Impact of moisture on in vitro germination of Metarhizium anisopliae and Beauveria bassiana and their activity on Triatoma infestans

The in vitro germination of 11 Metarhizium anisopliae and 11 Beauveria bassiana isolates originating from substrates collected in rural peridomestic areas in Central Brazil where triatomines are common was tested. Conidia completed germination up to 24 h after exposure to water activity of >0.99 a_w in all isolates tested. At lower 0.93 a_w germination was delayed but conidia of most isolates germinated at high rates (>98 %) within 216 h of incubation. Activities of 2 M. anisopliae and 2 B. bassiana isolates with different patterns of germination at 0.93 a_w were tested in Triatoma infestans third instar nymphs. There was no relationship between germination kinetics in vitro at 0.93 a_w and their activity in vivo at 98, 75 and 43 % relative humidity (rh). Isolates with accelerated germination at 0.93 a_w were not more virulent at 75 and 43 % rh compared with isolates with retarded or no germination. Highest mortalities were observed at 98 % rh, and they did not exceed 25 % after 25 d incubation at lower 75 and 43 % rh. Isolates that originated from a region with an extensive annual arid period showed no adaptation to lower humidity in their activity against T. infestans.     

The Influence of Water Activity and Temperature on Germination,Growth and Sporulation of <Emphasis Type="Italic">Stachybotrys chartarum</Emphasis> Strains

The objectives were to determine the influence of water activity (a_w, 0.997–0.92) and temperature (10–37°C) and their interactions on conidial germination, mycelial growth and sporulation of two strains of Stachybotrys chartarum in vitro on a potato dextrose medium. Studies were carried out by modifying the medium with glycerol and either spread plating with conidia to evaluate germination and germ tube extension or centrally inoculating treatment media for measuring mycelial growth rates and harvesting whole colonies for determining sporulation. Overall, germination of conidia was significantly influenced by a_w and temperature and was fastest at 0.997–0.98 a_w between 15 and 30°C with complete germination within 24 h. Germ tube extension was found to be most rapid at similar a_w levels and 25–30°C. Mycelial growth rates of both strains were optimal at 0.997 a_w between 25 and 30°C, with very little growth at 37°C. Sporulation was optimum at 30°C at 0.997 a_w. However, under drier conditions, this was optimum at 25°C. This shows that there are differences in the ranges of a_w x temperature for germination and growth and for sporulation. This may help in understanding the role of this fungal species in damp buildings and conditions under which immune-compromised patients may be at risk when exposed to such contaminants in the indoor air environment.     

In vitro studies of the effect of Environmental Conditions on the anthacnose pathogen of bananas, Colletotrichum musae

In vitro studies were undertaken to determine the effect of pH, temperature, water availability and carbon dioxide (CO₂) concentration on germination and growth of Colletotrichum musae, the causal pathogen of anthracnose of bananas. The optimum pH for germination and growth varied between 4·0 and 5·0 depending on temperature. At low pH (< 3·0) and 15°C, both germination and growth were significantly reduced, with a marked increase in the lag time, in days, prior to growth. C. musae germinated and grew over a wide range of water activities (a_w; 0·995−0·94 and 0·995−0·92, respectively) at 20, 25 and 30°C. In all cases where germination occurred appresoria were subsequently produced. Optimum growth occurred at 30°C and 0·995 a_w, although this changed to 0·98 a_w at 35°C. Increasing CO₂ concentration to 15% or reducing oxygen concentration to 1% resulted in a significant (P < 0·05) reduction in growth, but did not inhibit growth completely.     

