The production of dried figs involves some unique agricultural practices, which present significant risk of fungal infection of the fruit and subsequent mycotoxin contamination. The figs are allowed to ripen and shrivel on the tree, and after falling to the ground are collected daily, before being laid out for sun-drying for 5 days or more. During the stage of optimum water activity (around 0.8) for fungal growth and at temperatures from 25-30 °C, fungal infection can easily take place either through spore-contaminated dust or insect transmission to the fruit on the tree, or directly from the soil or during the course of subsequent sun-drying. A variety of different fungal species can infect dried figs and as a consequence, contamination can occur with a large number of secondary metabolites including aflatoxins B1, B2, G1 and G2, ochratoxin A, patulin, fumonisin B2 and kojic acid. This review describes the agricultural production of dried figs and summarises the available data on occurrence of fungi and mycotoxins in figs providing insights into possible infection routes.
The potent hepatotoxic mycotoxin rubratoxin B causes fatty liver. To elucidate the lipid accumulation mechanism, we investigated the type of lipid droplets accumulated and the activities of the lipogenic enzymes glucose-6-phosphate dehydrogenase (G6PD) and fatty acid synthase in mouse livers treated for 24 h with rubratoxin B. Oil Red O staining revealed numerous microvesicular lipid droplets in the liver cells of rubratoxin B-treated mice. In addition, treatment with rubratoxin B notably induced the activity of G6PD, a crucial member of the pentose phosphate cycle, which produces and supplies NADPH for fatty acid synthesis. Unexpectedly, rubratoxin B decreased the activity of fatty acid synthase, which facilitates fatty acid chain elongation. However, because fatty acid synthase is not a key enzyme in fatty acid synthesis, its activity level in rubratoxin B-treated mice may be sufficient for lipid accumulation.
Recent evidences in stem cell biology suggest that tumors contain a hierarchy of heterogeneous cell population in which cancer stem cells (CSCs) exist in the top and confer the maintenance ability of tumor formation on transformed cells. In this study, to elucidate the relationship between the expression level of CD133 and appearance of CSCs in hepatoma cells, we performed the expression analysis of CD133, which is a stem cell-specific cell surface marker and is utilized for detection of CSCs in various tumors. Reverse transcriptasepolymerase chain reaction (RT-PCR) and immunocytochemical analyses indicated that CD133-positive cells existed with high frequency in aflatoxin B1-induced rat hepatoma K2 and human hepatoma Huh7 cells. Down-regulation of CD133 in Huh7 cells with the micro-interfering RNA (miRNA) suppressed the expression of ATP-binding cassette transporter ABCG2 gene, which is specifically expressed in stem cells and implicated in cell growth and multidrug resistance of tumors. Thus, these results suggest that the deregulated expression of CD133 in hepatoma cells takes part in the generation of CSCs at least in part.
An inter laboratory study was conducted to evaluate an application of the AOAC Official Method of Analysis for the determination of ochratoxin A (OTA) in barley to brown rice as a new matrix. Four artificially OTA contaminated brown rice test samples (blind duplicates at two levels), six OTA spiked brown rice test samples (blind duplicates at three levels) and two blank brown rice (blind duplicates) were sent to 12 collaborators. These samples were analyzed by a designated method. From the results, the RSDr values ranged from 3.4 to 8.9 % at the OTA concentrations of 0.5 to 40 μg/kg and the RSDR values ranged from 9.6 to 21.3 % at the same OTA concentrations, respectively. The HorRat ranged from 0.4 to 1. These results allowed us to conclude that the AOAC Official Method of Analysis for OTA in barley (49.6.04) showed acceptable for brown rice at the OTA levels ranging of 0.5 to 40 μg/kg. Recently, as LC-MS analysis for determination of OTA was reported, LC-MS analysis fixed the LC-MS conditions in our laboratory was examined the same extracts obtained from the test samples by 8 of 12 collaborators. From the results of this test, LC-MS analysis was confirmed acceptable at the OTA levels ranging of 10 to 40 μg/kg.
