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This study investigated the potential of endophytic microorganisms in fermentative processes using agro-industrial residues as substrates. The aim was to explore sustainable biotechnological methods for producing valuable compounds from waste. Endophytic microorganisms were isolated from strawberry, lychee, and tangerine and used in fermentative processes with passion fruit, guava, and pineapple residues. Key methods included isolating and screening potent aroma-producing microorganisms, identifying them using MALDI-TOF MS, and analyzing volatile compounds by GC-MS. The results identified 37 endophytic microorganisms, with Kloeckera apiculata presenting the greatest aromatic potential. Fermentation with agro-industrial residues produced significant volatile compounds, identifying 27, 23, and 22 compounds from passion fruit, pineapple, and guava residues, respectively. The main conclusions highlighted the high production of ethyl acetate and 2-phenylethyl acetate, which contributed to fruity and floral aromas. The novelty of this study lies in the use of endophytic microorganisms isolated from tropical fruits to biotransform agro-industrial waste into high-value aromatic compounds, offering economic and environmental benefits. This research is significant, as it proposes a sustainable approach to valorize waste and produce natural aroma compounds through biotechnological processes. [ABSTRACT FROM AUTHOR]

Here, we present the case of an otherwise healthy patient, without risk factors, who developed a refractory case of onychomycosis caused by Kloeckera apiculata, an uncommon human pathogen. The diagnosis was ultimately confirmed by fungal nail plate culture, histopathology, and PCR. Whereas prior treatments with topical 5 % tavaborole solution, oral terbinafine, and oral fluconazole were ineffective, complete clinical and mycological cure was achieved with a 3-month course of oral itraconazole.

This study investigated the potential of endophytic microorganisms in fermentative processes using agro-industrial residues as substrates. The aim was to explore sustainable biotechnological methods for producing valuable compounds from waste. Endophytic microorganisms were isolated from strawberry, lychee, and tangerine and used in fermentative processes with passion fruit, guava, and pineapple residues. Key methods included isolating and screening potent aroma-producing microorganisms, identifying them using MALDI-TOF MS, and analyzing volatile compounds by GC-MS. The results identified 37 endophytic microorganisms, with Kloeckera apiculata presenting the greatest aromatic potential. Fermentation with agro-industrial residues produced significant volatile compounds, identifying 27, 23, and 22 compounds from passion fruit, pineapple, and guava residues, respectively. The main conclusions highlighted the high production of ethyl acetate and 2-phenylethyl acetate, which contributed to fruity and floral aromas. The novelty of this study lies in the use of endophytic microorganisms isolated from tropical fruits to biotransform agro-industrial waste into high-value aromatic compounds, offering economic and environmental benefits. This research is significant, as it proposes a sustainable approach to valorize waste and produce natural aroma compounds through biotechnological processes.

Here, we present the case of an otherwise healthy patient, without risk factors, who developed a refractory case of onychomycosis caused by Kloeckera apiculata , an uncommon human pathogen. The diagnosis was ultimately confirmed by fungal nail plate culture, histopathology, and PCR. Whereas prior treatments with topical 5 % tavaborole solution, oral terbinafine, and oral fluconazole were ineffective, complete clinical and mycological cure was achieved with a 3-month course of oral itraconazole., Competing Interests: There are none., (© 2024 The Authors.)

Biological control is an environmentally friendly, safe, and replaceable strategy for disease management. Genome sequences of a certain biocontrol agent could lay a solid foundation for the research of molecular biology, and the more refined the reference genome, the more information it provides. In the present study, a higher resolution genome of Kloeckera apiculata 34-9 was assembled using high-throughput chromosome conformation capture (Hi-C) technology. A total of 8.07 M sequences of K. apiculata 34-9 genome was anchored onto 7 pesudochromosomes, which accounting for about 99.51% of the whole assembled sequences, and 4,014 protein-coding genes were annotated. Meanwhile, the detailed gene expression changes of K. apiculata 34-9 were obtained under low temperature and co-incubation with Penicillium digitatum treatments, respectively. Totally 254 differentially expressed genes (DEGs) were detected with low temperature treatment, of which 184 and 70 genes were upregulated and downregulated, respectively. Some candidate genes were significantly enriched in ribosome biosynthesis in eukaryotes and ABC transporters. The expression of gene Kap003732 and Kap001595 remained upregulated and downregulated through the entire time-points, respectively, indicating that they might be core genes for positive and negative response to low temperature stress. When co-incubation with P. digitatum , a total of 2,364 DEGs were found, and there were 1,247 upregulated and 1,117 downregulated genes, respectively. Biosynthesis of lysine and arginine, and phenylalanine metabolism were the highest enrichment of the cluster and KEGG analyses of the co-DEGs, the results showed that they might be involved in the positive regulation of K. apiculata 34-9 response to P. digitatum. The completeness of K. apiculata 34-9 genome and the transcriptome data presented here are essential for providing a high-quality genomic resource and it might serve as valuable molecular properties for further studies on yeast genome, expression pattern of biocontrol system, and postharvest citrus storage and preservation. [ABSTRACT FROM AUTHOR]

