The European corn borer (ECB) plays an important role in promoting Fusarium verticillioides infections and in the consequent fumonisin contamination in maize grain in temperate areas. The objective of this study was to evaluate whether the ECB feeding activity could also affect the occurrence of emerging mycotoxins in maize kernels. During the 2008–10 period, natural infestation of the insect was compared, in field research, with the protection of infestation, which was obtained by using an entomological net. The ears collected in the protected plots were free from ECB attack, while those subject to natural insect attacks showed a damage severity that varied from 10% to 25%. The maize samples were analysed by means of an LC-MS/MS-based multi-mycotoxin method, which led to the detection of various metabolites: fumonisins (FUMs), fusaproliferin (FUS), moniliformin (MON), bikaverin (BIK), beauvericin (BEA), fusaric acid (FA), equisetin (EQU), deoxynivalenol (DON), deoxynivalenol-3-glucoside (DON-3-G), zearalenone (ZEA), culmorin (CULM), aurofusarin (AUR) and butenolide (BUT). The occurrence of mycotoxins produced by Fusarium spp. of Liseola section was affected significantly by the ECB feeding activity. The presence of ECB injuries increased the FUMs from 995 to 4694 µg kg^–1, FUS from 17 to 1089 µg kg^–1, MON from 22 to 673 µg kg^–1, BIK from 58 to 377 µg kg^–1, BEA from 6 to 177 µg kg^–1, and FA from 21 to 379 µg kg^–1. EQU, produced by F. equiseti section Gibbosum, was also increased by the ECB activity, by 1–30 µg kg^–1 on average. Instead, the content of mycotoxins produced by Fusarium spp. of Discolor and Roseum sections was not significantly affected by ECB activity. As for FUMs, the application of a strategy that can reduce ECB damage could also be the most effective solution to minimise the other mycotoxins produced by Fusarium spp. of Liseola section.

Azole fungicides have been reported to be the most effective active substances in the control ofFusarium Head Blight (FHB) and in the reduction of the main mycotoxins that occur in cereal grain, such as deoxynivalenol (DON). Four field experiments have been conducted in North West Italy, over a period of 2 growing seasons, in order to evaluate the effect of azole fungicide (prothioconazole) applications on the prevalence of emerging mycotoxins in common winter wheat under naturally-infected field conditions. Wheat samples have been analysed by means of a dilute-and-shoot multi-mycotoxin LC-MS/MS method. Twenty fungal metabolites were detected: enniatins, aurofusarin, moniliformin, equisetin, DON, deoxynivalenol-3-glucoside, culmorin, bikaverin, beauvericin, fumonisins, fusaric acid, 3-acetyldeoxynivalenol, 15-acetyldeoxynivalenol, nivalenol, zearalenone, decalonectrin, butenolide, tentoxin, alternariol and alternariol methyl ether. The most abundant fungal metabolites were DON and culmorin, with an average contamination in the untreated control of 1,360 μg/kg and 875 μg/kg, respectively, in the growing season with the highest disease pressure (2011-2012). On average, the results have shown that the fungicide application significantly reduced the enniatins (from 127 μg/kg to 46 μg/kg), aurofusarin (from 62 μg/kg to 21 μg/kg), moniliformin (from 32 μg/kg to 16 μg/kg), tentoxin (from 5.2 μg/kg to 2.5 μg/kg) and equisetin (from 0.72 μg/kg to 0.06 μg/kg) contents in all the experiments. However, DON, deoxynivalenol-3-glucoside and culmorin were only significantly reduced in the growing season with the highest disease pressure. The other fungal metabolites were mainly found in traces in the untreated plots. These results, which have been obtained in different environmental and agronomic conditions, have underlined for the first time that the fungicide usually applied to control the FHB and DON content, also consistently reduces the main emerging mycotoxins of winter wheat in temperate areas.

Maize is considered one of the most susceptible crops to mycotoxin-producing fungi throughout the world, mainly belonging to the Fusarium spp. and Aspergillus spp. Maize is mainly used as animal feeds in Italy, as well as for human consumption, being essential for all the protected designation of origin (DOP) products. Our study investigated the occurrence of regulated mycotoxins in 3769 maize grain samples collected from 88 storage centers by the National Monitoring Network over an 11-year period (2011–2021). Moreover, an in-depth survey over a 4-year period, characterized by extremely different meteorological conditions, was conducted to investigate the co-occurrence of regulated, masked, and emerging mycotoxins. The survey confirmed that Fusarium spp. was the most frequent fungi and fumonisins were the main mycotoxins that were constantly detected in the different years and areas. Moreover, the areas characterized by high fumonisin levels were also the most prone to contamination by emerging mycotoxins produced by the same Fusarium species of the Liseola section. On the other hand, as a result of climatic changes, maize grains have also been affected by the increased frequency of aflatoxin accumulation. Deoxynivalenol, zearalenone, and other emerging mycotoxins produced by the same Fusarium species as the Discolor section occurred more abundantly in some areas in Northern Italy and in years characterized by predisposing meteorological conditions.

Cereals can be contaminated by several mycotoxins, whose co-presence may represent an undervalued risk for humans and animals. Maize and wheat are the most contaminated cereals and in temperate areas could be affected in field conditions by several Fusarium and Aspergillus infections. To date, only B-fumonisins (FBs), aflatoxins (AFs), zearalenone (ZEA), deoxynivalenol (DON) and T-2 and HT-2 toxins have been regulated in cereals in European Union. The other fungal metabolites, are commonly referred to as “emerging” and “masked” mycotoxins, and more information on their occurrence in combination with the regulated mycotoxins, are needed to design combined toxicological and exposure assessments. This research intends to develop and compare two multiresidue HPLC-ESI-TQ-MS/MS methods for the simultaneous determination of the main regulated, emerging and masked mycotoxins in maize and wheat, among which: FB₁, FB₂, DON, ZEA, AFB₁, AFB₂, AFG₁, AFG₂, moniliformin (MON), deoxynivalenol-3-glucoside (DON-3-G), 3-acetyldeoxynivalenol (3-ADON), 15-acetyldeoxynivalenol (15-ADON), nivalenol (NIV), enniatins A, A₁, B, B₁ (ENNA, ENNA₁, ENNB, ENNB₁). The extraction was performed for both methods using a mixture of CH₃CN/H₂O/CH₃COOH (79/20/1, v/v/v), while the dilution/purification was carried out through two different procedures: (1) by the “dilute-and-shoot” technique diluting 1:2 the filtered extract with CH₃CN/H₂O/CH₃COOH (20/79/1, v/v/v) to reduce the matrix effect; (2) using the Oasis^® PRiME HLB clean-up columns. The analysis was carried out using CH₃OH and H₂O both acidified with 0.1% of CH₃COOH as eluents. The injection volume was 20 μL and the flow rate 200 μL min^-1. The analysis of two reference material (maize and wheat), was performed to evaluate the trueness and precision of the two methods by matrix-matched calibration curves. For all the regulated mycotoxins analyzed by both methods, the range of recovery percentage established by the Regulation (EC) No. 401/2006 was respected, except for ZEA by using the Oasis^® PRiME HLB clean-up columns. Nevertheless, the results suggest that the Oasis^® PRiME HLB clean-up columns, could be a valid alternative to the dilute-and-shoot method, although an additional cost for the clean-up has to be considered. In conclusion, both two analytical methods considerably reduce the analytical time and costs and therefore result to be promising and applicable for high-throughput routine multi-mycotoxins analysis by the use of a TQ.