INCIDENCE OF AFLATOXIGENIC ISOLATES OF ASPERGILLUS FLAVUS/PARASITICUS OBTAINED FROM GEORGIA CORN PROCESSING PLANTS

Two corn processing facilities within Georgia were evaluated in order to determine the incidence of Aspergillus flavus or A. parasiticus within the plant and in corn harvested and processed in 1984 and 1985. Conidia of A. flavus/parasiticus were found in all corn samples evaluated as well as in settled dust samples taken within these processing facilities. Isolates were obtained by using the differential/selective medium Aspergillus flavus/parasiticus agar. Upon subsequent culture only 55% of the selected isolates were confirmed as belonging to A. flavus/parasiticus group. Some of these isolates were randomly chosen and their ability to produce aflatoxins B₁, B₂, G₁, or G₂ evaluated. Thirty-two percent of the A. flavus/parasiticus isolates cultured for aflatoxin production were found to be aflatoxigenic.     

Aflatoxin-producing potential of Aspergillus flavus and Aspergillus parasiticus isolated from samples of smoked-dried meat

Over a period of three years 420 samples of various smoke-dried meat products, collected from individual households in different region of Croatia were analysed for the presence of aflatoxigenic strains of the Aspergillus flavus group. Strains of A. flavus and A. parasiticus were present in 17,8% of the samples, and aflatoxin-producing ability was tested in 75 strains. In relation to sequential method of aflatoxin detection, 5 of 8 isolates were found in the first step (fluorescence in aflatoxin-producing ability medium - APA) and all of them in the second step (extraction method from syntheses on moist shredded wheat - SW). A. flavus strains produced mainly aflatoxin B₁, and had various levels of toxigenicity (1.4–3.12 mg/kg). Some strains of A. parasiticus produced all four aflatoxins B₁ B₂ G₁ G₂, while the other ones produced AF B₁ + G₁ only, with concentrations of aflatoxins from 0.1 to 450 mg/kg.     

Mycotoxin production by molds isolated from ‘Greek-style’ black olives

Seventeen mold strains were isolated from ‘Greek-style’ black olives produced in Morocco. Eight of these isolates were identified as Aspergillus flavus, seven as Aspergillus petrakii, and two as Aspergillus ocharaceus Wilhelm. The A. flavus strains were tested for production of aflatoxins B₁, B₂, G₁, and G₂; and A. ochraceus and A. petrakii strains were tested for production of ochratoxin, penicillic acid, patulin, and citrinin. The organisms were tested for mycotoxin production on five different substrates, including rice powder-corn steep agar, autoclaved rice, yeast-extract sucrose broth (YES), potato dextrose agar (PDA), and fresh olive paste. All strains of A. flavus produced aflatoxins on all substrates except olive paste and PDA. In PDA, only two strains produced Aflatoxin B₁. Five A. ochraceus group isolates produced penicillic acid on one or more of the substrates, but only two out of the five produced penicillic acid on olive paste. None produced ochratoxin, patulin or citrinin. Quantities of aflatoxin B₁ produced in rice ranged from 5 to 14 μg/g of rice, and of penicillic acid 15–32 μg/g of rice. In olive paste, the concentrations of penicillic acid were 11.4 and 30.2 μg/g. Biological toxicity of extracts of mold cultures was confirmed using chicken embryos and a microbiological test. Crude extracts of cultures were also tested for mutagenicity using the Salmonella mutagenicity (Ames) Test, and some gave positive mutagenic responses.     

Spoilage moulds and aflatoxins from poultry feeds

Aflatoxin producing strains of Aspergillus flayus Link (IMI 280819) and A. oryzae (Ahlb.) Cohn (IMI 280831) were among the eleven spoilage moulds isolated from five types of poultry feeds. The recorded pH and moisture content values of the various feeds are conducive to mould deterioration. All the four principal aflatoxins (B₁, B₂, G₁, G₂) were detected in the analysed feeds though at toxicologically ‘safe’ levels for most farm animals. Significant quantities of aflatoxin B₁ were produced by the two fungal isolates in all the five classes of poultry feeds with A. flavus yielding the larger amounts. Optimum aflatoxin B₁ production and mycelial growth in chick mash infusion medium were recorded for both species at 30 and 35 °C, respectively and similarly on the 8th and 6th day respectively when cultures were incubated at 30 °C.     