Biological control is an environmentally friendly, safe, and replaceable strategy for disease management. Genome sequences of a certain biocontrol agent could lay a solid foundation for the research of molecular biology, and the more refined the reference genome, the more information it provides. In the present study, a higher resolution genome of Kloeckera apiculata 34-9 was assembled using high-throughput chromosome conformation capture (Hi-C) technology. A total of 8.07 M sequences of K. apiculata 34-9 genome was anchored onto 7 pesudochromosomes, which accounting for about 99.51% of the whole assembled sequences, and 4,014 protein-coding genes were annotated. Meanwhile, the detailed gene expression changes of K. apiculata 34-9 were obtained under low temperature and co-incubation with Penicillium digitatum treatments, respectively. Totally 254 differentially expressed genes (DEGs) were detected with low temperature treatment, of which 184 and 70 genes were upregulated and downregulated, respectively. Some candidate genes were significantly enriched in ribosome biosynthesis in eukaryotes and ABC transporters. The expression of gene Kap003732 and Kap001595 remained upregulated and downregulated through the entire time-points, respectively, indicating that they might be core genes for positive and negative response to low temperature stress. When co-incubation with P. digitatum, a total of 2,364 DEGs were found, and there were 1,247 upregulated and 1,117 downregulated genes, respectively. Biosynthesis of lysine and arginine, and phenylalanine metabolism were the highest enrichment of the cluster and KEGG analyses of the co-DEGs, the results showed that they might be involved in the positive regulation of K. apiculata 34-9 response to P. digitatum. The completeness of K. apiculata 34-9 genome and the transcriptome data presented here are essential for providing a high-quality genomic resource and it might serve as valuable molecular properties for further studies on yeast genome, expression pattern of biocontrol system, and postharvest citrus storage and preservation.

The antifungal activity against Botrytis cinerea, a pathogen of grape gray mold disease, was validated in vitro for 59 of 225 wild yeast strains isolated from naturally fermented wine samples (69 strains), our university campus plants (129 strains), and commercially available fruits (27 strains). Twenty strains showing strong antifungal activity were identified. The majority of the strains were Saccharomyces cerevisiae (16 of the 20 strains). The S. cerevisiae strains were differentiated by genetic polymorphism analyses with comparisons of their antifungal activities. As a result, it was clarified that each strain type showed different antifungal activity, and that automatic rRNA intergenic spacer analysis was extremely useful for the differentiation of S. cerevisiae strains when compared to other conventional polymerase chain reaction-based methods. [ABSTRACT FROM AUTHOR]

Lineage-specific genes (LSGs) are defined as genes with sequences that are not significantly similar to those in any other lineage. LSGs have been proposed, and sometimes shown, to have significant effects in the evolution of biological function. In this study, two sets of Hanseniaspora spp. LSGs were identified by comparing the sequences of the Kloeckera apiculata genome and of 80 other yeast genomes. This study identified 344 Hanseniaspora -specific genes (HSGs) and 109 genes ('orphan genes') specific to K. apiculata. Three thousand three hundred thirty-one  K. apiculata genes that showed significant similarity to at least one sequence outside the Hanseniaspora were classified into evolutionarily conserved genes. We analyzed their sequence features, functional categories, gene origin, gene structure and gene expression. We also investigated the predicted cellular roles and Gene Ontology categories of the LSGs using functional inference. The patterns of the functions of LSGs do not deviate significantly from genome-wide average. The results showed that a few LSGs were formed by gene duplication, followed by rapid sequence divergence. Many of the HSGs and orphan genes exhibited altered expression in response to abiotic stress. Studying these LSGs might be helpful for understanding the molecular mechanism of yeast adaption. [ABSTRACT FROM AUTHOR]

Graphical abstract Highlights • P. membranaefaciens and K. apiculata were effective in control of brown rot in plum. • P. membranaefaciens and K. apiculata were well-adapted to the wound environment. • P. membranaefaciens and K. apiculata could induce plant disease resistance. • P. membranaefaciens and K. apiculata could induce phenylpropanoid pathway in plum. Abstract Strains belonging to the species Pichia membranaefaciens and Kloeckera apiculata were tested as biocontrol agents (BCAs) against brown rot caused by Monilinia fructicola in plum fruit. The two yeast strains demonstrated significant antifungal activities at concentration of 5 × 108 cells/mL. Effective colonization in the wounds and high population size to compete for nutrition and space with pathogen contributed to the control mechanism of P. membranaefaciens and K. apiculata. Besides, P. membranaefaciens and K. apiculata could activate the phenylpropanoid pathway of plum to strengthen its cell wall and prevent pathogenic invasion. Both of the yeasts significantly accelerated activities of related enzymes such as phenylalanine ammonia lyase, cinnamic acid 4-hydroxylase, p-coumaroyl: CoA ligase and peroxidase. Those enzymes enhanced biosynthesis of some active metabolites of plum such as lignin, flavonoid and total phenolics. [ABSTRACT FROM AUTHOR]

