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Simple Summary: Ochratoxin A (OTA), a nephrotoxic mycotoxin categorized as a possible carcinogenic agent, is a current problem in the cheese manufacturing industry. The main OTA producers are Penicillium nordicum and Penicillium verrucosum. Therefore, we aimed to investigate different cheese ripening phases to hinder OTA production. We elaborated cheese analogues, inoculated both fungal species, and performed two different ripening phases, at 10 and 15 °C. Despite the fact that both were able to grow, they did not express genes related to OTA production, and no OTA was detected during two weeks of ripening. Therefore, we show that, when the first phases of ripening are maintained at <15 °C, the possibility of OTA production is dramatically minimized. This information is of utmost importance for industries to enhance the safety of their cheese products, which positively impacts human health. A lack of control of the technological abiotic parameters apparent during cheese manufacture, including temperature and relative humidity, results in this dairy product being prone to mold contamination. Sometimes, inoculant molds are used to obtain the characteristic sensory properties of this type of product. However, during the maturation process, some unwanted molds can colonize the ripening cheese and produce mycotoxins. Mycotoxigenic molds such as Penicillium nordicum and Penicillium verrucosum can colonize ripened cheeses, contaminating them with ochratoxin A (OTA), a nephrotoxic 2B toxin. Thus, the presence of OTA in cheeses could represent a hazard to consumers' health. This study has evaluated the growth and OTA production of P. nordicum and P. verrucosum on a cheese analogue under simulated ripening conditions of 10 and 15 °C and 0.96 water activity (aw). Ecophysiological, molecular, and analytical tools assessed the mold growth, gene expression, and OTA production under these environmental conditions. Both species were able to effectively colonize the cheese under these ripening conditions. However, neither species expressed the otapks and otanps biosynthetic genes or produced phenotypic OTA. Therefore, these results suggest a relatively low risk of exposure to OTA for consumers of this type of cheese product. The conditions used were thus appropriate for cheese ripening to minimize the potential for contamination with such mycotoxins. An appropriate adjustment of the technological ripening parameters during such cheese manufacture could contribute to OTA-free cheeses. [ABSTRACT FROM AUTHOR]

There is little information on the effect of temperature, relative humidity (RH) and vapour pressure deficit (VPD) on the viable populations of two commercial biocontrol strains, Bacillus subtilis QST 713 (recently classified as B. velezensis) and Gliocladium catenulatum J1446 (syn. Clonostachys rosea). The PMAxx‐qPCR molecular assay was used to quantify the viable biocontrol agent (BCA) populations on fully extended lettuce and strawberry leaves under a range of temperature and RH combinations. Overall, there was a small decline in the population size of viable cells for the two biocontrol organisms on strawberry and lettuce leaves within 10 days of application. However, for most experimental runs, which contained general UK agronomy climates, such a decline was not statistically significant. Moreover, for a few runs, the viable populations increased significantly in optimal BCA growth temperatures with high RH. Only temperature (ambient) and dew point significantly affected the rate of temporal changes in the viable biocontrol population size. Thus, an increasing temperature led to decreased daily mortality. It should be noted that much of the variability in the estimated daily mortality rates remains unaccounted for, thus unless under extreme conditions, the biocontrol efficacy in practice is less likely to be affected by the survival of biocontrol microbes but more likely by other factors that influence the density of biocontrol cells in the phyllosphere, especially dilution due to rapid host leaf expansion, spray coverage and rain wash‐off. [ABSTRACT FROM AUTHOR]

Apple rootstock genotypes confer different levels of tolerance to apple replant disease (ARD) and vigour to a newly replanted apple tree. A hybrid management system of rotating the rootstock genotype planted between successive generations and inter-row planting in the alleyways of orchards may minimise the severity of ARD symptoms. High-throughput sequencing of the fungal ITS and bacterial 16S rDNA regions was used to investigate the diversity, and differential taxa present in soils displaying symptoms of ARD. Candidate pathogens and beneficial microorganisms were correlated with the above-ground establishment of each rootstock genotype in a UK cider orchard. Our results suggest that the same rootstock or rootstock with closely related parentage to the previous rootstock had more severe ARD symptoms. Planting in the alleyway appeared an effective strategy to minimise the severity of symptoms irrespective of rootstock genotype. The planting location effect had a higher contribution to the variation in the rhizosphere microbiome than that of the rootstock genotype. No predicted causal agents for ARD could be identified to a taxonomic level to predict their function but two species associated with mycorrhizae, Pteridiospora spinosispora and Paraglomus laccatum were identified as inversely correlated with ARD severity and could be candidate beneficial species for apple, warranting further investigation and research. Our findings suggest that planting in the alleyways and planting rootstocks genetically dissimilar to the previously planted rootstock can be beneficial for tree establishment. We have also identified species inversely associated with ARD severity, making candidates for future research to test the antagonistic effect of the species against ARD pathogens in apple roots. [ABSTRACT FROM AUTHOR]

Aim We assessed the effect of exposing apple orchard soil to different temperatures and CO2 levels on the resident microbiome of soils from a conventionally managed and an organically managed apple orchard. The key difference between these two orchards was that synthetic fertilizers and pesticides are routinely used in the former one. Methods and results To investigate the effect of CO2 and temperature, soil samples from each site at two depths were exposed to either elevated temperature (29°C) at either 5000 or 10 000 ppm for five weeks or control conditions (25°C + 400 ppm). Both bacterial and fungal communities were profiled with amplicon-sequencing. The differences between the two orchards were the most significant factor affecting the bacterial and fungal communities, contributing to 53.7–14.0% of the variance in Bray–Curtis β diversity, respectively. Elevated CO2 concentration and increased temperature affected organic orchard microbial diversity more than the conventionally managed orchard. A number of candidate beneficial and pathogenic microorganisms had differential abundances when temperature and CO2 were elevated, but their effect on the plant is unclear. Conclusions This study has highlighted that microbial communities in bulk soils are most significantly influenced by crop management practices compared to the climate conditions used in the study. The studied climate conditions had a more limited effect on microbial community diversity in conventionally managed soil samples than in organically managed soils. [ABSTRACT FROM AUTHOR]

The objectives of this study were to compare the effect of different Stevia-based sugar substitutes (S1–S3), sucrose alone and a mixture of sucrose + S1 on: (a) humectant properties, (b) relative colonisation rates of sponge cake slices at 0.90 a w by strains of Aspergillus flavus , Eurotium amstelodami , Fusarium graminearum and Penicillium verrucosum at 20 and 25 °C and (c) shelf-life periods in days prior to visible growth. Results showed that sucrose, S1 commercial sugar substitute and the mixture of sucrose + S1 in water solutions were able to reach water activity levels similar to those of glycerol and glucose mixtures. The S2 and S3 commercial sugar substitutes were unable to reduce a w levels significantly. At 25 °C, colonisation of sponge cake slices by E. amstelodami , A. flavus and P. verrucosum occurred in all the treatments. Growth of F. graminearum only occurred on sponge cake slices containing S2 and S3 Stevia-based products at both temperatures. The best control of growth (30 days) was achieved in cake slices modified with sucrose or S1 Stevia treatments inoculated with A. flavus and in the sucrose treatment for E. amstelodami at 20 °C. F. graminearum growth was completely inhibited when sucrose alone, S1 or sucrose + S1 treatments were used at both temperatures. This study suggests that, as part of a hurdle technology approach, replacing sucrose with low calorie sugar substitutes based on Stevia glycosides needs to be done with care. This is because different products may have variable humectant properties and bulking agents which may shorten the potential shelf-life of intermediate moisture bakery products. [ABSTRACT FROM AUTHOR]

