Fungi and aflatoxins associated with spices in the Sultanate of Oman

One hundred and five samples of seven spices (cumin, cinnamon, clove, black pepper, cardamom, ginger, and coriander) were purchased from five popular companies in the Sultanate of Oman. The spices were surveyed for the mycoflora and aflatoxins. Twenty fungal species were isolated in which Aspergillus flavus , A. niger . Penicillium , Rhizopus , and Syncephalastrum racemosum were the most dominant. When colony forming units per gm (cfu/gm) of fungi were compared, significant differences were found among spices and companies. Of the seven spices studied, clove was found to be the least contaminated, while cumin was the most contaminated. None of the 15 selected samples of the spices contaminated by A. flavus were found to contain aflatoxins. Nevertheless, nine isolates (45%) of the twenty A. flavus strains screened for aflatoxins were aflatoxigenic. The moisture content of most of the spices was below the maximum standard limit. The results showed that the spices were contaminated by some fungi that might constitute health hazards for humans. This revised version was published online in June 2006 with corrections to the Cover Date.     

A mycological and bacteriological survey on feed ingredients and mixed poultry feeds in Reunion island

A survey was carried out in Reunion island to obtain data on the occurrence of fungi, aflatoxigenic strains of Aspergillus flavus, aflatoxins, total aerobic bacteria and salmonellae of 150 samples of mixed poultry feeds and raw materials. These were collected at five farms over a 3-month period during the warm rainy season.White corn and Brazilian soybean meal seemed to present a better microbiological quality than yellow corn and US soybean meal.Mixed poultry feeds presented a high total mold count reflecting the mold flora of raw materials. The most frequent and abundant fungi were Aspergillus flavus, A. glaucus group, Fusarium spp., Penicillium spp., A. candidus, Mucor spp., A. restrictus, Scopulariopsis spp., Cladosporium spp. and A. versicolor. Of the 118 A. flavus strains screened, 42 (35.6%) were aflatoxigenic. Yellow corn samples were the most frequently contamined with aflatoxigenic strains (54.5%), followed by mixed feeds (44%).Of the 66 samples tested, 24 (36%) contained aflatoxins (traces to 22 ng/g). A good correlation seemed to exist between presence of at least one aflatoxigenic strain per sample and presence of aflatoxins.     

Natural occurrence of aflatoxin andAlternaria mycotoxins in oilseed rape from Catalonia (Spain): Incidence of toxigenic strains

During an investigation of the mycoflora on oilseed rape, the predominant fungal species present in 20 samples collected from Catalonia (Spain) wereAlternaria alternata (Fries) Keissler,Penicillium spp. andAspergillus flavus. None of the 20 samples analyzed presented contamination byAlternaria mycotoxins (tenuazonic acid, alternariol, alternariol methyl ether, altertoxin I and altertoxin II). Only aflatoxin B₁ was detected in 1 of the 20 samples analyzed, with a concentration of 0.25 ppb. Of the 40Aspergillus flavus strains isolated from oilseed rape samples, only 3 revealed aflatoxigenic capacity. None of thePenicillium spp. isolated from oilseed rape samples revealed mycotoxigenic capacity (citreoviridin, griseofulvin, citrinin, patulin and penicillic acid).     

A Survey on Distribution of Aspergillus Section Flavi in Corn Field Soils in Iran: Population Patterns Based on Aflatoxins, Cyclopiazonic Acid and Sclerotia Production