Moulds and mycotoxins in rice from the Swedish retail market

A survey of moulds and mycotoxins was performed on 99 rice samples taken from the Swedish retail market. The main objective was to study the mould and mycotoxin content in basmati rice and rice with a high content of fibre. Samples of jasmine rice as well as long-grain rice were also included. The samples were analysed for their content of ochratoxin A (high-performance liquid chromatography (HPLC)), aflatoxin B₁, B₂, G₁, and G₂ (HPLC, RIDA®QUICK), and mould (traditional cultivation methods in combination with morphological analysis). The majority of samples were sampled according to European Commission Regulation 401/2006. Subsamples were pooled and mixed before milling and both mould and mycotoxin analyses were performed on milled rice. The results showed that the majority of basmati rice (71%) and many jasmine rice samples (20%) contained detectable levels of aflatoxin B₁ (level of quantification = 0.1 µg aflatoxin kg^?1 rice). Two samples of jasmine rice and ten basmati rice samples contained levels over the regulated European maximum limits of 2 µg kg^?1 for aflatoxin B₁ or 4 µg kg^?1 for total aflatoxins. Aspergillus was the most common mould genus isolated, but also Penicillium, Eurotium, Wallemia, Cladosporium, Epicoccum, Alternaria, and Trichotecium were found. The presence of Aspergillus flavus in 21% of the samples indicates that incorrect management of rice during production and storage implies a risk of mould growth and subsequent production of aflatoxin. Rough estimates showed that high rice consumers may have an intake of 2–3 ng aflatoxin kg^?1 bodyweight and day^?1 from rice alone. This survey shows that aflatoxin is a common contaminant in rice imported to Europe.     

Pre-concentration and Extraction of Aflatoxins from Rice Using Air-Assisted Dispersive Liquid–Liquid Microextraction

An easy and quick sample pretreatment method, air-assisted dispersive liquid–liquid microextraction (AA-DLLME), has been introduced for the isolation and enrichment of aflatoxins B₁, B₂, G₁, and G₂ from rice samples prior to HPLC-FLD analysis. The method has also been compared with immunoaffinity column clean-up (IAC) method. In the case of AA-DLLME, the dispersion of extractant is formed in the absence of a disperser solvent and with the help of air bubbles. Several crucial factors, including the volume and type of extraction solvent, the volume of aqueous phase, and the number of extraction cycles were studied to obtain the optimal conditions. Under the optimal conditions, the suggested technique showed linearity in the range of 0.08–10 ng/mL with low limits of detection (0.68, 0.68, 0.13, and 0.13 ng/g for aflatoxins B₁, G₁, B₂, and G₂, respectively) and good sensitivity with limits of quantification of 2.0, 2.0, 0.4, and 0.4 ng/g for aflatoxins B₁, G₁, B₂, and G₂, respectively. The proposed procedure was successfully used to evaluate aflatoxins (AFs) in rice samples and acceptable recoveries in the range of 76.0–109.3% with appropriate precision (RSD?<?14.2%) were obtained.     

西藏高原粮油作物曲霉菌污染及菌株产毒力研究

为探明西藏高原粮油作物曲霉菌污染状况及黄曲霉菌产毒能力,连续5年对西藏青稞、小麦、花生3种作物中曲霉菌污染情况进行分析,并对其分离到的黄曲霉菌株开展产毒力研究,结果表明,204份样品中,共分离出15种曲霉菌,曲霉菌污染率呈花生>青稞>小麦。青稞、小麦中曲霉属优势种均为黑曲霉(Aspergillus niger),真菌毒素主要为杂色曲霉毒素和赭曲霉毒素;花生优势种为黄曲霉(A.flavus);仅受黄曲霉毒素污染。来源于不同作物的黄曲霉菌,其产毒类型也有差异,麦类作物产毒菌株以产黄曲霉毒素B₁(AFB₁)、黄曲霉毒素B₂(AFB₂)为主;花生产毒菌株以产AFB₁、AFB₂、AFG₁、AFG₂为主。     

Distribution of aflatoxins in shelling and milling fractions of naturally contaminated rice

