Elucidating the interaction of carbon, nitrogen, and temperature on the biosynthesis of Aureobasidium pullulans antifungal volatiles.

The combined biochemical impact of carbon, nitrogen and temperature on the biosynthesis of the antifungal volatile organic compounds (VOCs): ethanol, 2-methyl-1-propanol, 3-methyl-1-butanol and 2-phenylethanol produced by Aureobasidium pullulans A1 and A3 was investigated using a Box-Behnken experimental design and response surface methodology (RSM). Normalized peak areas derived from solid phase micro extraction-gas chromatography-mass spectrometry (SPME-GC-MS) analysis, indicated that initial carbon content had a significant influence on the biosynthesis of ethanol and alcohols with greater than three carbon atoms. This result suggests a dominant activity of the A. pullulans anabolic pathway to biosynthesize three higher alcohols via de novo biosynthesis of amino acids from sugar metabolism. Low concentrations of carbon (3-13 g l-1 ) with nitrogen as both ammonium and amino acids in the growth medium resulted in a higher number of significant linear and quadratic relationships. Nitrogen availability and growth temperature had significant negative linear and quadratic correlations with VOCs biosynthesis in most instances. Isolate-dependant metabolic response was evident for all abiotic parameters tested on alcohol production. The findings of this study offer new perspectives to improve the production of key antifungal compounds by antagonists in biological control systems.

Secondary Metabolites Coordinately Protect Grapes from Excessive Light and Sunburn Damage during Development.

Sunburn is a physiological disorder that reduces grape quality and vineyard yield. It is the result of excessive sunlight and high temperatures. As climate change continues to increase air temperatures, reports of sunburn damage in vineyards worldwide are becoming more frequent. Grapes produce secondary metabolites (carotenoids, polyphenols and aroma compounds) to counter photooxidative stress and acclimate to higher radiation environments. This study evaluated changes in these compounds in during ripening when grapes were exposed post-flowering (ED) and at véraison (LD), and compared them to a nondefoliated control (ND). ND contained more α-terpineol and violaxanthin, and the defoliated treatments contained more zeaxanthin, ß-carotene, C6 compounds and flavonoids. ED berries adapted better to higher-light environments, displayed larger changes in secondary metabolite concentrations and lower levels of sunburn damage than LD berries did. The composition of berries with increasing sunburn damage was evaluated for the first time. Berries with no damage had the lowest concentrations of flavonoids and oxidized glutathione, and the highest concentrations of chlorophyll and α-terpineol. As damage increased, destruction of photosynthetic pigments, increase in polyphenols and loss of aroma compounds were evidenced. A significant effect of temperature and developmental stage on grape composition was also observed. This study provides a holistic overview of changes in secondary metabolites experienced by grape berries when exposed to excessive light, how these vary along development and how they affect sunburn incidence.

Volatile organic compounds produced by Aureobasidium pullulans induce electrolyte loss and oxidative stress in Botrytis cinerea and Alternaria alternata.

Aureobasidium pullulans is a yeast-like fungus that produces volatile organic compounds (VOCs) with antifungal properties. VOCs have the potential to trigger the production of intracellular reactive oxygen species (ROS), lipid peroxidation and electrolyte loss in microorganisms. The relationship among A. pullulans VOCs, induced ROS accumulation and electrolyte leakage was investigated in Botrytis cinerea and Alternaria alternata in vitro. Exposure to a mixture of A. pullulans VOCs: ethanol, 2-methyl-1-propanol, 3-methyl-1-butanol and 2-phenylethanol, resulted in electrolyte leakage in both B. cinerea and A. alternata. Fluorescence microscopy using 2',7'-dichlorofluorescein diacetate indicated triggered ROS accumulation in exposed fungal mycelia and the presence of the superoxide radical was evident by intense red fluorescence with dihydroethidium. Partial inhibition of enzymes of the mitochondrial respiratory chain complex I of B. cinerea and A. alternata by pre-treatment with rotenone reduced ROS accumulation in hypha exposed to A. pullulans VOCs and reversed the VOCs inhibition of fungal growth. Scanning electron micrographs revealed that B. cinerea and A. alternata hypha exposed to A. pullulans VOCs had altered cell wall structures. Our findings give insights into the potential mechanisms involved in the antifungal properties of A. pullulans in the suppression of B. cinerea and A. alternata growth in vitro.

Lower-alcohol wines produced by Metschnikowia pulcherrima and Saccharomyces cerevisiae co-fermentations: The effect of sequential inoculation timing.

