Sequential Inoculation Research Articles

Interaction studies of Serendipita indica and Zhihengliuella sp. ISTPL4 and their synergistic role in growth promotion in rice.

Rapid urbanization and globalization demand increasing agricultural productivity. Soil nutrient supply capacity is continuously decreasing due to soil erosion, degradation, salt deposition, undesired element, metal deposition, water scarcity, and an uneven nutrient delivery system. Rice cultivation requires a large amount of water which is becoming detrimental due to these activities. There is a need to increase its productivity. Microbial inoculants are becoming increasingly important in achieving sustainable agricultural production systems. The current study was conducted to investigate the interaction between the root endophytic fungus Serendipita indica (S. indica) and the actinobacterium Zhihengliuella sp. ISTPL4 (Z. sp. ISTPL4) and their synergistic effects on the growth of rice (Oryza sativa L). Both S. indica and Z. sp. ISTPL4 showed positive interactions. Growth of S. indica was observed at different days after Z. sp. ISTPL4 inoculation, and stimulated growth of S. indica was observed when Z. sp. ISTPL4 was inoculated at 5 dafi (days after fungal inoculation). Z. sp. ISTPL4 promoted the growth of S. indica as it increased spore germination. Furthermore, confocal and scanning electron microscopy (SEM) analyses showed a 27% increase in the spore size of S. indica in the presence of Z. sp. ISTPL4. In a liquid chromatography with tandem mass spectrometry (LC-MS/MS) analysis increased production of alanine and glutamic acid was observed in their sequential co-culture as compared with individual cultures. Sequential inoculation of S. indica and Z. sp. ISTPL4 significantly increased the biochemical and physical characteristics of rice as compared with their individual inoculum. Biochemical parameters such as chlorophyll content, total soluble sugar, and flavonoid content in the rice increased by up to 57%, 47%, and 39%, respectively, in the presence of the combined inoculum of S. indica and Z. sp. ISTPL4. This will be the first study, to the best of our knowledge, which shows the fungus and actinobacterium interaction and their synergistic roles in the growth promotion of rice. Furthermore, this novel combination can also be used to boost the growth of other crops to increase the agricultural yield.

Effect of Sequential Fermentation with Lachancea thermotolerans/S. cerevisiae on Aromatic and Flavonoid Profiles of Plavac Mali Wine.

In this study, the effects of sequential fermentation of Lachancea thermotolerans/S. cerevisiae on the production of Plavac Mali wines were investigated in comparison with the commonly used inoculation of the commercial Saccharomyces cerevisiae strain and spontaneous fermentation. A total of 113 aroma compounds and 35 polyphenolic compounds were analyzed. Sequential inoculation resulted in a decrease in alcohol content and pH (up to 0.3% v/v and 0.12 units, respectively) and an increase in total acidity (0.6 g/L, expressed as tartaric acid). The wines produced by spontaneous fermentation exhibited the greatest diversity of volatile compounds and the highest concentration of C13 norisoprenoids, lactones, and other compounds. These wines exhibited maximum hydroxycinnamic acids, prodelphinidin monomer units, epigallocatechin, B1, B3, and B4 dimers, and total flavan-3-ols. Sequential inoculation decreased the content of the aromas and polyphenols in the wines. The practical significance of this procedure lies in the selective effect on aroma compounds, the decrease in green aromas, undetectable volatile phenols, and the decrease in bitter and astringent compounds such as gallic acid, flavan-3-ol monomers (catechin and epicatechin), and dimers (B1, B2, B3, and B4). This work demonstrates the potential of sequential and spontaneous fermentation to improve the aromatic characteristics and overall quality of Plavac Mali wines.

