Inhibition Of Aflatoxin Production Research Articles

Phytochemicals analysis and aflatoxin B1 detoxification potential of leaves extract of Moringa oleifera and Calotropis procera

The present study aimed to identify the presence of certain classes of phytochemicals in the leaf extract of medicinal herbs viz. Moringa oleifera and Calotropis procera, using qualitative detection tests and explored the potential of aqueous and ethanolic extract to inhibit aflatoxin production by thin layer chromatography at 25 °C and pH (7) of different incubation times i.e. 0-, 1-, 3-, 6- and 24-h. Qualitative phytochemical analysis reported that the aqueous leave extracts of M. oleifera and C. procera contained tannins, phlobatannins, quinones, steroids, sugar, betacyanins, fatty acids, phenols, and volatile oils. Aflatoxin analysis reported that the ethanolic extract of M. oleifera was found more effective than detoxifying 100% of AFB1 after 24 h of incubation. In the case of C. procera, the aqueous extract reduced 96.5% of AFB1 and ethanolic extract reduced 96% of AFB1 after 24-h of incubation. The results revealed that natural plant products have a high potential to reduce AFB1 and could contribute to mitigation plans for AFB1. There is a need for further characterisation using techniques such as GC-MS, LC-MS, or NMR which would provide valuable information on the chemical composition of the extracts.

Inhibition of Aflatoxin Production in Aspergillus flavus by a Klebsiella sp. and Its Metabolite Cyclo(l-Ala-Gly).

During an experiment where we were cultivating aflatoxigenic Aspergillus flavus on peanuts, we accidentally discovered that a bacterium adhering to the peanut strongly inhibited aflatoxin (AF) production by A. flavus. The bacterium, isolated and identified as Klebsiella aerogenes, was found to produce an AF production inhibitor. Cyclo(l-Ala-Gly), isolated from the bacterial culture supernatant, was the main active component. The aflatoxin production-inhibitory activity of cyclo(l-Ala-Gly) has not been reported. Cyclo(l-Ala-Gly) inhibited AF production in A. flavus without affecting its fungal growth in a liquid medium with stronger potency than cyclo(l-Ala-l-Pro). Cyclo(l-Ala-Gly) has the strongest AF production-inhibitory activity among known AF production-inhibitory diketopiperazines. Related compounds in which the methyl moiety in cyclo(l-Ala-Gly) is replaced by ethyl, propyl, or isopropyl have shown much stronger activity than cyclo(l-Ala-Gly). Cyclo(l-Ala-Gly) did not inhibit recombinant glutathione-S-transferase (GST) in A. flavus, unlike (l-Ala-l-Pro), which showed that the inhibition of GST was not responsible for the AF production-inhibition of cyclo(l-Ala-Gly). When A. flavus was cultured on peanuts dipped for a short period of time in a dilution series bacterial culture broth, AF production in the peanuts was strongly inhibited, even at a 1 × 104-fold dilution. This strong inhibitory activity suggests that the bacterium is a candidate for an effective biocontrol agent for AF control.

Phytochemical Analysis and Antifungal Effects of the Extracts and Oils of Harjel (Solenostemma argel) Basil (Ocimum basilicum) and Parsley (Petroselinum crispum) on the Fungus Aspergillus flavus Growth and Aflatoxin Production, Sudan