The objective of this study was to determine the distribution of an economically important class of mycotoxins, the aflatoxins, in rice milling fractions. Rice plants grown under field production conditions are frequently infected with types of pathogenic fungi that produce toxic metabolites (mycotoxins). Paddy (seeds) rice from healthy plants in the field was collected and stored on a farm under humid, poorly ventilated conditions. Samples were milled into four fractions (hulls, brown rice, bran and white rice) and analysed for aflatoxins (B₁, B₂, G₁ and G₂) using a validated method. Rice fractions from healthy plants, which contained low levels of aflatoxins (less than 1?µg?kg^?1), were used to determine the efficiency of the extraction method. Seeds stored under poor conditions were found to be contaminated with aflatoxins B₁ and B₂ as were the fractions. The sums of AFB₁ and AFB₂ in stored paddy rice, hulls, brown rice, bran and white rice were 141, 39, 158, 367 and 56?µg?kg^?1, respectively. The ratio of aflatoxin B₁ and B₂ was about 10?:?1. AFG₁ and AFG₂ were less than 1?µg?kg^?1. Thus, brown rice contained 92.9% of the aflatoxins in paddy rice, whereas white rice contained only 27.9%.     

Effect of BHA, BHT, TBHQ and PG on Growth and Toxigenesis of Selected Aspergilli

The antifungal effect of butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), tertiary butylhydroquinone (TBHQ) and propyl gallate (PC) alone or in combination on three toxigenic strains of aspergilli (NRRL 2999, NRRL 4123, NRRL 5835) and three nontoxigenic strains of aspergilli (NRRL 5521, NRRL 5917, NRRL 5918) was examined in a solid medium and in salami. BHT and PG (0.001,0.005,0.01,0.02g per plate) did not inhibit growth, sporulation, and toxigenesis of all six cultures. Aflatoxin production by toxigenic aspergilli (B₁, B₂, G₁ and G₂) in the presence of BHA, TBHQ, and a combination of BHA and TBHQ was reduced significantly (P < 0.05). In salami BHA, TBHQ alone or in combination at 100 ppm significantly (P < 0.05) decreased the aflatoxin production by aspergilli when compared to control samples. A combination of BHA and TBHQ showed synergistic inhibition in both studies (solid medium and salami studies).     

贵州秋季栽培不同荞麦品种成熟期果实中黄曲霉及其毒素的分离与鉴定

本研究以贵阳秋季栽培的2个米苦荞(F.tataricum,贵米苦荞18-1号和贵黑米苦荞12号)、2个多苦荞(F.tatari-cymosum,贵多苦荞003C和贵多苦荞60)、2个甜荞(F.esculentum,贵红花甜荞2号和1412-1)、2个常规苦荞(F.tataricum,定苦荞1号和六苦2017)为材料。对其成熟期种子果壳和籽粒进行了黄曲霉分离鉴定,并采用高效液相色谱法对所有品种果壳和籽粒中分离出的黄曲霉菌株进行AFB₁、AFB₂、AFG₁和AFG₂毒素的检测。结果表明,所有品种果壳中均没有分离出黄曲霉菌落;4类荞麦籽粒中仅米苦荞分离出了黄曲霉菌落,共分离出4株黄曲霉菌株。其中贵米苦荞18-1号黄曲霉带菌率为1.56%,贵黑米苦荞12号黄曲霉带菌率为0.78%。分离菌株形态学和ITS序列扩增产物测序结果与已知黄曲霉菌序列完全一致。毒素检测结果表明不同品种之间产毒素差异显著,所有品种籽粒中只有米苦荞中检出4种毒素,贵米苦荞18-1号产AFB₁最高为(5.861±0.055) μg/kg、AFB₂最少为(1.605±0.052) μg/kg,贵黑米苦荞12号产AFB₁最高为(14.475±0.533) μg/kg、AFG₂最少为(3.393±0.151) μg/kg;籽粒产毒量远大于分离菌株产毒量;各分离出菌株之间产毒素能力差异显著,最大产AFT为(11.102±0.095) μg/kg、最小产AFT为(1.794±0.024) μg/kg。上述结果显示供试米苦荞籽粒带菌来源可能是由于果壳开裂籽粒外露后部分籽粒被直接侵染所致。所得结果可为米苦荞中黄曲霉抗性育种研究及荞麦种子的保存、运输、储藏等研究奠定基础。     

