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Kluyveromyces marxianus and Saccharomyces cerevisiae yeasts were compared for the production of Carboxypeptidase Y, a serine class glycoprotein exopeptidase (CPD-Y) using Bio calorimetry and Respirometric devices. The production medium components were optimized by Response Surface Methodology-based design of experiments. Bio calorimetric investigation combined with Respirometric studies suggested that the rate of heat evolution and respiration rate was directly linked to Dry Cell Weight (DCW), enzyme activity, and the consumption rate of the substrate. The Maximum yield obtained for K. marxianus and S. cerevisiae was 14 IU/mL and 8 IU/mL respectively which were threefold higher than the shake flask experiment. Yeast growth and nature of growth predicted using calorespirometric data fitted well with measured experimental results.

Efficient bioethanol production from lignocellulosic biomass requires thermotolerant yeasts capable of utilizing multiple sugars, tolerating inhibitors and fermenting at high temperatures. In this study, 98 thermotolerant yeasts were isolated from nine compost samples. We selected 37 yeasts that belonged to 11 species; 31 grew at 45°C; 6 strains grew at 47°C, while 9 yeasts could utilize multiple sugars. Many yeast isolates showed high ethanol production in the range of 12-24 g l-1 , with fermentation efficiencies of 47-94% at 40°C using 5% glucose. Kluyveromyces marxianus CSV3.1 and CSC4.1 (47°C), Pichia kudriavzevii CSUA9.3 (45°C) produced 21, 22 and 23 g l-1 of ethanol with efficiencies of 83, 87 and 90%, respectively, using 5% glucose. Among these yeasts, K. marxianus CSC4.1 and P. kudriavzevii CSUA9.3 exhibited high tolerance against furfural, 5-HMF, acetic acid and ethanol. These two strains produced high amounts of ethanol from alkali-treated RS, with 84 and 87% efficiency via separate hydrolysis and fermentation; 76 and 74% via simultaneous saccharification and fermentation at 47 and 45°C, respectively. Therefore, this study demonstrates compost as a potential anthropogenic niche for multiple sugar-utilizing, inhibitor-tolerant ethanologenic yeasts suitable for high-temperature ethanol production via SHF of rice straw.

Xylose is the second most abundant sugar in lignocellulosic biomass and on the planet. In the second-generation bioethanol production process, it is essential to metabolize all sugars obtained in the hydrolysis of lignocellulose to increase ethanol titers and conversion yields of biomass to biofuel and decrease production costs. Wild-type Saccharomyces cerevisiae strains, the workhorse microorganisms in ethanol production, are not able to ferment xylose. In this review, a comparison in bioethanol production from xylose was carried out between non-conventional yeasts and recombinant strains of Saccharomyces, highlighting the advantages and disadvantages according to the most important factors that influence sugar metabolism to ethanol, such as the assimilation pathway used, co-assimilation with glucose, oxygen levels, tolerance to fermentation inhibitors, and process temperature, among others. The review begins reviewing the metabolic pathways of xylose assimilation, followed by the associated cofactor imbalance, heterologous expression of xylose isomerase (XI), regulation of xylose reductase (XR) and xylitol dehydrogenase (XDH), and the relevance of genes involved in the pentose phosphate pathway (PPP). Likewise, the importance of other yeasts, such as Scheffersomyces stipitis, Candida shehatae, and Kluyveromyces marxianus, as non-conventional yeasts to produce bioethanol is highlighted, due to their ability to ferment a wide range of sugars, including xylose, compared to S. cerevisiae.

Enzyme engineering toward catalytic-tetrad residues usually results in activity loss. Unexpectedly, we found that a directed evolution campaign yielded a beneficial residue A100 in KmCR (a carbonyl reductase from Kluyveromyces marxianus ZJB14056), which is a residue of catalytic tetrad and conserved according to multiple sequence alignment. Inspired by this finding, we performed saturation mutagenesis on all the four residues of catalytic tetrad of KmCR. A number of variants with improved enzymatic activities were obtained. Among them, the variant KmCR_A100S exhibited increased catalytic efficiency (kcat /KM = 47.3 s-1 ·mM-1 ), improved stereoselectivity (from moderate selectivity (deP = 66.7%) to strict (S)-selectivity (deP > 99.5%)), and extended substrate scope, compared to those of KmCR_WT. In silico analysis showed that a relay system was rebuilt in KmCR via the beneficial residue S100. Furthermore, comparison of 11 protein engineering campaigns indicated that the beneficial position is easily overlooked due to the long distance (>10 A) from ketone substrates. Since CRs share similar catalytic mechanism, the knowledge gained from this study has universal significance to CR engineering.

Ethanol tolerance is one of the main limiting factors for scaling up its production. Bioprocess control can be performed by knowing the ethanol effects on cell growth, since it is a product associated with microbial growth and, therefore, its synthesis depends on the specific cell growth rate (µ). In this study, mathematical modeling based on the integration of effects was used to develop an ethanol inhibition model for Kluyveromyces marxianus CCT 7735 (UFV-3), at different ethanol concentrations, using glucose as a limiting substrate. A modified Monod model based on studies by Ghose & Tyagi was designed. This model was able to describe with high accuracy the µ parameter behavior as a function of sub-inhibitory glucose concentrations (R²=0.992), and inhibitory ethanol conditions (R²=0.999). The proposed model presented µmax = 0.673 h− 1, KS= 0.997 g.L− 1, and Pcrit= 6.78 % (v/v), whose validation had a mean error of 7.43 %. The µ parameter was mainly influenced by ethanol, showing a linear reduction with the ethanol increase in the culture medium, while glucose concentrations above 20 g.L− 1 had little influence on its value. Therefore, using a simple methodology, an efficient ethanol tolerance model was proposed, which can contribute to future studies of large-scale fermentation using K. marxianus strains.

Ergosterol, an important pharmaceutical intermediate, is the precursor of liposoluble vitamin D2 and cortisone. It is also a main sterol in yeast cells and responsible for structural features of membranes such as the integrity, fluidity, permeability and activity of membrane-bound enzymes. Kluyveromyces marxianus is able to utilize various sugars such as lactose, xylose and arabinose against Saccharomyces cerevisiae and is also thermotolerant. Based on these aforementioned characteristics, K. marxianus can be of great importance in the utilization of whey and lignocellulosic biomass. In this paper, the effect of four factors on the specific ergosterol content and yeast growth was investigated using two statistical experimental designs. The factors examined were initially added alcohol, temperature, salt concentration and pH. The initially added alcohol had a positive effect on the specific ergosterol content, resulted in 37 % specific ergosterol content increasement. The temperature had a negative effect on yeast growth reducing the biomass concentration by 50 % when increased from 25 °C to 30 °C. The pH had a significant effect only on the specific ergosterol content, having an optimum at pH 5.5. The salt concentration had no significant effect in either case. Based on the results, it is suggested that the setup which facilitates higher ergosterol content but does not slow down the growth of the yeast remarkably should be selected, which are 25 °C, pH 5.3 and 3 % of initial ethanol content.

Technology valuation of cellulosic ethanol production by Kluyveromyces marxianus CCT 7735 based on the concepts of circular economy was investigated. After an acid-base pretreatment followed by saccharification at 50 °C for 72 h, the sweet sorghum hydrolysate was used to produce ethanol at three temperatures: 37, 42, and 45 °C. About 17.83 g/L of cellulosic ethanol was reached at 42 °C in 24 h, with a yield of 2769.8 L/hasorghum, almost double that obtained for corn straw. This was the best cellulosic ethanol production from sweet sorghum bagasse achieved by K. marxianus strain and, therefore, it was used in the technology valuation. This analysis was carried out through the application of the profit equation, in which the revenue corresponded to ethanol sales in the global market in 2020, and the costs were defined by the activity-based costing method. An integration between 1G and 2G ethanol production, without mixing material streams, was considered to improve the technology valuation analysis. As a result, the equivalent to 17.2% of the profit obtained with the sugarcane juice ethanol was achieved for the sweet sorghum bagasse. Therefore, the technology valuation indicated the potential of sweet sorghum bagasse in the biorefinery ethanol production.

