Aflatoxin M<Subscript>1</Subscript> in human breast milk in southeastern Turkey

This study was performed to determine aflatoxin M₁ (AFM₁) in human breast milk samples collected in ?anl?urfa, located in Southeastern region of Turkey, and to investigate a possible correlation between AFM₁ occurrence (frequency and levels) and sampling seasons. Human breast milk samples collected in December 2014 and in June 2015 from a total of 74 nursing women, both outpatient and inpatient volunteers in hospitals located in ?anl?urfa, Turkey, were analyzed using competitive enzyme-linked immunosorbent assay (ELISA) for the presence of AFM₁. AFM₁ was detected in 66 (89.2%) out of 74 samples at an average concentration of 19.0 ± 13.0 ng/l (min.-max., 9.6–80 ng/l). There was a statistically significant difference between December and June concerning AFM₁ levels (p < 0.05). Further detailed studies will be needed to determine the main sources of aflatoxins in food, to establish protection strategies against maternal and infant exposure to these mycotoxins.     

Mycotoxins in South African foods: a case study on aflatoxin M<Subscript>1</Subscript> in milk

The contamination of cow’s milk at the farm level with aflatoxin M₁ was investigated in South Africa. Samples of feeds, forage, maize and milk were taken at nine dairy farms, and at the same time samples of the processed milk (retail milk) were collected from the respective dairies to which the farms delivered their milk. The feeds were analysed for aflatoxin B₁ and the milk samples for aflatoxin M₁ using high performance liquid chromatography (HPLC) and fluorescence detection. All milk samples from the dairy farms were positive for aflatoxin M₁, ranging from 0.02 μg/l to 1.5 μg/l. Retail milk was also frequently contaminated with AFM₁, at levels of 0.01-3.1 μg/l. High AFB₁ levels in feed materials on the farms supplying the raw milk indicate that various sources account for this contamination frequency in milk.     

Occurrence of aflatoxin M<Subscript>1</Subscript> in commercial pasteurized milk samples in Sari,Mazandaran province,Iran

The frequency and levels of aflatoxin M₁ (AFM₁) in pasteurized milk samples in Sari, located in Mazandaran province, Iran, were determined by enzyme immunoassay. Seventy-six samples of pasteurized milk from different retail stores were randomly collected over four seasons during the year 2015. AFM₁ contamination was detected in all milk samples. The mean concentration of aflatoxin M₁ was 65.8 ng/l, with a range of 11.7–106.6 ng/l. The highest AFM₁ level was detected in milk samples collected during spring. Forty-six (60.53 %) samples had AFM₁ levels that exceeded the maximum acceptable levels (50 ng/l) recommended by the European Union (EU). Comparison of these results with previously published data for AFM₁ in milk in Iran shows that the percentage of samples exceeding the EU maximum level is consistently high over the years, indicating a general problem related to AFB₁ contamination in dairy feedingstuff.     

Aflatoxin M<Subscript>1</Subscript> levels in different marketed milk products in Nairobi,Kenya

Milk is an important source of energy and nutrients, especially for children, and in Kenya, milk consumption is higher than other countries in the region. One major concern with milk is the risks of chemical contaminants, and reports of high levels of aflatoxin M₁ (AFM₁) in milk in Kenya has been causing public health concerns. This study collected marketed milk products every month during 1 year, just as a consumer would purchase them from retailers and traders in a low-income area, and a major supermarket in a middle/high-income area. In total, 291 sampled milk products (raw, pasteurised, UHT milk, yoghurt and lala) were collected and analysed for AFM₁ using a commercial ELISA kit. More than 50% of the samples exceeded 50 ng/kg (the level allowed in the EU), but only three samples exceeded 500 ng/kg (the level allowed in the USA). Geometric mean AFM₁ level was 61.9 ng/kg in the 135 samples from the low-income area while it was 36.1 ng/kg in the 156 from the higher income area (p?<?0.001). The levels varied significantly depending on the time of year, with lowest levels of milk in January. There were also differences between manufacturers and products, with UHT milk having lower levels. There was no difference depending on the price for all dairy products, but when only including milk, higher price was associated with lower levels of AFM₁. In conclusion, this study shows that milk purchased by a consumer is likely to contain AFM₁ above 50 ng/kg, and that further research is needed to find ways to mitigate AFM₁ contamination through working with farmers and milk processors both in the formal and informal sectors.     

