Distribution of plasma phosphatidylcholine molecular species in rabbits fed fish oil is modulated by dietary n-6 fatty acids

Natural occurrence of Fusarium mycotoxins (trichothecenes and fumonisins) and aflatoxin B1 (AFB1) were surveyed in 32 corn samples, harvested in 1993 and randomly sampled in 1994 in several districts of Hanoi, Vietnam. Corn samples were first milled into fine powder, extracted with methanol-water (3:1) and the crude extracts obtained from the same samples were used for the simultaneous analysis of the trichothecenes such as nivalenol (NIV), deoxynivalenol (DON), and T-2 toxin (T-2) by gas chromatography/mass spectrometry (GC/NS); fumonisins B1 (FB1), B2 (FB2), and B3 (FB3) by high-performance liquid chromatography (HPLC) with a flourescence detector; and AFB1 by and ELISA kit based on a monoclonal antibody. The data revealed that 14, 8, 4, 3, and 2 out of 15 corn kernel samples were positive for AFB1, FB1, FB2, FB3, and NIV with the average levels being 28, 1, 101, 276, 232, and 858 ppb, respectively, and neither DON nor T-2 were detected. As for the other 17 samples of corn powder, 13, 15, 12, 10, 4 and 2 were positive for AFB1, FB1, FB2, FB3, DON, and NIV with the average being 30, 780, 289, 176, 3, 170, and 1,365 ppb, respectively, and T-2 was not detected. Although their positive rates and levels fell in the ranges reported elsewhere, it was found for the first time that the Fusarium toxins (NIV, DON, and fumonisins) and an Aspergillus toxin (AFB1) were naturally co-contaminated in selected samples of corn produced in north Vietnam.     

Total hip arthroplasty cemented femoral component distal stem centralizer. Effect on stem centralization and cement mantle

Samples of wheat harvested from 1988 to 1990 and stored in elevators in the south of Brazil (12 Brazilian, 4 Argentinian and 2 Uruguayan) were analysed in 1990 for 14 mycotoxins: deoxynivalenol (DON), nivalenol, diacetoxyscirpenol (DAS), T-2 and HT-2 toxins, T-2 triol, T-2 tetraol, aflatoxins B1, B2, G1, G2, ochratoxin A (OCHRA A), zearalenone and sterigmatocystin. One sample (1988 harvest) was contaminated with OCHRA A (0.04 microgram/g) and three other samples (1990 harvest) were contaminated with DON (0.40 microgram/g), DAS (0.30 microgram/g), T-2 (two samples, 0.35 and 0.36 gamma g/g) and T-2 tetraol (1.68 micrograms/g). Fusarium graminearum Schwabe was found in the 1990 samples with a relative incidence ranging from 1 to 22% and predominated in Argentinian and Uruguayan wheat (1990 harvest). Fusarium dimerum Penzig (8-75%) was the main Fusarium sp. in Brazilian wheat from the 1990 harvest.     

Natural occurrence of Fusarium toxins in feedstuff

The mycotoxins diacetoxyscirpenol, deoxynivalenol, and zearalenone, produced by Fusarium roseum, were found naturally occuring in mixed feed samples. In all cases analyzed, deoxynivalenol occurred together with zearalenone. The natural occurrence of zearalenone in sesame seed is reported for the first time. Strains of F. roseum isolated in various parts of the world form feed implicated in animal mycotoxicosis produced monoacetoxyscirpenol, diacetoxyscirpenol, deoxynivalenol, and zearalenone.     

Evaluation of maxillofacial vascular abnormalities with Tc-99m RBC

Cassava bread was prepared by pre-gelling, battering and baking cassava flour to which were added, in moderate amounts, sugar, yeast solution and edible oil. Baking was at 215 degrees C for 40 min. No mould was isolated from the cassava bread and the mean value of aflatoxin B1 (AFB1) for the three subsamples of cassava bread was 0.03 microgram/kg. The cassava tuber (Manihot esculenta Crantz), which was used for the production of cassava bread had an initial AFB1 level of 1.91 micrograms/kg and the dominant mycoflora were Penicillium oxalicum, Aspergillus flavus, Fusarium spp and some unidentified fungi.     

