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AimWe 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 resultsTo 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.ConclusionsThis 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.

Aims: A relative quantification system (RQ-PCR) was used to monitor the correlations between the activity of the nor-1 (=aflD) gene of Aspergillus flavus using real-time PCR in relation to phenotypic aflatoxin B1 (AFB1) production and populations of A. flavus in stored peanuts at three water activity levels (aw, 0·95, 0·90 and 0·85) for 6 weeks. Methods and Results: Real-time PCR was used to amplify the nor-1 gene (target gene), and benA56 (β-tubulin gene) used as a control gene. Expression of three structural genes, nor-1 (=aflD), ver-1 (=aflM), and omtA (=aflP), and the regulatory gene aflR of the aflatoxin biosynthetic pathway were also assayed. There were significant differences between nor-1 gene expression at the three aw levels; higher expression at 0·90 aw in weeks 1–3, when compared to 0·95. In contrast, in the driest treatment (0·85 aw) none or very low nor-1 expression occurred. The populations of A. flavus colony-forming units (CFUs g−1) increased over time with the highest at 0·95 aw. Highest AFB1 production was at 0·90 and 0·95 aw from weeks 3–6. Aw had a significant effect on aflR transcription at 0·95 aw over the 6-week period, while at 0·90 aw, only in the last 2 weeks. Conclusions: Correlations between different factors showed that log AFB1 × log CFUs, log AFB1 × aw, and log CFUs × aw were statistically significant, while log CFUs × RQ-PCR and RQ-PCR × aw were not. The AflR gene may not have an important role in the regulation of nor-1 expression in food matrices (e.g. peanuts). Significance and Impact of the study: Determination of correlations between nor-1 expression and aflatoxin production by A. flavus in raw peanuts under different aw levels could be helpful to predict potential risk of aflatoxin production during storage of this hygroscopic food product and minimize contamination with the AFB1.

Aims: To develop and validate a logistic regression model to predict the growth and ochratoxin A (OTA) production boundaries of two Aspergillus carbonarius isolates on a synthetic grape juice medium as a function of temperature and water activity (aw). Methods and Results: A full factorial design was followed between the factors considered. The aw levels assayed were 0·850, 0·880, 0·900, 0·920, 0·940, 0·960, 0·980 and the incubation temperatures were 10, 15, 20, 25, 30, 35 and 40°C. Growth and OTA production responses were evaluated for a period of 25 days. Regarding growth boundaries, the degree of agreement between predictions and observations was >99% concordant for both isolates. The erroneously predicted growth cases were 3·4–4·1% false-positives and 0·7–1·4% false-negatives. No growth was observed at 10°C and 40°C for all aw levels assayed, with the exception of 0·980 aw/40°C, where weak growth was observed. Similarly, OTA production was correctly predicted with a concordance rate >98% for the two isolates with 0·7–1·4% accounting for false-positives and 2·0–2·7% false-negatives. No OTA production was detected at 10°C or 40°C regardless of aw, and at 0·850 aw at all incubation temperatures. With respect to time, the OTA production boundary shifted to lower temperatures (15–20°C) as opposed to the growth boundary that shifted to higher temperature levels (25–30°C). Using two literature datasets for growth and OTA production of A. carbonarius on the same growth medium, the logistic model gave one false-positive and three false-negative predictions out of 68 growth cases and 13 false-positive predictions out of 45 OTA production cases. Conclusions: The results of this study suggest that the logistic regression model can be successfully used to predict growth and OTA production interfaces for A. carbonarius in relation to temperature and aw. Significance and Impact of the Study: The proposed modelling approach helps the understanding of fungal-food ecosystem relations and it could be employed in risk analysis implementation plans to predict the risk of contamination of grapes and grape products by A. carbonarius.

