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[en] Effect of gamma-ray on the destruction of aflatoxin in peanut were studied. Forty peanut samples available in the market from different parts of the country were inoculated with A. flatus. After incubation at room temperature for a certain period of time, the inoculated peanut samples were irradiated with 6.4 kGy dose. The concentration of aflatoxin B1 in irradiated and non irradiated samples were determined. It was found that there was no significant differences on the average concentration of aflatoxin B1 between irradiated and non-irradiated samples. Furthermore, the concentration of aflatoxin B1 in 22 out of 40 peanut samples (55 per mille) were varied from 0 to 4834 ppb and mostly (42.5 per mille) found above the level of the standard value. Maximum microbial load and aflatoxin concentration were found on 9 day of incubation, approximately 1.04 x 1010 colonies and 1200 ppb respectively
[en] Highlights: • A novel QB-based ICA with two test lines was developed for the accurately quantitative detection of AFB1 and ZEN. • The SA-biotin system was innovatively introduced as the signal output of the C line instead of anti-mouse IgG antibodies. • This work can serve as a reference for the sensitive and accurate detection of multiple targets simultaneously. - Abstract: Immunochromatographic assay (ICA) is a promising technology for on-site detection. Nonetheless, the wide-scale application of ICA is hindered by several disadvantages, such as poor reproducibility, low sensitivity, and single-target detection. Thus, a novel quantum dot nanobead (QB)-based multiplexed ICA (QB-ICA) with multiple test lines was developed in this study for the simultaneous quantitative detection of aflatoxin B1 (AFB1) and zearalenone (ZEN), where QBs with high luminescence were used as labels to enhance the analytical sensitivity of the ICA. Moreover, a streptavidin (SA)-biotin system, which was undisturbed by the target mycotoxins, was introduced as the signal output for the control line. Consequently, stable and reliable T/C values (ratios of signals on the test line to that of the control line) were obtained as quantitative signals. The proposed QB-ICA demonstrated high sensitivity for the simultaneous detection of AFB1 and ZEN, of which the half-maximal inhibitory concentrations reached as low as 38.98 pg mL-1 and 1.23 ng mL-1, respectively. At 10% competitive inhibition concentration, the limit detections (LOD) were 1.65 and 59.15 pg mL-1 for AFB1 and ZEN, respectively. The average recoveries of the intra- and inter-assays ranged from 81.77% to 119.70% and from 94.18% to 111.4% for AFB1 and ZEN quantification, respectively, and the variation coefficients were less than 12%, thereby indicating that the proposed method is highly accurate and robust. These findings suggest that QB-ICA using SA-biotin system as the signal output of control line is an excellent point-of-care platform for the rapid screening of mycotoxins.
[en] Produced by several species of Aspergillus, Aflatoxin B1 (AFB1) is a carcinogenic mycotoxin contaminating many crops worldwide. The utilization of fungicides is currently one of the most common methods; nevertheless, their use is not environmentally or economically sound. Thus, the use of natural compounds able to block aflatoxinogenesis could represent an alternative strategy to limit food and feed contamination. For instance, eugenol, a 4-allyl-2-methoxyphenol present in many essential oils, has been identified as an anti-aflatoxin molecule. However, its precise mechanism of action has yet to be clarified. The production of AFB1 is associated with the expression of a 70 kB cluster, and not less than 21 enzymatic reactions are necessary for its production. Based on former empirical data, a molecular tool composed of 60 genes targeting 27 genes of aflatoxin B1 cluster and 33 genes encoding the main regulatory factors potentially involved in its production, was developed. We showed that AFB1 inhibition in Aspergillus flavus following eugenol addition at 0.5 mM in a Malt Extract Agar (MEA) medium resulted in a complete inhibition of the expression of all but one gene of the AFB1 biosynthesis cluster. This transcriptomic effect followed a down-regulation of the complex composed by the two internal regulatory factors, AflR and AflS. This phenomenon was also influenced by an over-expression of veA and mtfA, two genes that are directly linked to AFB1 cluster regulation.(author)
[en] The aim of our study was to gain starting information on the aflatoxin B1 and ochratoxin A levels in cereals and feed mixtures which are in poultry breeding. To ascertain the presence of mycotoxins, we examined the cultivars of cereals (maize and wheat) and the feed mixtures. The cereals came from different regions of eastern Slovakia. In all cereals examined, the low mycotoxin levels did not exceed the tolerance limit set by hygienic standard (Sv. 61, 1986, No. 69). In wheat, the contamination by aflatoxin B1 ranged from 0.028 to 0.125 μg·kg-1. In maize, the contamination by aflatoxin B1 ranged from 0.166 to 0.707 μg·kg-1. The results enhance our knowledge of feedstuff and feed mixture contamination in poultry breeding