Group specific PCR-detection of potential trichothecene-producing Fusarium-species in pure cultures and cereal samples

A PCR based assay (Tox5 PCR) which analyses Fusarium species potentially producing trichothecenes was developed using a pair of primers derived from the DNA-sequence of the trichodiene synthase gene (tri5). The primer pair was tested using DNA isolated from a variety of strains representing 64 species and varieties of Fusarium as well as from other fungi, bacteria and cereals. A 658 bp PCR fragment was specifically amplified with DNA isolated from strains of species belonging to the Fusarium sections Discolor, Sporotrichiella, Arthrosporiella, Gibbosum, and "Dlaminia". PCR products obtained were sequenced. Alignment to tri5 sequences given in the literature revealed a high degree of homology. Results of the PCR developed correlated well with literature data on the trichothecene producing capabilities of the respective species. Potential trichothecene producing fusaria were detected in contaminated cereals and malts using the Tox5 PCR assay. Intensity of the signals produced were well correlated with the concentration of deoxynivalenol (DON) in samples of wheat.     

The effects of resistance training on well-being and memory in elderly volunteers

In vitro affinity tests were conducted to test the effectiveness of 19 activated carbons (ACs), hydrates sodium calcium aluminosilicate (HSCAS) and sepiolite (S) in binding ochratoxin A (OA) and deoxynivalenol (DON) from solution. Relationships between adsorption ability and physicochemical parameters of ACs (surface area, iodine number, methylene blue index) were tested. When 5 ml of a 4-micrograms/ml aqueous solution of OA was treated with 2 mg of AC, the ACs adsorbed 0.80 to 99.86% of the OA. HSCAS and S were not effective in binding OA. In two saturation tests carried out with increased amounts of OA (5 ml of 10-and 50-micrograms/ml aqueous solutions of OA, respectively) three ACs also showed high adsorption ability (adsorbing 92.23 to 96.57% of the OA). When 5 ml of a 4-micrograms/ml aqueous solution of DON was treated with 10 mg of AC, ACs adsored 1.83 to 98.93% of the DON. HSCAS and S were not effective in binding DON. An overall relation of adsorption ability to the physicochemical parameters of ACs was observed. The methylene blue index was more reliable than iodine number and surface area in predicting ability of ACs to adsorb OA and DON. Based on the data observed on the xxxxx eh present study as well as on aflatoxin B1 and fumonisin B1 from previous studies, it is concluded that ACs have high in vitro affinity for chemically different mycotoxins, and can be considered as potential multi-mycotoxin-sequestering agents. However, the ability to bind the main mycotoxins singly or in combination should be confirmed by in vivo investigations. Moreover, information on the amounts of AC to be added to feeds, and on the possible long-term effect on absorption of essential nutrients are needed.     

Survey of the occurrence of aflatoxin M1 in dairy products marketed in Italy

During 1995, 159 samples of milk, 97 samples of dry milk for infant formula, and 114 samples of yogurt were randomly collected in supermarkets and drug stores in four large Italian cities and checked for aflatoxin M1 (AFM1) by immunoaffinity column extraction and HPLC. AFM1 was detected in 136 (86%) of the milk samples (in amounts ranging from < 1 ng/liter to 108.5 ng/liter; mean level: 10.19 ng/liter), in 81 (84%) of the dry milk samples (in amounts ranging from < 1 ng/liter to 101.3 ng/kg; mean level: 21.77 ng/kg), and in 91 (80%) of the yogurt samples (in amounts ranging from < 1 ng/liter to 496.5 ng/liter; mean level: 18.08 ng/liter). Altogether, only two samples of milk, two samples of yogurt, and one sample of dry milk had levels of AFM1 exceeding the Swiss legal limits, which are the most restrictive in the world. AFM1 contamination levels in milk and yogurt samples collected in the period of November to April were ca. four times as high as those in samples collected in the period of May to October. It is concluded that during 1995, despite the widespread occurrence of AFM1, the mean contamination levels in dairy products sold in Italy were not a serious human health hazard.     