Mycotoxin co-occurrence in rice,oat flakes and wheat noodles used as staple foods in Ecuador

The co-occurrence of aflatoxin B₁ (AFB₁), B₂ (AFB₂), G₁ (AFG₁) and G₂ (AFG₂), ochratoxin A (OTA), deoxynivalenol (DON), fumonisin B₁ (FB₁), zearalenone (ZEN), and HT-2 and T-2 toxins in the main Ecuadorian staple cereals (rice, oat flakes, and yellow and white wheat noodles) was evaluated. A ultra high performance liquid chromatography/time-of-flight mass spectrometry (UHPLC/TOFMS) method was developed and validated to screen for the presence of these mycotoxins in those cereal matrices. Matrix-matched calibration curves were used to compensate for ion suppression and extraction losses and the recovery values were in agreement with the minimum requirements of Regulation 401/2006/EC (70–110%). For most mycotoxins, the LODs obtained allowed detection in compliance with the maximum permitted levels set in Regulation EC/2006/1881, with the exception of OTA in all cereals and AFB1 in yellow noodles. Extra target analysis of OTA in oat flakes and wheat noodles was performed by HPLC with fluorescence detection. High rates of contamination were observed in paddy rice (23% DON, 23% FB₁, 7% AFB₁, 2% AFG₁ and 2% AFG₂), white wheat noodles (33% DON and 5% OTA) and oat flakes (17% DON, 2% OTA and 2% AFB₁), whereas the rates of contamination were lower in polished rice (2% AFG₁ and 4% HT-2 toxin) and yellow noodles (5% DON). Low rates of co-occurrence of several mycotoxins were observed only for white wheat noodles (5%) and paddy rice (7%). White noodles were contaminated with DON and/or OTA, while combinations of AFG₁, AFB₁, DON and FB₁ were found in paddy rice. Yellow noodles were contaminated with DON only; oat flakes contained DON, OTA or AFB₁, and polished rice was contaminated with AFG₁ and HT-2 toxin.     

Mycotoxic flora and mycotoxins in smoke-dried fish from Sierra Leone

Examination of 20 samples of smoke-dried fish of the Ethmolosa sp. commonly called “Bonga”, from homes and markets in Njala (Sierra Leone) revealed the presence of 4 Aspergilli species: A. flavus Links ex Fries, A. ochraceus Wilhelm, A. tamarii Kita and A. niger van Tieghem. Fresh or properly preserved smoke-dried fish showed no signs of fungal contamination. Mouldy fish extracts contained varying amounts of aflatoxins B₁, G₁, G₂ and ochratoxin A. Isolates of A. flavus grown on yeast extract sucrose (YES) medium, produced considerable amounts of aflatoxin B₁ and G₁ and trace amounts of G₂. On YES medium A. ochraceus produced large amounts of ochratoxin A but no penicillic acid.     

Identification and quantification of fungi and mycotoxins from Pu-erh tea

Pu-erh tea originates from the province of Yunnan in south-western China. As this tea is produced by so called Aspergillus post-fermentation the question arises which molds and mycotoxins may be found in this tea. In total 36 samples of Pu-erh tea were investigated for their content of filamentous fungi and the mycotoxins aflatoxins B₁, B₂, G₁, and G₂, fumonisins B₁, B₂, and B₃, and ochratoxin A. Fungi were isolated from all samples in a concentration of 1.0 × 10¹ to 2.6 × 10⁶ colony forming units (cfu)/g tea, all together 19 fungal genera and 31 species were identified. The most prevalent species were Aspergillus acidus and Aspergillus fumigatus, followed by Zygomycetes and Penicillium species. Aflatoxins and fumonisins were not found in the samples investigated, ochratoxin A was detected in 4 of 36 teas (11.1%).     