The use of antagonistic yeasts to control postharvest pathogens is a promising alternative to fungicides. The effect of the yeast antagonists Pichia membranaefaciens and Kloeckera apiculata for controlling brown rot of plums in postharvest and the antifungal activity of the two yeast strains in in vitro conditions against Monilinia fructicola were investigated. In in vivo trials, both P. membranaefaciens and K. apiculata showed a significant reduction of rot incidence produced by M. fructicola by 76.0% and 65.8%, respectively. In in vitro trials, spore germination and mycelial growth of M. fructicola were markedly inhibited by P. membranaefaciens and K. apiculata . In addition, the two antagonistic yeasts, especially K. apiculata , had attachment to the hyphae of M. fructicola . Moreover, P. membranaefaciens and K. apiculata secreted hydrolytic enzymes, such as chitinase (CHI) and β -1, 3-glucanase (GLU). According to these results, P. membranaefaciens and K. apiculata might have the ability to parasitize M. fructicola . The volatile organic compounds (VOCs) produced by P. membranaefaciens and K. apiculata had a significant inhibitory effect on M. fructicola in vitro and in vivo. VOCs in a simultaneous incubation (SI) trial showed the best inhibition. [ABSTRACT FROM AUTHOR]

Kloeckera apiculata, as the anamorphic state of Hanseniaspora uvarum from the Ascomycota phylum, plays an important role in the inhibition of fungal diseases in plants and spontaneous wine fermentation. This study was performed to sequence and analyze the whole genome of K. apiculata strain 34-9; This analysis provides further genomic features and assists functional research. The complete genome was determined using an Illumina HiSeq 2000 system applying paired-end and mate-pair methods to construct four reads libraries. The data assembly of all the reads resulted in a total genome size of 8.1 Mb, including 106 contigs, which were assembled into 41 scaffolds with a 31.95 % G+C content and a 234X sequence coverage. The performance of the gene prediction and functional annotation revealed that 2724 of 3786 protein-coding genes matched the KOG database, and 1127 genes were classified into GO categories. Further genome features analyses found 1066 microsatellite sites, 71 tRNAs, 3 rRNAs and 3 microRNAs in the genomic DNA. A prediction of the metabolic pathways identified potentially crucial genes for explaining the phenylalanine pathway involved in biocontrol. Comparisons with the typical yeasts Lachancea thermotolerans, Kluyveromyces lactis and Saccharomyces cerevisiae revealed the particularity and difference of K. apiculata strain 34-9. The genome alignments among Hanseniaspora vineae T02/19AF, K. apiculata DSM 2768 and 34-9 indicated numerous homologous regions distributed over the genomes between strain DSM2768 and 34-9. A SSR analysis identified that mono- and tri- nucleotide repeat types were more abundant in all six types, likely affecting the evolution of K. apiculata. [ABSTRACT FROM AUTHOR]

The current work examines the environmental conditions to control the expression of self-flocculation of Kloeckera apiculata mc1 isolated from Argentinian grapes with proven beneficial effect on wine aroma. Temperature had a direct effect on the expression of flocculation resulting in an increase in the number of zymolectin sites determined with a BSA-Gal-FITC probe and quantified with the Scatchard plot. Agitation had a positive effect on yeast flocculation and the effect did not correlate with an increase in the hydrophobic index. The addition of glucose, fructose or ethanol to yeast nitrogen base medium without amino acids or ammonium sulphate stimulated flocculation. Assimilative nitrogen sources had a negative impact on flocculation in the absence of an energy source. During winemaking, control of flocculation by environmental factors such as temperature and the presence of nutrients could be applied in order to optimize the fermentation process and hence the quality of the final product. Knowledge of the effect of different parameters on flocculation of the wine yeast K. apiculata mc1 allows prediction of the behaviour of this yeast during fermentation.