Intracellular bottlenecking permits no more than three tomato yellow leaf curl virus genomes to initiate replication in a single cell

Viruses are constantly subject to natural selection to enrich beneficial mutations and weed out deleterious ones. However, it remains unresolved as to how the phenotypic gains or losses brought about by these mutations cause the viral genomes carrying the very mutations to become more or less numerous. Previous investigations by us and others suggest that viruses with plus strand (+) RNA genomes may compel such selection by bottlenecking the replicating genome copies in each cell to low single digits. Nevertheless, it is unclear if similarly stringent reproductive bottlenecks also occur in cells invaded by DNA viruses. Here we investigated whether tomato yellow leaf curl virus (TYLCV), a small virus with a single-stranded DNA genome, underwent population bottlenecking in cells of its host plants. We engineered a TYLCV genome to produce two replicons that express green fluorescent protein and mCherry, respectively, in a replication-dependent manner. We found that among the cells entered by both replicons, less than 65% replicated both, whereas at least 35% replicated either of them alone. Further probability computation concluded that replication in an average cell was unlikely to have been initiated with more than three replicon genome copies. Furthermore, sequential inoculations unveiled strong mutual exclusions of these two replicons at the intracellular level. In conclusion, the intracellular population of the small DNA virus TYLCV is actively bottlenecked, and such bottlenecking may be a virus-encoded, evolutionarily conserved trait that assures timely selection of new mutations emerging through error-prone replication.

Starter cultures inoculation procedure changes microbial community structure during low-salt moromi fermentation

Soy sauce is a condiment made of soybeans fermented in high salt concentration (18-22% NaCl) called moromi. Efforts to prevent sodium overconsumption has led to salt reduction in moromi. However, it could alter the microbial community dynamics during fermentation, resulting in undesirable sensory changes. This study investigates the effect of moromi starter cultures, Tetragenococcus sp. and Zygosaccharomyces rouxii, on microbial population changes during reduced-salt moromi (6% NaCl) fermentation. Both microbes were inoculated at different times, concentrations, and ratios, and the changes in total bacterial and yeast population during fermentation were monitored by culture on tryptic soy agar (TSA) and potato dextrose agar (PDA), respectively. The results showed that the inoculum size could shorten the bacterial exponential growth as it increased. Moreover, increasing the proportion of Tetragenococcus or Z. rouxii by two-fold could enhance the yeast and bacterial population, respectively by ∼3 log CFU/mL. Simultaneous inoculation suppressed bacterial growth, while sequential inoculation of Z. rouxii could maximize bacterial growth up to 7.38 log CFU/mL. However, a synergistic growth in simultaneous inoculation was observed when 5% inoculum with an equal ratio was used. This study could help researchers and manufacturers determine appropriate inoculation procedures to control fermentation better.

Oenological characteristics of two indigenous Starmerella bacillaris strains isolated from Chinese wine regions.

To broaden knowledge about the oenological characteristics of Starmerella bacillaris, the influence of two Chinese indigenous S. bacillaris strains on the conventional enological parameters and volatile compounds of Cabernet Sauvignon wines were investigated under different inoculation protocols (single inoculation and simultaneous/sequential inoculation with the commercial Saccharomyces cerevisiae EC1118). The results showed that the two S. bacillaris strains could complete alcohol fermentation alone under high sugar concentrations while increasing the content of glycerol and decreasing the content of acetic acid. Compared with wines fermented by EC1118 single inoculation, S. bacillaris single inoculation and S. bacillaris/EC1118 sequential inoculation increased the contents of isobutanol, ethyl isobutanoate, terpenes, and ketones and decreased the contents of isopentanol, phenylethyl alcohol, fatty acids, acetate esters, and total ethyl esters. Furthermore, for S. bacillaris/EC1118 simultaneous inoculation, the concentrations of ethyl esters were increased, contributing to a higher score of "floral" and "fruity" notes in agreement with sensory analysis. KEY POINTS: • S. bacillaris single and simultaneous/sequential inoculation. • Conventional enological parameters and volatile compounds were investigated. • S. bacillaris/EC1118 simultaneous fermentation increased ethyl esters.

