Anomala Strain Research Articles

Evaluation of a phenolic-tolerant Wickerhamomyces anomalus strain for efficient ethanol production based on omics analysis

Lignocellulosic biomass is a highly economically viable substrate for ethanol fermentation, but the presence of phenolic compounds (PCs) in lignocellulosic hydrolysates severely hinders the growth of ethanologenic strains. This study aims to investigate the ethanol fermentation capabilities and PC tolerance of the newly isolated Wickerhamomyces anomalus CAP5. The strain demonstrated remarkable ethanol production, achieving 87.1 g/L with a yield of 0.47 g/g, exceeding 90 % of the theoretical maximum. To elucidate the mechanisms responsible for this high ethanol yield, comprehensive genome and transcriptome analyses were performed. These analyses identified key metabolic pathways, including acetate reabsorption and an incomplete glycerol synthesis pathway, as crucial factors contributing to the strain's superior performance. Moreover, W. anomalus CAP5 demonstrated notable resistance to PCs, particularly phenolic acids. In fermenting with coffee husk hydrolysate (CHH), which is rich in PCs, W. anomalus CAP5 achieved noteworthy ethanol production of 60.5 g/L, marking a record for ethanol production from CHH. Hence, W. anomalus CAP5 emerges as a promising strain for effective ethanol production from lignocellulosic hydrolysates enriched with PCs.

Exploring Non-Saccharomyces yeasts from Daqu for beer production

Daqu is an essential component in the solid-state fermentation of Chinese Baijiu, and non-Saccharomyces yeasts from Daqu significantly contribute to the complex flavors in Baijiu. However, the potential of non-Saccharomyces yeasts from Daqu in beer production has remained unexplored. In this study, 80 strains representing 14 non-Saccharomyces yeast species were isolated from various Daqu. These strains were scrutinized for their stress tolerance, flocculation, fermentation performance, and the production of flavor compounds. Subsequently, representative strains were chosen for beer production, and the resulting products from both single- and mixed-strain fermentations underwent sensory evaluation. Toruaspora delbrueckii TD2 exhibited excellent fermentation properties in wort. In compared to US-05, the non-Saccharomyces yeast strains tested significantly increased volatile favor compounds. Notably, the Wickerhamomyces anomalus strain showed a 13.86-fold increase in ethyl acetate production, while the Hanseniaspora osmophila strain displayed a 7-fold increase in phenylethyl acetate production. Co-fermentations of these strains with US-05 did not impact attenuation and ethanol concentration but significantly enhanced aromatic profiles. This study offers valuable insights into the fundamental attributes and brewing potential of non-Saccharomyces yeasts sourced from Daqu, broadening the spectrum of yeast selection for the creation of innovative beers.

Xylitol production by a Wickerhamomyces anomalus strain adapted for enhanced tolerance to sugarcane bagasse hemicellulosic hydrolysate with high content of fermentation inhibitors

BackgroundXylitol, a five-carbon polyalcohol, is used in the food and pharmaceutical industries and as a building block in the synthesis of high-value chemicals. It can be sustainably produced from renewable sources through xylose assimilating microbe fermentation. ResultsWe screened microbial strains for xylitol production and identified Wickerhamomyces anomalus Z1 as a key xylitol producer. Utilizing lignocellulosic biomass hydrolysates for xylitol production poses challenges due to microbial sensitivity to inhibitors from biomass pre-treatment. In this study, an adaptive laboratory evolution (ALE) of W. anomalus Z1 was performed by culturing the yeast in a mineral medium supplemented with gradual increases of sugarcane bagasse hemicellulosic hydrolysate (SCHH) obtained by intensified steam explosion pretreatment. The performance of the adapted yeast, named Wickerhamomyces anomalus ALE, was assessed in comparison to the wild-type strain regarding its capacity to produce xylitol using SCHH. The evolved yeast reached a xylitol yield of 0.11 g xylitol/g xylose whereas the wild-type strain could not produce xylitol. Removing acetic acid from SCHH enhanced W. anomalus ALE performance, with optimal results at 75% hydrolyzed hemicellulose, yielding 0.44 g xylitol/g xylose and 13.41 g/L xylitol. ConclusionsThis study demonstrates the potential of W. anomalus ALE in successfully valorizing the hemicellulosic fraction of sugarcane bagasse for sustainable xylitol production.How to cite: Bonfiglio F, Cagno M, Nuñez L, et al. Xylitol production by a Wickerhamomyces anomalus strain adapted for enhanced tolerance to sugarcane bagasse hemicellulosic hydrolysate with high content of fermentation inhibitors. Electron J Biotechnol 2024;71. https://doi.org/10.1016/j.ejbt.2024.05.004.