Dexamethasone treatment of lipopolysaccharide-induced meningitis in rabbits that mimics magnification of inflammation following antibiotic therapy

Natural occurrence of fumonisins B1 and B2, incidence of Fusarium species, and capacity to produce fumonisins by Fusarium isolates, were investigated in 50 corn-based samples from Spain destined for animal consumption. Forty-four samples (88%) were found to be contaminated with fumonisins. The levels of contamination were very low, with a mean of 400 ng/g in the samples. We investigated the capacity of 11 isolates of Fusarium moniliforme and 19 isolates of F. proliferatum to produce fumonisins. All F. proliferatum isolates and 8 out of the 11 F. moniliforme isolates assayed produced fumonisins on a corn medium. The FB1/FB2 ratio in the isolates ranged from 1.1 to 3.5.     

Measurement of antibacterial activities of T-2 toxin, deoxynivalenol, ochratoxin A, aflatoxin B1 and fumonisin B1 using microtitration tray-based turbidimetric techniques

Various mycotoxins were tested for their antibacterial activity by evaluating growth delays using a fully automated microturbidmetric method. Ten different strains of the genera Escherichia, Streptococcus, Staphylococcus, Yersinia, Salmonella, Erysipelothrix and Lactobacillus were used as test micro-organisms. T-2 toxin, deoxynivalenol (DON), ochratoxin A (OTA), aflatoxin B1 (AFB1) and fumonisin B1 (FB1) were used as representative mycotoxins. The inhibitory effect in vitro was defined as the difference between the growth rate without mycotoxins and the growth rate in the presence of a mycotoxin. Among the tested strains, Streptococcus agalactiae was found to be sensitive to all the toxins, with the exception of OTA. T-2 toxin and FB1 were the most effective in slowing down the growth of Staphylococcus aureus. AFB1 affected the growth of Yersinia enterocolitica. The growth rate of Escherichia coli and Salmonella infantis was decreased by FB1. Among the bacterial strains used in this study, only the growth of Erysipelothrix rhusiopathiae was inhibited by OTA. Thus, using appropriate tester strains it should be possible to set up a broad-range microtubidimetry assay for individual mycotoxin screening in vitro. We concluded that the microtitration technique provides a rapid, convenient and high-throughput capacity system to analyse bacteria-mycotoxin interactions.     

The occurrence of fungi along the Red Sea coast and variability among isolates of Fusarium as revealed by isozyme analysis

Nineteen identified species which belong to nine fungal genera were recovered from 14 samples collected from different sites of the Red Sea governorate. The aquatic fungal genera were Allomyces, Dictyuchus, Saprolegnia and Pythium while, the terrestrial fungal genera were Aspergillus, Penicillium, Fusarium, Neurospora and Rhizopus. Aspergillus was the most frequent genus, represented by seven species, of which A. niger, A. flavus and A. ustus were the most common. Penicillium was of occurred less frequently and was represented by two species, while Fusarium was isolated unfrequently and contributed four species. The remaining genera were unfrequent or rare and were each represented by one species. In addition, two electrophoretic isozyme patterns, esterase and glutamate oxalate transaminase (GOT), were determined to measure variability among 10 isolates of Fusarium. The results revealed that the tested fungi differed from each other in one or more esterase bands, except that F. moniliforme isolated from Safaga and from 40 Kilometers south of El-Kaussier yielded similar banding pattern. The activity of GOT was observed in the samples of F. solani and F. oxysporum and not detected in other isolates of Fusarium. The results indicated that F. solani differed from F. oxysporum in the isozymes of GOT, while no differences were observed between the isolated of the same species.     

