Growth Of Aspergillus Parasiticus Research Articles

Effects of Tropical Citrus Essential Oils on Growth, Aflatoxin Production, and Ultrastructure Alterations of Aspergillus flavus and Aspergillus parasiticus

Ethyl acetate extracts and hydrodistillated essential oils from five cultivars of tropical citrus epicarps were evaluated for their inhibitory activities against Aspergillus fumigatus, Aspergillus niger, Aspergillus flavus, Aspergillus parasiticus, and Penicillium sp. using disk diffusion and broth microdilution assays. Essential oils prepared from kaffir lime (Citrus hystrix DC) and acid lime (Citrus aurantifolia Swingle) epicarps exhibited stronger antifungal activity to all fungi than their ethyl acetate extracts with minimum inhibitory concentration and minimum fungicidal concentration values of 0.56 and 1.13 mg/ml (dry matter), respectively, against aflatoxin-producing A. flavus and A. parasiticus. The dominant components of the essential oil from kaffir lime were limonene, citronellol, linalool, o-cymene, and camphene, whereas limonene and p-cymene were major components of acid lime essential oil. Pure limonene, citronellal, and citronellol were five to six times less fungicidal than the natural essential oils, indicating the synergistic activity of many active compounds present in the oils. Kaffir and acid lime essential oils significantly reduced aflatoxin production of A. flavus and A. parasiticus, particularly lime essential oil, which completely inhibited growth and aflatoxin production of A. flavus at the concentration of 2.25 mg/ml. Target cell damage caused by acid lime essential oil was investigated under transmission electron microscopy. Destructive alterations of plasma and nucleus membrane, loss of cytoplasm, vacuole fusion, and detachment of fibrillar layer were clearly exhibited in essential-oil-treated cells.

Modelling the effect of temperature and water activity in the growth boundaries of Aspergillus ochraceus and Aspergillus parasiticus

The aim of this work was to model the growth of Aspergillus parasiticus and Aspergillus ochraceus, both mycotoxin producers, near to the growth/no growth boundaries and validate those models in sterile maize grain, peanuts and coffee beans. Malt extract agar was adjusted to six different water activities: 0.93, 0.91, 0.89, 0.87, 0.85 and 0.80. Plates were incubated at 10, 15, 20, 25, 30, 37 and 42 °C. For each of the 42 conditions, 10 Petri dishes were inoculated. Both kinetic and probability models were applied to colony growth data. The results of the present study indicate that the developed probability modelling approach could be satisfactorily employed to quantify the combined effect of temperature and water activity on the growth responses of A. ochraceus and A. parasiticus. However, validation of kinetic results led to poor goodness of prediction. In this study, the validation samples were placed near to the expected boundaries of the models in order to test them under the worst situation. Probability of growth prediction under extreme growth conditions was somewhat compromised, but it can be considered acceptable.

Modelling the effect of temperature and water activity on the growth of Aspergillus parasiticus on irradiated Argentinian flint maize

A full factorially designed experiment including storage temperature (10, 20, 30 and 37 °C) and water activity (0.88, 0.92 and 0.96) was undertaken to study the growth of Aspergillus parasiticus in maize samples. Kinetic parameters such as specific growth rate ( μ), lag phase duration and maximum logarithmic increase were determined by fitting the Modified Gompertz equation to the viable mould count data (N in CFU/g) as a function of time collected in twelve experiments. The average coefficient of determination ( R 2 ) was 0.987, being the mean standard deviation of the estimate of 0.216 in units of log 10 N. In the practical range of 10–30 °C, the relationship of the three kinetic parameters with temperature was described by second order polynomial expressions, whose parameters, in turn, depended on water activity. The combined or full model i.e., the Modified Gompertz model with its parameters expressed as a function of temperature and water activity, was able to predict log 10 N with an average percentage error of 4.3, so agreement with the experimental data was highly satisfactory. In a simulation exercise, the full model was able to predict the viable mould count, given an initial value and grain temperature and water activity histories, with promising results for maize storage.