Soil isolates of Aspergillus section Flavi from Mazandaran and Semnan provinces with totally different climatic conditions in Iran were examined for aflatoxins (AFs; B and G types), cyclopiazonic acid (CPA) and sclerotia production. A total of 66 Aspergillus flavus group strains were identified from three species viz. Aspergillus flavus, Aspergillus parasiticus and Aspergillus nomius in both locations. A. flavus (87.9%) was found to be the prominent species followed by A. nomius (9.1%) and A. parasiticus (3.0%). Only 27.5% of A. flavus isolates were aflatoxigenic (B₁ or B₁ and B₂), out of which approximately 75% were capable to producing CPA. All the A. parasiticus and A. nomius isolates produced AFs of both B (B₁ and B₂) and G (G₁ and G₂) types, but did not produce CPA. Sclerotia production was observed in only 4 isolates of A. flavus among all 66 isolates from three identified species. A. flavus isolates were classified into various chemotypes based on the ability to produce aflatoxins and CPA. In this study, a new naturally occurring toxigenic A. flavus chemotype comprising of two strains capable of producing more AFB₂ than AFB₁ has been identified. A relatively larger proportion of aflatoxigenic A. flavus strains were isolated from corn field soils of Mazandaran province which indicate a possible relationship between high levels of relative humidity and the incidence of aflatoxin-producing fungi. The importance of incidence of Aspergillus section Flavi in corn field soils regard to their mycotoxin production profiles and crop contamination with special reference to climatic conditions is discussed.     

广陈皮外源真菌的组成及产毒真菌分析

[背景]广陈皮为药食同源中药材,在高温、高湿且贮存不当的条件下容易发霉,从而产生毒素,严重威胁陈皮的质量安全.[目的]分析广陈皮表面外源真菌的组成及其产生毒素的真菌.[方法]采用平板稀释法分离广陈皮表面外源真菌,利用分生孢子形态特征及DNA序列分析进行真菌鉴定,采用高效液相色谱-三重串联四极杆质谱联用技术对青霉属和曲霉...     

Fungi associated with dried fish in Sri Lanka

The mycoflora of dried-salted fish from markets in Kandy, Sri Lanka was studied with emphasis on visibly spoiled dried fish. A total of 61 fungal isolates from 25 dried-fish were isolated and identified. The most prevalent fungus wasAspergillus niger. Species ofAspergillus flavus, A. fumigatus, A. glaucus, A. restrictus, Aureobasidium spp.Basipetospora halophila (a genuinely halophilic fungus)Cladosporium herbarum, Gliomastix, spp.,Penicillium chalybeum andPenicillium expansum were present. The isolated fungi did not grow in synthetic media containing more than 30% sodium chloride.Aureobasidium spp. andGliomastix spp. did not grow on dried-fish under laboratory conditions. The protective exoskeleton appeared to prevent fungal growth on dried shrimp. TheA. flavus strains isolated were not aflatoxigenic.     

Evaluation of the mycological status of luncheon meat with special reference to aflatoxigenic moulds and aflatoxin residues

The luncheon meat samples analyzed, which were produced locally by the two main luncheon meat producing companies in Egypt were relatively highly contaminated either by moulds and yeasts in general, aflatoxigenic species and aflatoxin residues in particular. The most frequently encountered fungi from the samples were yeasts, Aspergillus niger, A. flavus, Penicillium chrysogenum, Rhizopus stolonifer, Mucor circinelloides. Less common were Cladosporium sphaerospermum, Alternaria alternata, Mycosphaerella tassiana, P. aurantiogriseum and P. oxalicum. The most important aflatoxigenic species, A. flavus, was isolated frequently. It was 10% of the total fungal isolates from both samples of the two companies. Seven luncheon meat samples out of 50 analyzed were positive for aflatoxin B₁ or B₁ and G₁, while all samples were negative for aflatoxins B₂, G₂, M₁ and M₂. Aflatoxin B₁ was detected only in 4 and 3 samples out of 25 analyzed from each of company A and B, respectively. The highest detectable level, 11.1 ppb, was recorded in a sample from company B and the least, 0.5 ppb, in a sample from company A. Aflatoxin G₁, at concentration of 3.2 ppb, was detected in only one sample of the aflatoxin B₁ – contaminated 3 samples of company B: this sample also had the highest level of aflatoxin B₁. Some luncheon meat samples had higher numbers of aflatoxigenic A. flavus than others, however these samples were negative for aflatoxins. The hazardous potential of such contamination will be discussed. This revised version was published online in June 2006 with corrections to the Cover Date.     

