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Occurrence of Fusarium species and mycotoxins in nepalese maize and wheat and the effect of traditional processing methods on mycotoxin levels.

Maize (Zea mays) and wheat (Triticum aestivum) collected in the foothills of the Nepal Himalaya Mountains were analyzed for Fusarium species and mycotoxins: fumonisins, nivalenol (NIV), and deoxynivalenol (DON). Predominant species were Gibberella fujikuroi mating population A (F. moniliforme) in maize and F. graminearum in maize and wheat; G. fujikuroi mating population D (F. proliferatum), F. acuminatum, F. avenaceum, F. chlamydosporum, F. equiseti, F. oxysporum, F. semitectum, and F. torulosum were also present. Strains of G. fujikuroi mating population A produced fumonisins, and strains of F. graminearum produced NIV or DON. By immunoassay or high-performance liquid chromatography, fumonisins were >1000 ng/g in 22% of 74 maize samples. By immunoassay or fluorometry, NIV and DON were >1000 ng/g in 16% of maize samples but were not detected in wheat. Fumonisins and DON were not eliminated by traditional fermentation for producing maize beer, but Nepalese rural and urban women were able to detoxify contaminated maize by hand-sorting visibly diseased kernels.

Beer↗

Mycotoxins in ingredients of animal feeding stuffs: I. Determination of Alternaria mycotoxins in oilseed rape meal and sunflower seed meal.

A multi-toxin method was developed for the detection of some of the known Alternaria mycotoxins, altenuene, iso-altenuene, alternariol, alternariol monomethyl ether, tenuazonic acid and altertoxin I in oilseed rape meal and sunflower seed meal. The method involves extraction of the toxins with an acidified mixture of acetonitrile: aqueous potassium chloride solution, followed by liquid-liquid extraction and further purification using gel permeation chromatography. The final extract is then examined on a reverse phase high performance liquid chromatographic gradient system with both fluorescence and UV detection. The average recoveries found were 94, 84, 109, 85, 66 and 93% for spiked oilseed rape meal samples and 91, 89, 96, 75, 61 and 102% for spiked sunflower meal samples with limits of determination of about 40, 50, 50, 40, 350 and 200 micrograms/kg for the above toxins, respectively. Detection limits were about 30% of these values. Thirty samples of oilseed rape meal and 22 samples of sunflower meal were examined using the methods developed. Twenty of the oilseed rape products which had been grown in the UK were free from contamination while 10 contained one or more of tenuazonic acid, alternariol and alternariol monomethyl ether. In contrast, all of the sunflower meal samples, of Argentinean, Indian or EC origin, were contaminated with one or more of alternariol, alternariol monomethyl ether and tenuazonic acid. Average levels of alternariol, alternariol monomethyl ether and tenuazonic acid were 68, 55 and 730 micrograms/kg, respectively for the contaminated samples of oilseed rape meal and 180, 100 and 1900 micrograms/kg, respectively for the contaminated samples of sunflower seed meal.

Alternaria↗

Mycotoxins in cereal grain. Part VI. The effect of ochratoxin A on growth and tissue residues of the mycotoxin in broiler chickens.

Eight weeks experiment on effects of continuous feeding of mycotoxin graded levels (0; 9.5; 1.0; .5; 2.0 mg ochratoxin A (OA) per kilogram of feed) to male and female broiler chickens was carried out. Birds' mortality was similar in all groups including control. Macroscopical veterinary examination did not reveal any changes in livers and kidneys. Males were found to be more sensitive to OA than females. The depression of body weight was found to be proportional to level of OA. The reduction of body weight of males (30%) was more significant than that of females (20%). Feed consumption per one kilogram of body weight was dependent on OA concentration in feed. In livers and white muscles of birds fed 1.0; 1.5 and 2.0 mg OA/kg feed and in red muscles of birds fed 1.5 and 2.0 mg OA/kg feed residues of OA were observed. OA disappeared completely from all mentioned tissues after 4 days when birds were fed with OA free feed.

Animal Feed↗

Mycotoxins in cereal grain. Part IV. Inactivation of ochratoxin A and other mycotoxins during ammoniation.

