Fusarium mycotoxins: The major food contaminants

Abstract Mycotoxins, which are secondary metabolites produced by toxicogenic fungi, are natural food toxins that cause acute and chronic adverse reactions in humans and animals. The genus Fusarium is one of three major genera of mycotoxin‐producing fungi. Trichothecenes, fumonisins, and zearalenone are the major Fusarium mycotoxins that occur worldwide. Fusarium mycotoxins have the potential to infiltrate the human food chain via contamination during crop production and food processing, eventually threatening human health. The occurrence and development of Fusarium mycotoxin contamination will change with climate change, especially with variations in temperature, precipitation, and carbon dioxide concentration. To address these challenges, researchers have built a series of effective models to forecast the occurrence of Fusarium mycotoxins and provide guidance for crop production. Fusarium mycotoxins frequently exist in food products at extremely low levels, thus necessitating the development of highly sensitive and reliable detection techniques. Numerous successful detection methods have been developed to meet the requirements of various situations, and an increasing number of methods are moving toward high‐throughput features. Although Fusarium mycotoxins cannot be completely eliminated, numerous agronomic, chemical, physical, and biological methods can lower Fusarium mycotoxin contamination to safe levels during the preharvest and postharvest stages. These theoretical innovations and technological advances have the potential to facilitate the development of comprehensive strategies for effectively managing Fusarium mycotoxin contamination in the future.


INTRODUCTION
Fungi are a kind of eukaryote with a vegetative structure that can be filamentous or unicellular.Their cell walls are composed of chitin, chitosan, or polysaccharides, and they reproduce by spores that are produced asexually or sexually.Fungi are ubiquitous in the biosphere and the second most species-rich eukaryotic organisms after insects 1 .They play a pivotal role in the intricate processes of the nutrient cycle and ecosystem balance and are consumed as food and used in the production of industrial materials 2 .Similar to viruses and bacteria, fungi also have negative effects on humans.More than 500 fungal species are capable of infecting the human body, which causes at least 1.5 million deaths globally each year 3 .In plantation crop production, plant pathogenic fungi can cause economic losses of upward of hundreds of billions of dollars each year.Plant pathogenic fungi adversely affect crop growth, yield, and quality, and mycotoxin contamination is one of the most important causes of quality degradation. 4ycotoxins are natural toxic compounds with low molecular weight (often less than 1000 Da) that are synthesized by fungi and have the characteristics of nephrotoxicity, hepatotoxicity, carcinogenicity, teratogenicity, immune toxicity, neurotoxicity, genotoxicity, mutagenicity, cytotoxicity, reproductive toxicity, alimentary canal toxicity, dermal toxicity, and so on 5,6 .Mycotoxin exposure, whether in humans or in animals, even at low concentrations, can lead to acute or chronic diseases and, in certain instances, death.Cereals, nuts, fruits, spices, legumes, and their products or byproducts are most likely to contain mycotoxins.Based on a previous estimation by the Food and Agriculture Organization (FAO) of the United Nations, approximately 25% of cereals produced worldwide are contaminated by mycotoxins 7 .However, a recent study conducted by Eskola et al. revealed that the contamination rate could be 60%-80% 8 .Mycotoxins enter the food chain via fungal infections of crops, which can occur at any phase of crop production, including planting, harvesting, and storage.Mycotoxins can be introduced into the human body directly via the ingestion of contaminated foods or derived products, or indirectly through the consumption of eggs, edible offal, meat, milk, and related products from livestock that consume contaminated feed 8 .
Mycotoxins are not essential to the growth or development of fungi, but they seem to be a way for fungi to reduce the number of superfluous precursors 9 .It has also been suggested that mycotoxins contribute to the defensive tactics of mycotoxigenic fungi against other microorganisms.Moreover, mycotoxins exert a substantial influence on the pathogenicity, aggressiveness, and virulence of mycotoxigenic fungi 10 .More than 400 varieties of mycotoxins have been identified, but aflatoxin (AF), ochratoxins, fumonisins (FUMs), trichothecenes (TRIs), patulin, citrinin (CIT), and zearalenone (ZEA) are those most inextricably linked to agriculture, economics, and public health 5,6 .There are three dominant toxigenic fungal genera: Aspergillus, Fusarium, and Penicillium 6 .Fusarium species usually infect crops and produce mycotoxins before or immediately after harvest, while the Aspergillus and Penicillium species are more commonly associated with foods during drying and storage 5 .
Fusarium, a prominent genus of plant pathogenic and mycotoxin-producing fungi worldwide, targets various plant parts, including grains, seedlings, heads, roots, and stems (Figure 1), causing yield loss and quality reduction of crops 11 .For example, Fusarium head blight (FHB) is a highly destructive disease that affects cereals and results in both significant yield reduction and a negative influence on grain quality 12 .Currently, Fusarium comprises more than 300 phylogenetically distinct species and has been classified into 23 informal species complexes 13 .Different Fusarium species have various abilities to produce mycotoxins.The Fusarium sambucinum species complex includes many devastating plant pathogens, most of which are important mycotoxin producers (Figure 2); the three predominant classes of mycotoxins synthesized by Fusarium species are FUMs, TRIs, and ZEA.Moreover, a type of TRI analog, deoxynivalenol (DON), is consistently regarded as a significant concern in the realm of food safety due to its prevalent occurrence as a grain contaminant.Due to a lack of adequate understanding, other toxic Fusarium metabolites, which are commonly referred to as emerging mycotoxins and include beauvericin (BEA), enniatins, fusaproliferin, fusaric acid, fusarins, and moniliformin (MON), neither undergo routine determination nor are subject to legislative regulation 11 .Furthermore, Fusarium mycotoxins can be converted into modified forms, known as modified mycotoxins, by plants, microorganisms, and chemical or physical approaches during processing, such as acid, alkali, heat, pressure, and irradiation.The modified mycotoxins can coexist with their parent forms 14 .Among these, the modified mycotoxins generated as a result of plant defense mechanisms, primarily via glucosylation catalyzed by uridine diphosphate-glucosyltransferases, are known as masked mycotoxins 15 .The glucose conjugates of mycotoxins, the most commonly identified masked mycotoxins in current research, include DON-3-glucoside (DON-3G), T-2-toxin-3-glucoside, HT-2-toxin-3-glucoside, nivalenol-3-glucoside (NIV-3G), ZEA-14G, α-zearalenol-14glucoside (α-ZEL-14-G), and β-ZEL-14-G.Although the limited available information suggests that masked mycotoxins show lower toxicity than their parent forms, the free forms of masked mycotoxins can still cause unpredicted toxicity via hydrolysis by mammalian gut microorganisms 15 .
In this review, we outline the main types of Fusarium mycotoxins and elucidate their impact on food contamination as well as associated hazards.We then review the interactions between climate change (CC) and Fusarium mycotoxin risks.Finally, we focus on the detection technologies and management strategies for Fusarium mycotoxin contamination.This review provides a reliable reference source for Fusarium mycotoxin control.
In Fusarium, TRI biosynthetic enzymes are encoded by a total of 15 TRI genes, which are distributed across three different loci on different chromosomes: the single-gene TRI101 locus, the two-gene TRI1-TRI16 locus, and the 12-gene core TRI cluster (Figure 4A) 21,22 .In F. graminearum, the TRI16 homolog is rendered nonfunctional as a result of multiple insertions and deletions within its coding region.Table 1 shows the phenotypes of mutants with TRI gene disruption or deletion in F. sporotrichioides.TRI5 catalyzes the cyclization of farnesyl pyrophosphate (FPP) into (A) trichodiene (TDN), which represents the initial enzymatic step in TRI biosynthesis (Figure 4B).TDN is subsequently transformed into calonectrin (CAL) by TRI4, TRI101, TRI11, and TRI3.The catalytic reactions from FPP to CAL are conserved across Fusarium species that synthesize Type A and Type B TRIs.The allelic variants of TRI1 are responsible for the significant structural disparities between Type A and Type B TRIs.In F. sporotrichioides, TRI1 exclusively catalyzes   The production profiles of Fusarium mycotoxins were determined by liquid chromatography-mass spectrometry analysis of extracts and filtrates of Fusarium mycotoxin biosynthesis gene mutants.FUMs, fumonisins; PKS, polyketide synthase; TAS, 3,4,15-triacetoxyscirpenol; TRIs, trichothecenes; ZEA, zearalenone.Data were extracted from Alexander and colleagues 21,23,24 .
hydroxylation at C-8, resulting in Type A TRIs, while F. graminearum TRI1 mediates hydroxylation at both C-7 and C-8, resulting in Type B TRIs 17,21,22 .DON (also called vomitoxin) is one of the mycotoxins linked to FHB, and it is synthesized in F. graminearum, F. culmorum, and so on.The formal chemical name of DON is 3α,7α, 15-trihydroxy-12,13-epoxymonospora-9-ene-8-one 25 .It mainly contaminates cereal crops, such as maize, wheat, rice, and barley, as well as some cash crops.All over the world, DON contamination causes economic losses of billions of dollars each year.The Panel on Contaminants in the Food Chain, which is part of the European Food Safety Authority, examined 26,613 cereal samples from 21 European countries and found that the DON contamination rate was close to 50% 26 .Yan et al. collected 579 wheat samples and 606 maize samples from the major wheat-and maize-producing provinces in China and determined the co-occurrence of type-B TRIs; all of the wheat samples showed positive results for DON, whereas 99.83% of the maize samples were DON-positive 27 .In humans and animals, DON typically causes diarrhea, vomiting, and gastrointestinal inflammation.The chronic consumption caused by DON can result in immunesuppressive diseases, growth impairment, and abnormalities of the reproduction and nervous systems 28 .The FAO and World Health Organization (WHO) first identified DON as a highly hazardous food contaminant in the 1970s 25 .
By binding to the 60S ribosomal subunit, DON can disrupt the action of peptidyl transferase and inhibit protein synthesis 18,28 .Moreover, DON can modulate mitogen-activated protein kinase (MAPK) activity, which includes extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and p38 mitogen-activated protein kinase (p38), via the "ribotoxic stress response" by rapidly activating doublestranded RNA-associated protein kinase (PKR) and hematopoietic cell kinase (Hck 29 ; Figure 5A).DON-induced MAPK activation can mediate the upregulation of transcription factors (nuclear factor kappa-light-chain-enhancer of activated B cells [NF-κB], activating protein 1 [AP-1], and CCAAT/ enhancer-binding protein [C/EBP]) to promote the expression of proinflammatory cytokines, such as interleukin-6 (IL-6), IL-1β, and tumor necrosis factor-α (TNF-α), which cause inflammation 28,30 .DON causes oxidative stress by inhibiting the expression of glutathione (GSH), superoxide dismutase (SOD), catalase (CAT), and GSH peroxidase (GSH-PX), which engage in the repair of oxidative stress-induced damage, decreasing total antioxidant capacity and glutathione S-transferase levels 31 .However, DON can also induce nuclear factor-erythroid 2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) activation to remove excessive reactive oxygen species (ROS).These results show that DON not only elicits oxidative stress but also impedes its occurrence.The potential impact of DON may vary depending on the dosage, although the precise mechanism remains incompletely elucidated 32 .
The relationship between oxidative stress and inflammation is interrelated and interdependent.Specifically, the products of oxidative stress (primarily ROS) have been shown to increase proinflammatory responses.Additionally, inflammatory cells release significant amounts of ROS at sites of inflammation, exacerbating oxidative damage 33 .In most cell types, mitochondria are not only the largest contributors to intracellular ROS production but also the targets of cellular ROS.A feed-forward detrimental cycle exists between the generation of ROS and mitochondrial damage, wherein ROS-damaged mitochondria undergo dysfunctionality, subsequently leading to an exacerbation in intracellular ROS production 34 .In the case of oxidative stress, damaged mitochondria show a decrease in mitochondrial membrane potential, which is closely related to apoptosis 28,32 .The B-cell lymphoma 2 (Bcl-2) gene family regulates apoptosis through the mitochondrial pathway.Among this gene family, the Bcl-2-associated X (Bax) gene and Bcl-2 antagonist/killer-1 (Bak-1) gene are important proapoptotic genes, while Bcl-2 is an antiapoptotic gene.The augmented Bax/Bcl-2 ratio serves as a pivotal indicator of the initiation of apoptosis.Caspases (cysteine-aspartic proteases, cysteine aspartases, or cysteine-dependent aspartate-directed proteases), a group of protease enzymes, are crucial players in the process of apoptosis, with Caspase 3 being known as the executor of apoptosis 35 .DON-induced oxidative stress results in oxidation and damage of DNA and lipids through the elevation of ROS levels, finally leading to apoptosis through the mitochondrial pathway via activation of the JNK pathway, which increases the Bax/Bcl-2 ratio and induces the conversion of Caspase 9 into Caspase 3. In addition, this process triggers proinflammatory cytokine expression by upregulating NF-κB 32,36,37 .Moreover, DON can also induce apoptosis by regulating the Janus kinase 2/signal transducers and activators of transcription (JAK2/STAT) pathway and inhibiting the phosphatidylinositol 3-kinase/ threonine kinase (PI3K/AKT) pathway, the latter of which is important in inhibiting apoptosis because it regulates downstream effector molecules (e.g., suppressing Bax translocation) 36,[38][39][40] .In addition, DON can disturb normal cell cycle progression by upregulating ERK and JAK2/STAT to promote cyclin-dependent kinase inhibitor p21 expression 40 .Perturbations in endoplasmic reticulum (EnR) function resulting from enhanced protein synthesis or the accumulation of misfolded proteins give rise to a condition known as EnR stress 41 .The generation of ROS can elicit an elevation in the EnR stress level, while EnR stress can induce the production of ROS in the EnR and mitochondria 42 .EnR stress can induce cellular death, including apoptosis and autophagy, through three classical signaling pathways (protein kinase RNA-like endoplasmic reticulum kinase [PERK], activating transcription factor 6 [ATF6], and the inositol-requiring protein 1 [IRE1]-mediated signaling pathway).In EnR stress-induced apoptosis, the C/EBP homologous protein (CHOP) transcription factor plays a pivotal role in the PERK, ATF6, and IRE1 signaling pathways 43 .To date, studies have demonstrated only that 3ADON has the capability to induce apoptosis in a mouse liver by inducing EnR stress via the IRE1 pathway 44 .No similar study of DON yet exists.The DON-activated death receptor pathway of apoptosis includes the TNF-induced model and the Fas/FasL-mediated model (Fas, the first apoptosis signal, is also known as Apo-1 or CD95; FasL, Fas ligand), both of which are linked to TNF receptor (TNFR) family receptors that are connected to extrinsic signals.Although the exact mechanism is not fully understood, DON has demonstrated an ability to hinder DNA and RNA synthesis 18,28 .Moreover, You et al. 32 raised the presumption that DON and T-2 could achieve the "immune evasion" process to actively evade immune surveillance by immune cells.A recent study showed that T-2 could initiate the "immune evasion" process by activating the signaling pathway involving the programed cell death protein-1/ programmed cell death-ligand 1 45 .For DON, however, there is a dearth of related studies.

