Toxicity Potential of Nutraceuticals.

During the past few decades and especially during and after the COVID-19 pandemic, the use of nutraceuticals has become increasingly popular in both humans and animals due to their easy access, cost-effectiveness, and tolerability with a wide margin of safety. While some nutraceuticals are safe, others have an inherent toxic potential. For a large number of nutraceuticals, no toxicity/safety data are available due to a lack of pharmacological/toxicological studies. The safety of some nutraceuticals can be compromised via contamination with toxic plants, metals, mycotoxins, pesticides, fertilizers, drugs of abuse, etc. Knowledge of pharmacokinetic/toxicokinetic studies and biomarkers of exposure, effect, and susceptibility appears to play a pivotal role in the safety and toxicity assessment of nutraceuticals. Interaction studies are essential to determine efficacy, safety, and toxicity when nutraceuticals and therapeutic drugs are used concomitantly or when polypharmacy is involved. This chapter describes various aspects of nutraceuticals, particularly their toxic potential, and the factors that influence their safety.

Discovery of the Alternaria mycotoxins alterperylenol and altertoxin I as novel immunosuppressive and antiestrogenic compounds in vitro.

Alternaria mycotoxins may pose significant challenges to food safety and public health due to the wide spectrum of reported adverse effects. Despite this, critical information on the immunomodulatory and antiestrogenic properties of most of these contaminants is still lacking. The present study aimed to identify the mycotoxins responsible for the immunosuppressive and antiestrogenic effects of a complex extract of Alternaria mycotoxins (CE) obtained by growing an Alternaria alternata strain on rice. Through a toxicity-guided fractionation procedure involving the production of CE-fractions by supercritical fluid chromatography and mycotoxin quantification by LC-MS/MS, the mycotoxins alternariol (AOH), tenuazonic acid (TeA), altertoxin I (ATX-I), and alterperylenol (ALTP) were identified as potential toxicologically relevant constituents contributing to the in vitro effects exerted by the extract. The assessment of the immunomodulatory effects, performed by applying the NF-κB reporter gene assay in THP1-Lucia™ monocytes, revealed the scarce contribution of AOH to the effects exerted by the CE. TeA showed no effect on the NF-κB pathway up to 250 µM, whereas ATX-I and ALTP suppressed the LPS-mediated pathway activation at concentrations ≥ 1 µM. The evaluation of antiestrogenic effects, performed in Ishikawa cells by applying the alkaline phosphatase assay, revealed the ability of ALTP (≥ 0.4 µM) and ATX-I (≥ 2 µM) to suppress the estrogen-dependent expression of enzyme activity. Given the risk of detrimental impacts stemming from alterations in endocrine and systemic immune responses by the investigated mycotoxins, further studies are needed to elucidate their underlying mechanisms of action and comprehensively evaluate the health risks posed by these toxins.

Fumonisins in infant cereals marketed as complementary food in Argentina.

Infant cereals are typically the first foods introduced as complementary foods. Cereals used to elaborate complementary foods, such as wheat, maize and rice, are susceptible to mycotoxin contamination. Among mycotoxins, fumonisins have been epidemiologically associated, in humans, with oesophageal cancer, neural tube defects and stunting. Fumonisins have been found in maize and wheat grains in Argentina. In the present study, a survey was conducted to determine their occurrence in 82 wheat-based and multicereal-based infant cereal items collected from retail stores in Rio Cuarto, Argentina, using HPLC-MS. Of these samples, 84% showed FBs contamination with levels ranging from 0.05 to 992 µg/kg). Although FB1 was the most prevalent fumonisin, FB2 was found at higher levels. Most samples had levels below the limit of 200 µg/kg set for Argentinean cereal products for children. The outcome of this survey provides information on the naturally presence of fumonisin in infant cereal intended for children in Argentina, which can be helpful to consider relevant monitoring programmes.

Evaluation of mycotoxins, mycobiota and toxigenic fungi in the traditional medicine Radix Dipsaci.