A survey of aflatoxin B<Subscript>1</Subscript> and total aflatoxin contamination in baby food,peanut and corn products sold at retail in Indonesia analysed by ELISA and HPLC

Aflatoxin contamination has been well known as a world-wide health-threatening problem in tropical countries including Indonesia. This research was undertaken to determine the degree of aflatoxin contamination in different Indonesian foodstuffs. A preliminary survey was carried out to evaluate the level of total aflatoxin (AfT) and aflatoxin B₁ (AfB₁) contamination of baby foods, peanut products, and corn products, which were purchased from traditional markets and supermarkets in Indonesia during the year 2001-2002. Eighty two peanut products, 12 baby foods products, and 11 corn products from different brands were analysed for AfT and AfB₁ using the Enzyme-Linked Immunosorbent Assay (ELISA) method. The results indicate that, of the brands analysed, 35% of the peanut products were contaminated with aflatoxins at various levels (range 5 to 870 μg/kg). Peanut-chilli sauces had the highest percentage of AfT contamination 9/12 (75%), which was followed by traditional snacks 5/11 (45%), peanut butter 4/11 (40%), flour egg coated peanut 6/16 (37%), and peanut cake 3/10 (30%). Fried peanuts and roasted peanut were found to contain aflatoxin at relatively lower percentages of 9% and 8%, respectively. From the 12 analysed baby food samples, on the other hand, no sample was found to be contaminated with aflatoxins. Two of 11 samples (18%) of corn based products were contaminated with AfT, ranging between 5.8 and 12.4 μg/kg. Additionally, 30 selected samples in different concentration ranges were further analysed to verify the correlation between ELISA and HPLC techniques and results were compared.     

Aflatoxin M<Subscript>1</Subscript> contamination of raw cow’s milk in five regions of Kosovo during 2016

During February and August 2016 (winter and summer season), 192 samples of raw bulk tank milk have been collected from small dairy farms in five different regions of Kosova (Prishtina, Prizren, Peja, Mitrovica, and Gjilan). These samples were analyzed for aflatoxin M₁ (AFM₁) contamination level by ultra-high performance liquid chromatography coupled with electrospray ionization triple quadrupole tandem mass spectrometry (UHPLC–ESI–MS/MS). The limit of quantification for AFM1 was 0.02 μg/L and recovery at 0.1–0.5 μg/L was 90–95% (coefficient of variation 15–25%). A total of 74 milk samples (38%) were found to be contaminated with AFM₁ and 11 samples (5.7%) slightly exceeded the European Union maximum level of 0.05 μg/L. The maximum AFM₁ level was at 0.082 μg/L. There was no relevant difference between summer and winter with regard to AFM₁ frequency and levels. Although the frequency of AFM₁-positive samples varied between regions, from 7% (Prishtina) to 53% (Prizren), only minor regional differences were observed with regard to average and maximum toxin levels. The high percentage of milk samples which were non-compliant with AFM₁ maximum levels indicates that efforts to reduce the contamination levels of aflatoxin B₁ in cows feed in Kosovo are necessary.     

Occurrence and fumonisin B<Subscript>2</Subscript> producing potential of <Emphasis Type="Italic">Aspergillus</Emphasis> section <Emphasis Type="Italic">Nigri</Emphasis> in Brazil nuts

Bertholletia excelsa is the tree that produces Brazil nuts which have vast economic importance in the Amazon region and as an export commodity. The aim of this study was to assess the presence of Aspergillus section Nigri in Brazil nut samples at different stages of its production chain and to verify the toxigenic potential for fumonisin B₂ (FB₂) production of these isolates along with the presence of this mycotoxin in Brazil nut samples. The fungal infection ranged from 0 to 80% at the different stages of the harvest and processing chain and the water activity of the nuts from 0.273 to 0.994. A total of 1052 A. section Nigri strains were isolated from Brazil nuts and 200 strains were tested for their ability to produce FB₂: 41 strains (20.5%) were FB₂ producers with concentrations ranging from 0.09 to 37.25 mg/kg; 2 strains (1%) showed traces of FB₂, less than the detection limit (0.08 mg/kg); and 157 (78.5%) were not FB₂ producers. Although several samples showed high contamination by A. section Nigri, no sample was contaminated by FB₂.     

