Flavus Growth Research Articles

Aspergillus flavus growth and aflatoxin production as influenced by total lipid content during growth and development of cottonseed

ABSTRACTAspergillus flavus infects several food and feed crops, such as corn (Zea mays L.), cotton (Gossypium hirsutum L.), peanuts (Arachis hypogaea L.), and tree nut crops and contaminates the seed with carcinogenic aflatoxins. These susceptible crops contain rich reserves of lipids and fatty acids. The nature of relationship between lipids and the ability of the fungus to infect and produce aflatoxins in mature cottonseed, a protein-rich animal feed, has been addressed previously. In this study, we tracked lipid accumulation in developing cottonseed (15–35 days post-anthesis [DPA]) and also the ability of an aflatoxigenic strain and an isogenic non-aflatoxigenic strain to grow and produce aflatoxins in planta. The aflatoxigenic strain Af-70 green fluorescent protein (GFP) and the isogenic, non-aflatoxigenic strain SRRC 1500 (fungal collections maintained at the Southern Regional Research Center) did not differ much in infection and colonization of cottonseed. The non-aflatoxigenic strain did not produce aflatoxin at all stages of cottonseed development, whereas the aflatoxigenic strain Af-70 GFP produced copious amounts of aflatoxin and it coincided with the increasing levels of lipids, especially in mature cottonseed (30–35 DPA). Fungal growth, as quantified by the GFP expression in the fungus, was highly correlated with toxin production.

To evaluate the effects of secondary metabolites produced by Bacillus subtilis mutant M419‌ against Papilio demoleus L. and Aspergillus flavus

Bacillus subtilis inhibits growth of plant pathogen and causes mortality of plant pests by producing lipopeptide biosurfactans. In this study secondary metabolites produced by Bacillus subtilis mutant M419 was extracted and production of lipopeptide surfactants of the mutant was observed by hemolytic activity and oil spreading techniques. The effects of the biosurfactant on mortality of the first and second instars of Papilio demoleus Linnaeus was evaluated by leaves immersion method at 3600, 1800, 900, 450 and 0mg/l concentrations in laboratory. The LC50 and LC90 values after 4days of contact were measured 1172.145mg/l and 3336.999mg/l, respectively. Results indicated that, these values were 3.8-fold and 1.62-fold lower than that obtained after 2 and 3days, respectively. The crude biosurfactant retained the larvicidal activity even when autoclaved at 121°C for 15min. B. subtilis M419 was able to inhibit considerably Aspergillus flavus growth in a dual culture assay.

Aspergillus flavus growth inhibition by Lactobacillus strains isolated from traditional fermented Kenyan milk and maize products.

Certain strains of lactic acid bacteria have been reported to inhibit fungal growth and may so be potential as biocontrol agents. In this study, 171 LAB strains were isolated from traditional fermented Kenyan milk and maize products and tested against aflatoxin-producing A. flavus fungi. The three LAB strains showing highest antifungal activity were identified as Lactobacillus plantarum. None of the strains were able to completely inhibit fungal growth under conditions favorable for fungi and suboptimal for LAB. These conditions probably reduced the growth and metabolic activity of some LAB isolates, as several growth-related aspects like production of antifungal biomolecules and other metabolites contribute to the inhibiting activity. The results suggest that certain LAB strains could be employed in food to control the growth of aflatoxigenic fungi. Further studies to establish the efficacy of the potential LAB strains in fermented products are in progress.

Inhibition of aflatoxin B1 production by verbascoside and other olive polyphenols

In this study, the effects of pure olive phenolic compounds and olive mill wastewater (OMWW) (after membrane filtration treatments) onAspergillus flavus growth and aflatoxin B1 (AFB1) production, were investigated. Five OMWWs coming from Greek (Lianolia, Koroneiki andAsprolia) and Italian (Cellina di Nardò andCoratina) olive oil cultivars, opportunely filtered using a membrane system, were added at two concentrations (5 and 15%) to growth medium, in order to evaluate their effect onA. flavus growth and AFB1 production. The OMWW fractions treatment, after 6 days of incubation, did not inhibit the fungal growth rate, but at 15% concentration significantly reduced the AFB1 production (ranging from 88 to 100%). A similar approach was used for caffeic acid, hydroxytyrosol, tyrosol and verbascoside, the major pure phenolic compounds identified in OMWW fractions. They were evaluated at increasing doses (10, 50 and 100 µg/ml) following both AFB1 production and fungal growth. At the highest concentration (100 µg/ml) all pure compounds showed a reduction of about 99% of AFB1 production without any influence on fungal growth. This is the first time in which OMWWs and their main phenolics were used in the treatments against AFB1 production. The results obtained could provide possible new strategies for preventing AFB1 food contamination using olive polyphenols and OMWW fractions with anti-aflatoxigenic effect, and permitting to harness in a sustainable way an olive oil by-product.