Two new species of Aspergillus (A. aculeatinus, A. sclerotiicarbonarius) were previously isolated from coffee in Thailand. The objective of this study was to examine the effect of interacting environmental factors of water availability (water activity, aw) and temperature on lag phases prior to growth, growth and potential for ochratoxin A (OTA) production by three strains of each species on a green coffee-based medium for the first time. This showed that overall the growth of the three strains of each species was similar over the 20-37 °C and 0.85-0.99 aw ranges. The lag phase prior to growth was < 1 day at 0.95-0.98 aw and 25-37 °C and increased to 2-3 days at marginal temperatures and aw levels. The growth of strains of the uniseriate species A. aculeatinus was optimum at 0.98 aw and 30-35 °C. For the biseriate A. sclerotiicarbonarius strains this was 0.99 aw and 30 °C. This species was not able to grow at 37 °C. None of the strains of the two species grew at 0.85 aw, regardless of temperature. Integrated profiles based on the data from three strains of each species have been developed to show the optimum, maximum and marginal conditions of interacting aw and temperature conditions for growth. None of the strains produced OTA on a green coffee-based medium. This information is important as these species are part of the mycobiota of coffee and may influence OTA contamination by other ochratoxigenic species during coffee processing. [ABSTRACT FROM AUTHOR]

Abstract: The effects of temperatures (20–30 °C) and water activity (0.90–0.99 a w) on the lag phase duration, mycelial growth, and nutritional utilisation patterns of two toxigenic (AFL1+ & AFL2+) and three atoxigenic (AFL1−, AFL2−, & AFL3−) Aspergillus flavus strains were evaluated in vitro. Both temperature and a w and their interactions had a significant influence on the growth and nutritional utilisation patterns (p < 0.05). There were no significant differences between toxigenic and atoxigenic strains in terms of lag phase prior to growth and mycelial growth rates. Based on carbon source (CS) utilisation patterns, toxigenic and atoxigenic strains' niche size was greater at higher temperatures and in wetter conditions. Additionally, based on niche overlap indices (NOIs), regardless of temperature, when water was freely available, atoxigenic and toxigenic strains co-existed. However, under moisture stress, the nutritional competitiveness was variable. Temporal carbon utilisation sequences (TCUS) of toxigenic and atoxigenic strains were compared. At 0.99 a w most CS sources were utilised by the strains and the time to detection (TTD) of each strain was shortest on monosaccharides at the same level of a w. Conversely, under moisture stress the least number of CS was utilised. The current study has demonstrated that carbon utilisation patterns are equally important as are other determinants of competitiveness and that growth rate alone is not a key attribute which determines competitiveness. [Copyright &y& Elsevier]

Abstract: Molecular studies, especially in relation to the activity of secondary metabolite gene clusters, require the ability to extract good quality RNA from fungal biomass. This is often hindered by the cell wall structure and endogenous RNase activity in filamentous fungi. There is thus a need for rapid methods for the extraction of good quality RNA for use in microarrays and for quantitative PCR assays. The objective of this study was to examine the use of different systems for the high throughput method to extract intact RNA from filamentous fungi. Two bead beating systems with different motion patterns and speed capacities were tested in the development of the extraction protocol. They were evaluated based on the total RNA yield and overall RNA quality. The high speed bead beating with glass beads associated with an automated purification method gave more than three times higher total RNA yields with less than a quarter of the amount of mycelium required. Furthermore the integrity and overall quality was conserved, with RNA Quality Indicator (RQI) numbers consistently >7.5. This method also reduced cross contamination risks and kept RNA handling to a minimum while still being capable of multiple sample processing, reducing the time required to obtain RNA from filamentous fungi. [Copyright &y& Elsevier]

The temporal efficacy of three different chemical fungicides (Folicur®, Proline®, Sportak 45EW®) and a biocontrol bacterium (Serenade, B. subtilis) in reducing growth and toxin production by isolates of F. verticillioides and A. flavus was studied in vitro under different water activity regimes (0.99, 0.98 and 0.95). All the fungicides significantly inhibited mycelial growth compared with the control; the most effective treatment, both against F. verticillioides and A. flavus, was Sportak 45EW® (approx. 99%). The inhibitory effect of all fungicides generally improved with increasing concentration. Serenade always decreased fungal growth, with optimal results at concentrations of 104 and 106 (70-75% reduction). All the fungicide treatments resulted in a significant reduction in both FB1+FB2 and AFB1 production when compared to the control, at the end of the incubation period and with the 2 concentrations used (approx. 99%). A threshold concentration inoculum of at least 104 CFUs of B. subtilis per g was required to achieve a significant control of mycotoxin production. Sportak 45EW® and Serenade gave the best control of mycotoxin production with a reduction of 95% compared to the controls. Use of Serenade in the field should include due consideration to its sensitivity to low water activities, when compared to the target pathogens. [ABSTRACT FROM AUTHOR]

The effects of ecophysiological factors, temperature and solute potential, on both the growth and the regulation of the fumonisin biosynthetic FUM1 gene were studied and compared in one isolate each of the two closely related fumonisin-producing and maize pathogens Fusarium verticillioides and Fusarium proliferatum. The effect of solute potential and temperature was examined on in vitro mycelia growth and on the expression of the FUM1 gene, quantified by species-specific real-time reverse transcriptase-PCR assays. Although both isolates showed similar two-dimensional profiles of growth, for F. verticillioides, optimal growth conditions were maintained at higher temperatures and lower solute potential values. FUM1 gene expression was markedly induced at 20 °C in both isolates, under suboptimal conditions for growth; however, their expression patterns differed in relation to solute potential. Whereas FUM1 expression was induced in response to increasing water stress in the isolate of F. verticillioides, the F. proliferatum one showed a stable expression pattern regardless of water potential conditions. These results suggest a differential regulation of fumonisin biosynthesis in these isolates of the two species that might be related to their different host range, and play an ecological role. Additionally, environmental conditions leading to water stress (drought) might result in increased risk of fumonisin contamination of maize caused by F. verticillioides. [ABSTRACT FROM AUTHOR]

The quality of harvested wheat grain can deteriorate markedly during the post-harvest management stages. Biotic factors, such as grain type and ripeness, coupled with the prevailing abiotic factors, such as water content and temperature, and also preservative concentration will influence the safe storage life and the level of contamination with mycotoxins. These mycotoxins include deoxynivalenol (DON) produced pre-harvest and zearalenone (ZEA) produced post-harvest by Fusarium graminearum and Fusarium poae, respectively, ochratoxin (OTA) produced by Penicillium verrucosum post-harvest in cool damp northern European climates, and perhaps T-2 and HT-2 toxins produced by Fusarium langsethiae. This review presents recent data on the relationship between dry matter losses caused by F. graminearum under different environmental regimes (water activities, temperatures) and the level of contamination with DON. This is important as poor post-harvest drying and storage management may exacerbate DON contamination already present pre-harvest. It is thus critical to relate the environmental factors in stored wheat grain during storage, especially of intergranular relative humidity (RH) and temperature, to safe storage periods without spoilage or risk from increased DON contamination. The growth/no growth and DON/no DON (F. graminearum) and OTA/no toxin production (P. verrucosum) have been used to build a model with a simple interface to link temperature and RH values to the potential risk level which may allow growth or toxin production. This paper also considers the use of modified atmospheres, preservatives and biocontrol to minimise DON and OTA in moist wheat grain. These approaches together with clear monitoring criteria and hygiene could contribute to better post-harvest management of stored temperate cereals and ensure that mycotoxin contamination is minimised during this key phase in the food/feed chain. [ABSTRACT FROM AUTHOR]