Solvent-dependent transformation of aflatoxin B<Subscript>1</Subscript> in soil

To date, all studies of aflatoxin B₁ (AFB₁) transformation in soil or in purified mineral systems have identified aflatoxins B₂ (AFB₂) and G₂ (AFG₂) as the primary transformation products. However, identification in these studies was made using thin layer chromatography which has relatively low resolution, and these studies did not identify a viable mechanism by which such transformations would occur. Further, the use of methanol as the solvent delivery vehicle in these studies may have contributed to formation of artifactual transformation products. In this study, we investigated the role of the solvent vehicle in the transformation of AFB₁ in soil. To do this, we spiked soils with AFB₁ dissolved in water (93:7, water/methanol) or methanol and used HPLC-UV and HPLC-MS to identify the transformation products. Contrasting previous published reports, we did not detect AFB₂ or AFG₂. In an aqueous-soil environment, we identified aflatoxin B_2a (AFB_2a) as the single major transformation product. We propose that AFB_2a is formed from hydrolysis of AFB₁ with the soil acting as an acid catalyst. Alternatively, when methanol was used, we identified methoxy aflatoxin species likely formed via acid-catalyzed addition of methanol to AFB₁. These results suggest that where soil moisture is adequate, AFB₁ is hydrolyzed to AFB_2a and that reactive organic solvents should be avoided when replicating natural conditions to study the fate of AFB₁ in soil.     

Characterization of Myelin Sheath F<Subscript>o</Subscript>F<Subscript>1</Subscript>-ATP Synthase and its Regulation by IF<Subscript>1</Subscript>

F_oF₁-ATP synthase is the nanomotor responsible for most of ATP synthesis in the cell. In physiological conditions, it carries out ATP synthesis thanks to a proton gradient generated by the respiratory chain in the inner mitochondrial membrane. We previously reported that isolated myelin vesicles (IMV) contain functional F_oF₁-ATP synthase and respiratory chain complexes and are able to conduct an aerobic metabolism, to support the axonal energy demand. In this study, by biochemical assay, Western Blot (WB) analysis and immunofluorescence microscopy, we characterized the IMV F_oF₁-ATP synthase. ATP synthase activity decreased in the presence of the specific inhibitors (olygomicin, DCCD, FCCP, valynomicin/nigericin) and respiratory chain inhibitors (antimycin A, KCN), suggesting a coupling of oxygen consumption and ATP synthesis. ATPase activity was inhibited in low pH conditions. WB and microscopy analyses of both IMV and optic nerves showed that the Inhibitor of F₁ (IF₁), a small protein that binds the F₁ moiety in low pH when of oxygen supply is impaired, is expressed in myelin sheath. Data are discussed in terms of the role of IF₁ in the prevention of the reversal of ATP synthase in myelin sheath during central nervous system ischemic events. Overall, data are consistent with an energetic role of myelin sheath, and may shed light on the relationship among demyelination and axonal degeneration.     

Aflatoxin B<Subscript>1</Subscript> levels in groundnut products from local markets in Zambia

In Zambia, groundnut products (milled groundnut powder, groundnut kernels) are mostly sold in under-regulated markets. Coupled with the lack of quality enforcement in such markets, consumers may be at risk to aflatoxin exposure. However, the level of aflatoxin contamination in these products is not known. Compared to groundnut kernels, milled groundnut powder obscures visual indicators of aflatoxin contamination in groundnuts such as moldiness, discoloration, insect damage or kernel damage. A survey was therefore conducted from 2012 to 2014, to estimate and compare aflatoxin levels in these products (n = 202), purchased from markets in important groundnut growing districts and in urban areas. Samples of whole groundnut kernels (n = 163) and milled groundnut powder (n = 39) were analysed for aflatoxin B₁ (AFB₁) by competitive enzyme-linked immunosorbent assay (cELISA). Results showed substantial AFB₁ contamination levels in both types of groundnut products with maximum AFB₁ levels of 11,100 μg/kg (groundnut kernels) and 3000 μg/kg (milled groundnut powder). However, paired t test analysis showed that AFB₁ contamination levels in milled groundnut powder were not always significantly higher (P > 0.05) than those in groundnut kernels. Even for products from the same vendor, AFB₁ levels were not consistently higher in milled groundnut powder than in whole groundnut kernels. This suggests that vendors do not systematically sort out whole groundnut kernels of visually poor quality for milling. However, the overall contamination levels of groundnut products with AFB₁ were found to be alarmingly high in all years and locations. Therefore, solutions are needed to reduce aflatoxin levels in such under-regulated markets.     