Characterization of yeast mutant strains for starter culture in Arabica coffee fermentation

Arabica coffee is the most popular and best-selling type of coffee. During coffee fermentation, microorganisms are essential for the production of metabolites and volatile compounds that affect coffee flavor quality. This work aimed to study the mutation, selection, and characterization of the Wickerhamomyces anomalus strain YWP1-3 as a starter culture to enhance the flavor quality of Arabica coffee. The results revealed that six mutants could produce relatively high levels of the pectinase enzyme on pectin agar media and exhibited high activity levels, ranging from 332.35 to 415.88 U/ml in mucilage broth. Strains UV22-2, UV22-3, UV41-1 and UV32-1 displayed higher levels of amylase activity than did the wild type. The UV22-2 and UV22-3 mutants exhibited the highest pectin degradation indices of 49.22% and 45.97%, respectively, and displayed significantly enhanced growth rates in nitrogen yeast base media supplemented with various sugars; thus, these mutants were evaluated for their ability to serve as a starter for fermentation of Arabica coffee. The cupping scores of coffees derived from UV22-2 and UV22-3 were 83.5 ± 1.5 and 82.0 ± 2.14, respectively. The volatile compounds in the roasted coffee fermented by UV22-2 were analyzed by GC‒MS, which revealed higher levels of furfuryl alcohol and furfuryl acetate than did the other samples. These findings suggested that UV22-2 could be an influential starter culture for Arabica coffee fermentation.

Comparative metabolic profiling of Cabernet Sauvignon wines reveals the potential of different Wickerhamomyces anomalus co-fermented with commercial Saccharomyces cerevisiae

In this paper, five Wickerhamomyces anomalus accumulated 9.1–19.7% less released CO2 than S. cerevisiae in the synthetic must. Untargeted metabolomics was used to analyze co-fermented real Cabernet Sauvignon wines. The results showed that all W. anomalus strains significantly decreased the levels of malic acid and hypoxanthine in co-fermented wines, thereby promoting the process of TCA cycle and purine metabolism, respectively. Furthermore, WG5, WG26, and WG27 strains utilized more amino acid, thus favoring the enhancement of aroma compounds. WG27 strain also could biotransform more odorless metabolites into flavor active compounds, such as phenethyl alcohol, indole and indole-3-acetaldehyde. Notably, WG27 strain significantly up-regulated secondary metabolites by more than 400% compared to the other W. anomalus strains. These secondary metabolites were involved in phenylpropanoid and flavonoid metabolism, tryptophan metabolism and methionine metabolism. In addition, co-fermentation with the WG27 strain also resulted in an increase in antioxidant activities by more than 16.09% compared to the control wine. This suggests that the WG27 strain not only improves the flavor and style of the wine but also enhances its antioxidant properties. The findings can be helpful in expanding general understanding of W. anomalus and its potential applications in the food and beverage industry.

Tripartite interactions comprising yeast-endobacteria systems in the gut of vector mosquitoes.

It is shown that bacteria use yeast as a niche for survival in stressful conditions, therefore yeasts may act as temporary or permanent bacterial reservoirs. Endobacteria colonise the fungal vacuole of various osmotolerant yeasts which survive and multiply in sugar-rich sources such as plant nectars. Nectar-associated yeasts are present even in the digestive system of insects and often establish mutualistic symbioses with both hosts. Research on insect microbial symbioses is increasing but bacterial-fungal interactions are yet unexplored. Here, we have focused on the endobacteria of Wickerhamomyces anomalus (formerly Pichia anomala and Candida pelliculosa), an osmotolerant yeast associated with sugar sources and the insect gut. Symbiotic strains of W. anomalus influence larval development and contribute digestive processes in adults, in addition to exerting wide antimicrobial properties for host defence in diverse insects including mosquitoes. Antiplasmodial effects of W. anomalus have been shown in the gut of the female malaria vector mosquito Anopheles stephensi. This discovery highlights the potential of utilizing yeast as a promising tool for symbiotic control of mosquito-borne diseases. In the present study, we have carried out a large Next Generation Sequencing (NGS) metagenomics analysis including W. anomalus strains associated with vector mosquitoes Anopheles, Aedes and Culex, which has highlighted wide and heterogeneous EB communities in yeast. Furthermore, we have disclosed a Matryoshka-like association in the gut of A stephensi that comprises different EB in the strain of W. anomalus WaF17.12. Our investigations started with the localization of fast-moving bacteria-like bodies within the yeast vacuole of WaF17.12. Additional microscopy analyses have validated the presence of alive intravacuolar bacteria and 16S rDNA libraries from WaF17.12 have identified a few bacterial targets. Some of these EB have been isolated and tested for lytic properties and capability to re-infect the yeast cell. Moreover, a selective competence to enter yeast cell has been shown comparing different bacteria. We suggested possible tripartite interactions among EB, W. anomalus and the host, opening new knowledge on the vector biology.