Biodegradation of organochlorine pesticide, endosulfan, by a fungal soil isolate, Aspergillus niger

Endosulfan is a chlorinated pesticide widely used in India for the protection of cotton, tea, sugarcane and vegetables. The persistence of endosulfan in environment and toxic effects on biota necessitate its removal. The role of soil fungi in recycling organic matter prompted us to attempt biodegradation of endosulfan using fungi. This study aims at enrichment, isolation and screening of fungi capable of metabolizing endosulfan. In all, 16 fungal isolates were obtained by enrichment of soil samples that had seems exposed to endosulfan before. Isolates were screened by a gradient plate assay, and results were confirmed by broth assay. On the basis of tolerance to endosulfan, an isolate, identified as Aspergillus niger was selected for further studies. The culture could tolerate 400 mg ml⁻¹ of technical grade endosulfan. Complete disappearance of endosulfan was seen on 12 days of incubation. Evolution of carbon dioxide during endosulfan metabolism has indicated the complete mineralization of endosulfan. Change in pH of culture broth to acidic range supported the biological transformation. Thin layer chromography (TLC) analyses revealed the formation of various intermediates of endosulfan metabolism including endosulfan diol, endosulfan sulfate, and an unidentified metabolite. The toxic intermediate, endosulfan sulfate, was also metabolized, further resulting in complete mineralization of endosulfan. Direct desulfurization of endosulfan sulfate or a novel pathway could be the mechanism of endosulfan and endosulfan sulfate degradation in Aspergillus niger. The fungal strain isolated by us could prove valuable for bioremediation of endosulfan contaminated soils and waters.     

Aspergillus nomius,a new aflatoxin-producing species related to Aspergillus flavus and Aspergillus tamarii

Aspergillus nomius is described and represents a new aflatoxigenic species phenotypically similar to A. flavus. Strains examined were isolated from insects and agricultural commodities. Separation from A. flavus is based on the presence of indeterminate sclerotia and a lower growth temperature. Comparisons of DNA relatedness show A. nomius to have only relatively recently evolved from A. flavus and A. tamarii.     

Fungi associated with food and feed commodities from Ecuador

Freshly harvested soybean, rice and corn from farms and corn-based pelleted feeds were collected from ranches from the coastal and mountain regions in Ecuador during 1998, and assessed for fungal contamination. The most prevalent fungi on pelleted feed were Aspergillus flavus and Fusarium graminearum. The prevalent fungi recovered from soybean were F. verticillioides, F. semitectum, Aspergillus flavus and A. ochraceus. In rice, F. oxysporum was the most prevalent toxigenic fungal species recorded, followed by F. verticillioides and A. flavus. In corn, F. verticillioides was the most prevalent fungus isolated in both the coastal and mountain regions, with high isolation frequencies of A. flavus and A. parasiticus at the coast. Based on the toxigenic species recovered, ochratoxin A may pose a contamination risk for soybean. A higher probability of aflatoxin contamination of corn was found in the coastal samples compared to those of the mountain region, while a risk of fumonisin contamination of corn exists in both regions.This revised version was published online in October 2005 with corrections to the Cover Date.     

Aerobiological,biochemical and immunological studies on some of the dominant <Emphasis Type="Italic">Aspergillus</Emphasis> species of South Assam (India)

Two years atmospheric survey of air-borne Aspergillus was carried out in the environmental conditions of South Assam. The survey revealed a total of 16 different species of Aspergillus with marked seasonal and annual variations. Aspergillus fumigatus was found to be the dominant atmospheric fungal species followed by Aspergillus flavus, Aspergillus niger, etc. Among the sample extracts tested, highest quantity of soluble protein was recorded in Aspergillus fumigatus (95.0 mg/g) whereas highest quantity of soluble carbohydrate (40.8 mg/g) and free amino acid (135.0 mg/g) was recorded in the sample extract of Aspergillus niger per gram of dry weight, respectively. The highest numbers of protein polypeptide bands were detected in the sample extract of Aspergillus fumigatus followed by Aspergillus flavus and lowest in Aspergillus niger. The maximum numbers of immunoglobulin E binding protein fractions were found in Aspergillus fumigatus, followed by Aspergillus flavus, Aspergillus clavatus, etc.     

