Controversies in fumonisin mycotoxicology and risk assessment.

Fusarium verticillioides causes several animal diseases and the contamination maize suggests that it could adversely affect human health. The fumonisin B mycotoxins were characterized from the fungal culture material and shown to be the causative principle responsible for the major mycotoxicological effects of the fungus in experimental and farm animals. The main focus was on the toxicological effects in rats and mice, the outcome of which played an important role in setting risk assessment parameters for exposure of the fumonisins to humans. The International Agency for Research on Cancer characterized the fumonisins as Group 2B carcinogens. Several controversial findings regarding the toxicological effects of the culture material of the fungus, the genotoxicity and carcinogenicity of pure fumonisin B(1) (FB(1)) in rats have been reported that should be clarified prior to assessing the risk in humans. The underlying differences between the diets with the high protein levels are likely to sensitize the kidneys to FB(1)-induced toxic and carcinogenic effects. Several other dietary factors, such as plant extracts (antioxidants) and dietary Fe, could either stimulate or inhibit cancer induction of FB(1), which complicates the comparison of toxicological effects in experimental animals. Cognisance should be taken of the modulating role of dietary constituents as it will determine the outcome of toxicological assays and determine the threshold of an adverse effect in a specific target organ to be used in determining risk assessment parameters.

Resistance in Maize Inbred Lines to Fusarium verticillioides and Fumonisin Accumulation in South Africa.

Fusarium ear rot of maize, caused by Fusarium verticillioides, is an important disease affecting maize production worldwide. Apart from reducing yield and grain quality, F. verticillioides produces fumonisins which have been associated with mycotoxicoses of animals and humans. Currently, no maize breeding lines are known with resistance to F. verticillioides in South Africa. The objective of this study, therefore, was to evaluate 24 genetically diverse maize inbred lines as potential sources of resistance to Fusarium ear rot and fumonisin accumulation in field trials at Potchefstroom and Vaalharts in South Africa. After artificial silk channel inoculation with F. verticillioides, Fusarium ear rot development was determined at harvest and fumonisins B1, B2, and B3 quantified. A significant inbred line by location effect was observed for Fusarium ear rot severity (P ≤ 0.001), although certain lines proved to be consistently resistant across both locations. The individual inbred lines also differed considerably in fumonisin accumulation between Potchefstroom and Vaalharts, with differentiation between susceptible and potentially resistant inbred lines only being possible at Vaalharts. A greenhouse inoculation trial was then also performed on a subset of potentially resistant and highly susceptible lines. The inbred lines CML 390, CML 444, CML 182, VO 617Y-2, and RO 549 W consistently showed a low Fusarium ear rot (<5%) incidence at both Potchefstroom and Vaalharts and in the greenhouse. Two of these inbred lines, CML 390 and CML 444, accumulated fumonisin levels <5 mg kg-1. These lines could potentially act as sources of resistance for use within a maize breeding program.

Cancer initiating properties of fumonisin B1 in a short-term rat liver carcinogenesis assay.

The nature of cancer initiation by fumonisin B(1) (FB(1)) was investigated in rat liver by monitoring the effect of phenobarbital (PB) as cancer promoter and evaluating the involvement of spontaneously initiated cells. A PB promoting regimen (0.05% in the diet) stimulated the outgrowth of FB(1)-induced placental glutathione S-transferase (GSTP) positive initiated hepatocytes. Reversion of the FB(1)-induced GSTP(+) foci was noticed in the absence of a promoting regimen. Younger rats were shown to be more sensitive to the induction of GSTP(+) foci by FB(1). Cancer initiation by FB(1) was associated with a hepatotoxic effect, which was less pronounced in older rats presumably due to a reduced intake. A specific role of spontaneously initiated cells and their promotion by FB(1) into the development of eosinophilic clear cell foci could not be established under the present experimental conditions. The ability of different stimuli to selectively promote the outgrowth of FB(1) initiated cells further verifies the cancer initiating potency of this apparent non-genotoxic mycotoxin. The underlying mechanism(s) involved in the genesis of the initiated hepatocytes is not known at present.