Sudan is one of the largest countries in Africa and is rich in many valuable plants that produce materials important for human and animal health. Some of these were used in traditional medicine. The present study aims to investigate the antifungal activities and the phytochemical composition of the extracts and oils of three plants (Hargel et al.). The study was conducted at the Food Microbiology Laboratory, Faculty of Engineering and Technology, University of Gezira. The mycelial growth and spore germination methods were used for the antifungal tests, using a selective medium, while thin-layer chromatography was used for phytochemical tests and aflatoxin analysis. The effects of clove oil and some amino acids were used for comparison. The results of the fungal growth showed that the leaf extracts of Hargel were highly effective and gave complete inhibition (0.0 cm radial growth), while the stem extracts and oil of Hargel were less effective, given (0.3 mm). Rehan (Basil) oil was the most effective (0.0 cm) compared to (0.4cm) and (0.5cm) for the extracts of the leaf and stem, respectively. The oil, leaf, and stem extracts of Parsley and clove oil were all highly effective, giving (0.0 cm) radial growth. The results of the effects of the amino acids on radial growth indicated that they were all highly effective, giving complete (0.0 cm) at a higher concentration (100mg/ml). The results of the effect on mycelial weights showed that the oils of the three plants, as well as that of clove, gave complete inhibition (0.0 mg weight); in contrast, the extracts of leaf of Hargel, Parsley, and Basil showed (1.74, 3.26 and 3.34, mg respectively) while, the extracts of the stems gave (3.29, 1.86 and 1.86, mg respectively). Among the amino acids tested, only methionine, leucine, and arginine were adequate (7.0, 7.4, and 7.4 mg, respectively). Concerning the effects of the spore germination, it was found that all the oils tested gave complete inhibition (0.0 %, pore germination) and the stem extracts of the three plants. However, only the leaf extracts of Parsley were highly influential; those of Harjel and Rehan were less effective (7.69% and 15.0 % germination, respectively). All the amino acids tested were less effective, giving about (10.0 % germination). The leaf and stem extracts of Harjel and both Parsley and Clove oils inhibited aflatoxin production by the fungus at all concentrations. The stem extracts of Rehan, the leaf extracts of Parsley, and the oil of Harjel inhibited aflatoxin production at higher concentrations only. The stem extracts and Rehan's oil were found to be unable to inhibit aflatoxin production at all tested concentrations. The chemical analysis of the tested plants' extracts and oils revealed that they contained saponins, tannins, flavonoids, alkaloids, and phenolics but not Steroids. However, glutamic acid was the only amino acid that inhibited aflatoxin production at all concentrations. From the results, it could be concluded that the oils of the plants were more effective than the extracts, and the leaf and stem extracts of Parsley were better among the extracts. It could be recommended that more plant extracts be tested since some plants may be more effective.

Cocoa-associated filamentous fungi for the biocontrol of aflatoxigenic Aspergillus flavus.

Aflatoxin and other mycotoxin contamination are major threats to global food security and present an urgent need to secure the global food crop against spoilage by mycotoxigenic fungi. Cocoa material is noted for naturally low aflatoxin contamination. This study was designed to assess the potential for harnessing cocoa-associated filamentous fungi for the biocontrol of aflatoxigenic Aspergillus flavus. The candidate fungi were isolated from fermented cocoa beans collected from four cocoa-growing areas in Ghana. Molecular characterization included Internal Transcribed Spacer (ITS)-sequencing for identification and polymer chain reaction (PCR) to determine mating type. Effects of the candidate isolates on growth and aflatoxin-production by an aflatoxigenic A. flavus isolate (BANGA1) were assessed. Aflatoxin production was monitored by UV fluorescence and quantified by enzyme-linked immunosorbent assay (ELISA). Thirty-six filamentous fungi were cultured and identified as Aspergillus, Cladosporium, Lichtheimia, or Trichoderma spp. isolates. The isolates generally interacted negatively with BANGA1 growth and aflatoxin production. The Aspergillus niger and Aspergillus aculeatus biocontrol candidates showed the strongest colony antagonism (54%-94%) and reduction in aflatoxin production (12%-50%) on agar. In broth, the A. niger isolates reduced aflatoxin production by up to 97%. Metabolites from the A. niger isolates showed the strongest inhibition of growth by BANGA1 and inhibited aflatoxin production. Four of the candidate isolates belonged to the MAT1-1 mating type and 12 identified as MAT1-2. This may be indicative of the potential for genetic recombination events between fungi in the field, a finding which is particularly relevant to the risk posed by A. flavus biocontrol measures that rely on atoxigenic A. flavus strains.

Inhibition of Aflatoxin Production by Citrinin and Non-Enzymatic Formation of a Novel Citrinin-Kojic Acid Adduct.

