Activity of essential oil and its major compound, 1,8-cineole, from Eucalyptus globulus Labill., against the storage fungi Aspergillus flavus Link and Aspergillus parasiticus Speare

The essential oil from leaves of Eucalyptus globulus obtained by hydrodistillation, as well as its major compound 1,8-cineole, identified by gas chromatography coupled with a mass selective detector, were evaluated for their effectiveness against the storage fungi Aspergillus flavus and Aspergillus parasiticus. The evaluation was performed by compound dissolution in yeast extract sucrose (YES) medium and exposure to headspace volatiles. Complete fungal growth inhibition of both species was achieved with the essential oil by contact and volatile assays. Volatile exposure showed total inhibition at the lower level tested of 500 μL. The 1,8-cineole tested alone showed partial inhibition only at the highest level of 1.3492 μL. Aflatoxin B₁ production was reduced in headspace volatile assays and partial inhibition was observed at the 200 μL dose of the essential oil.     

Inhibitory effect of the essential oil of Curcuma longa L. and curcumin on aflatoxin production by Aspergillus flavus Link

Aflatoxins are highly toxic, mutagenic, teratogenic and carcinogenic mycotoxins. Consumption of aflatoxin-contaminated food and commodities poses serious hazards to the health of humans and animals. Turmeric, Curcuma longa L., is a native plant of Southeast Asia and has antimicrobial, antioxidant and antifungal properties. This paper reports the antiaflatoxigenic activities of the essential oil of C. longa and curcumin. The medium tests were prepared with the oil of C. longa, and the curcumin standard at concentrations varied from 0.01% to 5.0%. All doses of the essential oil of the plant and the curcumin standard interfered with mycotoxin production. Both the essential oil and curcumin significantly inhibited the production of aflatoxins; the 0.5% level had a greater than 96% inhibitory effect. The levels of aflatoxin B₁ (AFB₁) production were 1.0 and 42.7 μg/mL, respectively, for the samples treated with the essential oil of C. longa L. and curcumin at a concentration of 0.5%.     

Antifungal potential of some natural products against Aspergillus flavus in soybean seeds during storage

The inhibitory effect of cow-dung fumes, captan, leaf powder of Withania somnifera, Hyptis suaveolens, Eucalyptus citriodora, peel powder of Citrus sinensis, Citrus medica and Punica granatum, neem cake and pongamia cake on the growth of Aspergillus flavus in soybean seeds during storage was investigated. Soybean seed was treated with different natural products and the fungicide captan and was stored at ambient conditions for 6 months. Seed samples were withdrawn at monthly intervals and the incidence of seed-borne A. flavus and percentage germination of the seed was determined. Captan, neem cake, pongamia cake and peel powder of C. sinensis reduced the incidence of A. flavus. Leaf powder of W. somnifera, H. suaveolens, E. citriodora and peel powder of P. granatum also checked the frequency of A. flavus. All treatments maintained a high germination percentage of the soybean seeds over a storage period of 6 months. These natural products may be alternatives to chemical fungicides and provide an easy method to protect soybean and other agricultural commodities from A. flavus in storage.     

Effect of Capsicum carotenoids on growth and aflatoxins production by Aspergillus flavus isolated from paprika and chilli

The aim of this study was to determine the effect of a carotenoid mixture (Capsantal FS-30-NT), containing capsanthin and capsorubin, on growth and aflatoxins (AF) production of AF-producing Aspergillus flavus isolates.     

Characterization of nonochratoxigenic strains of Aspergillus carbonarius from grapes

Aspergillus carbonarius is the main responsible source of ochratoxin A (OTA) in food commodities such as wine, grapes or dried vine fruits from main viticultural regions worldwide. Besides, OTA production is a very consistent property of this species and for this reason atoxigenic isolates of A. carbonarius are very rarely found in natural environments. In the present study, for the first time, three nonochratoxigenic wild strains of A. carbonarius have been discovered, unambiguously identified, characterized in deep and compared to ochratoxigenic strains of the same species. In addition, polyketide synthase (pks) genes suggested to be involved in OTA biosynthesis were also screened in these strains. The identification of the strains was confirmed by ITS-5.8S rRNA, β-tubulin and calmodulin gene sequencing. The three atoxigenic strains did not produce OTA in a conducive culture medium at any of the temperatures and times of incubation tested. Five ketosynthase domains from pks genes previously described in A. carbonarius were detected both in ochratoxigenic and in nonochratoxigenic strains. Atoxigenic strains of A. carbonarius could be useful as biotechnological agents to be used in food industry and as biological agents for control of OTA production in vineyards and other crops.     