Production of fumonisins B<Subscript>2</Subscript> and B<Subscript>4</Subscript> in <Emphasis Type="Italic">Tolypocladium</Emphasis> species

Tolypocladium inflatum is known primarily for its production of the cyclosporines that are used as an immunosuppressive drug. However, we report here the production of the carcinogenic fumonisins B₂ and B₄ by this biotechnologically relevant fungal genus. These mycotoxins were detected in 11 strains tested from three species: Tolypocladium inflatum, T. cylindrosporum, and T. geodes. Production of fumonisins by Fusarium spp. and Aspergillus niger is highly medium- and temperature-dependent, so the effect of these parameters on fumonisin production by three T. inflatum strains was studied. Maximum production was achieved on media with high sugar content incubated at 25–30°C. Since these results demonstrate that fumonisin production could be widespread within the genus Tolypocladium, the potential contamination of commercial cyclosporine preparations with fumonisins needs to be investigated.     

Boric acid: a potential chemoprotective agent against aflatoxin b<Subscript>1</Subscript> toxicity in human blood

Aflatoxin B₁ is the most potent pulmonary and hepatic carcinogen. Since the eradication of Aflatoxin B₁ contamination in agricultural products has been difficult, the use of natural or synthetic free radical scavengers could be a potential chemopreventive strategy. Boric acid is the major component of industry and its antioxidant role has recently been reported. The present study assessed, for the first time, the effectiveness of boric acid following exposure to Aflatoxin B₁ on human whole blood cultures. The biochemical characterizations of glutathione and some enzymes have been carried out in erythrocytes. Alterations in malondialdehyde level were determined as an index of oxidative stress. The sister-chromatid exchange and micronucleus tests were performed to assess DNA damages in lymphocytes. Aflatoxin B₁ treatment significantly reduced the activities of antioxidants by increasing malondialdehyde level (30.53 and 51.43%) of blood, whereas, the boric acid led to an increased resistance of DNA to oxidative damage induced by Aflatoxin B₁ in comparison with control values (P < 0.05). In conclusion, the support of boric acid was especially useful in Aflatoxin-toxicated blood. Thus the risk on tissue targeting of Aflatoxin B₁ could be reduced ensuring early recovery from its toxicity.     

Exposure assessment and risk characterization of aflatoxin B<Subscript>1</Subscript> in Malaysia

Aflatoxin B₁ (AFB₁) was detected in 57% of the nuts and nut products marketed in Penang, Malaysia using the liquid chromatography tandem mass spectrometry. The contamination levels ranged from 0.40 to 222 μg/kg and 17 out of 128 samples (13.3%) contained AFB₁ above the European Commission permitted level (2 μg/kg). Estimated dietary exposure of AFB₁ in nuts and nut products were 0.36 ng per kg body weight and day and 8.89 ng per kg body weight and day, representing the low and high-level of exposure, respectively. Dose-response modelling resulted in benchmark dose lower confidence limit (BMDL₁₀) values of 0.305 ng per kg body weight and day, with the best fitted from the log-logistic model. The derived margin of exposure (MoE) values ranged from 34 to 847 suggested that AFB₁ would be of public health concern and might reasonably be considered as a high priority for risk management actions.     

