Fungi Aspergillus Parasiticus Research Articles

Proteomic and Biochemical Evidence Support a Role for Transport Vesicles and Endosomes in Stress Response and Secondary Metabolism in Aspergillus parasiticus

Aflatoxin is among the most potent naturally occurring carcinogens known. Previous studies demonstrated that endosomes in the filamentous fungus Aspergillus parasiticus carry enzymes that catalyze the final two steps in aflatoxin synthesis, and these structures also play a role in aflatoxin storage and export. We hypothesized that endosomes house a complete and functional aflatoxin biosynthetic pathway. To address this hypothesis, we purified a cellular fraction containing endosomes, transport vesicles, and vacuoles (V fraction) from A. parasiticus grown under aflatoxin inducing and noninducing conditions. We also added (fed) aflatoxin pathway intermediates to V fraction to test the functional status of aflatoxin pathway enzymes. High throughput LC-MS/MS analysis of proteins in V fraction detected 8 aflatoxin enzymes with high reliability and 8 additional enzymes at lower reliability, suggesting that most aflatoxin pathway enzymes are present. Purified V fraction synthesized aflatoxin and addition of the pathway intermediate versicolorin A increased aflatoxin synthesis, confirming that middle and late aflatoxin enzymes in V fraction are functional. Of particular significance, proteomic and biochemical analysis strongly suggested that additional secondary metabolic pathways as well as proteins involved in response to heat, osmotic, and oxidative stress are housed in V fraction.

Stress-related Transcription Factor AtfB Integrates Secondary Metabolism with Oxidative Stress Response in Aspergilli

In filamentous fungi, several lines of experimental evidence indicate that secondary metabolism is triggered by oxidative stress; however, the functional and molecular mechanisms that mediate this association are unclear. The basic leucine zipper (bZIP) transcription factor AtfB, a member of the bZIP/CREB family, helps regulate conidial tolerance to oxidative stress. In this work, we investigated the role of AtfB in the connection between oxidative stress response and secondary metabolism in the filamentous fungus Aspergillus parasiticus. This well characterized model organism synthesizes the secondary metabolite and carcinogen aflatoxin. Chromatin immunoprecipitation with specific anti-AtfB demonstrated AtfB binding at promoters of seven genes in the aflatoxin gene cluster that carry CREs. Promoters lacking CREs did not show AtfB binding. The binding of AtfB to the promoters occurred under aflatoxin-inducing but not under aflatoxin-noninducing conditions and correlated with activation of transcription of the aflatoxin genes. Deletion of veA, a global regulator of secondary metabolism and development, nearly eliminated this binding. Electrophoretic mobility shift analysis demonstrated that AtfB binds to the nor-1 (an early aflatoxin gene) promoter at a composite regulatory element that consists of highly similar, adjacent CRE1 and AP-1-like binding sites. The five nucleotides immediately upstream from CRE1, AGCC(G/C), are highly conserved in five aflatoxin promoters that demonstrate AtfB binding. We propose that AtfB is a key player in the regulatory circuit that integrates secondary metabolism and cellular response to oxidative stress.

Expression profiling of non-aflatoxigenic Aspergillus parasiticus mutants obtained by 5-azacytosine treatment or serial mycelial transfer.

Aflatoxins are carcinogenic secondary metabolites produced by the fungi Aspergillus flavus and Aspergillus parasiticus. Previous studies found that repeated serial mycelial transfer or treatment of A. parasiticus with 5-azacytidine produced colonies with a fluffy phenotype and inability to produce aflatoxins. To understand how these treatments affect expression of genes involved in aflatoxin production and development, we carried out expressed sequence tag (EST)-based microarray assays to identify genes in treated clones that are differentially expressed compared to the wild-type. Expression of 183 genes was significantly dysregulated. Of these, 38 had at least two-fold or lower expression compared to the untreated control and only two had two-fold or higher expression. The most frequent change was downregulation of genes predicted to encode membrane-bound proteins. Based on this result we hypothesize that the treatments cause changes in the structure of cellular and organelle membranes that prevent normal development and aflatoxin biosynthesis.

