Enhancing biological control of postharvest green mold in lemons: Synergistic efficacy of native yeasts with diverse mechanisms of action.

Argentina is among the most important lemon fruit producers in the world. Penicillium digitatum is the primary lemon fungal phytopathogen, causing green mold during the postharvest. Several alternatives to the use of synthetic fungicides have been developed, being the use of biocontrol yeasts one of the most promising. Although many of the reports are based on the use of a single yeast species, it has been shown that the combination of agents with different mechanisms of action can increase control efficiency through synergistic effects. The combined use of native yeasts with different mechanisms of action had not been studied as a biological control strategy in lemons. In this work, the mechanisms of action of native yeasts (Clavispora lusitaniae AgL21, Clavispora lusitaniae AgL2 and Clavispora lusitaniae AcL2) with biocontrol activity against P. digitatum were evaluated. Isolate AgL21 was selected for its ability to form biofilm, colonize lemon wounds, and inhibit fungal spore germination. The compatibility of C. lusitaniae AgL21 with two killer yeasts of the species Kazachstania exigua (AcL4 and AcL8) was evaluated. In vivo assays were then carried out with the yeasts applied individually or mixed in equal cell concentrations. AgL21 alone was able to control green mold with 87.5% efficiency, while individual killer yeasts were significantly less efficient (43.3% and 38.3%, respectively). Inhibitory effects were increased when C. lusitaniae AgL21 and K. exigua strains were jointly applied. The most efficient treatment was the combination of AgL21 and AcL4, reaching 100% efficiency in wound protection. The combination of AgL21 with AcL8 was as well promising, with an efficiency of 97.5%. The combined application of native yeasts showed a synergistic effect considering that the multiple mechanisms of action involved could hinder the development of green mold in lemon more efficiently than using single yeasts. Therefore, this work demonstrates that the integration of native yeasts with diverse modes of action can provide new insights to formulate effective microbial consortia. This could lead to the development of tailor-made biofungicides, allowing control of postharvest fungal diseases in lemons while remaining competitive with traditionally used synthetic chemicals.

Selection of Antarctic yeasts as gray mold biocontrol agents in strawberry.

The postharvest disease popularly known as gray mold is considered one of the most limiting factors strawberry fruit production. The most effective way to control this disease is still the use of chemical fungicides. However, other alternative sources of control are being explored. Among these, psychrophilic yeasts adapted to extreme conditions, such as those found in the Antarctic region, may have great potential for use as biocontrol agents. Thus, the present study aimed to select psychrotolerant yeasts obtained from Antarctic region and to evaluate their potential for biocontrol under gray mold, caused by Botrytis cinerea in strawberries stored at low temperature. For this, 20 potential antagonist yeasts were evaluated in vitro (thermotolerance and enzymatic) assays. Debaryomyces hansenii, Rhodotorula mucilaginosa and Dioszegia hungarica were selected for growing in strawberry juice. However, only D. hansenii was selected for in vivo studies and showed a reduction in the incidence of gray mold by 82% for the tests performed on injury and 86% for the tests on non-injured fruits treated by immersion bath. Thus, demonstrating that the selection of this cold-adapted Antarctic yeast can be a promising strategy as a biocontrol agent used to curb the development of gray mold in strawberry fruits.

Microalgae, Seaweeds and Aquatic Bacteria, Archaea, and Yeasts: Sources of Carotenoids with Potential Antioxidant and Anti-Inflammatory Health-Promoting Actions in the Sustainability Era.