The effect of Baccharis glutinosa extract on the growth of mycotoxigenic fungi and fumonisin B1 and aflatoxin B1 production

The aim of this study was to evaluate the effect of Baccharis glutinosa isolated extract on the growth of Aspergillus flavus and Aspergillus parasiticus, and their aflatoxin B1 production; and growth of Fusarium verticillioides, and their fumonisin B1 production. The three fungi were exposed to an antifungal fraction, designated as fraction F6-1, isolated from B. glutinosa by methanolic extraction followed by silica gel chromatography. The growth of the fungi was evaluated in kinetics of radial extension growth, kinetics of spores germination, length and diameter of hyphae, spores diameter, as well as in aflatoxin B1 and fumonisin B1 production. Fraction F6-1 caused radial growth inhibition of the three fungi mainly F. verticillioides. Spores germination of A. flavus and A. parasiticus was delayed in the early stage of the incubation time, although they completely germinated at 27 h. In contrast, spore germination of F. verticillioides was inhibited 87.7% up to 96 h. The lengths and diameters of hyphae, and spore diameters of the three fungi, were significantly smaller in comparison with those of the controls, and several morphological alterations were observed. Concerning aflatoxin B1 and fumonisin B1, fraction F6-1 did not show any inhibition effect at the concentration used. Fraction F6-1 was able to significantly inhibit the development of the three fungi, mainly F. verticillioides. The strong inhibitory effect of F6-1 on hyphae and spores suggests that it interacted with the fungi cell walls, which caused severe deformities. Nevertheless, this fraction was unable in inhibiting mycotoxin production from the three fungi at the concentration tested.

Effect of various surfactants (cationic, anionic and non-ionic) on the growth of Aspergillus parasiticus (NRRL 2999) in relation to aflatoxin production

The effect of surfactants (two cationic, one anionic and three non-ionic) at 0.001, 0.01, 0.1 and 1.0 % concentrations on aflatoxin production, ergosterol content and sugar consumption by Aspergillus parasiticus (NRRL 2999) in YES liquid culture medium is reported. At 0.01% concentration, the cationic surfactants, cetyl dimethyl ammonium bromide (CDAB) and dodecyl trimethyl ammonium bromide (DTAB), and the anionic surfactant, sodium dodecyl sulfate (SDS), completely inhibited spore germination, while DTAB also inhibited the production of ergosterol and toxin (p < 0.05). At a concentration of 0.001%, CDAB was found to enhance toxin production, while SDS was found to inhibit it when compared with other surfactants. Non-ionic surfactants, polyoxyethylene sorbitan monolaurate (Tween-20), polyoxyethylene lauryl ether (Brij-35) and ethoxylated p-tert-octylphenol (Triton X-100) delayed the spore germination up to day 5 at all concentrations and inhibited toxin and ergosterol production at 0.001% concentration. The affect was found to be dose-dependent from 0.001% to 1%, for Triton X-100 only. Positive correlation between ergosterol content and toxin production in the presence of different surfactants at various time periods (3, 5, 7, 9 and 12days) was found. Tween-20 was most effective in inhibiting toxin production on day 7, when aflatoxin production was found to be maximal in control group. Sugar consumption was directly proportional to the ergosterol content, showing a significant correlation with aflatoxin production.

Antiaflatoxigenic and antioxidant activity of an essential oil fromAgeratum conyzoidesL.

Aflatoxin contamination of various commodities can occur as a result of infection, mainly by Aspergillus flavus and Aspergillus parasiticus. Every year, almost 25% of the world's food supply is contaminated by mycotoxins. Aflatoxins B(1), B(2), G(1) and G(2), which occur naturally, are significant contaminants of a wide variety of commodities. A number of biological activities have been associated with Ageratum conyzoides. We have therefore investigated the antiaflatoxigenic, antioxidant and antimicrobial activity of essential oils of A. conyzoides. This could help to turn A. conyzoides, a nuisance weed, into a resource. The essential oil of Ageratum conyzoides L. shows the presence of 12 compounds when analyzed by gas chromatography-mass spectrometry. The growth and aflatoxin production of the toxigenic strain Aspergillus parasiticus was completely inhibited by essential oil. All the studied concentrations of the oil demonstrate a reduction in mycelia growth and decreased production of different aflatoxins in fungi, as revealed by liquid chomatographic-tandem mass spectrometric analysis. Volatiles from macerated green leaf tissue of A. conyzoides were also effective against A. parasiticus. The strongest antibacterial activity was observed against the bacteria Staphylococcus aureus and Bacillus subtilis in a disk diffusion bioassay. Essential oil and methanol extract of A. conyzoides L. were assayed for their antioxidant activity. Methanol extract showed the highest antioxidant activity in FRAP and DPPH assay, whereas essential oil showed greater lipid peroxidation inhibition than methanol extract. The plant's ethno-medicinal importance, antioxidant potential, inhibitory activity against the Aspergillus group of fungi and production of aflatoxins may add a new dimension to its usefulness in the protection of stored product.