Diets low in fermentable oligo-, di-, and monosaccharides and polyols (FODMAPs) can help reduce symptoms in 50 to 80% of patients suffering from irritable bowel syndrome. Patients are, therefore, often advised to avoid products contributing to FODMAP intake, such as cereal grain products. However, these products are nutritious staple foods and avoiding their consumption may result in nutritional deficiencies. The development of low-FODMAP, high-fiber cereal grain products is therefore desirable. This pilot-scale study shows that Kluyveromyces marxianus CBS6014 (K. marxianus) results in more fructan hydrolysis and a significantly lower final fructan level in white and whole-grain toast bread as well as in rye sourdough bread compared to a commercial Saccharomyces cerevisiae baking strain. Moreover, combined fructan and fructose levels in white and whole-grain bread prepared with K. marxianus remained well below the threshold concentration for low-FODMAP products. In addition to reducing fructan levels, K. marxianus in rye sourdough bread also positively impacted bread height. Whereas further follow-up studies are needed to assess the potential of K. marxianus for bread production fully, our study suggests that this yeast species may open exciting novel routes for the production of low-FODMAP, high-fiber products.

Seventeen samples of Calabrian ewe’s milk, ewe’s cheese (Pecorino del Poro) made with raw milk, goat’s milk, and goat’s cheese (Caprino d’Aspromonte) made with raw milk were used to obtain 124 yeast isolates. The most abundant species was Debaryomyces hansenii (61.3%), followed by Candida zeylanoides (32.3%) and Kluyveromyces marxianus (3.2%). The enzymatic profile of 25 selected yeast strains was determined. Lastly, they were studied for their interaction with eight dairy lactic acid bacteria – four coccal-shaped and four rod-shaped. The best strains may be used as adjunct cultures for cheese making.

Background: Natural products constitute more than half of all biomolecules lately being used in clinical settings. Mannoprotein derived from the yeast cell wall has found full biotechnological applications. Objective: This study was intended to investigate antioxidant, anticancer, and toxicological properties of Kluyveromyces marxianus mannoprotein (KM). Methods: The KM extract was obtained through a sequence of operations, including centrifugation for cell isolation, precipi-tation with potassium citrate/ sodium metabisulfite, and recovery and purification. Its antioxidant, growth inhibition, macro-phage mitogenic, and toxic activities were evaluated for its future use in the biomedical field. Results: Significantly inhibitory effects of KM were obtained on reactive species. It showed antiproliferative activity against HeLa (human cervical adenocarcinoma) and MCF-7 (human breast cancer) cell lines with no toxic effects on HUVECs (human umbilical vein endothelial cells). The in vitro model of CHO-K1 (Chinese hamster ovary) cell lines did not show the cytotoxic and genotoxic of KM. Also, it enhanced macrophage activity in terms of nitric oxide (NO) production and viabil-ity. No sign of acute toxicity was found in BALB/c mice, and body weight remained unchanged in guinea pigs over three months. Conclusion: Comprehensive biological evaluations in this study are expected to expand the potential of KM as a natural material.

Kluyveromyces marxianus is a yeast that could be identified from kefir and can use a broad range of substrates, such as glucose and lactate, as carbon sources. The lactate produced in kefir culture can be a substrate for K. marxianus. However, the complexity of the kefir microbiota makes the traits of K. marxianus difficult to study. In this research, we focused on K. marxianus cultured with lactate as the sole carbon source. The optimal growth and released protein in lactate culture were determined under different pH conditions, and the LC–MS/MS-identified proteins were associated with the tricarboxylic acid cycle, glycolysis pathway, and cellular stress responses in cells, indicating that autolysis of K. marxianus had occurred under the culture conditions. The abundant glyceraldehyde-3-phosphate dehydrogenase 1 (GAP1) was cocrystallized with other proteins in the cell-free fraction, and the low transcription level of the GAP1 gene indicated that the protein abundance under autolysis conditions was dependent on protein stability. These results suggest that lactate induces the growth and autolysis of K. marxianus, releasing proteins and peptides. These findings can be fundamental for K. marxianus probiotic and kefir studies in the future.

In recent years, increased rates of yeast infections in humans and animals have been recognized worldwide. Since animals may represent a source of yeast infections for humans, knowing the antifungal susceptibility profile of yeast isolates from milk and evaluating their pathogenic potential would be of great medical importance. Therefore, the aim of this survey was to study yeast diversity in milk samples, analyze the hemolytic and phospholipase activities of isolates and determine minimal inhibition concentration (MIC) for fluconazole, voriconazole and flucytosine. Out of 66 yeast isolates obtained from 910 individual raw milk samples from subclinically infected cows, 26 different yeast species were determined based on sequencing of the D1/D2 and ITS regions. Among them, Pichia kudriavzevii (formerly known as Candida krusei), Kluyveromyces marxianus (formerly known as Candida kefyr) and Debaryomyces hansenii (formerly known as Candida famata) were the most commonly identified. Hemolysin and/or phospholipase activity was observed in 66.7% of isolates. The elevated MIC for fluconazole was determined in 16 isolates from 11 species. The findings of this study demonstrate that yeast isolates from raw milk have the potential to express virulence attributes such as hemolysin and phospholipase, and additionally, some of these strains showed elevated MIC to fluconazole or to flucytosine. Lay summary We identified 66 yeast isolates, including 26 different yeast species from 910 individual milk samples. Our results indicate that individual milk samples may serve as a source of yeasts with the potential to trigger infection and may have reduced sensitivity to tested antifungal agents.

An industrial process is profitable when its individual unit operations are efficient and thus, this work shows a guideline for designing efficient fermentation-industrial processes for agave distilled production based on a sequential approach of optimization, beginning in the laboratory and followed by the adjustment of the variable values using the evolutionary operation method for successful process scaling. The results at the laboratory showed that a starter inoculum containing a 5 × 106 cells/mL mixture of Kluyveromyces marxianus, Clavispora lusitaniae, and Kluyveromyces marxianus var. drosophilarum strains in a bioreactor containing agave syrup with 120 g/L fermented sugar, processed at a constant temperature of 33 °C and 1.0 VVM aeration for 1.6 h, led to a fermented product with a 4.18% (v/v) alcohol content after 72 h of processing time. The scale-up process results showed that the best operating conditions at the pilot-plant level were a temperature of 35 °C and aeration at 1.0 VVM for 1.2 h, which led to a fermented product with a 4.22% (v/v) total alcohol content after 72 h of processing time. These represent similar performance values for both production processes, but each one worked with their specific values of process variables, which demonstrates that each level of production had its own specific values for process variables. The volatile compound analysis shows that both distilled products contained a similar profile of volatile components that provide fruity and ethereal aromatic notes pleasant to the palate. Therefore, the process design for agave spirit production at the semi-industrial level was successfully achieved.