Carotenoids are a large group of health-promoting compounds used in many industrial sectors, such as foods, feeds, pharmaceuticals, cosmetics, nutraceuticals, and colorants. Considering the global population growth and environmental challenges, it is essential to find new sustainable sources of carotenoids beyond those obtained from agriculture. This review focuses on the potential use of marine archaea, bacteria, algae, and yeast as biological factories of carotenoids. A wide variety of carotenoids, including novel ones, were identified in these organisms. The role of carotenoids in marine organisms and their potential health-promoting actions have also been discussed. Marine organisms have a great capacity to synthesize a wide variety of carotenoids, which can be obtained in a renewable manner without depleting natural resources. Thus, it is concluded that they represent a key sustainable source of carotenoids that could help Europe achieve its Green Deal and Recovery Plan. Additionally, the lack of standards, clinical studies, and toxicity analysis reduces the use of marine organisms as sources of traditional and novel carotenoids. Therefore, further research on the processing of marine organisms, the biosynthetic pathways, extraction procedures, and examination of their content is needed to increase carotenoid productivity, document their safety, and decrease costs for their industrial implementation.

Indigenous Yeasts for the Biocontrol of Botrytis cinerea on Table Grapes in Chile.

One hundred twenty-five yeast strains isolated from table grapes and apples were evaluated for the control Botrytis cinerea of in vitro and in vivo. Ten strains were selected for their ability to inhibit mycelial growth of B. cinerea in vitro. In the in vivo assays, these yeasts were tested at 20 °C on 'Thompson Seedless' berries for 7 days; only three were selected (m11, me99 and ca80) because they significantly reduced the incidence of gray mold. These three yeast strains were then evaluated at different concentrations (1 × 107, 1 × 108 and 1 × 109 cells mL-1) on 'Thompson Seedless' grape berries at 20 °C. The strains m11, me99 and ca80 reduced the incidence of B. cinerea to 11.9, 26.1 and 32.1%, respectively, when the berries were submerged in a yeast suspension at a concentration of 1 × 109 cells mL-1 24 h before inoculation with B. cinerea. The most favorable pH for antifungal activity was 4.6 in the three isolates. The three yeast strains secreted the hydrolytic enzymes chitinase and ß-1-glucanase, and two strains (me99 and ca80) produced siderophores. The three yeast strains exhibited low oxidative stress tolerance and only strain m11 had the ability to produce biofilms. The strains were identified using 5.8S-ITS rDNA PCR-RFLP and correspond to the Meyerozyma guilliermondii (m11) and Aureobasidium pullulans (me99 and ca80) species.

Characterization of kefir yeasts with antifungal capacity against Aspergillus species / Caracterización de levaduras de kéfir con capacidad antifúngica frente a especies de Aspergillus

Kefir is a fermented probiotic drink obtained by placing kefir granules in a suitable substrate. The kefir granules are a consortium of bacteria and yeasts embedded in a exopolysaccharide matrix. The aim of this research was the isolation and identification of yeasts from kefir of different origin, the evaluation of their antifungal capacity against Aspergillus spp., and the characterization of virulence related traits. Using RFLP of ITS1/ITS4 region, D1/D2 region sequencing, and RAPD techniques, 20 kefir isolates were identified as Geotrichum candidum, Pichia kudriavzevii, Pichia membranifaciens, Saccharomyces cerevisiae, and Candida ethanolica. Their antifungal capacity was evaluated by their conidia germination reduction, which allowed the selection of eight isolates with high to moderate conidia germination reduction against Aspergillus flavus and Aspergillus parasiticus. Furthermore, these selected isolates showed growth inhibition on contact in the dual culture assay for both Aspergillus species and 3 of them—belonging to S. cerevisiae and P. kudriavzevii species—generated volatile organic compounds which significantly affected the growth of both fungi. For the evaluation of virulence-related traits, growth at high temperatures, enzymatic activities, and the adhesion to Caco-2 cells were analyzed. The isolates did not present more than one positive virulence-related trait simultaneously. In particular, it is important to highlight that the adhesion capacity to the model of intestinal barrier was extremely low for all of them. According to the results obtained, further studies would be of interest for the possible use of these promising yeasts as biocontrol agents against fungi in food.(AU)

Characterization of kefir yeasts with antifungal capacity against Aspergillus species.