Medicinal herbs have been increasingly used for therapeutic purposes against a diverse range of human diseases worldwide. However, inevitable contaminants, including mycotoxins, in medicinal herbs can cause serious problems for humans despite their health benefits. The increasing consumption of medicinal plants has made their use a public health problem due to the lack of effective surveillance of the use, efficacy, toxicity, and quality of these natural products. Radix Dipsaci is commonly utilized in traditional Chinese medicine and is susceptible to contamination with mycotoxins. Here, we evaluated the mycotoxins, mycobiota and toxigenic fungi in the traditional medicine Radix Dipsaci. A total of 28 out of 63 Radix Dipsaci sample batches (44.4%) were found to contain mycotoxins. Among the positive samples, the contamination levels of AFB1, AFG1, AFG2, and OTA in the positive samples ranged from 0.52 to 32.13 µg/kg, 5.14 to 20.05 µg/kg, 1.52 to 2.33 µg/kg, and 1.81 to 19.43 µg/kg respectively, while the concentrations of ZEN and T-2 were found to range from 2.85 to 6.33 µg/kg and from 2.03 to 2.53 µg/kg, respectively. More than 60% of the contaminated samples were combined with multiple mycotoxins. Fungal diversity and community were altered in the Radix Dipsaci contaminated with various mycotoxins. The abundance of Aspergillus and Fusarium increased in the Radix Dipsaci contaminated with aflatoxins (AFs) and ZEN. A total of 95 strains of potentially toxigenic fungi were isolated from the Radix Dipsaci samples contaminated with mycotoxins, predominantly comprising Aspergillus (73.7%), Fusarium (20.0%), and Penicillium (6.3%). Through morphological identification, molecular identification, mycotoxin synthase gene identification and toxin production verification, we confirmed that AFB1 and AFG1 primarily derive from Aspergillus flavus, OTA primarily derives from Aspergillus westerdijkiae, ZEN primarily derives from Fusarium oxysporum, and T-2 primarily derives from Fusarium graminearum in Radix Dipsaci. These data can facilitate our comprehension of prevalent toxigenic fungal species and contamination levels in Chinese herbal medicine, thereby aiding the establishment of effective strategies for prevention, control, and degradation to mitigate the presence of fungi and mycotoxins in Chinese herbal medicine.

Antifungal activity of poly(lactic acid) nanofibers containing the essential oil from Corymbia citriodora Hook or the monoterpenes ß-citronellol and citronellal against mycotoxigenic fungi.

Food contamination by mycotoxigenic fungi is one of the principal factors that cause food loss and economic losses in the food industry. The objective of this work was to incorporate the essential oil from Corymbia citriodora Hook and its constituents citronellal and ß-citronellol into poly(lactic acid) nanofibers; to characterize the nanofibers by scanning electron microscopy (SEM), Fourier transform infrared (FTIR) spectroscopy and differential scanning calorimetry (DSC); to evaluate the antifungal activity by the fumigation method; to evaluate the antimycotoxigenic activity against Aspergillus carbonarius, Aspergillus ochraceus, Aspergillus westerdijkiae, Aspergillus flavus and Aspergillus parasiticus; and to evaluate the morphology of these microorganisms. All the nanofibers had a regular, smooth and continuous morphology. FTIR analyses confirmed that the active ingredients were incorporated into the polymer matrix. All samples exhibited antifungal and ochratoxigenic inhibitory activities of up to 100% and 99%, respectively, with the best results observed for (PLA + 30 wt% ß-citronellol) nanofibers and (PLA + 30 wt% citronellal) nanofibers. However, 100% inhibition of the production of aflatoxin B1 and B2 was not observed. The images obtained by SEM indicated that the nanofibers caused damage to the hyphae, caused a decrease in the production of spores, and caused deformation, rupture and non-formation of the conid head, might be an alternative for the control of mycotoxigenic fungi.

Low molecular weight acids differentially impact Fusarium verticillioides transcription.

Fusarium verticillioides is both an endophyte and pathogen of maize. During growth on maize, the fungus often synthesizes the mycotoxins fumonisins, which have been linked to a variety of diseases, including cancer in some animals. How F. verticillioides responds to other fungi, such as Fusarium proliferatum, Aspergillus flavus, Aspergillus niger, and Penicillium oxalicum, that coinfect maize, has potential to impact mycotoxin synthesis and disease. We hypothesize that low molecular weight acids produced by these fungi play a role in communication between the fungi in planta/nature. To address this hypothesis, we exposed 48-h maize kernel cultures of F. verticillioides to oxalic acid, citric acid, fusaric acid, or kojic acid and then compared transcriptomes after 30 min and 6 h. Transcription of some genes were affected by multiple chemicals and others were affected by only one chemical. The most significant positive response was observed after exposure to fusaric acid which resulted in >2-fold upregulation of 225 genes, including genes involved in fusaric acid synthesis. Exposure of cultures to the other three chemicals increased expression of only 3-15 genes. The predicted function and frequent co-localization of three sets of genes support a role in protecting the fungus from the chemical or a role in catabolism. These unique transcriptional responses support our hypothesis that these chemicals can act as signaling molecules. Studies with gene deletion mutants will further indicate if the initial transcriptional response to the chemicals benefit F. verticillioides.

