During different food processing technologies, including cooking, boiling, baking, frying, baking, and pasteurizing, most mycotoxins remain chemically and thermally stable. The result of contaminated feed is the presence of mycotoxins in animal foods such as meat, eggs, and milk, thereby leading to contamination of the human plate [
3]. Regulatory limits on significant levels of mycotoxins in food and feed are established by various authorities worldwide such as the United States (US) Food and Drug Administration (FDA), the World Health Organization (WHO), the Food Agriculture Organization (FAO), and the European Food Safety Authority (EFSA) [
5]. The International Agency of Research on Cancer (IARC) classifies some important mycotoxins into categories by examining the existence of sufficient human evidence for carcinogenicity, through toxicological studies [
19].
Table 1 lists the major mycotoxins, their IARC number, the main producers, and some commonly contaminated foodstuffs, along with the US FDA and European Union (EU) regulatory limits for mycotoxin levels both in food and in animal feed [
3,
19].
The Rapid Alert System for Food and Feed (RASFF) monitors the contamination of food and feed by mycotoxins on a weekly basis in Europe. Through RASFF, all EU member states can be informed via an information exchange system and take measures to ensure the safety of food and feed [
14]. Mycotoxins consistently constitute the highest risk category for notifications, and, every year, they are found among the “top ten” hazards reported annually by the RASFF. The records of the decade 2009–2018 [
23,
24,
25,
26,
27,
28,
29,
30,
31,
32] show that aflatoxins held the highest percentages (
Table 2), while, based on the latest RASFF report for the year 2018 in the EU (
Table 3), mycotoxin notifications amounted to 655, with aflatoxin notifications totaling 536, accounting for a significant proportion (82%) [
32].
Figure 1 presents the chemical structures of the main mycotoxins.
The agricultural industry has to deal with the presence of mycotoxins in food, as it is of global importance and a major threat [
33]. In developing countries, factors such as poor food quality control, hot climate, poor production technologies, and poor crop storage conditions favor the development of fungi and the formation of mycotoxins, resulting in the more frequent occurrence of mycotoxin-contaminated foods in these countries [
34]. Huge agricultural and industrial losses in billions of dollars occur annually because 25% of the world’s harvested crops are contaminated by mycotoxins [
19]. The increased cost of production, the lowered animal production, the decreased market values, the irregularity of production [
35], the regulatory enforcement, and the testing and other quality control measures [
9] are some of the significant sources of economic loss due to the occurrence of mycotoxins in food and feed.
2.1. Aflatoxins
Aflatoxins are secondary metabolites, and they belong to the category of difuranocoumarins [
44]. Under warm and humid conditions,
Aspergillus flavus, A. nomius, and
A. parasiticus produce AFs [
22,
45], commonly found in food and feeds. The species
A. flavus and
A. parasiticus are found worldwide in the soil and in the air [
46], preferring to grow at temperatures between 22 and 35 °C and a
w between 0.95 and 0.98 [
47]. Other species producing aflatoxins similar to
A. flavus are
A. zhaoqingensis and
A. bombycis, while those similar to
A. parasiticus are
A. toxicarius and
A. parvisclerotigenus. Moreover,
A. pseudotamarii,
A. ochraceoroseus,
A. rambellii,
A. toxicarius,
Emericella astellata,
E. olivicola, and
E. venezuelensis are some species of mycotoxin producers. Τwo other recently described aflatoxigenic species are
A. minisclerotigenes and
A. arachidicola [
48].