Doogh is an Iranian drink produced from yoghurt. Swelling is an undesirable change of Doogh, usually caused by Kluyveromyces marxianus. The aim of this study was the investigation of the effect of the thermosonication process on Kluyveromyces marxianus survival as well as the determination of the impact of this process on other quality parameters (viscosity and serum separation). For this reason, Kluyveromyces marxianus (about 105 cells per milliliter) inoculated into Doogh samples, thereafter samples being exposed to various thermosonication process conditions. Results showed that the thermosonication operation resulted in the reduction of the yeast population of Doogh samples. The lowest yeast population was observed in the sample subjected to acoustic density=1w/ml, T= 60°C and t=30min (1.7±0.01CFU/ml). Other operation conditions also had impact on yeast survival. Thermosonication also had significant effects on the viscosity of the Doogh samples. Serum separation of samples was also measured and observation implied changing serum separation magnitude between different samples in comparison with the control sample.

The demand for "preservative-free" food products is rising, and biopreservation seems to be a potential alternative to replace or reduce the use of chemical preservatives. This study's objective was to assess the antifungal activity of lactic acid bacteria (LAB) (n = 98) and the efficacy and applicability of the chosen bioprotective cultures against fungal spoilers in dairy products. First, 14 strains of antifungal strains were preliminarily screened by in vitro tests against Pichia pastoris D3, Aspergillus niger D1, Geotrichum candidum N1, Kluyveromyces marxianus W1, and Penicillium chrysogenum B1 and validated by challenge tests in yogurts, indicating that the fungal-inhibiting activity of LAB was species specific and yogurts fermented with antifungal LAB cultures were more effective in extending the shelf life. Secondly, the chosen 14 LAB strains were identified by the 16SrDNA sequence analysis and carbohydrate fermentation test. The results were as follows: 9 strains were Lactobacillus plantarum , 3 were Lactobacillus paracasei , 1 was Enterococus faecium , and 1 was Lactobacillus rhamnosus. Among them, active L. plantarum N7 was the chosen and studied factor that affects the antifungal activity using the response surface methodology (RSM). Finally, in situ tests were conducted to validate the activity of L. plantarum N7 in actual dairy products (whey beverage). Physicochemical and microbial indices of whey beverage during storage period exhibited that antifungal L. plantarum N7 could slow the fungal growth and be candidates of interest for industrial applications.

Kluyveromyces marxianus is an ascomycetous yeast which has shown promising results in cellulosic ethanol and renewable chemicals production. It can survive on a variety of carbon sources under industrially favorable conditions due to its fast growth rate, thermotolerance, and acid tolerance. K. marxianus, is generally regarded as a safe (GRAS) microorganism, is widely recognized as a powerhouse for the production of heterologous proteins and is accepted by the US Food and Drug Administration (USFDA) for its pharmaceutical and food applications. Since lignocellulosic hydrolysates are comprised of diverse monomeric sugars, oligosaccharides and potential metabolism inhibiting compounds, this microorganism can play a pivotal role as it can grow on lignocellulosic hydrolysates coping with vegetal cell wall derived inhibitors. Furthermore, advancements in synthetic biology, for example CRISPR-Cas9 (clustered regularly interspaced short palindromic repeats with Cas9)-mediated genome editing, will enable development of an engineered yeast for the production of biochemicals and biopharmaceuticals having a myriad of industrial applications. Genetic engineering companies such as Cargill, Ginkgo Bioworks, DuPont, Global Yeast, Genomatica, and several others are actively working to develop designer yeasts. Given the important traits and properties of K. marxianus, these companies may find it to be a suitable biocatalyst for renewable chemicals and fuel production on the large scale. This paper reviews the recent progress made with K. marxianus biotechnology for sustainable production of ethanol, and other products utilizing lignocellulosic sugars.

A produção do etanol como combustível é uma alternativa promissora a utilização dos combustíveis fósseis. Nesse contexto, os resíduos agroindustriais têm despontado como matérias-primas capazes de aumentar a produção de etanol sem competir com a produção de alimentos. Assim, tanto o soro de queijo, quanto subprodutos lignocelulósicos podem ser utilizados como substratos para a produção de etanol. No entanto, a levedura Saccharomyces cerevisiae é incapaz de metabolizar a lactose presente no soro de queijo e de crescer nas elevadas temperaturas requeridas no processo de Sacarificação e Fermentação Simultâneas (SSF) de biomassa lignocelulósica. Por outro lado, Kluyveromyces marxianusCCT7735 é uma linhagem desta levedura, que apresenta potencial para produzir etanol tanto a partir de soro de queijo quanto de subprodutos lignocelulósicos. Isto porque ela fermenta lactose e cresce em altas temperaturas. Todavia, ela tem o seu crescimento inibido em altas concentrações de etanol. Portanto, para viabilizar a produção de etanol, torna-se necessário compreender os mecanismos de resposta ao estresse por etanol em K. marxianus. Sendo assim, este trabalho teve como objetivo descrever e analisar os principais mecanismos de resposta ao estresse por etanol em leveduras,descritos na literatura, através de uma revisão bibliográfica, a fim de compreender como K. marxianus responde ao estresse etanólico. Nesse sentido, a abordagem utilizada nesta pesquisa foi do tipo exploratório, descritivo e bibliográfico.Desta forma, os artigos obtidos a partir das bases de dados eletrônicos foram analisados e sistematizados de acordo com a sua relevância para a compreensão dos mecanismos de respostas ao estresse por etanol em leveduras. A partir da análise dos mecanismos envolvidos na tolerância ao etanol descritos na literatura concluiu-se que eles contribuem para o entendimento das respostas moleculares ao estresse etanólico em K. marxianus. Entretanto, devido à complexidade destes mecanismos torna-se necessário fazer um estudo mais detalhado a nível molecular a fim de compreendê-los melhor.

Oil palm empty fruit bunch (OPEFB) has high potency to be converted into second generation bioethanol production. OPEFB pretreated liquid hot water (LHW) assisted maleic acid (MA) pretreatment was used as substrate in simultaneous saccharification fermentation (SSF). This pretreatment was conducted at 180 °C for 70 min by using 1.5% (w/v) MA acid concentration with material and liquid ratio of 1:14. Subsequently, the pretreated samples were fermented by addition of surfactant, polyethylene glycol (PEG) 4000 and amphiphilic lignin derivatives (A-LD) using Kluyveromyces marxianus InaCC Y119 yeast at temperature of 38 °C for 89 h. Before SSF, pretreated OPEFB was pre hydrolyzed by cellulase enzyme with enzyme loading 30 FPU/g at 50 °C for 3 h. Fermentation filtrate was taken at 0, 24, 48, 72, dan 89 h as much as 1.5 mL for reducing sugar and ethanol content determination. The surfactant addition can enhance reducing sugar and also ethanol content compared to control. The highest ethanol content of 1.251 g/L was found at fermentation of OPEFB pretreated MA with PEG 4000 surfactant at 89 h.

Yeasts are distributed in all environments and have been reported as potential biocontrol agents against various phytopathogenic fungi. To investigate their enzymatic and biological activities, 32 yeasts were isolated from 15 date vinegar samples. Evaluation of the antagonistic activities of isolated yeasts against the plant pathogens Fusarium oxysporium, Sclerotinia sclerotiorum, and Macrophomina phaseolina indicated that there are two yeasts had the highest inhibitory effect against plant pathogens, these yeasts identified as Kluyveromyces marxianus and Torulaspora delbrueckii using traditional and molecular methods. These yeast isolates were tested for fungal cell wall degrading enzymes (in vitro), and results indicated that the yeasts had strong protease and amylase enzyme activity and moderate chitinase and cellulase enzyme activity. The antagonistic activities of each yeast were evaluated using a dual culture technique. The results showed that K. marxianus inhibited the mycelial growth of F. oxysporium, S. sclerotiorum, and M. phaseolina by 70.5, 57.5, and 75.5%, respectively, whereas T. delbrueckii inhibited mycelial growth of F. oxysporum, S. sclerotiorum, and M. phaseolina by 55.3%, 66.2%, and 31.11%, respectively. The biofilm production assay indicated that the tested yeast could form biofilms as a mechanism of antagonistic activity against phytopathogenic fungi.