Composition and antimicrobial activity of the essential oil of Heracleum thomsonii (Clarke) from the cold desert of the western Himalayas

Volatile oil composition of hydro-distilled (HD) and supercritical carbon dioxide (SC-CO2) essential oil of freshly collected aerial parts of Heracleum thomsonii (Umbeliferae) from the western Himalayas was studied by GC-FID and GC-MS. Results revealed qualitative and quantitative dissimilarity in the composition of hydro-distilled and SC-CO2 extracted oils. Nineteen constituents, which accounted for 89.32% of total constituents in HD oil, represented by limonene (4.31%), (Z)-β-ocimene (3.69%), terpinolene (22.24%), neryl acetate (36.19%), nerol (9.51%) and p-cymene-8-ol (2.61%) were identified. In SC-CO2 extracted oil, 24 constituents representing 89.95% of total constituents were identified. Terpinolene (5.08%), germacrene D (2.17%), neryl acetate (51.62%), nerol (9.78%), geranyl acetate (2.06%), α-bisabolol (2.48%) and 1-nonadecanol (4.96%) were the dominating constituents. In vitro antimicrobial activity of hydro-distilled oil was conducted against microrobial strains including two Gram-positive (Staphylococcus aureus and Bacillus subtilis) and five Gram-negative (Burkholderia cepacia, Escherichia coli, Enterobacter cloacae, Klebseilla pneumoniae and Pseudomonas aeruginosa) bacteria as well as seven fungi (Candida albicans, Issatchenkia orientalis, Aspergillus flavus, Aspergillus niger, Aspergillus parasiticus, Aspergillus sydowii and Trichophyton rubrum) using broth microdilution method. The results of bioassay showed that the oil exhibited moderate to high antimicrobial activity against fungi C. albicans (MIC 625 µg ml−1), A. parasiticus (MIC 312.5 µg ml−1), A. sydowii (MIC 312.5 µg ml−1), T. rubrum (MIC 625 µg ml−1), Gram-positive bacteria B. subtilis (MIC 625 µg ml−1) and Gram-negative bacteria P. aeruginosa (MIC 312.5 µg ml−1).

Willow volatiles influence growth, development, and secondary metabolism in Aspergillus parasiticus

Aflatoxin is a mycotoxin and the most potent naturally occurring carcinogen in many animals. Aflatoxin contamination of food and feed crops causes a significant global burden on human and animal health. However, available methods to eliminate aflatoxin from food and feed are not fully effective. Our goal is to discover novel, efficient, and practical methods to control aflatoxin contamination in crops during storage. In the present study, we tested the effect of volatiles produced by willow (Salix acutifolia and Salix babylonica) and maple (Acer saccharinum) bark on fungal growth, development, and aflatoxin production by the fungus Aspergillus parasiticus, one economically important aflatoxin producer. S. acutifolia bark volatiles nearly eliminated aflatoxin accumulation (>90% reduction) by A. parasiticus grown on a minimal agar medium. The decrease in aflatoxin accumulation correlated with a twofold reduction in ver-1 (encodes a middle aflatoxin pathway enzyme) transcript level. Expression data also indicate that one histone H4 acetyltransferase, MYST3, may play a role in epigenetic control of aflatoxin gene transcription in response to volatile exposure. Volatiles derived from wood bark samples also increased fungal growth up to 20% and/or enhanced conidiospore development. Solid-phase microextraction-gas chromatographic-mass spectrometric analysis of bark samples identified sets of shared and unique volatile compounds that may mediate the observed regulatory effects on growth, development, and aflatoxin synthesis. This work provides an experimental basis for the use of willow industry by-products to control aflatoxin contamination in food and feed crops.