Comparison of Aflatoxigenic and Nonaflatoxigenic Isolates of Aspergillus flavus using DNA Amplification Fingerprinting Techniques

Aspergillus flavus is a filamentous fungus that produces mycotoxins in many food and feed crops, such as maize (Zea mays L.). Isolates were analyzed for toxin production by nucleic acid profiles in an attempt to differentiate aflatoxigenic from nonaflatoxigenic isolates. A total of 41 aflatoxigenic and 34 nonalfatoxigenic isolates were included in the study. The isolates were evaluated initially using DNA amplification fingerprinting (DAF) without clear resolution of the groups. A weak association of aflatoxigenic isolates was observed, as evidenced by their clustering in 18 of 81 trees recovered from maximum parsimony analysis of binary characters derived from arbitrary signatures from amplification profiles (ASAP) data; nonaflatoxigenic isolates exhibited a pattern of paraphyletic laddering. Up to five markers unambiguously supported the aflatoxigenic isolate grouping, but the presence of alternative conflicting topologies in equally parsimonious trees precluded the observation of meaningful statistical support. With additional markers for genome of A. flavus, this method could be used to resolve toxigenic from nontoxigenic strains. This additional work could resolve aflatoxigenic isolates of A. flavus present on maize plants using ASAP, which would reduce labor intense costs and potentially lead to faster determination of resistant cultivars in breeding efforts.     

Characterization of the pyruvate kinase-encoding gene (pki1) of Trichoderma reesei

The pyruvate kinase-encoding gene (pki1) from Trichoderma reesei was isolated by hybridization to the corresponding Aspergillus nidulans pkiA gene. The 1614-bp nucleotide (nt) sequence of the cloned gene codes for a 538-amino-acid protein. The coding sequence contains a single intron of 246 nt at a position identical to that of intron E in the A. nidulans gene. The PKI protein shows extensive homology to the PKIs of A. nidulans and A. niger (67%) and Saccharomyces cerevisiae (59%). The 5' non-coding sequence contains a number of motifs typical for yeast glycolytic genes, but so far only rarely found in filamentous fungi.     

Microbiological and physicochemical factors affecting <Emphasis Type="Italic">Aspergillus</Emphasis> section <Emphasis Type="Italic">Flavi</Emphasis> incidence in Cavendish banana (<Emphasis Type="Italic">Musa cavendishii</Emphasis>) chips production in Southern Philippines

Microbiological and physicochemical factors affecting the incidence of Aspergillus section Flavi in dried Cavendish banana (Musa cavendishii) chips production in Southern Philippines were examined. The average counts of Aspergillus section Flavi (AFC) in fresh and dried Cavendish bananas from 10 production batches of the Philippine Agro-Industrial Development Cooperative in Davao del Norte, Southern Philippines were 1.2 × 10² and 1.6 × 10² cfu/g, respectively. Isolates from both samples were identified to be Aspergillus flavus based on spore type and conidial structure of isolates. An increasing trend in the AFC of Cavendish bananas was observed during dried banana chips processing. Variability in the AFC between production batches was attributed to differences in aerobic and fungal populations and physicochemical characteristics of the fruits, peel damage of the raw materials, concentration of AFC in the air and food-contact surfaces of the production area, and temperature and relative humidity (RH) conditions of the environment during production and storage. Physicochemical characteristics of Cavendish bananas from the receipt of raw materials up to the first day of drying were within the reported range of values allowing growth and toxin production by aflatoxigenic fungi. Air-borne AFC varied depending on the section of the production area examined. The close proximity of the waste disposal area from the production operation to the preparation, drying and storage areas suggests that cross-contamination, probably air-borne or insect-borne was a likely occurrence. The hands of workers were also identified as AFC sources. Results of this study highlight the need for the development of strategies to control aflatoxigenic fungi and aflatoxin contamination in Philippine dried Cavendish bananas.     