بررسی تاثیر تلقیح کپک Aspergillus flavus بر تغییرات میزان اسیدهای چرب، اندیس پراکسید و تولید آفلاتوکسین در چهار واریته رایج بادام زمینی برداشت شده از سطح مزارع استان گلستان

بادام‌زمینی یکی از حساس ترین محصولات کشاورزی در برابر هجوم A. flavus و متعاقبا آلودگی به آفلاتوکسین محسوب می شود. در این پژوهش پس از جداسازی و تائید شناسایی A. flavus از نمونه های بادام‌زمینی آلوده شده به این کپک، مقادیر مشخصی از آن به چهار واریته بادام‌زمینی رایج استان گلستان به نام های گلی، محلی، چینی و هندی که در شرایط مناسب خشک و پوست‌گیری شدند تلقیح گردید. سپس تاثیر تلقیح کپک مذکور (پس از یک هفته رشد در دمای °C 26) بر روی تغییرات اسیدهای چرب اولئیک و لینولئیک، اندیس پراکسید و تولید آفلاتوکسین مورد بررسی قرار گرفت. بمنظور ارزیابی تغییرات اسیدهای چرب از کروماتوگرافی گازی و جهت بررسی میزان آفلاتوکسین B1 از الایزای رقابتی مستقیم استفاده شد. جهت ارزیابی میزان حساسیت ارقام بادام‌زمینی به رشد A. flavus آفلاتوکسین زا نیز میزان تشکیل پرگنه کپک بر روی مغز بادام‌زمینی و میزان اسپور تولیدی محاسبه گردید. نتایج حاصل، نشان داد که در بین واریته های مختلف، واریته گلی و واریته هندی به ترتیب کمترین و بیشترین حساسیت را به رشد A. flavus و تولید آفلاتوکسین B1 دارا بودند. همچنین با رشد A. flavus بر روی مغز بادام‌زمینی، بطور معنی داری (5% = α) میزان اسیدهای چرب اولئیک و لینولئیک، کاهش و اندیس پراکسید افزایش یافت.

Inhibition of the Aspergillus flavus Growth and Aflatoxin B1 Contamination on Pistachio Nut by Fengycin and Surfactin-Producing Bacillus subtilis UTBSP1.

In this study, the treatment of pistachio nuts by Bacillus subtilis UTBSP1, a promising isolate to degrade aflatoxin B1 (AFB1), caused to reduce the growth of Aspergillus flavus R5 and AFB1 content on pistachio nuts. Fluorescence probes revealed that the cell free supernatant fluid from UTBSP1 affects spore viability considerably. Using high-performance liquid chromatographic (HPLC) method, 10 fractions were separated and collected from methanol extract of cell free supernatant fluid. Two fractions showed inhibition zones against A. flavus. Mass spectrometric analysis of the both antifungal fractions revealed a high similarity between these anti-A. flavus compounds and cyclic-lipopeptides of surfactin, and fengycin families. Coproduction of surfactin and fengycin acted in a synergistic manner and consequently caused a strong antifungal activity against A. flavus R5. There was a positive significant correlation between the reduction of A. flavus growth and the reduction of AFB1 contamination on pistachio nut by UTBSP1. The results indicated that fengycin and surfactin-producing B. subtilis UTBSP1 can potentially reduce A. flavus growth and AFB1 content in pistachio nut.