Aims: To examine the efficacy of natamycin produced by Streptomyces natalensis against strains of Aspergillus carbonarius growth and ochratoxin A (OTA) production under different environmental factors on a grape juice-based medium. Methods and Results: Detailed studies in the range 0–20 ng ml−1 for control of growth and ochratoxin production by strains of A. carbonarius at 0·98, 0·96 and 0·94 water availabilities (aw) and 15–25°C on a fresh red grape extract medium were examined. Inhibition of growth was depending on temperature and aw level. At 15°C, 5–10 ng ml−1 natamycin was effective in reducing growth almost completely. However, at 20–25°C and all the three aw levels, growth was only slightly inhibited by 5–10 ng ml−1 natamycin. There were strain differences with regard to inhibition of OTA production. At 15°C and 0·98 aw, 10 ng ml−1 was required to inhibit production by >90%. However, at 0·96 and 0·94 aw, almost complete inhibition occurred. At 20°C, OTA production was only significantly inhibited by 10 ng ml−1 natamycin at 0·94 aw. At 0·96 and 0·98 aw, some inhibition occurred with 5–10 ng ml−1, but greater concentrations would be required for effective inhibition. At 25°C, 5 ng ml−1 was effective at all aw levels. However, at 15°C and 25°C and a wide range of aw levels, natamycin effectively controlled OTA production. Conclusions: Natamycin appears to be a very effective for controlling growth and OTA production by strains of A. carbonarius over a range of aw and temperature conditions on grape-based media. Significance and Impact of the Study: This is the first detailed study to demonstrate the impact of natamycin against A. carbonarius. This study suggests that use of natamycin at 50–100 ng ml−1 can give complete inhibition of growth of A. carbonarius and OTA production over a range of environmental conditions. Natamycin could be an important component of a system to prevent OTA contamination of wine as well during the drying and production of vine fruits.

Aims: To develop descriptive models for the combined effect of temperature (10–40°C) and water activity (0·850–0·980) on the growth of two ochratoxin A producing strains of Aspergillus carbonarius from Greek wine grapes on a synthetic grape juice medium. Methods and Results: Fungal growth was measured as changes in colony diameter on a daily basis. The maximum specific colony growth rates (μmax) were determined by fitting the primary model of Baranyi describing the change in colony diameter (mm) with respect to time (days). Secondary models, relating μmax with temperature and aw were developed and comparatively evaluated based on polynomial, Parra, Miles, Davey and Rosso equations. No growth was observed at 0·850 aw (water activity) regardless of temperature, as well as at marginal temperature levels assayed (10 and 40°C) regardless of water activity. The data set was fitted successfully in all models as indicated by the values of regression coefficients and root mean square error. Models with biological interpretable parameters were highly rated compared with the polynomial model, providing realistic cardinal values for temperature and aw. The optimum values for growth were found in the range 0·960–0·970 aw and 34–35°C respectively for both strains. The developed models were validated on independently derived data from the literature and presented reasonably good predictions as inferred by graphical plots and statistical indices (bias and accuracy factors). Conclusions: The effect of temperature and aw on the growth of A. carbonarius strains could be satisfactorily predicted under the current experimental conditions, and the proposed models could serve as a tool for this purpose. Significance and Impact of the Study: The results could be successfully employed as an empirical approach in the development and prediction of risk models of contamination of grapes and grape products by A. carbonarius.

Aims To develop a model for the combined effect of water activity (a(w)) and temperature on growth of strains of Aspergillus niger, and comparison with data on food spoilage moulds in the literature. Methods and results An extended combined model describing the growth of two strains of A. niger, as a function of temperature (25-30 degrees C) and a(w) (0.90-0.99) was developed. The growth rate (micro) was expressed as the increase in colony radial growth per unit of time. This extends the previous square root model showing the relationship between temperature and bacterial growth rate developed by Ratkowsky et al. (1983) and the parabolic relationship between the logarithm of the growth rate and a(w) developed by Gibson et al. (1994). A good correlation between the experimental data and the model predictions was obtained, with regression coefficients (r(2)) > 0.99. In addition, the use of this model allowed predictions of the cardinal a(w) levels: a(w(min)), and a(w(opt)). The estimation of the minimum a(w) levels (a(w(min))) was in accordance with data in the literature for similar and a range of other Aspergillus and related species, regardless of the solutes used for a(w) modification. The estimation of the optimal a(w) (a(w(opt))) and the optimal growth rate (micro(opt)) were in good agreement with the experimental results and data from the literature. Conclusions This approach enables accurate prediction of the combined effects of environmental factors on growth of spoilage fungi for rapid prediction of cardinal limits using surface response curves. Significance and impact of the study This approach is a rapid method for predicting optimal and marginal conditions for growth of a wide range of spoilage micro-organisms in relation to interacting environmental conditions and will have applications for improving shelf-life of intermediate moisture foods.