2013—2015年广州市市售普洱茶真菌毒素污染调查

目的 了解广州市市售普洱茶真菌毒素污染状况及评估健康风险。方法 2013—2015年,在广州市批发市场、零售店、超市、餐饮单位、网店和茶博会6类场所采集不同生产工艺、包装形态、生产年份、原料产地、价格的普洱茶样品432份,使用多功能净化柱净化-柱后衍生高效液相色谱法测定黄曲霉毒素(B₁、B₂、G₁、G₂),高效液相色谱-质谱法测定雪腐镰刀菌烯醇、脱氧雪腐镰刀菌烯醇和玉米赤霉烯酮。结果 432份普洱茶样品黄曲霉毒素(B₁、B₂、G₁、G₂)、雪腐镰刀菌烯醇、脱氧雪腐镰刀菌烯醇、玉米赤霉烯酮7个项目检测结果均低于所用检测方法的检出限。结论 本次对广州市市售普洱茶的调查研究,未检出真菌毒素。     

Effects of gamma irradiation on aflatoxins

Summary The effects of gamma irradiation on a mixture of aflatoxins B₁, B₂, G₁, G₂ were studied. Standard solutions of A and B were irradiated at 5, 10, and 20 kGy in a solution of water/DMSO (9+1, v/ v) by using a¹³⁷Cs source. The control (0 kGy) and irradiated samples were subjected to RP-HPLC analyses with methanol/water (4+5, v/v) as the mobile phase. Aflatoxin B₁ (AFB₁) was the most radio-sensitive of the four compounds. The radiosensitivity of the other aflatoxins, was in increasing order: G₂, B₂, G₁. Only about 5% of AFB₁ remained after irradiation of solution A at 5 kGy. When the concentration of solution B was increased two-fold, trace amounts of AFB₁ remained after irradiation doses of 10 and 20 kGy. Irradiation was found to be suitable for the destruction of aflatoxins in solution.

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Einfluß von Gammastrahlung auf die Aflatoxine

Zusammenfassung Der EinfluB von -Strahlen auf die Aflatoxinmischung B₁, B₂, G₁ und G₂ wurde untersucht. Standardlösungen A und B wurden mit -Strahlung von 5; 10; bzw. 20 kGy in der Lösung von Wasser/DMSO (9+1, v/v) unter Verwendung einer¹³⁷Cs-Quelle bestrahlt. Kontroll(0 kGy)-und bestrahlte Proben wurden der RP-HPLC Analyse unter Verwendung von Methanol/Wasser (4+5, v/v) als mobile Phase unterworfen. Aflatoxin B₁ (AFB₁) war die strahlenempfindlichste von den vier, die Strahlenempfindlichkeit der anderen Aflatoxine nahm bei 5 kGy in der Reihenfolge: G₂, B₂, G₁ zu. Nur ca. 5% von AFB₁ ist nach der Bestrahlung üibriggeblie-ben. Wenn sich die Konzentration zweifach erhöht (Lösung B), ist von AFB₁ nur eine Spur nach der 10 kGy- und 20 kGy-Bestrahlung übriggeblieben. Es wurde gefunden, daß die Bestrahlung zur Zerstörung der Aflatoxine in der Lösung geeignet ist.

     

Aflatoxin accumulation in whole crop maize silage as a result of aerobic exposure

BACKGROUND: Most of the maize silage stored in horizontal silos is exposed to air and can be spoiled by fungi. Potentially toxigenic fungi have been found in maize silage, and about 300 mycotoxins have been detected. Among these mycotoxins, the most harmful for feed and food safety are aflatoxins. The aim of the study was to set up a specific method to detect aflatoxins in maize silage, and to investigate whether aflatoxin contamination in maize silage depends on the level of field contamination of the crop, and whether the occurrence of aerobic spoilage during ensiling has any effect on the final contamination of the silage. RESULTS: A method for the determination of aflatoxin B₁, B₂, G₁ and G₂ in maize silage using high‐performance liquid chromagraphy with fluorescence detection has been developed and validated. Recoveries of aflatoxin B₁, B₂, G₁, and G₂ spiked over the 0.25 to 5 µg kg^?1 range averaged 74–94%. The results of laboratory scale and farm scale ensiling experiments indicated that aflatoxins could increase when silage is exposed to air during conservation or during the feed‐out phase. CONCLUSIONS: The method here proposed to detect aflatoxins in silages has proved to be sensitive and is able to detect levels of 0.1 and 0.5 ng mL^?1 for AFB₁ and AFG₁, and between 0.025 and 0.125 ng mL^?1 for AFB₂ and AFG₂. This study also provides evidence of aflatoxin accumulation in whole crop maize silage as a result of aerobic exposure. Copyright © 2011 Society of Chemical Industry     

