Yeast Cell Wall Research Articles

P4-ATPase subunit Cdc50 plays a role in yeast budding and cell wall integrity in Candida glabrata

BackgroundAs highly-conserved types of lipid flippases among fungi, P4-ATPases play a significant role in various cellular processes. Cdc50 acts as the regulatory subunit of flippases, forming heterodimers with Drs2 to translocate aminophospholipids. Cdc50 homologs have been reported to be implicated in protein trafficking, drug susceptibility, and virulence in Saccharomyces cerevisiae, Candida albicans and Cryptococcus neoformans. It is likely that Cdc50 has an extensive influence on fungal cellular processes. The present study aimed to determine the function of Cdc50 in Candida glabrata by constructing a Δcdc50 null mutant and its complemented strain.ResultsIn Candida glabrata, the loss of Cdc50 led to difficulty in yeast budding, probably caused by actin depolarization. The Δcdc50 mutant also showed hypersensitivity to azoles, caspofungin, and cell wall stressors. Further experiments indicated hyperactivation of the cell wall integrity pathway in the Δcdc50 mutant, which elevated the major cell wall contents. An increase in exposure of β-(1,3)-glucan and chitin on the cell surface was also observed through flow cytometry. Interestingly, we observed a decrease in the phagocytosis rate when the Δcdc50 mutant was co-incubated with THP-1 macrophages. The Δcdc50 mutant also exhibited weakened virulence in nematode survival tests.ConclusionThe results suggested that the lipid flippase subunit Cdc50 is implicated in yeast budding and cell wall integrity in C. glabrata, and thus have a broad influence on drug susceptibility and virulence. This work highlights the importance of lipid flippase, and offers potential targets for new drug research.

Anti-oxidative and anti-aging effects of mannoprotein-rich yeast cell wall enzymatic hydrolysate by modulating gut microbiota and metabolites in Caenorhabditis elegans

In this study, antioxidant and anti-aging studies were carried out by mannoprotein-rich yeast cell wall enzymatic hydrolysate (MYH) obtained by enzymatic hydrolysis of yeast cell wall through the Caenorhabditis elegans (C. elegans) model. It was found that MYH could improve the lifespan and anti-stress ability of C. elegans by increasing the activity of antioxidant enzymes such as T-SOD, GSH-PX and CAT, and reducing the levels of MDA, ROS and apoptosis. At the same time, through the verification expression of corresponding mRNA, it was found that MYH exerted antioxidant and anti-aging activities by up-regulating the translation of MTL-1, DAF-16, SKN-1 and SOD-3 mRNA, and down-regulating the translation of AGE-1 and DAF-2 mRNA. In addition, it was found that MYH could improve the composition and distribution of the gut microbiota of C. elegans, and significantly improve the level of metabolites through the sequencing of gut microbiota and untargeted metabolomic studies. It has contributed to studying the antioxidant and anti-aging activities of microorganisms such as yeast through the level of gut microbiota and metabolites and the development of related functional foods.

High-Purity Bioactive Ingredient-3S,3'S-Astaxanthin: A New Preparation from Genetically Modified Kluyveromyces marxianus without Column Chromatography and Gel Filtration.

A highly efficient methodology for bioactive ingredient 3S,3'S-astaxanthin (3S,3'S-AST) preparation from genetically modified yeast (Kluyveromyces marxianus) with a combination of enzyme-assisted extraction and salt-assisted liquid-liquid extraction (SALLE) was achieved. The highest yield of 3S,3'S-AST indicated that FoodPro® CBL for yeast cell walls hydrolysis could significantly enhance extraction and obtain, with the help of SALLE procedure, quantified 3S,3'S-AST over 99% in purity through cation chelation. In the oxygen radical antioxidant capacity (ORAC) assay, the antioxidant capacity of high-purity 3S,3'S-AST products were 18.3 times higher than that of the original raw material extract. This new combination preparation may replace previous methods and has the potential to be scaled up in the manufacture of high-purity 3S,3'S-AST from low-value bioresources of raw materials to high-value products in the food and/or drug industries with lower cost and simple equipment.