Quantitative PCR Assay as a Tool for the Detection of Lactobacilli in Sicilian Table Olives Produced at an Industrial Scale

Table olives are an important fermented product of the Mediterranean area consumed all over the world. In our era, the food industry requires a safe and stable final product with desirable characteristics for the consumer. In the present study, two different experimental fermentations (L, with Lactiplantibacillus plantarum strains, and LY, with L. plantarum strains and Wickerhamomyces anomalus strain) were conducted and monitored up to 180 days and compared with a spontaneous fermentation, used as control (C). The safety and stability of table olives were determined by applying a plate count and quantitative real-time PCR (qPCR) approach. Compared with the control sample (C), experimental fermentations showed a faster acidification and a good inhibition rate of spoilage bacteria, indicating the safety of the process. Quantitative PCR data confirmed the abundance of the Lactobacillus group in both experimental table olives, confirming the importance of the starter cultures for the stability of the final product. In conclusion, the use of starter cultures ensures the safety of industrially produced table olives, and the application of qPCR seems to be a promising tool to detect and quantify lactobacilli as a positive biomarker of table olive fermentation.

Sustainable Bioconversion of Glycerine Pitch into the Novel Wickerhamomyces anomalus Bio-preservative

Background: Yeasts with high protein content are used as single-cell proteins due to their nutritional values and rare pathogenicity. However, the nucleic acid of the yeasts must be removed before consumption to avoid hazards to health. Wickerhamomyces anomalus is an authorized bio-preservative with promising antifungal activity and safety. However, its high protein content associ-ated with high nucleic acid content results in high nitrogen content that imposes additional down-stream processing costs due to the nucleic acid removal step required to prevent uric acid precipitation which leads to various health concerns. Objectives: The objectives were to (i) isolate a novel Wickerhamomyces anomalus strain with low nitrogen content and (ii) to optimize the production of W. anomalus biomass through response surface methodology (RSM). Methods: The novel Wickerhamomyces sp. USMAST-TP1 with low nitrogen content was isolated from fermented food and its biomass was optimized through RSM. Results: Wickerhamomyces sp. USMAST-TP1 showed promising tolerance to glycerine pitch with a higher composition of growth-inhibiting impurities where its biomass was not negatively affected by excessive glycerine pitch supply. Upon optimization through RSM, 11.4 g/L biomass harboring pro-tein content of 13% was achieved. Conclusion: The >17-fold lower nitrogen content of 2% indicated low nucleic acid content compared to common W. anomalus strains, thus the costly nucleic acid purification steps can be excluded, easing applications in agro-food industries.

The Ability of the Yeast Wickerhamomyces anomalus to Hydrolyze Immunogenic Wheat Gliadin Proteins.

Gliadins proteins make up around 30% of total wheat flour proteins. They are involved in many immune disorders affecting an increasing number of people who eat foods made with wheat flour. The triggering factor is the accumulation in the gut of immunogenic peptides derived from incomplete degradation of gliadins by gastric proteases. Previous research has revealed the effectiveness of sourdough-fermentation technology or related lactic acid bacteria in reducing wheat flour allergenic proteins. However, there are no single yeast cultures for producing reduced allergenicity wheat products. This study evaluated sourdough-related yeast Wickerhamomyces anomalus strains for their ability to hydrolyze gliadin proteins. All yeast strains were able to degrade gliadins and use them as carbon and nitrogen sources. The proliferation of the yeast strains depended on the gliadin addition; complete hydrolysis was observed after 24 h. The strain showing higher proteolytic activity fermented, acceptably wheat flour dough. The gliadin content of the leavened dough was reduced by 50%. Bread made from the W. anomalus-fermented dough showed a 78% reduction in immunogenic α-gliadins. 50% of the decrease was attributed to the proteolytic activity of the yeast cells, and the other 35% to the baking process. These results show the potential of the yeast W. anomalus as a starter for reducing immunogenicity wheat products.