Screening for microorganisms that inhibit aflatoxin production from environments showed that Penicillium citrinum inhibited aflatoxin production by Aspergillus parasiticus. The inhibitory substance in the culture medium of P. citrinum was confirmed to be citrinin (CTN). RT-PCR analyses showed that CTN did not inhibit expressions of aflatoxin biosynthetic genes (aflR, pksL1, and fas-1) of A. parasiticus, whereas feeding experiments using A. parasiticus showed that CTN inhibited the in vivo conversion of dihydrosterigmatocystin to AFB2·AFG2. These results suggest that CTN inhibits a certain post-transcriptional step in aflatoxin biosynthesis. CTN in the culture medium of A. parasiticus was found to be decreased or lost with time, suggesting that a certain metabolite produced by A. parasiticus is the cause of the CTN decrease; we then purified, characterized, and then analyzed the substance. Physico-chemical analyses confirmed that the metabolite causing a decrease in CTN fluorescence was kojic acid (KA) and the resulting product was identified as a novel substance: (1R,3S,4R)-3,4-dihydro-6,8-dihydroxy-1-(3-hydroxy-6-(hydroxymethyl)-4-oxo-4H-pyran-2-yl)-3,4,5-trimethyl-1H-isochromene-7-carboxylic acid, which was named "CTN-KA adduct". Our examination of the metabolites' toxicities revealed that unlike CTN, the CTN-KA adduct did not inhibit aflatoxin production by A. parasiticus. These results indicate that CTN's toxicity was alleviated with KA by converting CTN to the CTN-KA adduct.

Chemical Composition and Bioactive Potential of Extracts from Diospyros capricornuta F. White against Aspergillus flavus and Aspergillus parasiticus

Diospyros capricornuta is an endemic species widely distributed along the coast of Tanzania that is used as food condiments and traditional medicine. The chemical compositions of Diospyros capricornuta leaves, stem-bark, and root-bark extracts; and their bioactive potentials against Aspergillus flavus and Aspergillus parasiticus were investigated. The leaves, stem-bark, and root-bark samples of D. capricornuta were extracted using Soxhlet apparatus and the resultant extracts were analyzed using Gas Chromatography-Mass Spectrometry (GC-MS). A total of 14 compounds were identified from the extracts, whereby 2,4-di-tert-butylphenol was the most abundant compound in all extracts. The growth and aflatoxin production inhibitions against A. flavus and A. parasiticus were determined via antifungal and antiaflatoxigenic bioassays of the extracts at the concentrations of 0.0, 62.5, 125.0, and 250.0 µg/mL using a poisoned-food method. The High-Performance Liquid Chromatography (HPLC) technique was used to quantify the aflatoxins after bioassays to evaluate aflatoxin inhibitions. The stem-bark extracts at the highest dose of 250.0 µg/mL inhibited aflatoxin production by A. flavus for over 99% and A. parasiticus for over 94%. Overall, the results show that the leaves, stem-bark, and root-bark extracts of D. capricornuta are potential inhibitors against A. flavus and A. parasiticus-the producers of aflatoxins.
 Keywords: Diospyros capricornuta; Growth inhibitions; Aflatoxin inhibitions; Aspergillus flavus; and Aspergillus parasiticus.

Intraspecific Growth and Aflatoxin Inhibition Responses to Atoxigenic Aspergillus flavus: Evidence of Secreted, Inhibitory Substances in Biocontrol.

The fungus Aspergillus flavus infects corn, peanut, and cottonseed, and contaminates seeds with acutely poisonous and carcinogenic aflatoxin. Aflatoxin contamination is a perennial threat in tropical and subtropical climates. Nonaflatoxin-producing isolates (atoxigenic) are deployed in fields to mitigate aflatoxin contamination. The biocontrol competitively excludes toxigenic A. flavus via direct replacement and thigmoregulated (touch) toxin inhibition mechanisms. To understand the broad-spectrum toxin inhibition, toxigenic isolates representing different mating types and sclerotia sizes were individually cocultured with different atoxigenic biocontrol isolates. To determine whether more inhibitory isolates had a competitive advantage to displace or touch inhibit toxigenic isolates, biomass accumulation rates were determined for each isolate. Finally, to determine whether atoxigenic isolates could inhibit aflatoxin production without touch, atoxigenic isolates were grown separated from a single toxigenic isolate by a membrane. Atoxigenic isolates 17, Af36, and K49 had superior abilities to inhibit toxin production. Small (<400 µm) sclerotial, Mat1-1 isolates were not as completely inhibited as others by most atoxigenic isolates. As expected for both direct replacement and touch inhibition, the fastest-growing atoxigenic isolates inhibited aflatoxin production the most, except for atoxigenic Af36 and K49. Aflatoxin production was inhibited when toxigenic and atoxigenic isolates were grown separately, especially by slow-growing atoxigenic Af36 and K49. Additionally, fungus-free filtrates from atoxigenic cultures inhibited aflatoxin production. Toxin production inhibition without direct contact revealed secretion of diffusible chemicals as an additional biocontrol mechanism. Biocontrol formulations should be improved by identifying isolates with broad-spectrum, high-inhibition capabilities and production of secreted inhibitory chemicals.