Evaluation of the effects of dietary mycotoxin-degrading adsorbent on juvenile turbot (Scophthalmus maximus L.) fed aflatoxin B1-contaminated diets

This eight-week feeding trial was conducted to investigate whether adding yeast cell wall extract mixed with bacteria producing mycotoxin-degrading enzymes (YEMDE) to an aflatoxin B1 (AFB1)-contaminated diet affects the growth performance, liver health, antioxidant status, and intestinal microbiota status of juvenile turbot. A total of 480 fish were divided into four groups (three replicate tanks each group): basal diet (Control); Control diet supplemented with 100 μg/kg AFB1 (AFB1); and AFB1 supplemented with 500 mg/kg or 1000 mg/kg YEMDE (AFY500 and AFY1000, respectively). The results showed that a minor variation in growth performance was observed among all groups (P > 0.05). Compared to the Control diet, the AFB1 diet significantly deceased the activities of triglyceride and catalase in the serum (P < 0.05); Compared to the Control diet, the AFB1 diet significantly increased the gene expression of cyp3a and apoptosis-related genes (p53, Bax, caspase3, caspase7 and caspase9) (P < 0.05), reduced the expression of catalase in the liver (P < 0.05); In addition, the AFB1 diet negatively changed the communities of intestinal microbiota and down-regulated the abundance of some potential probiotics Bifidobacterium and Stenotrophomonas (P < 0.05). However, both the AFY500 and AFY1000 diets markedly recovered the antioxidant capacity, inhibited the apoptosis of hepatocytes, and improved the communities of intestinal microbiota (P < 0.05). More positive effects were observed in the AFY1000 group. Compared to the AFB1 diet, the AFY1000 diet markedly increased the abundance of some potential beneficial bacteria in the intestine, such as Faecalibacterium and Lachnoanaerobaculum (P < 0.05). In conclusion, dietary YEMDE can effectively reduce the harmful impacts caused by AFB1 on turbot, and in particular, it had beneficial effects on the communities of intestinal microbiota. It is promising to use YEMDE to control AFB1 contamination in fish feeds.

Dietary yeast cell wall enhanced intestinal health of broiler chickens by modulating intestinal integrity, immune responses, and microbiota

This study was conducted to determine the effects of dietary yeast cell wall (YCW) on growth performance, intestinal health, and immune responses of broiler chickens. In a randomized completely block design (block: initial body weight), a total of 800 broilers (Ross 308; 45.18 ± 3.13 g of initial body weight) were assigned to 2 dietary treatments (40 birds/pen; 10 replicates/treatment) and fed for 5 wk: 1) a basal broiler diet based on corn-soybean meal (CON) and 2) CON+0.05% dietary YCW. Growth performance was measured at intervals in 3 phase feed program. On the final day of the study, one bird per pen was randomly selected and euthanized for sample collection. Broilers fed YCW had decreased (P < 0.05) feed conversion ratio during the grower phase compared with those fed CON. The YCW increased (P < 0.05) villus height to crypt depth ratio in the duodenum, jejunum, and ileum compared with the CON. In addition, the YCW tended to higher (P < 0.10) number of goblet cells in the duodenum than in the CON. Broilers fed YCW had increased (P < 0.05) serum TGF- β1, ileal gene expression of the claudin family, and relative abundance of Lactobacillus, Prevotella, and Enterococcus compared with the CON, but decreased serum TNF-α (P < 0.05), IL-1β (P < 0.05), and IL-6 (P < 0.10), ileal gene expression of IL-6 (P < 0.05), and relative abundance of Clostridium (P < 0.05). The present study demonstrated that the addition of dietary YCW in broiler diets enhanced the intestinal health of broiler chickens and may be associated with modulated intestinal morphology and integrity by upregulating tight junction-related protein gene expression and modifying the ileal microbiota. In addition, dietary YCW modulated immune responses and inflammatory cytokine gene expression in the ileum.

Candida parapsilosis cell wall proteins-CPAR2_404800 and CPAR2_404780-Are adhesins that bind to human epithelial and endothelial cells and extracellular matrix proteins.

One of the initial steps necessary for the development of Candida infections is the adherence to the host tissues and cells. Recent transcriptomic studies suggest that, in Candida parapsilosis-a fungal infectious agent that causes systemic candidiasis in immunosuppressed individuals-the adhesion is mediated by pathogen cell-exposed proteins belonging to the agglutinin-like sequence (Als) family. However, to date, the actual interactions of individual members of this family with human cells and extracellular matrix (ECM) have not been characterized in detail. In the current study, we focused attention on two of these C. parapsilosis Als proteins-CPAR2_404800 and CPAR2_404780-that were proteomically identified in the fungal cell wall of yeasts grown in the media suitable for culturing human epithelial and endothelial cells. Both proteins were extracted from the cell wall and purified, and using a microplate binding assay and a fluorescence microscopic analysis were shown to adhere to human cells of A431 (epithelial) and HMEC-1 (endothelial) lines. The human extracellular matrix components that are also plasma proteins-fibronectin and vitronectin-enhanced these interactions, and also could directly bind to CPAR2_404800 and CPAR2_404780 proteins, with a high affinity (KD in a range of 10-7 to 10-8 M) as determined by surface plasmon resonance measurements. Our findings highlight the role of proteins CPAR2_404800 and CPAR2_404780 in adhesion to host cells and proteins, contributing to the knowledge of the mechanisms of host-pathogen interactions during C. parapsilosis-caused infections.