Effect of temperature and water activity on spore germination and mycelial growth of three fungal biocontrol agents against water hyacinth (Eichhornia crassipes)

Aims: To determine the effect of water activity (a_w = 0·880–0·960) and temperature (15–35°C) on the percentage of viable conidia and mycelial growth of three biocontrol agents effective against water hyacinth in Mali: Alternaria sp. isolate Mlb684, Fusarium sacchari isolate Mln799 and Cadophora malorum isolate Mln715. Methods and Results: The fungi were grown in vitro on plates containing potato dextrose agar medium at different a_w values (glycerol being added to adjust the a_w). The percentage of viable conidia and radial growth rate decreased with decreasing water activity. Statistical analysis showed a significant effect of a_w, temperature and the a_w × temperature interaction on mycelial growth (P < 0·0001). Water activity emerged as the factor exerting the greatest influence. Differences were observed between the fungi tested, the C. malorum appearing more tolerant to low a_w and the F. sacchari more tolerant to high temperature (35°C). Growth models predicting the combined effect of a_w and temperature were developed and response surfaces generated, showing fairly good agreement with the experimental values. Conclusions: Our results confirm the previous finding that a_w has a greater influence than temperature on fungal growth. Under most conditions, variation of environmental factors has a detrimental influence on the percentage of viable conidia and mycelial growth rate of fungal isolates. Significance and Impact of the Study: The developed models may contribute to predicting the best environmental conditions for use of these fungi as effective biocontrol agents against water hyacinth.     

Studies on some fungi isolated from the rhizosphere of tomato plants and the consequent prospect for the control of Verticillium wilt

Summary Biological control of Verticillium wilt disease with antagonistic micro-organisms was studied. Antagonism of some fungi, isolated from tomato rhizosphere, toVerticillium albo-atrum R & B. was observedin vitro. A clearly defined zone, in which the growth of the pathogen was inhibited, was observed withPenicillium spp. (includingPenicillium chrysogenum Thom) andFusarium culmorum (S.G. Sm) Sacc., whileTrichoderma viride pers. ex Fries,Gliocladium spp. andPenicillium vermiculatum Dangeard, suppressed the growth ofV. albo-atrum by penetrating, and overgrowing it. OnlyT. viride andP. vermiculatum culture filtrate added to the Dox's agar, reduced the radial growth ofV. alboatrum. Root-dip application of culture filtrates ofT. viride andP. chrysogenum was found to be most effective in controlling the disease, followed by other species ofPenicillium andGliocladium spp. WhileFusarium culmorum provided no control. Improvement of plant height and vigour with a better yield due to culture filtrate treatment occurred. Root-dip application of antagonistic fungal propagules (T. viride, P. chrysogenum) to tomato seedlings was also very effective in controlling wilt in tomato plants grown inV. albo-atrum infested soil. Dedicated to the memory of the late Prof. Ivor Isaac with whom I had the pleasure of working     

Effects of temperature and water activity on <Emphasis Type="Italic">Lecanicillium</Emphasis> spp. conidia germination and growth,and mycosis of <Emphasis Type="Italic">Pissodes strobi</Emphasis>

Selecting entomopathogenic fungal isolates for use as biocontrol agents requires an assessment of their growth and virulence characteristics as affected by environmental conditions. Here we demonstrate a wide temperature and moisture range for colony growth, effective conidial germination and virulence against Pissodes strobi Peck (white pine weevil) of several isolates of Lecanicillium Gams and Zare, an entomopathogenic fungus distributed worldwide and indigenous to forests on Vancouver Island, British Columbia, Canada. In order to examine the potential Lecanicillium as a biological control agent, the pathogenicity of isolates collected from different geographical locations on P. strobi cadavers was assessed, and colony growth at different temperatures was evaluated. Colony growth was evident between 5 and 30°C, with optimal growth occurring at 25°C. Various combinations of water activity (0.55, 0.76, 0.85 and 0.99 a _w) and temperature (10, 15, 20, and 25°C) were also used to evaluate environmental impacts on conidial germination and cumulative mycosis of adult P. strobi. Certain Lecanicillium isolates displayed xerophilic (0.85 a _w) or psychrophilic (10°C) growth optima. Ultimately, identifying the abiotic limits of this entomopathogenic fungus will be used to determine which isolates have potential for future in situ biocontrol trials.     