We have summarized the results of the examinations for aflatoxins (AFs) conducted from 2002 to 2006 in approximately 2,300 to 3,000 annual samples of raw peanuts imported to Japan from major exporting countries. In peanuts from China, South Africa, the U.S. and Paraguay, a comparison was made between the proportion of products in which AFB1 and/or B2 was detected and that of products in which AFB1, B2, G1 and/or G2 was detected during the five years of survey. We also calculated the proportions of samples showing AF levels above the Japanese regulatory value (10 ppb for AFB1 only) and those showing levels above the international limit (a total of 15 μg/kg [ppb] for B- and G-group AFs) to all samples examined during the same survey period. The results showed that, in the Chinese peanuts which had large sample sizes, the proportion of products having AFB1 levels above the AFB1-focused regulatory value (0.4 % to 0.8 %) and that of products having AF levels above the limit of total B- and G-group AFs (0.4 % to 1.1 %) were very similar. Similarly, in South African peanuts, 0.3 % to 1.0 % and 0.3 to 1.2 % of the products showed AF levels above the AFB1-focused limit and the AF B- and G-group total limit, respectively. Isolates of Aspergillus section Flavi (the cause of AF contamination) from peanuts were identified for morphology and AF and cyclopiazonic acid productivity, and by heteroduplex panel analysis (HPA). It was found that the contamination of Chinese samples by B- and G-group AFs was caused mainly by A. parasiticus, whereas isolates from South African samples contaminated by B- and G-group AFs included, in addition to A. parasiticus, an atypical A. flavus isolate which forms numerous small sclerotia and produces B- and G-group AFs. This isolate was categorized into AfF4 by HPA of section Flavi.
4-(p-nitrobenzyl)pyridine (NBP) is often used for detection of trichothecenes developed on a thin-layer chromatography (TLC) plate, because this chromogenic reagent enables specific detection of the 12,13-epoxide group of trichothecenes. After the heat reaction with NBP and cooling at room temperature for several minutes, trichothecenes on the TLC plate are visualized as blue spots by alkalization with tetraethylenepentamine (TEPA). When the TLC plate was left overnight and then sprayed with TEPA, most trichothecenes yielded similar blue colors. However, dark yellowish green spots were given for 8-keto-15-hydroxytrichothecenes. This modified detection method can be used as an easy alternative of other instrumental analyses to distinguish between 8-keto-15-hydroxytrichothecenes and other trichothecenes that give similar Rf values on a TLC plate.
A food-derived, genotoxic hepatocarcinogen, 2-amino-3,8-dimethylimidazo[4,5-f ] quinoxaline (MeIQx) low-dose carcinogenicity was examined in the rat liver using a medium term bioassay for carcinogens. MeIQx was shown to form DNA adducts at very low doses, but did not induce 8-hydroxy-2′-deoxyguanosine formations, lacI gene mutations or an initiation activity. Glutathione S-transferase placental form (GST-P)-positive foci which are preneoplastic lesions occurred at higher dose. Furthermore, N-nitroso compounds, such as N-nitrosodiethylamine and N-nitrosodimethylamine were found not to induce GST-P-positive foci in the rat liver at very low doses. A genotoxic colon carcinogen, 2-amino-1-methyl-6-phenolimidazo [4,5-b] pyridine (PhIP) induced PhIP-DNA adducts at very low dose with no-effect level and aberrant crypt foci as a surrogate marker of preneoplastic lesions appeared at relatively high doses.These results imply the existence of thresholds, at least practical ones for the carcinogenicities of these genotoxic carcinogens. A non-genotoxic carcinogen, phenobarbital showed hormetic phenomenon in rat hepatocarcinogenicity, indicating the real threshold for the carcinogenicity.