Abstract: Embellisia allii results in the formation of a bulb canker and black soot on the surface of different alliums and it has been frequently detected on garlic bulbs together with the spoilage fungus, Fusarium oxysporum f. sp. cepae, which causes bulb basal plate rot. In this study, the influence of water activity (aw) and temperature on mycelial growth of E. allii and F. oxysporum f. sp. cepae, conidial size and sporulation of E. allii, interactions between E. allii and F. oxysporum f. sp. cepae, Index of Dominance (I D), and in situ virulence on garlic were examined. Mycelial growth of E. allii was optimal (5.97mm/day) at 0.995 aw and 25°C, slower at 30°C. However, almost no growth occurred at 0.937 aw/30°C. F. oxysporum f. sp. cepae grew faster than E. allii, (6.3–7.4mm/day) at 30°C. Interactions between E. allii and F. oxysporum f. sp. cepae were influenced by aw and temperature. Sporulation of E. allii was more abundant on PDA than on MEA, especially at high aw (0.995) and low temperature (20°C), but almost no sporulation occurred at 30°C regardless of nutritional medium or aw level. The spore length of E. allii was longer on PDA than MEA, and was significantly influenced by water availability. F. oxysporum f. sp. cepae was competitive against E. allii and had a higher I D value in comparison with E. allii especially at a higher temperature (30°C). In situ virulence tests showed that E. allii was weakly virulent on the garlic bulb cloves while that of F. oxysporum f. sp. cepae was highly dependent on aw. [Copyright &y& Elsevier]

Abstract: This study examined the utilization patterns of key carbon sources (CS, 24: including key sugars, amino acids and fatty acids) in maize by strains of Aspergillus flavus and Fusarium verticillioides under different water activity (a w, 0.87–0.98 a w) and temperature (20–35 °C) values and compared the niche overlap indices (NOI) that estimate the in vitro CS utilization profiles [Wilson, M., Lindow, S.E., 1994. Coexistence among epiphytic bacterial populations mediated through nutritional resource partitioning. Applied and Environmental Microbiology 60, 4468–4477.]. The ability to grow in these key CS in minimal media was studied for 120 h in 12 h steps. The NOI was calculated for inter-species (F. verticillioides–A. flavus) and for intra-species (A. flavus–A. flavus) using CS utilization patterns over the range of interacting environmental conditions. 30 °C, over the whole a w range examined, was found to be optimal for utilization of the maximum number of CS by A. flavus. In contrast, for F. verticillioides this was more so at 20 °C; 25 °C allowed a suboptimal usage of CS for both species. NOIs confirmed the nutritional dominance of A. flavus at 30 °C, especially at lower a w levels and that of F. verticillioides at 20 °C, mainly at 0.95 a w. In other conditions of a w, based on CS utilization patterns, the data indicated that A. flavus and F. verticillioides occupied different ecological niches. The variability in nutritional sources utilization between A. flavus strains was not related to their ability to produce aflatoxins (AFs). This type of data helps to explain the nutritional dominance of fungal species and strains under different environmental conditions. This could be useful in trying to find appropriate natural biocontrol microorganisms to compete with these mycotoxigenic species. [Copyright &y& Elsevier]

Abstract: This study has examined the effect of inoculation of soil microcosms with Trametes versicolor and Phanerochaete chrysosporium on wood chips on differential degradation of pesticides (simazine, trifluralin and dieldrin, 10mgkg−1 soil) at two water potentials (−0.7 and −2.8MPa) at 15°C. The soil microcosms were destructively sampled after 6/12 weeks and four extracellular enzymes quantified, respiration and pesticides measured with GC and HPLC. The fungal treatments produced extracellular enzymes in soil. Respiratory activity was significantly (P =0.05) enhanced in soil with the inocula, and higher in the pesticide mixtures. Cellulase/dehydrogenase increased in inoculated soil. Laccase increased significantly in the T. versicolor treatment. Degradation of the three pesticides by wood chip addition alone was enhanced (20–30%). T. versicolor increased degradation of simazine (27–46%), trifluralin (5–17%) and dieldrin (5–11%) and P. chrysosporium by 34–48%, 0–30% and 40–46%, respectively, when compared with controls after 12 weeks. This study has demonstrated that pesticide mixtures are differentially degraded by fungal inoculants and significant extracellular enzymes are produced in soil, even at −2.8MPa water potential. This suggests that effective bioremediation of xenobiotic mixtures using wood chips and fungal inoculants is achievable over a relatively wide water potential range when compared with that allowing plant growth (−1.4MPa). [Copyright &y& Elsevier]

The major hurdle in the production of commercial biocontrol agents (BCAs) has been the lack of production of appropriate formulations. Of particular importance is the conservation of viability and ecological competence after application. With this in mind studies were conducted to develop formulations of P. anomala which would have these attributes. Cells were grown in molasses-based medium modified with proline to different water availability levels (0.98 and 0.96) which significantly increased (up to 50%) the content of trehalose and arabitol in the yeast cells during liquid broth fermentation. The use of isotonic solutions for harvesting the yeast cells further increased the endogenous content of these compatible solutes as well as glycerol. Fluidised bed drying of cells at 30-80°C was carried out for 10 and 20 min and showed that viability was significantly decreased at 70-80°C. A temperature of 50°C for 20 min was found to be best for viability (70%) and moisture content of <10%. Several additives for conservation of viability showed that cotton seed flour+skimmed milk was the best treatment when dried at 50°C. The biocontrol efficacy of formulated P. anomala cells was tested in laboratory scale studies and this showed that they inhibited growth of Penicillium verrucosum and reduce ochratoxin A production in moist wheat grain under some combinations of water availability. Physiologically modified formulated yeast cells with increased levels of trehalose and arabitol gave similar efficacy as fresh cells. This suggests that ecophysiological manipulation of such BCAs can result in improved ecological competence of such formulations and effective biocontrol. [ABSTRACT FROM AUTHOR]

Systematic expression analysis of mycotoxin biosynthesis genes by real-time PCR and microarray was carried out to examine the relationship between growth and general expression patterns in relation to single environmental factors such as temperature, water activity ( aw) and pH and water activity × temperature interactions. For single parameters, one major peak of expression occurred close to optimum growth conditions. However, a second minor peak was observed under suboptimal growth conditions, when intermediate environmental stress was imposed on Aspergillus parasiticus ( afl genes), Penicillium verrucosum ( ota genes) and Fusarium culmorum ( tri genes). This expression profile pattern was more pronounced in relation to changes in temperature and aw than to pH. In a two-factorial experimental design with temperature × aw regimes, again two peaks of expression were observed for cluster genes after microarray analysis, one close to those giving optimal growth and one under imposed stress conditions. Interestingly, when the activity of single genes of the microarray data were plotted in relation to the two parameters, again a two-peak expression profile became obvious independently for both parameters. Expression of the mycotoxin biosynthesis genes was followed exactly by phenotypic mycotoxin production. This expression profile appears to be generic across the mycotoxigenic fungi examined. [ABSTRACT FROM AUTHOR]

Abstract: The impact of different concentrations of bisphenol A (BPA) was evaluated on growth of the white-rot basidiomycete, Trametes versicolor, and on the expression of genes encoding lignin-modifying enzyme (LME) activities. Effective doses (EDs) were obtained from fungal growth rate to monitor LME activities and the expression levels of their encoding genes. The fungus showed mycelial growth at concentrations of up to 300μgml−1 of BPA with an ED50 value of 185μgml−1. The LME activities were stimulated by BPA concentrations up to 300μgml−1. The lignin peroxidase (LIP) encoding gene may be sensitive to BPA stress. [Copyright &y& Elsevier]

