The action mechanism and biocontrol potentiality of novel isolates of Saccharomyces cerevisiae against the aflatoxigenic Aspergillus flavus.

Inhibition of Aspergillus flavus growth and its aflatoxins production using the biocontrol agent Saccharomyces cerevisiae as well as to explore its mode of action was studied. Eight strains of S. cerevisiae strains were able to suppress the growth of A. flavus Z103. The maximum growth inhibition of A. flavus Z103 was obtained by living cells of S. cerevisiae EBF101 and S. cerevisiae 117 with 85 and 83%, respectively. The sporulation inhibition and hyphae deterioration of A. flavus Z103 by S. cerevisiae cells adhesion were observed under SEM; up to 99·8% inhibition of aflatoxins biosynthesis by A. flavus Z103 was resulted when the fungus was treated by autoclaved extracellular crude of S. cerevisiae. Also, the tested strains are potential to produce exo-chitinase which could be suggested as another mode of action for its antifungal activity. GC-MS analysis of S. cerevisiae 117 extracellular secondary metabolites revealed the existence of 4-Hydroxyphenethyl alcohol (46·32%), 4, 4-Dimethyloxazole (9·14%) and 1,2-Benzenedicarboxylic acid dioctyl ester (2·8%). Significance and Impact of the Study: The use of Saccharomyces cerevisiae instead of chemical preservatives in fermented food, animal and fish feed and storage cereal grains could encourage the food industry to produce organic food free of chemical additives. Overall, our data suggest the possibility of using S. cerevisiae as an alternative treatment in the food industries to control the dispersion and aflatoxins production by Aspergillus flavus during storage. This method could provide an additional probiotic effect in the digestive tract of consumers after ingestion of the treated food. So, our study clarifies the exact mechanisms responsible for the reduction of the aflatoxin contents by S. cerevisiae.

Extracellular mycosynthesis of gold nanoparticles using Trichoderma hamatum: optimization, characterization and antimicrobial activity.

Synthesis of gold nanoparticles (AuNPs) has become a needed domain of applied science. Biological method for synthesis of AuNPs by Trichoderma hamatum SU136 aqueous mycelial extract was achieved. The culture filtrate of the fungus was exposed to three different concentrations of gold chloride. The culture filtrate of the fungus was exposed to three different concentrations of gold chloride (0·25, 0·5 and 1·0 mmol l-1 ). In all cases, the gold ions (Au3+ ) were reduced to Au0 , leading to the formation of stable AuNPs. The AuNPs were identified by UV-visible spectrometry, TEM and FT-IR. The presence of a surface plasmon band around 530 nm indicates AuNPs synthesis. Trichoderma hamatum SU136 synthesized 5-30 nm sized; spherical, pentagonal and hexagonal morphologies of AuNPs by TEM. The existence and binding of proteins with nanoparticles was approved by FT-IR study. Parameters optimization showed the smallest size of AuNPs was obtained with (0·5 mmol l-1 gold chloride, pH 7 at 38°C). Interestingly, AuNPs exhibited antimicrobial activity against four pathogenic bacterial strains in the presence of the standard antibiotic, streptomycin. SIGNIFICANCE AND IMPACT OF THE STUDY: Mycosynthesis of AuNPs by Trichoderma hamatum would provide some useful data for oriented biosynthesis of AuNPs. In addition, the applications of mycosynthesized AuNPs were studied against some pathogenic bacteria. Therefore, the gained results detect that these antimicrobial nanoparticles could be explored as hopeful candidates for a variety of biomedical and pharmaceutical applications. This study should provide a further prudence for the fungal-mediated synthesis of AuNPs.

Progesterone transformation as a biochemical aid in classification of the genus Emericella.