Integrated management of insect vectors of Aspergillus flavus in stored maize, using synthetic antioxidants and natural phytochemicals

The purpose of this study was to investigate the insecticidal activity of two benzoic acids 2(3)-tert-butyl-4 hydroxyanisole (BHA) and 2,6-di(tert-butyl)-p-cresol (BHT); two phenolic acids 3-phenyl-2-propenoic acid (CA) and trans-4-hydroxy-3-methoxycinnamic acid (FA) and two essential oils of Eugenia caryophyllata (clove tree) and Thymus vulgaris (thyme) against Sitophilus zeamais, Tribolium confusum and Rhyzopertha dominica, vector carriers of aflatoxigenic fungi in stored maize. The susceptibility of insects, the frequency of isolation of Aspergillus section Flavi in insects and maize, and the analysis of aflatoxin B₁ in maize were determined. BHA, BHT, BHA/BHT mixture and the natural phytochemicals AF and AF/AC mixture showed the highest insecticidal activity against S. zeamais, T. confusum and R. dominica after 120 days of incubation. The insecticidal efficacy of the volatile fraction of essential oils of clove and thyme showed less inhibition. There was no contamination of Aspergillus section Flavi in dead and live insects collected from maize treated with BHA. No aflatoxin B₁ accumulation was detected in the control and treatments. The information obtained shows that these substances have the potential to control pest insect vectors of aflatoxigenic fungi in stored maize in microcosms during 120 days.     

Modelling the effect of temperature and water activity on the growth of Aspergillus parasiticus on irradiated Argentinian flint maize

A full factorially designed experiment including storage temperature (10, 20, 30 and 37 °C) and water activity (0.88, 0.92 and 0.96) was undertaken to study the growth of Aspergillus parasiticus in maize samples. Kinetic parameters such as specific growth rate (μ), lag phase duration and maximum logarithmic increase were determined by fitting the Modified Gompertz equation to the viable mould count data (N in CFU/g) as a function of time collected in twelve experiments. The average coefficient of determination (R²) was 0.987, being the mean standard deviation of the estimate of 0.216 in units of log₁₀N. In the practical range of 10-30 °C, the relationship of the three kinetic parameters with temperature was described by second order polynomial expressions, whose parameters, in turn, depended on water activity. The combined or full model i.e., the Modified Gompertz model with its parameters expressed as a function of temperature and water activity, was able to predict log₁₀N with an average percentage error of 4.3, so agreement with the experimental data was highly satisfactory.In a simulation exercise, the full model was able to predict the viable mould count, given an initial value and grain temperature and water activity histories, with promising results for maize storage.     

A laboratory assessment of the potential of different strains of the entomopathogenic fungi Beauveria bassiana (Balsamo) Vuillemin and Metarhizium anisopliae (Metschnikoff) to control Callosobruchus maculatus (F.) (Coleoptera: Bruchidae) in stored cowpea