Schistosomiasis vaccine development--the current picture

The fate of three Fusarium mycotoxins, nivalenol (NIV), deoxynivalenol (DON) and zearalenone (ZEN), all common contaminants in New Zealand-grown maize, has been measured in fractions of maize after passage through a commercial wet-milling plant. Distribution of the three toxins follows a pattern reasonably expected from their physical solubility characteristics. The highly water-soluble mycotoxins, NIV and DON, were found at high concentrations (up to 8.8 mg/kg) in concentrated steep liquor (CSL) fractions, but at low levels (less than 0.3 mg/kg) in the solid (germ, fibre and gluten) fractions. The converse was true for ZEN, which is relatively insoluble in water. For ZEN, the maximum concentration found in CSL was 0.6 mg/kg compared with 2.2-4.8 mg/kg in germ, fibre and gluten fractions. Accordingly, an animal food byproduct composed mainly of pressed fibre and concentrated steep liquor was usually found to contain concentrations of all three mycotoxins above those existing in the input maize. A single sample of corn oil recovered during the study also had a high concentration (4.6 mg/kg) of ZEN. The analytical clean-up method used converts all trichothecenes present to parent alcohols, therefore results are indicative of total trichothecene content. HPLC analytical conditions suitable for the analysis of NIV and DON in complex process grain products are also described.     

Down-regulation of human FEN-1 gene expression during differentiation of promyelocytic leukemia cells

The presence of aflatoxin in corn and corn dust during relatively normal years and the increased risk of Aspergillus flavus infestation during drought conditions suggest that airborne agricultural exposures should be of considerable concern. Liquid extraction, thin layer chromatography, and high pressure liquid chromatography were used for the analysis of aflatoxin B1 in grain dust and bulk corn samples. A total of 24 samples of airborne dust were collected from 8 farms during harvest, 22 samples from 9 farms during animal feeding, and 14 sets of Andersen samples from 11 farms during bin cleaning. A total of 14 samples of settled dust and 18 samples of bulk corn were also collected and analyzed. The airborne concentration of aflatoxin B1 found in dust collected during harvest and grain unloading ranged from 0.04 to 92 ng/m3. Higher levels of aflatoxin B1 were found in the airborne dust samples collected from enclosed animal feeding buildings (5-421 ng/m3) and during bin cleaning (124-4849 ng/m3). Aflatoxin B1 up to 5100 ng/g were detected in settled dust collected from an enclosed animal feeding building; however, no apparent correlation was found between the airborne concentration of aflatoxin B1 and its concentration in settled dust or bulk corn. The data demonstrate that farmers and farm workers may be exposed to potentially hazardous concentrations of aflatoxin B1, particularly during bin cleaning and animal feeding in enclosed buildings.     

Direct synthesis of aflatoxin B1-N7 guanine adduct: a reference standard for biological monitoring of dietary aflatoxin exposure in molecular epidemiological studies