[en] Complete text of publication follows. The use of quantum dots (QDs) as fluorescent labels in the development of luminescent immunoassays is a powerful tool because of the exceptional optoelectronic properties of these nanoparticles compared with organic fluorescent dyes, such as narrow emission spectra, tuneable maximum wavelength of emission, less susceptibility to photobleaching, a broadband absorption spectra and intense and long-lasting photoluminescent emission. Moreover, the combination of these interesting optical properties with the selective detection of a given target molecule by its specific binding to an antibody justifies the great interest in the present research on QDs as a luminescent labels for the development of fluorescent immunoassays. However, to carry out these assays is necessary to ensure the separation between labeled and free antibody to guarantee that all the signal obtained corresponds to the antigen present in the sample and to avoid the interference of the free antibody. In this communication the results obtained for the investigation of two methods of purification of bio-conjugates and its application for the determination of Aflatoxin B1 (very toxic compound that might be present in several foods) will be shown. On the one hand, the bio-conjugates have been separated from the free antibodies and QDs excess by size exclusion chromatography (SEC), a sophisticated and well probed method (M. T. Fernandez-Argueelles et al. The Analyst, 133 (2008) 444-447) and on the other hand the separation is carried out by centrifugal filter of 100 KDa, an alternative and simplest method. The success of these purification methods is checked using an ELISA and fluorescence-based immunoassays to compare the response of the bio-conjugates purified by the two methods after knowing the concentration of these conjugates by a Bradford Test. Finally, a fraction of centrifugal filter bio-conjugate purified is also checked by SEC to demonstrate the success of this method of purification. As a result it can be affirmed that the centrifugal filter method could be used to purify the bio-conjugates but keeping in mind that the recovery and response of the conjugates purified by this method are worse than by the SEC method.
[en] In this study various poultry and fish feed samples were initially analyzed for presence of aflatoxin. All the samples were found contaminated with aflatoxin B I only. Contaminated samples were treated with different organic and inorganic chemicals to detoxify aflatoxin B 1 in poultry and fish feed samples. The maximum reduction in the aflatoxin Bl concentration was observed with 0.5% HCI as 14.20 ppb to 2.09 ppb (86.50%) in the poultry and 69.26 ppb to 10.46 ppb (84.89%) in fish feed samples.
[en] Highlights: • A versatile aptasensor was developed for simultaneous detection of OTA and AFB1. • This aptasensor has a wide detection range and low detection limit of pg mL-1 level. • The operation process is simple and convenient by using magnetic separation. • This strategy provides a new avenue for high throughput screen of multi-mycotoxins. - Abstract: Development of an efficient method for the simultaneous detection of two highly concerning mycotoxins, ochratoxin A (OTA) and aflatoxin B1 (AFB1), is of great significance on food safety monitoring. Herein, a magnetically controlled fluorescence aptasensor for simultaneous determination of OTA and AFB1 has been successfully developed. The working principle of the aptasensor is based on the specific aptamer-mycotoxin recognition and further leads to the partial release of two distinguishable fluorescence labels from the magnetic carriers. Through the magnetic separation, the reporter probes in the supernatant solution can be collected and converted into a sensitive fluorescence signal with dual emission peaks. This aptasensor provided a wide detection range of 2 pg mL-1 - 5 ng mL-1 for OTA and 5 pg mL-1 - 10 ng mL-1 for AFB1. The new easy-to-wash and simple-to-use approach offers a simultaneous and high selective detection with high sensitivity (limits of detection of 0.67 and 1.70 pg mL-1 for OTA and AFB1, respectively). Remarkable accuracy (relative standard deviation < 5.6%) during the mycotoxins determination as well as excellent quantitative recoveries (95–108%) during the analysis of the spiked corn samples were also achieved. This simple aptasensing scheme provides a new avenue for high throughput screen of dual mycotoxins due to its simple manipulation, short assay times, high selectivity and sensitivity.