A total of 65 isolates representing 13 Emericella taxa (5 isolates of each of 12 species and 1 variety) had the ability to transform progesterone into 11 alpha-hydroxyprogesterone. A systematic variation could be observed between the different tested Emericella taxa with respect to the transformation products. The isolates were divided according to the transformation types into six groups: (1) progesterone was hydroxylated into 6 beta-hydroxyprogesterone, 11 alpha-hydroxyprogesterone and 6 beta,11 alpha-dihydroxyprogesterone--found in Emericella acristata and E. dentata; (2) E. aurantio-brunna and E. parvathecia hydroxylated progesterone into 11 alpha-hydroxyprogesterone, 17 alpha-hydroxyprogesterone and 11 alpha,17 alpha-dihydroxyprogesterone; (3) E. nidulans and E. quadrilineata formed the same three products as members of group (2) and form epicortisol; (4) E. nidulans var. lata, E. bicolor and E. variecolor transformed progesterone into a mixture of mono-, di- and trihydroxy products; (5) E. striata and E. sublata exhibited an oxidative splitting of the progesterone side chain in position C-17 and hydroxylated progesterone into mono- and dihydroxy products; (6) E. rugulosa and E. unguis had the ability to degrade progesterone side-chain and to hydroxylate it into mono-, di- and trihydroxy products. This biochemical differentiation may supplement the morphological and other biochemical criteria used in the classification of the Emericella taxa.

Progesterone side-chain degradation by some species of Aspergillus flavus group.

Seventy isolates belonging to 6 species and one variety of A. flavus group were shown to degrade the progesterone side-chain to yield delta 4-androstene-3,17-dione and testosterone. The isolates of five species (A. flavo-furcatis, A. flavus, A. oryzae, A. parasiticus and A. tamarii) possessed enzyme systems catalyzing the opening of ring D and formed testololactone as final steroid metabolite in addition to their ability to produce the above mentioned two products. 11 beta-Hydroxy-delta 4-androstene-3,17-dione was formed by only A. flavus and A. tamarii while 11 beta-hydroxytestosterone was produced by A. flavo-furcatis, A. parasiticus and A. subolivaceus. The chromatographic resolution of the mixture products obtained (when the selective isolate of each species reacted with 1 g of progesterone) revealed that 60-75% of progesterone was converted into delta 4-androstene-3,17-dione (8-30%), testosterone (7-33%), testololactone (14-37%) and other products (3-40%). The most bioconversion activity was exhibited by A. oryzae, followed by A. parasiticus. The highest values of delta 4-androstene-3,17-dione (30% of added progesterone) and testosterone (33%) were formed by A. flavus var. columnaris while those of testololactone (37%) were produced by A. oryzae. A systematic variation could be observed between the different tested species of A. flavus group with respect to the transformation reactions of progesterone. Comparative biotransformation results showed that essential differences exist between the tested species in this group; this biochemical differentiation may supplement the morphological and other physiological criteria used in the identification of the different species in the A. flavus group.

Effect of selenite and tellurite on the morphological growth and toxin production of Aspergillus parasiticus var. globosus IMI 120920.

Aspergillus parasiticus var. globosus IMI 120920 was able to grow in presence of different concentrations tested (0.052-4.0%) of sodium selenite or concentrations up to 2.0% potassium tellurite. Growth of the fungus was decreased greatly by the increase of metals concentrations. Dark colour colony and black reverse were formed in presence of tellurite while reddish gray to grayish red colony colour and brownish red to orange red reverse were formed in presence of selenite. The fungal biomass was slightly decreased at lower concentrations and highly inhibited at higher concentrations of selenite or tellurite. Selenite slightly stimulated aflatoxin formation at lower concentrations and highly inhibited it at higher concentrations. Aflatoxin production was decreased greatly by increasing tellurite concentrations. Obvious malformations were observed in the morphological features of the fungus in presence of different levels of selenite or tellurite.

Aflatoxin formation and varietal difference of cow pea (Vigna unguiculata (L.) Walp.) and garden pea (Pisum sativum L.) cultivars.