FUMs
FUMs are a class of long-chain amino polyalcohols that are primarily synthesized by species in the Fusarium fujikuroi species complex, especially in Fusarium verticillioides and F. fujikuroi (Figure 2).FUMs occur primarily in cereals (rice, wheat, barley, maize, rye, oat, and millet).Over 28 FUM homologs are known, and particular emphasis has been given to the B series, notably FUM B 1 (FB 1 ), FB 2 , and FB 3   11   .FB 1 is composed of a diester comprising propane-1,2,3-tricarboxylic acids (TCA) and 2-amino-12,16-dime thyl-3,5,10,14,15-pentahydroxyleicosane, where hydroxyl groups at the C-14 and C-15 positions interact with the carboxyl groups of TCA to form an ester.Moreover, FB 2 and FB 3 can be regarded as the C-5 and C-10 dehydroxy analogs of FB 1 19 (Figure 3B).FUMs induce a range of deleterious effects on organisms, encompassing carcinogenicity, cytotoxicity, hepatotoxicity, immunotoxicity, nephrotoxicity, neurotoxicity, and reproductive toxicity 46 .In F. verticillioides, the biosynthesis of FUMs necessitates a sophisticated gene cluster spanning 42 kb with 17 coregulated genes (Figure 4A).Except for FUM20, the FUM production profiles of FUM gene deletion or disruption mutants were ascertained using liquid chromatography-mass spectrometry (LC-MS) analysis (Table 1).FUM1, which encodes a polyketide synthase (PKS), is the key gene in FUM biosynthesis.In the first step, the FUM1 protein (Fum1p) facilitates the condensation of 1 acetyl coenzyme A (acetyl-CoA), 8 malonyl-CoAs, and 2 S-adenosyl methionines (SAMs) to generate a linear 18-carbonlong polyketide.In the second step, Fum8p catalyzes the condensation between the linear polyketide and alanine, resulting in a 3-keto intermediate with a 20-carbon chain.In the third step, Fum6p is responsible for catalyzing the hydroxylation process of the polyketide-amino acid condensation product at C-14 and C-15.Then, the carbonyl group at C-3 is reduced into an alcohol group by Fum13p.The addition of a hydroxyl group to the C-10 carbon is catalyzed by Fum2p.The catalysis of esterification, leading to the hydroxylation at the C-14 and C-15 positions of the FUM backbone, is facilitated by Fum14p.In addition, the involvement of Fum7p, Fum10p, and Fum11p is also evident in the biosynthesis of the tricarballylate portion.The final step is the hydroxylation of the FUM backbone at C-5 by Fum3p (Figure 4C).Fum21p, a transcription regulator with a Zn(II)2Cys6 DNA-binding domain, can positively regulate the gene expression of FUM1 and FUM8 and is also required for FUM synthesis 21 .
FB 1 , the most typical FUM, has a structure similar to that of sphingolipids (SLs), and it has the capability to act as an inhibitor of ceramide synthase (CerS).Inhibition of CerS interferes with SL metabolism, causing the accumulation of free sphinganine (Sa) and sphinganine-1-phosphates (Sa-1-P) in cells, alterations in complex SLs, and reduced levels of ceramides 46 .FB 1 and its generated toxic products (Sa, Sa-1-P) induce oxidative stress by exacerbating peroxide production (ROS, H 2 O 2 , lipid peroxide, and lipid oxidation end products) and inhibiting the activity of antioxidants (SOD, CAT, GSH-PX, and GSH 46 ; Figure 5B).FB 1induced oxidative stress can induce JNK phosphorylation, activate P53 signaling, and upregulate the expression of proapoptotic factors (P53-upregulated modulator of apoptosis [PUMA] and Caspase 3) to cause apoptosis 41,47 .FB 1 -induced EnR stress leads to apoptosis through the JNK/p53/PUMA/ Caspase 3 pathway and autophagy through the IRE1/JNK pathway, which releases Beclin-1 (BECN1) from the Bcl-2-BECN-1 interaction to promote the conversion of microtubuleassociated protein 1 light chain 3 (LC3)-I into LC3-II 47 .In addition, FB 1 -induced EnR stress can induce autophagy by the PERK pathway and the AMP-dependent protein kinase (AMPK) pathway 47,48 .FB 1 exposure can also induce apoptosis by regulating the phosphatase and tensin homolog (PTEN)/PI3K/AKT signaling pathway via disruption in lipid raft formation 49 .Moreover, FB 1 exerts an epigenetic influence on the PTEN/PI3K/AKT signaling pathway to enhance DNA damage by inhibiting checkpoint kinase 1 (CHK1) activity through phosphorylation of its Ser280 residue, thereby impeding the repair process for damaged DNA 50 .It has also been demonstrated that the apoptotic pathway of FB 1 is linked to death receptor pathways, DON can efficiently induce the activation of MAPKs through a mechanism referred to as the "ribotoxic stress response" and hijack the JAK2/STAT-3 pathway to induce apoptosis, proinflammatory cytokine upregulation, and cell cycle arrest.DON also activates Caspase 8 to increase expression of Caspase 3 by the tumor necrosis factor-α and Fas/FasL signaling pathways, leading to apoptosis.(B) FB 1 disrupts sphingolipid metabolism, causing DNA damage and apoptosis through the PTEN/PI3K/AKT signaling pathway as well as oxidative stress, which further entrenches DNA damage and apoptosis and induces EnR stress.FB 1 -induced EnR stress not only causes apoptosis by the JNK/p53/PUMA/Caspase 3 pathway but also leads to autophagy via signaling pathways mediated by IRE1, PERK, and AMPK.IRE1 can activate JNK, which results in the subsequent elevation of BECN1, ATG5, and ATG7 and the conversion of LC3-I.PERK induces the expression of ATG5 and ATG7 to form the ATG5-ATG7 complex and promotes the conversion of LC3-I into LC3-II.Under EnR stress, AMPK inhibits the anabolic process of mTOR, thereby increasing the expression of autophagy-related genes and triggering autophagy.(C) ZEA and its derivatives can bind to ERs to elicit estrogen-like effects and promote cellular proliferation.ZEA-induced DNA damage can upregulate ATM and ATR expression and activate DNA damage checkpoints CHK1 and CHK2 to repair DNA damage.Next, the expression levels of CDC25A and CDC25C are upregulated, which promotes the expression of CCNB1 and CDK1, thus preventing the cell cycle from exiting the G2/M phase.Moreover, ZEA can induce apoptosis and autophagy similar to FB 1 and DON.Question marks indicate indeterminate roles.EnR, endoplasmic reticulum; MAPK, mitogen-activated protein kinase.
including the TNF pathway and the Fas pathway 46 .In the TNF pathway, the function of NF-κB is complex.Gopee et al. 51 suggested that FB 1 -induced apoptosis involved the activation of Caspase 3 in pig kidney epithelial cells (LLC-PK 1 ), which was correlated with the suppression of NF-κB.However, Chen et al. 52 suggested that FB 1 treatment resulted in the upregulation of both Caspase 3 and NF-κB in pig kidney (PK-15) cells.
In F. graminearum, the ZEA biosynthesis gene cluster contains four genes: PKS13, PKS4, ZEB1, and ZEB2 (Figure 4A).Disruption of PKS13, PKS4, or ZEB2 can result in a permanent halt in ZEA production, and the ZEB1 deletion mutant produces the ZEA derivative β-zearalenol (β-ZEL; Table 1).PKS4 can catalyze the synthesis of the hexaketide chain using one acetyl-CoA and five malonyl-CoA units.Then, the hexaketide is transferred to the nonreducing PKS13 to form nonaketide after completing three condensations.The nonaketide subsequently undergoes two successive intramolecular cyclization reactions, leading to the formation of an aromatic ring and a macrolide ring structure containing a lactone bond.Finally, β-ZEL is transformed to ZEA by ZEB1, which facilitates the transformation of the hydroxyl group on the macrolide into the ketone group 23 (Figure 4D).
ZEA and its metabolites have a three-dimensional (3D) structural similarity to estradiol and can exert estrogen-like effects.Estrogen regulates physiological processes via estrogen receptors (ERs), which are capable of initiating many signaling pathways.At low concentrations, ZEA usually induces the proliferation of cells through estrogen-like effects and carcinogenic properties 54 .ZEA and its metabolites can occupy and activate ERs and then mediate the expression of estrogen-responsive genes through the ERK signaling pathway 55 (Figure 5C).The modulation of physiological estrogen responses, such as endocrine disruption and cell proliferation, is attributed to the expression of genes regulated by estrogen 56 .The DNA damage caused by ZEA might lead to mutations or chromosome abnormalities, which can disturb the progression of the cell cycle and cause cell proliferation or cell cancerization 57,58 .Furthermore, ZEA downregulates the expression of tumor suppressor genes and upregulates the expression of oncogenes in TM3 cells, potentially promoting the conversion of normal cells into malignant cells 59 .Similarly, a change in oncogene expression might cause cell proliferation.Moreover, DNA damage caused by ZEA can also lead to cell cycle arrest.DNA damage causes the upregulated expression of ataxiatelangiectasia mutated serine/threonine kinase (ATM) and ataxia telangiectasia and Rad3-related protein (ATR), which activate CHK1 and CHK2, and cells begin to repair the damage.However, ZEA-exposed cells undergo arrest in the G2/M phase, during which there are upregulated expression of cell division cycle 25 phosphatases (CDC25) A and CDC25C.These two proteins subsequently enhance the expression of cyclin B1 (CCNB1) and cyclin-dependent kinase 1 (CDK1), thereby preventing exit from the G2/M phase of the cell cycle.The arrest of cell cycle progression triggers a halt in DNA replication and consequently inhibits cellular proliferation [60][61][62] .At high concentrations, ZEA causes mitochondrial dysfunction, EnR stress, apoptosis, and autophagy.ZEA reduces the protein expression of Nrf2 and HO-1 to further induce oxidative stress and cause cell apoptosis via the p38, JNK, and ERK MAPK pathways 63-65 .Bai et al. found that ZEA can induce apoptosis by modulating EnR stress though the PERK and ATF6 signaling pathways in porcine trophectoderm cells 66 .In addition to the classical signaling pathways, ZEA induces apoptosis through the ERK/p53/Caspase 3 signaling pathway and the Caspase 12 signaling pathway activated by Ca 2+ release from the EnR 67,68 .ZEA-induced EnR stress also causes autophagy through the PERK pathway 69 .Moreover, ZEA can cause apoptosis by activating death receptor pathways and inhibiting the PI3K/AKT pathway and can induce the expression of proinflammatory cytokines through the TNF/NF-κB pathway and MAPK/NF-κB pathway [70][71][72][73] .
ZEA can be biotransformed in the liver and bacterial gut flora of mammals by hydroxysteroid dehydrogenases (HSD).There are 4 reductive metabolites of ZEA in the reductive phase-I metabolism process: α-ZEL, β-ZEL, α-zearalanol (α-ZAL, also known as zeranol), and β-ZAL (Figure 3C).3α-and 3β-HSD can catalyze the hydroxylation of ZEA, resulting in stereoisomeric compounds αand β-ZEL, respectively.Furthermore, with the saturation of a double bond, αand β-ZEL can be further reduced into αand β-ZAL, respectively (Figure 4E).α-ZEL and α-ZAL show higher xenoestrogenic effects than ZEA, while β-ZEL and β-ZAL are just the opposite.α-ZAL has been extensively used as a growth enhancer to augment the fattening rates of cattle.Since 1985, the application of α-ZAL has been banned in the European Union. 74