Cross-genus inhibitory activity of polyoxins against aflatoxin production by Aspergillus parasiticus and fumonisin production by Fusarium fujikuroi.

Co-exposure to aflatoxin and fumonisin is a health concern where corn is a staple food, and a method to prevent co-contamination of these mycotoxins in foods is urgently needed. Polyoxins are chitin synthase inhibitors produced by Streptomyces cacaoi var. asoensis. The aflatoxin production inhibitory activity of a commercially available polyoxin D and four polyoxins purified from polyoxin AL water-soluble powder, an agricultural chemical containing polyoxins, was tested. The five polyoxins dose-dependently inhibited aflatoxin production by Aspergillus parasiticus and the IC50 values of polyoxin A, B, D, K and L were 16, 74, 110, 9 and 280µmol L-1, respectively. Polyoxins also inhibited fumonisin production by Fusarium fujikuroi, and the IC50 values of polyoxin B, D, K and L were 270, 42, 65 and 62µmol L-1, respectively. Polyoxins repressed the transcription of genes encoding proteins required for aflatoxin biosynthesis in A. parasiticus and fumonisin biosynthesis in F. fujikuroi. Polyoxin K and D also inhibited conidiation in A. parasiticus and F. fujikuroi, respectively. These results suggest that a mixture of polyoxins may effectively prevent co-contamination of aflatoxin and fumonisin in foods.

SUPPRESSION OF AFLATOXINS PRODUCTION IN ARTIFICIALLY INFESTED MAIZE GRAINS WITH ASPERGILLUS FLAVUS DURING STORAGE CONDITIONS

Maize is the one of important crops in Egypt. Aflatoxins (AFs) are the foremost cancer present compounds by Aspergillus flavus (A. flavus) and cause health risks to human and animals. This study aimed to suppression of aflatoxins production by A. flavus by using different concentrations of natural substances (carnation oil, lemongrass oil, propolis and beewax) and chemical substances (salicylic acid and potassium sorbate) on maize grains. The strains of A. flavus were isolated from local maize grains on Potato Dextrose Agar (PDA) and detect its ability of aflatoxins production on coconut agar media. Samples (100g) of sterilized maize grains were treated individually with different concentration of carnation oil, lemongrass oil, salicylic acid and potassium sorbate (0.25, 0.5, 1.0, 2.0, 4.0 and 6.0%) and at concentration 1% and 4% for propolis and beeswax each, then inoculated with A. flavus and stored for 30 days at 28±2˚C. AFs reduction was determined by using High Performance liquid Chromatography (HPLC). All the tested substances had active effect in inhibition of AFs production by A. flavus in stored maize grains. The production of Aflatoxin B1 and B2 (AFB1 and AFB2) was decreased to about 93% and 99% at concentration of 0.25% carnation oil. Lemongrass oil almost completely inhibited AFB1and AFB2 production (99.12% - 99.98% and 99.98% -99.99%, respectively) at concentration of 2% - 6%. Potassium sorbate and salicylic acid (0.25%- 6%) that were significantly effective controlling aflatoxins production on maize grains compared with control. While, the propolis and beewax found to be the most active to protect maize grains against fungi. Natural substances, carnation oil, lemongrass oil, beewax and propolis had higher active effect at low concentration on aflatoxin production more than salicylic acid and more safe for human used.