Different cultivars of cow pea and garden pea seeds were surveyed for susceptibility or resistance towards the toxigenic and aflatoxin-producing mould (Aspergillus flavus IMI 102135). The results show that aflatoxin production varied among the different cultivars of both cow pea and garden pea. Morphological and histological characters of the different cultivars tested did not show any relation between colour, shape and size of seeds and the amount of aflatoxin produced. The chemical analysis of the different constituents obtained from both seed coats and seed kernels with susceptible, partially resistant and resistant cow pea and garden pea cultivars revealed that the resistant cultivars of cow pea (namely: Balady cultivar) and garden pea (namely: Melting Sugar cultivar) contained lower levels of sodium and higher levels of phosphate and potassium.

Transformation reactions of progesterone by different species of Streptomyces.

A systematic study of transformation reactions of the genus Streptomyces with respect to the progesterone molecule was undertaken. The types of transformation reactions by different Streptomyces species were evaluated from the point of view of taxonomy. The isolates tested were divided according to the transformation types into six groups: (1) a group of species transform progesterone to 16 alpha-hydroxyprogesterone; (2) a group of species transform progesterone to 6 beta-hydroxyprogesterone; (3) a group of species transform progesterone to 6 beta, 11 alpha-dihydroxyprogesterone; (4) a group of species dehydrogenate progesterone in C1-2 position; (5) a group of species transform progesterone to 3 derivatives namely 6 beta-hydroxyprogesterone, 6 beta, 11 alpha-dihydroxyprogesterone and dehydrogenation in position C1-2; (6) a group of species with no capacity to transform progesterone into another steroid derivative. From the point of view of Streptomyces classification, the transformation reaction of progesterone fulfil all the requirements of taxonomic feature of Streptomyces. These appear to be specific properties and common to all the strains of individual Streptomyces species tested.

New sources of cytochalasins A and B from dematiaceous hyphomycetes.

One hundred isolates of 27 species belonging to 13 genera of dematiaceous hyphomycetes were screened for production of cytochalasins A and B. Most of these isolates (94) were obtained from Assiut University Culture Collection, Botany Department, Faculty of Science, Assiut University, Egypt; three isolates from CBS, The Netherlands; two isolates from DSM, Germany; and one isolate from IMI, UK. The results revealed that 10 isolates of six species representing five genera of fungi produced cytochalasins A and/or B. These species are Alternaria chlamydospora, Cochliobolus spicifer, Diplococcum spicatum, Phoma herbarum, Phoma multipora and Setosphaeria rostrata. This is the first report for the production of cytochalasins A and/or B by these species of dematiaceous hyphomycetes.

Based on biochemical and physiological behavior, where is Aspergillus egyptiacus better placed?

Physiological and biochemical properties were tested in 45 isolates of Aspergillus egyptiacus (16 isolates), Emericella nidulans (16) and Aspergillus versicolor (13). The three fungal species exhibited common and similar features. The big similarity between A. egyptiacus and E. nidulans was greater than between A. egyptiacus and A. versicolor. It included the inability to produce base either from sodium citrate or lactic acid media, growth at 45 degrees C (thermophilicity), and production of very similar pigmentations on Aspergillus flavus and parasiticus agar. A. egyptiacus is therefore better placed in the Aspergillus nidulans-Emericella assemblage.

Fungal flora and mycotoxins of six kinds of nut seeds for human consumption in Saudi Arabia.

A wide range of moulds representing several genera and species, was recorded in this study from 5 seed samples of each almond, cashew nut, chestnut, hazelnut, pistachio nut and walnut collected from different markets in Ar' Ar, Saudi Arabia. The total counts of fungi were widely fluctuated between 1960-7704 and 1948-7434 colonies/g dry seeds on glucose-Czapek's and glycerol agar media at 28 degrees C, respectively, and represented twenty genera, 53 species and 2 varieties of fungi. The prevalent fungi on the 2 agar media were Aspergillus flavus, A. niger and Penicillium chrysogenum. On glucose-Czapek's agar, Rhizopus stolonifer and Aspergillus flavus var. columnaris were isolated from all 6 kinds of nut, A. parasiticus from 5 kinds and A. fumigatus from 4 kinds with high frequencies. Eurotium species were completely absent on glucose-Czapek's agar but they were isolated in high frequency from all kinds of nut on glycerol agar medium. The different nut samples were analyzed by thin layer chromatography for the presence of aflatoxins B1, B2, G1 & G2, citrinin, ochratoxins, patulin, sterigmatocystin, diacetoxyscirpenol, T-2 toxin and zearalenone. Aflatoxins B1 & G1 were detected in 3 out of the 5 samples tested of chestnut at concentrations ranging between 20 to 60 micrograms/kg. All other samples of almond, cashew nut, hazelnut, pistachio nut, and walnut that were analyzed were mycotoxin free.