CC AND FUSARIUM MYCOTOXIN CONTAMINATION
On the basis of the Sixth Assessment Report of the Intergovernmental Panel on Climate Change, it is projected that global temperatures will continue to rise, while atmospheric concentrations of carbon dioxide ([CO 2 ]) are anticipated to undergo a twofold or even threefold increase within the next 25-50 years 75 .Adverse CC can lead to more frequent occurrences of extreme weather events, such as heat waves, cold waves, heavy rainfall, and drought.CC is a global challenge and is forecasted to have significant impacts on food security by influencing crop growth, the occurrence of pests and diseases, and mycotoxin contamination.CC may lead to transformations in the spatial distribution and manifestation of mycotoxigenic fungi, thereby causing alterations in both the geographical distribution and the occurrence pattern of mycotoxins.Studies have shown that temperature, water activity (a w ), [CO 2 ], or a combination of the three variables can also impact the growth, proliferation, and production of mycotoxins from mycotoxigenic fungi 76 (Figure 6).Although pathogenic Fusarium species are important producers of mycotoxins, there is no clear correlation, in general, between disease severity and mycotoxin contamination 77 .At Bottelare village, East Flanders province, Belgium, a positive correlation between FHB severity and DON content was observed during the growing seasons of 2001-2002 and 2002-2003; however, this relationship was not evident in the 2003-2004 season 78 .