Kefir is a fermented probiotic drink obtained by placing kefir granules in a suitable substrate. The kefir granules are a consortium of bacteria and yeasts embedded in a exopolysaccharide matrix. The aim of this research was the isolation and identification of yeasts from kefir of different origin, the evaluation of their antifungal capacity against Aspergillus spp., and the characterization of virulence related traits. Using RFLP of ITS1/ITS4 region, D1/D2 region sequencing, and RAPD techniques, 20 kefir isolates were identified as Geotrichum candidum, Pichia kudriavzevii, Pichia membranifaciens, Saccharomyces cerevisiae, and Candida ethanolica. Their antifungal capacity was evaluated by their conidia germination reduction, which allowed the selection of eight isolates with high to moderate conidia germination reduction against Aspergillus flavus and Aspergillus parasiticus. Furthermore, these selected isolates showed growth inhibition on contact in the dual culture assay for both Aspergillus species and 3 of them-belonging to S. cerevisiae and P. kudriavzevii species-generated volatile organic compounds which significantly affected the growth of both fungi. For the evaluation of virulence-related traits, growth at high temperatures, enzymatic activities, and the adhesion to Caco-2 cells were analyzed. The isolates did not present more than one positive virulence-related trait simultaneously. In particular, it is important to highlight that the adhesion capacity to the model of intestinal barrier was extremely low for all of them. According to the results obtained, further studies would be of interest for the possible use of these promising yeasts as biocontrol agents against fungi in food.

Production of microbial oils by the oleaginous yeast Rhodotorula graminis S1/2R in a medium based on agro-industrial by-products.

Biodiesel generated by transesterification of triglycerides from renewable sources is a clean form of energy that is currently used in many countries in blends with petrodiesel. It is mainly produced from food-grade vegetable oils obtained from oleaginous crops. High prices of these oils have made the sustainability of biodiesel production questionable. The use of nonedible feedstocks, such as intracellular triglycerides accumulated by oleaginous yeasts, appears as a feasible alternative. However, it has been demonstrated that an economically sustainable production of yeast oil could only be possible if low-cost media based on industrial subproducts, or wastes are used. In this work, we propose intracellular lipids production by a previously selected oleaginous yeast strain in a medium composed only by sugar cane vinasse and crude glycerol. Different culture strategies were studied. The highest biomass and lipid yields were obtained when the yeast R. graminis S1/2R was cultivated in batch without control of dissolved oxygen. The fatty acid methyl esters obtained under these conditions met the specification of international biodiesel standards.

Fusarium graminearum Species Complex: A Bibliographic Analysis and Web-Accessible Database for Global Mapping of Species and Trichothecene Toxin Chemotypes.

Fusarium graminearum is ranked among the five most destructive fungal pathogens that affect agroecosystems. It causes floral diseases in small grain cereals including wheat, barley, and oats, as well as maize and rice. We conducted a systematic review of peer-reviewed studies reporting species within the F. graminearum species complex (FGSC) and created two main data tables. The first contained summarized data from the articles including bibliographic, geographic, methodological (ID methods), host of origin and species, while the second data table contains information about the described strains such as publication, isolate code(s), host/substrate, year of isolation, geographical coordinates, species and trichothecene genotype. Analyses of the bibliographic data obtained from 123 publications from 2000 to 2021 by 498 unique authors and published in 40 journals are summarized. We describe the frequency of species and chemotypes for 16,274 strains for which geographical information was available, either provided as raw data or extracted from the publications, and sampled across six continents and 32 countries. The database and interactive interface are publicly available, allowing for searches, summarization, and mapping of strains according to several criteria including article, country, host, species and trichothecene genotype. The database will be updated as new articles are published and should be useful for guiding future surveys and exploring factors associated with species distribution such as climate and land use. Authors are encouraged to submit data at the strain level to the database, which is accessible at https://fgsc.netlify.app.

Fungal diversity in the coastal waters of King George Island (maritime Antarctica).