Inhibitory Effects of Ephedra major Host on Aspergillus parasiticus Growth and Aflatoxin Production

This study was undertaken to evaluate the effect of Ephedra major Host, an important medicinal plant with various biological activities, on growth and aflatoxin (AF) production by Aspergillus parasiticus NRRL 2999. The fungus was cultured in yeast extract-sucrose (YES) broth, a conductive medium that supports AF production, in the presence of various concentrations of essential oil (EO), hexanic and methanolic extracts of plant aerial parts, fruits, and roots using microbioassay technique. After incubating for 96 h at 28 degrees C in static conditions, mycelial dry weight was determined as an index of fungal growth, and aflatoxin B(1) (AFB(1)) was measured using HPLC technique. Based on the obtained results, EO of plant aerial parts significantly inhibited fungal growth at the highest concentration of 1000 microg/ml without any obvious effect on AFB(1) production at all concentrations used. Among plant extracts tested, only methanolic extract of aerial parts and roots were found to inhibit fungal growth and AFB(1) production dose-dependently with an IC(50) value of 559.74 and 3.98 microg/ml for AFB(1), respectively. Based on the GC/MS data, the major components of E. major EO were bis (2-ethylhexyl) phthalate (42.48%), pentacosane (20.94%), docosane (14.64%), citronellol (5.15%), heptadecan (4.41%), cis-3-Hexen-1-ol benzoate (4.07%), and 7-Octen-2-ol (3.25%). With respect to the potent inhibition of fungal growth and AF production by E. major, this plant may be useful in protecting crops from both toxigenic fungal growth and AF contamination.

Fungal Inhibition by Some Spice Essential Oils

The antifungal activities of essential oils of pickling herb (Echinophora tenuifolia spp. sibthorpiana), rosemary (Rosmarinus officinalis L.) and oregano (Origanum vulgare L.) plants growing wild in Turkey were tested against to the mycelial growth of Alternaria alternata, Aspergillus niger and Aspergillus parasiticus using Potato Dextrose Agar (PDA) in vitro. Essential oil of O. vulgare was the most effective on the mycelial growth of A. alternata compared with other two fungi. Oregano oil was the most active against all the moulds tested. Some concentrations of pickling herb and rosemary stimulated the mycelial growth of Aspergillus niger and Aspergillus parasiticus. It was concluded that the fungistatic effects of every oil increased with higher doses. The high fungistatic effect in the beginning of incubation decreased gradually towards the end of incubation. However, knowledge of how to protect the spoilage of food products from pathogens and saprophytic fungi can probably be gained from the different concentrations of spice derivatives.

Chemical composition and antiaflatoxigenic activity of Carum carvi L., Thymus vulgaris and Citrus aurantifolia essential oils

In order to find out plants useful to controlling aflatoxins (AFs) production, the essential oils (EOs) from 12 medicinal plants prepared by hydrodistillation were studied with special reference to the inhibition of Aspergillus parasiticus growth and AFs production. The toxigenic fungus was cultured in presence of various oils in 6-well microplates using a microbioassay technique. The mycelial mass was estimated as an index of fungal growth, while the aflatoxins B 1 (AFB 1) and G 1 (AFG 1) were measured by high performance liquid chromatography (HPLC). Among plants tested, Thymus vulgari and Citrus aurantifolia were found to inhibit both A. parasiticus and AF production. The EOs from Mentha spicata L., Foeniculum miller, Azadirachta indica A. Juss, Conium maculatum and Artemisia dracunculus were only inhibited fungal growth, while Carum carvi L. effectively inhibited AF production without any obvious effect on fungal growth. The other plants including Ferula gummosa, Citrus sinensis, Mentha longifolia and Eucalyptus camaldulensis had no effect on A. parasiticus growth and AF production at all concentrations used. The IC 50 values of T. vulgaris, C. aurantifolia and C. carvi for AF inhibition were reported as 93.5, 285.6, and 621.9 μg/ml for AFB 1, while they were calculated as 11.7, 50.1, and 56.0 μg/ml for AFG 1. These results indicate that the EOs of some medicinal plants may be considered as potential candidates to protect foods and feeds from toxigenic fungus growth and subsequent AF contamination.

