scholarly journals Trace analysis of emerging and regulated mycotoxins in infant stool by LC-MS/MS

2021 ◽  
Author(s):  
Magdaléna Krausová ◽  
Kolawole I. Ayeni ◽  
Lukas Wisgrill ◽  
Chibundu N. Ezekiel ◽  
Dominik Braun ◽  
...  
Keyword(s):  
Human Biomonitoring ◽  
Dry Weight ◽  
Monomethyl Ether ◽  
Food Contaminants ◽  
Alternariol Monomethyl Ether ◽  
Negative Effects ◽  
Toxicant Exposure ◽  
Chemical Structures ◽  
Stool Samples ◽  
Human Stool

AbstractInfants are sensitive to negative effects caused by food contaminants such as mycotoxins. To date, analytical methods assessing mycotoxin mixture exposure in infant stool are absent. Herein, we present a novel multi-mycotoxin LC-MS/MS assay capable of detecting 30+ analytes including the regulated mycotoxin classes (aflatoxins, trichothecenes, ochratoxins, zearalenone, citrinin), emerging Alternaria and Fusarium toxins, and several key metabolites. Sample preparation consisted of a ‘dilute, filter, and shoot’ approach. The method was in-house validated and demonstrated that 25 analytes fulfilled all required criteria despite the high diversity of chemical structures included. Extraction recoveries for most of the analytes were in the range of 65–114% with standard deviations below 30% and limits of detection between 0.03 and 11.3 ng/g dry weight. To prove the methods’ applicability, 22 human stool samples from premature Austrian infants (n = 12) and 12-month-old Nigerian infants (n = 10) were analyzed. The majority of the Nigerian samples were contaminated with alternariol monomethyl ether (8/10) and fumonisin B1 (8/10), while fumonisin B2 and citrinin were quantified in some samples. No mycotoxins were detected in any of the Austrian samples. The method can be used for sensitive human biomonitoring (HBM) purposes and to support exposure and, potentially, risk assessment of mycotoxins. Moreover, it allows for investigating potential associations between toxicant exposure and the infants’ developing gut microbiome. Graphical abstract

scholarly journals In vitro interactions of Alternaria mycotoxins, an emerging class of food contaminants, with the gut microbiota: a bidirectional relationship

2021 ◽  
Author(s):  
Francesco Crudo ◽  
Georg Aichinger ◽  
Jovana Mihajlovic ◽  
Elisabeth Varga ◽  
Luca Dellafiora ◽  
...  
Keyword(s):  
Gut Microbiota ◽  
Human Health ◽  
Monomethyl Ether ◽  
Bacterial Strains ◽  
Food Contaminants ◽  
Alternariol Monomethyl Ether ◽  
Human Gut ◽  
Alternaria Mycotoxins ◽  
In Vitro Effects

AbstractThe human gut microbiota plays an important role in the maintenance of human health. Factors able to modify its composition might predispose the host to the development of pathologies. Among the various xenobiotics introduced through the diet, Alternaria mycotoxins are speculated to represent a threat for human health. However, limited data are currently available about the bidirectional relation between gut microbiota and Alternaria mycotoxins. In the present work, we investigated the in vitro effects of different concentrations of a complex extract of Alternaria mycotoxins (CE; containing eleven mycotoxins; e.g. 0.153 µM alternariol and 2.3 µM altersetin, at the maximum CE concentration tested) on human gut bacterial strains, as well as the ability of the latter to metabolize or adsorb these compounds. Results from the minimum inhibitory concentration assay showed the scarce ability of CE to inhibit the growth of the tested strains. However, the growth kinetics of most of the strains were negatively affected by exposure to the various CE concentrations, mainly at the highest dose (50 µg/mL). The CE was also found to antagonize the formation of biofilms, already at concentrations of 0.5 µg/mL. LC–MS/MS data analysis of the mycotoxin concentrations found in bacterial pellets and supernatants after 24 h incubation showed the ability of bacterial strains to adsorb some Alternaria mycotoxins, especially the key toxins alternariol, alternariol monomethyl ether, and altersetin. The tendency of these mycotoxins to accumulate within bacterial pellets, especially in those of Gram-negative strains, was found to be directly related to their lipophilicity.

