Production of the 14 kDa trypsin inhibitor protein is important for maize resistance against Aspergillus flavus infection/aflatoxin accumulation

2016 ◽  
Vol 9 (2) ◽  
pp. 215-228 ◽  
Author(s):  
Z.-Y. Chen ◽  
M.L. Warburton ◽  
L. Hawkins ◽  
Q. Wei ◽  
Y. Raruang ◽  
...  
Keyword(s):  
Aspergillus Flavus ◽  
Trypsin Inhibitor ◽  
Aflatoxin Production ◽  
Field Resistance ◽  
Aflatoxin Contamination ◽  
Direct Role ◽  
Independent Events ◽  
Sequence Polymorphisms ◽  
Transgenic Lines ◽  
Aflatoxin Accumulation

Maize (Zea mays L.) is one of the major crops susceptible to Aspergillus flavus Link ex. Fries infection and subsequent aflatoxin contamination. Previous studies found the production of an antifungal 14 kDa trypsin inhibitor (TI) was associated with maize aflatoxin resistance. To further investigate whether the TI plays any direct role in resistance, a TI gene silencing vector was constructed and transformed into maize. Mature kernels were produced from 66 transgenic lines representing 18 independent events. A final total of twelve lines representing four independent events were confirmed positive for transformation, five of which showed significant reduction (63 to 88%) in TI transcript abundance in seedling leaf tissue and seven of which showed significant TI protein reduction (39-85%) in mature kernels. Six of the seven silenced transgenic lines supported higher levels of aflatoxin production compared to negative controls. To further confirm the role of TI in field resistance to aflatoxin accumulation, DNA sequence polymorphisms from within the gene or linked simple sequence repeats were tested in four quantitative trait loci (QTL) mapping populations for QTL effect, and three QTL with log of the odds scores of 11, 4.5, and 3.0 and possibly caused by the TI protein encoding gene were found. Sequence polymorphisms were also tested for association to aflatoxin levels in an association mapping panel, and three single nucleotide polymorphisms were found associated with aflatoxin accumulation (P<0.01). The data from both RNAi and genetic mapping studies demonstrated that production of the TI in maize is important for its resistance to A. flavus infection and/or aflatoxin production.

scholarly journals Interactions of Saprophytic Yeasts with anor Mutant of Aspergillus flavus

1999 ◽  
Vol 65 (6) ◽  
pp. 2738-2740 ◽  
Author(s):  
Sui-Sheng T. Hua ◽  
James L. Baker ◽  
Melanie Flores-Espiritu
Keyword(s):  
Aspergillus Flavus ◽  
Agar Plate ◽  
Aflatoxin Production ◽  
Aflatoxin Contamination ◽  
Orange Pigment ◽  
Yeast Strains ◽  
Norsolorinic Acid ◽  
Aflatoxin Accumulation ◽  
Food Commodities ◽  
Pigment Accumulation

ABSTRACT The nor mutant of Aspergillus flavus has a defective norsolorinic acid reductase, and thus the aflatoxin biosynthetic pathway is blocked, resulting in the accumulation of norsolorinic acid, a bright red-orange pigment. We developed a visual agar plate assay to monitor yeast strains for their ability to inhibit aflatoxin production by visually scoring the accumulation of this pigment of the nor mutant. We identified yeast strains that reduced the red-orange pigment accumulation in the normutant. These yeasts also reduced aflatoxin accumulation by a toxigenic strain of A. flavus. These yeasts may be useful for reducing aflatoxin contamination of food commodities.

scholarly journals Evaluation of Arachis Species and Interspecific Tetraploid Lines for Resistance to Aflatoxin Production by Aspergillus flavus