Temperature and rainfall
Although FUMs, DON, and ZEA have been detected all over the world, because of the different optimal production temperatures and a w values of Fusarium fungi (Table 2), the contaminants have clear differences in geographical distribution.FUMs are the main Fusarium mycotoxin contaminants in southern Europe, the Americas, the Middle East, Africa, and south and southeast Asia.DON contamination occurs mostly in northern, central, and eastern Europe, northern and central America, South Africa, and East Asia.Compared to those of FUMs and DON, the occurrence rate of ZEA is lower and is only over 50% in East Asia and sub-Saharan Africa 79 .
Against the backdrop of global warming, regions currently characterized by cooler climates will witness the prevalence of toxigenic fungi that show optimal growth and mycotoxin production under higher-temperature conditions.Conversely, areas already experiencing hot temperatures might observe a decrease in the occurrence of such fungi.With rising temperatures, lower crop yield and quality will occur in some regions that are currently considered warm.This might result in a reduction in total mycotoxin production due to the reduced crop quantity.However, due to the lower quality of crops, the mycotoxin content per unit weight of crops might increase 82 .Temperature is closely related to latitude and altitude.In most regions where maize is grown, FUM contamination tends to be higher in areas with lower latitudes and altitudes due to relatively warmer conditions compared to regions with higher latitudes or altitudes 83 .In the United States, the FUM risk is higher in Texas and the southeastern states than in the central states 84 .Shelby et al. 85 discovered a significant inverse relationship between latitude and FUM concentration in the United States.A similar pattern can also be found in Asia, north of the Tropic of Cancer.In the low-altitude maize production areas of central and south America, as well as southeast Asia, FUMs emerge as a significant risk factor.In Guatemala, a survey of maize samples gathered from fields between 2000 and 2003 showed that lowland maize exhibited significantly higher levels of FB 1 than highland maize did 86 .In Europe, the risk of FUM contamination is higher in Italy, Spain, and southern France.In Africa, all maizeproducing areas are at risk for FUMs, with severity depending on altitude 84 .A survey in Uganda found that the FUM contamination of maize was widespread and that maize from high altitudes showed the most significantly elevated levels of FUM content 87 .DON contamination is also sensitive to temperature changes.In northwestern European countries, according to the prediction model, the flowering and full maturation of wheat will advance with the relative increase in temperature, and most regions will show a substantial rise in DON contamination in the 2040s 88 .
Although numerous studies have demonstrated that Fusarium mycotoxin production requires high a w in vitro (Table 2) 80,81 , none have demonstrated monotonic relationships between rainfall and Fusarium mycotoxin contamination across different environments.In Ontario, Canada, a cool maize-growing region, FUMs are only present in drought-stressed fields 89 .In the United States, there is a significant negative correlation between June rainfall and FUM content at multiple locations 85 .Akello et al. 90 studied the FUM content of cereals in Zimbabwe during 2015 and 2017 and found that FUM contamination was higher in wet years than in dry years.According to existing meteorological data in the Philippines, Salvacion et al. 91 built a risk model of FUM contamination on corn using a fuzzy logic methodology.Due to the increased rainfall, they argued that a substantial proportion of the Philippines might be at a very high risk under prevailing circumstances, as well as under the anticipated CC scenarios for 2050.During the period 2012-2021, in both Serbia and Croatia, the highest mean contents of DON and ZEA in maize were observed in 2014, which might be related to the extreme precipitation during that year 92 .A 10-year (2008-2017) global survey showed that maize harvested in central and southern Europe exhibited elevated levels of DON and ZEA concentrations in 2014, which corresponded to higher rainfall in July 2014.In the primary maize-growing regions of China, DON and ZEA levels were relatively low in 2013, which might be related to the decrease in precipitation during August and September of that year 79 .
The period around flowering is considered to be the pivotal stage for Fusarium infection of cereals.Therefore, many studies have been dedicated to predicting the concentrations of Fusarium mycotoxins by the weather variables of the period around flowering, especially temperature and rainfall (Table 3).Campa et al. 93 built a model to predict the FUM concentration of maize by using data gathered from Argentina and the Philippines.The model showed that weather was the major variable in total FUM concentration; the temperature and precipitation of the four periods around silking were determined to be the key factors in the FUM concentration.Hooker et al. 94 identified a comprehensive set of weather variables and their temporal patterns for the prediction of DON contamination in mature wheat grain in Canada.They found that contamination showed a significant correlation with weather conditions during three pivotal periods around the heading stage.In the first period, 4-7 days before heading, DON generally decreased with the number of days with The optimal temperature and a w of FUMs and ZEA production were detected on Fusarium-infected maize grain.The optimal temperature and a w of DON production were detected on Fusarium-infected wheat grain.Data were extracted from Sanchis and colleagues 80,81 .a w , water activity; DON, deoxynivalenol.
temperatures below 10°C and increased with the number of days experiencing rainfall exceeding 5 mm.In the second period, 3-6 days after heading, DON levels showed an upward trend in correlation with an increase in the number of days with rainfall exceeding 3 mm and a downward trend when exposed to temperatures exceeding 32°C.In the third period, 7-10 days after heading, DON increased with an increase in days with rainfall exceeding 3 mm.In Schleswig-Holstein, northern Germany, Birr et al. 95 found that there were significant relationships between the two weather variables (cumulative precipitation and average temperature during the period of wheat flowering) and the concentrations of ZEA in wheat grain at harvest.Based on this finding, they derived weather-based forecasting models for predicting ZEA levels in wheat grain during the harvest stage for various Fusarium-susceptible wheat cultivars.Joo et al. 96 constructed a model to assess the influence of CC on ZEA contamination in rice grains cultivated in South Korea.The results indicated that increased temperature and relative humidity during the rice heading period and fluctuations in daily temperature throughout the harvest season can increase ZEA contamination in rice.The forecasts showed that ZEA contamination of rice could increase nationwide in both the 2030s and the 2050s, particularly in the western region of South Korea.

[CO 2 ] level
Although the impacts of elevated [CO 2 ] on crops have been studied in depth 97  ], the suppression of maize 13-lipoxygenase and jasmonic acid production was correlated with a decrease in terpenoid phytoalexins and an increase in susceptibility to the pathogen, while a reduction in 9-lipoxygenase, previously proposed to enhance mycotoxin production, was responsible for reduced FUMs per unit fungal biomass 98 .Further research showed that concurrent elevated [CO 2 ] and drought stress significantly augmented the susceptibility of maize to F. verticillioides infection, consequently leading to an escalated contamination of FUMs.However, the negative impacts of drought on the accumulation of maize phytohormones and metabolites were not mitigated by elevated [CO 2 ], and there was still no observed increase in FUMs per unit fungal biomass.Therefore, it is likely that the escalation in FUM contamination can be attributed to the greater F. verticillioides biomass 99 .F. graminearum infection produced similar results.Hay et al. 100 found that elevated [CO 2 ] can significantly increase F. graminearum biomass and DON accumulation in maize, but the DON per unit fungal biomass was unaffected.For wheat, it was a different story.In the F. culmorum single-floret inoculation treatment, the concentration of DON was significantly increased under elevated [CO 2 ].This result suggests that the DON content is not directly related to the level of infection with F. culmorum 101 .(2) 3 to 6 days after heading RAIN >3 mm TMAX > 32 °C (3) 7 to 10 days after heading RAIN >3 mm ZEA Wheat Flowering period The higher cumulative precipitation and average temperature [93]   Rice (1) Flowering period The higher average temperature and humidity [94]   (2) Harvest period The higher daily temperature changes a The risk of Fusarium mycotoxin contamination will increase with the number of days experiencing rainfall exceeding 2 mm.b The risk of Fusarium mycotoxin contamination will decrease with the number of days when the minimum temperature is below 15 °C.
In addition, acclimatization to elevated [CO 2 ] can impact the mycotoxin production of Fusarium fungi.A recent in vitro study demonstrated that, under elevated [CO 2 ] conditions, F. sporotrichioides had a greater ability to produce T-2 and HT-2 after 10 subculture generations than in the initial subculture of the strain 102 .

DETECTION OF FUSARIUM MYCOTOXINS
Typically, analysis methods for mycotoxins require three major steps: extraction, cleanup, and detection 103 .Mycotoxins can be detected using various techniques, mainly chromatographic methods, immunological methods, and biosensor technologies (Table 4).However, each approach has advantages and disadvantages.The selection of a particular method is contingent upon the specific detection requirements 20 .