scholarly journals Exposure to Mycotoxin-Mixtures via Breast Milk: An Ultra-Sensitive LC-MS/MS Biomonitoring Approach

2020 ◽  
Author(s):  
Dominik Braun ◽  
Chibundu N. Ezekiel ◽  
Doris Marko ◽  
Benedikt Warth
Keyword(s):  
Breast Milk ◽  
Matrix Effects ◽  
Biological Fluid ◽  
Monomethyl Ether ◽  
Natural Food ◽  
Published Data ◽  
Food Contaminants ◽  
Alternariol Monomethyl Ether ◽  
Milk Bank ◽  
Three Samples

Exposure to natural food contaminants during infancy may influence health consequences later in life. Hence, breast milk may serve as a vehicle to transport these contaminants, including mycotoxins, from mothers to their infants. Analytical methods mostly focused on single exposures in the past, thus neglecting co-occurrences and mixture effects. Here, we present a highly sensitive multi-biomarker approach by a sophisticated combination of steps during sample preparation including QuEChERS extraction followed by SPE cleanup and utilizing stable isotopes for compensating challenging matrix effects. The assay was validated in-house, reaching limits of detection (LOD) for all 34 analytes in the range of 0.1 to 300 ng/L with satisfying extraction efficiencies (75 - 109%) and stable intermediate precisions (1 - 18%) for most analytes. Compared a similar multi-mycotoxin assay for breast milk, LOD values were decreased by a factor of 2-60x enabling the assessment of chronic low-dose exposures. The new method was applied to a small set of Nigerian breast milk samples (n=3) to compare results with already published data. Concentration levels of samples that were found to be contaminated before could be confirmed. In addition, other mycotoxins were determined in all three samples, for example the newly investigated alternariol monomethyl ether (AME) was found for the first time in this biological fluid at concentrations up to 25 ng/L. Moreover, in a pooled Austrian sample obtained from a milk bank, trace amounts of multiple mycotoxins including AME (1.9 ng/L), beauvericin (5.4 ng/L), enniatin B (4.7 ng/L), enniatin B<sub>1</sub> (

Exposure to Mycotoxin-Mixtures via Breast Milk: An Ultra-Sensitive LC-MS/MS Biomonitoring Approach

2020 ◽  
Author(s):  
Dominik Braun ◽  
Chibundu N. Ezekiel ◽  
Doris Marko ◽  
Benedikt Warth
Keyword(s):  
Breast Milk ◽  
Matrix Effects ◽  
Biological Fluid ◽  
Monomethyl Ether ◽  
Natural Food ◽  
Published Data ◽  
Food Contaminants ◽  
Alternariol Monomethyl Ether ◽  
Safety Standards ◽  
Ultra Trace

Exposure to natural food contaminants during infancy may influence health consequences later in life. Hence, breast milk may serve as a vehicle to transport these contaminants, including mycotoxins, from mothers to their infants. Analytical methods mostly focused on single exposures in the past, thus neglecting co-occurrences and mixture effects. Here, we present a highly sensitive multi-biomarker approach by a sophisticated combination of steps during sample preparation including QuEChERS extraction followed by SPE cleanup and utilizing stable isotopes for compensating challenging matrix effects. The assay was validated in-house, reaching limits of detection (LOD) for all 34 analytes in the range of 0.1 to 300 ng/L with satisfying extraction efficiencies (75 - 109%) and stable intermediate precisions (1 - 18%) for most analytes. Compared to our first published multi-mycotoxin method LODs were decreased by a factor of 2-60x enabling the assessment of chronic low-dose exposures. The new method was applied to a small set of Nigerian breast milk samples to compare results with already published data. Concentration levels of samples determined to be contaminated before could be confirmed. The newly investigated alternariol monomethyl ether (AME) was found for the first time in this biological fluid at concentrations up to 25 ng/L. Moreover, in a pooled Austrian sample trace amounts of multiple mycotoxins confirmed co-occurrence and exposure even in countries with high food safety standards. In conclusion, the method facilitates the determination of mycotoxins at ultra-trace levels in breast milk, enabling the generation of occurrence data necessary for comprehensive co-exposure assessment.<br>