Dynamic Transcriptome Analysis Reveals Transcription Factors Involved in the Synthesis of Ethyl Acetate in Aroma-Producing Yeast.

Ethyl acetate is an important flavor element that is a vital component of baijiu. To date, the transcription factors that can help identify the molecular mechanisms involved in the synthesis of ethyl acetate have not been studied. In the present study, we sequenced and assembled the Wickerhamomyces anomalus strain YF1503 transcriptomes to identify transcription factors. We identified 307 transcription factors in YF1503 using high-throughput RNA sequencing. Some transcription factors, such as C2H2, bHLH, MYB, and bZIP, were up-regulated, and these might play a role in ethyl acetate synthesis. According to the trend of ethyl acetate content, heat map results and STEM, twelve genes were selected for verification of expression levels using quantitative real-time PCR. This dynamic transcriptome analysis presents fundamental information on the transcription factors and pathways that are involved in the synthesis of ethyl acetate in aroma-producing yeast. Of significant interest is the discovery of the roles of various transcription factor genes in the synthesis of ethyl acetate.

Comparison of potential Wickerhamomyces anomalus to improve the quality of Cabernet Sauvignon wines by mixed fermentation with Saccharomyces cerevisiae

Wine produced by mixed-yeast-starters of non-Saccharomyces strains and Saccharomyces cerevisiae proved to have distinctive aroma. To find more potential mixed-yeast-starters which can enhance sensory characteristics, five Wickerhamomyces anomalus strains were isolated in this paper. They were combined with S. cerevisiae respectively to co-ferment five wines. The aroma diversity and taste characteristics (intensity, astringency, complexity, and persistence) of three co-fermented wines were improved. Sc-WaG5 and Sc-WaG26 wines had higher concentrations of esters (acetate esters, ethyl esters and other esters). Sc-WaG27 wine showed highest contents of titratable acidity, total phenolic, total tannin and total anthocyanin. Additionally, it had higher level of norisoprenoids (ionone and damascenone), carbonyl compounds (2-octanone and nonanal), higher alcohols (isopentanol) and esters. Therefore, Sc-WaG27 wine possessed more complex aroma and highest sensory score. Our results highlighted that co-fermentation of the three W. anomalus strains with S. cerevisiae is a potential strategy to improve wine flavor and aroma, and enrich wine styles.

Efficacy and mechanism of action of food isolated yeasts in the control of Aspergillus flavus growth on pistachio nuts

Due to its nutritional characteristics, dried fruit and in particular pistachio is considered an important component in the daily diet. Unfortunately, pistachio nuts can be contaminated during storage with a wide range of pathogenic fungi, including Aspergillus flavus. The present work has evaluated how the use of two yeast strains belonging to the species Wickerhamomyces anomalus and Metschnikowia pulcherrima can inhibit the in vitro and in vivo growth of A. flavus. Both yeast strains demonstrated a good in vitro antifungal activity attributable to a specific mechanism of action, although higher efficacy was evidenced by W. anomalus strain. The production of volatile organic compounds (VOCs) and lytic enzymes was hypothesized as the main mechanisms of action exerted by W. anomalus, while the nutritional competition for iron was assumed as the main biocontrol mechanism for M. pulcherrima. Moreover, from the results of the in vivo test carried out on artificially infected pistachio seeds, it was clear as M. pulcherrima strain showed the same efficacy of W. anomalus in inhibiting the growth and sporulation of A. flavus mold, despite W. anomalus was the most effective strain during in vitro assay. Altogether, these results indicate that both strains could be considered as potential biocontrol agents against A. flavus fungal growth, notwithstanding it is always important considering the tritrophic interaction (yeast-mold- host), which could play a crucial role in determining the final results.