Effects of native Lachancea thermotolerans combined with Saccharomyces cerevisiae on wine volatile and phenolic profiles in pilot and industrial scale

Wines in the northwest region of China often suffer from poor color stability, low wine acidity, and inelegant aroma description. The Lachancea thermotolerans yeasts with a high yield of lactic acid and pleasant aroma compounds can ameliorate such wines. In this study, the performances of native L. thermotolerans CVE-LT1 co-inoculated with Saccharomyces cerevisiae in different ratios and strategies (simultaneous and sequential mode) were evaluated in pilot scale fermenter (60 L) and industrial scale fermenter (520 hL) of Cabernet Sauvignon grape must with high sugar/low acidity. Results evidenced that the mixed culture of L. thermotolerans and S. cerevisiae can significantly increase lactic acid level (up to 6.98 g/L) and decrease wine pH (up to 3.57). Meanwhile, the color parameters of a* and C* in mixed culture wines were improved. The higher concentrations of volatile compounds were mainly observed in sequential inoculation treatments, including the enhancement of ethyl lactate and 2-phenylethanol. As for phenolic compounds, L. thermotolerans promoted the formation of anthocyanins derivatives and phenolic acids in pilot and industrial fermentation. In conclusion, these results indicated that L. thermotolerans LT1 can be used in hot winery regions to improve the acidity, color indexes, phenolic compounds, and aroma quality of wines.

Improving acidity and flavors of citrus juice as well as its antioxidant activity by cofermentation with deacidification bacteria combination

The high acidity and poor taste of some citrus juice (CJ) affect its application value. In this study, a Hanseniaspora pseudoguilliermondii B4 was isolated and identified from lemon which could effectively degrade organic acids in CJ. Seven lactic acid bacteria (LAB) were selected to coferment with B4 in sequential and simultaneous inoculation on CJ to improve its taste and nutritional value. The physicochemical parameters, total phenolic content (TPC), total flavonoid content (TFC) and antioxidant activity were evaluated before and after mixed fermentation. The results revealed that cofermentation with B4 and Bifidobacterium lactis 20435 could not only decreased acidity by 96.05% but also increased TPC, TFC, DPPH and ABTS radical scavenging activity by 22.54%, 25.42%, 34.47% and 52.63%, respectively. In addition, 3-methyl-1-butanol, 2-methyl-1-butanol and phenylethyl alcohol were significantly (p < 0.05) increased, while terpenes were mostly decreased, compared to unfermented juice. The ester content increased and enhanced the flavor compounds of fermented CJ. This work provides a new strategy of using H. pseudoguilliermondii and LAB to adjust the acidity and taste of CJ as well as improve its bioactive substances and antioxidant activity.

Effects of Mixed Fermentation on the Aroma Compounds of ‘Italian Riesling’ Dry White Wine in Eastern Foothill of Helan Mountain

To study the effect of mixed fermentation of non-Saccharomyces strains and Saccharomyces cerevisiae on the aroma quality of ‘Italian Riesling’ wine in the eastern foothill of Helan Mountain and to determine the most optimum process of mixed fermentation, two selected non-Saccharomyces strains, including Hanseniaspora uvarum YUN268 and Pichia fermentans Z9Y-3, were inoculated with Saccharomyces cerevisiae in different proportions (10:1 or 1:1) and different stages (48 h in advance or simultaneously at the beginning) to ferment ‘Italian Riesling’ dry white wine. The oenological parameters and aroma indexes of the wine samples were evaluated. The results showed mixed fermentation can not only reduce the alcohol content of wine 0.24~0.71% vol but also increase the glycerol content to improve the taste of wine. The mixed fermentation effect of Pichia fermentans Z9Y-3 and Saccharomyces cerevisiae resulted in improvements, especially the high proportion (10:1) sequential inoculation and simultaneous inoculation of wine samples (WSP10 and WCP10), which not only produced more volatile aroma substances and glycerol content but also increased the total amount of ester substances by 49.4% and 56.5%, respectively, compared with the control. The sensory evaluation scores of WSP10 and WCP10 were significantly higher than the control (89.3 and 88.1 points, respectively). At the same time, it can also enhance the aroma of lemon, cream, almond, and others and increase the aroma complexity of wine. Therefore, these two methods of mixed fermentation inoculation are more suitable for the production of Italian Riesling wine in the eastern foothill of Helan Mountain. In conclusion, the mixed fermentation of Pichia fermentans Z9Y-3 and Saccharomyces cerevisiae 10:1 (simultaneous or sequential) inoculation is suitable for the production of Italian Riesling dry white wine in the eastern foothill of Helan Mountain.