In Latin, 'pulcherrima' is a superlative form of an adjective that translates as beautiful. Apart from being 'the most beautiful' yeast, Metschnikowia pulcherrima has a remarkable potential in production of wines with lower ethanol content. The oenological performance of six M. pulcherrima strains was hereby tested in sequential cultures with Saccharomyces cerevisiae. The best-performing strain MP2 was further characterised in fermentations with different S. cerevisiae inoculation delays in both white grape juice and Chemically Defined Grape Juice Medium (CDGJM). The analysis of main metabolites, undertaken prior to sequential inoculations and upon fermentation completion, highlighted metabolic interactions and carbon sinks other than ethanol in MP2 treatments. Depending on the inoculation delay, MP2 white wines contained between 0.6% and 1.2% (v/v) less ethanol than the S. cerevisiae monoculture, with even larger decreases detected in the CDGJM. The MP2 treatments also contained higher concentrations of TCA cycle by-products (i.e. fumarate and succinate) and glycerol, and lower concentrations of acetic acid. The analysis of volatile compounds showed increased production of acetate esters and higher alcohols in all MP2 wines, alongside other compositional alterations arising from the S. cerevisiae inoculation delay.

A matter of place: Sensory and chemical characterisation of fine Australian Chardonnay and Shiraz wines of provenance.

Establishing a fine wine image through regional typicality has been of interest to New World wine producing countries like Australia, but previous research mainly involved unoaked experimental wines, which were not reflective of the retail wine market. The regional typicality of commercially available fine Australian wines (FAW) was therefore explored, based on the hypotheses that sensory and chemical composition of varietal fine wines would discriminate by region, and further nuances within region would be explained by drivers of intraregional typicality. Chardonnay wines (2015 vintage) from Margaret River (MR, n = 16) and Yarra Valley (YV, n = 16); and Shiraz wines (2014 vintage) from Barossa Valley (BV, n = 16) and McLaren Vale (MV = 15), were selected for descriptive sensory analysis and underwent profiling of volatiles by gas chromatography-mass spectrometry (GC-MS). For both grape varieties, there was large variability in wine styles among wines from the same GI, such as fruity/crisp vs oaked Chardonnay and oaky/astringent vs savoury Shiraz. Consequently, human intervention seemed to be an important component of regional/sub-regional typicality, which therefore cannot be determined solely on geographic origin of the fruit. Using a combination of sensory markers and volatile profiles allowed the building of regional typicality models, which are promising, however, consumers may not perceive sub-regional differences based on sensory attributes. Undoubtedly, variation of wine styles emerging across wine regions, vintages, and viticultural and winemaking practices needs to be further explored, but this work created a preliminary sensory and volatile map for future research.

Sunburn in Grapes: A Review.

Sunburn is a physiological disorder that affects the visual and organoleptic properties of grapes. The appearance of brown and necrotic spots severely affects the commercial value of the fruit, and in extreme cases, significantly decreases yield. Depending on the severity of the damage and the driving factors, sunburn on grapes can be classified as sunburn browning (SB) or as sunburn necrosis (SN). Sunburn results from a combination of excessive photosynthetically active radiation (PAR) and UV radiation and temperature that can be exacerbated by other stress factors such as water deficit. Fruit respond to these by activating antioxidant defense mechanisms, de novo synthesis of optical screening compounds and heat-shock proteins as well as through morphological adaptation. This review summarizes the current knowledge on sunburn in grapes and compares it with relevant literature on other fruits. It also discusses the different factors affecting the appearance and degree of sunburn, as well as the biochemical response of grapes to this phenomenon and different potential mitigation strategies. This review proposes further directions for research into sunburn in grapes.

Oenological traits of Lachancea thermotolerans show signs of domestication and allopatric differentiation.

The yeast Lachancea thermotolerans (previously Kluyveromyces thermotolerans) is a species of large, yet underexplored, oenological potential. This study delivers comprehensive oenological phenomes of 94 L. thermotolerans strains obtained from diverse ecological niches worldwide, classified in nine genetic groups based on their pre-determined microsatellite genotypes. The strains and the genetic groups were compared for their alcoholic fermentation performance, production of primary and secondary metabolites and pH modulation in Chardonnay grape juice fermentations. The common oenological features of L. thermotolerans strains were their glucophilic character, relatively extensive fermentation ability, low production of acetic acid and the formation of lactic acid, which significantly affected the pH of the wines. An untargeted analysis of volatile compounds, used for the first time in a population-scale phenotyping of a non-Saccharomyces yeast, revealed that 58 out of 90 volatiles were affected at an L. thermotolerans strain level. Besides the remarkable extent of intra-specific diversity, our results confirmed the distinct phenotypic performance of L. thermotolerans genetic groups. Together, these observations provide further support for the occurrence of domestication events and allopatric differentiation in L. thermotolerans population.

Managing the excessive proliferation of glycogen accumulating organisms in industrial activated sludge by nitrogen supplementation: A FISH-NanoSIMS approach.

