PART III PROCESS SAFETY
2. Mycotoxins and Relevant Fungi
Filamentous fungi, taken as a whole, produce thousands of toxic compounds.
However, the more important mycotoxins belong to species of Aspergillus, Fusarium, and Penicillium. Only a small number of mycotoxins is considered of TABLE1. Historic overview on mycotoxicoses
Year Toxicosis Toxin Species
994 Ergotism Ergot alkaloids Claviceps purpurea
1890 Cardiac beri-beri Citreoviridin Penicillium citreonigrum
1913 & 1944 Alimentary toxic aleukia Trichotecenes Fusarium (mainly T-2 toxin) sporotrichioides 1952 Balkan endemic nephropathy Ochratoxins Penicillium verrucosum
1960 Turkey X disease Aflatoxins Aspergillus flavus
1989 Hole in the head syndrome Fumonisins Fusarium verticillioides
prime importance, namely, aflatoxins, ochratoxin A (OTA), patulin, deoxyni- valenol (DON), fumonisins, nivalenol (NIV), T2-toxin, and zearalenone (ZEA) (CAST, 2003). The chemical structures of selected compounds are demonstrated in Figure 1. Identification of the fungi is carried out by microscopic examination of the isolated fungus with the conidiophore (the sporing structure) being of par- ticular importance in this respect (Figures 2 and 3). This is perhaps not ade- quately appreciated as an important factor in mycotoxicology. However, it is relevant to the detection of areas within the commodity system where contami- nation occurs. Biochemical methods, including nucleic acid analysis, are being increasingly used to confirm existing identifications. Figure 4 demonstrates how fungal identifications are relevant to the field of mycotoxicology. A review of these issues is available, which should be consulted (Paterson et al., 2004). It con- siders predominately the penicillia. However, the points raised apply equally to other mycotoxigenic fungi.
Mycotoxins are classified by the International Agency for Research on Cancer (IARC) according to their toxicity, aflatoxins being the only ones proven to be carcinogenic to humans (IARC, 1993). Others have provided evidence that enables their classification in various other lesser categories (Table 2). Only the more important mycotoxins will be considered in this chapter.
O O OCH3
CH2OH O
O O
O O
H OH H H 8
O OH
Aflatoxin B1 Deoxynivalenol
Patulin Ochratoxin A
COOH O
CI N
H
O O
H CH3
HO O
O
OH O
FIGURE1. Chemical structure of some mycotoxins.
FIGURE2. Black Aspergillus on the surface of grapes.
2.1. Aflatoxins
Aspergillus flavus and A. parasiticus are the main producers of aflatoxins.
Aflatoxins B1and B2are produced by both species but only A. parasiticus is able to produce detectable amounts of G1 and G2. B and G refer to the blue and green colours observed after thin layer chromatography (TLC). The numerical FIGURE3. SEM image of a black Aspergillus conidiophore (sporing structure).
designation is for the different mobility (Rf value) of the compounds. Aflatoxin M1(M for milk) is a very important toxin in milk and products and is a minor metabolic product of these Aspergillus species. However, it apparently occurs mainly in dairy products as a transformation product by cows of the aflatoxin B1 contained in animal feeds. Aflatoxins were the first mycotoxins to have their con- tent regulated in foodstuff. In fact, in all countries where regulation exists for mycotoxins, there are tolerances concentrations for aflatoxins in foods or animal feedstuffs (CAST, 2003).