Contribution of Starmerella bacillaris and Oak Chips to Trebbiano d'Abruzzo Wine Volatile and Sensory Diversity.

In this study, six fermentation trials were carried out: co-inoculation and sequential inoculation of Saccharomyces cerevisiae and Starmerella bacillaris in the presence and absence of oak chips. Moreover, Starm. bacillaris strain was attached to the oak chips and co-inoculated or sequentially inoculated with S. cerevisiae. Wines fermented with Starm. bacillaris adhered to oak chips showed a higher concentration of glycerol (more than 6 g/L) than the others (about 5 g/L). These wines also showed a higher content of polyphenols (more than 300 g/L) than the others (about 200 g/L). The addition of oak chips induced an increase of yellow color (b* value of about 3). Oak-treated wines were characterized by a higher concentration of higher alcohols, esters and terpenes. Aldehydes, phenols and lactones were detected only in these wines, independently from the inoculation strategy. Significant differences (p < 0.05) were also observed in the sensory profiles. The fruity, toasty, astringency, and vanilla sensations were perceived as more intense in wines treated with oak chips. The white flower descriptor showed a higher score in wines fermented without chips. Oak surface-adhered Starm. bacillaris cells could be a good strategy to improve the volatile and sensory profile of Trebbiano d'Abruzzo wines.

Combined Use of Schizosaccharomyces pombe and a Lachancea thermotolerans Strain with a High Malic Acid Consumption Ability for Wine Production

The development of new fermentative strategies exploiting the potential of different wine-related species is of great interest for new winemaking conditions and consumer preferences. One of the most promising non-conventional approaches to wine fermentation is the combined use of deacidifying and acidifying yeasts. Lachancea thermotolerans shows several other properties besides lactic acid production; among them, high malic acid consumption is of great interest in the production of red wines for avoiding undesirable refermentations once bottled. The combination of a L. thermotolerans strain that is able to consume malic acid with a Schizosaccharomyces pombe strain helps to ensure malic acid elimination during alcoholic fermentation while increasing the final acidity by lactic acid production. To properly assess the influence of this alternative strategy, we developed combined fermentations between specific strains of L. thermotolerans and S. pombe under sequential inoculation. Both species showed a great performance under the studied conditions, influencing not only the acidity but also the aromatic compound profiles of the resulting wines. The new proposed biotechnological strategy reduced the final concentrations of ethanol, malic acid and succinic acid, while it increased the concentrations of lactic acid and esters.

Research on the Effect of Simultaneous and Sequential Fermentation with Saccharomyces cerevisiae and Lactobacillus plantarum on Antioxidant Activity and Flavor of Apple Cider