Antifungal effect of chitosan on the growth of Aspergillus parasiticus and production of aflatoxin B1

AbstractContamination of agricultural products by toxigenic fungi and the presence of mycotoxins cause serious economic damage, and toxic effects to humans and animals. In this study the effect of chitosan as a natural alternative for the control of the fungus Aspergillus parasiticus was evaluated. Chitosans were obtained from chitin previously extracted from shrimp heads using chemical (CS) and biological (CB) processes. The chitosan yields were 5.74 and 6.20% for CS and CB methods, respectively, with an amount of residual protein and ash below 1%, low molecular weight and degree of deacetylation of 80–82%. Chitosan showed fungistatic activity against A. parasiticus, with 6.71 and 10.66 g L−1 (CI50) of CS and CB, respectively, being required to delay fungal growth by 50% at 122 h. Chitosan (CI50) inhibited both radial growth up to 122 h, and germination of spores before 8 h higher than 50%. Chitosan also increased the average diameter of the spores and hyphae (P ≤ 0.05), reduced the septation process and increased the number of mitotic divisions of spores during germination. However, compared with control, chitosan decreased (P ≤ 0.05) aflatoxin B1 production at 8 days but increased (P ≤ 0.05) the total aflatoxins production by A. parasiticus. Copyright © 2011 Society of Chemical Industry

Essential oil of Origanum majorana L., Illicium verum Hook. f. and Cinnamomum zeylanicum Blume: chemical and antimicrobial characterization

Essential oils of Origanum majorana L. (marjoram), Illicium verum Hook. f. (star-anise) and Cinnamomum zeylanicum Blume (cinnamon) were obtained by steam distillation using a modified Clevenger device. The antimicrobial activity of each oil was evaluated against the bacteria Staphylococcus aureus, Escherichia coli and the fungi Aspergillus flavus and Aspergillus parasiticus by observing their growth and/or mycelial inhibition through comparison with the standard dish (without oil). The essential oils were analyzed using a gas chromatograph coupled to a mass spectrometer for identification and coupled to a flame ionization detector for quantification. The major constituents of marjoram, star-anise and cinnamon essential oils were 4-terpineol, trans-anetole and cinnamic aldehyde, respectively. In in vitro tests, essential oils of marjoram and cinnamon promoted an inhibitory effect on the bacteria S. aureus and E. coli, while the essential oil of star-anise presented activity only against E. coli. Marjoram, star-anise and cinnamon oils were effective against the studied fungi, presenting an inhibitory effect. The minimal inhibitory concentration for the mycelial growth of A. parasiticus was 1 and 0.01 µL mL-1 for star-anise and cinnamon oils, respectively. The minimal inhibitory concentration for A. parasiticus was 0.25, 2 and 2 µL mL-1 for cinnamon, star-anise and marjoram oils, respectively.

A comparative study of the antimicrobial activity of the leaf essential oils of chemo-varieties of Clausena anisata (Willd.) Hook. f. ex Benth

The emergence of multiple drug resistance to human pathogenic organisms has necessitated a search for new antimicrobial substances from various sources including plants. The present study was carried out as part of this search using the essential oils of the leaves of three chemo-varieties of Clausena anisata (Willd.) Hook. f. ex Benth. namely, estragole, trans-anethole and feniculin-containing chemo-varieties. The oils were screened against six bacteria ( Escherichia coli, Staphylococcus aureus, Salmonella typhi, Shigella sp., Proteus sp. and Pseudomonas aeruginosa) and three fungi ( Candida albicans, Aspergillus niger and Aspergillus parasiticus) isolated from clinical specimen using the disc sensitivity test. Microbes which showed significant sensitivity were further assayed with various concentrations of the active extracts in a dilution sensitivity test. The microorganisms were also assayed against seven broad spectrum antibiotics: penicillin G, amoxycillin, ampicillin, tetracycline, ceftizoxime, fosfomycin and urotractin. Results from the disc sensitivity test showed that the estragole-rich oil exhibited significant antimicrobial activity against E. coli (16.3 ± 0.3 mm) and Shigella sp. (17.2 ± 0.4 mm). The trans-anethole-rich oil exhibited less significant activity (11.4 ± 0.7 mm and 12.1 ± 0.3 mm respectively) whereas the feniculin-rich oil, acting alone and in combination with the trans-anethole-rich oil did not show any significant activity against the all microbes tested. Only the neat oils and their 1:2 dilutions showed visible inhibition of microbial growth in the dilution sensitivity test. The estragole-rich oil gave minimum inhibitory concentrations of 3.7, 6.7 and 13.2 mg/ml against C. albicans, S. aureus and E. coli respectively with corresponding ED 50 values of 1.3, 2.1 and 1.2 mg/ml. The trans-anethole-rich oil gave a minimum inhibitory concentration of 1.8 mg/ml against C. albicans with an ED 50 of 0.2 mg/ml. The findings suggest a significant antimicrobial activity of these plant essential oils though of lower efficacy compared to ampicillin. The results further suggest that such plant essential oils could potentially be exploited in the development of novel antibiotics.