Exposure to Mycotoxin-Mixtures via Breast Milk: An Ultra-Sensitive LC-MS/MS Biomonitoring Approach

2020 ◽  
Author(s):  
Dominik Braun ◽  
Chibundu N. Ezekiel ◽  
Doris Marko ◽  
Benedikt Warth
Keyword(s):  
Breast Milk ◽  
Matrix Effects ◽  
Biological Fluid ◽  
Monomethyl Ether ◽  
Natural Food ◽  
Published Data ◽  
Food Contaminants ◽  
Alternariol Monomethyl Ether ◽  
Safety Standards ◽  
Ultra Trace

Exposure to natural food contaminants during infancy may influence health consequences later in life. Hence, breast milk may serve as a vehicle to transport these contaminants, including mycotoxins, from mothers to their infants. Analytical methods mostly focused on single exposures in the past, thus neglecting co-occurrences and mixture effects. Here, we present a highly sensitive multi-biomarker approach by a sophisticated combination of steps during sample preparation including QuEChERS extraction followed by SPE cleanup and utilizing stable isotopes for compensating challenging matrix effects. The assay was validated in-house, reaching limits of detection (LOD) for all 34 analytes in the range of 0.1 to 300 ng/L with satisfying extraction efficiencies (75 - 109%) and stable intermediate precisions (1 - 18%) for most analytes. Compared to our first published multi-mycotoxin method LODs were decreased by a factor of 2-60x enabling the assessment of chronic low-dose exposures. The new method was applied to a small set of Nigerian breast milk samples to compare results with already published data. Concentration levels of samples determined to be contaminated before could be confirmed. The newly investigated alternariol monomethyl ether (AME) was found for the first time in this biological fluid at concentrations up to 25 ng/L. Moreover, in a pooled Austrian sample trace amounts of multiple mycotoxins confirmed co-occurrence and exposure even in countries with high food safety standards. In conclusion, the method facilitates the determination of mycotoxins at ultra-trace levels in breast milk, enabling the generation of occurrence data necessary for comprehensive co-exposure assessment.<br>

scholarly journals Exposure to Mycotoxin-Mixtures via Breast Milk: An Ultra-Sensitive LC-MS/MS Biomonitoring Approach

2020 ◽  
Author(s):  
Dominik Braun ◽  
Chibundu N. Ezekiel ◽  
Doris Marko ◽  
Benedikt Warth
Keyword(s):  
Breast Milk ◽  
Matrix Effects ◽  
Biological Fluid ◽  
Monomethyl Ether ◽  
Natural Food ◽  
Published Data ◽  
Food Contaminants ◽  
Alternariol Monomethyl Ether ◽  
Milk Bank ◽  
Three Samples

Exposure to natural food contaminants during infancy may influence health consequences later in life. Hence, breast milk may serve as a vehicle to transport these contaminants, including mycotoxins, from mothers to their infants. Analytical methods mostly focused on single exposures in the past, thus neglecting co-occurrences and mixture effects. Here, we present a highly sensitive multi-biomarker approach by a sophisticated combination of steps during sample preparation including QuEChERS extraction followed by SPE cleanup and utilizing stable isotopes for compensating challenging matrix effects. The assay was validated in-house, reaching limits of detection (LOD) for all 34 analytes in the range of 0.1 to 300 ng/L with satisfying extraction efficiencies (75 - 109%) and stable intermediate precisions (1 - 18%) for most analytes. Compared a similar multi-mycotoxin assay for breast milk, LOD values were decreased by a factor of 2-60x enabling the assessment of chronic low-dose exposures. The new method was applied to a small set of Nigerian breast milk samples (n=3) to compare results with already published data. Concentration levels of samples that were found to be contaminated before could be confirmed. In addition, other mycotoxins were determined in all three samples, for example the newly investigated alternariol monomethyl ether (AME) was found for the first time in this biological fluid at concentrations up to 25 ng/L. Moreover, in a pooled Austrian sample obtained from a milk bank, trace amounts of multiple mycotoxins including AME (1.9 ng/L), beauvericin (5.4 ng/L), enniatin B (4.7 ng/L), enniatin B<sub>1</sub> (

scholarly journals Differing Effects of Standard and Harsh Nucleic Acid Extraction Procedures on Diagnostic Helminth Real-Time PCRs Applied to Human Stool Samples