Addition of ammonia to final concentration 2% inactivates ochratoxin A, aflatoxin, citrinin, penicillic acid and partially zearalenon at temperature 20-50 degrees C. Detoxification of contaminated cereal grain (wheat, corn or barley) can be performed on a farm using ammoniation without special investment during 4 to 6 weeks. Ammoniation changes nutritional value of grain as feed in a small extent.

Ammonia↗

Mycoflora and mycotoxins of peanut (Arachis hypogaea L.) seeds in Egypt. 1--Sugar fungi and natural occurrence of mycotoxins.

Sixty-four species and 2 varieties which belong to nineteen genera of fungi were identified from 40 peanut seed samples collected from different places in Egypt by using a dilution-plate method on glucose-Czapek's medium. The most frequent genera were Aspergillus (21 species & 2 varieties), Penicillium (16 species) and Fusarium (6 species). A. flavus, A. fumigatus, A. niger, P. chrysogenum and F. oxysporum were the most common fungal species. Forty seven percent of the samples proved to be toxic to brine shrimp (Artemia salina) larvae. Thin-layer chromatographic analysis revealed that peanuts were contaminated by aflatoxins (11 samples), trichothecene-toxins (10 samples), zearalenone (one sample) and citrinin (one sample). We believe that this is the first report of the natural occurrence of zearalenone and trichothecene-toxins in peanuts.

Animals↗

Toxicity, metabolism, and impact of mycotoxins on humans and animals.

The worldwide contamination of foods and feeds with mycotoxins is a significant problem. Mycotoxins are secondary metabolites of molds that have adverse effects on humans, animals, and crops that result in illnesses and economic losses. Aflatoxins, ochratoxins, trichothecenes, zearelenone, fumonisins, tremorgenic toxins, and ergot alkaloids are the mycotoxins of greatest agro-economic importance. Some molds are capable of producing more than one mycotoxin and some mycotoxins are produced by more than one fungal species. Often more than one mycotoxin is found on a contaminated substrate. Factors influencing the presence of mycotoxins in foods or feeds include environmental conditions related to storage that can be controlled. Other extrinsic factors such as climate or intrinsic factors such as fungal strain specificity, strain variation, and instability of toxigenic properties are more difficult to control. Mycotoxins have various acute and chronic effects on humans and animals (especially monogastrics) depending on species and susceptibility of an animal within a species. Ruminants have, however, generally been more resistant to the adverse effects of mycotoxins. This is because the rumen microbiota is capable of degrading mycotoxins. The economic impact of mycotoxins include loss of human and animal life, increased health care and veterinary care costs, reduced livestock production, disposal of contaminated foods and feeds, and investment in research and applications to reduce severity of the mycotoxin problem. Although efforts have continued internationally to set guidelines to control mycotoxins, practical measures have not been adequately implemented.

Animal Feed↗

Sampling foods for mycotoxins.

It is difficult to obtain precise and accurate estimates of the true mycotoxin concentration of a bulk lot when using a mycotoxin-sampling plan that measures the concentration in only a small portion of the bulk lot. A mycotoxin-sampling plan is defined by a mycotoxin test procedure and a defined accept/reject limit. A mycotoxin test procedure is a complicated process and generally consists of several steps: (1) a sample of a given size is taken from the lot, (2) the sample is ground (comminuted) in a mill to reduce its particle size, (3) a subsample is removed from the comminuted sample, and (4) the mycotoxin is extracted from the comminuted subsample and quantified. Even when using accepted test procedures, there is uncertainty associated with each step of the mycotoxin test procedure. Because of this variability, the true mycotoxin concentration in the lot cannot be determined with 100% certainty by measuring the mycotoxin concentration in a sample taken from the lot. The variability for each step of the mycotoxin test procedure, as measured by the variance statistic, is shown to increase with mycotoxin concentration. Sampling is usually the largest source of variability associated with the mycotoxin test procedure. Sampling variability is large because a small percentage of kernels are contaminated and the level of contamination on a single seed can be very large. Methods to reduce sampling, sample preparation and analytical variability are discussed.

Food Analysis↗

Detecting mycotoxins in agricultural commodities.