The study examined the effect of Lactobacillus plantarum together with Saccharomyces cerevisiae on cider quality through simultaneous and sequential inoculation strategies to evoke malolactic fermentation. The antioxidant activities and flavor compound profiles of apple ciders fermented with mixed cultures of commercial wine yeast (S. cerevisia SY) and autochthonous bacteria (L. plantarum SCFF107 and L. plantarum SCFF200) were assessed. The antioxidant ability results indicated that apple ciders fermented with the simultaneous inoculation method had a higher DPPH radical scavenging rate and total antioxidant capacity, especially for SIL107 cider (simultaneous inoculation with S. cerevisiae SY and L. plantarum SCFF107), which exhibited the highest DPPH free radical scavenging activity (78.14% ± 0.78%) and the highest total antioxidant ability (255.92 ± 7.68 mmol/L). The results showed that ciders produced by mixed inoculation with L. plantarum improved flavor because of their higher contents of volatiles such as esters and higher alcohols and higher contents of non-volatile compounds like organic acids and polyphenols in comparison with the single culture of S. cerevisiae, especially for the simultaneous inoculation method. In addition, irrespective of the inoculation mode, compared to the single culture of cider, L-malic acid degraded dramatically in the presence of L. plantarum during alcoholic fermentation, accompanied by increases in lactic acid. What is more, sensory evaluation results demonstrated that ciders produced by mixed cultures gained higher scores than ciders fermented by the single culture of S. cerevisiae, especially in the simultaneous inoculation mode, in terms of the floral, fruity, and overall acceptability of the cider. Therefore, our results indicated that simultaneous inoculation with L. plantarum was found to compensate for some enological shortages of single S. cerevisiae fermented ciders, which could be a potential strategy to enhance the quality of cider products.

Sequential culture with aroma-producing yeast strains to improve the quality of Kyoho wine.

Despite many non-Saccharomyces yeasts being considered spoilage microorganisms, they can increase aroma and flavor diversity in alcoholic beverages. The purpose of this study was to investigate nontraditional inoculation strategies using aroma-producing yeast strains for Kyoho wine fermentation, followed by an instrumental analysis and sensory evaluation. The winemaking process was carried out using Saccharomyces cerevisiae Gr112, Hanseniaspora uvarum Pi235, and Pichia kluyveri Pe114. Multiple inoculation strategies were explored. In instrumental analysis results, mixed culture could promote the formation of esters (5.9-folds) and glycerol (1.3-folds) and reduce the content of ethanol (-0.5% [v/v]) in wine. The sensory analysis results suggested that the three yeast strains sequential inoculation treatment was associated with the aroma attributes "floral," "red fruity," and "tropical fruity." Co-cultivation contributed to an increase in complexity and aromatic intensity, with the three-strain inoculation treatment presenting a more distinctive appearance. PRACTICAL APPLICATION: The inoculation of S. cerevisiae improved the accumulation of volatile acids and esters by inhibiting the growth of non-Saccharomyces yeast strains. Inoculation of H. uvarum and P. kluyveri would effectively solve the defect of excessive content of higher alcohols in wines produced by S. cerevisiae. The suitable inoculation strategy between non-Saccharomyces yeasts could improve the overall quality of Kyoho wine whose starter might be widely used in fermentation industry.

Torulaspora delbrueckii May Help Manage Total and Volatile Acidity of Santorini-Assyrtiko Wine in View of Global Warming.

Non-Saccharomyces (NS) yeasts are gaining popularity in modern winemaking for improving wine quality. Climate change is one of the biggest challenges winegrowing now faces in warm regions. Here, Lachancea thermotolerans LtS1 and Torulaspora delbrueckii TdS6 combined with Saccharomyces cerevisiae ScS13 isolated from Assyrtiko grapes from Santorini island were evaluated in grape must fermentation with the aim to mitigate major consequences of temperature rise. Different inoculation protocols were evaluated, including simultaneous and sequential mixed-strain inoculations, displaying significant variation in the chemical and kinetic characteristics. Both LtS1 and TdS6 could raise the titratable acidity (TA). TdS6 also reduced the volatile acidity (VA) and was thus chosen for further evaluation in microvinifications and pilot-scale fermentations. Consistent with lab-scale trials, sequential inoculation exhibited the longest persistence of TdS6 resulting in minimum VA levels. Diethyl succinate, ethyl propanoate, and ethyl isobutyrate were significantly increased in sequential inoculations, although a decline in the net total ester content was observed. On the other hand, significantly higher levels of TA, succinic acid, and 2-methylpropanoic were associated with sequential inoculation. The overall performance of TdS6 coupled with a high compatibility with S. cerevisiae suggests its use in the fermentation of Santorini-Assyrtiko or other high sugar musts for the production of structured dry or sweet wines.