Corneal ulcer due to Neocosmospora vasinfecta in an immunocompetent patient.

We report a case of Neocosmospora vasinfecta keratitis in a 55-year-old man. While the patient did not recall any specific trauma or eye injury, he might have sustained a trivial wound during the course of his duties as a farmer. Direct examination of corneal scrapings revealed fungus filaments. As topical treatment with natamycin and econazole and subsequent systemic ketoconazole therapy failed, a full thickness therapeutic keratoplasty was performed. Post-operative treatment with amphotericin B and clotrimazole combined with cyclosporine resulted in a complete cure. The residual corneal infiltration in the recipient cornea became clear in a week. The fungal isolate was initially identified as a Fusarium species, but later reidentified through the use of morphological characteristics and sequence analysis of the internal transcribed spacer region as N. vasinfecta. The latters is a Hypocrealean fungus not hitherto reported as a causative agent of keratomycosis.

Sphingoid base levels in humans consuming fumonisin-contaminated maize in rural areas of the former Transkei, South Africa: a cross-sectional study.

High incidences of oesophageal cancer are associated with the consumption of subsistence-grown maize by rural populations in the former Transkei region of Eastern Cape Province, South Africa. This cross-sectional study was conducted in the north-eastern magisterial area of Bizana (a previously low oesophageal cancer incidence area) and the south-eastern area of Centane (a previously high incidence area). Plasma and urine samples of male and female participants were analysed for the sphingoid bases, sphinganine and sphingosine. Good home-grown and visibly mouldy maize samples, collected from the households of the participants, were analysed for fumonisin B(1), B(2) and B(3). Plasma sphinganine/sphingosine ratios in males and females were significantly lower (p < 0.05) due to lower sphinganine levels in Bizana compared to Centane. In contrast, the urinary female and combined (males + females) sphinganine/sphingosine ratios were significantly higher (p < 0.05) in Bizana due to the significantly lower (p < 0.05) urinary sphingosine levels. Interestingly, urinary sphingoid base levels were significantly lower (p < 0.05) in males than females within each area. Based on the mean total fumonisin levels in good maize, the estimated mean probable daily intake (PDI) was 5.8 microg kg(-1) body weight day(-1) in Bizana during 2000 and 4.4 and 6.7 5.8 microg kg(-1) body weight day(-1) in Centane during 1997 and 2000, respectively, exceeding the maximum tolerable daily intake proposed by JECFA. However, there was no significant difference in the mean total fumonisin levels in the maize between the magisterial areas. The observed differences in plasma and urinary sphingoid base levels could not be ascribed as a biomarker of fumonisin exposure and further studies at an individual level are required.

Exposure assessment for fumonisins in the former Transkei region of South Africa.

The fumonisins are mycotoxins produced mainly by Fusarium verticillioides and F. proliferatum in maize, the predominant cereal staple for subsistence farming communities in southern Africa. In order to assess exposure to these mycotoxins in the Bizana (now known as Mbizana) and Centane magisterial areas of the former Transkei region of the Eastern Cape Province of South Africa, the actual maize consumption by different age groups in these communities was measured. In the groups 1-9 years (n = 215) and 10-17 (n = 240) years, mean consumption (+/-standard error) was 246 +/- 10.8 and 368 +/- 10.3 g per person day(-1), respectively, with no significant difference (p > 0.05) between the magisterial areas. For adults (18-65 years) mean maize consumption in Bizana (n = 229) and Centane (n = 178) were significantly different (p < 0.05) at 379 +/- 10.5 and 456 +/- 11.9 g per person day(-1), respectively. An exposure assessment was performed by combining the maize consumption distribution with previously determined levels of total fumonisin (fumonisins B(1) and B(2) combined) contamination in home-grown maize in these two areas. Assuming an individual adult body weight of 60 kg, fumonisin exposure in Bizana, an area of relatively low oesophageal cancer incidence, was 3.43 +/- 0.15 microg kg(-1) body weight day(-1), which was significantly lower (p < 0.05) than that in Centane (8.67 +/- 0.18 microg kg(-1) body weight day(-1)), an area of high oesophageal cancer incidence. Mean fumonisin exposures in all age groups in both Bizana and Centane were above the provisional maximum tolerable daily intake (PMTDI) of 2 microg kg(-1) body weight day(-1) set by the Joint FAO/WHO Expert Committee on Food Additives.