The behaviour of the flocculent Kloeckera apiculata strain in coculture with the nonflocculent Saccharomyces cerevisiae strain, both yeasts isolated from wine, has been analyzed. Pure culture of Kloeckera apiculata (apiculate yeast) exhibits 75 % flocculation in the yeast extract/peptone/glucose (containing 20 g/L of glucose, YPG20) medium after 24-hour incubation at 28 °C. This yeast expresses a weak flocculent phenotype in glucose-poor medium (10 % of flocculated cells). Increasing the glucose concentrations in this medium induces yeast flocculation. When the apiculate yeast was co-inoculated with a nonflocculent strain of Saccharomyces cerevisiae (elliptic yeast), an increase of the number of elliptic cells that settled at the bottom of the culture was observed. Electron microscopy observations of the aggregates formed in the mixed culture confirmed that apiculate and elliptic yeasts can interact and establish a binding between them through homogeneous mucus. However, a lower percentage of Kloeckera apiculata flocculation with respect to the pure cultures was observed, this behaviour being correlated with a higher rate of glucose consumption by Saccharomyces cerevisiae. The prompt coflocculation of Kloeckera apiculata with Saccharomyces strains and the induction of this phenomenon by glucose could be considered an important biotechnological tool for the early decreasing of indigenous saccharomycetic flora from the media, before the inoculation of a selected starter strain to carry out a more controlled alcohol fermentation.

Background: The yeast Kloeckera apiculata strain 34-9 is an antagonist that shows biological control activity against the postharvest fungal pathogens of citrus. An antifungal compound, 2-phenylethanol (PEA), has been identified from the extract of K. apiculata. To better understand the molecular processes underlying the response of citrus fruit tissue to K. apiculata, the extract and PEA, microarray analyses were performed on navel oranges using an Affymetrix Citrus GeneChip. Results: As many as 801, 339 and 608 differentially expressed genes (DEGs) were identified after the application of K. apiculata, the extract and PEA, respectively. In general, K. apiculata induced the expression of defence-related genes. In addition to chitinase and β-1,3-glucanase, genes involved in ethylene (ET), jasmonic acid (JA), calcium signalling, MAPK signalling and phenylalanine metabolism were induced. In contrast, monodehydroascorbate reductase, superoxide dismutase (SOD), catalase (CAT), peroxidase (POD) and carotenoid biosynthesis genes were down-regulated. The expression profiles for the extract- and PEA-treated samples were similar to that found for yeast (sharing 57.4 % DEGs), with a significant increase in the transcript levels of defence-related genes. Conclusion: This study provides a global picture of the gene expression changes in navel oranges after the application of the antagonist yeast K. apiculata, its extract and PEA. The interpretation of the DEGs revealed new insight into the molecular processes that regulate the defence responses in orange tissue. [ABSTRACT FROM AUTHOR]

• K. apiculata reduced the colony diameter and mycelial weight of P. expansum. • Yeast parasitized on the mycelial, and culture showed maximum inhibition. • Yeast reduced the decay incidence and lesion diameter of apples. • Yeast roles involve nutrient and space competition, parasitism, induced resistance. Kloeckera apiculata is a yeast strain with excellent biocontrol capacity. The strain can effectively control a variety of postharvest diseases in fruit. However, the effect of the yeast control on blue mold caused by Penicillium expansum in apples and its related mechanisms have rarely been reported. In this study, the colony diameter and mycelial weight of P. expansum were reduced with varied doses of K. apiculata treatment. Compared with controls, the colony diameter and weight of P. expansum mycelial treated with 1 × 108 cell/mL of yeast were 63 % and 70 % lower, respectively. The original yeast culture treatment exhibited the highest inhibition, followed by the culture filtrate, while inactivated culture filtrate displayed no significant inhibitory effect. Results from scanning electron microscopy demonstrated that yeast parasitizes mycelial surfaces, thereby reducing survival and damaging mycelial integrity. Moreover, yeast rapidly colonized apple wounds, reducing decay and lesion diameter. The yeast treatment upregulated the activity of superoxide dismutase, catalase, peroxidase, polyphenol oxidase, and phenylalanine ammonia-lyase, and increased total phenols, flavonoids, and lignin content of the fruit. K. apiculata may directly inhibit the growth of P. expansum through competition for nutrients and space, and via parasitism. The yeast could also induce resistance against P. expansum by eliciting antioxidant enzymes and phenylpropanoid metabolism. [ABSTRACT FROM AUTHOR]

Background Green and blue mold decay, caused by Penicillium digitatum and P. italicum, respectively, are important postharvest diseases of citrus. Biocontrol by microbes is an alternative to synthetic fungicide application. In this study, the antagonistic yeast strain Kloeckera apiculata 34-9 was used to investigate the action mechanisms involved in the biocontrol of postharvest diseases. Results An antifungal substance, 2-phenylethanol (PEA), was isolated from K. apiculata and demonstrated to have antimicrobial activity against selected phytopathogenic fungi. Experiments on P. italicum cells identified the mitochondria and the nucleus as particularly sensitive to inhibition. Regulation of P. italicum gene expression was investigated using RNA-Seq. PEA up-regulated genes involved with the peroxisome, regulation of autophagy, phosphatidylinositol signaling system, protein processing in endoplasmic reticulum, fatty acid metabolism, and inhibited ribosome, RNA polymerase, DNA replication, amino acid biosynthesis, aminoacyl-tRNA biosynthesis and cell cycle. Inhibitory responses revealed by RNA-Seq suggest that PEA might compete for attachment on the active site of phenylalanyltRNA synthetase (PheRS). Conclusion This study provided new insight on the mode of action of biocontrol yeast agents in controlling postharvest pathogenic fungi. [ABSTRACT FROM AUTHOR]