Synthetic antimicrobial agents inhibit aflatoxin production.

Antimicrobial peptides (AMPs) are biologically active molecules that can eradicate bacteria by destroying the bacterial membrane structure, causing the bacteria to rupture. However, little is known about the extent and effect of AMPs on filamentous fungi. In this study, we synthesized small molecular polypeptides by an inexpensive heat conjugation approach and examined their effects on the growth of Aspergillus flavus and its secondary metabolism. The antimicrobial agents significantly inhibited aflatoxin production, conidiation, and sclerotia formation in A. flavus. Furthermore, we found that the expression of aflatoxin structural genes was significantly inhibited, and the intracellular reactive oxygen species (ROS) level was reduced. Additionally, the antimicrobial agents can change membrane permeability. Overall, our results demonstrated that antimicrobial agents, safe to mammalian cells, have an obvious impact on aflatoxin production, which indicated that antimicrobial agents may be adopted as a new generation of potential agents for controlling aflatoxin contamination.

Inhibition analysis of aflatoxin by in silico targeting the thioesterase domain of polyketide synthase enzyme in Aspergillus ssp

The spread of fungal growth causes enormous economic, agricultural, and health problems for humans, such as Aspergillus sp., which produce aflatoxins. Thus, the inhibition of aflatoxin production became a precious target. In this research, the thioesterase (TE) domain from Polyketide synthase enzyme was selected to employ the in silico docking, using AutoDock Vina, against 623 natural compounds from the South African natural compound database (SANCDB), to identify potential inhibitors that can selectively inhibit thioesterase domain. The top ten inhibitors components were pinocembrin, typhaphthalide, p-coumaroylputrescine, dilemmaone A, 9-angelylplatynecine, 2,4,6-octatrienal, 4,8-dichloro-3,7-dimethyl-, (2e,4z,6e)-, lilacinobiose, 1,3,7-octatriene, 5,6-dichloro-2-(dichloromethyl)-6-methyl-, [r*,s*-(e)]-(-)- (9ci), lilacinobiose, 1,3,7-octatriene, 5,6-dichloro-2-(dichloromethyl)-6-methyl-, [r*,s*-(e)]-(-)- (9ci), 1,3,7-octatriene, 1,5,6-trichloro-2-(dichloromethyl)-6-methyl-, [r*,s*-(z,e)] and 9-angelylhastanecine and that depending on the lowest binding energy, the best chemical interactions and the best drug-likeness. The results of those components gave successful inhibition with the thioesterase domain. So, they can be used for inhibition and controlling aflatoxin contamination of agriculture crop yields, specially, pinocembrin which gave promising results. Communicated by Ramaswamy H. Sarma

Dioctatin Activates ClpP to Degrade Mitochondrial Components and Inhibits Aflatoxin Production

Aflatoxin contamination of crops is a serious problem worldwide. Utilization of aflatoxin production inhibitors is attractive, as the elucidation of their modes of action contributes to clarifying the mechanism of aflatoxin production. Here, we identified mitochondrial protease ClpP as the target of dioctatin, an inhibitor of aflatoxin production of Aspergillus flavus. Dioctatin conferred uncontrolled caseinolytic capacity on ClpP of A.flavus and Escherichia coli. Dioctatin-bound ClpP selectively degraded mitochondrial energy-related proteins invitro, including a subunit of respiratory chain complex V, which was also reduced by dioctatin in a ClpP-dependent manner invivo. Dioctatin enhanced glycolysis and alcohol fermentation while reducing tricarboxylic acid cycle metabolites. These disturbances were accompanied by reduced histone acetylation and reduced expression of aflatoxin biosynthetic genes. Our results suggest that dioctatin inhibits aflatoxin production by inducing ClpP-mediated degradation of mitochondrial energy-related components, and that mitochondrial energy metabolism functions as a key determinant of aflatoxin production.