Carcinogenic Crops: Analyzing the Effect of Aflatoxin on Global Liver Cancer Rates

Tree nuts and groundnuts, along with maize and other grains, can harbor aflatoxins, naturally occurring fungal metabolites that have been identified as risk factors for developing liver cancer. This association has most often been seen in people infected with hepatitis B virus (HBV). A new study examines the af latoxin/HBVrelationship to offer the first quantitative risk assessment of the number of liver cancer cases worldwide caused by aflatoxin [EHP 118:818–824; Liu and Wu]. Although a relatively rare malignancy in developed countries, liver cancer is a common health threat in developing regions of the world including Southeast Asia, China, and sub-Saharan Africa. These same regions have higher prevalence of HBV infection as well as higher levels of aflatoxin contamination in food due to a lack of resources to control the fungi Aspergillus flavus and Aspergillus parasiticus, which infiltrate crops and produce aflatoxin. Research has shown that individuals with chronic HBV infection and aflatoxin exposure are up to 30 times more at risk for liver cancer than uninfected individuals exposed to aflatoxin. In the current study, researchers analyzed information on food consumption patterns, aflatoxin biomarker levels in serum and urine, HBV prevalence, and population size in different world regions to quantify the subsequent risk of developing liver cancer. The investigators found that consumption of maize and ground-nuts was higher overall in African and Asian countries than in wealthier, more developed nations, leading to increased aflatoxin exposure. However, risk of aflatoxin-induced liver cancer could vary widely within a given nation: urban populations with more diverse diets had lower aflatoxin exposures than their rural counterparts, and there also was a lower HBV prevalence in urban populations. The authors concluded that uncontrolled exposure to aflatoxin may cause 4.6–28.2% of all liver cancer cases globally, with China, Southeast Asia, and sub-Saharan Africa bearing the brunt of the burden. This broad range reflects the uncertainty and variability of the available data on aflatoxin exposure and HBV prevalence. One thing does seem certain, they write: if more interventions to control aflatoxin and its health risks (for instance, improved storage protocols and vaccination for HBV) were administered in regions where they are most needed, liver cancer incidence could be significantly reduced worldwide.

Global Burden of Aflatoxin-Induced Hepatocellular Carcinoma: A Risk Assessment

BackgroundHepatocellular carcinoma (HCC), or liver cancer, is the third leading cause of cancer deaths worldwide, with prevalence 16–32 times higher in developing countries than in developed countries. Aflatoxin, a contaminant produced by the fungi Aspergillus flavus and Aspergillus parasiticus in maize and nuts, is a known human liver carcinogen.ObjectivesWe sought to determine the global burden of HCC attributable to aflatoxin exposure.MethodsWe conducted a quantitative cancer risk assessment, for which we collected global data on food-borne aflatoxin levels, consumption of aflatoxin-contaminated foods, and hepatitis B virus (HBV) prevalence. We calculated the cancer potency of aflatoxin for HBV-postive and HBV-negative individuals, as well as the uncertainty in all variables, to estimate the global burden of aflatoxin-related HCC.ResultsOf the 550,000–600,000 new HCC cases worldwide each year, about 25,200–155,000 may be attributable to aflatoxin exposure. Most cases occur in sub-Saharan Africa, Southeast Asia, and China where populations suffer from both high HBV prevalence and largely uncontrolled aflatoxin exposure in food.ConclusionsAflatoxin may play a causative role in 4.6–28.2% of all global HCC cases.