AFs are the best known among all mycotoxins, because of their serious impact on human and animal health. Four main types of aflatoxins are the most studied among more than 20 known ones; these are aflatoxin AFB1, AFB2, AFG1, and AFG2, named after the fluorescence they display in UV light (B for blue and G for green). The hydroxylated metabolites of AFB1 and AFB2 are aflatoxin M1 (AFM1) and aflatoxin M2 (AFM2), which are present in the meat of animals that consumed aflatoxin-contaminated feed, as well as animal products such as eggs, milk, and cheese [
22]. Aflatoxin B1 is a carcinogenic substance (according to the classification by the IARC in 1987) (category 1A), while AFM1 is a potentially carcinogenic substance (category 2B) [
49], with a toxicity range of B1 > G1 > B2 > G2 [
50]. AFB1 is considered to be the most potent carcinogenic toxin known in mammals [
51], and food contamination should be reduced to the lowest possible level, since no food or health organization established a tolerable daily intake for humans (tolerable daily intake, TDI) [
52]. Exposure to chronic hepatitis B virus infection and aflatoxin may increase liver cancer risk by up to 30 times compared to the risk in individuals exposed to aflatoxin only [
53]. The risk of exposure to contaminated foods with varying levels of AFs worldwide exists for more than 4.5 billion people [
54]. At present, levels of AFs in food and feed are established in approximately 100 countries [
55]. The EU legal limit for AFB1 in processed cereal foods is 0.02 µg/kg [
56]. Different maximum upper limits are set worldwide for AFM1 in milk or milk products, with Codex Alimentarius and the EU setting the limit to 0.05 µg/kg AFM1, whereas the US and some Latin American countries set it to 0.5 µg/kg [
57].
Aflatoxins are the first mycotoxins to be initially classified as toxic, following research into the deaths of 100,000 poultry (mainly turkeys) in England. Originally, the causes of the strange disease at that time were unknown; thus, the disease was named “X disease “, that is, turkey X disease. It was later found that the cause was the growth of
A. flavus in poultry feed. This led to a breakthrough in the research on the field of mycotoxins, which in turn led to the intensive and systematic checks on any moldy product and setting maximum limits [
9].
Pre-harvest and post-harvest factors are related to the production of AFs. Thus, pre-harvest weather conditions associated with periods of drought and heat stress during flowering and fruit growth were reported to be the main factors responsible for the increased infection with AFs produced by
A. flavus and
A. parasiticus in maize, pistachio, cotton, and nuts [
58]. Furthermore, other stress factors in plants, such as inadequate nutrition, insect nutrition from growing fruits, weed competition, overgrowth of plants, and plant diseases, facilitate fungal infection and the production of AFs [
58]. After harvesting, higher concentrations of AFs are observed due to improper storage of the products, such as storage with inadequate moisture content and inappropriate temperature. If the product is quickly dried and stored under appropriate conditions, and the a
w value does not exceed 0.78, aflatogenic molds do not grow well. The biosynthesis of AFs is inhibited at an a
w of less than 0.8334. Because the production of AFs depends on the a
w interaction with temperature, maintaining the temperature in the storage area below 15 °C leads to minimum a
w for the production of mycotoxins at 0.934. Moreover, the formation of AFs can cause damage to the products [
59].
Aflatoxins are linked to various diseases, such as aflatoxicosis, in animals, pets, and humans around the world [
44], and they are considered to be particularly harmful as they have carcinogenic, mutagenic (DNA damaging), teratogenic, and immunosuppressive effects [
51]. Symptoms of acute aflatoxicosis in humans include vomiting, abdominal pain, jaundice, pulmonary edema, coma, convulsions, and death [
5,
60], while chronic aflatoxicosis occurs via cancer, immune system inhibition, and liver damage. There are significant differences in species sensitivity, with the size of the reaction depending on a variety of factors, such as age, sex, weight, nutrition, metabolism, exposure to infectious agents, and the occurrence of other mycotoxins [
5], as well as the type of toxin, mechanism of action, and levels of intake [
61]. In many areas of the world, where liver cancer occurs in large numbers in the population (e.g., in southeast Asia and sub-Saharan Africa), chronic hepatitis C infection and aflatoxin exposure are considered important risk factors, since they are likely to interact synergistically [
62].