Aflatoxin B1-N7-guanine and aflatoxin B1-human serum albumin adducts have been established as biomarkers of dietary aflatoxin exposure in epidemiological studies. Earlier chemical oxidants were used to synthesize aflatoxin B1-8,9-epoxide in vitro and its subsequent interaction with DNA or synthetic oligodeoxynucleotide was used as a source of authentic aflatoxin B1-N7-guanine adduct. In the present communication we report a simple single step procedure for the synthesis of aflatoxin B1-N7-guanine adduct using free guanine and m-chloroperbenzoic acid as the chemical oxidant for the production of AFB1-8,9-epoxide. At a molar ratio of 1:1 of AFB1-8,9-epoxide and guanine the recovery of the AFB1-N7-guanine adduct was found to be 60% while at higher molar ratios (1:2 and 1:4) of guanine the recovery of the AFB1-N7-guanine adduct was found to be low (30-40%). HPLC analysis of the AFB1-N7 guanine adduct showed a retention time identical with the retention time of the AFB1-N7-guanine adduct synthesized using calf thymus DNA. TLC-fluorodensitometric analysis indicated that the Rf of the AFB1-N7-guanine adduct was zero. Spectral analysis of the adduct synthesized showed an excitation wavelength of 360 nm and emission wavelength at 440 nm in phosphate buffer (100 mM, pH 7.4). Further, the formation of the AFB1-N7-guanine adduct was confirmed by perchloric acid treatment resulting in the destruction of the adduct. The AFB1-N7-guanine adduct thus synthesized was stable in both acidic as well as lyophilized conditions over a period of 2 weeks. The antibody capture assay showed that the antibodies produced against the antigen BSA-guanine-N7-AFB1 also cross-reacted with calf thymus DNA-AFB1 adduct, indicating specificity to the guanine-N7-AFB1 moiety. The method developed may find immediate application as a source of authentic reference standard in molecular epidemiological studies.     

Effect of feeding blends of Fusarium mycotoxin-contaminated grains containing deoxynivalenol and fusaric acid on growth and feed consumption of immature swine

Experiments were conducted to determine the effect of feeding diets containing combinations of the Fusarium metabolites deoxynivalenol (DON) and fusaric acid (FA) to starter swine. In all experiments, pigs of approximately 8.2 kg initial weight were fed diets containing blends of mycotoxin-contaminated corn, wheat, and barley for 21 d with growth and feed consumption determined weekly. In the first experiment, diets were determined to contain 0 microgram DON/g + 58.9 micrograms FA/g (control), 4.4 micrograms DON/g + 57.1 micrograms FA/g, 6.0 micrograms DON/g + 48.6 micrograms FA/g, and 7.5 micrograms DON/g + 57.4 micrograms FA/g. The feeding of all diets containing DON caused significant linear depressions in growth and feed intake after only 1 wk. Lower concentrations of DON and FA were fed in the second experiment with diets containing 0 microgram DON/g + 16.3 micrograms FA/g (control), .5 microgram DON/g + 14.3 micrograms FA/g, 1.1 micrograms DON/g + 14.1 micrograms FA/g, and 1.9 micrograms DON/g + 13.6 micrograms FA/g. There was a significant linear reduction in feed intake after 1 wk with increasing levels of dietary DON. Weight gains declined significantly only after 3 wk. Increasing amounts of FA combined with relatively constant amounts of DON were fed in the third experiment. By analysis, diets contained .5 micrograms DON/g + 2.9 micrograms FA/g (control), 2.2 micrograms DON/g + 12.2 micrograms FA/g, 2.5 micrograms DON/g + 15.6 micrograms FA/g, and 2.4 micrograms DON/g + 15.9 micrograms FA/g. In the 1st wk, the feeding of increasing amounts of fusaric acid combined with a relatively constant amount of DON caused a significant linear depression in weight gain. We concluded that a toxicological synergism exists between DON and FA when fed to immature swine and that FA concentrations in feeds should be determined whenever DON analysis is conducted.     

Beauvericin production by Fusarium species

Beauvericin is a cyclohexadepsipeptide mycotoxin which has insecticidal properties and which can induce apoptosis in mammalian cells. Beauvericin is produced by some entomo- and phytopathogenic Fusarium species (Fusarium proliferatum, F. semitectum, and F. subglutinans) and occurs naturally on corn and corn-based foods and feeds infected by Fusarium spp. We tested 94 Fusarium isolates belonging to 25 taxa, 21 in 6 of the 12 sections of the Fusarium genus and 4 that have been described recently, for the ability to produce beauvericin. Beauvericin was produced by the following species (with the number of toxigenic strains compared with the number of tested strains given in parentheses): Fusarium acuminatum var. acuminatum (1 of 4), Fusarium acuminatum var. armeniacum (1 of 3), F. anthophilum (1 of 2), F. avenaceum (1 of 6), F. beomiforme (1 of 1), F. dlamini (2 of 2), F. equiseti (2 of 3), F. longipes (1 of 2), F. nygamai (2 of 2), F. oxysporum (4 of 7), F. poae (4 of 4), F. sambucinum (12 of 14), and F. subglutinans (3 of 3). These results indicate that beauvericin is produced by many species in the genus Fusarium and that it may be a contaminant of cereals other than maize.     