Defluviicoccus vanus-related glycogen accumulating organisms (GAO) regularly proliferate in industrial wastewater treatment plants handling high carbon but nitrogen deficient wastes. When GAO dominate, they are associated with poor performance, characterised by slow settling biomass and turbid effluents. Although their ecophysiology has been studied thoroughly in domestic waste treatment plants, little attention has been paid to them in aerobic industrial systems. In this study, the effect of nitrogen addition on GAO carbon metabolism was investigated during an 8h cycle. Activated sludge dominated by GAO from a winery wastewater sequencing batch reactor was incubated under different carbon to nitrogen (COD:N) ratios (100:1, 60:1 and 20:1) using 13C - acetate and 15N - urea. GAO cell assimilation was quantified using FISH-NanoSIMS. The activated sludge community was assessed by 16S rRNA gene profiling, DNA and storage polymer production. Carbon and nitrogen quantification at the cellular level by NanoSIMS revealed that low (COD:N of 100:1) or null nitrogen concentrations enhanced GAO carbon uptake. COD:N ratios of 60:1 and 20:1 reduced GAO carbon uptake and promoted whole microbial community DNA production. Nitrogen dosing at COD:N ratios of 60:1 or higher was demonstrated as feasible strategy for controlling the excessive GAO growth in high COD waste treatment plants.

Exploring the Effects of Geographical Origin on the Chemical Composition and Quality Grading of Vitis vinifera L. cv. Chardonnay Grapes.

The relationship between berry chemical composition, region of origin and quality grade was investigated for Chardonnay grapes sourced from vineyards located in seven South Australian Geographical Indications (GI). Measurements of basic chemical parameters, amino acids, elements, and free and bound volatiles were conducted for grapes collected during 2015 and 2016. Multiple factor analysis (MFA) was used to determine the sets of data that best discriminated each GI and quality grade. Important components for the discrimination of grapes based on GI were 2-phenylethanol, benzyl alcohol and C6 compounds, as well as Cu, Zn, and Mg, titratable acidity (TA), total soluble solids (TSS), and pH. Discriminant analysis (DA) based on MFA results correctly classified 100% of the samples into GI in 2015 and 2016. Classification according to grade was achieved based on the results for elements such as Cu, Na, Fe, volatiles including C6 and aryl alcohols, hydrolytically-released volatiles such as (Z)-linalool oxide and vitispirane, pH, TSS, alanine and proline. Correct classification through DA according to grade was 100% for both vintages. Significant correlations were observed between climate, GI, grade, and berry composition. Climate influenced the synthesis of free and bound volatiles as well as amino acids, sugars, and acids, as a result of higher temperatures and precipitation.

Comprehensive Study of Volatile Compounds in Two Australian Rosé Wines: Aroma Extract Dilution Analysis (AEDA) of Extracts Prepared Using Solvent-Assisted Flavor Evaporation (SAFE) or Headspace Solid-Phase Extraction (HS-SPE).

Two rosé wines, representing a tropical and a fruity/floral style, were chosen from a previous study for further exploration by aroma extract dilution analysis (AEDA) and quantitative analysis. Volatiles were extracted using either liquid-liquid extraction (LLE) followed by solvent-assisted flavor evaporation (SAFE) or a recently developed dynamic headspace (HS) sampling method utilizing solid-phase extraction (SPE) cartridges. AEDA was conducted using gas chromatography-mass spectrometry/olfactometry (GC-MS/O) and a total of 51 aroma compounds with a flavor dilution (FD) factor ≥3 were detected. Quantitative analysis of 92 volatiles was undertaken in both wines for calculation of odor activity values. The fruity and floral wine style was mostly driven by 2-phenylethanol, ß-damascenone, and a range of esters, whereas 3-SHA and several volatile acids were seen as essential for the tropical style. When extraction methods were compared, HS-SPE was as efficient as SAFE for extracting most esters and higher alcohols, which were associated with fruity and floral characters, but it was difficult to capture volatiles with greater polarity or higher boiling point that may still be important to perceived wine aroma.

Factors influencing the aroma composition of Chardonnay wines.

Chardonnay is one of the oldest and most widely distributed wine grape cultivars and is of commercial importance for the world's wine-producing nations. It is an extremely flexible variety that has adapted to different regions with varied weather and soil characteristics. Somewhat uniquely among white wines, Chardonnay lends itself to a wide variety of production styles, which can be tailored to the target market. Techniques such as skin maceration, barrel and stainless steel fermentation, use of selected or indigenous yeasts, malolactic fermentation, and aging in barrels with or without lees are all applicable and lead to different compositional outcomes. A number of research papers have been published with a view to understanding Chardonnay composition and quality as well as the impact of different enological techniques on the final product. This review summarizes current knowledge, explaining the influence of viticultural and production techniques on aroma composition, and poses directions for further research into Chardonnay wines.