Abstract: The potential of an electronic nose (e-nose) consisting of a hybrid gas sensor array system has been examined for species discrimination and strain identification of dermatophytes which are causative pathogens for human and animal infection. Temporal volatile production patterns have been studied at a species level for a Microsporum species, two Trichophyton species and at a strain level for the two Trichophyton species. After about 120h principal component analysis (PCA) and cluster analysis showed possible discrimination between the species from controls. Data analysis also indicated probable differentiation between the strains of T. rubrum. The same could not however be achieved for the strains of T. mentagrophytes during preliminary experiments for the same time period, signifying a good similarity between the strains of this particular species based on their volatile fingerprints. This study suggests that volatile production patterns shows promise for species and strain identification of these dermatophytic fungi thereby facilitating early diagnosis and early management of patients. [Copyright &y& Elsevier]

Abstract: Contamination of cereal commodities by moulds and mycotoxins results in dry matter, quality, and nutritional losses and represents a significant hazard to the food chain. Most grain is harvested, dried and then stored on farm or in silos for medium/long term storage. Cereal quality is influenced by a range of interacting abiotic and biotic factors. In the so-called stored grain ecosystem, factors include grain and contaminant mould respiration, insect pests, rodents and the key environmental factors of temperature, water availability and intergranular gas composition, and preservatives which are added to conserve moist grain for animal feed. Thus knowledge of the key critical control points during harvesting, drying and storage stages in the cereal production chain are essential in developing effective prevention strategies post-harvest. Studies show that very small amounts of dry matter loss due to mould activity can be tolerated. With <0.5% dry matter loss visible moulding, mycotoxin contamination and downgrading of lots can occur. The key mycotoxigenic moulds in partially dried grain are Penicillium verrucosum (ochratoxin) in damp cool climates of Northern Europe, and Aspergillus flavus (aflatoxins), A. ochraceus (ochratoxin) and some Fusarium species (fumonisins, trichothecenes) on temperate and tropical cereals. Studies on the ecology of these species has resulted in modelling of germination, growth and mycotoxin minima and prediction of fungal contamination levels which may lead to mycotoxin contamination above the tolerable legislative limits (e.g. for ochratoxin). The effect of modified atmospheres and fumigation with sulphur dioxide and ammonia have been attempted to try and control mould spoilage in storage. Elevated CO2 of >75% are required to ensure that growth of mycotoxigenic moulds does not occur in partially dried grain. Sometimes, preservatives based on aliphatic acids have been used to prevent spoilage and mycotoxin contamination of stored commodities, especially feed. These are predominantly fungistats and attempts have been made to use alternatives such as essential oils and anti-oxidants to prevent growth and mycotoxin accumulation in partially dried grain. Interactions between spoilage and mycotoxigenic fungi and insect pests inevitably occurs in stored grain ecosystems and this can further influence contamination with mycotoxins. Effective post-harvest management of stored commodities requires clear monitoring criteria and effective implementation in relation to abiotic and biotic factors, hygiene and monitoring to ensure that mycotoxin contamination is minimised and that stored grain can proceed through the food chain for processing. [Copyright &y& Elsevier]

Abstract: Much attention has been focused on the production of musty-aroma compounds such as geosmin and their impact on the quality of fresh water and water-cultured raised fish and sea-food. At present, there are no efficient means of removing these off-flavours from water. Therefore, the rapid detection of geosmin-producing microorganisms, in particular the genus Streptomyces, at early stages of differentiation, is still the best option in preventing deterioration of water quality. We investigated the potential of an electronic nose consisting of an array of 14 conducting polymer sensors for the rapid and early detection of Streptomyces spores in reverse osmosis and tap water. Geosmin solutions in water at different concentrations were also prepared for headspace analysis in order to study the e-nose potential for detecting geosmin production in this environment. Normalised divergence data were analysed using principal component analysis (PCA) and discriminant function analysis (DFA). Data indicates that an e-nose could be employed to detect and monitor early activity of Streptomyces in water at different stages of differentiation, as well as to discriminate between different species based on their volatile production patterns. It also suggests that it could be used for monitoring geosmin production in water and possibly set threshold odour levels, as a routine task for specific water-screening purposes. [Copyright &y& Elsevier]

The aim of this study was the production of high numbers of M. anisopliae blastospores with enhanced germination efficiency under conditions of water-stress (ecological fitness). Different nitrogen sources and concentrations were screened for their ability to induce blastospore production while keeping pH and water activity ( a w ) at fixed levels (6.8, and 0.98 a w , respectively). After optimum nitrogen status was determined (cornsteep solid (CS) + yeast extract (YE); or cottonseed flour (CF) + YE), the effect of interaction of nitrogen source, pH (3.5, 5, 6.8, 8, 9 and 10) and solutes for a w adjustment (KCl, NaCl, PEG 200) on blastospore production, endogenous polyol content (glycerol, erythritol, arabitol and mannitol) and total protein, were determined. For both ionic (NaCl, KCl) and non-ionic solutes (PEG 200), optimum blastospore production (between 4×10 7 and 2×10 8 blastospores ml -1 ) and growth occurred in the pH range 6.8–8, with the CF + YE nitrogen profile giving higher yields than CS + YE. Optimum conditions for high erythritol and total protein endogenous concentrations (40.37–73.44 and 14.33–18.90 mg g -1 fresh weight, respectively) occurred between pH 6.8 and 8 by ionic a w modification (KCl, NaCl) and with the CF + YE nitrogen profile. Germination of blastospores produced under these cultural conditions was between 62 and 89% under conditions of water-stress (0.96 a w ). On the other hand, blastospores with lower amounts of erythritol and total protein content had decreased germination (8–67%). These results could have significant implications for developing a liquid fermentation medium for the production of high numbers of fungal propagules with enhanced efficacy under non-optimum environmental conditions. [ABSTRACT FROM AUTHOR]

The major species of fungi responsible for ochratoxin production (OTA) in a range of commodities are Penicillium verrucosum , Aspergillus ochraceus and Aspergillus section Nigri , especially A. carbonarius. P. verrucosum is particularly important in northern Europe where damp cooler conditions occur and where drying regimes need to be efficient and effective for preventing post-harvest contamination with OTA. A. ochraceus can infect cereals including barley, maize, coffee, cocoa and edible nuts. A. carbonarius has been identified as the key species responsible for OTA contamination of grapes, wine and vine fruits. Recent studies have identified the environmental regimes, especially of temperature and humidity, which are conducive to growth and OTA production by these species pre- and post-harvest and during transport. The optimum and marginal interacting conditions for growth and OTA contamination often vary considerably. This has to be borne in mind when effective preventative management strategies are being implemented. Recent studies with cereals have suggested that isolation frequency can be related to OTA contamination of cereals. A logistical model has been developed and identified that 1000&ThickSpace;CFUs&ThickSpace;g -1 grain of P. verrucosum (CFUs) is the threshold limit at which the probability of exceeding the EU legislative limit of 5&ThickSpace;µg&ThickSpace;kg -1 in cereal grain can be predicted under different storage regimes. Controlled atmospheres&ThickSpace;>50% C0 2 is required to effectively prevent OTA accumulation in damp cereals. With regard to grapes, preharvest contamination with A. carbonarius has been shown to be a good indicator for regional risk in southern Europe from OTA contamination. The ecological conditions for optimum growth and OTA production have been shown to differ with the optima being 30–35°C and 15–25°C and 0.98–0.99 and 0.93–0.95 water activity, respectively. Studies on vine fruits (drying currants) suggest that OTA contamination and increase contamination levels occur during this drying process of 7–14 days. This will be influenced by prevailing weather conditions and drying rates. Minimizing OTA contamination in these and other commodities including coffee and cocoa require clear guidelines on safe moisture and temperature regimes pre- and post-harvest for the development of effective management strategies based on ecological criteria. [ABSTRACT FROM AUTHOR]