Aims: This study investigated the in vitro effects of water activity (aw; 0·85–0·987) and temperature (10–40°C) on growth and ochratoxin A (OTA) production by two strains of Aspergillus carbonarius isolated from wine grapes from three different European countries and Israel on a synthetic grape juice medium representative of mid-veraison (total of eight strains). Methods and Results: The synthetic grape juice medium was modified with glycerol or glucose and experiments carried out for up to 56 days for growth and 25 days for OTA production. The lag phase prior to growth, growth rates and ochratoxin production were quantified. Statistical comparisons were made of all factors and multiple regression analysis used to obtain surface response curves of aw × temperature for the eight strains and optimum growth and OTA production by A. carbonarius. The lag phase increased from 20 days at marginal temperatures and water availabilities. Generally, most A. carbonarius strains grew optimally at 30–35°C, regardless of solute used to modify aw, with no growth at

Aims: The effect of osmotic and matric potential stress on growth and sugar alcohols (polyols: glycerol, erythritol, arabitol and mannitol) and sugars (trehalose and glucose) accumulation in toxigenic and nontoxigenic colonies of Aspergillus flavus and A. parasiticus was evaluated. Methods and Results: Growth of Aspergillus section Flavi with significant reductions at 20 and 30°C was more sensitive to changes in matric potential, between 60 and 100% in the range of −7 to −14 MPa. No significant differences were found between toxigenic and nontoxigenic strains for both species. Total polyol accumulation in unamended maize meal agar medium (−0·75 MPa water potential) was higher at 30 than 20°C. The major change in concentrations of endogenous sugars and total polyols was in matrically amended medium (with PEG 8000) at −7 and −10 MPa. Accumulation of glucose, arabitol, mannitol and erythritol content of A. flavus and A. parasiticus mycelial colonies was greater in normal unstressed maize meal agar medium (−0·75 Mpa) at 20°C. This was modified by solute and matric stress. Conclusions: The data showed relative sensitivity to osmotic and matric potential, and temperature, and the impact on growth rates, polyol and sugar accumulation in mycelia of A. flavus and A. parasiticus. Significance and Impact of the Study: The matric potential effects on growth may be of particular importance for growth and survival in environments with low-matric potential stress. The tolerance of spoilage fungi such as Aspergillus section Flavi to such modifications could increase the potential for spoilage and mycotoxin production in such substrates. This knowledge is important for understanding the relative ecological fitness of these aflatoxigenic species and in the development of prevention strategies for their control.

Aims: Early detection of spoilage fungi (two Eurotium spp., a Penicillium chrysogenum species) in bread analogues over periods of 72 h at 25∞C and 0AE95 water activity was evaluated using volatile production patterns, hydrolytic enzyme production, and changes in fungal populations. Methods and Results: Using an electronic nose system it was possible to differentiate between uninoculated controls and samples contaminated with P. chrysogenum within 28 h. After 40‐48 h it was possible to differentiate between the Eurotium spp., P. chrysogenum and the control using Principal Component Analysis (PCA). Cluster analyses could differentiate between the control, P. chrysogenum and the Eurotium spp. after 40 h. Of seven hydrolytic enzymes examined after 48 h, the specific activities of three were significantly different from uninoculated control bread. There were also differences between the mould species in production of three enzymes. Penicillium chrysogenum populations increased fastest, from about 10 3 cfu g ‐1 to 10 6 ‐10 7 cfu g ‐1 after 72 h. For the Eurotium spp. this increase was slower. Conclusions: Overall, this study suggests, for the first time, that an electronic nose system using a surface polymer sensor array is able to detect qualitative changes in volatile production patterns for the early detection of the activity of spoilage moulds in bakery products. Significance and Impact of the Study: Potential exists for application of such systems for microbial quality assurance in intermediate moisture food products.