Polymerase chain reaction-based assay for the early detection of aflatoxigenic fungi on maize kernels

Objectives The aim of this work was to set up a DNA-based method for detecting aflatoxigenic fungi Aspergillus flavus and Aspergillus parasiticus on whole maize kernels using species-specific primers. Methods In the experiments, two aflatoxigenic strains, A. flavus NRRL 3357 and A. parasiticus NRRL 2999, were used. Further, Aspergillus niger 7096, OTA producer strain, and other strains of Aspergillus spp., Penicillium spp. and Fusarium spp. isolated from maize kernels were analyzed. Twelve commercial maize hybrids employed in the food and feed industries, not inoculated and inoculated with the different fungi, at different water activities, were used in the polymerase chain reaction-based assay. Three primer pairs were used for amplifying fragments of Afl P (omt 1), Afl M (ver 1), Afl R, genes present in the aflatoxin biosynthesis cluster. Results DNA amplification was achieved only with DNA from fungal strains of A. parasiticus and A. flavus and from maize kernels inoculated with A. flavus or A. parasiticus. Amplification was never observed with DNA of the other fungal species. Amplification was evident in maize starting from 12 h of incubation after inoculation, when mycelium is not yet visible by stereomicroscope analysis. Conclusion The results indicate that these highly specific and sensitive polymerase chain reaction-based methods are able to discriminate maize kernels infected with A. flavus or A. parasiticus from uninfected ones. This polymerase chain reaction method could be a real, effective alternative to traditional diagnostic methods for the early detection of aflatoxigenic fungi in food commodities.

Activity of essential oil and its major compound, 1,8-cineole, from Eucalyptus globulus Labill., against the storage fungi Aspergillus flavus Link and Aspergillus parasiticus Speare

The essential oil from leaves of Eucalyptus globulus obtained by hydrodistillation, as well as its major compound 1,8-cineole, identified by gas chromatography coupled with a mass selective detector, were evaluated for their effectiveness against the storage fungi Aspergillus flavus and Aspergillus parasiticus. The evaluation was performed by compound dissolution in yeast extract sucrose (YES) medium and exposure to headspace volatiles. Complete fungal growth inhibition of both species was achieved with the essential oil by contact and volatile assays. Volatile exposure showed total inhibition at the lower level tested of 500 μL. The 1,8-cineole tested alone showed partial inhibition only at the highest level of 1.3492 μL. Aflatoxin B 1 production was reduced in headspace volatile assays and partial inhibition was observed at the 200 μL dose of the essential oil.

Liver cancer: Descriptive epidemiology and risk factors other than HBV and HCV infection

The incidence of liver cancer is high in all low-resource regions of the world, with the exception of Northern Africa and Western Asia. The estimated worldwide number of new cases of liver cancer in 2002 is 600,000, of which 82% are from developing countries. Given the poor survival from this disease, the estimated number of deaths is similar to that of new cases. Hepatocellular carcinoma (HCC) is the main form of liver cancer. A part from chronic infections with Hepatitis B and Hepatitis C viruses, which are the main causes of HCC, contamination of foodstuff with aflatoxins, a group of mycotoxins produced by the fungi Aspergillus flavus and Aspergillus parasiticus, is an important contributor to HCC burden in many low-income country. Alcoholic cirrhosis is an important risk factor for HCC in populations with low prevalence of HBV and HCV infection, and the association between tobacco smoking and HCC is now established. Diabetes is also related to an excess risk of HCC and the increased prevalence of overweight and obesity likely contributes to it. The second most important type of liver cancer is cholangiocarcinoma, whose main known cause is infestation with the liver flukes, Opistorchis viverrini and Clonorchis sinensis, which is frequent in some areas in South-East Asia. Angiosarcoma is a rare form of liver cancer whose occurence is linked to occupational exposure to vinyl chloride.

Sexual reproduction and recombination in the aflatoxin-producing fungus Aspergillus parasiticus

The fungal phylum Ascomycota comprises a large proportion of species with no known sexual stage, despite high genetic variability in field populations. One such asexual species, Aspergillus parasiticus, is a potent producer of carcinogenic and hepatotoxic aflatoxins, polyketide-derived secondary metabolites that contaminate a wide variety of agricultural crops. In this study, individuals of A. parasiticus from a population showing an evolutionary history of recombination were examined for sexual reproduction. Crosses between strains with opposite mating-type genes MAT1-1 and MAT1-2 resulted in the development of ascospore-bearing ascocarps embedded within stromata. Sexually compatible strains belonged to different vegetative compatibility groups. Recombination through the independent assortment of chromosomes 3 and 6 was detected using loci for mating type, aflatoxin gene cluster, and a protein-encoding gene. Our discovery of the sexual stage in A. parasiticus has important implications for current biological control strategies using nontoxigenic strains to reduce aflatoxin contamination in crops.