Extraction and precleaning methods
Primary extraction is essential for the determination of mycotoxins in various sample types.The cleanup step can eliminate interference from the extract and concentrate the analyte, and it is essential for the analysis of mycotoxins, especially at trace levels 102 .Currently, common approaches include solid phase extraction (SPE), multifunctional cleanup columns, liquid-liquid extraction (LLE), solid-liquid extraction (SLE), and immunoaffinity column (IAC) 170 .The selection of a method for mycotoxin extraction depends on the types of analytes.However, some methods can incur high costs, intricate procedures, and/or substantial time and solvent consumption.To minimize the sample treatment but prevent exposure to matrix effects, the "quick, easy, cheap, effective, rugged, and safe" (QuEChERS) sample preparation approach is a viable alternative.The QuEChERS method has been used for the extraction of mycotoxins from food samples, including dried fruits and cereals, as well as liquid samples, such as wine and beer 171 .

Chromatographic methods
There are many kinds of chromatographic analytical methods for mycotoxin analysis, such as thin-layer chromatography (TLC), high-performance TLC (HPTLC), gas chromatography (GC), high-performance liquid chromatography (HPLC), and ultra-performance liquid chromatography (UPLC).HPLC has emerged as the most widespread technique for mycotoxin analysis.By coupling to detectors, such as mass spectrometry (MS), ultraviolet (UV) detectors, visible detectors, and fluorescence (FL) detectors (FLDs), the compounds separated by chromatography can be further identified 20 .Currently, LC-MS and liquid chromatography FL detection (LC-FLD) are widely recognized as the standard methods for detecting Fusarium mycotoxins.There is no doubt that FLD is the most sensitive among all LC detectors.The sensitivity, precision, and accuracy of LC-MS may vary depending on the mycotoxins, matrix, ionization technique, and sensitivity of the process.Due to ion suppression and matrix effects, LC-MS often causes undesirable results for the quantitative measurement of mycotoxins.Tandem mass spectrometry (MS/MS) is the preferred detection method over FLD due to its ability to identify a wide range of both fluorescent and nonfluorescent mycotoxins, making it a cost-effective choice 103 .With the ongoing development of technology, high-throughput determination methods of single or multiple matrices using LC-MS/MS have been reported.Steiner et al. 172 developed a pioneering multiclass quantitative method for the analysis of over 1200 biotoxins, pesticides, and veterinary drugs in complex feeds by LC-MS/MS.In the past decade, liquid chromatography coupled with high-resolution mass spectrometry (LC-HRMS) has turned from a research-only technique into a costly tool for routine testing and high-throughput food analysis in laboratories.Although these methods were initially developed for pesticide detection, mycotoxins are now the primary focus of LC-HRMS method development 173 .High resolution, in combination with the fast generation of product ion spectra, has the potential to minimize indistinct outcomes and streamline peak detection, but there remains a disparity between LC-HRMS and LC-MS/MS regarding the limits of detection (LOD) and of quantitation for most analytes.To address the gap for most analytes and enhance the applicability of mycotoxin trace analysis, various strategies for analyte enrichment, including SPE, have been incorporated into experimental protocols.However, these additional procedures invariably lead to a substantial augmentation in manual laboratory tasks and expenses, thereby diminishing the potential advantages of HRMS systems 173,174 .Mateus et al. 175 developed a UHPLC-HRMS multianalyte method for pistachio nuts.They evaluated different approaches to dispersive SPE for high-lipid matrices; eventually, two procedures were validated.One involved the addition of enhanced matrix removal-lipid for the detection of FUMs, and the other used a zirconium-based material to achieve a slightly heightened sensitivity in analyzing G-type AFs without including FUMs.
HPLC, UHPLC, and GC coupled with non-MS detectors are also reference methods for Fusarium mycotoxin analysis.The current trend for the chromatographic analysis of Fusarium mycotoxins with non-MS detection involves the advancement of multitoxin analysis with FLD, UV detection, photodiode array (PDA) detection, and so forth.Pi et al. 176 developed a novel method for the simultaneous determination of nine mycotoxins based on ultrasonic-assisted aqueous two-phase extraction coupled with solidifying organic drop-dispersible liquid-liquid microextraction by HPLC with a diode array detector and FLD in series.The methodology effectively identified various mycotoxins in multiple foods.Furthermore, it can be utilized to effectively perform regular and extensive analyses of numerous mycotoxins within various samples.Lee et al. 177 built a simple and reliable HPLC-UV method for the simultaneous determination of DON, NIV, DON-3G, and NIV-3G.This method involves a straightforward sample extraction with IAC purification and was successfully applied to analyze 31 different baby formulas and Korean rice wines available on the Korean market.Ref.

MIP-EC biosensor
The negligible CRs with AFB

Immunological methods
Immunological detection methods rely on the antibody-antigen (Ab-Ag) binding relationship and vary from enzyme-linked immunosorbent assay (ELISA) and lateral flow immunoassay (LFI) to advanced immunosensors.ELISA is a technique utilized to detect the presence and quantity of Ag binding in biological samples based on the principle of Ag-Ab interactions.In recent decades, numerous ELISA kits have been effectively commercialized for the detection of Fusarium mycotoxins.ELISA tests are portable, simple, fast, and do not require expensive analytical equipment.This continues to make ELISA tests popular.Nevertheless, ELISAs often show limited precision at low concentrations, and structurally similar mycotoxins or matrices can impede conjugate and Ab binding, causing errors in quantifiable mycotoxin ELISA measurements 103 .
LFI is a simple one-step immunochromatographic paper assay that does not require complex instruments.It can be classified into two modes, competitive and sandwich; typically, LFI for mycotoxin detection adopts the competitive type.The basic LFI equipment consists of sample coating pads, conjugate-release pads, absorbent pads, and membranes (also called detection pads) 178 .For the user, LFI is strikingly simple: after simply adding the sample onto a single paper lateral flow strip and a short incubation time, the qualitative or semiquantitative result of the test is revealed by the appearance of a test line, and quantification can then be conducted using an optical reader 179 .The exceptional advantages of LFI in terms of convenience, affordability, and rapidity make it particularly suitable for on-site monitoring and rapid testing for Fusarium mycotoxin contamination in foods.In recent years, the advancement of novel nanomaterials has broadened the types of labels available for LFI of Fusarium mycotoxins.Colored nanoparticle (NP)-based LFI is simpler and more convenient, and it shows significant potential for on-site detection 180 .Due to the intricate nature of mycotoxin co-occurrence, there is a growing need for simultaneous detection of multiple mycotoxins.To date, several multiplex LFIs for mycotoxins with excellent performance have been effectively developed.For example, Liu et al. 181 devised an innovative LFI integrated with gold nanoparticles (AuNPs) and time-resolved FL microspheres.Their LFI is a smartphone-based quantitative dual detection for multiplex mycotoxins in cereals, such as aflatoxin B 1 (AFB 1 ), ZEA, DON, T-2, and FB 1 .