The thermotolerant yeast Kluyveromyces marxianus is a promising bioethanol producer, but its fermentation arrested earlier at high temperatures. Untargeted metabolomic analysis was used to explore K. marxianus’s stress responses during high-temperature fermentation. Fermentation experiments were conducted at 45 °C, with a group conducted at 30 °C as control. Samples were collected from both groups at 14 and 22 h and were subjected to metabolomic analysis. The results show that pathways related to biosynthesis of amino acids, metabolism of purine, pyridoxine and riboflavin, etc. were induced at high temperature. More oleic acid, less palmitic acid and less palmitoleic acid were synthesized at 45 °C than at 30 °C, indicating that K. marxianus can adapt to high temperature by increasing the mean fatty acid chain length. In addition, most of the metabolic pathways stopped after the fermentation arrest, but pathways related to amino acids metabolism, riboflavin metabolism, etc. were still active. This is the first study about comprehensive metabolomic profiling of K. marxianus during high-temperature fermentation. The findings in this study offer deeper insight in mechanisms of K. marxianus’s stress responses during high-temperature fermentation at the metabolome level, and provide potential target pathways for further metabolic engineering towards improved stress tolerance and efficient bioethanol production.

Biopreservation is defined as using microbes, their constituents, or both to control spoilage while satisfying consumer demand for clean-label products. The study objective was to investigate the efficacy of bacterial cultures in biopreserving cottage cheese against postprocessing fungal contamination. Cottage cheese curd and dressing were sourced from a manufacturer in New York State. Dressing was inoculated with 3 different commercial protective cultures—PC1 (mix of Lacticaseibacillus spp. and Lactiplantibacillus spp.), PC2 (Lacticaseibacillus rhamnosus), and PC3 (Lactic. rhamnosus)—following the manufacturer recommended dosage and then mixed with curd. A control with no protective culture was included. Nine species of yeast (Candida zeylanoides, Clavispora lusitaniae, Debaryomyces hansenii, Debaryomyces prosopidis, Kluyveromyces marxianus, Meyerozyma guilliermondii, Pichia fermentans, Rhodotorula mucilaginosa, and Torulaspora delbrueckii) and 11 species of mold (Aspergillus cibarius, Aureobasidium pullulans, Penicillium chrysogenum, Penicillium citrinum, Penicillium commune, Penicillium decumbens, Penicillium roqueforti, Mucor genevensis, Mucor racemosus, Phoma dimorpha, and Trichoderma amazonicum) were included in the study. Fungi strains were previously isolated from dairy processing environments and were inoculated onto the cheese surface at a rate of 20 cfu/g. Cheese was stored at 6 ± 2°C. Yeast levels were enumerated at 0, 7, 14, and 21 d postinoculation. Mold growth was visually observed on a weekly basis through d 42 of storage and imaged. Overall, the protective cultures were limited in their ability to delay the outgrowth in cottage cheese, with only 8 of the 20 fungal strains showing an effect of the cultures compared with the control. The protective cultures were not very effective against yeast, with only PC1 able to delay the outgrowth of 3 strains: D. hansenii, Tor. delbrueckii, and Mey. guilliermondii. The efficacy of these protective cultures against molds in cottage cheese was more promising, with all protective cultures showing the ability to delay spoilage of at least 1 mold strain. Both PC1 and PC2 were able to delay Pen. chrysogenum and Pho. dimorpha outgrowth, and PC1 also delayed Pen. commune, Pen. decumbens, and Pen. roqueforti to different extents compared with the controls. This study demonstrates that commercial lactic acid bacteria cultures vary in their performance to delay mold and yeast outgrowth, and thus each protective culture should be evaluated against the specific strains of fungi of concern within each specific dairy facility.

Ethyl acetate can be synthesized from acetyl-CoA and ethanol via a reaction by alcohol acetyltransferases (AATase) in yeast. In order to increase the yield of acetyl-CoA, different terminators were used to optimize the expressions of acetyl-CoA synthetase (ACS1/2) and aldehyde dehydrogenase (ALD6) to increase the contents of acetyl-CoA in Saccharomyces cerevisiae. ATF1 coding AATase was coexpressed in expression cassettes of ACS1/ACS2 and ALD6 to promote the carbon flux toward ethyl acetate from acetyl-CoA. Further to improve ethyl acetate production, four heterologous AATase including HuvEAT1 (Hanseniaspora uvarum), KamEAT1 (Kluyveromyces marxianus), VAAT (wild strawberry), and AeAT9 (kiwifruit) were introduced. Subsequently mitochondrial transport and utilization of pyruvate and acetyl-CoA were impeded to increase the ethyl acetate accumulation in cytoplasm. Under the optimal fermentation conditions, the engineered strain of PGAeΔPOR2 produced 1.69 g/L ethyl acetate, which was the highest value reported to date by metabolic engineering methods.

Xylan, second most abundant natural polysaccharide after cellulose, has tremendous applications in various industrial processes. There are several xylan degrading catalysts available, but, still, there is continuous quest among the scientific community to search for new hyperactive xylanases. In the present study, the catalytic potential of xylanase from Bacillus sp. XPB-11 was improved using various mutagens. 5-Bromo-Uracil proved to be the best mutagen resulting in stable and hyperactive mutant BU-7 that showed maximum xylanase activity, i.e., 154.16 ± 2.23 U/mg protein. Further, optimizing the various parameters resulted in 20% enhancement and overall twofold increase in the xylanase activity of mutant, i.e.188.24 ± 1.80 U/mg protein. This hyperactive mutant released 26.8 ± 2.17 mg/ml of C-5 sugar from microwave + 0.5 N NaOH pretreated pine needle waste biomass that on fermentation using Kluyveromyces marxianus MTCC 1498 produced 5.34 g/l of bioethanol with 3.89% yield and 93.5% purity. The hyperactive xylanase from Bacillus sp. XPB-11 mutant BU-7, developed in the present study to release C-5 sugar from pine needle waste biomass and its subsequent biotransformation into bioethanol, is of commercial significance to utilize one of the problematic waste biomass from Himalayan Region for its conversion into second-generation biofuel i.e. bioethanol.

This work presents a study using coconut cultivation wastes as carbon sources to produce lignocellulosic ethanol, such as green coconut shell (GCS), coconut-tree leaflet (CLL) and coconut-tree leaf stalk (CLS). The biomasses were submitted to hydrothermal (HPT) and acidic (APT) pretreatment. Enzymatic hydrolysis was performed using Cellic® Ctec2 supplemented with 10% Cellic® Htec, mainly based on the identification of glucose and xylose. It was found that enzymatic hydrolysis after HPT promoted better results when compared to APT. The glucose concentrations obtained for the biomasses submitted to the HPT were: 31.85 g/L for the GCS, 21.31 g/L for the CLL and 45.39 g/L for the CLS. For xylose, the higher concentration obtained was 7.93 g/L. During the fermentation step applying the yeast Kluyveromyces marxianus, ethanol concentrations (g/L) from HPT and APT hydrolyzed liquors were: 8.83 and 9.71 g/L for GCS, 10.26 and 7.01 g/L for CLL and 12.99 and 7.44 g/L for CLS, respectively. This study identified the energy potential of coconut residues in the production of biofuels, specifically ethanol, from K. marxianus, species which is little explored commercially and able to ferment glucose and xylose.