In India, the most serious outbreak of human hepatitis was recorded in 1974, when 108 of 397 patients died after consuming heavily contaminated maize with AFs at levels of 0.25–15 mg/kg [
63], while the largest and most serious case of acute aflatoxin poisoning in humans worldwide, recorded in April 2004 in Kenya, resulted in 125 out of 317 patients losing their lives (mortality rate, 39.4%) after eating infected maize, with aflatoxin levels of 5–20 mg/kg [
64]. A smaller-scale epidemic occurred in Kenya in 2005, causing 16 deaths [
65], while, in the same country in 1981, 12 deaths were recorded from consumption of contaminated maize at levels of 3.2–12 mg/kg with AFB1 [
64]. In addition, encephalopathy and visceral degeneration in children are symptoms of Reye’s syndrome, which is linked to aflatoxin toxicity [
19].
Aflatoxin contamination was reported in various countries such as Argentina [
66], Brazil [
67], China [
68], Italy [
69], Portugal [
70], Spain [
71], and Tanzania [
72]. In food analysis, the presence of AFB1 is often the highest in the AF mixture [
19]. AFs are mainly detected in cereals (barley, corn, rice, wheat, oat) [
33] and their derivatives (bread, flour, breakfast products, cornflakes and pasta) [
4], in nuts (almonds, pecans, pistachios, walnuts, cashews, and Brazil nuts) and peanuts [
73], in species and herbs [
6,
74], in edible vegetable oils [
55], in wines [
75], in sugarcane [
76], in cottonseed [
40], in dried fruits [
77], and in animal food products such as milk [
78], eggs [
79], cured meat [
80], and animal tissues [
81]. Some detailed recent studies on AF occurrence in foods are presented in
Table 4. The problem of AFs is very important around the world and particularly in Africa, where aflatoxin contamination is reported in raw cereals with 50% incidence, with infestation reaching 1642 μg/kg in rice [
33].
2.2. Ochratoxin A
Among the ochratoxin categories A, B, and C, OTA is the most abundant and harmful mycotoxin that contaminates foods [
93]. OTA was first identified in South Africa, from the fungus
A. ochraceus, from which it derives its name. It is chemically known as the phenylalanyl derivative of a substituted iso-coumarin (
R)-
N-[5-chloro-3,4-dihydro-8-hydroxy-3-methyl-1-oxo-1
H-2-benzopyran-7-y1] carbonyl]-
l phenylalanine [
94].
Aspergillus and
Penicillium are the two main genera of OTA producers. The main producing species belong to the
Aspergillus section
Circumdati,
Aspergillus section
Nigri,
P. verrucosum,
P. thymicola, and
P. nordicum [
19]. The non-chlorinated analogue, ochratoxin B, which is much less toxic, sometimes co-occurs with OTA in food and feed [
53]. Although OTA produced by
Aspergillus can likely occur pre-harvest, recent studies [
95] pointed to OTA in grains as mainly a storage issue.
Ochratoxin is linked to immunotoxic, genotoxic, neurotoxic, carcinogenic, nephrotoxic, and teratogenic effects, considered the most toxic ones among the ochratoxin family members. Moreover, it is classified by the IARC as a possible human carcinogen (Group 2B) [
20], but the specific mechanism of toxicity is not fully understood. Increased incidence of testicular cancer in animals is associated with ingestion of OTA [
19]. Although OTA is liable to decomposition in the rumen, it was found in cow’s milk [
96]. The mutagenic capacity of AFB1 could be increased in cases of co-occurrence with OTA in some crops [
97]. A provisional tolerable weekly intake (PTWI) of 112 ng/kg body weight (b.w.) was proposed by the Joint FAO/WHO Expert Committee on Food Additives (JEFCA) [
98].