Occurrence of aflatoxins, fumonisin B1, and zearalenone in foods and feeds in Botswana

Sorghum and maize form the main dietary staple foods in Botswana. Other products such as peanuts, peanut butter, phane (an edible larval stage of an emperor moth Imbrasia belina Westwood), and pulses (cowpeas and beans) are also widely used as food and for the manufacture of feeds. These important food and feed commodities were analyzed for the presence of aflatoxins, fumonisin B1, and zearalenone. Aflatoxins were detected in 40% of the samples analyzed. The concentration of total aflatoxins ranged from 0.1 to 64 microg/kg. The mean concentration ranged from 0.3 microg/kg in sorghum to 23 microg/kg in peanut butter. Peanut butter samples were the most contaminated (71%). No aflatoxins were detected in maize. Fumonisin B1 was detected in 36% of the samples. Maize samples were the most contaminated (85% of the samples) with the concentration ranging from 20 to 1,270 microg/kg. No fumonisin B1 was detected in peanuts, phane, and beans. Zearalenone was only found in 2.6% of the samples analyzed at 40 microg/kg. Aflatoxins were the most common toxins detected in foods and feeds in Botswana. However, fumonisin B1 was more prevalent in maize than aflatoxins or zearalenone.     

Immunochemical approaches to AGE-structures: characterization of anti-AGE antibodies

An extractionless method for determining aflatoxin M1 (AFM1), a major metabolite of aflatoxin B1 (AFB1), in human urine was developed. The biological fluid is injected directly into the chromatographic system after simple dilution and centrifugation. A pre-column, packed with a cation-exchange phase and coupled on-line to a column-switching liquid chromatography (LC) system, is used for sample pre-treatment and concentration. The analytes are non-selectively desorbed with the LC eluent and cleaned by means of a column-switching procedure. Pre-treatment and analysis were performed within 40 min. Average AFMI recovery reached 97% in the 10-100 ng/l range of urine. The detection limit of AFM1 in urine and milk was 2.5 ng/l for 1 ml of injected sample. A comparison with an immunoaffinity column clean-up and LC method was performed. The method was applied to determine AFM1 in the urine of AFB1 gavaged rats, and in the urine of both potentially exposed and supposedly unexposed workers. The method was also extended to milk.     

The MBP fusion protein restores the activity of the first phosphatase domain of CD45

Thirty samples of paddy rice and twenty-five of milled rice were obtained from processing centers located in two northern Provinces of Argentina and one southern Province of Paraguay. Contaminating fungi were enumerated by direct plating on dichloran rose bengal chloramphenicol agar and oxytetracycline glucose yeast extract agar before and after surface disinfection. All fungi were isolated and identified to the genus level and percentage infection of samples calculated. Those belonging to the genera Penicillium, Aspergillus and Fusarium were identified to species level. The surface mycoflora was dominated by storage fungi, notably Penicillium citrinum (73% of samples), P. islandicum (60% of samples), Aspergillus niger, A. flavus and Fusarium semitectum. The major fungi found as internal contaminants of paddy rice were, again, Penicillium citrinum (66% of samples) and P. islandicum (50% of samples). Milled rice showed a lower level of contamination, but with a similar species distribution, Penicillium citrinum and P. islandicum again being the main contaminants. The presence of these species suggests a potential for mycotoxin production. Further studies are needed to establish the mycotoxin quality of rice from this region.     