Evaluation of bio-acidifying yeast Lachancea thermotolerans as a strategy to reduce the effects of climate change in Tempranillo grape must vinification

The effects of climate change are posing major challenges for winemaking, especially in warm regions. Rising temperatures are leading to a considerable increase in sugar content and a reduction in the acidity of grape juices. This has prompted the search for new chemical and biological tools to reduce these effects in winemaking. Lachancea thermotolerans represent an interesting tool for wine improvement, due to their effects on lactic acid, ethanol, and volatile acidity. In this study, we evaluated the sequential inoculation of Lachancea thermotolerans (AEB Italia) + Saccharomyces cerevisiae as an alternative to the addition of water and tartaric acid in the vinification of over-ripened wine must (Tempranillo cv.). To do so, we conducted a micro vinification experiment that included three experimental treatments (replicated four times); AAS = Addition of water + tartaric acid fermented with Saccharomyces cerevisiae, ALS = sequential fermentation with Lachancea thermotolerans followed by Saccharomyces cerevisiae and SAC = Only inoculation with Saccharomyces cerevisiae (control). We measured the lactic acid, malic acid, acetic acid, total acidity, pH, glucose + fructose, and ethanol concentration of the resulting wines. We found a higher lactic acid content and total acidity in ALS wines (2.2 g/L y 6.65 g/L) compared with AAS (0.13 g/L y 5.75 g/L) and SAC wines (0.18 g/L y 5.37 g/L). ALS wines showed a significantly lower pH (4.00) than SAC wines (4.19) but did not differ significantly from AAS wines (3.19). Finally, acetic acid was higher in ALS wines (0.89 g/L) than AAS wines (0.64 g/L) but did not differ significantly from SAC wines (1.02 g/L). Our results point out that the use of Lachancea thermotolerans in a sequential inoculation with Saccharomyces cerevisiae is an alternative to the addition of water and tartaric acid in the vinification of over-ripened Tempranillo musts.

Effects of Torulaspora delbrueckii co-fermented with Saccharomyces cerevisiae on physicochemical and aromatic profiles of blueberry fermented beverage

To investigate the effects of mixed fermentation with T. delbrueckii on aroma profiles of blueberry fermented beverage, five fermentations were conducted: monoculture of T. delbrueckii and S. cerevisiae, respectively; co-inoculation of two strains; sequential inoculation of two strains at time intervals of 24 h and 48 h, respectively. Compared with pure S. cerevisiae fermentation, ethanol level was decreased by up to 1.1% vol., while total anthocyanins were increased by 27.7%-85.0% in mixed fermentations. Marker aroma compounds in different fermentations with relative odor activity values higher than 1were identified. T. delbrueckii significantly decreased volatile acid content (especially acetic acid) by 22.2%-83.3%. Ethyl 3-methylbutanoate, ethyl hexanoate and ethyl octanoate, in pure T. delbrueckii fermentation were significantly decreased, while their concentrations were increased by 1.6–4.4 folds in sequential fermentations. Besides, linalool, rose oxide, benzeneacetaldehyde were significantly increased by sequential fermentation, which was associated with the enhancement of fruity and sweet notes.

Indigenous yeast can increase the phenolic acid and volatile ester compounds in Petit Manseng wine.