Volatile profiles and hydrolytic enzyme production by one non-mycotoxigenic and three mycotoxigenic strains of Fusarium moniliforme and F. proliferatum, grown in vitro for up to 96 h on a grain medium at 25 degrees C/0.95 water activity, were examined for differentiation of isolates. After spore lawn inoculation, measurements were made after 48, 72 and 96 h by sampling the head space above cultures with an electronic nose system using a 14 sensor surface polymer array, and by extraction and quantification of hydrolytic enzymes. There was good reproducibility of volatile patterns between replicates of the same treatment. Principal component analysis indicated that discrimination could be achieved between the uninoculated controls, the non-mycotoxigenic strain and the mycotoxin-producing strains for both species after 48 h. The total and specific activity of three out of seven enzymes (beta-D-glucosidase, alpha-D-galactosidase and N-acetyl-beta-D-glucosaminidase) were found to increase significantly in the non-mycotoxigenic when compared with the toxigenic strains of both species after 72 h. Activities of the others (beta-D-fucosidase, alpha-D-mannosidase, beta-D-xylosidase and N-acetyl-alpha-D-glucosaminidase) were not significantly different between strains. The study has shown for the first time that it is possible to differentiate between mycotoxigenic and non-mycotoxigenic strains of such spoilage fungi based on their volatile production patterns using an electronic nose system. These results have significance in the development of methods for the early detection of toxin-producing spoilage moulds in the food industry.

N. T EIXIDO ´, I. VINAS, J. USALL, V. SANCHIS AND N. MAGAN. 1998.The growth responses of the biocontrol agent Candida sake to changes in water activity (aw), temperature and pH and their interactions, and accumulation of sugars (glucose, trehalose) and sugar alcohols (glycerol, erythritol, arabitol and mannitol) were determined in vitro in nutrient yeast dextrose based media. The aw temperature profile for growth was between 0·995 and 0·90 and 4‐37 °C with the non-ionic solute glycerol, and between 0·995 and 0·92 and 10‐30 °C with the ionic solute NaCl. Regardless of solute, there was a longer lag time prior to growth as aw was reduced, and at marginal temperatures for growth. Relative growth rates were compared at different aw levels and temperatures and it was found that aw, temperature, solute and two and three‐way interactions were statistically significant. By contrast, C. sake was tolerant of a wide range of pH levels (3‐7) regardless of aw, although growth rates were reduced at marginal temperatures and aw. In non-stressed basal NYDB, glucose and arabitol were the predominant endogenous reserves accumulated in the cells of C. sake. However, when nutrient status was diluted (75%) and stressed by the addition of glycerol or NaCl (0·98 and 0·96 aw), significant changes in the accumulation of sugars and sugar alcohols occurred. In glycerol-stressed media, glucose and glycerol were the major compounds accumulated, with markedly lower arabitol and little trehalose or mannitol present. With NaCl-stressed media, glycerol was the only sugar alcohol accumulated, with very low levels of the sugars and other sugar alcohols. This study has defined the ecological niche within which C. sake would be active as a biocontrol agent for the first time and suggests that endogenous reserves can be significantly modified by nutrient and aw stress; these changes may be useful for improving environmental competence of such micro-organisms in the environment.