The inhibitory effect of toxic byproducts mainly derived from the pretreatment of lignocellulosic biomass is one of the limiting factors hindering industrial application. In this work, a simple yet practicable aeration strategy was proposed to enhance microbial tolerance to inhibitors, including formic acid, acetic acid, furfural, and 5-hydroxymethyl furfural (5-HMF). Batch fermentation by Kluyveromyces marxianus CICC 1727−5 using xylose and glucose alone and in combination under different aeration conditions coupled with multiple inhibitors was performed. The fermentation time and residual xylose significantly decreased with high xylitol production (0.7 g/g) and high ethanol production (0.42 g/g) at an aeration rate of 0.5 vvm. The postglucose effect was effectively relieved during co-fermentation by the microaeration strategy. Transcriptomics analysis was performed to determine the mechanism of inhibitor resistance. Transcription of genes related to DNA replication, protein processing, carbon metabolism, biosynthesis of the ribosome, amino acids, peroxidase, sterols, and unsaturated fatty acids were significantly upregulated accompanied by downregulated expression of genes related to autophagy under aeration conditions in the presence of multiple inhibitors. These results provide a valuable reference for future studies on biofuel fermentation from lignocellulosic biomass by a facultative fermenting microbe.

In the food industry, the β-galactosidase enzyme is widely used, being applied mainly in the elaboration of products for the lactose intolerant, can carry out the hydrolysis of lactose. To date, it...

We present a case of Kluyveromyces marxianus keratitis nine months after Descement's membrane endothelial keratoplasty (DMEK) in a patient with Fuchs endothelial disease. Endothelial scraping revealed this rare yeast infection at the interface between graft and host cornea. Immediate antifungal treatment with intracameral and corneal intrastromal injections of voriconazole and amphotericin B remained unsuccessful, requiring penetrating keratoplasty. This case highlights the challenging management of keratomycosis in patients with endothelial grafts.

International audience; The production of 2-phenylethanol (2-PE), a high-valued aroma, has been performed naturally through the bioconversion of Lphenylalanine by the Ehrlich pathway in yeast. Nevertheless, one of the main limitations of this bioconversion is the inhibition of yeast growth by 2-PE. This work evaluated the intensification of 2-PE production in a batch culture using a conventional bioreactor coupled to an in-situ product removal (ISPR) process, by means of membrane-based solvent extraction (MBSE). The culture of the native yeast Kluyveromyces marxianus ITD0090, using a shaker flask stirred at 150 rpm, produced a maximum concentration of 2-PE of 0.7 g L −1 after 24 hours. When the yeast was cultivated in a controlled aerated-stirred bioreactor operated at 0.8 vvm and 350 rpm, the maximum production of 2-PE after 48 h reached 1.49 g L −1. Intensification of the bioprocess was achieved by coupling the controlled bioreactor with a MBSE system, which allowed the continuous recovery of the 2-PE produced, reaching a final titre of 3.02 g L −1 after 56 h.

This study was intended to investigate physico-chemical, rheological, and emulsifying properties of oil-in-water emulsions prepared from the Kluyveromyces marxianus mannoprotein (KMM). Also, the stress-response function of the KMM emulsions was compared with that of the whey protein concentrate (WPC) emulsions in terms of zeta potential, size, and rheology. The stress experiments were conducted at different pH (3 to 9), ionic composition (0 to 500 mM NaCl), and temperatures (30 to 90 °C). The extracted KMM with a molecular weight of 107.2 kDa had 28.8% proteins and 68.22% carbohydrates. With increasing the KMM concentration to 1.5% (w/w), the zeta potential, droplet size, and apparent viscosity of the emulsions reached -35 mV, ∼1 μ, and ∼9 mPa·s, respectively. After applying pH, ionic composition, and temperature, the KMM emulsions were more stable than the WPC emulsions. In conclusion, KMM can be used as a bioemulsifier and be more effective in stabilizing emulsions than WPC. PRACTICAL APPLICATION: Yeasts are a rich source of natural materials. In this study, we extracted mannoproteins from the yeast cell wall and evaluated their functional properties to be used as an emulsifier in oil-in-water emulsions. The results of this study confirm that the yeast-derived mannoproteins are good at stabilizing these emulsions either in the presence or absence of different environmental conditions.

Mutant sugar transporter ScGAL2-N376F was overexpressed in Kluyveromyces marxianus for efficient utilization of xylose, which is one of the main components of cellulosic biomass. K. marxianus ScGal2_N376F, the ScGAL2-N376F-overexpressing strain, exhibited 47.04 g/l of xylose consumption and 26.55 g/l of xylitol production, as compared to the parental strain (24.68 g/l and 7.03 g/l, respectively) when xylose was used as the sole carbon source. When a mixture of glucose and xylose was used as the carbon source, xylose consumption and xylitol production rates were improved by 195% and 360%, respectively, by K. marxianus ScGal2_N376F. Moreover, the glucose consumption rate was improved by 27% as compared to that in the parental strain. Overexpression of both wild-type ScGAL2 and mutant ScGAL2-N376F showed 48% and 52% enhanced sugar consumption and ethanol production rates, respectively, when a mixture of glucose and galactose was used as the carbon source, which is the main component of marine biomass. As shown in this study, ScGAL2-N376F overexpression can be applied for the efficient production of biofuels or biochemicals from cellulosic or marine biomass.

Yeast communities associated with cacti were studied in three ecosystems of Southeast, Central and North Brazil. A total of 473 yeast strains belonging to 72 species were isolated from 190 samples collected. Cactophilic yeast species were prevalent in necrotic tissues, flowers, fruits and insects of cacti collected in Southeast and North Brazil. Pichia cactophila, Candida sonorensis and species of the Sporopachydermia complex were the most prevalent cactophilic species in Southeast and Central regions. Kodamaea nitidulidarum, Candida restingae and Wickerhamiella cacticola were frequently associated with cactus flowers and fruits. The diversity of yeasts associated with the substrates studied was high. Twenty-one novel species were found. One is described here as Kluyveromyces starmeri sp. nov. based on 21 isolates obtained from necrotic tissues, flowers, fruits and associated insects of the columnar cacti Cereus saddianus, Micranthocereus dolichospermaticus and Pilosocereus arrabidae in two different ecosystems in Brazil. Phylogenetic analyses of sequences encoding the gene of the small subunit (SSU) rRNA gene, the internal transcribed spacer, the 5.8S rRNA gene and the D1/D2 domains of the large subunit (LSU) rRNA showed that the species is related to Kluyveromyces dobzhanskii, Kluyveromyces lactis and Kluyveromyces marxianus. Phylogenomic analyses based on 1264 conserved genes shared among the new species and 19 other members of the Saccharomycetaceae confirmed this phylogenetic relationship. The holotype is K. starmeri sp. nov. CBS 16103T (=UFMG-CM-Y3682T ). The Mycobank number is MB 836817.