Fungi have been reported as common inhabitants of the maritime waters in Antarctica by studies based on culture-dependent methods. More recently, results obtained using DNA sequencing technologies, revealed that fungal diversity worldwide has been underestimated by culture methods. The present study provides the first characterization of fungal communities in the coastal waters of King George Island (maritime Antarctica) using both culture-dependent and high-throughput sequencing (HTS) methods. HTS demostrated a higher level of fungal diversity than the obtained by culture methods. A high prevalence of basidiomycetous yeasts and ascomycetous filamentous fungi was confirmed by both methods, however, Chythriomycota, Rozellomycota, lichenized fungi and Malassezia spp. were detected only by HTS. Correspondingly, members of some genera, such as Metschnikowia, were only found by culture-dependent methods. Our results confirm that culturing and HTS, should be seen as complementary approaches that enable one to obtain a more comprehensive picture of the composition of microbial communities.

Global analysis of the apple fruit microbiome: are all apples the same?

We present the first worldwide study on the apple (Malus × domestica) fruit microbiome that examines questions regarding the composition and the assembly of microbial communities on and in apple fruit. Results revealed that the composition and structure of the fungal and bacterial communities associated with apple fruit vary and are highly dependent on geographical location. The study also confirmed that the spatial variation in the fungal and bacterial composition of different fruit tissues exists at a global level. Fungal diversity varied significantly in fruit harvested in different geographical locations and suggests a potential link between location and the type and rate of postharvest diseases that develop in each country. The global core microbiome of apple fruit was represented by several beneficial microbial taxa and accounted for a large fraction of the fruit microbial community. The study provides foundational information about the apple fruit microbiome that can be utilized for the development of novel approaches for the management of fruit quality and safety, as well as for reducing losses due to the establishment and proliferation of postharvest pathogens. It also lays the groundwork for studying the complex microbial interactions that occur on apple fruit surfaces.

Debaryomyces hansenii F39A as biosorbent for textile dye removal.

Many industries generate a considerable amount of wastewater containing toxic and recalcitrant dyes. The main objective of this research was to examine the biosorption capacity of Reactive Blue 19 and Reactive Red 141 by the Antarctic yeast Debaryomyces hansenii F39A biomass. Some variables, including pH, dye concentration, amount of adsorbent and contact time, were studied. The equilibrium sorption capacity of the biomass increased with increasing initial dye concentration up to 350mg/l. Experimental isotherms fit the Langmuir model and the maximum uptake capacity (qmax) for the selected dyes was in the range of 0.0676-0.169mmol/g biomass. At an initial dye concentration of 100mg/l, 2g/l biomass loading and 20±1°C, D. hansenii F39A adsorbed around 90% of Reactive Red 141 and 50% of Reactive Blue 19 at pH 6.0. When biomass loading was increased (6g/l), the uptake reached up to 90% for Reactive Blue 19. The dye uptake process followed a pseudo-second-order kinetics for each dye system. As seen throughout this research study, D. hansenii has the potential to efficiently and effectively remove dyes in a biosorption process and may be an alternative to other costly materials.

Yeasts and Bacterial Consortia from Kefir Grains Are Effective Biocontrol Agents of Postharvest Diseases of Fruits.

Fungal pathogens in fruits and vegetables cause significant losses during handling, transportation, and storage. Biological control with microbial antagonists replacing the use of chemical fungicides is a major approach in postharvest disease control, and several products based on single antagonists have been developed but have limitations related to reduced and inconsistent performance under commercial conditions. One possible approach to enhance the biocontrol efficacy is to broaden the spectrum of the antagonistic action by employing compatible microbial consortia. Here, we explore commercial kefir grains, a natural probiotic microbial consortium, by culture-dependent and metagenomic approaches and observed a rich diversity of co-existing yeasts and bacterial population. We report effective inhibition of the postharvest pathogen Penicillium expansum on apple by using the grains in its fresh commercial and milk-activated forms. We observed few candidate bacteria and yeasts from the kefir grains that grew together over successive enrichment cycles, and these mixed fermentation cultures showed enhanced biocontrol activities as compared to the fresh commercial or milk-activated grains. We also report several individual species of bacteria and yeasts with biocontrol activities against Penicillium rots on apple and grapefruit. These species with antagonistic properties could be further exploited to develop a synthetic consortium to achieve enhanced antagonistic effects against a wide range of postharvest pathogens.