Aflatoxin-producing Aspergillus spp. and aflatoxin levels in stored cassava chips as affected by processing practices

Cassava chips (cassava balls, and cassava pellets) are derived cassava products traditionally produced by farmers in sub-Saharan Africa following fermentation, and drying of fresh roots of cassava, and are widely consumed in Cameroon. Once produced, this food commodity can be stored for more than two months and contaminated by a wide array of harmful microbes. In order to assess persistence of toxigenic fungi in cassava chips, aflatoxin-producing fungi ( Aspergillus flavus, Aspergillus nomius, and Aspergillus parasiticus) and aflatoxins were contrasted at regular intervals in home-stored cassava chips collected in two locations of southern Cameroon throughout a two-month monitoring period. Three hundred and forty-six isolates of aflatoxin-producing fungi were found to be associated with all samples. A. flavus contaminated more samples in both types of chips (267 isolates in 53 samples), followed by A. nomius (58 isolates in 15 samples), whereas A. parasiticus was rarest. A direct competitive Enzyme-linked immunosorbent assay (ELISA)-based method was implemented to quantify the content in aflatoxins. Eighteen of the samples contained some aflatoxins at detectable levels whereas 54 did not. The levels of aflatoxin ranged between 5.2 and 14.5 ppb. The distribution of aflatoxin in positive samples depended on 8 parameters including pH, moisture content, storage duration, types of chips, level of contamination by aflatoxin-producing fungi, processing practices and storage facilities. From analysis of variance results, only pH ( p < 0.01), duration of storage ( p < 0.01), population of aflatoxin-producing species (0.0001) and the chip type ( p < 0.05) were significantly related to aflatoxin in positive samples. A stepwise regression analysis (forward selection procedure) indicated that aflatoxin levels were significantly ( p < 0.01) correlated with processing practices, storage facilities, and storage duration of the chips.

Iturin produced by Bacillus pumilus HY1 from Korean soybean sauce ( kanjang) inhibits growth of aflatoxin producing fungi

A new strain of Bacillus pumilus, designated HY1, was isolated from Korean soybean sauce ( kanjang). This classification was based on morphological, physiological, and chemotaxonomic features of the organism that identified it as a Gram-positive bacillus, and confirmed by 16S rDNA based phylogenetic analysis. Strain HY1 showed strong antifungal activity against the aflatoxin-producing fungi Aspergillus flavus and Aspergillus parasiticus, two common contaminants of fermented soybean foods. MALDI-TOF mass analysis revealed that the antifungal compound was similar to the known lipopeptide iturin. Iturin purified from strain HY1 had three isoforms with protonated masses of m/z 1,043.4, 1,057.4, and 1,071.4, and different structures in combination with Na + ion using MALDI-TOF MS. Purified iturin from HY1 also exhibited antifungal activity against A. flavus and A. parasiticus.

The 'omics' tools: genomics, proteomics, metabolomics and their potential for solving the aflatoxin contamination problem

Aflatoxins are highly carcinogenic secondary metabolites produced primarily by the fungi Aspergillus flavus and Aspergillus parasiticus. Aflatoxin contamination of food and feed is an age old problem of particular concern over the last four decades. Now, for the first time control measures for this problem appear within reach. For practical and sustainable control of aflatoxin contamination to be realised, however, additional information is needed rather rapidly, particularly for understanding the specific molecular factors (both in the plant and the fungus) involved during host plant-fungus interaction. The information derived from the use of novel tools such as genomics, proteomics and metabolomics provides us with the best and the quickest opportunity to achieve a clear understanding of the survival of toxigenic fungi in the field, the ability of the fungus to invade crops, and the process of toxin contamination under various environmental conditions. Significant progress has been made recently in understanding the genomic makeup of the most significant aflatoxin producing field fungus, namely Aspergillus flavus. Progress also has been made in the study of host crop resistance to fungal invasion through the use of proteomics. The information available on production of aflatoxin and other metabolites by Aspergillus flavus is reasonably extensive, although the application of metabolomics as a tool in this study is relatively new. In this review there is a discussion of the use of genomics, proteomics and metabolomics in deriving the requisite information for developing effective strategies to interrupt the machinery in the fungus for production of these toxins, as well as to assist in the development of host-resistance against fungal invasion and aflatoxin contamination of crops.