Highlights: [•] Antagonistic yeast was a safe and effective measure for postharvest diseases. [•] Effects of antagonist on fruit wax production and gene expression were evaluated. [•] Esters, fatty acids and fatty alcohols changed in antagonist treatment. [ABSTRACT FROM AUTHOR]

Se ha estudiado la influencia de la aireación en la actividad fermentativa de Kloeckera apiculata RIVE 9-2-1 con el objetivo de evaluar la producción de metabolitos de la fermentación. Para ello, la cepa se cultivó en frascos Erlenmeyer conteniendo jugo de manzana esterilizado y sin aroma, y los compuestos químicos producidos durante la fermentación en cultivo agitado (200 min-1) y estático (sin agitación) se determinaron. Los resultados mostraron que la agitación del medio de cultivo incrementa la producción de alcoholes superiores (hasta 591,0 mg/L) comparado al cultivo estático, mientras que por el contrario, la producción de ácido acético, etil acetato y glicerol (260,0 ± 11,0 mg/L, 196,0 ± 10,0 mg/L y 2,6 ± 0,2 g/L) resultaron ser mayores que en cultivo agitado (222,0 ± 8,0 mg/L, 96,0 ± 4,5 mg/L y 1,8 ± 0,2 g/L) respectivamente. Cultivos bacth realizados en biorreactor con un flujo de aire de 25 L/h reportaron una tasa de crecimiento (µ) de 0,17 h-1, producción de etanol (12,5 ± 2,0 g/L) y otros compuestos típicamente producidos durante la fermentación alcohólica. La concentración de oxígeno disuelto en el medio de fermentación afecta su metabolismo así, cantidades insuficientes de oxígeno provocaría un metabolismo respirofermentativo. Los mejores resultados en términos de calidad organoléptica de la bebida fermentada en lo referente al aroma, sabor y olor se obtuvieron en la fermentación en cultivo estático. El control de la aireación del medio de fermentación puede ser usado para controlar la síntesis de compuestos químicos de impacto sensorial en la producción de bebidas fermentadas.

Highlights: [•] The evaluated yeast is capable of reducing the degree of blue mold decay of citrus. [•] The yeast competes with the phytopathogen for nutrients. [•] Vitamins are necessary for biological biocontrol of Kloeckera apiculata. [Copyright &y& Elsevier]

The current work examines the environmental conditions to control the expression of self-flocculation of Kloeckera apiculata mcl isolated from Argentinian grapes with proven beneficial effect on wine aroma. Temperature had a direct effect on the expression of flocculation resulting in an increase in the number of zymolectin sites determined with a BSA-Gal-FITC probe and quantified with the Scatchard plot. Agitation had a positive effect on yeast flocculation and the effect did not correlate with an increase in the hydrophobic index. The addition of glucose, fructose or ethanol to yeast nitrogen base medium without amino acids or ammonium sulphate stimulated flocculation. Assimilative nitrogen sources had a negative impact on flocculation in the absence of an energy source. During wine-making, control of flocculation by environmental factors such as temperature and the presence of nutrients could be applied in order to optimize the fermentation process and hence the quality of the final product. Knowledge of the effect of different parameters on flocculation of the wine yeast K. apiculata mcl allows prediction of the behavior of this yeast during fermentation. [ABSTRACT FROM AUTHOR]

Abstract: The aim of this study was to determine the influence of different yeasts isolated from fresh blue plum fruits (Aureobasidium sp.) and spontaneously fermenting plum musts (Kloeckera apiculata and Saccharomyces cerevisiae), as well as commercial wine and distillery strains, on the fermentation and chemical composition of plum brandies. Gas chromatography methods were used to detect major volatile components. The most rapid fermentation occurred in musts inoculated with S. cerevisiae. However, the highest concentration of ethanol was detected in samples after spontaneous fermentation (8.40% v/v). Plum brandies obtained after distillation contained from 66.3 (K. apiculata) up to 74.3% v/v ethanol (spontaneous fermentation). The samples after spontaneous fermentation were distinguished by a high content of acetoin, ethyl acetate and total esters, accompanied by a low level of methanol and fusel alcohols. Non-Saccharomyces yeasts were responsible for higher concentrations of esters and methanol, while S. cerevisiae strains resulted in increased levels of higher alcohols. It was also found that isolated indigenous strains of S. cerevisiae synthesized relatively low amounts of higher alcohols compared to commercial cultures. Samples obtained using the distillery strain of S. cerevisiae received the highest score (18.2) during sensory analysis and were characterized by a well-harmonised taste and aroma. [Copyright &y& Elsevier]