Biocontrol of Aspergillus flavus in Ensiled Sorghum by Water Kefir Microorganisms.

The capacity of microorganisms from water kefir (WK) to control Aspergillus flavus growth during the aerobic phase of ensiled sorghum grains was determined. Sorghum inoculated with A. flavus was treated with filter-sterilized and non-sterilized water kefir, ensiled, and incubated 7 days at 25 °C. A. flavus growth was quantified by qPCR after incubation. Mold growth was inhibited in the presence of water kefir while no inhibition was observed when filter-sterilized water kefir was applied, demonstrating the relevant role of the microorganisms in the kefir water in the biocontrol process. Fungal and bacterial diversity in treated sorghum mini-silos was analyzed by high-throughput sequencing. Firmicutes was the predominant bacterial phyla and Lactobacillus represented the most abundant genus, while Ascomycota was the predominant fungal phyla with Saccharomyces and Pichia as the major genera. Bacterial and yeast counts before and after incubation indicated that the microbial community obtained from WK was able to grow in the sorghum mini-silos in the presence of A. flavus. Results of the present work indicate that the use of a mixed inoculum of microorganisms present in WK may represent an alternative management practice to avoid the growth of A. flavus in ensiled sorghum grains and the concomitant contamination with aflatoxins.

Effect of moisture on wheat grains lipid patterns and infection with Fusarium graminearum.

Suitable conditions of temperature and humidity are required to maintain wheat grains quality, but during processing and storage, the grains can be exposed to adverse environmental conditions and presence of infectious fungi. Fusarium graminearum, the main causal agent of Fusarium head blight on wheat, affects crop yields and grain quality by alteration of their biochemical components and mycotoxin contamination, which reduces the possibilities of wheat end use and compromises food safety. Lipid degradation by hydrolytic, oxidative and microbial deterioration is the predominant cause of the loss of sensory acceptability, nutritional value and baking quality. The aim of this research was to determine the influence of adverse environmental conditions -as the increasing moisture - on lipid patterns of whole wheat flours contaminated with F. graminearum in relation to the infection degree. In vitro cultures of F. graminearum were carried out on wheat grains under different degrees of relative humidity (11, 50, 75 and 100%) throughout 45 days of incubation at 28 °C. The fungal biomass measured by q-PCR increased proportionally with the humidity. A decrease in the signals of saturated (palmitic and estearic) and unsaturated (oleic, linoleic and linolenic) fatty acids, analyzed as fatty acid methyl esters (FAMEs) by GC-MS, was observed in relation with the humidity and infection degree. The degradation rate of the lipids was high during the first 15 days of incubation, reaching the fatty acids content, values around 20-40% of those found in the control. From that moment on, the rate of degradation was slower or even null. It was observed that in all treatments, the linolenic acid reached the highest degradation ratio in comparison with the other fatty acids, which may be caused by the action of lipoxygenases. The lipase activity and the content of deoxynivalenol were also determinate on the flours. The lipase activity increased until day 25 of incubation reaching twice the initial value. The deoxynivalenol content also increased along incubation while fatty acids decreased. Our results demonstrated that the magnitude in the signal of fatty acids in whole wheat flours varied in relation to the degree of humidity and fungal infection of the grains from which they were obtained. Otherwise, lipids and their oxidation products are related with the pathogenesis and production of mycotoxins. These observations highlight the importance of an adequate manipulation of wheat grains on the processing chain to prevent quality changes and mycotoxins contamination.

Evaluation of different hurdles on Penicillium crustosum growth in sponge cakes by means of a specific real time PCR.