Temperature and moisture requirements for conidial germination of an isolate of Beauveria bassiana,pathogenic to Rhodnius prolixus

The effects of temperature, relative humidity and water activity on germination of conidia of an isolate of Beauveria bassiana (Bals.) Vuill. pathogenic to the triatomine vector of Chagas' disease, Rhodnius prolixus Stål., were investigated in vitro. Germination occurred at temperatures between 15 °C and 35 °C under saturated atmosphere and the optima ranged from 25 °C to 3O °C. At the extreme temperatures tested (15 °C and 35 °C) the germination process was delayed, but germination rates reached more than 95%. Germination of B. bassiana conidia was strongly affected by moisture conditions. The availability of water, in both atmospheric and liquid conditions, caused changes in germination times as well as in germination rates. For example, at 25 °C + O.5 °C, germination took place within 20 h at 95.5% RH, whereas it needed 72 h of incubation at 90% RH. Germination times increased as the water activity declined from 0.96 a_w to 0.92 a_w. Below 0.92 a_w, o germination was observed after a 72 h incubation time.     

Occurrence and moisture requirements of microbial growth in building materials

Microbial growth was studied in six damp buildings. Mesophilic fungi, especially Penicillium spp., yeasts, and species of Cladosporium and Aspergillus, occurred most abundantly on building constructions. Thermophilic fungi and mesophilic actinomycetes were occasionally found. A toxigenic fungus, Stachybotrys sp., was also detected on cellulose-based materials. In a cytotoxicity test, 23% of samples were positive. Spore counts varied considerably on materials, but no correlation between counts and the substrate or its water activity (a_w) was observed. In experiments a rapid increase in CO₂ production and spore propagule count was observed in all materials incubated at a relative humidity (RH) (RH=0·01^*water activity) of 96–98°. Some differences were noted between materials in CO₂ evolved, but not in propagule counts.     

In vitro and in vivo antagonism of biocontrol agents against Colletotrichum lindemuthianum causing bean anthracnose

Bean anthracnose caused by Colletotrichum lindemuthianum is a serious seed borne disease. For devising an effective management strategy, the efficacy of different bioagents, viz. Trichoderma viride, Trichoderma harzianum, Trichoderma hamatum and Gliocladium virens conducted under in vitro and in vivo conditions revealed maximum inhibition of mycelial growth in dual culture (59.48%) and inverted plate (55.98%) with T. viride. All the bioagents overgrew the pathogen and the principal mechanism of mycoparisitism observed was coiling, brusting and disintegration of pathogen hyphae. Culture filtrate from T. viride was found best as it completely inhibited radial growth at 25 and 50% concentration and reduced the spore germination of test fungus significantly. However, lower concentrations of culture filtrate from all bioagents showed little effect on spore germination. Seed application of bioagents was found better as compared to soil application. A maximum increase in seed germination and inhibition of seed borne infection was observed with T. viride followed by T. harzianum under pot culture conditions. T. viride has the maximum potentiality to suppress the spore germination, mycelial growth, seed borne infection of C. lindemuthianum and increased seed germination when compared with the other biocontrol agents.     

Impact of some environmental factors on growth and production of ochratoxin A of/by <Emphasis Type="Italic">Aspergillus tubingensis,A. niger</Emphasis>, and <Emphasis Type="Italic">A. carbonarius</Emphasis> isolated from moroccan grapes

The effects of temperature, water activity (a_w), incubation time, and their combinations on radial growth and ochratoxin A (OTA) production of/by eight Aspergillus niger aggregate strains (six A. tubingensis and two A. niger) and four A. carbonarius isolated from Moroccan grapes were studied. Optimal conditions for the growth of most studied strains were shown to be at 25°C and 0.95 a_w. No growth was observed at 10°C regardless of the water activity and isolates. The optimal temperature for OTA production was in the range of 25°C∼30°C for A. carbonarius and 30°C∼37°C for A. niger aggregate. The optimal a_w for toxin production was 0.95∼0.99 for A. carbonarius and 0.90∼0.95 for A. niger aggregate. Mean OTA concentration produced by all the isolates of A. niger aggregate tested at all sampling times shows that maximum amount of OTA (0.24 μg/g) was produced at 37°C and 0.90 a_w. However, for A. carbonarius, mean maximum amounts of OTA (0.22 μg/g) were observed at 25°C and 0.99 a_w. Analysis of variance showed that the effects of all single factors (a_w, isolate, temperature and incubation time) and their interactions on growth and OTA production were highly significant.     