The behaviour of the flocculent Kloeckera apiculata strain in coculture with the nonflocculent Saccharomyces cerevisiae strain, both yeasts isolated from wine, has been analyzed. Pure culture of Kloeckera apiculata (apiculate yeast) exhibits 75 % flocculation in the yeast extract/peptone/glucose (containing 20 g/L of glucose, YPG20) medium after 24-hour incubation at 28 °C. This yeast expresses a weak flocculent phenotype in glucose-poor medium (10 % of flocculated cells). Increasing the glucose concentrations in this medium induces yeast flocculation. When the apiculate yeast was co-inoculated with a nonflocculent strain of Saccharomyces cerevisiae (elliptic yeast), an increase of the number of elliptic cells that settled at the bottom of the culture was observed. Electron microscopy observations of the aggregates formed in the mixed culture confirmed that apiculate and elliptic yeasts can interact and establish a binding between them through homogeneous mucus. However, a lower percentage of Kloeckera apiculata flocculation with respect to the pure cultures was observed, this behaviour being correlated with a higher rate of glucose consumption by Saccharomyces cerevisiae. The prompt coflocculation of Kloeckera apiculata with Saccharomyces strains and the induction of this phenomenon by glucose could be considered an important biotechnological tool for the early decreasing of indigenous saccharomycetic flora from the media, before the inoculation of a selected starter strain to carry out a more controlled alcohol fermentation. [ABSTRACT FROM AUTHOR]

We have evaluated the induction of the flocculent phenotype of Kloeckera apiculata by glucose mc1 and propose a pathway involved in carbohydrate flocculation induction. Pulses of glucose were given to cells growing in glucose-poor medium (2 g l−1) and the flocculation percentage was measured. To elucidate the mechanism involved in flocculation induction, cycloheximide was injected into the cultures 120 min before the glucose pulse. 2,4-Dinitrophenol or cAMP was added to the media instead, or simultaneously with glucose, while a protein kinase A (PKA) inhibitor was added 30 min before the glucose pulse. With 20 and 50 g l−1 glucose pulse, the yeast flocculation percentage arises to 55 and 65%, respectively. The quantity of proteins and the reflocculating capacity of a lectinic protein extract from the yeast cell wall increase as the concentration of glucose pulse was higher. Cycloheximide prevented the glucose-induced flocculation, while cAMP or 2,4-dinitrophenol increased it 4- and 5-fold, respectively. PKA inhibitor completely prevented the glucose induction flocculation. The flocculent phenotype of K. apiculata mc1 was induced by glucose and the mechanism seems to imply de novo protein (lectin) synthesis via the PKA transduction pathway. This work contributes to the elucidation of the mechanism involved in flocculation induction by glucose of a non- Saccharomyces wine yeast, K. apiculata, which has not been reported. The induction of flocculation by glucose could be a biotechnological tool for the early removal of the indigenous microorganisms from the grape must before the inoculation of a selected starter strain to conduct the alcohol fermentation. [ABSTRACT FROM AUTHOR]

The efficacy of the yeast Kloeckera apiculata, strain 34-9, in controlling postharvest decay of citrus fruit was evaluated in small-scale and pilot tests in commercial packinghouse. Kloeckera apiculata grew efficiently on different media and maintained its antagonistic activity against spore germination of Penicillium italicum. In small-scale experiments with citrus fruits dipped in the yeast cell suspension, the development of decay in citrus was effectively inhibited. The yeast was compatible with a mixture of low concentration of a commonly chemical fungicide. In packinghouse tests, combining the yeast with 40 mg/kg Carbendazim (MBC) resulted a reduction in the incidence of decay to a level equal to that of the commercial treatment of 200 mg/kg MBC. The efficacy of the strain 34-9 could also be maintained under packinghouse conditions at a cell concentration of the yeast antagonist as low as 106 cells/ml. No significant difference in the efficacy of K. apiculata was found in either the drench or the spray application systems tested in citrus packinghouse. Scanning electron microscopy revealed attachment of the yeast cells to the pathogen hyphae. The high antagonistic activity of strain 34-9 against citrus blue mould may be related to its capability to compete with Penicillium italicum, for space and nutrients and /or involvement of directly antagonist of the yeast on the fungus. [ABSTRACT FROM AUTHOR]

