent

Resistance or susceptibility of maize inbreds to infection by Aspergillus flavus was evaluated by the kernel screening assay. A green fluorescent protein-expressing strain of A. flavus was used to measure fungal spread and aflatoxin levels in real-time following fungal infection of kernels. Among the four inbreds tested, MI82 showed the most resistance and Ga209 the least. TZAR101 was also resistant to fungal infection, whereas Va35 was susceptible to fungal infection. However, Va35 produced lower aflatoxin levels compared to the susceptible line Ga209. Fluorescence microscopy indicated that the site of entry of the fungus into the kernel was consistently through the pedicel. Entry through the pericarp was never observed in undamaged kernels. In view of these results, incorporation or overexpression of antifungal proteins should be targeted to the pedicel and basal endosperm region in developing kernels. Once the fungus has entered through the pedicel, it spreads quickly through the open spaces between the pericarp and the aleurone layer, ultimately colonising the endosperm and scutellum and, finally, the embryo. A clear correlation was established between fungal fluorescence and aflatoxin levels. This method provides a quick, reliable means of evaluating resistance to A. flavus in undamaged kernels and provides breeders with a rapid method to evaluate maize germplasm.

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Resistance to Aflatoxin Accumulation in Kernels of Maize Inbreds Selected for Ear Rot Resistance in West and Central Africa

Journal of Food Protection ◽

10.4315/0362-028x-64.3.396 ◽

2001 ◽

Vol 64 (3) ◽

pp. 396-400 ◽

Cited By ~ 42

Author(s):

ROBERT L. BROWN ◽

ZHI-YUAN CHEN ◽

ABEBE MENKIR ◽

THOMAS E. CLEVELAND ◽

KITTY CARDWELL ◽

...

Keyword(s):

Fungal Infection ◽

Natural Infection ◽

Polyacrylamide Gel Electrophoresis ◽

Sodium Dodecyl ◽

Central Africa ◽

Aflatoxin Contamination ◽

Ear Rot ◽

Screening Assay ◽

West And Central Africa ◽

Aflatoxin Accumulation

Thirty-six inbred lines selected in West and Central Africa for moderate to high resistance to maize ear rot under conditions of severe natural infection were screened for resistance to aflatoxin contamination using the previously established kernel screening assay. Results showed that more than half the inbreds accumulated aflatoxins at levels as low as or lower than the resistant U.S. lines GT-MAS:gk or MI82. In 10 selected aflatoxin-resistant or aflatoxin-susceptible inbreds, Aspergillus flavus growth, which was quantified using an A. flavus transformant containing a GUS-β-tubulin reporter gene construct, was, in general, positively related to aflatoxin accumulation. However, one aflatoxin-resistant inbred supported a relatively high level of fungal infection, whereas two susceptibles supported relatively low fungal infection. When kernels of the 10 tested lines were profiled for proteins using sodium dodecyl sulfate-polyacrylamide gel electrophoresis, significant variations from protein profiles of U.S. lines were observed. Confirmation of resistance in promising African lines in field trials may significantly broaden the resistant germplasm base available for managing aflatoxin contamination through breeding approaches. Biochemical resistance markers different from those being identified and characterized in U.S. genotypes, such as ones inhibitory to aflatoxin biosynthesis rather than to fungal infection, may also be identified in African lines. These discoveries could significantly enhance the host resistance strategy of pyramiding different traits into agronomically useful maize germplasm to control aflatoxin contamination.

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A Novel Niosome-Encapsulated Essential Oil Formulation to Prevent Aspergillus flavus Growth and Aflatoxin Contamination of Maize Grains During Storage

Toxins ◽

10.3390/toxins11110646 ◽

2019 ◽

Vol 11 (11) ◽

pp. 646 ◽

Cited By ~ 6

Author(s):

García-Díaz ◽

Patiño ◽

Vázquez ◽

Gil-Serna

Keyword(s):

Aspergillus Flavus ◽

Fungal Growth ◽

Storage Conditions ◽

Aflatoxin Contamination ◽

Small Scale ◽

Low Concentrations ◽

Oil Formulation ◽

Scale Test ◽

Encapsulation Method

Aflatoxin (AF) contamination of maize is a major concern for food safety. The use of chemical fungicides is controversial, and it is necessary to develop new effective methods to control Aspergillus flavus growth and, therefore, to avoid the presence of AFs in grains. In this work, we tested in vitro the effect of six essential oils (EOs) extracted from aromatic plants. We selected those from Satureja montana and Origanum virens because they show high levels of antifungal and antitoxigenic activity at low concentrations against A. flavus. EOs are highly volatile compounds and we have developed a new niosome-based encapsulation method to extend their shelf life and activity. These new formulations have been successfully applied to reduce fungal growth and AF accumulation in maize grains in a small-scale test, as well as placing the maize into polypropylene woven bags to simulate common storage conditions. In this latter case, the antifungal properties lasted up to 75 days after the first application.