In this study we examined the extracellular enzymatic activity of two white rot fungi (Phanerochaete chrysosporiumandTrametes versicolor) in a soil extract broth in relation to differential degradation of a mixture of different concentrations (0–30 p.p.m.) of simazine, dieldrin and trifluralin under different osmotic stress (−0.7 and−2.8 MPa) and quantified enzyme production, relevant to P and N release (phosphomonoesterase, protease), carbon cycling (β-glucosidase, cellulase) and laccase activity, involved in lignin degradation. Our results suggest thatT. versicolorandP. chrysosporiumhave the ability to degrade different groups of pesticides, supported by the capacity for expression of a range of extracellular enzymes at both−0.7 and−2.8 MPa water potential.Phanerochaete chrysosporiumwas able to degrade this mixture of pesticides independently of laccase activity. In soil extract,T. versicolorwas able to produce the same range of enzymes asP. chrysoporiumplus laccase, even in the presence of 30 p.p.m. of the pesticide mixture. Complete degradation of dieldrin and trifluralin was observed, while about 80% of the simazine was degraded regardless of osmotic stress treatment in a nutritionally poor soil extract broth. The capacity of tolerance and degradation of high concentrations of mixtures of pesticides and production of a range of enzymes, even under osmotic stress, suggest potential bioremediation applications. [ABSTRACT FROM AUTHOR]

The use of volatile production patterns produced by Mycobacterium tuberculosis and associated bacterial infections from sputum samples were examined in vitro and in situ using an electronic nose based on a 14 sensor conducting polymer array. In vitro, it was possible to successfully discriminate between M. tuberculosis (TB) and control media, and between M. tuberculosis and M. avium, M. scrofulaceum and Pseudomonas aeruginosa cultures in the stationary phase after 5–6 h incubation at 37 °C based on 35 samples. Using neural network (NN) analysis and cross-validation it was possible to successfully identify 100% of the TB cultures from others. A second in vitro study with 61 samples all four groups were successfully discriminated with 14 of 15 unknowns within each of the four groups successfully identified using cross-validation and discriminant function analysis. Subsequently, lipase enzymes were added to 46 sputum samples directly obtained from patients and the head space analysed. Parallel measurements of bacterial contamination were also carried out for confirmation using agar media. NN analysis was carried out using some of the samples as a training set. Based on the NN and genetic algorithms of up to 10 generations it was possible to successfully cross-validate 9 of 10 unknown samples. PCA was able to discriminate between TB infection alone, the controls, M. avium, P. aeruginosa and a mixed infection. These findings will have significant implications for the development of rapid qualitative systems for screening of patient samples and clinical diagnosis of tuberculosis. [Copyright &y& Elsevier]

Contamination of cereal commodities with mycotoxins represents a significant hazard to consumer health and has thus received increasing attention from food safety authorities and legislators. For trichothecenes, and deoxynivalenol (DON) in particular, the imminent implementation of legislative limits has focused attention on ways to prevent entry of such mycotoxin contaminants into the food and feed chains. Knowledge of the pre- and post-harvest stages in the cereal production chain and in particular information on where prevention strategies can be implemented is being used to develop quality assurance systems for improving food safety. Information on the ecology of Fusarium species, breeding for resistance, more effective fungicides, potential for biological control and effective drying and storage and preservation systems, are all helping to develop effective preventative strategies for minimising consumer exposure to trichothecenes and other mycotoxins. [Copyright &y& Elsevier]

This review focuses on the fumonisin-producing Fusarium species and the ecophysiology of these species. The effects of environmental biotic and abiotic factors on germination, growth, and fumonisin B1 production by Fusarium verticillioides and Fusarium proliferatum have been investigated under laboratory, field, and storage conditions. An understanding of the factors involved in production of fumonisins is the first step in preventing accumulation of these toxins. [ABSTRACT FROM AUTHOR]

Studies have been carried out to determine the potential for the detection of different microbial species (Enterobacter aerogenes, Escherichia coli, Pseudomonas aeruginosa), alone and in the presence of low concentrations of different heavy metals (As, Cd, Pb and Zn) in bottled, reverse osmosis (RO) and tap water, using an electronic nose. Studies show that it is possible to discriminate control water samples from water contaminated with 0.5 ppm of a mixture of metals. The presence of heavy metals may modify the activity of microorganisms and thus the volatile production patterns. Bacterial species at 102–104 colony forming units (CFUs) ml−1 could be detected after 24 h of incubation. Work is in progress to identify the limits of detection for a range of other microorganisms, including, fungi and cyanobacteria, and chlorinated phenols using electronic nose technology. [Copyright &y& Elsevier]

The use of volatile production patterns produced by bacterial contaminants in urine samples were examined using electronic nose technology. In two experiments 25 and 45 samples from patients were analysed for specific bacterial contaminants using agar culture techniques and the major UTI bacterial species identified. These samples were also analysed by incubation in a volatile generation test tube system for 4–5 h. The volatile production patterns were then analysed using an electronic nose system with 14 conducting polymer sensors. In the first experiment analysis of the data using a neural network (NN) enabled identification of all but one of the samples correctly when compared to the culture information. Four groups could be distinguished, i.e. normal urine, Escherichia coli infected, Proteus spp. and Staphylococcus spp. In the second experiment it was again possible to use NN systems to examine the volatile production patterns and identify 18 of 19 unknown UTI cases. Only one normal patient sample was mis-identified as an E. coli infected sample. Discriminant function analysis also differentiated between normal urine samples, that infected with E. coli and with Staphylococcus spp. This study has shown the potential for early detection of microbial contaminants in urine samples using electronic nose technology for the first time. These findings will have implications for the development of rapid systems for use in clinical practice. [Copyright &y& Elsevier]

Examines the growth and conidial physiology of the entomopathogenic fungi Beauveria bassiana, Metarhizium anisopliae and Paecilomyces farinosus under different conditions. Determination of the effects of culture age, temperature and pH; Effects of culture age on polyols and trehalose.

Reports that the manipulation of intracellular glycerol and erythritol enhances germination of conidia at low water availability. Entomopathogenic fungi; Germ tube extension; Polyol and trehalose content of conidia.

Determines the effects of changing the concentration of glycerol, glucose, trehalose or starch in solid media. Culture conditions, growth assessment and collection of conidia; Extraction and detection of polyols and trehalose.