Inhibitory effect of Capsicum chinense and Piper nigrum fruits, capsaicin and piperine on aflatoxins production in Aspergillus parasiticus by downregulating the expression of aflD, aflM, aflR, and aflS genes of aflatoxins biosynthetic pathway

Aflatoxins produced by Aspergillus parasiticus are toxic and carcinogenic metabolites. The biosynthesis of this mycotoxins is a complex process and involves at least 30 genes clustered within an approximately 82 kB gene cluster. In the present study, the effect of Capsicum chinense and Piper nigrum fruits on Aspergillus parasiticus growth and aflatoxin production were studied in relation to the expression of aflD, aflM, aflR, and aflS four; key genes of aflatoxins biosynthesis pathway. GC-EIMS analysis identified capsaicin (66,107 µg g−1) and piperine (1,138 µg g−1) as the most abundant compounds in C. chinense and P. nigrum fruits, respectively. The antifungal and anti-aflatoxigenic assays showed that C. chinense, P. nigrum, capsaicin, and piperine inhibited A. parasiticus growth and aflatoxins production in a dose-dependent manner. The piperine at 300 µg mL−1 produced higher radial growth inhibition (89%) and aflatoxin production inhibition (69%). The expression of aflatoxin biosynthetic genes was evaluated by quantitative real-time PCR (qRT-PCR) and revealed that aflatoxin inhibition occurring via downregulating the aflS and aflR, and subsequently aflD and aflM genes. These results will improve our understanding of the mechanism of aflatoxin regulation by C. chinense, P. nigrum, capsaicin, and piperine, and provides a reference for further study.

GARLIC (Allium sativum L.) ESSENTIAL OIL AGAINST GROWTH AND AFLATOXIN PRODUCTION OF Aspergillus parasiticus

&lt;p&gt;&lt;strong&gt;Background: &lt;/strong&gt;&lt;em&gt;Aspergillus parasiticus&lt;/em&gt; is a mold to produce aflatoxins, and growth in maize; the used&lt;strong&gt; &lt;/strong&gt;synthetic fungicides for their inhibition are risk to health and the environment. However, garlic essential oil to have antifungal activity and can be alternative for control &lt;em&gt;A. parasiticus&lt;/em&gt;. &lt;strong&gt;Objective/hypothesis:&lt;/strong&gt; Essential garlic oil can inhibit the &lt;em&gt;in vitro&lt;/em&gt; growth of &lt;em&gt;A. parasiticus&lt;/em&gt; and its aflatoxins. &lt;strong&gt;Methodology:&lt;/strong&gt; The &lt;em&gt;in vitro&lt;/em&gt; antifungal activity of garlic (&lt;em&gt;Allium sativum&lt;/em&gt; L.) essential oil was evaluated against &lt;em&gt;A. parasiticus &lt;/em&gt;(radial growth and spore germination). In addition, the mean inhibitory concentrations (IC&lt;sub&gt;50&lt;/sub&gt;) and minimum inhibitory (MIC) were calculated. Aflatoxins were tested on maize with High-Performance Thin-Layer Chromatography (HPTLC). &lt;strong&gt;Results:&lt;/strong&gt; The mean inhibitory concentration (IC&lt;sub&gt;50&lt;/sub&gt;) and minimum inhibitory (MIC) obtained by Probit analyses with radial growth were 971 and 2875 ppm of garlic essential oil, respectively, in &lt;em&gt;A. parasiticus&lt;/em&gt;. These concentrations inhibited spore germination until 24 h. Starting from 100 ppm of garlic essential oil, mycelial inhibition was shown. Evidence of inhibition of aflatoxin production by &lt;em&gt;A. parasiticus&lt;/em&gt; in maize was not possible at concentrations of 971 and 2875 ppm garlic essential oil. &lt;strong&gt;Implications:&lt;/strong&gt; The concentrations 971 and 2875 ppm of garlic essential oil inhibited the growth &lt;em&gt;in vitro&lt;/em&gt; of &lt;em&gt;A. parasiticus&lt;/em&gt;. However, they did not inhibit aflatoxin production by &lt;em&gt;A. parasiticus&lt;/em&gt; into maize. &lt;strong&gt;Conclusion:&lt;/strong&gt; Garlic essential oil inhibit the &lt;em&gt;in vitro&lt;/em&gt; growth of &lt;em&gt;A. parasiticus&lt;/em&gt; on growth radial and germination spores; but not inhibit aflatoxins production on maize to concentrations 971 and 2875 ppm of garlic essential oil.&lt;/p&gt;