Ochratoxin A status at birth is associated with reduced birth weight and ponderal index in rural Burkina Faso.

BACKGROUND: Mycotoxin exposure during pregnancy has been associated with adverse birth outcomes and poor infant growth. We assessed multiple biomarkers and metabolites of exposure to mycotoxins at birth and their associations with birth outcomes and infant growth in 274 newborns in rural Burkina Faso. METHODS AND FINDINGS: Whole blood microsamples were analyzed for mycotoxin concentrations in newborns in the Biospecimen sub-study nested in the MISAME-III trial using ultra performance liquid chromatography coupled to tandem mass spectrometry. Unadjusted and adjusted associations between mycotoxin exposure, and birth outcomes and infant growth at 6 months were estimated using linear regression models for continuous outcomes and linear probability models with robust variance estimation for binary outcomes. Infant growth trajectories from birth to 6 months were compared by exposure status using mixed-effects models with random intercept for the individual infant and random slope for the infant's age. Ochratoxin A (OTA) exposure was detected in 38.3% of newborns, with other mycotoxins being detected in the range of 0.36% and 4.01%. OTA exposure was significantly associated with adverse birth outcomes, such as lower birth weight (ß (95% CI): -0.11 kg (-0.21, 0.00); p = 0.042) and ponderal index (ß (95% CI): -0.62 gm/cm3 (-1.19, -0.05); p = 0.034), and a marginally significant lower length growth trajectories during the first 6 months (ß (95% CI): -0.08 cm/mo (-0.15, 0.0); p = 0.057). CONCLUSIONS: OTA exposure was prevalent among newborns and also associated with lower growth at birth and during the first 6 months. The results emphasize the importance of nutrition-sensitive strategies to mitigate dietary OTA, as well as adopting food safety measures in Burkina Faso during the fetal period of development.

Effect of zearalenone in sugar beet products on zootechnical and reproductive performance and lesions of sows and piglets.

Following the use of sugar beet pulp that was retrospectively found to be predominantly contaminated with zearalenone (ZEN) in diets of reproducing sows largely exceeding the EU-guidance value for critical ZEN concentration of 0.25 mg/kg, farmers did not report any changes in the reproductive performance of sows. Thus, the aim of the study was to verify this guidance value in a dose-response setup by using sugar beet pulp as a ZEN source hitherto not considered a risky feedstuff additionally characterized by comparatively low levels of deoxynivalenol. A total of 90 sows was equally allocated to one of the three feed groups during experimental lactation 1 and up to 40 days after insemination: CON with a minimal ZEN concentration, ZEN1 with a target concentration of 250 µg ZEN/kg feed, and ZEN2 with a target concentration of 500 µg ZEN/kg feed. Thereafter, all sows received the same feed without ZEN for the rest of gestation, and the following lactation for testing of putative carry-over effects resulting from previous ZEN exposure. Exposure of sows to ZEN with blood serum as an indicator was linearly related to dietary ZEN concentrations. Reproductive and zootechnical performances of sows were only affected by ZEN exposure at weaning weight. Clinical-chemical parameters indicated no clear effect of ZEN exposure. An influence of ZEN on the occurrence of tail and ear injuries (not necrosis) in piglets and lesions on the mammary complexes in sows is possible. The influence of a ZEN concentration above the EU guidance value on the study farm can therefore not be neglected.

Multiple mycotoxin exposure assessment through human biomonitoring in an esophageal cancer case-control study in the Arsi-Bale districts of Oromia region of Ethiopia.