In this study, we aimed to assess the feasibility of the lactic acid bacterium Lactobacillus kefiranofaciens DN1 (LKF_DN1) and the yeast Kluyveromyces marxianus KU140723-05 (KMA5), recently isolated from kefir, as probiotics. Specifically, we evaluated the effect of early administration of these 2 microbes on the inhibition of Salmonella Enteritidis (SE) colonization in neonatal chicks. We also examined the effects of exposure of chicks to probiotics before SE exposure on the reduction in the number of gut SE. A total of 108 1-day-old specific-pathogen-free male layer chicks were used for 3 independent experiments. The experimental chicks were randomly divided into 6 groups (negative control: basal diet [BD] without probiotics and SE; positive control: BD; probiotic group [PG] 1: BD + LKF_DN1; PG2: BD + KMA5; PG3: BD + LKF_DN1 + KMA5; and PG4: BD+ a commercial product IDF-7), all of which, except negative control, were coadministered with SE strain resistant to rifampicin (SERR). We found that the administration of LKF_DN1 and/or KMA5 reduced the number of viable cells of the SERR strain in chicks by up to 1.90 log10, relative to positive control chicks. Compared with late administration (day [D] 10 and D11), early administration (D1 and D2) of the probiotics was more effective in reducing SERR cell numbers in the gut. Furthermore, we detected no significant difference in the reduction of gut SERR cell numbers in chicks from the same groups exposed to the probiotics at D10 and D11 before and after administration with SERR. Collectively, our findings indicate that, as dietary additives, LKF_DN1 and KMA5 showed potential probiotic activity in chicks. Moreover, the combination of the lactic acid bacteria and/or yeast strain was found to rapidly reduce SE numbers in the chick gut and showed a prolonged inhibitory effect against SE colonization. We, thus, propose that the administration of these 2 probiotics, as early as possible after hatching, would be considerably effective in controlling SE colonization in the guts of chicks.

In the study, Lactobacillus paracasei H4‐11, Lactobacillus fermentum D1‐1, Lactobacillus casei H1‐8, Lactobacillus reuteri H2‐12, and Kluyveromyces marxianus L1‐1 were screened from traditional fermented rice acid based on several indicators: L‐lactic acid production capacity (13.46 ~ 19.69 g/kg), antioxidant capacity (DPPH clearance ability of 35.36 ~ 56.89%), and savory flavor indicators. Glutinous rice, quinoa, barley rice, and brown rice were selected to carry out rice acid fermentation. Different viable lactic acid bacteria and yeasts were screened, respectively, in the saccharification, acidification, alcoholization, and late acidification stages. Rice acid fermented with L. paracasei H4‐11 and K. marxianus L1‐1 in glutinous rice showed the high L‐lactic acid content (23.09 g/kg). The DPPH free radical scavenging ability in rice acid fermented with L. fermentum D1‐1 and L. paracasei H4‐11, respectively, reached 34.27% and 33.05% in 96 hr. Although quinoa rice acid had the highest L‐lactic acid content (33.74 g/kg) and the DPPH free radical scavenging ability (60.10%), it had the poor taste due to the high astringency intensity and bitter intensity. Rice acid fermented with both L. paracasei H4‐11 and K. marxianus L1‐1 in glutinous rice showed the highest savory flavor and had the lowest astringency and bitter. L. paracasei H4‐11 and K. marxianus L1‐1 were the potential strains for the fermentation of rice acid. These results promote the industrial development of Chinese rice acid. Practical applications Rice acid as a functional nondairy fermented product is widely accepted by Chinese consumers. However, rice acid fermentation is still a traditional spontaneous process, which causes instability in its flavor and quality. Lactobacillus paracasei H4‐11 and Kluyveromyces marxianus L1‐1 screened in this study contributed to the improvements in the flavor and antioxidant capacity of rice acid. In addition, glutinous rice was confirmed as the suitable fermentation material of rice acid in the study., The Lactobacillus paracasei H4‐11 and Kluyveromyces marxianus L1‐1 were isolated in this study contribute to the improvement of flavor and antioxidant capacity of rice acid. In addition, the mechanism of glutinous rice suitable for growth in rice acid system was verified, making our study particularly relevant. These results would aid in the development of industrial‐scale Chinese acidic fermented rice soup.

BACKGROUND Exoinulinase catalyzes the successive removal of individual fructose moiety from the non-reducing end of the inulin molecule, which is useful for biotechnological applications like producing fructan-based non-grain biomass energy and high-fructose syrup. In this study, an exoinulinase (KmINU) from Kluyveromyces marxianus DSM 5418 was tailored for increased catalytic activity and acidic adaptation for inulin hydrolysis processes by rational site-directed mutagenesis. RESULTS Three mutations, S124Y, N158S and Q215V distal to the catalytic residues of KmINU were designed and heterologously expressed in Pichia pastoris GS115. Compared to the wild-type, S124Y shifted the pH-activity profile towards acidic pH values and increased the catalytic activity and catalytic efficiency by 59% and 99% to 688.4 ± 17.03 s-1 and 568.93 L mmol-1 s-1 , respectively. N158S improved the catalytic activity under acidic pH conditions, giving a maximum value of 464.06 ± 14.06 s-1 on inulin at pH 4.5. Q215V markedly improved the substrate preference for inulin over sucrose by 5.56-fold, and showed catalytic efficiencies of 208.82 and 6.88 L mmol-1 s-1 towards inulin and sucrose, respectively. Molecular modeling and computational docking indicated that structural reorientation may underlie the increased catalytic activity, acidic adaptation and substrate preference. CONCLUSIONS The KmINU mutants may serve as industrially promising candidates for inulin hydrolysis. Protein engineering of exoinulinase here provides a successful example of the extent to which mutating non-conserved substrate recognition and binding residues distal to the active site can be used for industrial enzyme improvements. © 2020 Society of Chemical Industry.

Background: Yeasts play diverse roles in human life. Since ancient times, these micro organisms have been used to produce food products and beverages including bread and beer. Nowadays, the biotechnological products of yeast are some of the main components of commercial products. Objective: Some species of yeast such as Saccharomyces cerevisiae and Saccharomyces boulardii are recognized as probiotic yeast with extensive applications in the food and drug industries. However, certain species like Kluyveromyces marxianus are still not recognized as probiotic micro organisms despite their widespread industrial usage. In this study, the application of K. marxianus in preparing food and the medicinal product was reviewed in terms of its beneficial or harmful effects. Methods: Pub Med, Google Scholar, Scopus, and Science Direct databases were searched by using “Probiotics”, “Yeast”, and “Kluyveromyces marxianus”. Results: The findings suggest that K. marxianus can be recognized as a probiotic yeast species. Conclusion: It can be concluded that K. marxianus may be considered as a probiotic micro organism with a variety of commercial and medical applications.

Purpose: The aim of this study was to investigate the enhancement of very high gravity (VHG) bioethanol production using co-substrates of cassava chip hydrolysates and molasses by Kluyveromyces marxianus DMKU-KS07. Methods: The factors effecting cassava chip hydrolysis by raw starch degrading enzyme (RSDE) from the thermophilic bacterium Laceyella sacchari LP175 and commercial glucoamylase were investigated. The obtained raw cassava chip hydrolysate was co-fermented with molasses by the thermotolerant K. marxianus DMKU-KS07 at 42°C under nonsterile system. Results: Factors affecting sugar syrup production from raw cassava chips by synergistic hydrolysis and enzyme saccharification were optimized at temperature, substrate concentration and agitation rate of 50 °C, 250 g/L and 200 rpm, respectively. A yield of 98.6 g/L was obtained at 6 h of incubation, equivalent to 45.9% saccharification and 51.4% hydrolysis of starch, respectively. High bioethanol concentration at 118 g/L, with highest productivity of 2.19 g/L/h and ethanol yield (YP/S) at 0.44 g EtOH/g total sugar, equivalent to 86.3% theoretical yield, was obtained by modified simultaneous saccharification and fermentation (Modified SSF) with co-fermentation of substrates from the enzymatic hydrolysates of raw cassava chips at 42 °C for 12 h. Subsequent addition of molasses increased the final concentration to 100 g total sugar/L at 36 h. Conclusions: Co-fermentation of raw cassava chip hydrolysates with molasses enhanced production of bioethanol at VHG condition, and showed potential for application in ethanol production by enhancing the fermentation process and reducing energy consumption.