Biosensors
Biosensors are bioanalytical devices that incorporate biological recognition elements to bind target molecules and a signal transducer for converting the biorecognition event into a quantifiable signal.Biorecognition elements, including Abs, Ags, nucleic acids, and enzymes, are used for the identification and detection of target analytes.Based on a variety of bioinspired recognition elements, biosensors can be classified into immunosensors, aptasensors, and molecularly imprinted polymer (MIP)-based sensors 179 .According to the principle of signal transduction, biosensors can be divided into electrochemical (EC) biosensors, optical biosensors, mass-sensitive biosensors, thermal biosensors, and so on 182 .Compared to the other analytical methods mentioned above, biosensors offer a much simpler and more efficient means of dynamically monitoring reaction changes in real-time with digital outputs.Biosensors not only reduce detection time but also enhance sensitivity, simplicity, robustness, and reusability, enabling the development of cost-effective, highthroughput screening methods for mycotoxins 179 .
Immunosensors, as novel and widely utilized analytical instruments, use Ab as the recognition element and a transducer to convert the Ag-Ab binding event into a quantifiable physical signal.Due to the superior specificity of the Ag-Ab immunoreaction, immunosensors show superior selectivity and sensitivity 183 .Various immunosensors have been developed based on the different mechanisms of signal variations, including FL, colorimetric, chemiluminescence, electrochemiluminescence, surface plasmon resonance (SPR), surface-enhanced Raman spectroscopy (SERS), and EC immunosensors.By combining the exceptional specificity of Abs and the remarkable sensitivity of FL detection, FL immunosensors have recently emerged as highly favored contenders for mycotoxin detection 183 .With advancements and breakthroughs in nanotechnology, a diverse array of nanomaterial semiconductors, including semiconductor quantum dots (QDs), QD nanobeads (QBs), carbon dots (CDs), fluorescent metallic NPs, and upconversion nanoparticles (UCNPs), with unique photostability, bright FL, and good biocompatibility have garnered immense attention regarding their use in the construction of FL immunosensors 184 .Yang et al. 185 devised a novel FL immunosensor detection platform that integrates multicolor UCNP barcoding technology with smartphone-based portable devices for simultaneous analysis of multiple mycotoxins (AFB 1 , ochratoxin A [OTA], and ZEA).The quantitative detection platform demonstrated feasibility and reliability, with a LOD of 1 ng that surpassed the values obtained from standard assays.SERS is a surface-sensitive vibrational spectroscopy technique for the detection and characterization of analytes that are adsorbed on or close to the surface of plasmonic nanostructures.SERS integrates the advantages of the molecular specificity of Raman spectroscopy and the optical sensitivity of plasmonic nanostructures to boost the Raman signal, greatly extending the role and application field of standard Raman spectroscopy 183 .SERS significantly amplifies the Raman signal, thereby expanding the scope and applicability of conventional Raman spectroscopy in a profound manner.SERS immunosensors, which integrate the SERS labeling technique with Ag-Ab specific interactions, have emerged as novel immunosensing devices for mycotoxins.Li et al. 186 developed a SERS immunosensor to simultaneously detect AFB 1 , ZEA, and OTA by using AuNPs labeled with 5,5-dithiobis(succinimidyl-2-nitrobenzoate) as a Raman reporter.The SERS immunosensor determination method demonstrated results that are consistent with conventional instrumental analysis.
Aptamers are novel recognition elements with exceptional affinity and specificity that have the potential to serve as recognition molecules in aptasensors (aptamer-based biosensors) for the efficient and swift identification of various targets 187 .Aptamers can be divided into two categories, DNA/ RNA-based aptamers and peptide aptamers.Currently, aptamers for the detection of mycotoxins are a class of singlestranded DNAs or RNAs that undergo screening through the systematic evolution of ligands by exponential enrichment, which can selectively bind to different ligands through noncovalent bonds 188 .The functional similarity of aptamers to Abs makes them widely applicable in the field of biosensors.Compared to Abs, aptamers are low cost, have a longer shelf life and good stability, even at elevated temperatures, and are easy to modify and synthesize 189 .As with immunosensors, according to the signal variation mechanism, aptasensors can also be classified as FL aptasensors, EC aptasensors, SPR aptasensors, SERS aptasensors, CEL aptasensors, and so on.EC aptasensors use electrodes as sensing units and electrochemical workstations as signal transformation systems 190 .Aptamers are typically immobilized on the surface of the electrode; the specific molecular interactions between molecules on the electrode surface result in the conversion of target binding into electrical signals.These electrical signals, such as current, resistance, potential, or capacitance, are transmitted to a computer for quantitative or qualitative analysis of the target 187 .Zhang et al. 191 devised an electrochemical aptasensor-based target-induced strand displacement strategy to achieve highly sensitive detection of T-2.The aptasensor was highly specific, stable, and suitable for T-2 detection in real samples.SPR is a phenomenon in which the electrons in a metal surface layer are excited by photons of incident light with a certain angle of incidence and then propagate parallel to the metal surface.With a constant light source wavelength and a thin metal surface, the angle that triggers SPR depends on the refractive index of the material near the metal surface.The affinity binding interaction on the surface of thin metal films can cause a small change in the reflective index of the sensing medium, which can hinder the occurrence of SPR.SPR aptasensors can detect those changes on the optical transducer surface, and optical transduction can directly convert the molecular binding event into a physically measurable signal, which is proportional to the concentration of analyte molecules 192 .As a label-free analytical strategy, SPR aptasensors have the capability to detect multiple mycotoxins simultaneously with real-time monitoring, high sensitivity, good specificity, minimal sample preparation requirements, and high-throughputdetection 192 .Wei et al. 144 effectively and simultaneously detected OTA, DON, AFB 1 , and ZEA in wheat and corn using SPR aptasensors that demonstrated high sensitivity, good linearity, and specificity.
MIPs, also referred to as "plastic antibodies," are bespoke synthetic receptors that use the molecular imprinting technique.Following the general molecular imprinting method, imprint molecules (templates) are added along with functional monomers and cross-linkers, which are polymerized under appropriate conditions.After copolymerization, the templates are extracted, leaving the 3D matrix with cavities that possess a matching shape, size, and chemical functionality with the template.The resulting cavities possess the ability to selectively reassociate with the template upon subsequent exposure and become synthetic receptors 174,193 .The interactions between the polymers and the templates are similar to those between Abs and Ags.MIPs provide a costeffective and easily prepared approach for targeted template recognition with outstanding biocompatibility, repeatability, and broad utility.MIPs are particularly robust in extreme environments, such as elevated pressures, fluctuations in pH levels, and exceptionally high or low temperatures.By integrating MIPs with various sensing reporter systems (electrical, EC, optical methods, etc.), a valuable device can be developed for monitoring or screening purposes 194 .Utilizing a blend of MIP membranes (ZEA-selective urethane acrylate MIP membranes) and the Spotxel ® Reader smartphone application (Sycasys Software GmbH), a miniature sensor was used to analyze the natural FL of ZEA in cereals in the field 195 .NPs show a high level of sensitivity, yet their selectivity is limited.Therefore, the combination of MIPs and NPs can result in MIP composites with high sensitivity and selectivity.To date, several NPs, including QDs, UCNPs, carbon NPs (CNPs), AuNPs, magnetic NPs, and metal-organic frameworks, have played important roles in mycotoxin analysis with MIPs 193 .The molecularly imprinted polymer nanoparticlebased assay (MINA) was developed for the determination of FB 1 -the molecularly imprinted nanoparticles replace the primary Ab used in a competitive ELISA.The MINA showed a high level of specificity and did not show any cross-reactivity with other mycotoxins, including AFB 1 , CIT, DON, ZEA, and FB 2 .The results of the MINA agreed with those obtained using traditional ELISA and HPLC methods 196 .Calahorra-Rio et al. 197 developed a new molecularly imprinted magnetic nanobead that can specifically extract ZEA from river and tap water for further analysis using HPLC-FLD.Their findings indicate that the magnetic nanobead showed exceptionally accurate and consistent ZEA detection in real liquid samples.

THE MANAGEMENT STRATEGY FOR FUSARIUM MYCOTOXIN CONTAMINATION
Fusarium mycotoxins can be produced by several fungi in several stages of several crops, so a single management strategy cannot fully control Fusarium mycotoxin contamination.However, a series of practices, including agronomic, chemical, physical, and biological methods, can be implemented to avoid the spread of mycotoxins and to minimize their frequency in food products during preharvest and postharvest stages (Figure 7).

Agronomic method
Fusarium fungi can survive in plant residues and on wild grasses, existing as mycelia, conidia, or perithecia.Therefore, residue management, tilling, and deep plowing can reduce the primary Fusarium inoculum that causes infections and the presence of mycotoxins 12 .Reduced or no tillage practices can contribute to increased DON contamination of wheat and maize 198 .Proper crop rotation can substantially mitigate Fusarium disease and mycotoxin contamination.An unfavorable practice in crop rotation is the consecutive cultivation of cereal plants 12 .Qiu et al. 199 collected 90 wheat samples in China during 2013 and 2014, revealing that DON contamination was more prevalent in the rice-wheat rotation, while ZEA accumulation was found to be higher in the maize-wheat rotation.Crop rotation with noncereals can be a more favorable method for limiting Fusarium mycotoxin contamination; legumes, brassicas, and root crops might be better forecrops 12 .Moreover, the implementation of intercropping is a good method for controlling Fusarium mycotoxin contamination.Drakopoulos et al. 200 found that the use of white mustard or Indian mustard as an intercrop with maize can reduce DON in winter wheat compared to maize grown as a sole crop in maize-wheat rotation fields under reduced tillage.The cultivation system, conventional or organic, also affects Fusarium mycotoxin contamination of cereals.Bernhoft et al. 201 conducted a comprehensive analysis of available published studies (1991-2017) of controlled field experiments, comparing DON, ZEA, and T-2/HT-2 in grains from organic and conventional cultivation systems.Summary of the results revealed that organic production showed lower mycotoxin levels in 24 cases, no significant difference in 16 cases, and higher levels in only two cases.

Host resistance
The most economical and sustainable approach to controlling Fusarium disease and mycotoxin contamination is through the cultivation of varieties that show resistance or partial resistance to Fusarium species, whether because of traditional breeding or through use of transgenic technology.Taking the case of FHB in wheat, there are five types of resistance: Type I (resistance to initial infection), Type II (resistance to disease spread within infected heads), Type III (resistance to mycotoxin accumulation), Type IV (resistance to kernel damage), and Type V (tolerance) 202 .These FHB resistance traits of wheat are governed by multiple quantitative trait loci (QTLs).Over 250 QTLs for FHB have been found on all 21 chromosomes in various wheat genotypes 203 .Fhb1 on chromosome 3BS is the most important QTL.The Chinese wheat cultivar Sumai 3, known for carrying the Fhb1 gene, is widely acknowledged as the superior source of FHB resistance and has been extensively utilized as a parent in numerous breeding programs 204 .Transgenic germplasms can offer novel supplementary resources for FHB management.Wheat resistance to F. graminearum could be significantly enhanced by overexpression of genes that encode defense signaling pathway-related proteins, cell wall-degrading enzyme inhibitors, and detoxification proteins 22 .McLaughlin et al. 205 showed that the overexpression of AtLTP4.4,a nonspecific lipid transfer protein-encoding gene of Arabidopsis thaliana, in transgenic wheat, can significantly reduce F. graminearum infection and DON contamination in the field.Based on RNA silencing, plants naturally have a defense system to stave off viral invasion.This feature has been harnessed to advance host-induced gene silencing (HIGS) technology to control other plant pathogens by silencing pathogen genes in plants during infection.HIGS is also an effective control strategy against FHB in wheat.Recently, many researchers have successfully silenced important genes encoding cytochrome P450, chitin synthase, protein kinases, and others in F. graminearum, Fusarium oxysporum, F. culmorum, and F. verticillioides, resulting in reduced mycotoxin production 206 .