Generally, climate conditions in Japan are considered to be unsuitable for aflatoxin production. On the other hand, natural contamination of Fusarium mycotoxins, patulin, ochratoxins and citrinin in domestic agricultural products can occur. Recently, it was revealed that some mycotoxins naturally occur in Japan. Therefore detail survey and development of countermeasure against mycotoxin contamination are matter of great urgency, now. In this paper, the situation and control of natural occurrence of mycotoxins in Japan will be presented and discussed. Natural contamination of deoxynivalenol and nivalenol in wheat and patulin in apple definitely occur in Japan, and that of patulin in grape and aflatoxin in crude sugar probably occur. Natural contamination of ochratoxin in cereals possibly occur in Japan. In 2003, high level of patulin was found in a part of domestic apples as raw material for juice. The incidence of patulin in apples stored for long term was relatively high. The fact indicated that apples were contaminated with patulin in storage at low temperature. To reduce the risk of patulin, the effort to guard apples from infection with P. expansum and to avoid long-term storage of damaged apples is necessary.
Damaged fruit was collected from the apple orchard at the National Institute of Fruit Tree Science and tests were conducted to isolate the apple blue mold pathogen Penicillium expansum. Blue mold pathogen was isolated from fruit with split skin or that had fallen to ground. The pathogen was also isolated from stored fruit raw material from a few domestic apple juice plants. The type of damage on the fruit surface and the occurrence of the disease were then investigated. All fruit with damage great enough to expose the interior of fruit was found to be infected. Fruit not damaged by human activity was observed to be infected by contact with the decay starting from stem-end fruit cracking, rough fruit skin and lenticels. Measures to prevent blue mold disease, including the use of biological agrochemicals and improvement of storage methods, were considered. Of the many commercial biological agrochemicals, Erwinia carotovora preparation was found to be highly effective in controlling this disease. CA (controlled atmosphere) storage was effective for controlling blue mold disease outbreaks. In the past few years “freshness preserving film” has been used to create the same type of atmosphere as CA storage when used in MA (modified atmosphere) packaging. Therefore, the effectiveness of MA packaging for preventing blue mold disease was investigated. Apple fruit inoculated with blue mold pathogen was placed in bags made with freshness preserving film (below MA-bags) After the passage of a set time, the size of decay areas and the amount of the mycotoxin patulin accumulating in the decay areas were measured. Storage in MA bags controlled the progress of fruit decay and markedly controlled patulin accumulation.
There is no report in Japan about the frequency of the ochratoxin producing strains on Aspergillus section Nigri and the distribution of the producing species. The surveillance study is needed for ochratoxin producing species based on the detailed classification of Aspergillus section Nigri. As a result of the investigation in Yamanashi, although most of A. niger (DNA type; AN-D-5 and AN-D-7) did not produce ochratoxin, a strain of AN-D-5 did slightly produce ochratoxin. A. carbonarius is a main ochratoxin produce species in Aspergillus section Nigri, it produce much ochratoxin A and show DNA type AN-D-4. However, some strains did not produce ochratoxins accoding to culture same conditions. Most of vineyard soil and the fungus of air born were A. japonicus (DNA type AN-D-1 and AN-D-2), and A. niger (DNA type AN-D-5 and AN-D-7), A. carbonarius of DNA type AN-D-4 was one share among 129 strains from soil. This isolate frequency was very low. If management after harvest is performed proper, it will be thought that there is very little contamination to food.
Ministry of Agriculture, Forestry and Fisheries (MAFF) developed “the Standard Operating Procedure for Food Safety Risk Management in Ministry of Agriculture, Forestry and Fisheries and Ministry of Health, Labour and Welfare” which provides the standard risk management procedure for food safety. MAFF considers food safety policies in accordance with this SOP. Risk profiles and a priority list of chemical contaminants including mycotoxin, prepared in the preliminary stages of risk management, are published online. As for mycotoxin, MAFF has been working on activities such as providing guidance on risk management measures, conducting surveillance of domestic agricultural products and promoting researches based on political needs. Moreover, Japanese code of practice summarizing measures to prevent and reduce mycotoxin contamination in wheat and barley is now under development by MAFF.