Indigenous yeasts are generally found in grapes, vineyards, and natural environments. Sequential inoculation and fermentation with non-Saccharomyces cerevisiae yeast (H30) and Saccharomyces cerevisiae (YT13) also improve the flavor of wine. This study sequentially inoculated fermented Petit Manseng and natural grape juice with native H30 and YT13 selected from vineyards in Yantai, China. The sensory characteristics of Petit Manseng wine were evaluated by detecting the primary organic acids, phenolic acid compounds, and volatile ester compounds. The results showed that the lactic acid content of the natural wine fermented sequentially with H30 and YT13 increased by 490 μg/L compared with the control group, while the ferulic acid content was 1.4 times that of the single-yeast fermentation group. Furthermore, butyrolactone and anthocyanidin propionate were present in the mixed fermentation group, increasing the aroma complexity of Petit Manseng wine and providing high-quality yeast resources that increase the regional characteristics when producing dry white wine.

Effect of Hanseniaspora uvarum-Saccharomyces cerevisiae Mixed Fermentation on Aroma Characteristics of Rosa roxburghii Tratt, Blueberry, and Plum Wines.

Hanseniaspora uvarum, a non-Saccharomyces cerevisiae species, has a crucial effect on the aroma characteristics of fruit wines, thus, attracting significant research interest in recent years. In this study, H. uvarum-Saccharomyces cerevisiae mixed fermentation was used to ferment Rosa roxburghii Tratt, blueberry fruit wine, and plum fruit wines using either a co-inoculated or a sequentially inoculated approach. The three fruit wines' volatile aroma characteristics were analyzed by headspace-solid-phase microextraction-gas chromatography-mass spectrometry (HS-SPME-GC-MS). The results showed that the mixed inoculation of H. uvarum and S. cerevisiae reduced the alcoholic content of Kongxinli fruit wine. Moreover, H. uvarum-S. cerevisiae fermented Rosa roxburghii Tratt, blueberry, and plum fruit wines and further enriched their flavor compounds. The overall flavor characteristics of sequentially inoculated fruit wines differed significantly from those fermented with S. cerevisiae alone, although several similarities were also observed. Sequential inoculation of H. uvarum and S. cerevisiae positively affected the mellowness of the wine and achieved a better harmony of the overall wine flavors. Therefore, H. uvarum-Saccharomyces cerevisiae mixed fermentation can improve the complexity of the wines' aromatic composition and empower them with a unique identity. In particular, H. uvarum-Saccharomyces cerevisiae blueberry wine produced by mixed fermentation had the widest variety and content of aroma compounds among the fermented wines. Therefore, H. uvarum-Saccharomyces cerevisiae mixed-fermentation inoculation in the three fermented fruit wines significantly increased the aroma compound variety and content, thus, enriching their aroma richness and complexity. This study is the first comparative evaluation of the aroma characteristics of different fruit wines fermented with a mixed inoculation of H. uvarum and S. cerevisiae and provides a preliminary guide for these fruit wines produced with non-Saccharomyces yeast.

Evolution of polyphenolic, anthocyanin, and organic acid components during coinoculation fermentation (simultaneous inoculation of LAB and yeast) and sequential fermentation of blueberry wine.

This research aims to investigate the effects of both sequential fermentation and coinoculation fermentation with yeast and lactic acid bacterial (LAB) on the dynamics of changes in basic quality parameters and organic acid, anthocyanin, and phenolic components as well as antioxidant activity during the fermentation of blueberry. The coculture-fermented blueberry wineshowed significant decreases in total phenolics, flavonoids, and anthocyanins,by 23.9%, 15.9%, and 13.7%, respectively, as compared with those before fermentation Fermentation changed the contents of organic acids in each group, with a more than 7-fold increase in lactic acid contents as well as a more than 4-fold reduction in quinic acid and malic acid contents. The content of all investigated anthocyanins first increased and then decreased. Moreover, different fermentation strategies exerted a profound influence on the dynamic change in phenolic components during fermentation; specifically, most of the phenolic acids showed a trend of increasing first, then decreasing, and finally increasing. Gallic acid, p-coumaric acid, quercetin, and myricetin were increased by 116.9%, 130.1%, 127.2% and 177.6%, respectively, while syringic acid, ferulic acid, cinnamic acid, and vanillic acid were decreased by 49.5%, 68.5%, and 37.1% in sequentially fermented blueberry wine. Coinoculation fermentation with yeast and LAB produces faster dynamic variations and higher organic acid, anthocyanin, and phenolic profiles than sequential inoculation fermentation. PRACTICAL APPLICATION: In this work, brewing technology of sequential fermentation and coinoculation fermentation with yeast and LAB (Lactobacillus plantarum SGJ-24 and Oenococcus oeni SD-2a) was adopted to ferment blueberry wine. This is an innovative technology of fruit wine brewing technology to produce wine products. Compared with traditional sequential brewing, simultaneous inoculation brewing can significantly accelerate the brewing process of fruit wine and slightly improve the quality of fruit wine in terms of active ingredients.