A Chemical Approach Against Aspergillus paraciticus and Aflatoxins Production for Feed Safety During Storage

The anti-aflatoxigenic activity of citric acid, acetic acid, ascorbic acid and sodium bicarbonate were investigated in Aspergillus parasiticus inoculated broiler feed. The commercial broiler feed was treated with these synthetic additives (1%, 3% and 5% w/w) and stored for the period of six months at ambient conditions of 16% moisture and 28oC. Aflatoxins (B1, B2, G1 and G2) levels were analyzed at the end of each month by HPLC technique and compared with control. Sodium bicarbonate (1%) inhibited aflatoxins production up to 100% during six months of storage, whereas ascorbic acid (5%) inhibited aflatoxins production up to 4 months. Results revealed that Sodium bicarbonate is a potential source of anti-aflatoxigenic activity, which could be employed to inhibit the aflatoxins production of feed and other commodities during storage.

Illicium verum essential oil, a potential natural fumigant in preservation of lotus seeds from fungal contamination

The antifungal properties of Illicium verum essential oil (IV-EO) against a toxigenic strain of Aspergillus flavus (A. flavus) were analyzed in order to determine its use in the preservation of lotus seeds. GC/MS analysis of IV-EO revealed that its main components are trans-anethole (91.32%), D-Limonene (2.0%) and estragole (2.0%). The antifungal activity of IV-EO against A. flavus infection was analyzed in vitro upon A. flavus with the minimum inhibitory concentration (MIC) and minimum fungicidal concentration (MFC) determined to be 2.0 and 4.0 μL/mL, respectively. IV-EO exhibited strong inhibitory effects on the mycelial growth and spore production of A. flavus, and caused a significant reduction in dry mycelium weight and aflatoxin B1 and B2 (AFB1 and AFB2) biosynthesis. Complete inhibition was observed at an IV-EO concentration of 3.6 μL/mL. Analysis in an in vivo (food system) demonstrated that IV-EO inhibited AFB1 and AFB2 accumulation in lotus seeds to varying degrees depending on its concentration, and completely inhibited aflatoxin production at concentrations above 6.0 μL/g. Morphological studies carried out using SEM revealed that ultra-structural changes in the hyphae and conidiophore structures following IV-EO treatment. Therefore, IV-EO has demonstrated its practical efficacy as a new biological fumigant in the preservation of lotus seeds.

Thiosemicarbazone nano-formulation for the control of Aspergillus flavus.

Nanoparticles are widely studied for applications in medical science. In recent years, they have been developed for agronomical purposes to target microbial pest such as bacteria, fungi, and viruses. Nanoparticles are also proposed to limit the use of pesticides, whose abuse is causing environmental impact and human health concerns. In this study, nanoparticles were obtained by using poly-(ε-caprolactone), a polyester chosen for its biocompatibility and biodegradability properties. Poly-(ε-caprolactone) nanoparticles were formulated by using poly(vinyl alcohol) or Pluronic® F127 as non-ionic surfactants, and then loaded with benzophenone or valerophenone thiosemicarbazone, two compounds that inhibit aflatoxin production by Aspergillus flavus. The different types of nanoparticles were compared in terms of size, polydispersity index, morphology, and drug loading capacity. Finally, their effects were investigated on growth, development, and aflatoxin production in the aflatoxigenic species Aspergillus flavus, a ubiquitous contaminant of maize, cereal crops, and derived commodities. Aflatoxin production was inhibited to various extents, but the best inhibitory effect was obtained with respect to sclerotia production that was most effectively suppressed by both benzophenone and valerophenone thiosemicarbazone-loaded nanoparticles. These data support the idea that it is possible to use such nanoparticles as an alternate to pesticides for the control of mycotoxigenic sclerotia-forming fungi.