Structure-function relationships of human liver cytochromes P450 3A: aflatoxin B1 metabolism as a probe

Cytochromes P450 3A4 and 3A5, the dominant drug-metabolizing enzymes in the human liver, share >85% primary amino acid sequence identity yet exhibit different regioselectivity toward aflatoxin B1 (AFB1) biotransformation [Gillam et al., (1995) Arch. Biochem. Biophys. 317, 374-384]. P450 3A4 apparently prefers AFB1 3alpha-hydroxylation, which results in detoxification and subsequent elimination of the hepatotoxin, over AFB1 exo-8,9-oxidation. In contrast, P450 3A5 is incapable of appreciable AFB1 3alpha-hydroxylation and converts it predominantly to the exo-8,9-oxide which is genotoxic. To elucidate the structural features that govern the regioselectivity of the human liver 3A enzymes in AFB1 metabolism and bioactivation, a combination of approaches including sequence alignment, homology modeling, and site-directed mutagenesis was employed. Specifically, the switch in AFB1 regioselectivity was examined after individual substitution of the divergent amino acids in each of the six putative substrate recognition sites (SRSs) of P450 3A4 with the corresponding amino acid of P450 3A5. Of the P450 3A4 mutants examined, P107S, F108L, N206S, L210F, V376T, S478D, and L479T mutations resulted in a significant switch of P450 3A4 regioselectivity toward that of P450 3A5. The results confirmed the importance of some of these residues in substrate contact in the active site, with residue N206 (SRS-2) being critical for AFB1 detoxification via 3alpha-hydroxylation. Moreover, the P450 3A4 mutant N206S most closely mimicked P450 3A5, not only in its regioselectivity of AFB1 metabolism but also in its overall functional capacity. Furthermore, the other SRS-2 mutant, L210F, also resembled P450 3A5 in its overall AFB1 metabolism and regioselectivity. These findings reveal that a single P450 3A5 SRS domain (SRS-2) is capable of conferring the P450 3A5 phenotype on P450 3A4. In addition, some of these P450 3A4 mutations that affected AFB1 regioselectivity had little influence on testosterone 6beta-hydroxylation, thereby confirming that each substrate-P450 active site fit is indeed unique.     

Absence of mutations in the p53 tumor suppressor gene in woodchuck hepatocellular carcinomas associated with hepadnavirus infection and intake of aflatoxin B1

Infection with hepadnaviruses and exposure to aflatoxin B1 (AFB1) are considered major risk factors in the development of hepatocellular carcinoma (HCC) in humans and in animals. A high rate of mutations in the p53 tumor suppressor gene in hepatocellular carcinomas of predominantly hepatitis B virus (HBV) carrier patients has been recently related to dietary aflatoxin. Another member of the hepadnavirus family, the woodchuck hepatitis virus (WHV), infects woodchucks in a manner similar to that of HBV in humans. Therefore, it was of particular interest to determine whether the p53 gene in woodchuck HCCs associated with hepadnavirus infection and with exposure to AFB1 is affected in the same manner as in human HCCs. By direct PCR-sequencing, we analyzed exons 4-9 of the p53 gene in 13 HCCs from 12 woodchucks (two uninfected, ten WHV carriers). Six WHV carrier and two uninfected woodchucks were treated with AFB1. None of the analyzed HCC samples exhibited mutations, either in p53 gene exons 4-9, or in splicing donor-acceptor sites. The present data are consistent with our previous study that indicated a low rate of p53 mutations in HCCs of AFB1-treated ground squirrels, either infected or not infected with ground squirrel hepatitis virus, and in WHV carrier woodchucks not exposed to AFB1. Overall, our findings indicate that in woodchucks and in ground squirrels exposure to aflatoxin may affect the development of p53 mutations less than in humans.