Diversity of Volatile Aroma Compound Composition Produced by Non-Saccharomyces Yeasts in the Early Phase of Grape Must Fermentation.

There is a lack of studies evaluating the metabolic contribution of non-Saccharomyces yeasts in early fermentation phases. This study aimed to investigate the volatile aroma profiles produced by various non-Saccharomyces yeasts just before sequential inoculation with Saccharomyces cerevisiae to provide an insight into the particular effects they induce at this stage. The grape must of Malvazija istarska was inoculated with monocultures of Torulaspora delbrueckii, Metschnikowia pulcherrima, Pichia kluyveri, Lachancea thermotolerans, and Schizosaccharomyces pombe, alongside a S. cerevisiae control. Eighty volatile compounds were quantified via headspace solid-phase microextraction and gas chromatography–mass spectrometry, and the data were statistically elaborated. Volatile profiles of non-Saccharomyces yeasts differed significantly from the S. cerevisiae control. Most treatments caused increases in linalool and β-damascenone, decreases in higher alcohols and fatty acids, and improved synthesis of odoriferous esters. Torulaspora delbrueckii and M. pulcherrima produced compounds not commonly found in S. cerevisiae fermented wines. Multivariate statistical analysis linked the investigated yeasts to specific, particularly abundant compounds. Future studies should explore to what degree these contributions persist after sequential inoculation with S. cerevisiae in diverse grape must matrices.

A conserved glutamate residue in RPM1-INTERACTING PROTEIN4 is ADP-ribosylated by the Pseudomonas effector AvrRpm2 to activate RPM1-mediated plant resistance.

Gram-negative bacterial plant pathogens inject effectors into their hosts to hijack and manipulate metabolism, eluding surveillance at the battle frontier on the cell surface. The effector AvrRpm1Pma from Pseudomonas syringae pv. maculicola functions as an ADP-ribosyl transferase that modifies RESISTANCE TO P. SYRINGAE PV MACULICOLA1 (RPM1)-INTERACTING PROTEIN4 (RIN4), leading to the activation of Arabidopsis thaliana (Arabidopsis) resistance protein RPM1. Here we confirmed the ADP-ribosyl transferase activity of another bacterial effector, AvrRpm2Psa from P. syringae pv. actinidiae, via sequential inoculation of Pseudomonas strain Pto DC3000 harboring avrRpm2Psa following Agrobacterium-mediated transient expression of RIN4 in Nicotiana benthamiana. We conducted mutational analysis in combination with mass spectrometry to locate the target site in RIN4. A conserved glutamate residue (Glu156) is the most likely target for AvrRpm2Psa, as only Glu156 could be ADP-ribosylated to activate RPM1 among candidate target residues identified from the MS/MS fragmentation spectra. Soybean (Glycine max) and snap bean (Phaseolus vulgaris) RIN4 homologs without glutamate at the positions corresponding to Glu156 of Arabidopsis RIN4 are not ADP-ribosylated by bacterial AvrRpm2Psa. In contrast to the effector AvrB, AvrRpm2Psa does not require the phosphorylation of Thr166 in RIN4 to activate RPM1. Therefore, separate biochemical reactions by different pathogen effectors may trigger the activation of the same resistance protein via distinct modifications of RIN4.