Study of the Ochratoxin A effect on Aspergillus parasiticus growth and aflatoxin B 1 production

Aflatoxin B 1 (AFB 1) is a carcinogenic metabolite produced by certain Aspergillus species. Ochratoxin A (OTA) is a metabolite of Aspergillus ochraceus and Penicillium verrucosum. AFB 1 and OTA are amongst the most frequent combinations of mycotoxins found in plant products. Thus, synergistic effects or interactions between the two mycotoxins could be taking place. The aim of the present study was to investigate the effect of OTA on Aspergillus parasiticus growth and AFB 1 production in yeast extract sucrose (YES) medium at concentrations of 0.16, 1.6 and 16 ng OTA flask −1. The AFB 1 extracted from cultures and purified with immunoaffinity columns was then quantitated by HPLC. The recovery and detection limit of the method were 95.3% and 0.02 ng AFB 1 mL −1, respectively. Maximum AFB 1 productions in cultures with OTA were observed from 9 to 12 days (76.09–82.52 ng AFB 1 flask −1) while in control cultures (without OTA) maximum production (197.2 ng AFB 1 flask −1) was observed on 14th day. Maximum AFB 1 levels in cultures with OTA were reduced by a percentage of 58–61% compared to control cultures. Furthermore AFB 1 levels in cultures with OTA were practically (92%) degraded after 18 days of incubation. Conclusively when OTA is present the production of AFB 1 by A. parasiticus in YES medium is inhibited.

Antimycotoxigenic characteristics of Rosmarinus officinalis and Trachyspermum copticum L. essential oils

Aflatoxin B 1 (AFB 1) is a highly toxic and carcinogenic metabolite produced by Aspergillus species on food and agricultural commodities. Natural products may regulate the cellular effects of aflatoxins and evidence suggests that aromatic organic compounds of spices can control the production of aflatoxins. With a view to controlling aflatoxin production, the essential oils from Rosmarinus officinalis and Trachyspermum copticum L. were obtained by hydrodistillation. Antifungal activities of the oils were studied with special reference to the inhibition of Aspergillus parasiticus growth and aflatoxin production. Minimal inhibitory (MIC) and minimal fungicidal (MFC) concentrations of the oils were determined. T. copticum L. oil showed a stronger inhibitory effect than R. officinalis on the growth of A. parasiticus. Aflatoxin production was inhibited at 450 ppm of both oils with that of R. officinalis being stronger inhibitor. The oils were analyzed by GC and GC/MS. The major components of R. officinalis and T. copticum L. oils were Piperitone (23.65%), α-pinene (14.94%), Limonene (14.89%), 1,8-Cineole (7.43%) and Thymol (37.2%), P-Cymene (32.3%), γ-Terpinene (27.3%) respectively. It is concluded that the essential oils could be safely used as preservative materials on some kinds of foods to protect them from toxigenic fungal infections.