Modelling the effect of temperature, water activity and pH on the growth of Serpula lacrymans

Aims: To predict the risk factors for building infestation by Serpula lacrymans, which is one of the most destructive fungi causing timber decay in buildings. Methods and Results: The growth rate was assessed on malt extract agar media at temperatures between 1·5 and 45°C, at water activity (a_w) over the range of 0·800–0·993 and at pH ranges from 1·5 to 11·0. The radial growth rate (μ) and the lag phase (λ) were estimated from the radial growth kinetics via the plots radius vs time. These parameters were then modelled as a function of the environmental factors tested. Models derived from the cardinal model (CM) were used to fit the experimental data and allowed an estimation of the optimal and limit values for fungal growth. Optimal growth rate occurred at 20°C, at high a_w level (0·993) and at a pH range between 4·0 and 6·0. The strain effect on the temperature parameters was further evaluated using 14 strains of S. lacrymans. The robustness of the temperature model was validated on data sets measured in two different wood‐based media (Quercus robur L. and Picea abies). Conclusions: The two‐step procedure of exponential model with latency followed by the CM with inflection gives reliable predictions for the growth conditions of a filamentous fungus in our study. The procedure was validated for the study of abiotic factors on the growth rate of S. lacrymans. Significance and Impact of the Study: This work describes the usefulness of evaluating the effect of physico‐chemical factors on fungal growth in predictive building mycology. Consequently, the developed mathematical models for predicting fungal growth on a macroscopic scale can be used as a tool for risk assessment of timber decay in buildings.     

Impact of a Streptomyces (AS1) strain and its metabolites on control of Aspergillus flavus and aflatoxin B1 contamination in vitro and in stored peanuts

Six actinomycetes were isolated from peanuts in Egypt. Of these, a Streptomyces strain (AS1) was found in in vitro assays to inhibit directly or via secondary metabolites both germination and growth of Aspergillus flavus. Tests of the AS1 cells for direct control of A. flavus populations or aflatoxin B₁ (AFB₁) production on stored peanuts was unsuccessful over 14-day storage periods. However, crude extracts of AS1 metabolites at 50 and 100 ppm completely inhibited spore germination of conidia of A. flavus in vitro over 48 h. Comparison of solvents for extracting the metabolites showed that the ethyl acetate extract was most effective. This gave greater than 85% inhibition of mycelial growth at these concentrations at different water availabilities (water activity; a _w; 0.95, 0.92, and 0.89) and 25°C. Doses of 50, 200, and 500 ppm of AS1 metabolites significantly inhibited populations of A. flavus on stored peanuts at two water stress levels (0.90, 0.93 a _w) at 25°C over 14-day storage periods. The amounts of AFB₁ produced by A. flavus on peanuts stored at 0.90 a _w were significantly decreased by AS1 metabolites for only 7 days. However, at 0.93 a _w doses of 200 and 500 ppm significantly controlled AFB₁ accumulation in peanuts for 14 days.     

In vitro Antifungal Activity of Essential oils and Major Components against Fungi Plant Pathogens

The indiscriminate use of synthetic fungicides has caused several problems to the environment, which place human and animal health at risk. Due to this fact, the search for natural alternatives to control phytopathogenic fungi growth has increased. This study aims to evaluate the antifungal activity of two essential oils (EOs) and three major components of EOs on the radial growth and spore germination of Fusarium verticillioides and Alternaria tenuissima. Minimum and half‐maximal inhibitory concentrations (CMI and CI₅₀) at 96 h for each treatment were calculated. Lemongrass EO and citral caused the highest inhibition for A. tenuissima (CMI of 1000 μl/l and CI₅₀ of 10 μl/l). For F. verticillioides, the most effective component was geraniol (CMI and CI₅₀ of 1000 and 250 μl/l, respectively). Spore germination rate was delayed by the EOs and major components tested. The use of EOs was effective to control these two fungal species in their different grow stages.     