[en] Between 2002 and 2003, an outbreak of a trout's mass death occurred at the intensive fish culture a Peruvian rural town (Marcara, Huaraz, Peru) where 15,000 from 20,000 fish died. Our objective in the present study was to investigate the high mortality of the trout biomass occurred in period of two months. This study was conducted after the peak of the outbreak has occurred. We collected samples of fishes, water and fish foodstuff which were examined for aflatoxin, metals, toxics and bacteria. We interviewed people who administered the feed pellet. Feed sample preparation, transport and storage. The processing of fish feed was at room temperature which was below 16 deg C. Once prepared the diet it was keep under an appropriate room for a few days before sending to Marcara town. Fishes. 20,000 immature trout larval of rainbow trout (Oncorhynchus mykiss) was acquired from an official Peruvian fish culture. The fishes were fed twice a day. Adjusted of feed ration was based from the monthly sample weight. Pellet sample analysis. The samples were analyzed for aflatoxin Bl (AFB1) according to the method previously published. The sensitivity is 0.1 μg per 1 kg of sample. During the fish development until the peak of the outbreak, the foodstuff to fishes was maintained in plastic bags. At this time the storage room temperature was 18-20 deg. C between 1.00-2.00 P.M. and the humidity rose close to 90 % at the Marcara facilities. Mortality development and Effect on survival. The fishes maintained in 4 pods had a normal surviving until end of November, less than 10 specimen dead by month. The fish outbreaks started the first week of December and continuing until the fourth week of January totalizing 15,000 dead fish from 20,000. The survival of the fish at the first month was less than 50 %. The mortality continues throughout January totalizing 15,000 dead fish and leaving only 25% survival. Laboratory data. The collected samples for analysis were frozen and transported in dry ice to the analysis laboratory. We took the samples on January 23 and it was analyzed on January 25. Aflatoxin Bl was detected in three samples of fish muscle and in the 3 samples of fish feed but it was negative in the 3 water samples. The AFB1 concentration was 10 times in the fish feed than in the fish muscle. In spite of heavy metal residues (lead, mercury and arsenic) were found in the fish samples, those concentrations were below the permissible levels. Volatile toxic residues were negative in water, fish and feed. Only the fish feed samples were contaminated by bacteria (Staphylococcus aureaus). Under favourable conditions of temperature and humidity, the Aspergillus flavus grows on certain foods and feeds, resulting in the production of aflatoxin Bl. For the trout, the highest admissible amount of AFBI in feed is 0.1 μg per kg. The data showed suggest that an improper handling of fish foodstuff (18-20 deg. C and 90 % humidity) was the cause growing of mould and/or spores and consequently it produced an increased concentration of AFBI in fish feed. Liver is strategically located between intestinal tract and general circulation. As AFBI concentration ranged in liver between 10 and 100 ppb, this level is capable to produce an acute hepatotoxicity in the fish stocks. (author)
[en] Fourteen fungal species were isolated from six kinds of spices (namely black pepper, chilli, cumin, aniseed, caraway and cinnamon) belonging to the genera aspergillus, penicillium, paecilomyces, epicoccum and trichoderma. The production of aflatoxins were detected for all fungal isolates (121 isolates) in synthetic medium (sabouroud's yeast extract medium) by thin layer chromatography and biological method. A. flavus (isolated from black pepper) A. niger (isolated from chilli) A. ochracues (isolated from cumin) and penicillium notatum (isolated from caraway) produced aflatoxins B2, B1,B2 and G1 respectively. The highest amount of toxins was produced by A. flavus. It has been found that irradiation dose 5 kGy completely inactivated the fungal flora contaminated all kinds of spices, Whereas the irradiation dose 3 kGy was the inhibitory dose for growth of toxigenic fungi and completely inhibited the production of aflatoxins. On the other hand the inhibitory effect of tested spices on the growth of toxigenic fungus A. flavus as well as on the production of aflatoxins was also investigated. It has been found that 2% of cinnamon or cumin completely inhibited the production of aflatoxin produced by A. flavus
[en] Whole-body autoradiography in rainbow trout (Oncorhynchus mykiss) after oral and intravenous administration of 3H-labelled aflation B1 showed labelling of several extrahepatic tissues, such as the uveal melanin and the vitreous humour of the eyes, the trunk and head kidney, the olfactory rosettes and the pyloric caecae. Liquid chromatography of extracts of the vitreous humour showed that unmetabolized 3H-AFB1 was the main labelled material present at this site. Liquid chromatography of extracts of the uveal melanin showed presence of aflatoxicol and aflatoxin B1 in proportions of about 3:1. The binding to the pigment is probably due to a hydrophobic type of interaction with the melanin. Microautoradiography showed that melanin-containing cells in the trunk and head kidney and in the olfactory rosettes also accumulated high amounts of radioactivity. In the trunk kidney there was, in addition, a labelling of the second segment of the proximal tubules and of the distal tubules and the collecting ducts. Studies in vitro with microsomal and 12,000xg supernatant preparations of the trunk kidney showed formation of DNA- and protein-bound metabolites from the aflatoxin B1. It is probable that the bioactivation of the aflatoxin B1 is confined to the second segment of the proximal tubules. The labelling of the distal tubules and the collecting ducts, which was confined to the cytoplasm of the cells, may be related to excretion and/or absorption processes. Microautoradiography of the olfactory rosettes, showed labelling of the sensory epithelium, but not the indifferent epithelium. A low formation of protein-bound aflatoxin B1 -metabolites was found in incubations with microsomal preparations of this tissue. The same observation was made in incubations with microsomal preparations of the head kidney. In the pyloric caeca bound metabolites were observed in vivo at a level comparable to that found in the trunk kidney. Our results suggest that retention and metabolism in some extrahepatic tissues might be of importance as concerns the toxicologic potential of aflatoxin B1 in the rainbow trout. (au)