Ochratoxin production is observed in the a
w range of 0.92–0.99, with the maximum concentration being in the range 0.95–0.99 depending on the strains. The optimum temperature for OTA production is 20 °C, followed by the temperature of 15 °C, with significantly lower production at 30–37 °C [
99]. Taking into account that
Aspergillus and
Penicillium responsible for the production of OTAs have a temperature range of 12–37 °C for
A. ochraceus and 0–31 °C for
P. verrucosum, OTA can be produced in all agricultural areas of the world [
33].
Ochratoxin was reported in cereals [
28], in species [
6], in alcoholic beverages such as in wines [
75] and in beer [
100], in dried vine fruits [
40], in coffee [
101], in cocoa and chocolate [
102,
103], in meat [
104], and in milk [
96]. Among foods, cereals occupy the first position of the total exposure to OTAs with 60% [
105]. The maximum limits of 5 ng/g OTA in raw cereal grains, 3.0 ng/g in cereal-processed products, 10 ng/g in coffee and dried fruits, 2 µg/L in wine, and 0.5 ng/g in cereal-based baby foods are set by European commission (EU) [
106].
Pig’s blood, kidney, liver muscle, and adipose tissue are some of the tissues where OTA was detected with rather high levels found in animals suffering from porcine nephropathy, especially in countries of the Balkan Peninsula [
53]. In a human disease of kidney referred to as Balkan endemic nephropathy, OTA is implicated. The disease is characterized by tubule interstitial nephritis, and OTA is associated with a high incidence of kidney, pelvis, ureter, and urinary bladder tumors in some eastern European countries [
51]. It is also suspected that the inhalation of OTs via air and dust caused by the opening of Egyptian tombs may have led to the deaths of archaeologists [
6].
Recent OTA studies in food commodities are presented in
Table 5.
Risk assessments were carried out based on OTA occurrence data in Brazil [
118], Benin, Cameroon, Mali, Nigeria [
119], and Paraguay [
103]. According to these studies, the majority of the population did not exceed the TDI.
The impact of OTA in recent research shows the importance of reinforcing OTA control strategies in food production. Although mycotoxins are extremely stable during food processing, there are several factors that can affect their stability [
6]. At different stages of food processing such as baking, roasting, frying, brewing, canning, and peeling, OTA cannot be completely deactivated [
120]. Moreover, it was reported that OTA can be transferred into beer and wine samples from contaminated grains [
121]. Feeding animals with OTA-infected bread can cause its accumulation in the meat of animals intended for human consumption [
122].
2.3. Fumonisins
Fumonisins belong to a large group of toxins referred to as
Fusarium toxins, which occur in cereals originating from pathogenic fungi, mostly
Fusarium verticillioides and
Fusarium proliferatum [
123]. In addition,
Aspergillus niger can produce fumonisins on grapes and raisins [
124]. The group of 28 analogues of FBs is divided into four main groups: fumonisin A, B, C, and P. In addition to A, B, C, and P, there is also Fumonisins “Py” that can occur [
123]. The fumonisin B (FB) analogues, which include FB1, FB2, and FB3, occur in nature with the highest frequency, whereas FB1 is usually found at the highest concentrations [
125]. Fumonisins cause health effects in animals, especially in the liver and kidney, although data for the health effects of fumonisins in humans remain limited [
126]. FB1 can cause leukoencephalomalacia in horses [
127], and pulmonary edema syndrome and hydrothorax in pigs [
128].
The IARC classifies FB1 and FB2 as possibly carcinogenic to humans (Group 2B) [
20], and the JECFA set a provisionally maximum tolerable daily intake (PMTDI) of 2 µg/kg b.w./day for FB1, FB2, and FB3 alone or in combination [
129]. Acceptable upper limits of 800–4000 and 2000–4000 µg/kg FB1 and FB2, respectively, were set by the European Union [
38] (EU Regulation 1126/2007) and the US, in cereal-based products [
130].
Fumonisins contaminate cereals [
131] and their derivatives [
132,
133], and they also exist in maize and maize-based products [
134], asparagus [
135], grapes, and raisins [
124].