Heavy metal tolerant Pseudomonas protegens isolates from agricultural well water in northeastern Algeria with plant growth promoting, insecticidal and antifungal activities

The application of plant growth promoting bacteria (PGPB) with biocontrol activities as inoculants of crop plants against phytopathogenic fungi and insect pests provide a biological alternative to the use of agrochemicals. Two Pseudomonas protegens strains were isolated from agricultural well water in a heavy metal contaminated area near Bejaia, northeastern Algeria. The isolates S4LiBe and S5LiBe had 16S rRNA gene sequence similarities of 99.4%–99.7% with P. protegens CHA0T and other P. protegens strains. The phenotypic profiles tested with BIOLOG-GN2-microplates showed differences in 12 of 95 carbon sources tested, as compared to the type strain P. protegens CHA0T. The isolates S4LiBe and S5LiBe showed plant growth promoting potential which is commonly associated with the production of the phytohormone indole acetic acid and siderophores and the solubilization of insoluble phosphate. In addition, they produce chitinase and other polymer degrading enzymes. As the strains S4LiBe and S5LiBe were isolated from heavy metal polluted well water, they are resistant against several heavy metals (2.0 mM K2Cr2O7 and 3.0 mM CoSO4, HgSO4, CdSO4 8H2O and PbCl2), while the reference strain P. protegens CHA0T was very sensitive to Hg2+ and Cd2+ and had lower tolerance towards Co2+ and Pb2+. The isolates S4LiBe and S5LiBe were active in mycelial growth inhibition assays against Botrytis cinerea, Verticillium dahliae, Fusarium graminearum, Aspergillus niger and Aspergillus flavus (growth inhibition between 88% and 48%). Furthermore, S4LiBe and S5LiBe showed effective insecticidal activities, when tested in the Galleria injection assay and they were tested positive for the insect toxin gene fitD alike the reference strain CHA0T. Finally, inoculation of barley seeds with S5LiBe in non-polluted agricultural soil significantly stimulated the germination rate and growth of seedlings, with increased shoot length (11.96 cm ± 0.59), shoot and root fresh weight (0.10 g ± 0.009, 0.04 g ± 0.006), shoot and root dry weight (0.075 g ± 0.003, 0.03 g ± 0.007) as compared to non-inoculated plants (10.23 cm ± 0.84, 0.06 g ± 0.007, 0.025 g ± 0.006, 0.047 g ± 0.006, and 0.016 g ± 0.004, respectively). In heavy metal contaminated soil, inoculation with strain S5LiBe resulted in similar increase of germination rate and growth parameters of barley like in the non-polluted soil, while P. protegens CHA0T inoculated plants were not stimulated. Thus, the heavy metal tolerant isolates S4LiBe and S5LiBe have a potential as beneficial bacteria for agricultural application even in heavy metal polluted soils, e.g. for the stimulation of biomass crops. The demonstration of successful isolation from agricultural well water may open more ready access for a wide variety of this kind of beneficial bacteria for agricultural application.

Description of Aspergillus flavus growth under the influence of different factors (water activity, incubation temperature, protein and fat concentration, pH, and cinnamon essential oil concentration) by kinetic, probability of growth, and time-to-detection models

A Box-Behnken design was used to determine the effect of protein concentration (0, 5, or 10g of casein/100g), fat (0, 3, or 6g of corn oil/100g), aw (0.900, 0.945, or 0.990), pH (3.5, 5.0, or 6.5), concentration of cinnamon essential oil (CEO, 0, 200, or 400μL/kg) and incubation temperature (15, 25, or 35°C) on the growth of Aspergillus flavus during 50days of incubation. Mold response under the evaluated conditions was modeled by the modified Gompertz equation, logistic regression, and time-to-detection model. The obtained polynomial regression models allow the significant coefficients (p<0.05) for linear, quadratic and interaction effects for the Gompertz equation's parameters to be identified, which adequately described (R2>0.967) the studied mold responses. After 50days of incubation, every tested model system was classified according to the observed response as 1 (growth) or 0 (no growth), then a binary logistic regression was utilized to model A. flavus growth interface, allowing to predict the probability of mold growth under selected combinations of tested factors. The time-to-detection model was utilized to estimate the time at which A. flavus visible growth begins. Water activity, temperature, and CEO concentration were the most important factors affecting fungal growth. It was observed that there is a range of possible combinations that may induce growth, such that incubation conditions and the amount of essential oil necessary for fungal growth inhibition strongly depend on protein and fat concentrations as well as on the pH of studied model systems. The probabilistic model and the time-to-detection models constitute another option to determine appropriate storage/processing conditions and accurately predict the probability and/or the time at which A. flavus growth occurs.