Dominant fungi in the rhizosphere of established tea bushes and their interaction with the dominant bacteria under in situ conditions

Species of Penicillium and Trichoderma were found to dominate the rhizosphere of established tea bushes in a detailed study conducted from various tea growing locations in India. Penicillium erythromellis, P. janthinellum, P. raistrickii, Trichoderma pseudokoningii and T. koningii were found to be closely associated with tea roots. While seasonal fluctuation was observed in the case of Penicillium spp., the population of Trichoderma spp. showed less variation during the year. Both species were sensitive to low temperatures. In general, fungi associated with the tea rhizosphere were found to prefer a mesophillic temperature range (15 °C to 35 °C). The dominant species of Penicillium and Trichoderma also exhibited tolerance to lower temperatures, i.e., 5 to 10 °C on agar plates. Most fungi were able to grow in a wide range of pH (4 to 12). Lowering of soil pH in the rhizosphere of tea bushes was positively correlated with the age of the bush and may have affected the development of a specific microbial community in the rhizosphere.The populations of Penicillium and Trichoderma species were inversely correlated with the populations of two most dominant rhizosphere bacteria, Bacillus subtilis and B. mycoides. Both Bacillus species have been shown to have antagonistic activity against these two fungi under in vitro conditions. The present study demonstrates the existence of a similar antagonism under in situ conditions in the rhizosphere of established tea bushes.     

黄曲霉功能基因组学研究

黄曲霉（Aspergillus flavus）是一种常见的腐生真菌和条件致病菌,其次生代谢产物黄曲霉毒素（Aflatoxin,AFT）具有高度的致癌性和致畸性,严重危及人类和动物健康。近年来,功能基因组学研究发展迅速,在真菌生长发育、挖掘真菌次级代谢产物以及研究包括黄曲霉毒素在内的真菌毒素等方面得到了广泛的应用。功能基因组学在研究黄曲霉与宿主之间的相互作用以及黄曲霉与其他曲霉之间的相互作用方面具有巨大的潜力。然而,黄曲霉功能基因组学受到细胞壁难以破除、耐药性高、筛选标记少、缺陷型菌株构建费力耗时等因素的影响而发展缓慢。概述了黄曲霉的选择标记、遗传转化方法和黄曲霉毒素以及环匹阿尼酸（cyclopiazonic acid, CPA）生物合成的研究进展,并讨论了在提高黄曲霉基因操作效率方面的潜在策略。例如,构建缺乏非同源末端连接（NHEJ）途径的菌株、Cre-loxP重组系统、CRISPR-Cas9等方法,为深入开展黄曲霉遗传学研究提供参考。     

Survey of Vietnamese Peanuts,Corn and Soil for the Presence of <Emphasis Type="Italic">Aspergillus flavus</Emphasis> and <Emphasis Type="Italic">Aspergillus parasiticus</Emphasis>

Aspergillus flavus and Aspergillus parasiticus cause perennial infection of agriculturally important crops in tropical and subtropical areas. Invasion of crops by these fungi may result in contamination of food and feed by potent carcinogenic aflatoxins. Consumption of aflatoxin contaminated foods is a recognised risk factor for human hepatocellular carcinoma (HCC) and may contribute to the high incidence of HCC in Southeast Asia. This study conducted a survey of Vietnamese crops (peanuts and corn) and soil for the presence of aflatoxigenic fungi and used microsatellite markers to investigate the genetic diversity of Vietnamese Aspergillus strains. From a total of 85 samples comprising peanut (25), corn (45) and soil (15), 106 strains were isolated. Identification of strains by colony morphology and aflatoxin production found all Vietnamese strains to be A. flavus with no A. parasiticus isolated. A. flavus was present in 36.0% of peanut samples, 31.1% of corn samples, 27.3% of farmed soil samples and was not found in virgin soil samples. Twenty-five per cent of the strains produced aflatoxins. Microsatellite analysis revealed a high level of genetic diversity in the Vietnamese A. flavus population. Clustering, based on microsatellite genotype, was unrelated to aflatoxin production, geographic origin or substrate origin.     