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In Vitro Activity of Balanites aegyptiaca and Tamarindus indica Fruit Extracts on Growth and Aflatoxigenicity of Aspergillus flavus and A. parasiticus

Journal of Food Research ◽

10.5539/jfr.v2n4p68 ◽

2013 ◽

Vol 2 (4) ◽

pp. 68 ◽

Cited By ~ 1

Author(s):

Saifeldin Ahmed El-nagerabi ◽

Abdulkadir E. Elshafie ◽

Mohamed R. Elamin

Keyword(s):

Aspergillus Flavus ◽

Fungal Growth ◽

Aflatoxin Contamination ◽

Tamarindus Indica ◽

Balanites Aegyptiaca ◽

Total Aflatoxin ◽

Animal Products ◽

Fruit Extracts ◽

Aflatoxin B

Aflatoxin and especially aflatoxin B1 (AFB1) is a carcinogenic secondary metabolite synthesized by certain Aspergillus species. They contaminate natural and processed agricultural and animal products which render them unfit for consumption. The aim of this study was to evaluate the in vitro effects of Balanites aegyptiaca and Tamarindus indica fruit extracts on the growth and aflatoxin secretion of Aspergillus flavus (SQU21) and A. parasiticus (CBS921.7) strains. The two fruit extracts significantly (P < 0.05) reduced aflatoxin and did not inhibit mycelial dry weights of the two Aspergillus strains. At different concentrations of balanites (2.5-10%), the inhibition of total aflatoxin was 49.9-84.8% for A. flavus (SQU21) and 32.1-84.4% for A. parasiticus (CBS921.7), whereas the inhibition of aflatoxin Bwas 38.2-81.4% and 32.8-80.6% for the two strains. Tamarind fruit extract (2.5-7.5%) caused 28.8-84.2% and 40.7-85.5% reductions in total aflatoxin and 37.1-83.5% and 33.9-85.9% in aflatoxin B for the two strains, respectively. None of these extracts inhibited the fungal growth or detoxified synthetic aflatoxin B1. We have concluded that these fruits contain various inhibitors to aflatoxin biosynthesis and secretion. Therefore, they can be used in combination as safe green biopreservatives to combat aflatoxin contamination of food.

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No Effect of Soybean Lipoxygenase on Aflatoxin Production in Aspergillus flavus–Inoculated Seeds

Journal of Food Protection ◽

10.4315/0362-028x-65.12.1984 ◽

2002 ◽

Vol 65 (12) ◽

pp. 1984-1987 ◽

Cited By ~ 6

Author(s):

J. E. MELLON ◽

P. J. COTTY

Keyword(s):

Aspergillus Flavus ◽

Fungal Pathogens ◽

Seed Viability ◽

Soybean Seed ◽

Fungal Growth ◽

Aflatoxin Production ◽

Aflatoxin Contamination ◽

Soybean Seeds ◽

Heat Treated ◽

Significant Difference

Soybean lines lacking lipoxygenase (LOX) activity were compared with soybean lines having LOX activity for the ability to support growth and aflatoxin B1 production by the fungal seed pathogen Aspergillus flavus. Whole seeds, broken seeds, and heat-treated (autoclaved) whole seeds were compared. Broken seeds, irrespective of LOX presence, supported excellent fungal growth and the highest aflatoxin levels. Autoclaved whole seeds, with or without LOX, produced good fungal growth and aflatoxin levels approaching those of broken seeds. Whole soybean seeds supported sparse fungal growth and relatively low aflatoxin levels. There was no significant difference in aflatoxin production between whole soybean seeds either with or without LOX, although there did seem to be differences among the cultivars tested. The heat treatment eliminated LOX activity (in LOX+ lines), yet aflatoxin levels did not change substantially from the broken seed treatment. Broken soybean seeds possessed LOX activity (in LOX+ lines) and yet yielded the highest aflatoxin levels. The presence of active LOX did not seem to play the determinant role in the susceptibility of soybean seeds to fungal pathogens. Seed coat integrity and seed viability seem to be more important characteristics in soybean seed resistance to aflatoxin contamination. Soybean seeds lacking LOX seem safe from the threat of increased seed pathogen susceptibility.