Effects of zinc chelating nutrients on Aflatoxin production in Aspergillus flavus

Aflatoxins are carcinogenic metabolites produced by Aspergillus and Penicillium spp. Aflatoxin contamination of food is a serious health hazard. Some metal ions (such as Zn2+) affect Aspergillus growth and aflatoxin biosynthesis. Presence of zinc in the growth medium incites aflatoxin production. This study investigates the effect of zinc binding amino acids and peptides on aflatoxin synthesis in indigenous toxigenic Aspergillus species isolated from agro-ecological zones in Northern Iran. Zinc (II) chelating nutrients (such as Histidine (His), Cysteine (Cys), Histidine-Cysteine (His-Cys), and triple peptide (Asn-Cys-Ser) were added to the growth medium of toxigenic Aspergillus isolates and incubated at temperature range of 25–40 °C. Aflatoxin production on different culture media was tested using ELISA. Addition of cysteine to Sabouraud dextrose broth (SDB) medium significantly reduced aflatoxin production, which could be related to its zinc chelating property. Aflatoxin production was drastically restrained at high concentration of His, especially in combination with Cys, at high pH values and incubation temperature (pH = 7.5, temperature = 40 °C). Aflatoxin production was low in presence of triple peptide (Asn-Cys-Ser) at concentration of 500 mg/L. From the application perspective, natural zinc chelators can be used as harmless aflatoxin-production inhibitors.

Influence of Neighboring Clonal-Colonies on Aflatoxin Production by Aspergillus flavus.

Aspergillus flavus is an ascomycete fungus that infects and contaminates corn, peanuts, cottonseed, and treenuts with acutely toxic and carcinogenic aflatoxins. The ecological function of aflatoxin production is not well understood; though not phytotoxic, aflatoxin may be involved in resisting oxidative stress responses from infection or drought stress in plants. Observation of aflatoxin stimulation in 48-well plates in response to increasing inoculated wells sparked an investigation to determine if A. flavus volatiles influence aflatoxin production in neighboring colonies. Experiments controlling several culture conditions demonstrated a stimulation of aflatoxin production with increased well occupancy independent of pH buffer, moisture, or isolate. However, even with all wells inoculated, aflatoxin production was less in interior wells. Only one isolate stimulated aflatoxin production in a large Petri-dish format containing eight small Petri dishes with shared headspace. Other isolates consistently inhibited aflatoxin production when all eight Petri dishes were inoculated with A. flavus. No contact between cultures and only shared headspace implied the fungus produced inhibitory and stimulatory gases. Adding activated charcoal between wells and dishes prevented inhibition but not stimulation indicating stimulatory and inhibitory gases are different and/or gas is inhibitory at high concentration and stimulatory at lower concentrations. Characterizing stimulatory and inhibitory effects of gases in A. flavus headspace as well as the apparently opposing results in the two systems deserves further investigation. Determining how gases contribute to quorum sensing and communication could facilitate managing or using the gases in modified atmospheres during grain storage to minimize aflatoxin contamination.

Antifungal activity and inhibition of aflatoxins production by Zingiber officinale Roscoe essential oil against Aspergillus flavus in stored maize grains

ABSTRACT: Essential oils are possible alternatives to the use of synthetic pesticides for control of fungal contamination. Ginger (Zingiber officinale) essential oil (GEO) is known for having antifungal and antiaflatoxigenic properties, but its use as a fumigant in situ has not been studied yet. The aim of this study was to evaluate GEO’s effects upon Aspergillus flavus as a fumigant agent in stored maize grains. The main compounds reported in GEO were α-zingiberene (23.85%) and geranial (14.16%), characterized by gas chromatography-mass spectrometry and nuclear magnetic resonance. The GEO was used as a fumigant in irradiated maize grains in concentrations ranging from 5 to 50 µg/g and the resulting effects were compared to a synthetic antifungal agent (carbendazim and thiram), an antifungal traditionally used for seed treatment. The antifungal efficacy of GEO against A. flavus has been proven in a dose-dependent manner through in situ (maize grains) test. The GEO inhibited aflatoxin production at concentrations 25 and 50 µg/g and controlled fungal growth. Therefore, GEO can be used as an effective and non-toxic alternative to conventional treatments in stored maize grains for the natural control of A. flavus.