Apyap1 affects aflatoxin biosynthesis during Aspergillus parasiticus growth in maize seeds

It is demonstrated that, in fungal cells grown in synthetic media, the Apyap1 gene is implicated in the modulation of aflatoxin biosynthesis following the perturbation of redox balance. This study suggests that an association between oxidative stress and aflatoxin biosynthesis also occurs in maize seeds. We used ΔApyap1, a strain in which the gene Apyap1 was disrupted, to verify whether this oxidative stress-related transcription factor, by affecting cell redox balance, can have a role in the modulation of aflatoxin synthesis. The amount of hydroperoxides (ROOH) produced by wild type (WT) and ΔApyap1, both grown in potato dextrose broth, was assayed in the filtrate. In maize seeds (30 g), inoculated with WT and ΔApyap1conidia and incubated at 30°C for 15 days, lipoxygenase activity (LOX), lipoperoxides (LOOH) production, fungal growth and aflatoxin biosynthesis was analysed. It was observed that ΔApyap1 released more hydroperoxides in the culture media and more aflatoxins in seeds, possibly through stronger stimulation of LOX, which, in turn led to greater LOOH production in the seeds. On the basis of the results, a hypothesis regarding strategies to control aflatoxin synthesis is formulated.

Inhibitory effect of phenolics extracted from sorghum genotypes on Aspergillus parasiticus (NRRL 2999) growth and aflatoxin production

AbstractThe inhibitory activity of bioactive polyphenols present in six sorghum genotypes—two red (AON 486 and IS 620), two yellow (LPJ and IS 17779) and two white (SPV 86 and SPV 462) varieties—on Aspergillus parasiticus (NRRL 2999) growth and aflatoxin production was evaluated. In the first experiment the production of aflatoxins in the six sorghum genotypes after removal of surface phenolics by acidic methanol treatment was studied and compared with that in untreated grains. Aflatoxin production was found to be fourfold higher in treated grains. The total phenols and bioactive polyphenols extracted by acidic methanol were quantified using the Folin–Denis method and the bovine serum albumin–benzidine conjugate procedure respectively. In the second experiment the effect of extracted sorghum phenolics under in vitro conditions on fungal growth and aflatoxin production was studied at two concentrations (0.01% and 0.1%) of phenolics. Extracted phenolics added to yeast extract sucrose (YES) medium at 0.1% concentration showed an inhibitory effect on aflatoxin production. At 0.01% phenolic concentration, aflatoxin production was minimal on day 3 after infection. At other time points the aflatoxin content was similar to that in the control. At 9 days after infection the fungal biomass in IS 620 was significantly lower than that in the control. At 0.1% phenolic concentration, aflatoxin production was minimal and the red genotype IS 620 showed maximum resistance. Fungal biomass was lowest at all growth stages in IS 620 as compared with the control. Polyphenol oxidase (PPO) activity was not detected in A. parasiticus grown on YES medium (control). PPO activity was not induced in A. parasiticus by the addition of phenolics to the liquid culture medium (no PPO activity was detected in the culture medium). The inhibitory activity of bioactive polyphenols could be attributed to the lack of PPO enzyme in this fungus. Copyright © 2007 Society of Chemical Industry

Inhibitory Effects of Akacid®plus on Growth and Aflatoxin Production by Aspergillus parasiticus

The effects of Akacid plus, a novel guanidine-based polymer first introduced as a biocidal and disinfectant agent were studied on Aspergillus parasiticus growth and its aflatoxin (AF) productivity. The fungus was cultured on yeast extract-sucrose (YES) broth in presence of various twofold serial dilutions of 25% Akacid plus (1.5-96 microL/50 mL medium) and then incubated in shaking condition with 150 rev./min at 28 degrees C for 96 h. Based on obtained results, Akacid plus was found to significantly inhibit both growth and aflatoxin B1 (AFB1) synthesis in very low concentrations in a dose-dependent manner. Fungal growth inhibition was determined in the range of 9.6-99.6% in mycelia exposed to the total concentration range of 1.5-48 microL. A final concentration of 96 microL was necessary to completely inhibit the growth of fungus. Under similar conditions, AFB1 synthesis was found to be strongly inhibited by 8.1-98.0% in presence of 1.5-24 microL Akacid plus with a maximum of 100% by 48 microL concentration. With respect to the unique physico-chemical properties of Akacid plus, its marked inhibitory effects on A. parasiticus growth and its AFB1 synthesis shown for the first time in this study make it a promising candidate for application in prevention programmes of AF contamination of susceptible crops.