To develop descriptive models for the combined effect of temperature (10-40 degrees C) and water activity (0.850-0.980) on the growth of two ochratoxin A producing strains of Aspergillus carbonarius from Greek wine grapes on a synthetic grape juice medium.Fungal growth was measured as changes in colony diameter on a daily basis. The maximum specific colony growth rates (mu(max)) were determined by fitting the primary model of Baranyi describing the change in colony diameter (mm) with respect to time (days). Secondary models, relating mu(max) with temperature and a(w) were developed and comparatively evaluated based on polynomial, Parra, Miles, Davey and Rosso equations. No growth was observed at 0.850 a(w) (water activity) regardless of temperature, as well as at marginal temperature levels assayed (10 and 40 degrees C) regardless of water activity. The data set was fitted successfully in all models as indicated by the values of regression coefficients and root mean square error. Models with biological interpretable parameters were highly rated compared with the polynomial model, providing realistic cardinal values for temperature and a(w). The optimum values for growth were found in the range 0.960-0.970 a(w) and 34-35 degrees C respectively for both strains. The developed models were validated on independently derived data from the literature and presented reasonably good predictions as inferred by graphical plots and statistical indices (bias and accuracy factors).The effect of temperature and a(w) on the growth of A. carbonarius strains could be satisfactorily predicted under the current experimental conditions, and the proposed models could serve as a tool for this purpose.The results could be successfully employed as an empirical approach in the development and prediction of risk models of contamination of grapes and grape products by A. carbonarius.

To examine the efficacy of natamycin produced by Streptomyces natalensis against strains of Aspergillus carbonarius growth and ochratoxin A (OTA) production under different environmental factors on a grape juice-based medium.Detailed studies in the range 0-20 ng ml(-1) for control of growth and ochratoxin production by strains of A. carbonarius at 0.98, 0.96 and 0.94 water availabilities (a(w)) and 15-25 degrees C on a fresh red grape extract medium were examined. Inhibition of growth was depending on temperature and a(w) level. At 15 degrees C, 5-10 ng ml(-1) natamycin was effective in reducing growth almost completely. However, at 20-25 degrees C and all the three a(w) levels, growth was only slightly inhibited by 5-10 ng ml(-1) natamycin. There were strain differences with regard to inhibition of OTA production. At 15 degrees C and 0.98 a(w), 10 ng ml(-1) was required to inhibit production by90%. However, at 0.96 and 0.94 a(w), almost complete inhibition occurred. At 20 degrees C, OTA production was only significantly inhibited by 10 ng ml(-1) natamycin at 0.94 a(w). At 0.96 and 0.98 a(w), some inhibition occurred with 5-10 ng ml(-1), but greater concentrations would be required for effective inhibition. At 25 degrees C, 5 ng ml(-1) was effective at all a(w) levels. However, at 15 degrees C and 25 degrees C and a wide range of a(w) levels, natamycin effectively controlled OTA production.Natamycin appears to be a very effective for controlling growth and OTA production by strains of A. carbonarius over a range of a(w) and temperature conditions on grape-based media.This is the first detailed study to demonstrate the impact of natamycin against A. carbonarius. This study suggests that use of natamycin at 50-100 ng ml(-1) can give complete inhibition of growth of A. carbonarius and OTA production over a range of environmental conditions. Natamycin could be an important component of a system to prevent OTA contamination of wine as well during the drying and production of vine fruits.

Aims: Aspergillus carbonarius is an important ochratoxin A (OTA)-producing fungus which is responsible for toxin contamination of grapes and wine. The objectives of this study were to examine the partitioning of OTA in mycelium and conidia of a range of A. carbonarius strains on artificial grape juice and defined media, to determine the excretion patterns of OTA from these spores, and the effect of organic acids used in wine production on OTA excretion from conidia. Methods and Results: The results showed that 60–70% of the OTA was accumulated in the conidia of a number of different isolates of A. carbonarius. Calculations showed that on different defined media, an amount of 0·011- to 0·1-pg OTA was present per conidium. The OTA in spores was found to be rapidly excreted into the medium during the initial few hours after conidial germination leading to an increase of OTA in must during maceration for wine production. The presence of tartaric acid inhibited OTA production, but malic acid enhanced this production during mycelial growth. These acids were also shown to affect the time course of germination and the rate of OTA excretion from conidia during germination. Conclusions: This study is the first to examine and show the partitioning of OTA into spores of strains of A. carbonarius and that rapid excretion of OTA from spores could be a reason for OTA accumulation in musts during wine production. Significance and Impact of the Study: Conidia of A. carbonarius could be a major source of OTA contamination of grapes used in wine production. This information could help in the development of effective prevention strategies to minimize wine contamination with this important mycotoxin.