Twelve indigenous and exotic isolates of Beauveria bassiana and Metarhizium anisopliae were evaluated for their virulence and their ability to suppress populations of Callosobruchus maculatus in stored cowpea. LT₅₀ values ranged from 3.11 to 6.12 days following immersion in aqueous suspensions containing 1×10⁸ conidia ml⁻¹. Indigenous isolates that had been recovered from C. maculatus were more virulent in laboratory bioassays than exotic isolates from other insects. The two isolates with the shortest LT₅₀ values were compared in dose-response assays by immersion and by exposure to cowpea grains treated with dry conidia. In both assays B. bassiana 0362 was consistently more virulent than M. anisopliae 0351. By immersion, LC₅₀ values on day 6 post-treatment were 9.10×10⁴ and 7.10×10⁵ conidia ml⁻¹ for B. bassiana 0362 and M. anisopliae 0351, respectively. Exposure to treated grains gave LC₅₀ values of 1.15×10⁷ and 4.44×10⁷ conidia g⁻¹ grain for B. bassiana 0362 and M. anisopliae 0351, respectively. In 1 kg batches of cowpea stocked with 50 adult C. maculatus, B. bassiana 0362 at both 1×10⁷ and 1×10⁸ conidia g⁻¹ grain led to significant adult mortality and reduced F₁ emergence relative to untreated populations. At 1×10⁸ conidia g⁻¹ the effect of the fungus persisted into the F₁ generation. The net reproductive rates, R₀, measured 26 days after insects were released were 5.16 and 7.32 for the high and low doses compared to 9.52 for the untreated control.     

Peanut antioxidants: Part 1. Genotypic variation and genotype-by-environment interaction in antioxidant capacity of raw kernels

The potential to select for antioxidant traits in peanut (Arachis hypogaea L.) kernels by conventional plant breeding was investigated by oxygen radical absorbance capacity (ORAC) assay of (a) 32 full-season maturity genotypes grown at Kairi Research Station in 2008/09; and (b) a subset of ten genotypes with diverse antioxidant capacities grown in five different production regions in Queensland in 2009/10 and 2010/11. There were significant levels of variation of more than 25% relative standard deviation (RSD) in antioxidant capacity among the 32 tested genotypes; and that genotype, environment, and genotype-by-environment (G × E) interaction all significantly (P < 0.05) affected trait expression. Analysis of the G × E data confirmed the heritability of antioxidant capacity, which has not been previously quantified, whereby genotype explained 44% of the phenotypic variation on a plot basis and 82% on an entry mean basis. This indicated that there was substantial genetic control of antioxidant capacity in peanut kernels, but also that it will be important to characterize environmental interaction to enable plant/seed selection in the Breeding Program. The research also highlighted the utility of the ORAC assay for screening germplasm and provided practical advice on design of varietal assessments using antioxidant screenings.     

Effects of cold storage, rearing temperature, parasitoid age and irradiation on the performance of Trichogramma evanescens Westwood (Hymenoptera: Trichogrammatidae)

In this study, the effects of cold storage, rearing temperature, parasitoid age, and irradiation on the performance of the egg parasitoid Trichogramma evanescens were investigated. Pupae of T. evanescens can be stored at 4 °C for up to 3 weeks without much loss of performance. The longevity and walking speed of adults emerging from chilled pupae significantly decreased after longer storage periods. The F₁ generation of adults which emerged from pupae stored up to 3 weeks was able to parasitize as well as the control. The parasitization rate was similar at 24, 27, and 30 °C, but significantly decreased at 33 and 36 °C. Although T. evanescens developed to the pupal stage at 36 °C, no adult emergence was observed at this temperature. Developmental periods were longer at 24 °C than at higher temperatures. The optimum age for T. evanescens to successfully parasitize host eggs ranged from 24 to 90 h. The parasitization frequency of the 56-78 h aged females was higher than for the other age groups. The daily egg laying pattern of female T. evanescens adults was similar when they were reared on Ephestia kuehniella or Plodia interpunctella eggs. Gamma- or ultraviolet-irradiated and unirradiated host eggs were equally preferred by adult females.     

Diversity of culturable microorganisms and occurrence of Listeria monocytogenes and Salmonella spp. in Tuber aestivum and Tuber melanosporum ascocarps