Chemical method
Currently, due to the absence of efficacious disease-resistant cultivars, the utilization of chemical fungicides remains the primary strategy for Fusarium disease management.There are several common types of fungicides for controlling Fusarium diseases, including benzimidazoles (benomyl, carbendazim, thiophanate-methyl, and thiabendazole), triazoles (triadimefon, tebuconazole, diniconazole, and propiconazole), and strobilurins (azoxystrobin and pyraclostrobin).Strobilurin treatment can induce DON formation, so strobilurins are unlikely to be effective 207 .Triazoles, a group of demethylation inhibitor fungicides, are the gold standard for controlling FHB in wheat worldwide, while carbendazim is the most widely used fungicide in China for this purpose 207,208 .In addition, phenamacril, a myosin I inhibitor and cyanoacrylate fungicide, is an efficacious and highly speciesspecific fungicide for FHB in China.Phenamacril is not only capable of inhibiting the mycelial growth of a few Fusarium species, including F. graminearum, Fusarium asiaticum, F. verticillioides, and F. oxysporum, but also significantly disrupts DON-toxisome formation to hinder DON biosynthesis 209 .Although spraying fungicides is a very effective method, the use of chemical fungicides comes with its own set of challenges.In addition to the well-known environmental problems caused by fungicides, fungicide-resistant population of Fusarium species are also an important concern.Due to prolonged and intensive use of fungicides, the benzimidazole-and triazole-resistant strains of F. graminearum have become feared throughout world agriculture.There was a high frequency of carbendazim-resistant F. graminearum in China.In response, the local government of Jiangsu Province, where large carbendazim-resistant F. graminearum population had appeared, even proposed a fallow subsidy policy 210 .Regarding triazoles, tebuconazole-resistant F. graminearum strains have been found in Argentina, China, Germany, and the United States 207 .Therefore, the development of new fungicides for the management of Fusarium diseases and mycotoxin contamination is urgently needed.
Chemical preservatives can be added to stored grain, particularly in the case of damp grains intended for animal feed.Common chemical preservatives include mixtures of aliphatic acid salts and antioxidants.The former has demonstrated efficacy in controlling FB 1 production of F. proliferatum on irradiated maize kernels 211 .The two antioxidants, butylated hydroxyanisol and propyl paraben, were shown to reduce both the growth and FUM production of F. verticillioides and F. proliferatum in culture media and maize grain 212 .Recently, considering the adverse effects of synthetic preservatives, there has been a growing trend toward utilizing plant essential oils (EOs) as viable alternatives for mycotoxin management.

Biological control method
Biological control is an eco-friendly approach that uses living organisms or their derivatives to control pests.The useful biological control agents (BCAs) for Fusarium diseases and mycotoxin contamination include beneficial bacteria, fungi, actinomycetes, and mycoviruses.BCAs can protect crops against pathogenic Fusarium fungi before harvest, thereby effectively mitigating the risk of mycotoxin contamination in the food chain.The mechanisms of BCAs encompass competition for space and nutrients, mycoparasitism, synthesis of antifungal metabolites, cross-protection, promotion of plant growth, and induction of host plant resistance 213 .
Among the bacterial BCAs, two genera that secrete antifungal compounds, Bacillus (such as Bacillus velezensis, Bacillus megaterium, and Bacillus subtilis) and Pseudomonas (including Pseudomonas aeruginosa, Pseudomonas frederiksbergensis, Pseudomonas fluorescens, and Pseudomonas simiae), are the most widely used and can effectively reduce Fusarium infection and mycotoxin contamination under field conditions 214 .Bacillus contain three important antifungal lipopeptides that inhibit Fusarium pathogens, including Bacillomycin D, surfactin, and fengycin (synonymous with plipastatin) 215 .Bacillomycin D and fengycin can inhibit the growth of hyphae, cause morphological alterations or destruction of cell walls and plasma membranes, and trigger the cell lysis of F. graminearum and Fusarium moniliforme through the accumulation of ROS.Moreover, Iturin A, a kind of Bacillomycin D, can also control the T-2 toxin synthesis of F. oxysporum by inhibiting TRI5 expression 216 .Regarding F. moniliforme, surfactin can inhibit hyphal growth and induce ROS, damaging DNA and protein in living cells 217 .In addition to direct interactions, bacterial BCAs can also protect plants against Fusarium pathogens in indirect ways.Chen et al. 218 suggested that B. velezensis LM2303 has various biocontrol mechanisms against F. graminearum, such as enhancing wheat systemic resistance, facilitating wheat plant growth, and outcompeting for space and nutrient competition via efficient colonization and antibacterial metabolites.
The genera Aureobasidium, Cladosporium, Clonostachys, Cryptococcus, Sarocladium, and Trichoderma are the representative fungal BCAs for Fusarium disease management.Trichoderma is a notable example of a fungal antagonist, as it not only inhibits the growth and reproduction of Fusarium fungi through mycoparasitism, antibiotics, promotion of plant growth, and the induction of defense responses in host plants, but also has the ability to control the biosynthesis of Fusarium mycotoxins and directly degrade Fusarium mycotoxins 219 .Błaszczyk et al. 220 reported that Trichoderma atroviride shows significant inhibitory effects on the biosynthesis of mycotoxins (DON, 3ADON, 15ADON, NIV, ZEA, BEA, and MON) of Fusarium avenaceum, Fusarium cerealis, F. culmorum, F. graminearum, and Fusarium temperatum.Tian et al. 221,222 indicated that Trichoderma fungi can convert ZEA into ZEA sulfate and ZEL sulfate by sulfation and glycosylate TRIs into glycosylated forms.Galletti et al. 223 found that Trichoderma gamsii B21 can remove FUMs from liquid cultures, but the mechanisms were unclear.Recently, the utilization of endophytic fungi as BCAs has become regarded as a compelling strategy for controlling plant disease.Kemp et al. 224 found that the wheat endophytic fungus Sphingobacterium zeae NRRL 34560 can induce defense responses in wheat, effectively controlling FHB and DON contamination in a greenhouse.
After harvest, beneficial microorganisms can be used to control Fusarium mycotoxin contamination.Lactic acid bacteria (LAB), which are frequently used as biological preservatives, not only can prevent Fusarium fungal growth but can also control mycotoxin contamination in food and feed.Strains of Lactobacillus, Bifidobacterium, Lactococcus, Leuconostoc, Pediococcus, Propionibacterium, and Streptococcus have been used to control mycotoxin contamination 229 .LAB produce many antifungal metabolites, such as organic acids, hydrogen peroxide, diacetyl, rutherin, reutericyclin, acetoin, bacteriocins, and bacteriocin-like inhibitory substances 230 .In addition to its antifungal activity, LAB possess the ability to adsorb, degrade, or detoxify Fusarium mycotoxins.On one hand, LAB can assimilate mycotoxins toward their cell wall operative groups; on the other hand, LAB can degrade mycotoxins via metabolic apparatus or enzymes 229 .
A drop in the utilization of synthetic preservation has occurred due to its side effects, such as resistance development in pests, nonbiodegradable characteristics, and toxic effects on nontargeted organisms.Therefore, as green preservatives, plant EOs have become widely used substitutes for synthetic preservatives.In the past decade, cinnamon, clove, eucalyptus, fennel, oregano, rosemary, palmarosa, and thyme were the most frequently used EOs to control mycotoxigenic fungi and their mycotoxins.EOs can prevent fungal infection and mycotoxin biosynthesis in many ways, such as the inhibition of fungal growth, destruction of cell permeability, disruption of the electron transport chain, and modulation of gene expression patterns and metabolic processes 231 .Due to the development of nanotechnology, nanoencapsulation of EOs has emerged as a novel strategy to increase the stability of EO constituents, extend their applicability, and overcome their major limitations by controlled release.Singh et al. 232 fabricated clove oil nanoemulsions and found that they can control the growth of F. proliferatum and reduce the FB 1 and FB 2 contents of maize.

Physical method
Physical methods can be used to eliminate mycotoxins during postharvest storage and processing; these include separation, debranning and milling, heat treatment, radiation, and adsorption.Mycotoxins are mainly found in the moldy, fragmented, and discolored parts of grains, and the specific gravity of mycotoxin-contaminated cereals is lower than that of uncontaminated cereals 233 .These characteristics enable Fusarium-damaged grains to be segregated by aspiration, image processing techniques, gravity separation, and photoelectric separation.In general, the external layer of grains tends to show higher levels of mycotoxin contamination.Partial debranning of 10% of the wheat grain weight can result in a 64% reduction in DON content 234 .Therefore, debranning and milling procedures are important for the decontamination of Fusarium mycotoxins.After polishing, the Fusarium mycotoxin content of white rice can be markedly decreased 235 .Regarding the milling process, Fusarium mycotoxins tend to accumulate in the outer fractions of wheat grains (bran, flour shorts, screenings, and middlings), which are primarily utilized as animal feeds, and lower concentrations are found in the inner fractions (flour or semolina) intended for human consumption 236 .
There are a series of innovative physical methods to control Fusarium mycotoxin contamination that are usually based on nonthermal techniques: cold atmospheric plasma (CAP), electron beam irradiation, pulsed light, and so on.The CAP process makes use of an ionized gas at near room temperature to inactivate microorganisms 237 .CAP is also successful at degrading mycotoxins due to the oxidizing potential of plasma.Compared to conventional and other nonthermal approaches, CAP shows rapid efficacy in controlling fungi and mycotoxins, exerts minimal influence on product quality, and requires only low energy consumption 238 .Feizollahi et al. 239 found that CAP treatment can reduce DON in barley by 49% in 6 min, and Wielogorska et al. 240 reported a significant decrease (66%) in FB 1 in maize following a 10-min treatment with CAP.So, CAP technology represents remarkable progress in the food industry.Unfortunately, prior studies have only been successful on the laboratory scale.Therefore, additional research is warranted to demonstrate the effectiveness within wide-scale food production and different food matrices, as well as of different types of gasses 233 .