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Aflatoxin Contamination in Agricultural Commodities

Indian Journal of Pharmaceutical and Biological Research ◽

10.30750/ijpbr.1.4.25 ◽

2013 ◽

Vol 1 (04) ◽

pp. 148-151 ◽

Cited By ~ 5

Author(s):

P. N. Rajarajan ◽

K. M. Rajasekaran ◽

N. K. Asha Devi

Keyword(s):

Aspergillus Flavus ◽

Aflatoxin B1 ◽

Animal Health ◽

Aflatoxin Contamination ◽

Aflatoxin M1 ◽

Organic Substrates ◽

Naturally Occurring ◽

Food And Feed ◽

Hydroxylated Metabolite ◽

Carcinogenic Compound

Aflatoxin is a naturally occurring Mycotoxin produced by Aspergillus flavus and Aspergillus parasiticus. Aspergillus flavus is common and widespread in nature and is most often found when certain grains are grown under stressful conditions such as draught. The mold occurs in soil, decaying vegetation, hay and grains undergoing microbiological deterioration and invades all types of organic substrates whenever and wherever the conditions are favourable for its growth. Favourable conditions include high moisture content and high temperature.The aflatoxin group is comprised of aflatoxin B1,B2,G1 and G2. In addition , aflatoxin M1 (AFM1), a hydroxylated metabolite of AFB1, is excreted in the milk of dairy cows consuming an AFB1-contaminated ration. Aflatoxin B1 a prototype of the aflatoxins, is widely recognized as the most potent hepato carcinogenic compound and along with other certain members of the group, possess additional toxic properties including mutagenicity, tetrogenicity, acute cellular toxicity and it suppresses the immune system. Aflatoxin contamination of food and feed has gained global significance as a result of its deleterious effects on human as well as animal health. The marketability of food products is adversely affected by aflatoxin contamination.

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Living Maize Embryo Influences Accumulation of Aflatoxin in Maize Kernels

Journal of Food Protection ◽

10.4315/0362-028x-56.11.967 ◽

1993 ◽

Vol 56 (11) ◽

pp. 967-971 ◽

Cited By ~ 65

Author(s):

ROBERT L. BROWN ◽

PETER J. COTTY ◽

THOMAS E. CLEVELAND ◽

NEIL W. WIDSTROM

Keyword(s):

Aspergillus Flavus ◽

Fungal Growth ◽

Aflatoxin Contamination ◽

Population Variation ◽

Maize Populations ◽

Susceptible Control ◽

Maize Embryo ◽

Metabolic Activities ◽

Maize Kernels ◽

Two Populations

Kernels from two maize populations, MAS:gk and MAS:pw,nf, showed significant postharvest resistance to aflatoxin contamination by Aspergillus flavus but showed no significant inter-population variation for this resistance. Growth of A. flavus on both populations was significantly less than on susceptible control lines. Kernels from the resistant populations retained resistance when wounded through the pericarp prior to inoculation with A. flavus, despite the fact that the exposed endosperm supported good fungal growth. Kernels from these populations also retained resistance when they were acetone washed before inoculation. Resistance to aflatoxin contamination was lost in kernels that were autoclaved, crushed, or embryo wounded. All assays were incubated under conditions favorable to kernel germination. Results suggest that postharvest resistance to aflatoxin contamination in these two populations is related to metabolic activities of the living com embryo.

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Inhibition of Aflatoxin Production by Paraquat and External Superoxide Dismutase in Aspergillus flavus

Toxins ◽

10.3390/toxins11020107 ◽

2019 ◽

Vol 11 (2) ◽

pp. 107 ◽

Cited By ~ 4

Author(s):

Tomohiro Furukawa ◽

Shohei Sakuda

Keyword(s):

Superoxide Dismutase ◽

Aspergillus Flavus ◽

Regulatory Protein ◽

Fungal Growth ◽

Aflatoxin Production ◽

Aflatoxin Contamination ◽

Sod Activity ◽

Fungal Cell ◽

Cell Extracts ◽

Ic50 Value

Aflatoxin contamination of crops is a worldwide problem, and elucidation of the regulatory mechanism of aflatoxin production, for example relative to the oxidative–antioxidative system, is needed. Studies have shown that oxidative stress induced by reactive oxygen species promotes aflatoxin production. However, superoxide has been suggested to have the opposite effect. Here, we investigated the effects of the superoxide generator, paraquat, and externally added superoxide dismutase (SOD) on aflatoxin production in Aspergillus flavus. Paraquat with an IC50 value of 54.9 µM inhibited aflatoxin production without affecting fungal growth. It increased cytosolic and mitochondrial superoxide levels and downregulated the transcription of aflatoxin biosynthetic cluster genes, including aflR, a key regulatory protein. The addition of bovine Cu/ZnSOD to the culture medium suppressed the paraquat-induced increase in superoxide levels, but it did not fully restore paraquat-inhibited aflatoxin production because bovine Cu/ZnSOD with an IC50 value of 17.9 µg/mL itself inhibited aflatoxin production. Externally added bovine Cu/ZnSOD increased the SOD activity in fungal cell extracts and upregulated the transcription of genes encoding Cu/ZnSOD and alcohol dehydrogenase. These results suggest that intracellular accumulation of superoxide impairs aflatoxin production by downregulating aflR expression, and that externally added Cu/ZnSOD also suppresses aflatoxin production by a mechanism other than canonical superoxide elimination activity.