Table 6 presents representative studies on the occurrence of FUs (μg/kg) in food samples worldwide during 2014–2019.
2.5. Zearalenone
The fungi of the
Fusarium genus produce ZEN; in particular, fungal species of
F.
graminearum (
Gibberella zeae),
F.
culmorum,
F.
crookwellense, F.
semitectum, and
F.
equiseti are the major producers of ZEN, infecting cereals and food worldwide, mainly in temperate climates [
1,
183]. While contamination with ZEN is low in grains in the field, it increases in storage conditions with moisture of more than 30%–40% [
1]. Currently, the limits for ZEN in cereals vary between countries and range from 50 to 1000 µg/kg [
184]. A TDI for ZEN of 0.25 μg/kg b.w./day was established by the EFSA [
40]. Moreover, maximum permissible limits for ZEN should be within the range of 100–200 μg/kg in unprocessed cereals, 75 μg/kg for processed cereals, 20 μg/kg in processed cereal foods, and 50 μg/kg in cereal snacks according to EU legislations [
185]. Risk assessments were performed on the basis of ZEN exposure data in France, Germany, Finland, China, and India.
In only a few cases, the possible ZEN intake was found to exceed the TDI, and almost all studies agreed that the majority of the population did not exceed the TDI value given by the EU [
183]. ZEN is soluble in aqueous alkali, acetone, acetonitrile, benzene, methyl chloride, alcohols, and ethers, but insoluble in water [
98]. The IARC classifies ZEA as a Group 3 carcinogen [
49]. ZEN is a non-steroidal estrogenic mycotoxin and works by mimicking the effects of the female estrogen hormone, affecting conception, ovulation, and fetal development at concentrations above 1 mg/kg [
166]. ZEN can lead to hyperestrogenism, mainly affecting reproduction. The most susceptible species to ZEN infection are prepubertal swine. Swelling of the vulva, increases in uterine size and secretions, mammary gland hyperplasia and secretion, prolonged estrus, anestrus, increased incidence of pseudopregnancy, infertility, decreased libido, and secondary complications of rectal and vaginal prolapses, stillbirths, and small litters are some of the typical clinical symptoms of hyperestrogenism [
1].
Occurrence of ZEN is reported both in various developed countries like Germany [
186], and Japan [
187] and in developing countries like Egypt [
188], Thailand [
189], Iran [
190] Croatia [
154], and the Philippines [
161]. It is worth mentioning that zearalenone is a synthetic nonsteroidal estrogen of the resorcylic acid lactone group related to mycoestrogens found in fungi in the
Fusarium genus, and it is used mainly as an anabolic agent in veterinary medicine, where it can also contribute to related exposure/toxicity [
191].
Table 8 presents representative studies on the occurrence of ZEN (μg/kg) in food samples worldwide during 2014–2019.
2.6. Emerging Fusarium Mycotoxins (Fusaproliferin, Moniliformin, Beauvericin, NX-2 Toxin, and Enniatins)
In recent years, emerging mycotoxins became a major issue due to their high occurrence in cereals and their products [
120]. In a more recent paper, emerging mycotoxins were defined as “mycotoxins, which are neither routinely determined, nor legislatively regulated; however, the evidence of their incidence is rapidly increasing” [
195]. Currently, an opinion on the presence of ENNs and BEA in food and feed was reported by the EFSA without a risk assessment due to the lack of relevant toxicity data. Moreover, until now, there are no maximum levels for emerging
Fusarium mycotoxins [
196].
Fusaproliferin is a bicyclic sesterterpene produced by
Fusarium species such as
F. proliferatum F. subglutinans, and
F. verticillioides [
197]. FUs showed toxicity on chicken embryos and brine shrimp larvae [
197].