CONCLUDING REMARKS
In this review, we elucidate the effects of Fusarium mycotoxin contamination on food safety and summarize the countermeasures to prevent or mitigate harm to humans.In addition, we discuss how CC influences Fusarium mycotoxin contamination-many models have been constructed on historical or current datasets of climatic conditions to anticipate the interactions of Fusarium mycotoxins with CC.However, most models lack the [CO 2 ] factor, which exerts a significant influence on the growth of both crops and fungi.The ultimate goal of controlling Fusarium mycotoxin contamination is to protect human health.Nevertheless, few studies have estimated the transfer of mycotoxins from fields to human foods or their eventual impact on human health under different CC scenarios.
Fusarium mycotoxins cannot be completely eliminated worldwide.All we can do is to reduce contamination to acceptable levels that do not threaten human health.Therefore, a number of detection technologies and management strategies have been developed.Both agricultural production and industrial food processing are economic activities; therefore, scholars and practitioners should consider both the economic benefits and the costs of the application of Fusarium mycotoxin countermeasures to producers.With the improvement of living standards, environmental stability begins to take precedence over production efficiency and economic benefits.The development and recommended use of farm chemicals is a representative example.So, the development of Fusarium mycotoxin countermeasures should be based on a balance among these three factors.Good agricultural practices (GAPs) are a set of guidelines for producing safe and healthy food and nonfood agricultural products through on-farm and post-production practices, with due consideration of the sustainability of economic, social, and environmental aspects 241 .With regard to the control of Fusarium mycotoxins, GAP is of great significance and has been applied in many countries 242 .Nevertheless, it is not easy to change the agricultural practices of smallholder farmers.These changes may require more communication, education, and support from local governments, especially in developing countries.

Figure 2 .
Figure 2. The varieties of mycotoxin production in Fusarium species.On the left side, the cladogram depicts phylogenetic relationships among the chosen set of Fusarium species.On the right side, the circle represents the mycotoxin production abilities of Fusarium species.The cladogram was constructed by maximum likelihood analysis using the RPB1 + RPB2 dataset described by O'Donnell et al. 16 in RAxML 8.2.10.Support for branches was determined through bootstrap analysis (1000 replicates), and only values of more than 60 are shown.The distribution of mycotoxin production was based on the work of Munkvold et al. 11 .White space indicates that there are no chemical or genetic evidence for mycotoxin production.

Figure 4 .
Figure 4.The formation approaches of Fusarium mycotoxins.(A) Mycotoxin biosynthetic genes and gene clusters in Fusarium.Arrows denote the location and transcriptional orientation of genes, while the corresponding gene name is provided adjacent to each arrow.(B) The proposed pathway of TRI biosynthesis.The 3,4,15-triacetoxyscirpenol is an important branching point.One path leads to a Type A TRI, and the other leads to a Type B TRI.In F. sporotrichioides, FsRI1 exclusively catalyzes hydroxylation of 3,4,15-triacetoxyscirpenol at C-8, resulting in Type A TRIs, while in F. graminearum, FgTRI1 mediates hydroxylation of 3,4,15-triacetoxyscirpenol at both C-7 and C-8, resulting in Type B TRIs. Dashed arrows denote steps where gene assignment has not been established.(C)The proposed pathway of FUM biosynthesis.Fum11p, a tricarboxylate transporter, transports tricarboxylic acid precursors out of the inner mitochondrial lumen for FUM biosynthesis, while Fum7p and Fum10p catalyze the conversion of tricarboxylic acid precursors into acetyl CoA-activated tricarballylic acid.This product is then esterified to the polyketide backbone at C-14 and C-15 by Fum14p to produce either FB 3 or FB 4 .Finally, the hydroxylation of FB 3 and FB 4 by Fum3p yields FB 1 and FB 2 , respectively.(D) The proposed pathway of ZEA biosynthesis.The biosynthesis begins with PKS4, which facilitates the condensation of carbons derived from one acetyl-CoA and five malonyl-CoAs.PKS13 completes the polyketide backbone as an extender unit and is responsible for cyclization and aromatization.As the final step, β-ZEL is converted into ZEA by ZEB1.(E) The formation pattern of metabolites of ZEA.The C6 keto group of ZEA is reduced to yield α-ZEL and β-ZEL, followed by a subsequent reduction at the C11-C12 double bond that results in the formation of α-ZAL and β-ZAL, respectively.Those proposed biosynthesis and metabolites pathways were derived from the works of McCormick et al.17 , Alexander et al21 , Nahle et al23 and EFSA Panel on Contaminants in the Food Chain (CONTAM) et al74 .CAL, calonectrin; FgTRI1, F. graminearum TRI1; FsTRI1, F. sporotrichioides TRI1; FUM, fumonisin; PKS, polyketide synthase; SAM, S-adenosyl methionine; TRI, trichothecenes; ZEA, zearalenone.

Figure 5 .
Figure 5.The toxicological mechanisms of the main Fusarium mycotoxins.(A) DON-induced oxidative stress can cause DNA damage, EnR stress, and the suppression of the PI3K/AKT pathway.Consequently, it can lead to a decrease in Bcl-2 gene expression, an increase in Bax gene expression, and elevated levels of CytoC expression.It enhances the upregulated expression of Caspase 9 and Caspase 3 and induces apoptosis.DON can efficiently induce the activation of MAPKs through a mechanism referred to as the "ribotoxic stress response" and hijack the JAK2/STAT-3 pathway to induce apoptosis, proinflammatory cytokine upregulation, and cell cycle arrest.DON also activates Caspase 8 to increase expression of Caspase 3 by the tumor necrosis factor-α and Fas/FasL signaling pathways, leading to apoptosis.(B) FB 1 disrupts sphingolipid metabolism, causing DNA damage and apoptosis through the PTEN/PI3K/AKT signaling pathway as well as oxidative stress, which further entrenches DNA damage and apoptosis and induces EnR stress.FB 1 -induced EnR stress not only causes apoptosis by the JNK/p53/PUMA/Caspase 3 pathway but also leads to autophagy via signaling pathways mediated by IRE1, PERK, and AMPK.IRE1 can activate JNK, which results in the subsequent elevation of BECN1, ATG5, and ATG7 and the conversion of LC3-I.PERK induces the expression of ATG5 and ATG7 to form the ATG5-ATG7 complex and promotes the conversion of LC3-I into LC3-II.Under EnR stress, AMPK inhibits the anabolic process of mTOR, thereby increasing the expression of autophagy-related genes and triggering autophagy.(C) ZEA and its derivatives can bind to ERs to elicit estrogen-like effects and promote cellular proliferation.ZEA-induced DNA damage can upregulate ATM and ATR expression and activate DNA damage checkpoints CHK1 and CHK2 to repair DNA damage.Next, the expression levels of CDC25A and CDC25C are upregulated, which promotes the expression of CCNB1 and CDK1, thus preventing the cell cycle from exiting the G2/M phase.Moreover, ZEA can induce apoptosis and autophagy similar to FB 1 and DON.Question marks indicate indeterminate roles.EnR, endoplasmic reticulum; MAPK, mitogen-activated protein kinase.

Figure 6 .
Figure 6.The impact of climate change on Fusarium mycotoxin contamination.Temperature, rainfall (or water activity), and carbon dioxide concentration ([CO 2 ]) are the main known climate drivers affecting Fusarium mycotoxin contamination.Climate factors not only influence the growth and development of crops and Fusarium fungi but also affect the infection process of Fusarium fungi through changes in crop susceptibility, virulence of Fusarium fungi, and environmental factors at critical stages of infection.

Figure 7 .
Figure 7. Proper practices to minimize Fusarium mycotoxin contamination.There are a series of management strategies that can be utilized to control Fusarium mycotoxins during the preharvest (sowing and growing) and postharvest stages (storing and processing).During the preharvest stage, these strategies place emphasis on preventing infection by Fusarium fungi.During the postharvest stage, management strategies are more focused on controlling the production of new mycotoxins and the detoxification and removal of existing mycotoxins. 17

Table 1 .
Function of Fusarium mycotoxin biosynthesis genes and phenotypes of their mutants.

Table 2 .
The optimal temperature and a w of Fusarium mycotoxin production.

Table 3 .
The conditions for predicting the impact of CC on Fusarium mycotoxin contamination.

Table 4 .
The detection methods of main Fusarium mycotoxins.