It is difficult to obtain precise and accurate estimates of the true mycotoxin concentration of a bulk lot when using a mycotoxin-sampling plan that measures the concentration in a small portion of the bulk lot. A mycotoxin-sampling plan is defined by a mycotoxin test procedure and a defined accept/reject limit. A mycotoxin test procedure is a complicated process and generally consists of several steps: (a) a sample is taken from the lot, (b) the sample is ground (comminuted) in a mill to reduce particle size, (c) a subsample is removed from the comminuted sample, and (d) the mycotoxin is extracted from the comminuted subsample and quantified. Even when using accepted test procedures, there is variability associated with each step of the mycotoxin test procedure. Because of this variability, the true mycotoxin concentration in the lot cannot be determined with 100% certainty by measuring the mycotoxin concentration in a sample taken from the lot. The variability for each step of the mycotoxin test procedure, as measured by the variance statistic, is shown to increase with mycotoxin concentration. Sampling is usually the largest source of variability associated with the mycotoxin test procedure. Sampling variability is large because a small percentage of kernels are contaminated and the level of contamination on a single seed can be very large. Methods to reduce sampling, sample preparation, and analytical variability are discussed.

Aflatoxins↗

Combined toxic effects of mycotoxins.

It is known for many years that several food items, derived from plants infected by fungi in the field during growing of the plant or during harvest and storage of the food item, can contain concomitantly different mycotoxins. As these combined mycotoxins occur simultaneously in the food item, consumption of the food will lead to a combined intake depending on the absorption rates of the different mycotoxins. Therefore, the question is justified whether such a combined intake of mycotoxins would lead to a possible higher risk for adverse health effects than the intake of one of these mycotoxins alone. It will be dealt with on the basis of some practical cases of such combined intake of mycotoxins of which research data are available. This is the case for citrinin and ochratoxin A, but as the workshop focuses on trichotecenes and so this paper concentrates on these. When the mycotoxins are of similar structure and of the same species, or of the same families, it is likely to expect that the mode of action of the mycotoxins and or the toxicity profiles will be quite similar. This indicates that such related mycotoxins are likely to exert only additive effects, which is important to know. In terms of risk assessment, these mycotoxins could be dealt with by establishing a group daily tolerable intake (TDI) or a provisional tolerable weekly intake (PTWI). In terms of risk assessment those mycotoxins which interact in synergistic manner are of more concern. It is concluded that, at present tools are not fully developed to establish the type of interaction or whether there is any interaction at all.

Animals↗

A review of mycotoxins in indoor air.

Mycotoxins are chemical compounds, produced by a variety of fungi, that can cause illness in humans and animals. This paper is a review of literature on mycotoxins with emphasis on mycotoxins in indoor air. Consideration is given to specific mycotoxins identified in indoor air, indoor sources of the mycotoxins, factors affecting mycotoxin production, potential health effects indicated by animal laboratory studies, and case studies of possible human inhalation health effects of these mycotoxins. Historically, mycotoxicoses have been associated with consumption of moldy grain. In recent years, some attention has been given to mycotoxins in dust from agricultural environments, but relatively few studies have examined mycotoxins or mycotoxin-producing molds in indoor environments. The few indoor studies suggest that mycotoxicoses may occur in some indoor environments. More studies are needed to understand the potential for mycotoxin occurrence and significance in indoor environments.

Air Pollutants, Occupational↗

Solvent comparison in the isolation, solubilization, and toxicity of Stachybotrys chartarum spore trichothecene mycotoxins in an established in vitro luminescence protein translation inhibition assay.