Structurally, MON is a 1-hydroxycyclobut-1-ene-3,4 dion, a small molecule, soluble in water, which can be produced by
F. verticillioides,
F. begoniae,
F. denticulatum,
F. lactis,
F. nisikadoi,
F. phyllophilum,
F. pseudocircinatum,
F. pseudonygamai,
F. ramigenum,
F. tricinctum,
F. acutatum,
F. anthophilum,
F. bulbicola,
F. concentricum,
F. diaminii,
F. fujikuroi,
F. napiforme,
F. nygamai,
F. proliferatum,
F. pseudoanthophilum,
F. sacchari,
F. subglutinans,
F. thapsinum,
F. beomiforme,
F. oxysporum,
F. redolens,
F. chlamydosporum,
F. arthrosporiodes,
F. avenaceum, and
F. acuminatum [
198], and recently proven to be a metabolite of
Penicillium melanoconidium [
197].
Structurally, BEA is cyclic hexadepsipeptide consisting of an alternating sequence of three
d-a-hydroxy-
iso-valeryl- and
N-methyl-
l-phenylalanyl residues [
199]. BEA was first isolated from
Beauveria bassiana, a fungus that causes diseases in insects [
199], but it is frequently found in corn and corn-based foods and feeds infected by
Fusarium spp. BEA occurs in cereal and cereal-based products not only in different European countries such as Romania [
200], Spain [
201], Italy [
112], and Czech Republic [
92], but also throughout the world in countries such as Japan [
202], Tanzania, Rwanda [
203], Iran [
204], and Morocco [
205]. BEA has antibacterial, antifungal, and insecticidal activities and causes toxic effects such as induction of apoptosis, increased concentration of cytoplasmic calcium, and DNA fragmentation in mammalian cell lines [
199].
A new trichothecene mycotoxin, named NX-2, was recently characterized in rice cultures. NX-2 is similar in structure and similar in toxicity to 3-ADON, but lacks the keto group at C-8; hence, it is a type A trichothecene [
206].
The
Fusarium species identified as producers of ENNs are
F. merismoides,
F. acuminatum,
F. arthrosporioides,
F. avenaceum,
F. compactum,
F. culmorum,
F. equiseti,
F. kyushuense,
F. langsethiae,
F. lateritium,
F. oxysporum,
F. poae,
F. sambucinum,
F. scirpi,
F. sporotrichioides,
F. torulosum,
F. tricinctum, and
F. venenatum [
207]. Fusarium species capable of producing ENNs can be found in different geographical areas, and the extent of seed contamination is only occasionally as high as mg/kg [
208]. ENNs contaminate not only cereal grains but also many kinds of foods including vegetable oil, beans, dried fruits, tree nuts, and coffee [
196]. The ENNs most detected in foods and feed are enniatin A, (ENA), enniatin A1 (ENA1), enniatin B (ENB), and enniatin B1 (ENB1) [
209]. With regard to enniatins, there is relatively little to indicate that it is of concern to humans and animals; however, it may play a role in pronouncing the impact of other
Fusarium toxins (i.e., DON) by inhibiting cellular export [
210].
Due to their high prevalence in feed and food, possibly at high concentrations, as well as their potential toxicity to animals and humans, research interest in emerging mycotoxins increased [
199].
Table 9 presents representative studies on the occurrence of emerging
Fusarium mycotoxins (μg/kg) in food commodities worldwide during 2014–2019.
2.7. Ergot Alkaloids
The EA group of mycotoxins is derived from the genus
Claviceps, which is a phytopathogen, with effects known from the Middle Age (ergotism, the human disease historically known as St. Anthony’s Fire) [
42], and it is classified as a tryptophan-derived alkaloid [
213]. Fungal structures of
Claviceps species are produced instead of kernels on grain ears or seeds on grass heads, with large and dark sclerotia representing the final stage of the disease, known as “ergots” [
213]. Psychological and physiological effects in humans can occur by ergot poisoning, affecting blood supply to the extremities and central nervous system, while, in animal health, there are problems associated with reduced productivity, diarrhea, and internal bleeding [
44,
214].