Aims: To examine the effect of interactions between water, temperature and gas composition on growth and ochratoxin A (OTA) production by isolates of Penicillium verrucosum in vitro and in situ on grain-based media and wheat grain. Methods and Results: Three isolates of P. verrucosum were examined in relation to radial growth rate and OTA production, and to interacting conditions of water activity (aw), temperature and gas composition on a milled wheat medium. Subsequently, detailed temporal studies were carried out on gamma irradiated wheat grain over the range 0·75–0·995 aw, 10–25°C and air, 25 or 50% CO2. This showed that optimum growth of P. verrucosum was at 0·98 aw in vitro at 25°C, but at 0·95 aw and 25°C on wheat grain. The aw minimum for growth was about 0·80 aw, although no OTA was produced under this condition even after 56 days. Significant inhibition of growth and OTA production occurred with 50% CO2, and 0.90–0·995 aw at 25°C. Conclusions: The optimum and marginal conditions for growth and OTA production on wheat grain have been identified. At least 50% CO2 is needed to inhibit growth and OTA production by >75% in moist grain (0·90–0·995 aw). Significance and Impact of the Study: First detailed identification of optimal and marginal interacting conditions of water/temperature and gas composition on growth and OTA production by P. verrucosum on wheat grain. This is a critical component of the postharvest management strategy for minimizing contamination by this important mycotoxin and predicting risk, based on environmental conditions, during drying and storage.

This study compared the effect of temperature (5-45 degrees C), water availability (water activity, aw; 0.995-0.75) and their interactions on the temporal rates of germination and mycelial growth of three mycotoxigenic strains of Aspergillus ochraceus and one isolate each of A. flavus, A. niger, Penicillium aurantiogriseum and P. hordei in vitro on a maize extract medium. Germination was very rapid at0.90 aw with an almost linear increase with time for all species. However, at0.90 aw, the germination rates of A. flavus and P. hordei were slower. The aw minima for germination were usually lower than for growth and varied with temperature. The effect of aw x temperature interactions on the lag phases (h), prior to germination, and on the germination rates (h(-1)), were predicted for the first time for these fungi using the Gompertz model modified by Zwietering. This showed that A. flavus, A. niger and the two Penicillium spp. had very short lag times between 0.995-0.95 aw over a wide temperature range. At marginal temperatures, these were significantly higher, especially at10 degrees C for Aspergillus spp. and30 degrees C for Penicillium spp. There were also statistically significant differences between lag phases and germination rates for three different isolates of A. ochraceus. The Aspergillus spp. also germinated faster than the Penicillium spp. The temperature x aw profiles for mycelial growth varied considerably between species, both in terms of rates (mm d(-1)) and tolerances. Predictions of the effects of important environmental factors such as temperature, aw and their interactions on lag times to germination, germination rates and mycelial growth are important in the development of hurdle technology approaches to predicting fungal spoilage in agricultural and food products.