Limited shelf life of bakery products, caused by microbial deterioration, is a concern for industries due to economic losses. Fungal spoilage of sponge cakes industrially produced in Montevideo was caused mainly by Penicillium species, in particular by Penicillium crustosum. The combination of different hurdles was studied to inhibit P. crustosum growth in sponge cakes. A full factorial design was performed to study the effect of the concentration of potassium sorbate, pH, packaging atmosphere and storage time. The results showed that packaging atmosphere and storage time were the significant factors in the ranges tested. No growth was detected in cakes stored in modified atmosphere packaging (MAP) (N2:CO2 50:50) at room temperature (25 °C) for 15 days. The effect of MAP on P. crustosum growth in cakes at room temperature was compared with the effect of air-packaging and storage at low temperature (4 °C) for 30 days. P. crustosum growth was not detected in cakes packaged in MAP, whereas it was detected after 20 days in cakes packaged in air and stored at 4 °C. This growth was quantified by a specific real time PCR developed in this work. Specific primers were designed using the sequence of ß-tubulin gene of P. crustosum as a target and PCR conditions were adjusted to ensure specificity. PCR efficiency was 107%, with a detection limit of 0.0014 ng of DNA. The qPCR method presented here, resulted specific and sensitive enough to detect the growth of P. crustosum even before biodeterioration signs were visible.

Process conditions for a rapid in situ transesterification for biodiesel production from oleaginous yeasts

BACKGROUND: Microbial oils produced by diverse microorganisms are being considered as alternative sources of triglycerides for biodiesel production. However, the standalone production of biodiesel from microorganisms is not currently economically feasible. In case of yeasts, the use of low-value nutrient sources in microbial production and the implementation of cost-efficient downstream processes could reduce costs and make microbial lipids competitive with other commodity-type oils in biodiesel production. Industrial biodiesel synthesis from oleaginous seeds is currently based on a multistep process. However, a simple process called in situ transesterification (ISTE), which takes place within the biomass without a previous lipid extraction step, is receiving increasing interest. In this work, the optimal conditions for an ISTE process to obtain biodiesel from previously selected oleaginous yeast (Rhodotorula graminis S1/S2) were defined using the response surface methodology (RSM). RESULTS: Using the RSM approach, the optimal conditions for the maximum yield with minimum reaction time included a methanol-to-biomass ratio of 60:1, 0.4 M H2SO4, and incubation at 70°C for 3 h. The optimized in situ process yield was significantly higher (123%) than that obtained with a two-step method in which fatty acids from saponifiable lipids were first extracted and then esterified with methanol. The composition of the fatty acid methyl ester mixture obtained from R. graminis S1/S2 by ISTE met Uruguayan standards for biodiesel. CONCLUSION: The characteristics achieved by the optimized method make microbial oil a potential alternative for biodiesel production from yeast at an industrial scale.

Species composition, toxigenic potential and aggressiveness of Fusarium isolates causing Head Blight of barley in Uruguay.

Fusarium Head Blight (FHB) is a major constraint to barley production that substantially reduces yield and grain quality. FHB is also a major food safety concern because FHB pathogens contaminate grain with trichothecenes and other mycotoxins. DNA sequence-based analyses and in-vitro toxin assessments were used to characterize the species and trichothecene chemotype composition of FHB pathogens on barley in Uruguay. F. graminearum was the dominant species (89.7%), and three other members of the F. graminearum species complex (FGSC) were identified as FHB pathogens of barley in Uruguay for the first time. Other minor contributors to FHB species diversity included F. poae, F. avenaceum, F. pseudograminearum and an unnamed species from the F. incarnatum-equiseti species complex (FIESC). Most isolates (89.7%) had the 15-acetyldeoxynivalenol (15-ADON) trichothecene type. However, the results expanded the known area of occurrence within Uruguay for the nivalenol (NIV) toxin type, which was observed among isolates from three species of the FGSC, F. pseudograminearum, and F. poae. Isolates with the 3-acetyldeoxynivalenol (3-ADON) or NX-2 toxin types were not observed, although a previously published multilocus genotyping assay was updated to identify NX-2 strains. Analyses of population structure and comparisons with FHB isolates from wheat in Uruguay indicated that F. graminearum constitutes a single genetic population with no evidence of population differentiation related to the sampled hosts. Inter and intraspecific differences were observed in aggressiveness toward four barley genotypes with different levels of resistance to FHB, and in general nivalenol producers were the least aggressive isolates. Sensitivity to metconazole was approximately 10 times higher than was detected for tebuconazole. This is the first report regarding tebuconazole and metconazole sensitivity for Fusarium species causing FHB in barley in Uruguay, and constitutes an important starting point for monitoring temporal or spatial changes in FGSC sensitivity, which is critical to define FHB management practices.