2004 ◽  
Vol 31 (2) ◽  
pp. 134-141 ◽  
Author(s):  
H. Q. Xue ◽  
T. G. Isleib ◽  
H. T. Stalker ◽  
G. A. Payne ◽  
G. OBrian
Keyword(s):  
Aspergillus Flavus ◽  
Insect Pests ◽  
Low Cost ◽  
Aflatoxin Production ◽  
Management Program ◽  
Aflatoxin Contamination ◽  
Sources Of Resistance ◽  
Resistant Parent ◽  
Arachis Species ◽  
Aflatoxin Accumulation

Abstract Anatoxins are carcinogenic and extremely toxic secondary metabolites produced primarily by two fungi, Aspergillus flavus Link ex Fries and A. parasiticus Speare. Elimination of aflatoxin contamination in peanut (Arachis hypogaea L.) is a high priority of the peanut industry. Resistant cultivars should be an effective and low-cost part of an integrated aflatoxin management program. To date, no cultivated peanut has been reported with stable high levels of resistance to aflatoxin production. Arachis species and interspecific tetraploid lines have been evaluated for resistance to several peanut diseases and insect pests, and highly resistant accessions have been reported. Seven accessions of A. cardenasii Krapov. and W.C. Gregory, 29 of A. duranensis Krapov. and W.C. Gregory, and 17 interspecific tetraploid lines derived from A. hypogaea × A. cardenasii were inoculated with A. flavus strain NRRL 3357 and analyzed for aflatoxin content after incubation. On average, A. duranensis and A. cardenasii accumulated significantly less aflatoxin than A. hypogaea checks. The mean difference between the two wild species was not significant. Arachis duranensis accessions PI 468319 (GKBSPSc 30073), PI 468200 (GKBSPSc 30064), and PI 262133 (GKP 10038 sl.); and A. cardenasii accessions PI 262141 (GKP 10017) and PI 475997 (KSSc 36018) had reduced levels of aflatoxin accumulation and should be valuable sources of resistance to aflatoxin contamination. Of the interspecific tetraploid lines, only GP-NC WS 2 supported aflatoxin production not significantly different from resistant parent A. cardenasii GKP 10017, and it appears to be a line with reduced capacity for aflatoxin accumulation.

Modelling the colonisation of maize by toxigenic and non-toxigenic Aspergillus flavus strains: implications for biological control

2008 ◽  
Vol 1 (3) ◽  
pp. 333-340 ◽  
Author(s):  
H. Abbas ◽  
R. Zablotowicz ◽  
H. Bruns
Keyword(s):  
Biological Control ◽  
Aspergillus Flavus ◽  
Growth Parameters ◽  
Biological Control Agent ◽  
Cyclopiazonic Acid ◽  
Aflatoxin Contamination ◽  
Tween 20 ◽  
Lag Phase ◽  
Gompertz Equation ◽  
Aflatoxin Accumulation

To successfully exploit biological control it is desirable to understand how the introduced agent colonises the host and interferes with establishment of the pest. This study assessed field colonisation of maize by Aspergillus flavus strains as biological control agents to reduce aflatoxin contamination. Maize (corn, Zea mays L.) ears were inoculated with A. flavus using a pin-bar technique in 2004 and 2005. Non-aflatoxigenic strains K49 (NRRL 30797) & CT3 (NRRL 30798) and toxigenic F3W4 (NRRL 30798) were compared against a carrier control (0.2% aqueous Tween 20). Ten ears were sampled over 12 to 20 days, visually assessed, and curves fit to a three compartment Gompertz equation or other best appropriate regressions. Aflatoxin was determined by HPLC and cyclopiazonic acid (CPA) by LC/MS. The Gompertz model describes growth parameters, e.g. growth constant, lag phase and maximum colonisation characterised patterns of maize colonisation for most inoculated treatments. Aflatoxin accumulation in maize inoculated with F3W4 was about 35,000 ng/g in 2004 and 2005, with kinetics of aflatoxin accumulation in 2005 well described by the Gompertz equation. Less than 200 ng/g was observed in maize inoculated with strains CT3 & K49 and accumulation was described by a linear or logistic model. Maize inoculated with strains CT3 and F3W4 accumulated a maximum of 220 and 169 µg/kg CPA, respectively, compared to 22 and 0.2 µg/kg in the control and K49 inoculated, respectively. This technique can be used to elucidate colonisation potential of non-toxigenic A. flavus in maize in relation to biological control of aflatoxin. The greatest reduction of aflatoxin and CPA in maize inoculated with strain K49 and Gompertz parameters on colonisation indicates its superiority to CT3 as a biological control agent. The dynamics of maize colonisation by A. flavus strains and subsequent mycotoxin accumulation generated by using the pin-bar technique has implications for characterising the competence of biocontrol strains for reducing aflatoxin contamination.