Abstract Aims: To characterize and analyze the flocculation phenomenon of Kloeckera apiculata mc1 from Agentinian wine to understand the cell–cell interaction pattern. Methods and Results: Kloeckera apiculata mc1 possess intense cell–cell interactions in MYPG medium (0·5% malt extract, 1% yeast extract, 2% glucose, 2% peptone), pH 5·5 by shaking at 25°C. Optimum flocculation is observed at pH 4·5 in the presence of 3 mmol l-1 Ca2+ . The flocculation is induced by peptone and malt extract and not by yeast extract and is reversed by 50 mmol l-1 galactose or lactose. The flocculation is highly susceptible to pronase, chymotrypsine and proteases types IV and XXVII and is partially resistant to trypsin. The electronic microscopy shows that the cells are attached to each other along their sides by fine hair-like threads. Conclusions: The mechanism of flocculation of K. apiculata mc1 is mediated by protein–carbohydrate interaction, stabilized by Ca2+ . Significance and Impact of the Study: The use of selected pure yeast inocula of known ability is preferred to wine elaboration, therefore the indigenous flora must be avoided and the flocculation of K. apiculata could be an economic method to do it. [ABSTRACT FROM AUTHOR]

Abstract Biocontrol activities of a total of 103 yeast isolates were tested against postharvest diseases of peaches. Seven isolates, with the best efficacy in reducing the number of infected wounds or lesion sizes of Penicillium expansum and Botrytis cinerea , were selected for further large-scale experiments. In large-scale experiments, all selected isolates were significantly effective (P ≤ 0.05) in reducing the number of infected wounds and the diameter of lesions caused by P. expansum , B. cinerea , and Monilinia fructicola . DR52 was significantly superior to all the other yeasts in effectiveness against all three pathogens. The efficacy of the other yeast antagonists against B. cinerea and P. expansum was almost equal, while the control of M. fructicola was inferior. DR52 was selected for further storage experiment because its efficacy was higher against the three pathogens and it also showed a different random amplified polymorphic DNA-polymerase chain reaction pattern compared with other isolates. DR52 was identified by Centraalbureau voor Schimmeelcultures (Baarn, The Netherlands) as Kloeckera apiculata . K. apiculata completely controlled both pathogens after 30 days of storage. Its efficacy declined to an 83.4% reduction in B. cinerea incidence and an 87.5% reduction in P. expansum incidence after 45 days of storage. [ABSTRACT FROM AUTHOR]

The behaviour of Kloeckera apiculata, Candida pulcherrima and Saccharomyces cerevisiae was studied with pure and mixed cultures in grape juice. The non-Saccharomyces species showed significant growth and survival in mixed cultures, although S. cerevisiae dominated the fermentation. Ethanol production was higher with S. cerevisiae in both cultures. K. apiculata and C. pulcherrima formed the lowest amounts of ethanol with high residual sugars and the highest contents of ethyl acetate in pure cultures. The concentrations of ethyl acetate produced in mixed cultures were higher than those produced using only S. cerevisiae. Moreover, other volatile compounds were not formed in undesirable levels in mixed cultures. [Copyright &y& Elsevier]

A rapid and efficient analytical method for the determination of glycerol in wines is described. This method utilizes high-performance thin layer chromatography (HPTLC) plates coupled with an automated multiple development system with an elution gradient based on acetonitrile–acetone–hexane on silica gel layers. The absence of clean-up procedures, sometimes only centrifugation, makes this method suitable also for the large-scale control of alcoholic beverages. In particular the capacity of different wine yeast species ( Saccharomyces cerevisiae, Zygosaccharomyces bailii, Kloeckera apiculata and Saccharomycodes ludwigii) to produce glycerol was determined. Generally, the strains of S. cerevisiae produced elevated amounts of glycerol together with Z. bailii, whereas K. apiculata strains formed the lowest amounts of glycerol, exhibiting also a great strain variability. [ABSTRACT FROM AUTHOR]

One of the important factors affecting wine fermentation is temperature. The influence of elevated temperatures from 10 to 25 °C at 5 °C intervals on yeast growth and fermentation products were studied in mixed cultures of Kloeckera apiculata and Saccharomyces cerevisiae in grape juice. In the experiments carried out at 10 and 15 °C, K. apiculata grew and survived longer compared to trials conducted above 20 °C. In most cases, higher temperatures stimulated the production of higher alcohols but lowered the formation of esters and acetaldehyde. [ABSTRACT FROM AUTHOR]

This work deals with biogenic amine production by yeast strains isolated from grapes and wines. A total of 50 strains were tested for their capacity to produce biogenic amines in wine. In general, all the species produced very low or non-detectable amounts of histamine, whereas methylamine and agmatine were formed by all the species considered. The highest concentration of total biogenic amines was formed by Brettanomyces bruxellensis, with an average value of 15 mg/l, followed by Saccharomyces cerevisiae with an average of 12.14 mg/l. The other species formed less than 10 mg of total biogenic amines per litre. Wines fermented with the most fermentative strains of S. cerevisiae species had the highest contents of ethanolamine, from 2.3 to 16 mg/l, and of agmatine, from 3.1 to 7.5 mg/l. The strains of the other species, which exhibited a low fermentative ability, Kloeckera apiculata, B. bruxellensis and Metschnikowia pulcherrima, varied in the production of agmatine and phenylethylamine. A significant variability in the production of cadaverine was characteristic of Candida stellata strains, which varied also in ethanolamine production. Our results emphasize the importance of using selected strains of S. cerevisiae, not only for the expression of desirable technological traits, but also to avoid potentially negative effects on human health. Therefore, the characterization of strains of S. cerevisiae for the 'production of biogenic amines' becomes of applicative interest. [ABSTRACT FROM AUTHOR]