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Comparison of Different Image Processing Methods for Segregation of Peanut (Arachis hypogaea L.) Seeds Infected by Aflatoxin-Producing Fungi

Agronomy ◽

10.3390/agronomy11050873 ◽

2021 ◽

Vol 11 (5) ◽

pp. 873

Author(s):

Peyman Ziyaee ◽

Vahid Farzand Ahmadi ◽

Pourya Bazyar ◽

Eugenio Cavallo

Keyword(s):

Image Processing ◽

Arachis Hypogaea ◽

Aspergillus Flavus ◽

Animal Feed ◽

Animal Health ◽

Fungal Growth ◽

Support Vector ◽

Computational Techniques ◽

Inference System ◽

Arachis Hypogaea L

Fungi such as Aspergillus flavus and Aspergillus parasiticus are molds infecting food and animal feed, are responsible for aflatoxin contamination, and cause a significant problem for human and animal health. The detection of aflatoxin and aflatoxigenic fungi on raw material is a major concern to protect health, secure food and feed, and preserve their value. The effectiveness of image processing, combined with computational techniques, has been investigated to detect and segregate peanut (Arachis hypogaea L.) seeds infected with an aflatoxin producing fungus. After inoculation with Aspergillus flavus, images of peanuts seeds were taken using various lighting sources (LED, UV, and fluorescent lights) on two backgrounds (black and white) at 0, 48, and 72 h after inoculation. Images were post-processed with three different machine learning tools: the artificial neural network (ANN), the support vector machine (SVM), and the adaptive neuro-fuzzy inference system (ANFIS) to detect the Aspergillus flavus growth on peanuts. The results of the study show that the combination of LED light and a white background with ANN had 99.7% accuracy in detecting fungal growth on peanuts 72 h from infection with Aspergillus. Additionally, UV lights and a black background with ANFIS achieve 99.9% accuracy in detecting fungal growth on peanuts 48 h after their infection with Aspergillus.

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Inhibition of Aspergillus flavus growth and aflatoxin B1 production by natamycin

World Mycotoxin Journal ◽

10.3920/wmj2020.2620 ◽

2021 ◽

pp. 1-16

Author(s):

P. Chang ◽

B. Tai ◽

M. Zheng ◽

Q. Yang ◽

F. Xing

Keyword(s):

Aspergillus Flavus ◽

Animal Health ◽

Fungal Growth ◽

Outer Wall ◽

Aflatoxin Production ◽

Ergosterol Biosynthesis ◽

Dependent Manner ◽

Conidia Germination ◽

Streptomyces Spp ◽

Inhibitory Mechanisms

Aspergillus flavus causes huge crop losses, reduces crop quality and has adverse effects on human and animal health. A large amount of food contaminated with aflatoxin can greatly increase the risk of liver cancer. Therefore, prevention and control of aflatoxin production have aroused attention of research in various countries. Natamycin extracted from Streptomyces spp. has been widely used in production practice due to its good specificity and safety. Here, we found that natamycin could significantly inhibit fungal growth, conidia germination, ergosterol and AFB1 production by A. flavus in a dose-dependent manner. Scanning electron microscope analysis indicated that the number of conidia was decreased, the outer wall of conidia was destroyed, and the mycelia were shrivelled and tangled by natamycin. RNA-Seq data indicated that natamycin inhibited fungal growth and conidia development of A. flavus by significantly down-regulating some genes involved in ergosterol biosynthesis, such as Erg13, HMG1 and HMG2. It inhibited conidia germination by significantly down-regulating some genes related to conidia development, such as FluG and VosA. After natamycin exposure, the decreased ratio of aflS/aflR caused by the down-regulation of all the structural genes, which subsequently resulted in the suppression of AFB1 production. In conclusion, this study served to reveal the inhibitory mechanisms of natamycin on fungal growth and AFB1 biosynthesis in A. flavus and to provide solid evidence for its application in controlling AFB1 contamination.

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