The attempts toward addition of biocontrol agents in dairy products have gained popularity. Here, we worked on analysing the antifungal activity of binary and ternary combinations of three Lactic Acid Bacteria (LAB) against five spoilage yeasts in yogurt. The yogurt samples were characterized in terms of pH, acidity, WHC, textural parameters, viscosity, survivability and antifungal activity of LAB and sensorial properties during cold storage. The results showed that the inoculation of LAB in yogurt gave rise in significant reduction of pH throughout cold storage while titrable acidity and WHC decreased (p

The yeast species Saccharomyces cerevisiae and Kluyveromyces marxianus are associated with fermentation of West African indigenous foods. The aim of this study was to characterize potential probiotic properties of S. cerevisiae and K. marxianus isolates from the West African milk products lait caille and nunu and a cereal-based product mawe. The strains (14 in total) were identified by 26S rRNA gene sequencing and characterized for survival at gastrointestinal stress (bile salts and low pH) and adhesion to Caco-2 intestinal epithelial cells. Selected yeast isolates were tested for their effect on the transepithelial electrical resistance (TEER), using the intestinal epithelial cell line Caco-2 and for maintenance of intracellular pH (pHi ) during perfusion with gastrointestinal pH (3.5 and 6.5). All tested yeasts were able to grow in bile salts in a strain-dependent manner, exhibiting a maximum specific growth rate (μmax ) of 0.58-1.50 h-1 . At pH 2.5, slow growth was observed for the isolates from mawe (μmax of 0.06-0.80 h-1 ), whereas growth of yeasts from other sources was mostly inhibited. Yeast adhesion to Caco-2 cells was strain specific and varied between 8.0% and 36.2%. Selected strains of S. cerevisiae and K. marxianus were able to maintain the pHi homeostasis at gastrointestinal pH and to increase TEER across the Caco-2 monolayers, indicating their potential to improve intestinal barrier functions. Based on overall results, strains of K. marxianus and S. cerevisiae from mawe exhibited the highest probiotic potential and might be recommended for further development as starter cultures in West African fermented products.

Thermostable β-xylosidase produced by Aspergillus fumigatus (A. fumigatus) XC6 can be efficiently extracted and purified via an aqueous two-phase system consisting of 20% (m/v) PEG 2000 and 25% (m/v) NaH2PO4 with a purification factor of approximately 16. Maximum activity of the purified β-xylosidase was observed at 75 °C and pH 4.5 using p-nitrophenyl-β-Dxylopyranoside (pNPXyl) as a substrate. The thermal stability of purified β-xylosidase is higher than that of compared fungal strains. Thermotolerant kluyveromyces marxianus was directly added to a fermentation system composed of 50 g L−1 rice straw and 10 g L−1 (NH4)2SO4, but prehydrolysis with A. fumigatus XC6 pellets for 12 h yielded 11.2 g L−1 ethanol within 36 h, an amount 80% of the theoretical value. The results indicated that A. fumigatus XC6 β-xylosidase is a thermostable enzyme with high xylose production activity, and may have promising industrial application for cost-effective lignocellulosic bioethanol production.

The effective utilization of okara (soybean residue) has become a considerable challenge in recent years. In this paper, the potential advantages of β-glucosidase production from okara fermented by Kluyveromyces marxianus were evaluated and the properties of the β-glucosidase were also characterized. The results showed that okara can significantly induce the production of β-glucosidase from K. marxianus. The β-glucosidase activity was up to 4.5 U/mg under optimized fermentation conditions. The optimal parameters were as follows: fermentation temperature 35 °C, cultivation time 98 h, inoculum concentration 10%, and 30 g/L of okara. After two steps of purification using ammonium sulfate precipitation and Sephadex G-75 column chromatography, the activity of β-glucosidase was 71.4 U/mg. The native enzyme was an approximately 66 kDa dimer consisting of two different subunits (22 and 44 kDa). The kinetic parameters of the K. marxianus β-glucosidase, using pNPG as substrate, were Vmax 8.34 μmol min−1 mg−1 and Km 7.42 mM. The β-glucosidase showed high thermostability and acid–alkali tolerance as well as low inhibition by DMSO (10–50%). In conclusion, this study supports the notion that okara fermentation by K. marxianus could be a useful process to produce β-glucosidase.

Pseudotrachydium kotschyi is a native medicinal plant used as a local medicine from Lorestan province of western Iran having numerous biological properties. After GC−MS analysis of Pseudotrachydium kotschyi essential oils (PKEOs), some biological activities were investigated. Additionally, TNF-α and TGF-β1 were studied in Leishmania-infected macrophages (in vitro) and Carrageenan-induced paw edema in rats. Finally, anti-inflammatory and wound-healing potential of PKEOs were investigated in vivo in mice. Total components identified by GC−MS contained 53 molecules, among which Z-α-trans-Bergamotol (23.25%), Durylaldehyde (16.07%), and α-Bergamotene (10.48%) existed in the highest amount in PKEOs. Antimicrobial assay of PKEOs showed maximum growth inhibition against Listeria monocytogenes and Kluyveromyces marxianus at the dose of 625 μg/ml. PKEOs exhibited a high cytotoxicity on lung cancer cell line A549 at a dose of 4 mg/ml. Anti-Leishmania activity of PKEOs showed concentration of 5000 µg/ml that had similar potency of glucantime at 125 µg/ml. The highest anti-inflammatory effect of PKEOs for Carrageenan-induced paw edema was observed at 100 concentrations, with paw inflammation reduced by 83.70% after using PKEO ointment. The present study reports broad-spectrum biological functions for essential oil from P. kotschyi. However, these biological activities, including anti-proliferative, anti-Leishmania, anti-inflammatory, antimicrobial and wound-healing potential, were not studied in detail. Therefore, it may be attractive for investigating the detailed mechanistic and molecular aspects of biological activities of PKEOs. Considering that the present study is the first to report the biological activity of P. kotschyi, further studies on their metabolites are needed to replicate the present findings.

To overcome the economic losses associated with fungi and their toxic metabolites, environmentally safe and efficient approaches are needed. To this end, biological control using yeasts and safe bacterial strains and their products are being explored to replace synthetic fungicides. In the present study, the biocontrol effect of a yeast strain of Kluyveromyces marxianus, QKM-4, against the growth and mycotoxin synthesis potential of key toxigenic fungi was evaluated. In vitro assays were performed to find the application of yeast volatile organic compounds (VOCs) against fungal contamination on important agricultural commodities. The removal of ochratoxin A (OTA) and deoxynivalenol (DON) by living and heat-inactivated yeast cells was also explored. VOCs produced by strain QKM-4 were able to significantly limit the fungal growth of 17 fungal species belonging to genera Aspergillus, Penicillium, and Fusarium. Yeast VOCs were able to reduce OTA biosynthesis potential of Penicillium verrucosum and Aspergillus carbonarius by 99.6 and 98.7%, respectively. In vivo application of QKM-4 VOCs against Fusarium oxysporum and A. carbonarius infection on tomatoes and grapes, respectively, determined a complete inhibition of fungal spore germination. GC/MS-based analysis of yeast VOCs identified long-chain alkanes, including nonadecane, eicosane, docosane, heptacosane, hexatriacontane, and tetracosane. In vitro testing of the mycotoxin-binding potential of the living and heat-inactivated QKM-4 cells showed a reduction of OTA and DON up to 58 and 49%, respectively, from artificially contaminated buffers. Our findings clearly demonstrate the strong antifungal potential of K. marxianus QKM-4 and propose this strain as a strong candidate for application in agriculture to safeguard food and feed products.