In Europe, a series of EAs, such as ergocryptine, ergocristine, ergotamine, ergosine, ergometrine, ergocornine, and their corresponding epimers can be detected in the sclerotia after the contamination of
Claviceps purpurea, which is the major producer of EAs [
68].
C. purpurea is widespread throughout the world and infects many monocotyledonous plants, like cereal grains and forage grasses [
42,
215]. Main affected crops by EAs are cereals like rye, barley, wheat, millet, oats, and triticale [
216]. Among cereals, rye has the highest rates of fungal contamination by
C. purpurea as it is a cross-pollinator with large open florets [
217]. As a result, ergot alkaloids appear more in rye and rye-based foods as compared to other cereals [
217]. Even if ergot bodies are removed by a hand-cleaning procedure, EAs could remain in grains [
214].
Ergocristine, ergosine, ergotamine, ergometrine, ergocornine, and ergocryptine are the EAs with the highest frequency of detection [
214]. In the EU, no maximum permitted levels of EAs are set for feed or food, and the only available standard (EU) sets a limit of 0.5 g/kg for the sum of ergot alkaloids in unprocessed cereals, except for maize and rice [
218]. Based on toxicological studies on their vasoconstrictive effects and following an estimate of human and animal dietary exposure by the EFSA, a group TDI of 0.6 µg/kg b.w./day was derived [
213]. The maximum permissible level of 300 mg of ergot bodies/kg grain was set by the United States and Canada, while 0.01% is the limit in China of the total ergot alkaloid content in cereals [
217].
Table 10 presents representative studies on the occurrence of EAs (μg/kg) in food samples worldwide during 2014–2019.
2.8. Alternaria Toxins (Altenuene, Alternariol, Alternariol Methyl Ether, Altertoxin, Tenuazonic Acid, Tentoxin)
The
Alternaria species can be found everywhere and in many ecosystems such as plants, seeds, agricultural commodities, atmosphere, and soil [
220]. They produce
Alternaria toxins that contaminate foods in storage [
221], with AOH, AME, TeA, ALT, ATXs, and TeA being the most important. More than 70 secondary metabolites are produced by the toxin-producing
Alternaria [
222], including species such as
A. alternata,
A. brassicae,
A. dauci,
A.
japonica,
A. solani,
A. tenuissima, and
A. triticina [
220]. Moreover, over 30 mycotoxins were isolated belonging to different classes based on chemical structure [
221]. The genus
Alternaria includes saprophytic, endophytic, and pathogenic species, and it is a cosmopolitan fungal genus found in natural and anthropogenic environments [
220]. Among the
Alternaria species,
A. alternata is the most common in harvested fruit and vegetables, and it is the most important species producing mycotoxins [
222]. AOH, AME, and ALT belong to dibenzo-α-pyrone derivatives, while ATXs belong to perylene quinone derivatives [
223]. TeA belongs to tetramic acid derivatives with antibacterial and phytotoxic activities and acute toxicity for mice, chicken, and dogs, as well as hematological disorders in human [
224].
According to the EFSA, among ATs, the most frequently studied are AOH, AME, and TeA [
222]. Although most ATs exhibit only low acute toxicity, AOH and AME are the most toxic due to mutagenic, carcinogenic, cytotoxic, and genotoxic effects, with evidence from in vitro toxicological studies using bacterial and mammalian cells [
225]. AOH is more genotoxic than AME in human carcinoma colon cells [
224]. Currently, there are no regulatory limits or monitoring guidelines established for ATs in food worldwide. The risk assessment for
Alternaria toxins entitled “Dietary exposure assessment to
Alternaria toxins in the European population” for four of the known ATs, namely, AOH, AME, TeA, and TEN, with the highest exposure to AOH, AME, and TeA in “toddlers” and “other children”, was recently published by the EFSA. The TTC approach (toxicological concern threshold) was implemented by the EFSA as there is little or no toxicity data on ATs in order to assess the levels of concern for human health. For genotoxic ATs (AOH and AME), a TTC value of 2.5 ng/kg b.w./day was set, whereas, for non-genotoxic ATs (TeA and TEN), a TTC value of 1500 ng/kg b.w./day was set, and these exposure estimates are unlikely to be a concern for human health [
44]. A major need for the assessment of exposure of humans and animals to potential health risks is the acquisition of additional toxicological data on the contamination of food and feed with ATs [
222].