BACKGROUND: Esophageal cancer (EC) is a malignancy with a poor prognosis and a five-year survival rate of less than 20%. It is the ninth most frequent cancer globally and the sixth leading cause of cancer-related deaths. The incidence of EC has been found to vary significantly by geography, indicating the importance of environmental and lifestyle factors along with genetic factors in the onset of the disease. In this work, we investigated mycotoxin exposure in a case-control study from the Arsi-Bale districts of Oromia regional state in Ethiopia, where there is a high incidence of EC while alcohol and tobacco use - two established risk factors for EC - are very rare. METHODS: Internal exposure to 39 mycotoxins and metabolites was assessed by liquid chromatography-tandem mass spectrometry in plasma samples of EC cases (n = 166) and location-matched healthy controls (n = 166) who shared similar dietary sources. Demographic and lifestyle data were collected using structured questionnaires. Principal Component Analysis and machine learning models were used to identify the most relevant demographic, lifestyle, and mycotoxin (co-)exposure variables associated with EC. Multivariate binary logistic regression analysis was used to assess EC risk. RESULT: Evidence of mycotoxin exposure was observed in all plasma samples, with 10 different mycotoxins being detected in samples from EC cases, while only 6 different mycotoxins were detected in samples from healthy controls. Ochratoxin A was detected in plasma from all cases and controls, while tenuazonic acid was detected in plasma of 145 (87.3%) cases and 71 (42.8%) controls. Using multivariable logistic regression analysis, exposure to tenuazonic acid (AOR = 1.88 [95% CI: 1.68-2.11]) and to multiple mycotoxins (AOR = 2.54 [95% CI: 2.10-3.07]) were positively associated with EC. CONCLUSION: All cases and controls were exposed to at least one mycotoxin. Cases were exposed to a statistically significantly higher number of mycotoxins than controls. Exposure to tenuazonic acid and to multiple mycotoxins were associated with increased risk of EC in the study population. Although aflatoxin B1-lysine and the ratio of sphinganine to sphingosine (as a biomarker of effect to fumonisin exposure) were not assessed in this study, our result emphasizes the need to characterize the effect of mycotoxin co-exposure as part of the exposome and include it in risk assessment, since the current mycotoxin safety levels do not consider the additive or synergistic effects of mycotoxin co-exposure. Moreover, a prospective study design with regular sampling should be considered in this high incidence area of EC in Ethiopia to obtain conclusive results on the role of mycotoxin exposure in the onset and development of the disease.

Mutual interaction of the entomopathogenic and endophytic fungus Metarhizium anisopliae with zearalenone as a native component of crude Fusarium extract.

The present study revealed the consequences of the interaction of a widely used bioinsecticide and endophyte Metarhizium anisopliae with the hazardous mycotoxin zearalenone (ZEN) as a pure substance and with ZEN as a native component of a crude Fusarium extract. In the environment, microorganisms encounter a mixture of metabolites secreted by other organisms living in the same area, not single substances. The obtained results suggest that M. anisopliae, exposed to a variety of active substances produced by Fusarium graminearum, is able to eliminate ZEN. Within 14 days, M. anisopliae biotransformed 90.8% and 85.8% of ZEN as a pure substance and ZEN as a native component of the F. graminearum extract from Rice Medium (E-Fg-RM), respectively, through reduction predominantly to α-epimers of zearalenols and zearalanols, considered more estrogenic than ZEN, which can raise concerns. Compared to pure ZEN, E-Fg-RM significantly affected the production of Metarhizium secondary metabolites by increasing the destruxins amount by approximately 20-25% and reducing the swainsonine content by 96.2%. All these findings provide a possible picture of the interaction of M. anisopliae with ZEN in the wild, mainly as a result of the use of crude extract from Fusarium, which contained a mixture of different metabolites.

Identification and Differentiation of the Fusarium graminearum NX-2 Chemotype Using High-Resolution Melting (HRM).

Fusarium head blight causes significant yield losses in wheat and other cereals and contaminates grain products with trichothecene mycotoxins. F. graminearum isolates are classified into different chemotypes depending on the type of mycotoxin produced, including the type B trichothecenes 3-acetyl deoxynivalenol (3-ADON), 15-acetyl deoxynivalenol (15-ADON), nivalenol (NIV), and the recently identified type A trichothecene NX-2. Molecular tools to differentiate NX-2 producers from other chemotypes have remained relatively laborious and time consuming. In this study, we developed and validated a high-resolution melting (HRM) assay that can identify NX-2 producers quickly and cost-effectively. By analyzing TRI1 coding sequences from 183 geographically diverse isolates representing all four F. graminearum chemotypes, we selected a 75-base pair region containing four non-synonymous single nucleotide polymorphisms (SNPs) that are specific to the NX-2 genotypes. The amplicon generated two HRM profiles, one of which was specific for only NX-2. We confirmed that the assay is robust across qPCR platforms and unambiguously differentiates NX-2 from other chemotypes using a panel of 72 diverse isolates previously collected from North America. The HRM assay was also successful in identifying NX-2 producers directly from DNA extracted from infected wheat spikes with varying levels of disease severity and fungal DNA. The assay can detect as little as 0.01 ng of fungal DNA in a background of 50 ng of plant DNA. This new diagnostic assay can be used for high-throughput molecular detection of the NX-2 chemotype of F. graminearum from infected plant samples and culture collections, thus making it a valuable tool for surveys of contemporary and historical FHB pathogen populations.