Development of a PCR-RFLP method based on the transcription elongation factor 1-α gene to differentiate Fusarium graminearum from other species within the Fusarium graminearum species complex.

Fusarium head blight (FHB) is a destructive disease of cereals crops worldwide and a major food safety concern due to grain contamination with trichothecenes and other mycotoxins. Fusarium graminearum, a member of the Fusarium graminearum species complex (FGSC) is the dominant FHB pathogen in many parts of the world. However, a number of other Fusarium species, including other members of the FGSC, may also be present for example in Argentina, New Zealand, Ethiopia, Nepal, Unites States in cereals such as wheat and barley. Proper species identification is critical to research aimed at improving disease and mycotoxin control programs. Identification of Fusarium species is are often unreliable by traditional, as many species are morphologically cryptic. DNA sequence-based methods offer a reliable means of species identification, but can be expensive when applied to the analyses of population samples. To facilitate identification of the major causative agent of FHB, this work describes an easy and inexpensive method to differentiate F. graminearum from the remaining species within the FGSC and from the other common Fusarium species causing FHB in cereals. The developed method is based on a PCR-RFLP of the transcription elongation factor (TEF 1-α) gene using the restriction enzyme BsaHI.

Pectinolytic yeasts from cold environments: novel findings of Guehomyces pullulans, Cystofilobasidium infirmominiatum and Cryptococcus adeliensis producing pectinases.

One hundred and three yeasts isolated from soil samples from King George Island and Tierra del Fuego province were screened in relation with their capability to produce pectinolytic enzymes. Although all the yeasts showed well-developed colonies at 20 °C, only eight showed a clear halo around the colony, indicative of pectin degradation. A secondary screening demonstrated that only four yeasts were capable to produce pectinases at low temperatures (8 °C). It could be seen that the selected yeasts were able to grow and produce high levels of polygalacturonase activity when submerged fermentations were performed using pectin-containing fruit wastes as substrates. None of the strains produced neither lyase nor rhamnogalacturonan hydrolase activities. Regarding pectin esterase activity, it was only produced in lower amounts by G. pullulans 8E (0.022 U ml-1). A TLC analysis of the substrate cleavage pattern of the pectinolytic systems was consistent with an endo-type activity. The clarification of apple juice was only accomplished by G. pullulans pectinolytic system, with a clarification of 80% (%T650) using 4 U/ml of enzyme at 20 °C. As far as we concern this work describes for the first time the production of pectinases by the cold-adapted yeasts species Cystofilobasidium infirmominiatum, Cryptococcus adeliensis and G. pullulans.

Occurrence and biodiversity of Aspergillus section Nigri on 'Tannat' grapes in Uruguay.

Ochratoxin A (OTA) is a nephrotoxic mycotoxin which has been found worldwide as a contaminant in wines. It is produced on grapes mainly by molds from Aspergillus section Nigri. This study has demonstrated for the first time the occurrence of black aspergilli on Tannat grapes from Uruguay, in a two year survey. Aspergillus uvarum (uniseriate) and Aspergillus welwitschiae (from Aspergillusniger aggregate) were the prevalent species whereas Aspergillus carbonarius which is considered the main OTA producing species was not detected. OTA production in culture medium was evaluated for native isolates from A. niger aggregate and compared to levels produced by a type strain of A. carbonarius. This work also includes the development of quick and easy molecular methods to identify black aspergilli to species level, avoiding sequencing.