Substrate composition, temperature, pH, and a
w are the most important biotic and abiotic parameters affecting the biosynthesis of mycotoxins and, thus, the biosynthesis of ATs. In particular, a
w and pH affect most the biosynthesis of
A. alternata [
224].
Table 11 shows representative studies on the occurrence of ATs (μg/kg) in food samples worldwide during 2014–2019.
The studies were conducted in dried fruits, in wheat and wheat-based products, in fresh and dried tomatoes, in juice samples, and in red wine. ATs that are the focus of most studies are AOH, AME, TEN, TeA, and ALT [
223]. The occurrence of ATs is reported in various countries like Germany [
223], Argentina [
224], Canada [
221,
229], China [
108], the Netherlands [
230], and Italy [
227]. ATs are detected in a large range of foodstuff commodities such as dried fruit [
108], wheat, bran, flour [
224,
228,
229,
230], fresh and dried tomatoes [
228], peppers [
231], wine, vegetable juices, fruit juices [
223], beer [
232], cereal-based products such as rice and oat flake [
233], sunflower seeds, and sunflower oil [
222].
2.9. Patulin
Structurally, PAT is a heterocyclic lactone (4-hidroxi-4
H-furo [3,2-c]piran-2(6
H)-ona). It has a molecular weight of 154.12 g/mol and low volatility [
233,
234]. In total, 60 different fungal species such as
Penicillium expansum (
P. leucopus),
P. crustosum,
P. patulum (
P. urticae and
P. griseofulvum), and
A. clavatus produce PAT, whereas
P. expansum is the most common PAT producer [
233]. The strain significantly determines the amount of patulin produced. Neurotoxicity, immunotoxicity, carcinogenesis, teratogenicity, and mutagenicity are acute and chronic effects exerted by patulin on cell cultures [
235]. Patulin causes immunotoxic and neurotoxic effects in animals, and there is no clear evidence that it is carcinogenic to humans [
224].
The EU, US Food and Drug Administration, and Chinese legislation all set the upper limit of 50 µg/L/kg patulin in apple and fruit juices [
234]. For fruit juices, concentrated fruit juices such as reconstituted and fruit nectars, and spirit drinks, cider, and other fermented drinks derived from apples or containing apple juice, the EU established maximum levels of 50 µg/kg. For solid apple products, including apple compote and apple puree intended for direct consumption, the EU established maximum levels of 25 µg/kg. For apple juice and solid apple products, including apple compote and apple puree, for infants and young children and labeled and sold as such, the EU established maximum levels of 10 µg/kg [
235]. The JECFA implemented a PMTDI of 0.4 mg/kg b.w./day for PAT [
224] in 1995.
PAT is found in fruits and vegetables, especially apples and apple products in various parts of the world, and occasionally in other fruits such as pears, oranges, grapes, and their products [
236,
237,
238,
239,
240]. If rotten fruits, especially apples, are not removed during fruit juice processing, patulin is transferred to juices [
224]. PAT was initially studied as a potential antibiotic, but subsequent research showed human toxicity, including nausea, vomiting, ulceration, and hemorrhage [
3]. The EU, US, and China present major problems of PAT contamination as they are the main producers of apples and apple products [
234].
Table 12 presents representative studies on the occurrence of PAT (μg/kg or μg/L) in food samples worldwide during 2014–2019. The studies were conducted in dried fruits (figs, raisins), juices (apple and mixed fruit), and jam.