The study evaluated the dose–response relationship of two commercial microbial biocontrol agents, Bacillus subtilis and Gliocladium catenulatum, against Botrytis cinerea both in vitro and in vivo. Inoculum doses, formulation, temperature and foliar leaf part all affected the control achieved by the two BCAs. In vitro competition assays on modified PDA plates tested a range of BCA doses (log10 3–10 CFUs or spores/droplet) at 4, 10 and 20 °C on the development of B. cinerea colonies. The dose–response relationship was influenced by both the BCA formulation and temperature. In vivo studies on lettuce plants in semi-commercial greenhouses examined the BCA dose (log10 5–9 CFUs or spores/mL) for controlling B. cinerea with a high inoculum (log10 6 spores/mL). Leaf disc assays showed that the dose–response relationship was influenced by the leaf parts sampled. These results suggest that the dose–response relationship between a BCA and specific pathogen will be significantly influenced by environmental conditions, formulation and plant phyllosplane tissue. [ABSTRACT FROM AUTHOR]

Simple Summary: There is a need to be able to track the viable populations of biocontrol agents when applied on the foliar surfaces of plants. We have developed a molecular-based method for the quantification of viable cells of two commercial biocontrol agents—a bacterium (Bacillus subtilis) and a fungus (Gliocladium catenulatum). The method has been tested on the leaf surfaces of lettuce plants to examine the changes in viable population over 10–12 days for the first time. Optimising the use of biocontrol agents (BCAs) requires the temporal tracking of viable populations in the crop phyllosphere to ensure that effective control can be achieved. No sensitive systems for quantifying viable populations of commercially available BCAs, such as Bacillus subtilis and Gliocladium catenulatum, in the phyllosphere of crop plants are available. The objective of this study was to develop a method to quantify viable populations of these two BCAs in the crop phyllosphere. A molecular tool based on propidium monoazide (PMA) (PMAxx™-qPCR) capable of quantifying viable populations of these two BCAs was developed. Samples were treated with PMAxx™ (12.5–100 μM), followed by 15 min incubation, exposure to a 800 W halogen light for 30 min, DNA extraction, and quantification using qPCR. This provided a platform for using the PMAxx™-qPCR technique for both BCAs to differentiate viable from dead cells. The maximum number of dead cells blocked, based on the DNA, was 3.44 log10 for B. subtilis and 5.75 log10 for G. catenulatum. Validation studies showed that this allowed accurate quantification of viable cells. This method provided effective quantification of the temporal changes in viable populations of the BCAs in commercial formulations on lettuce leaves in polytunnel and glasshouse production systems. [ABSTRACT FROM AUTHOR]

Pistachio nuts can become colonized by mycotoxigenic fungi, especially Aspergillus flavus, resulting in contamination with aflatoxins (AFs). We examined the effect of gaseous O3 (50–200 ppm; 30 min; 6 L/min) on (a) in vitro germination, (b) mycelial growth, and (c) aflatoxin B1 (AFB1) production on a milled pistachio nut-based medium at different water activity (aw) levels and at 30 °C. This was complimented with in situ studies exposing raw pistachio nuts to 50–200 ppm of O3. Exposure of conidia to gaseous O3 initially resulted in lower germination percentages at different aw levels. However, 12 h after treatment, conidial viability recovered with 100% germination after 24–48 h. Growth rates of mycelial colonies were slightly decreased with the increase of the O3 dose, with significant inhibition only at 0.98 aw. The production of AFB1 after O3 treatment and storage for 10 days was stimulated in A. flavus colonies at 0.98 aw. Raw pistachio nuts inoculated with A. flavus conidia prior to O3 exposure showed a significant decrease in population after 20 days of storage. However, AFB1 contamination was stimulated in most O3 treatments. The relationship between exposure concentration, time and prevailing aw levels on toxin control needs to be better understood for these nuts. [ABSTRACT FROM AUTHOR]

Maize grown in both North and South America are now predominantly genetically modified (GM) cultivars with some resistance to herbicide, pesticide, or both. There is little information on the relative colonisation and aflatoxin B 1 (AFB 1) production with maize meal-based nutritional matrices based on kernels of non-GM maize and isogenic GM-ones by strains of Aspergillus flavus. The objectives were to examine the effect of interacting conditions of temperature (25–35 °C) and water availability (0.99–0.90 water activity, a w) on (a) mycelial growth, (b) AFB 1 production and (c) develop contour maps of optimum and marginal conditions of these parameters for four strains of A. flavus on three different non-GM and isogenic GM-maize based nutritional media. The growth of the four strains of A. flavus (three aflatoxigenic; one non-aflatoxigenic) was relatively similar in relation to the temperature × a w conditions examined on both non-GM and GM-based matrices. Optimum growth overall was at 30–35 °C and 0.99 a w for all four strains. Under water stress (0.90 a w) growth was optimum at 35 °C. Statistically: non-GM, GM cultivars, temperature and a w all significantly affected growth rates. For AFB 1 production, all single and interacting factors were statistically significant except for non-GM × GM cultivar. In conclusion, colonisation of GM- and non-GM nutritional sources was similar for the different A. flavus strains examined. The contour maps will be very useful for understanding the ecological niches for both toxigenic and non-toxigenic strains in the context of the competitive exclusion of those producing aflatoxins. • Aspergillus flavus strains grew similarly on GM/non-GM nutritional media under interacting abiotic factors • Contour maps of optimum and marginal conditions for growth were developed for 4 strains of A. flavus • Aflatoxin B 1 production was optimal at 0.99 water activity and 30 °C, regardless of strain. • Optimum and marginal toxin production contour maps were determined for toxigenic strains [ABSTRACT FROM AUTHOR]

The objective was to screen and evaluate the anti-fungal activity of lactic acid bacteria (LABs) isolated from Malaysian fermented foods against two Trichophyton species. A total of 66 LAB strains were screened using dual culture assays. This showed that four LAB strains were very effective in inhibiting growth of T. rubrum but not T. interdigitale. More detailed studies with Lactobacillus plantarum strain HT-W104-B1 showed that the supernatant was mainly responsible for inhibiting the growth of T. rubrum. The minimum inhibitory concentration (MIC), inhibitory concentration, the 50% growth inhibition (IC50) and minimum fungicide concentration (MFC) were 20 mg/mL, 14 mg/mL and 30 mg/mL, respectively. A total of six metabolites were found in the supernatant, with the two major metabolites being L-lactic acid (19.1 mg/g cell dry weight (CDW)) and acetic acid (2.2 mg/g CDW). A comparative study on keratin agar media showed that the natural mixture in the supernatants predominantly contained L-lactic and acetic acid, and this significantly controlled the growth of T. rubrum. The pure two individual compounds were less effective. Potential exists for application of the natural mixture of compounds for the treatment of skin infection by T. rubrum. [ABSTRACT FROM AUTHOR]

Raspberry production is under threat from the emerging fungal pathogenic genus Cladosporium. We used amplicon‐sequencing, coupled with qPCR, to investigate how fruit age, fruit location within a polytunnel, polytunnel location and sampling date affected the fruit epiphytic microbiome. Fruit age was the most important factor impacting the fungal microbiome, followed by sampling date and polytunnel location. In contrast, polytunnel location and fruit age were important factors impacting the bacterial microbiome composition, followed by the sampling date. The within‐tunnel location had a small significant effect on the fungal microbiome and no effect on the bacterial microbiome. As fruit ripened, fungal diversity increased and the bacterial diversity decreased. Cladosporium was the most abundant fungus of the fruit epiphytic microbiome, accounting for nearly 44% of all fungal sequences. Rotorod air samplers were used to study how the concentration of airborne Cladosporium inoculum (quantified by qPCR) varied between location (inside and outside the polytunnel) and time (daytime vs. nighttime). Quantified Cladosporium DNA was significantly higher during the day than the night and inside the polytunnel than the outside. This study demonstrated the dynamic nature of epiphytic raspberry fruit microbiomes and airborne Cladosporium inoculum within polytunnels, which will impact disease risks on raspberry fruit. [ABSTRACT FROM AUTHOR]