Evaluation of measurement uncertainties of an analytical method for the determination of aflatoxin M<Subscript>1</Subscript> in milk and milk powder

The mycotoxin aflatoxin M₁ (AfM₁) is a serious food safety hazard for which the European Commission has already established a maximum permissible level of 0.05 μg/kg AfM₁ in milk and products thereof. For control analysis laboratories are increasingly asked to submit full uncertainties of their analytical results.The evaluation of measurement uncertainties of an analytical method for the determination of AfM₁ in milk and milk powder on the basis of ‘in-house’ validation data in compliance with the ‘Guide to the Expression of Uncertainty in Measurement (GUM)’ [1] and the ‘EURACHEM Guide’ [2] is described. A similar approach will be used to assess the performance of methods employed by laboratories participating in the certification of reference materials for AfM₁ in milk powder.     

Molecular analysis of <Emphasis Type="Italic">Aspergillus</Emphasis> section <Emphasis Type="Italic">Flavi</Emphasis> isolated from Brazil nuts

Brazil nuts are an important export market in its main producing countries, including Brazil, Bolivia, and Peru. Approximately 30,000 tons of Brazil nuts are harvested each year. However, substantial nut contamination by Aspergillus section Flavi occurs with subsequent production of aflatoxins. In our study, Aspergillus section Flavi were isolated from Brazil nuts (Bertholletia excelsa), and identified by morphological and molecular means. We obtained 241 isolates from nut samples, 41% positive for aflatoxin production. Eighty-one isolates were selected for molecular investigation. Pairwise genetic distances among isolates and phylogenetic relationships were assessed. The following Aspergillus species were identified: A. flavus, A. caelatus, A. nomius, A. tamarii, A. bombycis, and A. arachidicola. Additionally, molecular profiles indicated a high level of nucleotide variation within β-tubulin and calmodulin gene sequences associated with high genetic divergence from RAPD data. Among the 81 isolates analyzed by molecular means, three of them were phylogenetically distinct from all other isolates representing the six species of section Flavi. A putative novel species was identified based on molecular profiles.     

Theoretical study of the novel sandwich compound [Au<Subscript>3</Subscript>Cl<Subscript>3</Subscript>Tr<Subscript>2</Subscript>]<Superscript>2+</Superscript>

A theoretical study of a sandwich compound with a metal monolayer sheet between two aromatic ligands is presented. A full geometry optimization of the [Au₃Cl₃Tr₂]²⁺ (1) compound, which is a triangular gold(I) monolayer sheet capped by chlorines and bounded to two cycloheptatrienyl (Tr) ligands was carried out using perturbation theory at the MP2 computational level and DFT. Compound (1) is in agreement with the 18–electron rule, the bonding nature in the complex may be interpreted from the donation interaction coming from the Tr rings to the Au array, and from the back-donation from the latter to the former. NICS calculations show a strong aromatic character in the gold monolayer sheet and Tr ligands; calculations done with HOMA, also report the same aromatic behavior on the cycloheptatrienyl fragments giving us an insight on the stability of (1). The Au –Au bond lengths indicate that an intramolecular aurophilic interaction among the Au(I) cations plays an important role in the bonding of the central metal sheet. Figure (a) Ground state geometry of complex 1; (b) Top view of compound 1 and Wiberg bond orders computed with the MP2/B1 computational method; (c) Lateral view of compound 1 and NICS values calculated with the MP2/B1 method; the values in parenthesis were obtained at the VWN/TZP level     

The influence of Sc doping on structural,electronic and optical properties of Be<Subscript>12</Subscript>O<Subscript>12</Subscript>, Mg<Subscript>12</Subscript>O<Subscript>12</Subscript> and Ca<Subscript>12</Subscript>O<Subscript>12</Subscript> nanocages: a DFT study

Density functional theory (DFT) calculations were used to study the effect of scandium doping on the structural, energetic, electronic, linear and nonlinear optical (NLO) properties of Be₁₂O₁₂, Mg₁₂O₁₂ and Ca₁₂O₁₂ nanoclusters. Scandium (Sc) doping on nanoclusters leads to narrowing of their E _g, which enhances their conductance greatly. Also, the polarizability (α) and first hyperpolarizability (β₀) of nanoclusters were dramatically increased as Be, Mg or Ca atoms are substituted with a Sc atom. Among all clusters, α and β₀ values for Sc-doped Ca₁₂O₁₂ were the largest. Consequently, the effect of the doping atom, as well as of cluster size, on electronic and optical properties was explored. Time dependent (TD)-DFT calculations were also carried out to confirm the β₀ values; the results show that the higher value of first hyperpolarizability belongs to Sc-doped Ca₁₂O₁₂, which has the smallest transition energy (ΔEgn). The results obtained show that these clusters can be candidates for using in electronic devices and NLO materials in industry.     