Occurrence of Ochratoxin A-Producing Fungi in Commercial Corn Kernels in Argentina

Potentially ochratoxigenic Aspergillus and Penicillium species were identified and the natural occurrence of ochratoxin A (OTA) in corn kernels was evaluated. Likewise, the capacity to produce OTA by Aspergillus section Nigri and Circumdati was investigated. A total of 50 corn samples for human consumption was collected in the south of Córdoba Province. The surface-disinfected method for mycobiota determination was used. The OTA detection was performed by HPLC. OTA production was tested in strains belonging to section Nigri and Circumdati. Statistical analysis demonstrated that the specie A. flavus was isolated in higher frequency (p<0.01) from corn kernels in DRBC and DG18 media. The percentage of corn kernels contaminated by A. niger var. niger was similar in DRBC and DG18 media. The frequency of grains contaminated by A. flavus and A. niger var. awamori was higher than A. niger var. niger and A. japonicus var. japonicus (p<0.01) in DG18 media. The other potentially ochratoxigenic species, A. ochraceus, was isolated between 5% and 10% of the corn kernels in DG18 and DRBC media, respectively. The OTA producing species P. verrucosum was not isolated. All samples of corn were OTA negative (<1 ng g⁻¹). Thirty strains (25%) of the black Aspergillus were OTA producers. From four strains of A. ochraceus isolated, only one produced OTA. Due to the storage variable conditions could not be adequate in this substrate, the presence of ochratoxigenic strains of section Nigri and OTA needs to be evaluated for a longer time to establish the toxicological risk for human beings. The contamination of stored corn kernels with A. flavus and Aspergillus section Nigri was significant.     

Discrepancies between MIC and MLC values of amphotericin B against isolates ofAspergillus species

There is little information addressing the phenomena of discrepancy between minimal inhibitory concentrations (MIC) and minimal lethal concentrations (MLC) values of amphotericin B (AMB) to clinical isolates of fungi. This study assessed in vitro activity of AMB against 70 clinical isolates of aspergilli: 30 strains ofAspergillus fumigatus, 20 strains ofAspergillus flavus and 20 strains ofAspergillus niger. Susceptibility tests were accomplished using a macro broth dilution procedure, with special emphasis on ascertainment of MLCs. AMB exhibited low MIC values against all clinical isolates. While we did not identify any AMB resistant isolates among 70Aspergillus spp. studied as judged by MIC levels, analysis of the data demonstrated a clear discrepancy between the MIC and MLC levels of AMB obtained against clinical isolates ofAspergillus spp. The MLC values of AMB were significantly higher than the MIC values with MIC 50 and MIC 90 of 0.29 and 0.5 µg/ml, respectively, at the second reading time, and MLC 50 and MLC 90 of 2.31 and 9.24 µg/ml, respectively (p<0.001). Additionally, minimal lethal concentrations in 36/70 (51%) of aspergillal isolates studied produced drug concentrations above those which can usually be sustained in patient plasma or tissue.     

Aspergillus on tree nuts: incidence and associations

California exports tree nuts to countries where they face stringent standards for aflatoxin contamination. Trade concerns have stimulated efforts to eliminate aflatoxins and Aspergillus flavus from almonds, pistachios and walnuts. Incidence of fungi on tree nuts and associations among fungi on tree nuts were studied. Eleven hundred pistachios, almonds, walnuts and brazil nuts without visible insect damage were plated on salt agar and observed for growth of fungi. Samples came both from California nut orchards and from supermarkets. To distinguish internal fungal colonization of nuts from superficial colonization, half the nuts were surface-sterilized before plating. The most common genera found were Aspergillus , Rhizopus and Penicillium . Each species of nut had a distinct mycoflora. Populations of most fungi were reduced by surface sterilization in all except brazil nuts, suggesting that they were present as superficial inoculum on (rather than in) the nuts. In general, strongly positive associations were observed among species of Aspergillus ; nuts infected by one species were likely to be colonized by other species as well. Presence of Penicillium was negatively associated with A. niger and Rhizopus in some cases. Results suggest that harvest or postharvest handling has a major influence on nut mycoflora, and that nuts with fungi are usually colonized by several fungi rather than by single species. This revised version was published online in June 2006 with corrections to the Cover Date.