Maize is prone to infection by Aspergillus flavus , which can contaminate the product with aflatoxins. The objective of this study was to examine the impact that interactions between water activity (a w ) and temperature may have on growth, the expression of a biosynthetic regulatory gene ( aflR ) and aflatoxin B 1 (AFB 1 ) production by a strain of A. flavus on a maize-based medium. Results showed that there were some differences between lag phases and growth rates of A. flavus . The optimum growth rate for A. flavus was at 30 °C and 0.99 a w . No growth occurred at 0.90 a w and 20 °C. Both temperature and a w had an influence on the relative aflR gene expression and AFB 1 production by A. flavus ; however, the results for AFB 1 production were not consistent with the effects on gene expression or growth. These findings provide data that are useful to better understand the conditions which represent higher risks from AFB 1 production. However, the aflR expression was not a good indicator of AFB 1 production alone. Thus, further molecular studies of other AF-related genes should be done. These results are discussed in the context of harvesting and storage of maize and the prevailing environmental conditions to minimise AFB 1 contamination. [ABSTRACT FROM AUTHOR]

Fungi threaten the security of food supply to human populations on several fronts. They destroy up to 30 % of crop products through disease and spoilage processes, while mycotoxin-producing fungi and opportunistic pathogens endanger food safety. Control of these fungi is vital for improving food security, but current measures are inadequate and further challenges due to human-population growth and climate change are escalating. Investment and innovation in research on strategies to control fungal growth, harnessed through international, inter-disciplinary collaboration across socio-economic boundaries, provides one key to rising to this challenge. [ABSTRACT FROM AUTHOR]

This study examined the effect of different elicitors (seven, different concentrations) and environmental factors (water activity (a w ), pH) on taxol production by strains of two endophytic fungi, Paraconiothyrium variabile and Epicoccum nigrum, isolated from temperate yew trees. A defined liquid broth medium was modified with elicitors, solute a w depressors at different pH values. For P. variabile , the best elicitor was salicylic acid at 50 mg/l which gave a taxol yield of 14.7 ± 4.8 μg/l. The study of synergistic effects between elicitor, a w and pH on taxol production showed that the highest yield of taxol (68.9 ± 11.9 μg/l) was produced under modified ionic stress of 0.98 a w (KCl) at pH 5 when supplemented with 20 mg/l of salicylic acid. For E. nigrum, serine was the best elicitor which increased yield significantly (29.6 fold) when KCL was used as the a w depressor (0.98 a w ) at pH 5.0 with 30 mg/l of serine. The maximum taxol yield produced by E. nigrum was 57.1 ± 11.8 μg/l. Surface response models were used to build contour maps to determine the conditions for maximum and marginal conditions for taxol yield in relation to the best elicitor and a w , and the best pH for the first time. This will be beneficial for identifying key parameters for improvement of taxol yields by endophytic fungi. [ABSTRACT FROM AUTHOR]

Alternaria species are common saprophytes or pathogens of a wide range of plants pre- and post-harvest. This review considers the relative importance of Alternaria species, their ecology, competitiveness, production of mycotoxins and the prevalence of the predominant mycotoxins in different food products. The available toxicity data on these toxins and the potential future impacts of Alternaria species and their toxicity in food products pre- and post-harvest are discussed. The growth of Alternaria species is influenced by interacting abiotic factors, especially water activity (aw), temperature and pH. The boundary conditions which allow growth and toxin production have been identified in relation to different matrices including cereal grain, sorghum, cottonseed, tomato, and soya beans. The competitiveness of Alternaria species is related to their water stress tolerance, hydrolytic enzyme production and ability to produce mycotoxins. The relationship between A. tenuissima and other phyllosphere fungi has been examined and the relative competitiveness determined using both an Index of Dominance (ID) and the Niche Overlap Index (NOI) based on carbon-utilisation patterns. The toxicology of some of the Alternaria mycotoxins have been studied; however, some data are still lacking. The isolation of Alternaria toxins in different food products including processed products is reviewed. The future implications of Alternaria colonization/infection and the role of their mycotoxins in food production chains pre- and post-harvest are discussed. [ABSTRACT FROM AUTHOR]

The effect of water activity (0.90, 0.94, and 0.98 a w ) and temperature (15, 20, and 25 °C) on the in vitro interactions between three ochratoxigenic strains of Aspergillus carbonarius (Ac-28, Ac-29, and Ac-33) and eleven non ochratoxigenic grape-associated fungal strains was assessed in this study. Fungal strains were allowed to grow in dual cultures on Synthetic Grape-juice Medium (SGM) for 15 days and fungal interactions were given a numerical score to obtain an Index of Dominance ( I D ) for each fungus. Results showed that in most cases A. carbonarius toxigenic strains were dominant against other fungal species. However, A. carbonarius presented mutual antagonism with A. section Nigri strains regardless of water activity ( a w ) and temperature. Moreover, interactions with Penicillium spinulosum and Cladosporium spp. at 15 °C, as well as Botrytis cinerea at 20 °C, showed that the antagonists were more competitive against A. carbonarius . In some cases, growth rates of A. carbonarius strains were either slightly stimulated or inhibited after interaction in dual cultures, depending on temperature, a w and competing species. Regarding OTA production, the presence of other species sometimes decreased the production or slightly enhanced it, depending on fungal competitor and environmental conditions. Overall, OTA production was higher at 15 °C/0.98 a w and 20 °C/0.98 a w for all target strains and at 20 °C/0.94 a w for Ac-33 strain only, but decreased at higher temperatures regardless of a w and interacting species. [ABSTRACT FROM AUTHOR]

Grey mould rot (Botrytis cinerea) development in vitro or in eggplant (Solanum melongena L.) fruit was evaluated after treatment with dittany (Origanum dictamnus L.) oil (DIT) and storage at 12°C and 95% relative humidity during or following exposure to the volatiles. DIT volatiles used in different concentration (0-50-100-250 μL/L) and times of exposure (up to 120 h) examined the effects on pathogen development as well as fruit quality parameters. In vitro, fungal colony growth was inhibited with the application of DIT oil (during or after exposure) and/or time of application. Continuous exposure to oils reduced conidial germination and production with fungistatic effects observed in 250 μL/L. In vivo, fungal lesion growth and conidial production reduced in DITtreated fruits. Interesting, in fruits preexposed to volatiles before fungal inoculation, DIT application induced fruit resistance against the pathogen, by reduced lesion growth and conidial production. Conidial viability reduced in >100 μL/L DIT oil. Fruits exposed to essential oil did not affect fruit quality related attributes in general, while skin lightness (L value) increased in 50 and 100 μL/L DIT oil. The results of the current study indicated that dittany volatiles may be considered as an alternative food preservative, eliminating disease spread in the storage/transit atmospheres. [ABSTRACT FROM AUTHOR]

The potential of three atoxigenic strains from different geographical origins in Africa were examined forin vitroandin situcompetitiveness against two toxigenic strains ofAspergillus flavusand subsequent inhibition of aflatoxin B1(AFB1) production under different environmental conditions. Temperature, water activity (aw) and substrate influenced the types of interaction between the three AFL−and two AFL+strains. The competitiveness and AFB1reduction ability of the three atoxigenic strains when interacting with the two toxigenic strains were evaluated by inoculation of 100, 25:75, 50:50, 75:25 and 100% ratios of mixed spore suspensionsin vitroon malt extract and milled maize agars over 28 days andin situon stored maize grain for 14 days, respectively at 0.99, 0.96 and 0.90aw. For all the treatments, the effect ofawand inoculum ratio and their interaction was highly significant. Toxin inhibition was >80%in vitroat both 0.99 and 0.96aw.In situAFB1reduction was influenced by the toxigenic strain assayed,awand the inoculum ratio. Where control was achieved, it was more variable at 0.96aw, while with more stringent water stress conditions (0.90aw) the percentage inhibition was up to 77.2%. The study shows the importance of including environmental factors in screening and identifying effective atoxigenic strains for control of AFs (aflatoxins). [ABSTRACT FROM AUTHOR]