Chemoprevention by thyme oils of Aspergillus parasiticus growth and aflatoxin production

The essential oils from Thymus eriocalyx and Thymus X-porlock obtained by hydrodistillation were analyzed by GC/MS. The major components of T. eriocalyx and T. X-porlock oils were thymol (63.8, 31.7%), β-phellandrene (13.30, 38.7%), cis-sabinene hydroxide (8.1, 9.6%), 1,8-cineole (2, 1.7%), and β-pinene (1.31, 2%), respectively. Antifungal activities of the oils were studied with special reference to the inhibition of Aspergillus parasiticus growth and aflatoxin production. Minimal inhibitory (MIC) and minimal fungicidal (MFC) concentrations of the oils were determined. Static effects of the above oils against A. parasiticus were at 250 ppm and lethal effects of T. eriocalyx and T. X-porlock were 500 and 1000 ppm of the oils, respectively. Aflatoxin production was inhibited at 250 ppm of both oils with that of T. eriocalyx being stronger inhibitor. Transmission electron microscopy (TEM) of A. parasiticus exposed to MIC level (250 ppm) of the oils showed irreversible damage to cell wall, cell membrane, and cellular organelles. It is concluded that the essential oils could be safely used as preservative materials on some kinds of foods at low concentrations to protect them from fungal infections.

Inhibition of growth and mycotoxin production of Aspergillus flavus and Aspergillus parasiticus by extracts of Agave species

In this work, the effect of ethanolic, methanolic and aqueous extracts of Agave asperrima and Agave striata on growth and production of aflatoxin (in A&M medium) and cyclopiazonic acid (CPA; in Czpaek-Dox medium) and on growth in corn under storage conditions was determined. Aspergillus strains were inoculated (10 6 conidia per ml of medium or per 6 g of corn), then plant extracts were added and incubated without shaking at 28 °C for 8 days (for aflatoxin-producing analysis) or for 12 days (for CPA-producing analysis). Aflatoxin was assayed by HPLC and cyclopiazonic acid by absorbance at 580 nm using the Erlich reagent. The extracts that most effectively inhibited growth were those from the flowers of both plants. These exhibited an MIC from 0.5 to 2 mg/ml in culture media. Extracts from scape showed an MIC from 15 to 30 mg/ml in culture media. The MIC of the flower extracts was higher (>30 mg/g) when examined in corn. However, concentrations lower than the MIC drastically inhibited production of aflatoxins in culture medium or in corn. Half of the MIC inhibited 99% of the production of aflatoxins and 85% of cyclopiazonic acid.

Effect of some chemical and herbal compounds on growth of Aspergillus parasiticus and aflatoxin production

The anti-fungal properties of a series of chemical and herbal compounds at different levels was tested on potato dextrose agar. Among the chemical compounds, propionic acid at 0.1–0.5%, ammonia at 0.5%, copper sulphate at 0.08–0.5% and benzoic acid at 0.1–0.5% completely inhibited Aspergillus parasiticus growth. Urea, citric acid and sodium propionate had moderate anti-fungal properties (36–64% reduction). Among the herbal compounds, clove oil at 0.5% completely inhibited fungal growth. Compounds which inhibited fungal growth by at least 20% were selected to test their efficacy to inhibit fungal growth and aflatoxin production in feeds. All the selected chemical and herbal compounds reduced ( P<0.01) fungal growth (i.e. fungal spore count) and aflatoxin production. Propionic acid at 0.05–0.5%, sodium propionate at 0.1–0.5%, benzoic acid at 0.2% and ammonia at 0.5% completely inhibited aflatoxin production. Reduction in aflatoxin production was high with 0.5–1% citric acid (91–94%), 0.1–0.5% urea (93–96%) and 0.08% copper sulphate (85%). Clove oil at 0.5–1% inhibited aflatoxin production completely. Moderate reduction in toxin production occurred with 0.2–1% turmeric (63–84%), 0.1–1% onion (64–76%) and 0.2–1% garlic (71–84%). Among the chemical compounds tested in feeds, propionic acid, sodium propionate, benzoic acid and ammonia were the best anti-fungal compounds, followed by urea and citric acid. Among the herbal compounds, clove oil was the best followed by turmeric, garlic and onion.