The aim of this study was to investigate the total mesophilic microorganisms, Pseudomonas genus, Enterobacteriaceae family, mold and yeast counts and the presence of Listeria monocytogenes and Salmonella spp on Tuber aestivum and Tuber melanosporum ascocarps. The results confirmed that the major percentage of the microorganisms, approximately 9.0 log ufc/g, were present in the peridium, the glebas of healthy truffles being practically free of microorganisms. The predominant microbial group was the Pseudomonas averaging 8.3 and 8.4 log cfu/g on T. aestivum and T. melanosporum whole ascocarps, respectively. The Enterobacteriaceae also achieved high populations, especially in T. aestivum truffles, with 6.3 log cfu/g. Molds and yeasts never exceeded 5.0 log cfu/g. The characterization of the isolates revealed that the fluorescens pseudomonads were the most prevalent. Raoultella terrigena and Enterobacter intermedius were the dominant Enterobacteriaceae. The identification of the yeast isolates revealed five species: Debaryomyces hansenii, Issatchenkia scutulata, Rhodotorula aurantiaca, Saccharomyces dairensis and Trichosporon beigelii subspecies A and B. The mold genera detected in both species of truffles were Aspergillus, Cladosporium, Penicillium and Fusarium, Trichoderma being present only in T. aestivum. L. monocytogenes was found in 10% of the samples of T. aestivum analysed but Salmonella spp. was not detected. Knowledge of the microbial population in terms of possible food borne and pathogen microorganisms is very useful for establishing successful disinfection and storage methods to prolong the shelf-life of ascocarps of T. aestivum and T. melanosporum.     

Effect of sublethal concentrations of propionic or butyric acid on growth and aflatoxin production by Aspergillus flavus

Propionic or butyric acid was added at sublethal doses (0.1–2 mg/ml) to a growth medium supporting growth of Aspergillus flavus and subsequent aflatoxin production. A reduction in growth and aflatoxin production occurred when the acids were added at the time of inoculation. Addition of the acids to cultures at different times resulted in little effect on growth but production of aflatoxin after 12 days was reduced with earlier time of application for both propionic and butyric acid. When the acids were added to rough rice with a moisture content of 21% and inoculated with A. flavus fungal growth and aflatoxin production were reduced relative to non-inoculated controls. Early application of acids resulted in lower yields of aflatoxin.     

Evolution of aroma volatiles during storage of sourdough breads made by mixed cultures of Kluyveromyces marxianus and Lactobacillus delbrueckii ssp. bulgaricus or Lactobacillus helveticus

Two mixed starter cultures were used for sourdough bread making to evaluate their ability to improve quality and increase bread shelf-life: Lactobacillus delbrueckii ssp. bulgaricus or Lactobacillus helveticus mixed with the lactose fermenting yeast Kluyveromyces marxianus as alternative baker’s yeast. Control sourdough breads (K. marxianus) without the addition of bacteria, were also prepared. The changes on the headspace aroma volatiles during storage were assessed using solid-phase microextraction (SPME) GC–MS analysis. The effect of these changes on bread flavour was evaluated by consumer preference evaluations and the results were co-evaluated with those from the GC–MS analysis. The obtained results showed differences in the volatile composition of the different types of breads examined, as well as dramatic decreases of the number and the amount of volatiles after five days of storage. The sourdough breads made with K. marxianus and L. bulgaricus, had a more complex aroma profile, longer shelf-life and achieved the highest scores in the sensory tests.     

Predation by Blattisocius tarsalis (Berlese) (Acari: Ascidae) on eggs of Ephestia kuehniella Zeller (Lepidoptera: Pyralidae)

The mean predation rate of female B. tarsalis on eggs of E. kuehniella was determined at 15°C, 21°C, and 27°C with prey densities of 3, 5, 10 and 15 eggs. Young eggs less than 24 h old were used as prey. Under all experimental conditions most eggs were only partially consumed, confirming the general theory that predatory mites leave most of the food content unutilized. A mean predation rate of 2.2-7.0 eggs destroyed in 24 h was found, reflecting that a large proportion of the available prey was destroyed. The result supports the view that the predatory mite B. tarsalis could be a relevant biological control agent against E. kuehniella in flour mills. It was further demonstrated that frozen moth eggs can be considered as an appropriate food source.     