Aims: To examine the effect of butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), trihydroxybutyrophenone and propylparaben (PP) (at concentrations of 1-20 mmol l(-1)) on growth of and fumonisin production by Argentinian strains of Fusarium verticillioides and F. proliferatum. Methods and Results: Studies on lag phases prior to growth, relative growth rates and fumonisin concentrations were carried out in vitro in relation to water activity (0.995-0.93 a(w)) and temperature (18 and 25 degrees C) on a maize meal agar. Overall, PP was the antioxidant which was most effective at inhibiting strains of both species. The lag phase prior to growth and growth rates were significantly decreased by PP and BHA at 10 and 20 mmol l(-1), regardless of the temperature or a(w) level tested. Total fumonisin production was higher at 0.98 a(w) and decreased by about 45-50% at 0.995 and 0.95 a(w). Overall, BHT only inhibited fumonisin production at 0.95 a(w) at 10 and 20 mmol l(-1), while BHA was effective at most a(w) levels tested at 10 and 20 mmol l(-1). Propylparaben completely inhibited fumonisin production by both F. verticillioides and F. proliferatum at > 1 mmol l(-1), regardless of the temperature or a(w) level. Small interstrain differences in the levels of inhibition by the antioxidants were observed for three F. verticillioides and four F. proliferatum strains at 0.995, 0.98 and 0.95 a(w). Propylparaben and BHA completely inhibited the growth of both species at the concentrations evaluated, regardless of the a(w) level. Conclusions: Two antioxidants show promise for the control of growth of and fumonisin production by these species over a wide range of environmental conditions. Significance and Impact of the Study: Potential exists for using such food-grade preservatives for prevention of mycotoxigenic fungi and their toxins entering the food chain.

Aim: To evaluate the effect of modifications of water activity (a(w) 0.996-0.92) of a molasses medium with different solutes (glycerol, glucose, NaCl, proline or sorbitol) on growth, intracellular water potentials (psi(c)) and endogenous accumulation of polyols/sugars in the biocontrol yeast Candida sake. Methods and Results: Modification of solute stress significantly influenced growth, psi(c) and accumulation of sugars (glucose/trehalose) and polyols (glycerol, erythritol, arabitol and mannitol) in the yeast cells. Regardless of the solute used to modify a(w), growth was always decreased as water stress increased. Candida sake cells grew better in glycerol- and proline-amended media, but were sensitive to NaCl. The psi(c) measured using psychrometry showed a significant effect of solutes, a(w) and time. Cells from the 0.96 a(w) NaCl treatment presented the lowest psi(c) value (-5.20 MPa) while cells from unmodified media (a(w) = 0.996) had the highest value (-0.30 MPa). In unmodified medium, glycerol was the predominant reserve accumulated. Glycerol and arabitol were the major compounds accumulated in media modified with glucose or NaCl. In proline media, the concentration of arabitol increased. In glycerol- and sorbitol-amended media, the concentration of glycerol rose. Some correlations were obtained between compatible solutes and psi(c). Conclusions and Significance: This study demonstrates that subtle changes in physiological parameters significantly affect the endogenous contents of C. sake cells. It may be possible to utilize such physiological information to develop biocontrol inocula with improved quality.

D. ALDRED, N. MAGAN and B.S. LANE.1999.This study investigated the effects of temperature, nutrient status and water activity (aW) on the production of squalestatin S1 by a Phoma sp. The fungus was grown on malt extract (MEA), wheat extract (WEA), oat extract (OEA) and oil seed rape extract (OSREA) agars at 15, 20 and 25 °C and 0·998, 0·995, 0·990, 0·980 and 0·960 aW levels. The growth rate and secondary metabolite formation were followed over a total of 30 d. The maximum growth rate was observed at 25 °C and 0·998–0·990 aW for all media types, which was significantly reduced (P= 0·05) for most media at 0·96 aw. The growth rate was greatest for WEA and OEA but the growth form was an effuse exploitative type compared with the dense assimilative type on the richer MEA. The lipid-based OSREA appeared to be a poor growth substrate for this fungus. In contrast to the growth rate data, squalestatin S1 production was maximal for all media types at slightly reduced aw in the range 0·990–0·980. There was greater production of the secondary metabolite under significant water stress (0·960 aW) compared with that with freely available water (0·998 aW). Maximum production was observed in WEA. Production began earlier in WEA and OEA compared with MEA. Squalestatin S1 production was not significantly affected by incubation temperature (P= 0·05). This study has shown that nutritionally depleted substrates may be usefully employed in the production of squalestatin S1 and perhaps also for other secondary metabolites.