Impact of Non-Saccharomyces Wine Yeast Strains on Improving Healthy Characteristics and the Sensory Profile of Beer in Sequential Fermentation.

The use of non-Saccharomyces yeasts in brewing is a useful tool for developing new products to meet the growing consumer demand for innovative products. Non-Saccharomyces yeasts can be used both in single and in mixed fermentations with Saccharomyces cerevisiae, as they are able to improve the sensory profile of beers, and they can be used to obtain functional beers (with a low ethanol content and melatonin production). The aim of this study was to evaluate this capacity in eight non-Saccharomyces strains isolated from Madrid agriculture. For this purpose, single fermentations were carried out with non-Saccharomyces strains and sequential fermentations with non-Saccharomyces and the commercial strain SafAle S-04. The Wickerhamomyces anomalus strain CLI 1028 was selected in pure culture for brewing beer with a low ethanol content (1.25% (v/v)) for its fruity and phenolic flavours and the absence of wort flavours. The best-evaluated strains in sequential fermentation were CLI 3 (Hanseniaspora vineae) and CLI 457 (Metschnikowia pulcherrima), due to their fruity notes as well as their superior bitterness, body, and balance. Volatile compounds and melatonin production were analysed by GC and HPLC, respectively. The beers were sensory-analysed by a trained panel. The results of the study show the potential of non-Saccharomyces strains in the production of low-alcohol beers, and as a flavour enhancement in sequential fermentation.

Effect of Dough-Related Parameters on the Antimold Activity of Wickerhamomyces anomalus Strains and Mold-Free Shelf Life of Bread

The aim of the present study was to assess the antimold capacity of three Wickerhamomyces anomalus strains, both in vitro and in situ, and to identify the responsible volatile organic compounds. For that purpose, two substrates were applied; the former included brain heart infusion broth, adjusted to six initial pH values (3.5, 4.0, 4.5, 5.0, 5.5, 6.0) and supplemented with six different NaCl concentrations (0.0%, 0.5%, 1.0%, 1.5%, 2.0%, 2.5%), while the latter was a liquid dough, fortified with the six aforementioned NaCl concentrations. After a 24 h incubation at 30 °C, the maximum antimold activity was quantified for all strains at 5120 AU/mL, obtained under different combinations of initial pH value and NaCl concentration. A total of twelve volatile compounds were detected; ethanol, ethyl acetate, isoamyl alcohol and isoamyl acetate were produced by all strains. On the contrary, butanoic acid-ethyl ester, acetic acid-butyl ester, ethyl caprylate, 3-methyl-butanoic acid, 2,4-di-tert-butyl-phenol, benzaldehyde, nonanal and octanal were occasionally produced. All compounds exhibited antimold activity; the lower MIC was observed for 2,4-di-tert-butyl-phenol and benzaldehyde (0.04 and 0.06 μL/mL of headspace, respectively), while the higher MIC was observed for butanoic acid-ethyl ester and ethyl caprylate (5.14 and 6.24 μL/mL of headspace, respectively). The experimental breads made with W. anomalus strains LQC 10353, 10346 and 10360 gained an additional period of 9, 10 and 30 days of mold-free shelf life, compared to the control made by commercially available baker’s yeast. Co-culture of the W. anomalus strains with baker’s yeast did not alter the shelf-life extension, indicating the suitability of these strains as adjunct cultures.

Killer yeasts isolated from olive brines: Technological and probiotic aptitudes

Table olive brines, inoculated with six different starters of lactic acid bacteria (LAB) or spontaneously fermented, have been used as isolating source of killer yeasts throughout the fermentation process (120 d). Killer yeast isolates were identified and evaluated for technological and probiotic traits. Although the count of yeast population did not markedly vary among the different vessels and over time, the killer yeast phenotype was mainly present in yeast strains isolated from spontaneous fermentation; the number of killer isolates decreased over fermentation time. Killer phenotype was found in species identified as Pichia kluyveri, Zygoascus hellenicus, Wickerhamomyces anomalus, Pichia membranifaciens, Candida boidinii, Candida diddensiae and Saccharomyces cerevisiae. Among all tested isolates, W. anomalus strains evidenced the widest spectrum of enzymatic activities and the highest β-glucosidase and phtytase activity. These strains evidenced also the best growth at low pH and increasing bile salt concentration, when grown at 37 °C, as well as the most constant viability index (%) during in vitro digestion.