Pathogens ◽  
2021 ◽  
Vol 10 (2) ◽  
pp. 188
Author(s):  
Tanja Hoffmann ◽  
Andreas Hahn ◽  
Jaco J. Verweij ◽  
Gérard Leboulle ◽  
Olfert Landt ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Real Time ◽  
Real Time Pcr ◽  
Acid Extraction ◽  
Nucleic Acid Extraction ◽  
Pcr Assay ◽  
Bead Beating ◽  
Stool Samples ◽  
Pcr Assays ◽  
Human Stool

This study aimed to assess standard and harsher nucleic acid extraction schemes for diagnostic helminth real-time PCR approaches from stool samples. A standard procedure for nucleic acid extraction from stool and a procedure including bead-beating as well as proteinase K digestion were compared with group-, genus-, and species-specific real-time PCR assays targeting helminths and nonhelminth pathogens in human stool samples. From 25 different in-house and commercial helminth real-time PCR assays applied to 77 stool samples comprising 67 historic samples and 10 external quality assessment scheme samples positively tested for helminths, higher numbers of positive test results were observed after bead-beating-based nucleic acid extraction for 5/25 (20%) real-time PCR assays irrespective of specificity issues. Lower cycle threshold values were observed for one real-time PCR assay after the standard extraction scheme, and for four assays after the bead-beating-based scheme. Agreement between real-time PCR results after both nucleic acid extraction strategies according to Cohen’s kappa ranged from poor to almost perfect for the different assays. Varying agreement was observed in eight nonhelminth real-time PCR assays applied to 67 historic stool samples. The study indicates highly variable effects of harsh nucleic acid extraction approaches depending on the real-time PCR assay used.

scholarly journals Characterization of Alternaria Species Associated with Heart Rot of Pomegranate Fruit

2021 ◽  
Vol 7 (3) ◽  
pp. 172
Author(s):  
Francesco Aloi ◽  
Mario Riolo ◽  
Simona Marianna Sanzani ◽  
Annamaria Mincuzzi ◽  
Antonio Ippolito ◽  
...  
Keyword(s):  
Elongation Factor ◽  
Monomethyl Ether ◽  
Alternariol Monomethyl Ether ◽  
Translation Elongation ◽  
Microscopic Features ◽  
Alternaria Species ◽  
Rot Disease ◽  
Multigene Phylogenetic Analysis ◽  
Pomegranate Fruit

This study was aimed at identifying Alternaria species associated with heart rot disease of pomegranate fruit in southern Italy and characterizing their mycotoxigenic profile. A total of 42 Alternaria isolates were characterized. They were obtained from pomegranate fruits with symptoms of heart rot sampled in Apulia and Sicily and grouped into six distinct morphotypes based on macro- and microscopic features. According to multigene phylogenetic analysis, including internal transcribed spacer (ITS), translation elongation factor 1-α (EF-1α), glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and a SCAR marker (OPA10-2), 38 isolates of morphotypes 1 to 5 were identified as Alternaria alternata, while isolates of morphotype 6, all from Sicily, clustered within the Alternaria arborescens species complex. In particular, isolates of morphotype 1, the most numerous, clustered with the ex-type isolate of A. alternata, proving to belong to A. alternata. No difference in pathogenicity on pomegranate fruits was found between isolates of A. alternata and A. arborescens and among A. alternata isolates of different morphotypes. The toxigenic profile of isolates varied greatly: in vitro, all 42 isolates produced tenuazonic acid and most of them other mycotoxins, including alternariol, alternariol monomethyl ether, altenuene and tentoxin.

scholarly journals Antimycotoxigenic Activity of Beetroot Extracts against Altenaria alternata Mycotoxins on Potato Crop