Human dietary exposure to fumonisin B1 from Iranian maize harvested during 1998-2000.

Fumonisin B(1) (FB(1)) is the most abundant of the fumonisin mycotoxins, mainly produced in maize by F. verticillioides and F. proliferatum. A previous study on the FB1 contamination of maize harvested in Mazandaran and Isfahan Provinces of Iran in 1998 and 1999 demonstrated contamination in both provinces. This present study was undertaken to further investigate the variation in levels of contamination and to estimate possible levels of human exposure to fumonisins in Iran. The mean level of FB(1) in 49 visually healthy maize samples collected from Mazandaran Province during 2000 was 6.14 mg/kg, which is higher than that found during 1998 and 1999 (2.27 and 3.18 mg/kg, respectively). Although these levels are higher than the Iranian legislative limits for fumonisins in maize intended for humans, the relatively low estimated consumption of maize in Iran (3.3 g/person/day) implies that average exposures (0.011 and 0.215 microg/kg body weight/day in Isfahan and Mazandaran, respectively) are within the provisional maximum tolerable daily intake of 2 microg/kg body weight/day set by the Joint FAO/WHO Expert Committee on Food Additives. Nevertheless, certain sections of the population who may consume higher amounts of maize or who may replace all or some of their consumption of other cereals with maize, could well exceed this limit.

Impact of mechanical shelling and dehulling on Fusarium infection and fumonisin contamination in maize.

Mechanical shelling and dehulling methods were tested to evaluate their impact on Fusarium infection and fumonisin contamination in maize. All shelling methods which were tested were found to damage the grains. The IITA sheller caused the highest level (up to 3.5%) of damage. Fusarium populations were higher on damaged grains, the highest being recorded from grains damaged by the IITA sheller (2,533.3 cfu g(-1)). Fumonisin levels were higher in damaged grains, the highest being in maize shelled with the IITA sheller (2.2 mg kg(-1)). Fumonisin levels were positively and significantly correlated with the percentage of damage caused by the shelling methods, and with the number of Fusarium colonies in maize. Mechanical dehulling methods significantly reduced fumonisin levels in maize, resulting in a mean reduction of 62% for Mini-PRL, 65% for Engelberg, and 57% for the attrition disc mill. It is important for farmers to choose appropriate shelling methods to reduce mycotoxin contamination. Dehulling should be widely promoted for the reduction of mycotoxins in maize.

Mango malformation disease and the associated fusarium species.

ABSTRACT Mango malformation disease (MMD) occurs in Asia, Africa, and the Americas and was first reported in India in 1891. The vegetative form of MMD was first reproduced in 1966 with Fusarium moniliforme and the floral form with isolates of F. moniliforme var. subglutinans from both vegetative shoots and floral tissue. The fungi were subsequently recognized as F. subglutinans. In 2002, a new species, F. mangiferae, was established based on nuclear and mitochondrial DNA sequences; it included strains of F. subglutinans from Egypt, Florida, Israel, Malaysia, and South Africa, some of which had been shown to cause MMD by artificial inoculation. At least three additional taxa have been associated with MMD: F. sterilihyphosum from Brazil and South Africa, and Fusarium sp. nov. and F. proliferatum (teleomorph: Gibberella intermedia) from Malaysia. To date, Koch's postulates have not been completed with them. In the future, gene sequencing will be essential to identify the Fusarium spp. that are associated with MMD. Work remains to be done on the morphology, sexual compatibility, pathogenicity, and toxigenicity of these taxa.