Effect of antioxidants and competing mycoflora on Fusarium verticillioides and F. proliferatum populations and fumonisin production on maize grain

This study was carried out to determine the temporal effect of the antioxidants butylated hydroxyanisol (BHA) and propyl paraben (PP) at doses of 500 and 1000 μg/g on the growth of Fusarium verticillioides and F. proliferatum inoculated on natural maize grain in the presence of the competing mycoflora and fumonisin production at 0.98 and 0.95 water activity (a_w) over a 28-day storage period. The reduction in the log colony forming units (CFU) of Penicillium, Aspergillus and Fusarium populations was 10-100 fold depending on dose of BHA or PP, a_w and time. However, the populations of all three groups were higher at 0.98 a_w than 0.95 a_w. BHA at 500 μg/g and 0.95 a_w reduced the fumonisin content by 82% after 7-14 days incubation, but at the end of the experimental period the reduction was only 32%. A higher reduction in the level of fumonisin produced (77%) was achieved with BHA at 1000 μg/g after 28 days. PP at 500 and 1000 μg/g decreased fumonisin production throughout the incubation period in the drier treatment, but at 0.98 a_w control of toxin production was only achieved after 7-14 days. The reduction in the fumonisin levels could be due to the combined effect of antioxidants, and the competing mycoflora, mainly Aspergillus and Penicillium species.     

The potential of Cycloclasticus and Altererythrobacter strains for use in bioremediation of petroleum-aromatic-contaminated tropical marine environments

Cycloclasticus sp. A5, which has been suggested to be a major degrader of petroleum aromatics spilled in temperate seas, showed higher degrading activities for petroleum aromatics, at both 25 °C and tropical sea temperature 30 °C, than the novel aromatic-degrading isolates, related to Altererythrobacter epoxidivorans (97.5% similarity in the almost full-length 16S rRNA gene sequence) and Rhodovulum iodosum (96.3% similarity), obtained after enrichment on crude oil in a continuous supply of Indonesian seawater. Cycloclasticus A5 degraded petroleum aromatics at a similar rate or faster at 30 °C as compared to 25 °C, but its growth on acetate was severely inhibited at 30 °C. These results suggest that, although their abundance would be low in tropical seas not contaminated with aromatics, the Cycloclasticus strains could be major degraders of petroleum aromatics spilled in tropical seas. The 16S rRNA gene of the Cycloclasticus strains has been identified from Indonesian seawater, and the gene fragments showed 96.7−96.8% similarities to that of Cycloclasticus A5. Introducing Cycloclasticus A5 may be an ecologically advantageous bioremediation strategy for petroleum-aromatic-contaminated tropical seas because strain A5 would disappear at 30 °C after complete consumption of the aromatics. Altererythrobacter and Rhodovulum-related isolates grew well on pyruvate in 10% strength marine broth at 30 °C whereas Cycloclasticus A5 did not grow well on acetate in the broth at 30 °C. These growth results, along with its petroleum-aromatic-degrading activity, suggest that the Altererythrobacter isolate could be an important petroleum-aromatic degrader in and around nutrient-rich tropical marine environments.     

Biodiversity of Aspergillus section Nigri populations in Argentinian vineyards and ochratoxin A contamination

Aspergillus section Nigri are described as the main source of ochratoxin A (OTA) contamination in grapes and wine worldwide. The knowledge of the factors affecting grape contamination by species included in this section and OTA production is essential to be able to reduce their presence, not only to improve wine quality, but also to maintain their safety. Therefore, the aims of this study were to determine the incidence of Aspergillus section Nigri species harvested in different grape-growing regions from Argentina, their ability to produce OTA, to correlate with meteorological conditions and geographical coordinates with their prevalence and to evaluate the OTA natural occurrence in grapes and wines. The morphological identification showed that Aspergillus niger aggregate species were the most prevalent ones, followed by Aspergillus carbonarius and Aspergillus uniseriate. These populations were confirmed through using AFLP markers and sequencing and, Aspergillus tubingensis was separated from A. niger aggregate. Climatic factors, altitude, longitude and latitude have influenced on the distribution of species included in the section. A. carbonarius and A. niger were OTA producers but differed in their OTA producing ability. Temperature was the factor which influenced the most over the highest incidence of A. carbonarius in La Rioja and San Juan regions. The trellis system in vineyards and drip irrigation also influenced the species isolation. The OTA levels detected in grapes and wines were low, but grape variety was more important in susceptibility to fungal infection and OTA levels.