Identification of hydrolytic activities expressed by Aspergillus flavus grown on cotton carpel tissue

To identify proteins important for invasion into host plant tissue, Aspergillus flavus was cultured on medium containing cotton carpel tissue as the sole carbon source. We identified several hydrolases suggesting they are important as A. flavus virulence factors for plant colonization. Specifically, Aspergillus flavus AF13 secreted at least two endoxylanase activities and a pectolytic activity when grown on the cotton carpel tissue medium. A concentrated sample derived from the A. flavus growth medium (6-day) was subjected to gel filtration chromatography on a BioGel P-30 column. A major endoxylanase activity was separated from the other fungal-secreted proteins. Additional fungal secreted proteins were partially resolved by gel filtration chromatography on a BioGel P-60 column. Multiple proteins with molecular weights in the 20 to 70 kD range were present in the harvested fungal growth medium. Analysis of these fungal-secreted proteins by liquid chromatography/tandem mass spectrometry identified both endo- and exo-glucanase proteins, α-l-arabinofuranosidase, glucoamylase, α-amylase A, pectate lyase A, xylanase F1, acetylxylan esterase, glutaminase A, as well as conserved hypothetical proteins of unknown function. These proteins likely assist A. flavus in the maceration of plant cell walls, allowing for pathogenic entry and accession of host nutrient resources. Pectolytic and xylanolytic hydrolases, as well as glucanases, appear to be important A. flavus virulence factors.

Effects of temperature, water activity and incubation time on fungal growth and aflatoxin B1 production by toxinogenic Aspergillus flavus isolates on sorghum seeds

Sorghum, which is consumed in Tunisia as human food, suffers from severe colonization by several toxigenic fungi and contamination by mycotoxins. The Tunisian climate is characterized by high temperature and humidity that stimulates mold proliferation and mycotoxin accumulation in foodstuffs. This study investigated the effects of temperature (15, 25 and 37°C), water activity (aw, between 0.85 and 0.99) and incubation time (7, 14, 21 and 28d) on fungal growth and aflatoxin B1 (AFB1) production by three Aspergillus flavus isolates (8, 10 and 14) inoculated on sorghum grains. The Baranyi model was applied to identify the limits of growth and mycotoxin production. Maximum diameter growth rates were observed at 0.99aw at 37°C for two of the isolates. The minimum aw needed for mycelial growth was 0.91 at 25 and 37°C. At 15°C, only isolate 8 grew at 0.99aw. Aflatoxin B1 accumulation could be avoided by storing sorghum at low water activity levels (≤0.91aw). Aflatoxin production was not observed at 15°C. This is the first work on the effects of water activity and temperature on A. flavus growth and AFB1 production by A. flavus isolates on sorghum grains.

Estimation of Aspergillus flavus Growth under the Influence of Different Formulation Factors by Means of Kinetic, Probabilistic, and Survival Models

A Box-Behnken design was conducted to determine the effect of casein concentration (0, 5, or 10%), corn oil (0, 3, or 6%), aw (0.900, 0.945, or 0.990), pH (3.5, 5.0, or 6.5), concentration of cinnamon essential oil (CEO: 0, 200, or 400ppm), and incubation temperature (15, 25, or 35°C) on the growth of A. flavus during 50 days of incubation. Potato dextrose agars were adjusted to the different levels of tested factors and poured into Petri dishes, once solidified were inoculated with mold spores and incubated at studied temperatures. Mold response was modeled using Gompertz and quadratic polynomial equations. The obtained polynomial regression model (allowed the significant (p<0.05) for linear, quadratic, and interaction effects for the Gompertz equation coefficients’ parameters to be identified) adequately described (R2>0.97) mold growth. Additionally, in order to describe growth/not-growth boundary, collected data after 50 days of incubation were classified according to the observed response as 1 (growth) or 0 (not growth), then a binary logistic regression was used to model growth interface. Mold growth probability strongly depend on casein, oil, temperature, and aw, as well as variations of pH and CEO concentration, being lower for those systems with higher content of CEO (>180ppm). Furthermore, survival analysis using failure time was utilized to estimate the time at which mold growth began. The time to fail was directly related to the temperature and CEO concentration; for systems formulated with more than 200ppm of CEO, time to fail was >30 days for low protein and fat contents. The three tested approaches to describe A. flavus response, adequately predicted growth rate and lag time, or growth probability, or the time in which growth will occur. The use and selection of any of these approaches will depend on the intended application.