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

2002 ◽  
Vol 65 (12) ◽  
pp. 1984-1987 ◽  
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.

scholarly journals The Frequency of Sex: Population Genomics Reveals Differences in Recombination and Population Structure of the Aflatoxin-Producing Fungus Aspergillus flavus

mBio ◽  
2020 ◽  
Vol 11 (4) ◽  
Author(s):  
Milton T. Drott ◽  
Tatum R. Satterlee ◽  
Jeffrey M. Skerker ◽  
Brandon T. Pfannenstiel ◽  
N. Louise Glass ◽  
...  
Keyword(s):  
Aspergillus Flavus ◽  
Population Genomics ◽  
Population Expansion ◽  
Aflatoxin Production ◽  
The United States ◽  
Aflatoxin Contamination ◽  
Deleterious Mutations ◽  
Content Type ◽  
Sexual And Asexual Reproduction ◽  
Potent Carcinogen

ABSTRACT The apparent rarity of sex in many fungal species has raised questions about how much sex is needed to purge deleterious mutations and how differences in frequency of sex impact fungal evolution. We sought to determine how differences in the extent of recombination between populations of Aspergillus flavus impact the evolution of genes associated with the synthesis of aflatoxin, a notoriously potent carcinogen. We sequenced the genomes of, and quantified aflatoxin production in, 94 isolates of A. flavus sampled from seven states in eastern and central latitudinal transects of the United States. The overall population is subdivided into three genetically differentiated populations (A, B, and C) that differ greatly in their extent of recombination, diversity, and aflatoxin-producing ability. Estimates of the number of recombination events and linkage disequilibrium decay suggest relatively frequent sex only in population A. Population B is sympatric with population A but produces significantly less aflatoxin and is the only population where the inability of nonaflatoxigenic isolates to produce aflatoxin was explained by multiple gene deletions. Population expansion evident in population B suggests a recent introduction or range expansion. Population C is largely nonaflatoxigenic and restricted mainly to northern sampling locations through restricted migration and/or selection. Despite differences in the number and type of mutations in the aflatoxin gene cluster, codon optimization and site frequency differences in synonymous and nonsynonymous mutations suggest that low levels of recombination in some A. flavus populations are sufficient to purge deleterious mutations. IMPORTANCE Differences in the relative frequencies of sexual and asexual reproduction have profound implications for the accumulation of deleterious mutations (Muller’s ratchet), but little is known about how these differences impact the evolution of ecologically important phenotypes. Aspergillus flavus is the main producer of aflatoxin, a notoriously potent carcinogen that often contaminates food. We investigated if differences in the levels of production of aflatoxin by A. flavus could be explained by the accumulation of deleterious mutations due to a lack of recombination. Despite differences in the extent of recombination, variation in aflatoxin production is better explained by the demography and history of specific populations and may suggest important differences in the ecological roles of aflatoxin among populations. Furthermore, the association of aflatoxin production and populations provides a means of predicting the risk of aflatoxin contamination by determining the frequencies of isolates from low- and high-production populations.

scholarly journals Resistance to Aspergillus flavus in Corn Kernels Is Associated with a 14-kDa Protein