Modelling the effect of water activity and temperature on growth rate and aflatoxin production by two isolates of Aspergillus flavus on paddy

Aims: This study was conducted to characterize the growth of and aflatoxin production by Aspergillus flavus on paddy and to develop kinetic models describing the growth rate as a function of water activity (a_w) and temperature. Methods and Results: The growth of A. flavus on paddy and aflatoxin production were studied following a full factorial design with seven a_w levels within the range of 0·82–0·99 and seven temperatures between 10 and 43°C. The growth of the fungi, expressed as colony diameter (mm), was measured daily, and the aflatoxins were analysed using HPLC with a fluorescence detector. The maximum colony growth rates of both isolates were estimated by fitting the primary model of Baranyi to growth data. Three potentially suitable secondary models, Rosso, polynomial and Davey, were assessed for their ability to describe the radial growth rate as a function of temperature and a_w. Both strains failed to grow at the marginal temperatures (10 and 43°C), regardless of the a_w studied, and at the a_w level of 0·82, regardless of temperature. Despite that the predictions of all studied models showed good agreement with the observed growth rates, Davey model proved to be the best predictor of the experimental data. The cardinal parameters as estimated by Rosso model were comparable to those reported in previous studies. Toxins were detected in the range of 0·86–0·99 a_w with optimal a_w of 0·98 and optimal temperature in the range of 25–30°C. Conclusions: The influences of a_w and temperature on the growth of A. flavus and aflatoxin production were successfully characterized, and the models developed were found to be capable of providing good, related estimates of the growth rates. Significance and Impact of the Study: The results of this study could be effectively implemented in minimizing the risk of aflatoxin contamination of the paddy at postharvest.     

Influence of water activity on the enzyme biosynthesis and enzyme activities produced by Trichoderma viride TS in solid-state fermentation

Influence of water activity (a_w) on biosynthesis of polygalacturonase, d-xylanase and β-glucosidase in solid culture system of Trichoderma viride TS was studied. It was found that the production of enzymes was strongly affected by water activity of substrate and nature of a_w depressor used. The polygalacturonase and d-xylanase production were maximized at a_w = 0.995 whereas β-glucosidase formation was favored at a_w = 0.96–0.98. The influence of water activity on catalytic effect of enzymes using sodium chloride, glycerol and sorbitol as a_w depressor was also investigated. It was observed that sorbitol improved the thermal stability of polygalacturonase and d-xylanase.     

Comparison of the effects of temperature and water activity on growth rate of food spoilage moulds

The influence of temperature (T) and water activity (a _w) on the growth rate (μ) of seven moulds (Alternaria alternata, Aspergillus flavus, Cladosporium cladosporioides, Mucor racemosus, Penicillium chrysogenum, Rhizopus oryzae and Trichoderma harzianum) was assessed in suboptimal conditions. Firstly, the dependence of fungal growth on temperature, at a _w 0.99, was modelled through an approach described previously for bacteria. A dimensionless growth rate variable: μ _dimα=μ/μ _optα depended on the following normalised temperature: T _dim=(T−T _min)/(T _opt− T _min) according to a power function: μ _dimα=[T _dim] ^α , where α was an exponent to be estimated. Secondly, the same approach was used to describe the influence of a _w on fungal growth, at the respective optimum temperatures for each mould. Similarly, μ _dimβ=μ/μ _optβ depended on the following normalised water activity: a _wdim=(a _w−a _wmin)/(a _wopt−a _wmin) according to a power function: μ _dimβ=[a _wdim]^β. Results show: (i) for each mould, the α-value is significantly less than the β-value, confirming that water activity has a greater influence than temperature on fungal development; (ii) the α-values and the β-values depend on the mould; (iii) the α-value is less than 1 for the mesophilic mould A. flavus, whereas the other moulds are characterised by higher α-values ranging from 1.10 to 1.54; (iv) the mesophilic A. flavus exhibits a low β-value, 1.50, compared to the hydrophilic T. harzianum, β=2.44, while β-values are within the range (1.71–2.37) for the other moulds. Journal of Industrial Microbiology & Biotechnology (2002) 28, 311–315 DOI: 10.1038/sj/jim/7000248 Received 27 June 2001/ Accepted in revised form 04 February 2002     