Biosensing strategies using recombinant luminescent proteins and their use for food and environmental analysis.

Progress in synthetic biology and nanotechnology plays at present a major role in the fabrication of sophisticated and miniaturized analytical devices that provide the means to tackle the need for new tools and methods for environmental and food safety. Significant research efforts have led to biosensing experiments experiencing a remarkable growth with the development and application of recombinant luminescent proteins (RLPs) being at the core of this boost. Integrating RLPs into biosensors has resulted in highly versatile detection platforms. These platforms include luminescent enzyme-linked immunosorbent assays (ELISAs), bioluminescence resonance energy transfer (BRET)-based sensors, and genetically encoded luminescent biosensors. Increased signal-to-noise ratios, rapid response times, and the ability to monitor dynamic biological processes in live cells are advantages inherent to the approaches mentioned above. Furthermore, novel fusion proteins and optimized expression systems to improve their stability, brightness, and spectral properties have enhanced the performance and pertinence of luminescent biosensors in diverse fields. This review highlights recent progress in RLP-based biosensing, showcasing their implementation for monitoring different contaminants commonly found in food and environmental samples. Future perspectives and potential challenges in these two areas of interest are also addressed, providing a comprehensive overview of the current state and a forecast of the biosensing strategies using recombinant luminescent proteins to come.

The Ecological Strategy Determines the Response of Fungi to Stress: A Study of the 2,4-diacetylphloroglucinol Activity Against Aspergillus and Fusarium Species.

Aspergillus and Fusarium are two economically important genera of fungi. They cause significant yield losses and contamination of crops with mycotoxins. In this study we aimed to evaluate the impact of 2,4-diacetylphloroglucinol (2,4-DAPG) on Aspergillus and Fusarium fungi. It is hypothesized that two fungal genera, which have different ecological strategies, react differently to stress caused by a secondary metabolite produced by rhizosphere Pseudomonas species. We found that 2,4-DAPG was able to reduce biofilm formation of Aspergillus and Fusarium, as reflected in biomass and its chemical composition. Furthermore, subinhibitory concentrations of 2,4-DAPG increased the levels of ergosterol and polysaccharides (α- and ß-glucans, chitin) in the cell membrane and cell wall of Aspergillus, while decreasing them in Fusarium. 2,4-DAPG altered the production of secondary metabolites, especially mycotoxins and extracellular proteases. The production of ochratoxin A was decreased in A. ochraceus, and T-2 toxin and zearalenone, on the contrary, were increased in F. culmorum and F. sporotrichioides, respectively. Thus, using 2,4-DAPG we demonstrated that the ecological role of fungi determines their reaction to antibiotic substances produced by the plant microbiome. Our data contributes to understanding the molecular mechanisms behind symbiotic relationships in natural communities, which are mediated by the biosynthesis of antibiotics.

A worldwide systematic review of ochratoxin A in various coffee products - human exposure and health risk assessment.

Coffee is one of the most commonly consumed beverages worldwide, so assessing its quality for potential health risks is essential. Therefore, this review aimed to determine the levels of ochratoxin A (OTA) in coffee worldwide and then estimate its human intake and health risks. The systematic search took place from June 1997 to April 2024 and 40 of 254 articles were selected based on the selection criteria. The results showed significant differences in average levels of OTA between countries, continents and coffee types (p < 0.001). Of 3256 samples, OTA was detected in 1778, accounting for 54.6% of the total, with the percentage of positive results varying between 7.5% and 100%. Only two studies reported OTA average levels in roasted coffee exceeding the maximum limit (ML) set by the European Commission (ML-EC = 5 µg/kg). The average OTA in soluble coffee was lower than ML-EC (10 µg/kg) in all studies, and in instant coffee, the level of OTA was higher than ML-EC (10 µg/kg) only in one study. The estimated daily intake (EDI) of OTA in all coffee types was lower than the provisional tolerable daily intake (PTDI) values set by joint FAO/WHO Expert Committee on Food Additives (JECFA) (14 ng/kg bw/day) and proposed by the European Food Safety Authority (EFSA) (17 ng/kg bw/day). Non-carcinogenic risk assessment through coffee consumption indicated that the hazard quotient (HQ) was below the acceptable level, HQ = 1. The Margin of Exposure (MoE) for neoplastic effects was acceptable and unacceptable for non-neoplastic effects (NNE) in 4.5% (one of 22 cases) of the roasted and soluble coffees, but acceptable for all instant coffees. In conclusion, the study shows that the OTA content of coffee is not toxic to consumers worldwide. However, preventative measures should be taken, including inhibiting fungal growth and reducing OTA-producing fungal growth.