Natural occurrence of Fusarium and subsequent fumonisin contamination in preharvest and stored maize in Benin, West Africa.

The natural occurrence of Fusarium and fumonisin contamination was evaluated from 1999 to 2003 in both preharvest and stored maize produced by small-scale farmers in four agroecological zones of Benin. Mycological analyses revealed a predominance of both Fusarium and Aspergillus in maize samples compared to other genera. The two Fusarium species most commonly isolated from maize were Fusarium verticillioides (68%) and Fusarium proliferatum (31%). Atypical isolates of F. verticillioides with some characteristics of Fusarium andiyazi but apparently closer to F. verticillioides, because the isolates were all high fumonisin producers, were also found only on preharvest maize. Study of F. verticillioides strains showed the presence of extremely high fumonisin producers in Benin with total fumonisin levels ranging from 8240 to 16,690 mg/kg. Apart from 2002-2003, Fusarium occurrence was not significantly different from one zone to another, although a slight decrease was observed from south, humid, to north, drier. Fusarium occurrence varied somewhat from one season to another. It significantly decreased over the 6 months of storage. Widespread fumonisin occurrence in maize was observed. Most of the maize samples collected were found positive for fumonisin with levels ranging from not detected to 12 mg/kg in 1999-2000, 6.7 mg/kg in 2000-2001 and 6.1 mg/kg in 2002-2003. Fumonisin levels in maize were found to be significantly higher in the two southern zones during all the surveys. The highest mean total fumonisin level was detected in 1999-2000 in maize samples from the southern Guinea Savannah (SGS) (12 mg/kg), whereas in both 2000-2001 and 2002-2003, it was in samples from the forest mosaic savannah (FMS) (6.7 and 6.1 mg/kg, respectively). Fumonisin levels varied from one season to another and, throughout the storage time, showing a decreasing trend in each zone. However, this decrease was not significant every season. An increasing trend was observed during some seasons in the SGS and northern Guinea Savannah (NGS) zones. The results of this study emphasise that farmers and consumers, not only in Benin but also in other West African countries, should be alerted to the danger of fumonisin contamination in maize.

Fate of aflatoxins and fumonisins during the processing of maize into food products in Benin.

The fate of aflatoxins and fumonisins, two mycotoxins that cooccur in maize, was studied through the traditional processing of naturally contaminated maize in mawe, makume, ogi, akassa, and owo, maize-based foods common in Benin, West Africa. Levels of total aflatoxin and fumonisin were measured at the main unit operations of processing, and the unit operations that induce significant reduction of mycotoxin level were identified. Overall reduction of mycotoxin level was more significant during the preparation of makume (93% reduction of aflatoxins, 87% reduction of fumonisins) and akassa (92% reduction of aflatoxins, 50% reduction of fumonisins) than that of owo (40% reduction of aflatoxins, 48% reduction of fumonisins). Sorting, winnowing, washing, crushing combined with dehulling of maize grains were the unit operations that appeared very effective in achieving significant mycotoxin removal. Aflatoxins and fumonisins were significantly recovered in discarded mouldy and damaged grains and in washing water. Fermentation and cooking showed little effect. During the preparation of ogi and akassa, reduction of fumonisin levels measured in food matrix was lower (50%) compared to mawe and makume, probably due to significant fumonisin release in ogi supernatant. Consequently, the use of ogi supernatant for preparing beverages or traditional herbal medicines could be harmful as it is likely to be contaminated with mycotoxin from the raw maize.

Fumonisin contamination of a corn sample associated with the induction of hepatocarcinogenesis in rats-role of dietary deficiencies.