Inhibition of Aspergillus flavus growth and detoxification of Aflatoxin B1 by the medicinal plant leaf extract- Cymbopogon citratus

Inhibition of Aspergillus flavus growth and detoxification of Aflatoxin B1 by the medicinal plant leaf extract- Cymbopogon citratus

Maize peroxidase Px5 has a highly conserved sequence in inbreds resistant to mycotoxin producing fungi which enhances fungal and insect resistance.

Mycotoxin presence in maize causes health and economic issues for humans and animals. Although many studies have investigated expression differences of genes putatively governing resistance to producing fungi, few have confirmed a resistance role, or examined putative resistance gene structure in more than a couple of inbreds. The pericarp expression of maize Px5 has previously been associated with resistance to Aspergillus flavus growth and insects in a set of inbreds. Genes from 14 different inbreds that included ones with resistance and susceptibility to A. flavus, Fusarium proliferatum, F. verticillioides and F. graminearum and/or mycotoxin production were cloned using high fidelity enzymes, and sequenced. The sequence of Px5 from all resistant inbreds was identical, except for a single base change in two inbreds, only one of which affected the amino acid sequence. Conversely, the Px5 sequence from several susceptible inbreds had several base variations, some of which affected amino acid sequence that would potentially alter secondary structure, and thus enzyme function. The sequence of the maize peroxidase Px5 common to inbreds resistant to mycotoxigenic fungi was overexpressed in maize callus. Callus transformants overexpressing the gene caused significant reductions in growth for fall armyworms, corn earworms, and F. graminearum compared to transformant callus with a β-glucuronidase gene. This study demonstrates rarer transcripts of potential resistance genes overlooked by expression screens can be identified by sequence comparisons. A role in pest resistance can be verified by callus expression of the candidate genes, which can thereby justify larger scale transformation and regeneration of transgenic plants expressing the resistance gene for further evaluation.

Inhibitory Effect of Cinnamaldehyde, Citral, and Eugenol on Aflatoxin Biosynthetic Gene Expression and Aflatoxin B1 Biosynthesis in Aspergillus flavus.

In order to reveal the inhibitory effects of cinnamaldehyde, citral, and eugenol on aflatoxin biosynthesis, the expression levels of 5 key aflatoxin biosynthetic genes were evaluated by real-time PCR. Aspergillus flavus growth and AFB1 production were completely inhibited by 0.80 mmol/L of cinnamaldehyde and 2.80 mmol/L of citral. However, at lower concentration, cinnamaldehyde (0.40 mmol/L), eugenol (0.80 mmol/L), and citral (0.56 mmol/L) significantly reduced AFB1 production with inhibition rate of 68.9%, 95.4%, and 41.8%, respectively, while no effect on fungal growth. Real-time PCR showed that the expressions of aflR, aflT, aflD, aflM, and aflP were down-regulated by cinnamaldehyde (0.40 mmol/L), eugenol (0.80 mmol/L), and citral (0.56 mmol/L). In the presence of cinnamaldehyde, AflM was highly down-regulated (average of 5963 folds), followed by aflP, aflR, aflD, and aflT with the average folds of 55, 18, 6.5, and 5.8, respectively. With 0.80 mmol/L of eugenol, aflP was highly down-regulated (average of 2061-folds), followed by aflM, aflR, aflD, and aflT with average of 138-, 15-, 5.2-, and 4.8-folds reduction, respectively. With 0.56 mmol/L of citral, aflT was completely inhibited, followed by aflM, aflP, aflR, and aflD with average of 257-, 29-, 3.5-, and 2.5-folds reduction, respectively. These results suggest that the reduction in AFB1 production by cinnamaldehyde, eugenol, and citral at low concentration may be due to the down-regulations of the transcription level of aflatoxin biosynthetic genes. Cinnamaldehyde and eugenol may be employed successfully as a good candidate in controlling of toxigenic fungi and subsequently contamination with aflatoxins in practice.