Formulation and Safety Tests of a Wickerhamomyces anomalus-Based Product: Potential Use of Killer Toxins of a Mosquito Symbiotic Yeast to Limit Malaria Transmission.

Wickerhamomyces anomalus strain WaF17.12 is a yeast with an antiplasmodial property based on the production of a killer toxin. For its symbiotic association with Anopheles mosquitoes, it has been proposed for the control of malaria. In an applied view, we evaluated the yeast formulation by freeze-drying WaF17.12. The study was carried out by comparing yeast preparations stored at room temperature for different periods, demonstrating that lyophilization is a useful method to obtain a stable product in terms of cell growth reactivation and maintenance of the killer toxin antimicrobial activity. Moreover, cytotoxic assays on human cells were performed, showing no effects on the cell viability and the proinflammatory response. The post-formulation effectiveness of the killer toxin and the safety tests indicate that WaF17.12 is a promising bioreagent able to impair the malaria parasite in vector mosquitoes.

Assessment of chemical constitution and aroma properties of kiwi wines obtained from pure and mixed fermentation with Wickerhamomyces anomalus and Saccharomyces cerevisiae.

To improve the aroma of kiwi wine through the utilization of Wickerhamomyces anomalus, kiwi juice was fermented using a selected W. anomalus strain in pure culture and mixed fermentations with Saccharomyces cerevisiae, which was inoculated simultaneously and sequentially. The physicochemical indices, volatile compounds and aroma properties of the kiwi wines were assessed. The study suggested that the ethanol, color indices and organic acids of the wines were closely related to the method of inoculation. Compared with the pure S. cerevisiae fermentation, the mixed fermentations produced more varieties and concentrations of volatiles. The sequential fermentations increased the concentrations of esters and terpenes, improving the flower and sweet fruit notes of the wines. The simultaneous inoculation enhanced the contents of esters and aldehydes, intensifying the flower, sweet and sour fruit of the wines. Partial least-squares regression analysis showed that esters and terpenes contributed greatly to the flower and sweet fruit aroma, whereas aldehydes were the major contributors to the sour note. Based on our results, the mixed fermentations not only enriched the types and concentrations of volatiles, but also had better sensory properties. © 2021 Society of Chemical Industry.

Improvement of whole wheat dough fermentation for steamed bread making using selected phytate-degrading Wickerhamomyces anomalus P4

Whole wheat steamed bread is highly recommended because of its health benefits. However, the presence of phytate negatively influences dough properties, because it binds minerals and proteins. In this study, a yeast strain with phytate-degrading activity, namely, Wickerhamomyces anomalus P4 was isolated from whole wheat dough fermented with Jiaozi. The yeast's ability to degrade phytate during whole wheat dough fermentation and its suitability for steamed bread making process were evaluated. W. anomalus P4 showed strong phytate degradation ability, and more than 45% of the initial phytate was degraded after 12 h of whole wheat dough fermentation. Degradation of phytate by W. anomalus P4 was fast in the first 4 h and then slowed gradually with time. The use of the selected W. anomalus P4 as a starter improved dough microstructure and increased mineral solubilization in comparison with commercial Saccharomyces cerevisiae. W. anomalus P4 fermented steamed bread had higher lightness and height/diameter ratio, and similar protein digestibility as the steamed bread prepared by commercial S. cerevisiae. Therefore, phytate-degrading W. anomalus strain P4 could potentially provide benefits by decreasing the phytate content in whole wheat dough for steamed bread making.

Isolation, Identification and Function of Pichia anomala AR2016 and Its Effects on the Growth and Health of Weaned Pigs.