Behavior of Aflatoxin during the Manufacture, Ripening and Storage of Manchego-type Cheese

The behavior of aflatoxins B₁, B₂, G₁, G₂, and M₁ was investigated during manufacture, ripening, refrigeration, and frozen storage of Manchego-type cheese. More aflatoxins (per weight or volume) occurred in curd than in whey, although total quantity was greater in whey. Aflatoxins B₁ B₂, and M₁ remained stable during ripening, but G₁ and G₂ levels increased. Concentration of B₁, B₂, and M₁ decreased after 15 and 30 days refrigeration storage, but after 60 days 100% recovery occurred in the inner portion and only 60% in the outer. Concentrations of G₁ and G₂ also decreased during the first 30 days, but on day 60 an increase was observed, 200% in the inner and 120% in the outer portions. Aflatoxins were stable after 90 days in frozen storage.     

High performance liquid chromatographic determination of aflatoxins in chilli,peanut and rice using silica based monolithic column

A simple and rapid high performance liquid chromatographic with fluorescence detection method for the determination of the aflatoxin B₁, B₂, G₁ and G₂ in peanuts, rice and chilli was developed. The sample was extracted using acetonitrile:water (90:10, v/v%) and then purified by using ISOLUTE® multimode solid phase extraction. After the pre-column derivatisation, the analytes were separated within 3.7 min using Chromolith® performance RP-18e (100–4.6 mm) monolithic column. To assess the possible effects of endogenous components in the food items, matrix-matched calibration was used for the quantification and validation. The recoveries of aflatoxins that were spiked into food samples were 86.38–104.5% and RSDs were <4.4%. The method was applied to the determination of aflatoxins in peanut (9), rice (5) and chilli (10) samples. Liquid chromatography–tandem mass spectrometry analysis using triple quadruple analyser and operated in the multiple reaction monitoring modes on the contaminated samples was performed for confirmation.     

Aflatoxins in Foods and Feedstuffs. Part 1. The Fluorescence Micro-method of Aflatoxin Determination in solution

The fluorescence micromethod of aflatoxins B₁, B₂, G₁ and G₂ determination in solution is described. The amount about 0.1 μg of aflatoxins B₁ and G₁, and 0,01 μg of aflatoxins B₂ and G₂ in one spot after elution with methanol can be determined, with accuracy about 5%. The results of described fluorescence method are in agreement with results of spectrophotometric, fluorodensitometric and visual determinations. A very strong influence of light on results of aflatoxins B₁ and G₁ determination was stated. Photodecomposition of aflatoxins B₁ and G₁ to several photoproducts was observed.     

Aflatoxin (B1, B2, G1, and G2) Contamination in Rice of Mexico and Spain,from Local Sources or Imported

Rice is an important cereal but it is often contaminated with aflatoxins (AFs). The purpose of this study was to identify and quantify AF (B₁, B₂, G₁, and G₂) in 67 rice samples cultivated in Mexico and Spain, and from imported crops collected in 2008 and 2009. The methodology was validated, the rice samples were concentrated and purified with immunoaffinity columns and were quantified by high‐pressure liquid chromatography (HPLC). The average total AF (AFt) in the Spanish rice was 37.3 μg/kg, the range was from 1.6 to 1383 μg/kg, the most contaminated samples being from San Juan de Aznalfarache, Sevilla (AFt = 138.6 μg/kg), from Tortosa, Tarragona (AFt = 104.6 μg/kg), and Calasparra, Murcia (AFt = 103.9 μg/kg). The rice imported from France to Spain had AFt of 26.6 μg/kg and from Pakistan AFt of 18.4 μg/kg, showing less AF contamination than the local one. The rice which originated from Mexico contained (AFt = 16.9 μg/kg), and those imported from the United States (AFt = 14.4 μg/kg) and Uruguay (AFt = 15.6 μg/kg). The imported rice had better quality in terms of the presence of AFs.