2.2. Ochratoxin A
Ochratoxin A (OTA) is produced by particular Aspergillus and Penicillium species (Table 3). The penicillia are probably the more important producers on grains (CAST, 2003). In other commodities, the most common are A. carbonarius,
1. Identify non-mycotoxigenic disease and pest fungi
1. Identify non-mycotoxigenic biodeterioration fungi
1. Identify mycotoxigenic fungi 2. Identify interacting fungi 1. Infrastructure for identifications, e.g.
funds, equipment, culture collections, training and research 2. Revenue from identifications 3. Costs and consequences from inaccurate identifiations or none 1. Ensure mycotoxin fungi eliminated
2. Identifications of fungal biocontrol agents to ensure they prevail 3. Collection of non-mycotoxigenic strains for biocontrol, food, and biotechnology
Control sytem
Commodity system
Spoilage system
Mycotoxin system Socio-econimic System
FUNGAL IDENTIFICATIONS
FIGURE4. Relevance of mycotoxigenic fungi identification to the control of mycotoxins.
(adapted from Paterson et al., 2004).
TABLE2. Classification of the principal mycotoxins according to their toxicity
Carcinogenic toa IARC
Mycotoxin Human Animal Groupb
Aflatoxins C C 1
Fumonisins I C 2B
OTA I C 2B
ZEA - L -
Patulin - I 3
a (C) clear evidence; (L) limited evidence; (I) insufficient evidence
b Group 1 -carcinogenic agent to humans; Group 2A -probably a carcinogenic agent to humans; Group 2B -possibly a carcinogenic agent to humans; Group 3 -not classifiable
A. ochraceus and P. verrucosum. OTA has been associated with Balkan endemic nephropathy, which is potentially fatal. However, its significance has been enhanced by it being classified as possibly carcinogenic to humans by the IARC (group 2B – Table 2).
2.3. Patulin
Patulin is produced by different species of Aspergillus and Penicillium (Paterson, 2004a). Byssochlamys and Paecilomyces can also be involved. P. expansum is the most well known for causing a blue rot of apples and is associated with patulin production in apple products. Byssochlamys spp. are noteworthy as they may pro- duce heat resistant spores which survive food processing with the potential to pro- duce patulin in processed foods. The significance of this mycotoxin has been increased by the recent statutory European Union (EU) regulations in fruit prod- ucts and its presence in baby foods is of particular concern (Commission Regulation EC n˚ 1425/2003).
2.4. Fumonisins, trichothecenes, and zearalenone
Fumonisins, trichothecenes, and zearalenone are the most important mycotoxins produced by Fusarium, a producer of a wide range of toxic compounds.
Tricothecenes are related compounds and the more important are grouped into type A or type B. Type A are more toxic than type B. However, the latter are more common and therefore more relevant to food safety. Type B contains DON and NIV as the main examples, whilst a type A example is T-2 toxin. T-2 toxin TABLE3. The main mycotoxigenic fungi and most relevant associated mycotoxins
Filamentous fungi Mycotoxins
Aspergillus
A. carbonarius Ochratoxin A
A. flavus Aflatoxin B1, B2
A. ochraceus Ochratoxin A
A. parasiticus Aflatoxin B1, B2, G1, G2
Byssochlamys
B. fulva Patulin
B. nivea Patulin
Fusarium
F. cerealis Nivalenol
F. culmorum Deoxynivalenol, Nivalenol
F. equiseti Zearalenone
F. graminearum Deoxynivalenol, Nivalenol
F. poae Nivalenol
F. sporotrichioides T-2 toxin
F. verticillioides (=F. moliniforme) Fumonisin B1 Penicillium
P. expansum Patulin
P. verrucosum Ochratoxin A
was probably the cause of “alimentary toxic aleukia” which affected thousands of people in Siberia during the Second World War. It is responsible for haemor- rhagic disease in animals and is associated with the formation of oral lesions and neurotoxic effects in poultry. However the most significant effect of this and other trichothecenes is the immunosuppressive activity, which is probably linked to the inhibitory effect on the biosynthesis of macromolecules (e.g., DNA, proteins).
Fumonisins are a group of very closely related compounds (fumonisin B1, B2and B3), produced predominately in maize by F. verticillioides (F. moniliforme).
Another important Fusarium toxin is zearalenone (ZEA), which has oestrogenic properties.