Simple SummaryPichia anomala is a simple pseudomycelium with a round cell shape, product killer factors, β-1,3-glucanase, and phytase. P. anomala had the functions of anticorrosion, improving the nutritive value of feed, increasing the protein content, and reducing the concentration of nutritive substances. In this study, P. anomala AR2016 is isolated and identified from solid wine koji, and its culture conditions are optimized. Heat tolerance, bile salt tolerance, and artificial gastric and intestinal juice tolerance are evaluated. In our methodology, thirty weaned pigs were randomly divided into three groups with 10 barrows in each, and fed a maize-soybean meal diet and orally administered 0.85% saline (CK), 1 mL 1 × 109 cfu/mL Candida utilis (C. utilis), and 1 mL 1 × 109 cfu/mL P. anomala once daily for 28 days (P. anomala). Our results show that P. anomala AR2016 grew best in yeast extract peptone dextrose medium with pH 5.0 at 28 °C, 180 r/min and could tolerate 45 °C for 0.5 h, 0.2% pig bile salts, simulated gastric fluid, and 1.0% simulated intestinal fluid. Our study indicates that P. anomala AR2016 can tolerate high acidity and high bile salts, and has high survivability in the artificial gastric intestinal juice environment. In conclusion, oral administration of P. anomala AR2016 improves the growth performance, reduces the incidence of diarrhea, enhances intestinal barrier function, and improves microflora in weaned pigs.This study isolates and identifies Pichia anomala (P. anomala) AR2016, and studies its effect on the growth and health of weaned pigs. A P. anomala strain from solid wine koji is isolated and identified using 26S rDNA analysis, and its culture conditions are optimized. Heat tolerance, bile salt tolerance, artificial gastric, and intestinal juice tolerance are evaluated. In our methodology, thirty 28 d Large White × Landrace × Rongchang weaned pigs were randomly divided into three groups with 10 barrows in each, and fed a maize-soybean meal diet and orally administered 0.85% saline (CK), 1 mL 1 × 109 cfu/mL Candida utilis (C. utilis), and 1 mL 1 × 109 cfu/mL P. anomala once daily for 28 days. A P. anomala strain was identified and named P. anomala AR2016. P. anomala AR2016 grew best in yeast extract peptone dextrose medium with pH 5.0 at 28 °C, 180 r/min and could tolerate 45 °C for 0.5 h, 0.2% pig bile salts, simulated gastric fluid, and 1.0% simulated intestinal fluid. Our results show that compared with the CK group, orally administered P. anomala AR2016 increases average daily gain, the ileal villus height, the ileal mucosal concentrations of occludin and zonula occluens-1, the serum glucose and total protein concentration, total superoxide dismutase, glutathione peroxidase, and total antioxidative capacity activity, the trypsin and lipase activity in jejunal and ileal contents, the jejunal and ileal mucosa mRNA levels of ALP, TNF-α, and TLR-2, and the relative abundance of Bacteroidetes, Actinobacteria, Succinivibrionaceae, Lachnospiraceae, and Prevotellaceae (p < 0.05). Compared with the CK group, oral administration of P. anomala AR2016 decreased the incidence of diarrhea, aspartate aminotransferase activity, alanine amino-transferase-activity, malondialdehyde, D-lactic acid and endotoxin content in serum, the mRNA level of aminopeptidase N of ileum mucosa, and the relative abundance of Proteobacteria, Clostridiaceae, Campylobacteraceae, Vibrionaceae, Bacillus, and Pseudon (p < 0.05). Collectively, the study indicates that P. anomala AR2016 can tolerate high acidity and high bile salts, and has high survivability in the artificial gastric intestinal juice environment. Oral administration of P. anomala AR2016 improves the growth performance, reduces the incidence of diarrhea, enhances intestinal barrier function, and improves microflora in weaned pigs.

Microbial Application to Improve Olive Mill Wastewater Phenolic Extracts.

Olive mill wastewater (OMW) contains valuable and interesting bioactive compounds, among which is hydroxytyrosol, which is characterized by a remarkable antioxidant activity. Due to the health claims related to olive polyphenols, the aim of this study was to obtain an extract from OMW with an increased level of hydroxytyrosol by means of microbial enzymatic activity. For this purpose, four commercial adsorbent resins were selected and tested. The beta-glucosidase and esterase activity of strains of Wickerhamomyces anomalus, Lactiplantibacillus plantarum, and Saccharomyces cerevisiae were also investigated and compared to those of a commercial enzyme and an Aspergillus niger strain. The W. anomalus strain showed the best enzymatic performances. The SP207 resin showed the best efficiency in selective recovery of hydroxytyrosol, tyrosol, oleuropein, and total phenols. The bioconversion test of the OMW extract was assessed by using both culture broths and pellets of the tested strains. The results demonstrated that the pellets of W. anomalus and L. plantarum were the most effective in hydroxytyrosol increasing in phenolic extract. The interesting results suggest the possibility to study new formulations of OMW phenolic extracts with multifunctional microorganisms.