1998 ◽  
Vol 88 (4) ◽  
pp. 276-281 ◽  
Author(s):  
Z.-Y. Chen ◽  
R. L. Brown ◽  
A. R. Lax ◽  
B. Z. Guo ◽  
T. E. Cleveland ◽  
...  
Keyword(s):  
Aspergillus Flavus ◽  
Trypsin Inhibitor ◽  
Aflatoxin Contamination ◽  
Inhibitor Protein ◽  
Trypsin Activity ◽  
Terminal Sequence ◽  
High Concentration ◽  
Resistant Varieties ◽  
Corn Kernels

Corn genotypes resistant or susceptible to Aspergillus flavus were extracted for protein analysis using a pH 2.8 buffer. The profile of protein extracts revealed that a 14-kDa protein is present in relatively high concentration in kernels of seven resistant corn genotypes, but is absent or present only in low concentration in kernels of six susceptible ones. The N-terminal sequence of this 14-kDa protein showed 100% homology to a corn trypsin inhibitor. The 14-kDa protein purified from resistant varieties also demonstrated in vitro inhibition of both trypsin activity and the growth of A. flavus. This is the first demonstration of antifungal activity of a corn 14-kDa trypsin inhibitor protein. The expression of this protein among tested genotypes may be related to their difference in resistance to A. flavus infection and subsequent aflatoxin contamination.

Atoxigenic Aspergillus flavus biological control of aflatoxin contamination: what is the mechanism?

2015 ◽  
Vol 8 (2) ◽  
pp. 235-244 ◽  
Author(s):  
K.E. Damann Jr.
Keyword(s):  
Biological Control ◽  
Aspergillus Flavus ◽  
Population Biology ◽  
Aflatoxin Production ◽  
Infection Process ◽  
Aflatoxin Contamination ◽  
Signal Molecules ◽  
Nutrient Competition ◽  
Toxigenic Strain ◽  
Aflatoxin Inhibition

The term ‘competitive exclusion’ involving physical blockage of growth or access of the toxigenic strain to the seed target has been used to describe the mechanism of biological control of aflatoxin contamination. However, recent evidence suggests that a form of intraspecific aflatoxin inhibition requiring growth of the competing strains together during the infection process in such a way that hyphae physically interact or touch is the trigger for preventing induction of aflatoxin synthesis. This direct touch-based inhibition of aflatoxin synthesis is posited to be the mechanistic basis of biological control in this system. Evidence for this idea comes from the published observations that co-culture of toxigenic and atoxigenic strains in a suspended disc system, in which the hyphae physically interact, prevents aflatoxin production. However, growth of the same strains in the same medium in the two compartments of a filter insert plate well system, separating the atoxigenic and toxigenic strains with a 0.4 μm or 3.0 μm filter, allows aflatoxin production approaching that of the toxigenic strain alone. When the strains are mixed and placed in both the insert and the well compartments, the intraspecific aflatoxin inhibition occurs as it did in the suspended disc culture system. This further suggests that neither nutrient competition nor soluble signal molecules, which should pass through the filter, are involved in intraspecific aflatoxin inhibition. When the two strains are separated by a 12 μm filter that would allow some passage of conidia or hyphae between the compartments the aflatoxin synthesis is approximately half that of the toxigenic strain alone. This phenomenon could be termed ‘competitive inclusion’ or ‘competitive phenotype conversion’. Work of others that relates to understanding the phenomenon is discussed, as well as an Aspergillus flavus population biology study from the Louisiana maize agro-ecosystem which has biological control implications.