This study examined the relationship between storage environmental factors (water activity (aw) (0.89–0.97) and temperature (15°C–30°C)), colonisation of wheat and maize by Fusarium graminearum and F. verticillioides respectively and the dry matter losses (DMLs) caused and quantified by contamination with deoxynivalenol (DON), zearalenone (ZEA) and fumonisins (FUMs) during storage. Fungal growth was assessed by the amount of CO2 produced under different interacting conditions of aw and temperature. DMLs were quantified using the cumulative CO2 data, and these were shown to increase as temperature and aw increased. The amount of DON, ZEA (wheat for human consumption) and FUMs (feed maize) produced was significantly affected by the storage conditions. The three toxins however showed different patterns of production. Optimum for DON was at the wettest conditions (0.97aw) and the highest temperature assessed (30°C), whereas for ZEA this shifted to 25°C. FUMs were produced in higher amounts in maize at 30°C and 0.97aw; however, at intermediate aw levels (0.955aw), the highest production occurred at 25°C followed by 20°C. Polynomial models were developed for the effect of the storage factors on DMLs and toxin production. DMLs under different environmental conditions were significantly correlated with DON and FUMs. DON contamination was above the EU limits in at least 80% of the wheat samples with DMLs >1%, whereas at least 70% of the same samples contained ZEA above the respective EU legislative limits. Similarly, at least 75% of the maize samples with DMLs ≥ 0.9% exceeded the EU limits for the sum of FUMs in feed. These results show that it may be possible to use temporal CO2 production during storage of grains as an indicator of the level of contamination of the grain with mycotoxins. [ABSTRACT FROM AUTHOR]

Abstract: A key step in the infection cycle by Aspergillus flavus in maize is sporulation of sclerotia present in soil or in crop debris. However, little information is available on this critical and important phase. This study included experiments on artificial (Czapek Dox Agar (CZ)) and natural (maize stalks) substrates under different conditions of temperature (T; from 5 to 45°C) and water activity (a w; from 0.50 to 0.99) levels to quantify sporulation from sclerotia. The mean numbers of spores were higher on defined nutritional medium in vitro on CZ agar than on maize stalks (4.5×106 spores/sclerotium versus 4.2×104 spores/sclerotium) with production initiated after 6 and 24h, respectively. Surprisingly, the optimal temperature was found at 30–35°C for CZ agar (9.23×106 spores/sclerotium) and to be 20–25°C for maize stalks (6.26×104 spores/sclerotium). Water stress imposition only reduced sporulation at ≤0.90 a w. With more available water no significant differences were found between 0.90 and 0.99 a w. This type of data is critical in the development of a mechanistic model to predict the infection cycle of A. flavus in maize in relation to meteorological conditions. [Copyright &y& Elsevier]

A membrane based enzyme linked immunosorbent assay (MELISA) for the detection of a common herbicide, isoproturon is described. A heterogeneous competitive ELISA was the format chosen for isoproturon detection. An immunoassay system with a horseradish peroxidase (HRP) labeled polyclonal antibody preparation was developed and characterized before suitable sensitivity and selectivity for isoproturon were attained. After development as a microtiter plate immunoassay, the system was transferred to an affinity membrane sorbent based ELISA where the isoproturon/ovalbumin conjugate was immobilized on commercial membranes. Different porosities and immobilization conditions were utilized to optimize the MELISA, including sensitivity, selectivity, and stability studies. This enabled detection of isoproturon in the range 0.5 ng ml−1–20 µg ml−1, with an LLD90 of 0.5 ng ml−1. The use of acetonitrile extracts from soil samples was found to not overly impair the performance of the MELISA. Good correlation between ELISA and HPLC was obtained for extracts from spiked soil samples. [ABSTRACT FROM PUBLISHER]

Abstract: A rapid method for screening antifungal compounds and performing ecophysiological studies with filamentous fungi has been developed by the use of specific semi-solid media and spectrophotometric/turbidimetric measurements using the Bioscreen C with 2×100 microtitre well plates. The medium composition and preparation, inoculum size, medium volumes, and incubation parameters for measuring initial germination and growth dynamics have been optimised. These have been applied to assess the effectiveness of 18 concentrations of propyl propane thiosulfinate (PTS) against Aspergillus flavus in YES medium under different environmental regimes. Minimum inhibitory concentrations (MIC), and non-inhibitory concentration (NIC) values, and a newly developed MIC50, have been calculated for the efficacy of these concentrations of PTS under four different environmental conditions (0.995 and 0.95 water activity (a w); 20 and 25°C) over 7d periods with automated measurements every 20min. Data were modelled using the Lambert–Pearson Model, a mathematical modelling approach previously used for bacterial inhibition. Results have been compared with traditional growth rate data and Lethal Dose50 (LD50) values. This approach could have major implications for rapid screening assays for growth and secondary metabolite production by filamentous organisms and has major advantages in terms of time reduction, culture medium volumes required, measurement and the ability to integrate and model key parameters for comparing efficacy. [Copyright &y& Elsevier]

Abstract: A membrane based heterogeneous competitive enzyme-linked immunosorbent assay (ELISA) was used in this work to develop an immunosensor for the detection of a common herbicide, isoproturon. A screen-printed carbon working electrode with carbon counter and silver–silver chloride pseudo-reference electrode was utilized incorporating a membrane fixed into intimate contact with the working electrode to facilitate signal transduction. The membrane containing an immobilized isoproturon–ovalbumin conjugate was laminated onto the carbon working electrode and horseradish peroxidase (HRP) labeled polyclonal antibody was then applied for the competitive assay. Two different amperometric systems, hydroquinone and o-phenylenediamine (OPD) mediation reduction were utilised and the properties of the resultant sensors were compared. A flow injection apparatus was also developed utilising the immunosensor. Limits of detection for isoproturon (LLD90) were found to be as low as 0.84ngmL−1. The senor was also validated using spiked extracted soil samples and also isoproturon contaminated samples. [Copyright &y& Elsevier]

Abstract: The objective of this work was to study the effect of ecophysiological factors on trichothecene gene expression and growth in Fusarium graminearum. The effect of non-ionic solute water potentials and temperature was examined on in vitro mycelial growth rates and on expression of the TRI5 gene, involved in trichothecene biosynthesis, quantified by real time RT–PCR. This study showed optimal values of 25°C and −2.8MPa (0.982 a w) for growth. Marginal temperatures such as 15°C and 30–35°C, particularly in combination with water potentials below −2.8MPa, drastically reduced growth. The expression of TRI5 was reasonably constant although some induction was observed between 20 and 30°C, the most favourable temperatures for growth, depending on the water potential imposed, particularly at −7.0MPa. A temporal kinetic experiment at 25°C examined the effect of ionic solute stress on TRI5 gene expression and growth rate. The results indicated independence of growth rate and TRI5 expression, as the fungal biomass increased with time while the gene expression remained constant. This suggested that favourable conditions for growth will result in higher trichothecene production, and that toxin production would always accompany the colonization process at a steady rate while the conditions for growth are permissive. Quantification of key biosynthetic toxin genes by real time RT–PCR was shown to be a valuable tool to gain knowledge of the ecophysiological basis for trichothecene biosynthesis and enable better control strategies to be developed during the life cycle of this important mycotoxigenic pathogen of cereals. [Copyright &y& Elsevier]