Inhibitory effects of Thyme oils on growth and aflatoxin production by Aspergillus parasiticus

Inhibition of Aspergillus parasiticus growth and its aflatoxin production exposed to the essential oils extracted from two varieties of Thyme i.e. Thymus eriocalyx and Thymus x-porlock were studied. The disc diffusion method was used to evaluate the zone of fungal growth inhibition at various concentrations of the oils. Minimal inhibitory concentration (MIC) and minimal fungicidal concentration (MFC) of the oils were determined and compared with each other. The oils from the above plants were found to be strongly fungicidal and inhibitory to aflatoxin production. The oils analyzed by GC and GC/MS lead to identification of 18 and 19 components in T. eriocalyx and T. x-porlock oils respectively. The profile of the oil components from T. eriocalyx was similar to that of T. x-porlock in almost all the compounds but at different concentrations. The major components of T. eriocalyx and T. x-porlock oils were Thymol (64.3, 30.7%), β-phellandrene (13.2, 39.4%) and cis-sabinene hydroxide (8.4, 9.7%) respectively. Substitution of currently used antifungal and aflatoxin inhibiting chemicals by natural compounds such as thyme is recommended.

Black olives as substrate for Aspergillus parasiticus growth and aflatoxin B 1 production

Aflatoxin B1 (AFB 1) is a highly toxic and carcinogenic metabolite produced by certain Aspergillus species on agricultural commodities. Molds isolated from black olives are potentially toxigenic and present a potential health hazard. Olive oil originating from contaminated olives with AFB 1 might also be contaminated. The aim of this study was to investigate A. parasiticus growth and AFB 1 production in black damaged olives inoculated with 100 conidia flask −1 (1) treated with NaOCl 1.25%, (2) autoclaved at 110°C for 2 min, in comparison to the mold growth and AFB 1 production into the yeast extract sucrose (YES) medium under the same conditions of incubation and inoculation. AFB 1 extracted from cultures or olives and purified with immunoaffinity columns, was derivatized to its hemiacetal AFB 2a and then quantitated by HPLC using fluorescence detector. The recoveries and detection limits from YES and olives were 99.2%, 0.02 ng AFB 1 ml −1 and 94%, 0.15 ng AFB 1 g −1, respectively. Results showed that mycelia growth was not observed in olives during the 15 days of observation. The maximum growth of A. parasiticus on YES medium was shown on the sixth day. The AFB 1 production for both treated with NaOCl and autoclaved olives inoculated or not with A. parasiticus was not significantly different. On the other hand, AFB 1 levels produced in olives treated with NaOCl were significantly higher as compared with the autoclaved. The range of contamination of all olive samples inoculated or not for the whole period of observation was 0.15–2.3 ng AFB 1 g −1. The production of AFB 1 in YES medium on the third, the ninth and the 15th day was ∼1000-, ∼2500- and 10 000-fold higher, respectively, compared with the production in olives thus showing that black damaged olives of Greek origin are not a substrate favorable for AFB 1 biosynthesis at hazardous levels. Nevertheless, the production is possible at detectable amounts even after a little contamination that could happen randomly.

Spice Oils for the Control of Co-Occurring Mycotoxin-Producing Fungi

The effect of nine different oils was evaluated on the growth of Aspergillus parasiticus and Fusarium moniliforme. The experimental design to examine the inhibition of mycotoxins involved the incorporation of each of seven oils into broth and patty cultures. The fungal mycotoxin was identified by high-pressure liquid chromatography. Clove oil (eugenol) was the most inhibitory to the growth of A. parasiticus and F. moniliforme, followed by cinnamon (cinnamic aldehyde), oregano (thymol and carvacol) and mace oils (myristin). Neem and eucalyptus oil (cineole) did not affect fungal growth. The feasibility of implementing the results of this study to control mycotoxin toxicity was examined by costoring whole and ground cloves with mycotoxin-infected grain. Addition of both whole and ground cloves markedly reduced the aflatoxin contamination of the grain. These results clearly suggest that commonly occurring mycotoxigenic fungi can be controlled with clove oil (eugenol), thus spice oil successfully inhibited the growth of A. parasiticus and F. moniliforme, regulated the production of fumonisins, and prevented the formation of aflatoxins. The social implication of this finding is that rural communities can prevent the formation of fungal toxins in contaminated grain by simple measures.