The red seaweed Gracilaria verrucosa has been used for the production of bioethanol. Pretreatment for monosaccharide production was carried out with 12% (w/v) G. verrucosa slurry and 500 mM HNO3 at 121°C for 90 min. Enzymatic hydrolysis was performed with a mixture of commercial enzymes (Cellic C-Tec 2 and Celluclast 1.5 L; 16 U/ml) at 50°C and 150 rpm for 48 h. G. verrucosa was composed of 66.9% carbohydrates. In this study, 61.0 g/L monosaccharides were obtained from 120.0 g dw/l G. verrucosa. The fermentation inhibitors such as hydroxymethylfurfural (HMF), levulinic acid, and formic acid were produced during pretreatment. Activated carbon was used to remove HMF. Wildtype and adaptively evolved Saccharomyces cerevisiae, Candida lusitaniae, and Kluyveromyces marxianus were used for fermentation to evaluate ethanol production.

The wastewater from the dumping site usually contains high pollutant levels. Biological process and physico-chemical treatments are among several technologies for wastewater treatment. Using microorganisms in the treatment of landfill leachate is an emerging research issue. Furthermore, bioremediation is a feasible approach for pollutants removal from landfill leachate which would provide an efficient way to resolve the issue of landfill leachate. In this study, the performance of yeast and bacteria isolated from kefir grains was assessed for landfill leachate treatment. Kefir grains microbial composition was evaluated by molecular approaches (Rep-PCR and 16S rRNA gene sequencing). The obtained outcomes denoted that high concentrations of lactic acid bacteria and yeast populations (over 107 CFU/ml) were found in the kefir grains and were essentially composed of Lactococcus lactis, Lactobaccillus kefirien, bacillus sp., L. lactis, and Kluyveromyces marxianus. The co-culture with 1% of inoculum size was demonstrated as the most efficient in the degradation of different contaminants. The overall abatement rate of chemical oxygen demand (COD), ammonium nitrogen ( NH 4 + - N ), and salinity were 75.8%, 85.9%, and 75.13%, respectively. The bioremediation process resulted in up of 75% removal efficiency of Ni and Cd, and a 73.45%, 68.53%, and a 58.17% removal rates of Cu, Pb, and Fe, respectively. The research findings indicate the performance of L. lactis and K. marxianus co-culture isolated from kefir grains for the bioremediation of LFL. PRACTITIONER POINTS: Isolation and identification of microorganisms from kefir grains was carried out. Biological treatment of LFL using monoculture of (Lactoccocus lactis; Kluyveromyces marxianus) and co-culture (5% of L. lactis and 5% K. marxianus) has been performed. Biological treatment using co-culture strain is an effective approach to remove organic matter, NH 4 + - N and heavy metals.

В работе представлены результаты исследований влияния эндополигалактуроназы дрожжей рода Kluyveromyces marxianus на процессы отделения и осветления сусла, а также качество коньячных виноматериалов и дистиллятов. Проблемой виноделия является высокое содержание полисахаридов в виноградной ягоде, что снижает выход сусла, затрудняет процессы его осветления, способствует окислению и обогащению нежелательными компонентами, ухудшающими качество готового продукта. Использование ферментных препаратов пектолитического действия в коньячном производстве ограничено из-за риска увеличения содержания метанола, который образуется при действии на пектины винограда пектинэстеразы. Перспективным направлением является использование в коньячном производстве эндополигалактуроназы, продуцируемой дрожжами вида Kluyveromyces marxianus . Установлено, что обработка сусла и мезги опытным ферментным препаратом, полученном при культивировании штамма дрожжей вида Kluyveromyces marxianus , способствует увеличению выхода сусла, снижению содержания взвесей и объема образующегося осадка при осветлении сусла, повышению его качества. Показано, что физико-химический состав виноматериалов и дистиллятов, полученных из обработанного сусла, практически не изменяется. Массовая концентрация метанола в опытных коньячных виноматериалах и дистиллятах не превышала уровня контрольных образцов. Установлена эффективность использования препарата в коньячном производстве с целью повышения выхода и качества сусла. The article presents the results of study on the influence of endopolygalacturonase yeast species Kluyveromyces marxianus on the processes of separation and clarification of must, as well as the quality of brandy wine materials and distillates. The problem of winemaking is the high content of polysaccharides in grape berries, which reduces the yield of must, complicates the process of clarification, contributes to the oxidation and enrichment with undesirable components degrading the quality of the finished product. Use of pectolytic enzymes in brandy production is limited due to the risk of increase in the content of methanol, appearing when pectin esterase acts on grape pectins. Use of endopoligalacturonase rendered by the yeast species Kluyveromyces marxianus is a promising direction in brandy production. It was found that treatment of the must and pulp with experimental enzyme preparation obtained by culturing a yeast strain of the species Kluyveromyces marxianus contributes to an increase in the yield of the must and a decrease in the content of suspensions and volume of the sediment formed in the process of must clarification, and quality improving. It was shown that the physicochemical composition of wine materials and distillates obtained from the treated must remained permanent. Mass concentration of methanol in the experimental brandy wine materials and distillates did not exceed the level of control samples. The effectiveness of use of the preparation in brandy production was established in order to increase the product yield and quality of the must.

Different strategies for simultaneous saccharification and fermentation (SSF) process were developed to enhance the productivity of ethanol by Kluyveromyces marxianus ATCC36907 as a part of biorefinery concept using cashew apple bagasse (CAB) as biomass. CAB was pretreated with 4.3% (v/v) alkaline hydrogen peroxide at pH 11.5 (CAB-AHP). Batch SSF conducted at 45 °C using 10% (w/v) CAB-AHP reported the highest concentration and ethanol yield. In the fed-batch SSF, the highest production (7.91 g ethanol/100gCAB) and ethanol yield (84.69%) were achieved using 10% (w/v) CAB-AHP initial solids load, 4% solid feeding at 24 h, and the feeding of enzymes only in the beginning of the process. The recycling of pretreatment inputs decreased 50% of the batch-SSF efficiencies, saving the consumption of water, H2O2, and NaOH in 66.67%, 48.44%, and 66.67%, respectively. Lignin was extracted from the hydrolysate with yield of 39.2%. Thus, the proposed biorefinery concept utilizes CAB to produce ethanol with high efficiency by fed-batch SSF, pretreatment input recycling, and the utilization of the extracted lignin for future applications.

Kefir grains are a microbial consortium of different genera of bacteria and yeasts. In this study, the performance of Tunisian Kefir grains during the biological treatment of a mixture of Gouda cheese whey and white wastewaters (GCW) in ratio 1:1 with very high organic matter concentration is investigated. The biological process was evaluated and optimized through the response surface methodology. Under the optimum conditions, Kefir grains concentration of 1.02%, temperature at 36.68 °C, and incubation time of 5.14 days, the removal efficiencies of COD, PO43−, and NO3− were 87, 37.48, and 39.5%, respectively. Interestingly, the reusability tests of the grains proved not only their high resistance to harsh environmental conditions but also their great potential for more practical applications. Particularly, different strains were isolated from the grains and identified as Kluyveromyces marxianus, Lactoccocus lactis, Lactobacillus kefiri, and Bacillus spp. using 16S rDNA sequence analysis and rep-PCR fingerprinting. At the biological level, the raw GCW (RGCW) has a negative impact on the Hordeum vulgare both on seed germination, and on the growth parameters of seedlings. Interestingly, after Kefir grains treatment, the treated GCW (TGCW) allow a seedlings growth and germination rate similar to those soaked in water.