Antifungal properties and inhibitory effects upon aflatoxin production by Zingiber officinale essential oil in Aspergillus flavus

SummaryIn this study, the efficacy of ginger (Zingiber officinale Roscoe) essential oil (GEO) in reducing A. flavus growth and aflatoxin production was investigated. Gas chromatography coupled to mass spectrometry and nuclear magnetic resonance spectroscopy showed that the major components of GEO were α‐zingiberene (23.85%) and geranial (14.16%). Mycelial growth of Aspergillus flavus was reduced significantly at a GEO concentration of 150 μg mL−1, and complete inhibition of conidial germination was observed at a concentration of 10 μg mL−1. Statistically significant inhibition of ergosterol biosynthesis was detected at a GEO concentration of 10 μg mL−1. GEO was capable of fully inhibiting aflatoxin production by A. flavus at a concentration of 15 μg mL−1. The results suggest that low concentrations of GEO are capable of inhibiting aflatoxin production; such ability could be valuable in the upcoming future for agricultural companies to better control aflatoxigenic fungi in agricultural products.

Controlling Aspergillus flavus and Aspergillus parasiticus growth and aflatoxin production in poultry feed using carvacrol and trans-cinnamaldehyde

Aflatoxins (AF) are toxic metabolites primarily produced by molds, Aspergillus flavus and Aspergillus parasiticus. Contamination of poultry feed with AF is a major concern to the poultry industry due to severe economic losses stemming from poor performance, reduced egg production, and diminished egg hatchability. This study investigated the inhibitory effect of 2 generally regarded as safe (GRAS), natural plant compounds, namely carvacrol (CR) and trans-cinnamaldehyde (TC), on A. flavus and A. parasiticus growth and AF production in potato dextrose broth (PDB) and in poultry feed. In broth culture, PDB supplemented with CR (0%, 0.02%, 0.04% and 0.08%) or TC (0%, 0.005%, 0.01% and 0.02%) was inoculated with A. flavus or A. parasiticus (6 log CFU/mL), and mold counts and AF production were determined on days 0, 1, 3, and 5. Similarly, 200 g portions of poultry feed supplemented with CR or TC (0%, 0.4%, 0.8%, and 1.0%) were inoculated with each mold, and their counts and AF concentrations in the feed were determined at 0, 1, 2, 3, 4, 8, and 12 weeks of storage. Moreover, the effect of CR and TC on the expression of AF synthesis genes in A. flavus and A. parasiticus (aflC, nor1, norA, and ver1) was determined using real-time quantitative PCR (RT-qPCR). All experiments had duplicate samples and were replicated 3 times. Results indicated that CR and TC reduced A. flavus and A. parasiticus growth and AF production in broth culture and chicken feed (P < 0.05). All tested concentrations of CR and TC decreased AF production in broth culture and chicken feed by at least 60% when compared to controls (P < 0.05). In addition, CR and TC down-regulated the expression of major genes associated with AF synthesis in the molds (P < 0.05). Results suggest the potential use of CR and TC as feed additives to control AF contamination in poultry feed.

Fungicidal activity and constituents of Ageratum conyzoides essential oil from three regions in São Paulo state, Brazil

The chemical composition and antifungal activity of the Ageratum conyzoidesessential oils, obtained from the leaves collected in Ibiúna (1), Ribeirão Pires (2) and Campinas (3) in the São Paulo state, Brazil, were investigated. The essential oils were obtained from A. conyzoidesleaves by hydrodestilation and analyzed by GC/MS. The chemical composition of the A. conyzoides oil collected in Ribeirão Pires and Ibiúna consisted mainly of precocene I and II. The essential oil from leaves collected in Campinas had only traces of precocene II and a highest proportion of precocene I, α-humulene and (E)-caryophyllene than the oils (1) and (2). The Aspergillus flavus growth was inhibited by essential oils (1) and (2) at 60 and 64%, respectively, and the oil (3) was inactive. On the other hand, the three essential oils inhibited the sporulation of the fungus for more than 120 days. The essential oils of leaves collected in sites that show similarities in the latitude, altitude and average temperatures, as Ribeirão Pires and Ibiúna, showed chemical composition and antifungal activity similar, either, which showed the importance of geo-ecological factors in production of metabolites of the plant.

Chemical Composition and Antifungal Activity of Ocimum basilicum L. Essential Oil.