The Mycotoxins T-2 and Deoxynivalenol Facilitate the Translocation of Streptococcus suis across Porcine Ileal Organoid Monolayers.

Mycotoxins have the potential to increase the risk of airway or intestinal infection due to their effects on epithelial integrity and function. The bacterium Streptococcus suis (S. suis) is often carried in pigs and can cause outbreaks of invasive disease, leading to sepsis and meningitis in postweaning piglets. In this study, we tested the effect of two Fusarium mycotoxins (deoxynivalenol (DON) and T-2) on the integrity of the intestinal epithelium and their interaction with S. suis. Porcine ileal organoids were exposed to DON and T-2 individually or in combination and co-cultured with or without S. suis. Both DON and T-2 were toxic for ileal organoid monolayers at a concentration of 1 µM but not S. suis, even at a higher concentration of 4 µM. To mimic sub-clinical exposures on farms, DON was tested at a concentration of 0.1 µM and T-2 at a concentration of 0.01 µM. The mycotoxins alone did not affect cell permeability, but in combination with S. suis there was an increase in epithelial permeability. Furthermore, DON and T-2 together decreased the transepithelial electrical resistance and increased bacterial translocation.

Application and antagonistic mechanisms of atoxigenic Aspergillus strains for the management of fungal plant diseases.

This review covers, for the first time, all methods based on the use of Aspergillus strains as biocontrol agents for the management of plant diseases caused by fungi and oomycetes. Atoxigenic Aspergillus strains have been screened in a variety of hosts, such as peanuts, maize kernels, and legumes, during the preharvest and postharvest stages. These strains have been screened against a wide range of pathogens, such as Fusarium, Phytophthora, and Pythium species, suggesting a broad applicability spectrum. The highest efficacies were generally observed when using non-toxigenic Aspergillus strains for the management of mycotoxin-producing Aspergillus strains. The modes of action included the synthesis of antifungal metabolites, such as kojic acid and volatile organic compounds (VOCs), secretion of hydrolytic enzymes, competition for space and nutrients, and induction of disease resistance. Aspergillus strains degraded Sclerotinia sclerotiorum sclerotia, showing high control efficacy against this pathogen. Collectively, although two Aspergillus strains have been commercialized for aflatoxin degradation, a new application of Aspergillus strains is emerging and needs to be optimized.

Limited Effectiveness of Penicillium camemberti in Preventing the Invasion of Contaminating Molds in Camembert Cheese.

Mold-ripened cheese acquires a distinctive aroma and texture from mold cultures that mature on a fresh cheese wheel. Owing to its high moisture content (aw = 0.95) and pliability, soft cheese is prone to contamination. Many contaminating mold species are unable to grow at colder temperatures, and the lactic acid produced by the cheese bacteria inhibits further infiltration. Thus, Camembert cheese is generally well protected against contamination by a wide range of species. In this study, cocultures of Penicillium camemberti and widely distributed mycotoxin-producing mold species were incubated on different types of agars, and purchased Camembert samples were deliberately contaminated with mycotoxin-producing mold species capable of growing at both 25 °C and 4 °C. The production of mycotoxins was then monitored by the extraction of the metabolites and their subsequent measurement by means of a liquid chromatography-tandem mass spectrometry (LC-MS/MS) based targeted metabolite profiling approach. The production of cyclopiazonic acid (CPA) was highly dependent on the species cocultivated with Penicillium camemberti, the temperature and the substrate. Contamination of Camembert cheese with Penicillium chrysogenum, Mucor hiemalis, or Penicillium glabrum induced CPA production at 25 °C. Although mold growth on cheese was not always evident on biofilms for certain cultures, except for Penicillium citrinum, which stained the monosaccharide agar yellow, mycotoxins were detected in many agar and cheese samples, as in all monosaccharide agar samples. In conclusion, cheese should be immediately discarded upon the first appearance of mold.