2021 ◽  
Vol 11 (9) ◽  
pp. 4239
Author(s):  
Nesrine H. Youssef ◽  
Sameer H. Qari ◽  
Said I. Behiry ◽  
Eldessoky S. Dessoky ◽  
Ehab I. El-Hallous ◽  
...  
Keyword(s):  
Potato Crop ◽  
Reduction Rate ◽  
Acid Methyl Ester ◽  
Ethanol Extract ◽  
Monomethyl Ether ◽  
Morphological Identification ◽  
Hexadecanoic Acid ◽  
Alternariol Monomethyl Ether ◽  
Alternaria Species ◽  
Main Components

Alternaria species, mainly air-borne fungi, affect potato plants, causing black spots symptoms. Morphological identification, pathogenicity assessment, and internal transcribed spacer (ITS) molecular identification confirmed that all isolates were Alternaria alternata. The annotated sequences were deposited in GenBank under accession numbers MN592771–MN592777. HPLC analysis revealed that the fungal isolates KH3 (133,200 ng/g) and NO3 (212,000 ng/g) produced higher levels of tenuazonic acid (TeA) and alternariol monomethyl ether (AME), respectively. Beet ethanol extract (BEE) and beet methanol extract (BME) at different concentrations were used as antimycotoxins. BME decreased the production of mycotoxins by 66.99–99.79%. The highest TeA reduction rate (99.39%) was reported in the KH3 isolate with 150 µg/mL BME treatment. In comparison, the most effective AME reduction rate (99.79%) was shown in the NO3 isolate with 150 µg/mL BME treatment. In the same way, BEE application resulted in 95.60–99.91% mycotoxin reduction. The highest TeA reduction rate (99.91%) was reported in the KH3 isolate with 150 µg/mL BEE treatment, while the greatest AME reduction rate (99.68%) was shown in the Alam1 isolate with 75 µg/mL BEE treatment. GC-MS analysis showed that the main constituent in BME was the antioxidant compound 1-dodecanamine, n,n-dimethyl with a peak area of 43.75%. In contrast, oxirane, methyl- (23.22%); hexadecanoic acid, methyl ester (10.72%); and n-hexadecanoic acid (7.32%) were the main components in BEE found by GC-MS. They are probably antimicrobial molecules and have an effect on the mycotoxin in general. To our knowledge, this is the first study describing the antimycotoxigenic activity of beet extracts against A. alternata mycotoxins-contaminated potato crops in Egypt, aimed to manage and save the environment.

scholarly journals Multicenter Evaluation of an ELISA for the Detection of Cryptosporidium spp. Antigen in Clinical Human Stool Samples

2021 ◽  
Vol 9 (2) ◽  
pp. 209
Author(s):  
Romy Razakandrainibe ◽  
Célia Mérat ◽  
Nathalie Kapel ◽  
Marc Sautour ◽  
Karine Guyot ◽  
...  
Keyword(s):  
Large Scale ◽  
Cryptosporidium Oocyst ◽  
Clinical Importance ◽  
Epidemiological Studies ◽  
Elisa Test ◽  
Enzyme Linked Immunosorbent Assay ◽  
Cryptosporidium Spp ◽  
Stool Samples ◽  
Conventional Microscopy ◽  
Human Stool

Human cryptosporidiosis remains underdiagnosed, and rapid/accurate diagnosis is of clinical importance. Diagnosis of the Cryptosporidium oocyst in stool samples by conventional microscopy is labor-intensive, time-consuming, and requires skillful experience. Thus, we aimed to evaluate the usefulness of a coproantigen enzyme-linked immunosorbent assay (ELISA) test in detecting Cryptosporidium spp. from fecal specimens. For this aim, we evaluated the performances of a commercial ELISA (CoproELISA Cryptosporidium kit, Savyon Diagnostics, Israel) for the detection of Cryptosporidium spp. in random clinical stool samples through a multicenter study. The sensitivity and specificity for coproantigen ELISA were 98.86% and 94.32%, respectively. The coproantigen ELISA results indicate that the simple, rapid, reliable, and standardized immunoassay test is sensitive and specific for routine diagnosis, and may be useful for large-scale epidemiological studies of cryptosporidiosis.

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