It is well known that non-viable mold contaminants such as macrocyclic trichothecene mycotoxins of Stachybotrys chartarum are highly toxinigenic to humans. However, the method of recovering native mycotoxin has been without consensus. Inconsistencies occur in the methods of isolation, suspension, preparation, and quantitation of the mycotoxin from the spores. The purpose of this study was to provide quantitatively comparative data on three concurrent preparations of 10(6)S. chartarum spores. The experiments were designed to specifically evaluate a novel method of mycotoxin extraction, solubilization, and the subsequent inhibitory effect in an established in vitro luminescence protein translation assay from 30 day-old spores. The mycotoxin-containing spores swabbed from wallboard cultures were milled with and without glass beads in 100% methanol, 95% ethanol, or water. Milled spore lysates were cleared of cell debris by filter centrifugation followed by a second centrifugation through a 5000 MWCO filter to remove interfering proteins and RNases. Cleared lysate was concentrated by centrivap and suspended in either alcohol or water as described. The suspensions were used immediately in the in vitro luminescence protein translation assay with the trichothecene, T-2 toxin, as a control. Although, mycotoxin is reported to be alcohol soluble, the level of translation inhibition was not reliably satisfactory for either the methanol or ethanol preparations. In fact, the methanol and ethanol control reactions were not significantly different than the alcohol prepared spore samples. In addition, we observed that increasing amounts of either alcohol inhibited the reaction in a dose dependent manner. This suggests that although alcohol isolation of mycotoxin is desirable in terms of time and labor, the presence of alcohol in the luminescence protein translation reaction was not acceptable. Conversely, water extraction of mycotoxin demonstrated a dose dependent response, and there was significant difference between the water controls and the water extracted mycotoxin reactions. In our hands, water was the best extraction agent for mycotoxin when using this specific luminescence protein translation assay kit.

Ethanol↗

Induction of Hsp 70 in Vero cells in response to mycotoxins cytoprotection by sub-lethal heat shock and by Vitamin E.

This paper analysed the toxicity mechanisms of several mycotoxins using Hsp 70 expression, cytoprotection of Vero cells by sub-lethal heat shock (sub-LHS) and Vitamin E. Our aim was (i) to determine whether Citrinin (CTN), Zearalenone (ZEN) and T2 toxin (T2) could induce the expression of Hsp 70, (ii) to check whether or not elevated levels of Hsp and Vitamin E pre-treatment could provide cytoprotection from these mycotoxins, and finally (iii) to emphasize the eventual involvement of oxidative stress on mycotoxin's toxicity. Our study demonstrated that the three examined mycotoxins induced Hsp 70 expression in a dose-dependent manner. A cytoprotective effect of Hsp 70 was obtained when Vero cells were exposed to sub-lethal heat shock followed by a 12h recovery prior to mycotoxins treatment and evidenced by a reduction of their cytolethality. This cytoprotection suggested that Hsp 70 might constitute an important cellular defence mechanism. A cytoprotective action was also obtained although at lesser extent, when cells were pre-treated with an antioxidant agent, the Vitamin E before mycotoxins treatment. This Vitamin E cytoprotection evoked the involvement of oxidative stress in mycotoxins induced toxicity, which was further, confirmed by the reduction of Hsp 70 expression when cells were pre-treated with Vitamin E prior to mycotoxins. Our data clearly shows that oxidative stress is certainly involved in the toxicity of the three studied mycotoxins, Citrinin, Zearalenone and T2 toxin and may therefore constitutes a relevant part in their toxicities; however, at variable extent from one mycotoxin to another.

Animals↗

Stachybotrys, a mycotoxin-producing fungus of increasing toxicologic importance.

BACKGROUND: Stachybotrys as a fungus has been implicated as a source of mycotoxins. While the toxicity of several well-known mycotoxins (aflatoxins) is well documented, recent studies on Stachybotrys have raised the question that mycotoxins produced by this fungus may be responsible for the health effects of occupants in water-damaged buildings. METHODS: Published articles regarding Stachybotrys-related mycotoxins were reviewed with particular focus on human toxicity. RESULTS: A critical review of papers, reports, and studies on Stachybotrys mycotoxins revealed only descriptive reports of suspected animal and human poisoning secondary to consumption of mold-contaminated food products. No studies of good toxicologic and epidemiologic designs answer whether airborne mycotoxins produced by Stachybotrys could produce specific human toxicity. CONCLUSIONS: Current data on the toxicology of mycotoxins produced by Stachybotrys demonstrate that this group of mycotoxins is capable of producing immunosuppression and inflammatory insults to gastrointestinal and pulmonary systems. Case control study and case reports have suggested a possible association with environmental exposure to Stachybotrys mycotoxins, although a firm causal relationship has not been firmly established. Additional studies are needed to document that humans with sufficient exposure to these mycotoxins develop compatible clinical and pathologic pictures as demonstrated in animal models.

Animals↗