Influence of age of mycelium and water activity of the medium on aroma production by Trichoderma viride grown on solid substrate

The production of 2-heptanone (cheese aroma) by Trichoderma viride TS cultivated on agar media was evaluated using headspace gas chromatographic analysis. The radial growth rate of the filamentous fungi increased with high water activity values, but the mycelial density was higher for lower water activity. Maximum aroma production of a culture was obtained at a_w = 0.96. An apparatus intended to measure the aroma production of different areas of a mycelial colony was set up. The study of the aroma production of these areas showed that the production values were greatly different and were evolving with time. It was shown that the mycelium aroma production was maximum when mycelia were about 3.5 to 6.5 d old.     

不同浓度壳聚糖对炭黑曲霉和硫色镰刀菌生长和发育的影响

【背景】壳聚糖是广泛存在于甲壳动物的一种多糖，具有广谱的抗真菌活性，但壳聚糖是否影响炭黑曲霉(Aspergillus carbonarius)和硫色镰刀菌(Fusarium sulphureum)生长和发育尚未见报道。【目的】明确不同浓度壳聚糖对A. carbonarius和F. sulphureum生长和发育的影响。【方法】通过在PDA培养基中添加不同浓度壳聚糖，测定两种真菌的菌落直径、生物量和菌丝干重，观察产孢量、孢子萌发和芽管长度，比较抑菌的差异。【结果】壳聚糖处理可显著改变两种真菌的菌落形态，处理浓度越高菌落皱缩和变形越明显；壳聚糖还可以有效抑制两种真菌的菌落生长、菌丝干重和菌丝生物量，抑制效果呈明显的浓度依赖，对F. sulphureum的抑制效果更好。壳聚糖可抑制两种真菌的产孢量、孢子萌发和芽管伸长，处理浓度越高抑制效果越好，对F. sulphureum的抑制效果更为明显。壳聚糖对A. carbonarius和F. sulphureum的EC₅₀值分别为0.12 mg/mL和0.075 mg/mL。【结论】壳聚糖可有效抑制A. carbonarius和F. sulphureum的生长发育，抑制效果呈浓度依赖，F. sulphureum对壳聚糖更为敏感。     

Effect of water activity adjusted with different solutes on growth and lactic acid production byLactobacillus helveticus

Effect of water activity (a _w) adjusted with NaCl or glycerol on the growth and metabolism ofLactobacillus helveticus var.pragensis at 40°C was demonstrated by growth curves (generation time) and changes in the degree of acidity of a milk medium expressed as lactic acid content. NaCl-regulateda _w of 0.970 inhibited growth and acid production completely. The same, glycerol-regulateda _w of 0.970 increased the generation time only from 33 min (a _w=0.994) to 67 min which was less than the generation time at the highera _w of 0.982 adjusted with NaCl (103 min). Glycerol-regulateda _w of 0.970 did not decrease the acid production (2.6% lactic acid). Ata _w of 0.951, the acid production was decreased by 39% compared with the values found in milk media with the original, unadjusteda _w of 0.994, after the same time of incubation. Media witha _w adjusted to the same values with NaCl or glycerol do not influence the growth and acid production ofL. helveticus in the same way. In contrast to glycerol, NaCl has a strong effect.     

Studies on the ability of mycotoxins to diffuse in bread

Summary The moulds Aspergillus parasiticus (aflatoxins B₁, B₂, G₁, G₂, and M₁), A. ochraceus (ochratoxin A) and Penicillium chrysogenum (citrinin) were grown on whole wheat bread either in the presence or absence of oxygen. In the presence of oxygen, both A. parasiticus and A. ochraceus developed dense colonies and formed considerable amounts of mycotoxins whereas Penicillium chrysogenum only grew and produced citrinin on the surface of the bread. In the absence of oxygen fungal growth did not occur and most of the toxins were undetectable even in regions of bread immediately adjacent to the moulds although a very slight diffusion of the aflatoxins B₁ and G₁ through 1 cm was observed. It is concluded that diffusion of the tested mycotoxins from hyphae into bread is not a problem for food safety.