Forty-nine strains of Kloeckera apiculata, isolated from the Friuli region in Italy, were differentiated on the basis of fermentation behaviour and production of secondary compounds in two different grape musts at 18 °C. The isolates exhibited a controlled production of acetic acid, only in a few cases more that 1 g/l. In Moscato grape must the strains exhibited a more uniform behaviour for the production of higher alcohols, ethyl acetate and acetoin than in red grapes. In general, higher levels of ethanol, glycerol and acetic acid were produced in red grape must fermentation. Apiculate strains behaved differently in the two musts, with different metabolic phenotypes dominating the fermentation process. The existence of different metabolic phenotypes correlated with the must composition underlines the need to perform a selection of indigenous apiculate yeasts to obtain the desired consistent products. [ABSTRACT FROM AUTHOR]

The (R)/(S) ratios of acetoin were always higher in wines obtained by Saccharomyces cerevisiae than in those obtained by Kloeckera apiculata. A significantly different behaviour was determined between the two species as regards contents and ratios of 2,3-butanediols: S. cerevisiae produced more (R,R)-2,3-butanediol (about 80%), whereas K. apiculata produced more meso-form (about 90%). [ABSTRACT FROM AUTHOR]

The article presents a study which analyzed aroma compounds of wines resulting from fermentation of sterile grape musts from Monastrell variety inoculated with pure and mixed cultures of apiculate and Saccharomyces yeasts. Gas chromatography with flame ionization and mass spectrometry were used. The initial presence of apiculate yeasts in fermentations improved the production of some compounds and in general, increased the higher alcohol and acid total amounts.

A total of 37 strains of Kloeckera apiculata was isolated during the spontaneous fermentation of star fruit must. Each strain was differentiated from the others on the basis of its capacity to produce acetaldehyde, ethyl acetate, higher alcohols, acetoin and acetic acid. All the strains were characterized by the low production of higher alcohols and the high production of ethyl acetate, whereas consistent differences in the production of acetaldehyde, acetoin and acetic acid served to differentiate star fruit apiculate strains into six different phenotypes, present at different stages of the fermentation process. The metabolic strain diversity found can be interpreted as a natural consequence of environmental conditions, which influenced the frequency and selection of specific apiculate strains. From the biotechnological point of view the different metabolic biotypes represent an important source of strains for potential use as starter cultures for star fruit fermentation. [ABSTRACT FROM AUTHOR]

The capacity to produce 2,3-butanediol by 90 strains of four different species of wine yeasts (Kloeckera apiculata, Saccharomyces cerevisiae, Saccharomycodes ludwigii, Zygosaccharomyces bailii) was tested in grape must by automated multiple development HPTLC. The total amount of 2,3-butanediol produced varied between 23mg l−1 and 857.7mg l−1 according to the yeast species. S. cerevisiae and Z. bailii behaved similarly, producing elevated amounts of 2,3-butanediol. K. apiculata and Sc. ludwigii, in contrast, were low producers. When considerable amounts of 2,3-butanediol were found, little acetoin was present; the amounts of butanediol and acetoin were characteristic of the individual species. [ABSTRACT FROM AUTHOR]

Partially (6-fold) purified plasma membrane ATPase from an ethanol-sensitive yeast, Kloeckera apiculata, had an optimum pH of 6.0, an optimum temperature of 35°C, a K of 3.6 m m ATP and a V of 11 μmol P/min.mg protein. SDS-PAGE of the semi-purified plasma membrane showed a major band of 106 kDa. No in vivo activation of the ATPase by glucose was observed. Although 4% (v/v) ethanol decreased the growth rate by 50% it did not affect the ATPase. Concentrations of ethanol ≥2% (v/v) did, however, inhibit the enzyme in vitro. The characteristics of the enzyme did not change during growth in the presence of ethanol. [ABSTRACT FROM AUTHOR]

Apple pomace was used in solid-substrate fermentation with the yeasts Kloeckera apiculata or Candida utilis Y15. A total crude protein content of 7.5% (w/w) was achieved after 72 h for each yeast. The concentration of essential amino acids in the modified apple pomace was more than twice that in the control, enhancing its nutritive value as a stock-feed supplement. The fermentation of pomace as described can be used to reprocess this waste material into a useful value-added product for the agricultural sector. [ABSTRACT FROM AUTHOR]