Aflatoxin B<Subscript>1</Subscript> Induced Systemic Toxicity in Poultry and Rescue Effects of Selenium and Zinc

Among many challenges, exposure to aflatoxins, particularly aflatoxin B₁ (AFB₁), is one of the major concerns in poultry industry. AFB₁ intoxication results in decreased meat/egg production, hepatotoxicity, nephrotoxicity, disturbance in gastrointestinal tract (GIT) and reproduction, immune suppression, and increased disease susceptibility. Selenium (Se) and zinc (Zn), in dietary supplementation, offer easy, cost-effective, and efficient ways to neutralize the toxic effect of AFB₁. In the current review, we discussed the impact of AFB₁ on poultry industry, its biotransformation, and organ-specific noxious effects, along with the action mechanism of AFB₁-induced toxicity. Moreover, we explained the biological and detoxifying roles of Se and Zn in avian species as well as the protection mechanism of these two trace elements. Ultimately, we discussed the use of Se and Zn supplementation against AFB₁-induced toxicity in poultry birds.     

Expression of human α<Subscript>1</Subscript>-proteinase inhibitor in <Emphasis Type="Italic">Aspergillus niger</Emphasis>

Background  

Human α₁-proteinase inhibitor (α₁-PI), also known as antitrypsin, is the most abundant serine protease inhibitor (serpin) in plasma. Its deficiency is associated with development of progressive, ultimately fatal emphysema. Currently in the United States, α₁-PI is available for replacement therapy as an FDA licensed plasma-derived (pd) product. However, the plasma source itself is limited; moreover, even with efficient viral inactivation steps used in manufacture of plasma products, the risk of contamination from emerging viruses may still exist. Therefore, recombinant α₁-PI (r-α₁-PI) could provide an attractive alternative. Although r-α₁-PI has been produced in several hosts, protein stability in vitro and rapid clearance from the circulation have been major issues, primarily due to absent or altered glycosylation.     

The subcommissural organ expresses D<Subscript>2</Subscript>, D<Subscript>3</Subscript>, D<Subscript>4</Subscript>, and D<Subscript>5</Subscript> dopamine receptors

Dopamine receptors have been found in certain populations of non-neuronal cells in the brain, viz., discrete areas of ciliated ependyma and the ependymal cells of the choroid plexus. We have studied the presence of both tyrosine-hydroxylase-immunoreactive nerve fibers and dopamine receptors in the subcommissural organ (SCO), an ependymal brain gland that is located in the roof of the third ventricle and that secretes, into the cerebrospinal fluid, glycoproteins that aggregate to form Reissners fiber (RF). Antibodies against D₂, D₃, D₄, and D₅ dopamine receptors were used in immunoblots of bovine striatum, fresh SCO, and organ-cultured SCO, and in immunocytochemistry of the bovine, rat, and mouse SCO. Only a few tyrosine-hydroxylase fibers appeared to reach the SCO. However, virtually all the secretory ependymal and hypendymal cells of the SCO immunoreacted with antibodies against D₂, D₄, and D₅ receptors, with the last-mentioned rendering the strongest reaction, especially at the ventricular cell pole of the secretory ependymocytes, suggesting that dopamine might reach the SCO via the cerebrospinal fluid. The antibodies against the four subtypes of receptors revealed corresponding bands in immunoblots of striatum and fresh SCO. Although the cultured SCO displayed dopamine receptors, dopamine had no apparent effect on the expression of the SCO-spondin gene/protein or on the release of RF-glycoproteins (SCO-spondin included) by SCO explants, suggesting that dopamine affects the function(s) of the SCO differently from the secretion of RF-glycoproteins.Financial support was provided by grants PI 030756 and Red CIEN, Instituto de Salud Carlos III, Spain (to J.M.P.F.), and 1030265 from Fondecyt, Chile (to E.M.R.)