BACKGROUND:The leaves of Ocimum basilicum L. (basil) are used in traditional cuisine as spices; its essential oil has found a wide application in perfumery, dental products as well as antifungal agents.AIM:To assess the chemical composition as well as the in vitro antifungal activity of O. basilicum L. essential oil against Aspergillus flavus fungal growth and aflatoxin B1 production.MATERIAL AND METHODS:The essential oil of O. basilicum was obtained by hydrodistillation and analysed using gas chromatography (GC) and GC coupled with mass spectrometry (GC/MS). The essential oil was tested for its effects on Aspergillus flavus (A. flavus) mycelial growth and aflatoxin B1 production in Yeast Extract Sucrose (YES) growth media. Aflatoxin B1 production was determined by high performance liquid chromatography (HPLC).RESULTS:Nineteen compounds, representing 96.7% of the total oil were identified. The main components were as follows: linalool (48.4%), 1,8-cineol (12.2%), eugenol (6.6%), methyl cinnamate (6.2%), α-cubebene (5.7%), caryophyllene (2.5%), β-ocimene (2.1%) and α-farnesene (2.0%). The tested oil showed significant antifungal activity that was dependent on the used oil concentration. The complete inhibition of A. flavus growth was observed at 1000 ppm oil concentration, while marked inhibition of aflatoxin B1 production was observed at all oil concentrations tested (500, 750 and 1000 ppm).CONCLUSION:These results confirm the antifungal activities of O. basilicum L. oil and its potential use to cure mycotic infections and act as pharmaceutical preservative against A. flavus growth and aflatoxin B1 production.

Inhibition of Fungal Aflatoxin B1 Biosynthesis by Diverse Botanically-Derived Polyphenols

Purpose: To identify and characterize the capacity of diverse botanically-derived polyphenols to inhibit aflatoxin B1 (AFB1) production by Aspergillus flavus.Methods: A tea-derived polyphenol mixture and numerous individual polyphenols were tested for their effects on A. flavus growth and AFB1 production. Fungal spores were cultured for 60 h with polyphenols (range 0 ‒ 1,000 μg/mL). The fungi were enumerated by hemocytometry, and AFB1 in culture supernatants was quantified by high-performance liquid chromatography (HPLC).Results: Neither the tea-derived polyphenol mixture nor individual polyphenol compound, except quercetin, inhibited A. flavus growth. Quercetin detectably inhibited growth at 800 μg/mL; none of the remaining polyphenols inhibited fungal proliferation, even at 1,000 μg/mL. However, catechin mixture and all individual polyphenols differentially inhibited fungal AFB1 biosynthesis. Non-ester catechin derivatives revealed stronger inhibitory activity than ester derivatives.Conclusion: Quercetin exhibits the strongest inhibitory effect on AFB1 production and is the only test compound that also inhibits fungal proliferation. Botanically-derived polyphenols are, therefore, promising reagents for controlling fungal contamination and associated toxic aflatoxin deposition in harvested crops and in food processing operations.Keywords: Polyphenols, Quercetin, Aflatoxin B1, Inhibition, Antioxidation

An improved simulation model to predict pre-harvest aflatoxin risk in maize

Aflatoxin is a potent carcinogen produced by Aspergillus flavus, which frequently contaminates maize (Zea mays L.) in the field between 40° north and 40° south latitudes. A mechanistic model to predict risk of pre-harvest contamination could assist in management of this very harmful mycotoxin. In this study we describe an aflatoxin risk prediction model which is integrated with the Agricultural Production Systems Simulator (APSIM) modelling framework. The model computes a temperature function for A. flavus growth and aflatoxin production using a set of three cardinal temperatures determined in the laboratory using culture medium and intact grains. These cardinal temperatures were 11.5°C as base, 32.5°C as optimum and 42.5°C as maximum. The model used a low (≤0.2) crop water supply to demand ratio—an index of drought during the grain filling stage to simulate maize crop's susceptibility to A. flavus growth and aflatoxin production. When this low threshold of the index was reached the model converted the temperature function into an aflatoxin risk index (ARI) to represent the risk of aflatoxin contamination. The model was applied to simulate ARI for two commercial maize hybrids, H513 and H614D, grown in five multi-location field trials in Kenya using site specific agronomy, weather and soil parameters. The observed mean aflatoxin contamination in these trials varied from <1 to 7143ppb. ARI simulated by the model explained 99% of the variation (p≤0.001) in a linear relationship with the mean observed aflatoxin contamination. The strong relationship between ARI and aflatoxin contamination suggests that the model could be applied to map risk prone areas and to monitor in-season risk for genotypes and soils parameterized for APSIM.