AFLATOXIN—RECOGNITION, UNDERSTANDING, AND CONTROL WITH PARTICULAR EMPHASIS ON THE ROLE OF THE AGRICULTURAL RESEARCH SERVICE

Aflatoxin, a metabolite of the fungi Aspergillus flavus and Aspergillus parasiticus, has been considered one of the world's most serious food safety problems since it was first identified in the 1960s following large numbers of turkey deaths in Britain. Imported peanut meal was targeted as the cause, leading to identification of mold hyphae, the causative fungus A. flavus, and isolation and identification of aflatoxins as the causative mycotoxins. The Agricultural Research Service (ARS) quickly joined the scientific effort in the 1960s to control aflatoxin and to assure food safety by toxicity testing and developing methodology for identification, quantification, and detoxification of aflatoxin. As National Program Leader for Food Safety at the ARS, I initiated a national aflatoxin research program to provide efficacious, workable preharvest controls to decrease losses and help assure human safety by preventing aflatoxin from occurring in susceptible commodities, peanut, corn, cottonseed, and tree nuts. A series of Aflatoxin Elimination Workshops was established to bring together the ARS scientists, university scientists, and industry representatives in a cooperative effort to control aflatoxin. The first success was methodology to reduce contamination of Arizona cottonseed by applying an atoxigenic strain of A. flavus, AF 36, to outcompete native toxigenic strains. This methodology is being extended to peanuts, corn, and tree nuts. Other lines of investigation include the discovery of natural compounds that prevent aflatoxin biosynthesis; understanding the role of oxidative stress in aflatoxin formation; and the isolation and identification of aflatoxin biosynthetic pathway synthesis and regulatory genes.

Amino acid supplementation reveals differential regulation of aflatoxin biosynthesis in Aspergillus flavus NRRL 3357 and Aspergillus parasiticus SRRC 143

Aflatoxins are toxic and carcinogenic secondary metabolites produced by the fungi Aspergillus flavus and Aspergillus parasiticus. To better understand the molecular mechanisms that regulate aflatoxin production, the biosynthesis of the toxin in A. flavus and A. parasticus grown in yeast extract sucrose media supplemented with 50 mM tryptophan (Trp) were examined. Aspergillus flavus grown in the presence of 50 mM tryptophan was found to have significantly reduced aflatoxin B(1) and B(2) biosynthesis, while A. parasiticus cultures had significantly increased B(1) and G(1) biosynthesis. Microarray analysis of RNA extracted from fungi grown under these conditions revealed 77 genes that are expressed significantly different between A. flavus and A. parasiticus, including the aflatoxin biosynthetic genes aflD (nor-1), aflE (norA), and aflO (omtB). It is clear that the regulatory mechanisms of aflatoxin biosynthesis in response to Trp in A. flavus and A. parasiticus are different. These candidate genes may serve as regulatory factors of aflatoxin biosynthesis.

Study on the characterization of lead (II) biosorption by fungus Aspergillus parasiticus

The lead (II) biosorption potential of Aspergillus parasiticus fungal biomass has been investigated in a batch system. The initial pH, biosorbent dosage, contact time, initial metal ion concentrations and temperature were studied to optimize the biosorption conditions. The maximum lead (II) biosorption capacity of the fungal biosorbent was found as 4.02 x 10(-4) mol g(-1) at pH 5.0 and 20 degrees C. The biosorption equilibrium was reached in 70 min. Equilibrium biosorption data were followed by the Langmuir, Freundlich and Dubinin- Radushkevich (D-R) isotherm models. In regeneration experiments, no significant loss of sorption performance was observed during four biosorption-desorption cycles. The interactions between lead (II) ions and biosorbent were also examined by FTIR and EDAX analysis. The results revealed that biosorption process could be described by ion exchange as dominant mechanism as well as complexation for this biosorbent. The ion exchange mechanism was confirmed by E value obtained from D-R isotherm model as well.