Filamentous fungi and mycotoxin detected in coconut.

Fifty-nine species and one variety belonging to 25 genera of fungi were isolated from 25 coconut samples on glucose-Czapek's (25 genera and 55 species + 1 variety) and dichloran-glycerol (8 genera and 32 species + 1 variety) agar media at 28 degrees C. The common fungi on both media used were Aspergillus flavus, A. niger, Penicillium chrysogenum and Cladosporium cladosporioides. On glucose-Czapek's agar, A. flavus var. columnaris, P. oxalicum, Alternaria alternata, Rhizopus stolonifer and Trichoderma hamatum were recorded as common fungi while A. sydowii and Eurotium chevalieri were isolated with high frequency only on dichloranglycerol medium. Chromatographic analysis of the chloroform extracts of the coconut samples revealed that 5 out of 25 samples tested were naturally contaminated with aflatoxin B1 (15-25 micrograms/kg) and 3 samples contaminated with ochratoxin A (50-205 micrograms/kg).

Survey of mycoflora and mycotoxins of some dried fruits in Egypt.

Fifty-five species and two varieties appertaing to 23 genera were collected and identified from 4 samples of dried fig and 3 dried samples of each of apricot, plum and raisin. Forty-nine species and two varieties belonging to 20 genera were isolated on 1% glucose-Czapek's while 31 species and one variety belonging to 16 genera were isolated on 40% sucrose-Czapek's agar medium at 28 +/- 2 degrees C. Penicillium, Aspergillus and cladosporium were the common genera on the two types of media used. Alternaria and Pleospora were common on 1% glucose-Czapek's agar only while the genus of Eurotium was isolated with high occurrence on 40% sucrose-Czapek's agar. The common species on the two types of media used were Penicillium chrysogenum, Aspergillus niger and cladosporium cladosporioides. On 1% glucose-Czapek's agar, P. aurantiogriseum, A. fumigatus, A. flavus, Alternaria tenuissima and Pleospora herbarum were isolated with high occurrence while A. versicolor, A. wentii, Eurotium amstelodami and E. chevalieri were common on 40% sucrose-Czapek's agar. The different dried fruit samples were assayed for the natural occurrence of aflatoxins B1, B2, G1 & G2, citrinin, ochratoxins, patulin, sterigmatocystin, diacetoxyscirpenol, T-2 toxin and zearalenone by thin layer chromatographic analysis. Ochratoxin A was detected in all samples tested of apricot (50-110 micrograms/kg), fig (60-120 micrograms/kg) and plum (210-280 micrograms/kg). The other mycotoxins under investigation were not detected. All samples examined of raisin proved to be mycotoxin free.

Mycotoxin production on different cultivars and lines of broad bean (Vicia faba L.) seeds in Egypt.

One hundred different cultivars and lines of broad bean (Vicia faba L.) seed samples were inoculated with Aspergillus flavus Link (CMI 102135) to determine varietal differences which may support or resist aflatoxin production. Thin-layer chromatographic analysis of the chloroform extracts of the different seed samples revealed that 11 cultivars/lines were highly resistant to seed invasion and aflatoxin production while 9 cultivars/line showed partial resistance. The remaining 80 samples were susceptible to the establishment of A. flavus and aflatoxin accumulation. All the resistant cultivars/lines seed samples were inoculated also with three local isolates of fungi namely; Stachybotrys chartarum (Ehrenb. ex Link) Hughes, Aspergillus ochraceus Wilhelm, and Fusarium oxysporum Schlecht. The resistant seed samples were also resistant for colonization with these fungi and mycotoxin formation.