Study on the Key Environmental Factors Aspergillus flavus Growth and Aflatoxin Production

2014 ◽  
Vol 668-669 ◽  
pp. 1550-1553 ◽  
Author(s):  
Chu Shu Zhang ◽  
Qin Zhao ◽  
Jian Xiong Feng ◽  
Jie Sun ◽  
Li Na Yu ◽  
...  
Keyword(s):  
Environmental Factors ◽  
Aspergillus Flavus ◽  
Prevention And Control ◽  
Aflatoxin Production ◽  
Aflatoxin Contamination ◽  
Key Factors ◽  
Culture Time ◽  
Burman Design ◽  
And Control ◽  
Plackett Burman Design

Single factor and Plackett-Burman design were used to determine the key environmental factors for Aspergillus flavus growth and aflatoxin production. The results showed that the key factors were culture time, moisture content. So Controling of moisture was crucial for Aflatoxin contamination prevention and control.

scholarly journals Assessment of the Effect of Satureja montana and Origanum virens Essential Oils on Aspergillus flavus Growth and Aflatoxin Production at Different Water Activities

Toxins ◽  
2020 ◽  
Vol 12 (3) ◽  
pp. 142 ◽  
Author(s):  
Marta García-Díaz ◽  
Jessica Gil-Serna ◽  
Belén Patiño ◽  
Esther García-Cela ◽  
Naresh Magan ◽  
...  
Keyword(s):  
Essential Oil ◽  
Essential Oils ◽  
Aspergillus Flavus ◽  
Aflatoxin Production ◽  
Aflatoxin Contamination ◽  
Activity Levels ◽  
Control Methods ◽  
Mycotoxin Biosynthesis ◽  
Satureja Montana

Aflatoxin contamination of foodstuffs poses a serious risk to food security, and it is essential to search for new control methods to prevent these toxins entering the food chain. Several essential oils are able to reduce the growth and mycotoxin biosynthesis of toxigenic species, although their efficiency is strongly influenced by the environmental conditions. In this work, the effectiveness of Satureja montana and Origanum virens essential oils to control Aspergillus flavus growth was evaluated under three water activity levels (0.94, 0.96 and 0.98 aw) using a Bioscreen C, a rapid in vitro spectrophotometric technique. The aflatoxin concentrations at all conditions tested were determined by HPLC-FLD. Aspergillus flavus growth was delayed by both essential oil treatments. However, only S. montana essential oil was able to significantly affect aflatoxin production, although the inhibition percentages widely differed among water activities. The most significant reduction was observed at 0.96 aw, which is coincident with the conditions in which A. flavus reached the highest levels of aflatoxin production. On the contrary, the treatment with S. montana essential oil was not effective in significantly reducing aflatoxin production at 0.94 aw. Therefore, it is important to study the interaction of the new control compounds with environmental factors before their application in food matrices, and in vitro ecophysiological studies are a good option since they provide accurate and rapid results.

scholarly journals Aflatoxin Production in Soybean Varieties Grown in Argentina

1991 ◽  
Vol 54 (7) ◽  
pp. 542-545 ◽  
Author(s):  
VIRGINIA E. FERNÁNDEZ PINTO ◽  
GRACIELA VAAMONDE ◽  
SILVIA B. BRIZZIO ◽  
NICOLÁS APRO
Keyword(s):  
Aspergillus Flavus ◽  
Aflatoxin Production ◽  
Aflatoxin Contamination ◽  
Fungal Isolate ◽  
Natural Occurrence ◽  
Laboratory Conditions

Natural occurrence of aflatoxin contamination on soybeans was investigated. Thirty four samples from the 1986 crop (5.8% positive samples) and 60 samples harvested in 1987 (11.6% positive samples) were analyzed. Aflatoxin levels in positive samples were low, ranging from traces to 36 μg/kg of total aflatoxins. Aflatoxin production by three isolates of Aspergillus flavus/A. parasiticus in 13 soybean varieties has been studied. Soybeans used for cultures were surface disinfected. Levels of aflatoxin formed were dependent both on the toxicogenic potential of the fungal isolate and on the variety of soybeans. Under laboratory conditions variety Prata was the most susceptible and variety SRF the less susceptible to aflatoxin formation.

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