A corn sample associated with a field outbreak of equine leukoencephalomalacia in Pennsylvania, USA, during 1983/1984 and induced hepatotoxic and hepatocarcinogenic effects when fed to male Fischer rats was analyzed mycologically and chemically for the presence of fumonisins (FB), hydrolysed FB derivatives and aflatoxins (AFB). Fusarium verticillioides was found to be the predominant fungal contaminant in the corn sample but Aspergillus flavus was also present. Trace amounts (0.1 microg/kg) of AFB(1) and AFB(2) and a total FB level of 33.5 mg/kg (FB(1):FB(2):FB(3) ratio of 9:2.3:1) were found. No hydrolysed FB derivatives or AFG(1) and AFG(2) were detected. Based on the chemical stability of the fumonisins in different corn cultures of F. verticillioides kept at 4 degrees C over a period of 13-20 years, a level of approximately 55 mg/kg of total FB is estimated in the original corn sample. A possible role of certain dietary constituents such as the high protein content and deficiencies in certain micronutrients is evaluated to address differences in the organ-specific toxicity of FB(1) in rats using commercial, semi-purified, purified and corn-only diets.

Fusarium dimerum as a cause of human eye infections.

Fusarium dimerum, typically a soil fungus, was isolated from an adult male suffering from a corneal ulcer following an injury to the eye. This fungus has not been described to cause human infections in South Africa and has not been recorded from soil, plant or organic material in this country. The macro- and microscopic characteristics of the isolate were found to be indistinguishable from described strains. Its authenticity was confirmed by comparing it to other human isolates from the eye obtained in the USA, thus rendering this the first report of F. dimerum from an eye infection in a human in South Africa.

Fumonisin B(1) in maize harvested in Iran during 1999.

The fumonisin B(1) (FB(1)) contamination of maize collected in two areas of Iran during 1999 was determined. The 20 maize samples from Mazandaran Province, situated on the Caspian littoral of Iran, consisted of random samples of farmers' lots and were all contaminated with FB(1) at a mean level of 3.18 mg kg(-1) (range 0.68-7.66 mg kg(-1)). The 10 samples (of the same maize cultivar) from Isfahan Province in central Iran were purchased as maize cobs in local retail markets and had mean FB levels of 0.22 mg kg(-1) (mean of all samples, 6/10 samples positive, range <0.01-0.88 mg kg(-1)). The FB levels in Mazandaran, an area of high oesophageal cancer, were significantly (p < 0.0001) higher than the FB levels found in maize from Isfahan, an area of low oesophageal cancer in Iran.

Interaction of fumonisin B(1) and aflatoxin B(1) in a short-term carcinogenesis model in rat liver.

The co-existence of the fumonisin and aflatoxin mycotoxins in corn merited studies to investigate their possible synergistic toxicological and carcinogenic effects. When utilising a short-term carcinogenesis model in rat liver, both the compounds exhibited slow cancer initiating potency as monitored by the induction of foci and nodules stained positively for the placental form of gluthatione-S-transferase (GSTP(+)). However, when rats were treated in a sequential manner with AFB(1) and FB(1) the number and size of GSTP(+) lesions significantly increased as compared to the separate treatments. Histopathological analyses indicated that the individual treatments showed far less toxic effects, including occasional hepatocytes with dysplastic nuclei, oval cell proliferation and, in the case of FB(1), a few apoptotic bodies in the central vein regions. The sequential treatment regimen induced numerous foci and dysplastic hepatocyte nodules, and with oval cells extending from the periportal regions into the centrilobular regions. This would imply that, in addition to the cancer promoting activity of FB(1) of AFB(1)-initiated hepatocytes, the AFB(1) pre-treatment enhanced the FB(1) initiating potency, presumably by rendering the liver more susceptible to the toxic effects of FB(1). The co-occurrence of AFB(1) and FB(1) in corn consumed as a staple diet could pose an increased risk and should be included in establishing risk assessment parameters in humans.