Improvement of Probiotic Viability by Mixing with Ultrasound-Treated Yeast Cells and Spray Drying.

The objective of the study was to determine the efficacy of ultrasound-treatmentSaccharomyces cerevisiae and spray drying in preserving the viability of Lactiplantibacillus plantarum. The combination of ultrasound-treated S. cerevisiae and L. plantarum was evaluated. Next, the mixture was blended with maltodextrin and either Stevia rebaudiana-extracted fluid, prior to undergoing spray drying. The L. plantarum viability was assessed after the spray drying process, during storage, and in simulated digestive fluid (SDF) conditions. The results showed that the impact of ultrasound caused the crack and holes in the yeast cell wall. Besides, the moisture content values were not significantly different in all samples after spray drying. Although the amount of powder recovery in stevia-supplemented samples was not higher than that of the control sample, the L. plantarum viability was significantly improved after the spray drying process. The density of L. plantarum tended to be stable during the first 30days of storage and decreased more rapidly after that. The results reveal that there was no statistically significant difference in the trend of the samples before and after storage. In the SDF test, the L. plantarum viability mixing with ultrasound-treated yeast cells in the spray drying samples was significantly improved. Besides, the presence of Stevia showed positive efficiency on the L. plantarum viability. The L. plantarum viability mixing with ultrasound-treated yeast cells and stevia-extracted fluid by spray drying process showed potential application due to making powder form which helped to improve the L. plantarum stability during the storage time.

Fluorescence activated cell sorting and fermentation analysis to study rumen microbiome responses to administered live microbials and yeast cell wall derived prebiotics.

Rapid dietary changes, such as switching from high-forage to high-grain diets, can modify the rumen microbiome and initiate gastrointestinal distress, such as bloating. In such cases, feed additives, including prebiotics and live microbials, can be used to mitigate these negative consequences. Bio-Mos® is a carbohydrate-based prebiotic derived from yeast cells that is reported to increase livestock performance. Here, the responses of rumen bacterial cells to Bio-Mos® were quantified, sorted by flow cytometry using fluorescently-labeled yeast mannan, and taxonomically characterized using fluorescence in situ hybridization and 16S rRNA sequencing. Further, to evaluate the effects of bovine-adapted Bacteroides thetaiotaomicron administration as a live microbial with and without Bio-Mos® supplementation, we analyzed microbial fermentation products, changes to carbohydrate profiles, and shifts in microbial composition of an in vitro rumen community. Bio-Mos® was shown to be an effective prebiotic that significantly altered microbial diversity, composition, and fermentation; while addition of B. thetaiotaomicron had no effect on community composition and resulted in fewer significant changes to microbial fermentation. When combined with Bio-Mos®, there were notable, although not significant, changes to major bacterial taxa, along with increased significant changes in fermentation end products. These data suggest a synergistic effect is elicited by combining Bio-Mos® and B. thetaiotaomicron. This protocol provides a new in vitro methodology that could be extended to evaluate prebiotics and probiotics in more complex artificial rumen systems and live animals.

Fluorescence activated cell sorting and fermentation analysis to study rumen microbiome responses to administered live microbials and yeast cell wall derived prebiotics

Fluorescence activated cell sorting and fermentation analysis to study rumen microbiome responses to administered live microbials and yeast cell wall derived prebiotics

Effects of dietary yeast cell wall supplementation on growth performance, intestinal Campylobacter jejuni colonization, innate immune response, villus height, crypt depth, and slaughter characteristics of broiler chickens inoculated with Campylobacter jejuni at d 21

A study was conducted to assess the effects of a dietary yeast cell wall (YCW) with and without a Campylobacter jejuni (CJ) challenge. A total of 2,240-day-old Ross 708 males were randomly assigned within 8 treatments with a 4×2 factorial design, with 4 diets (negative control, positive control, YCW constant dose (400 g/ton), and YCW step-down dose (800/400/200 g/ton in the starter/grower/finisher diets, respectively) and with and without d 21 CJ oral gavage challenge at 5.2×107 CFU/mL. At d 0, 14, 28, and 41 body weights and feed consumption were measured to determine performance. At d 14, 28, and 42, 8 jejunal and ileal histology samples per treatment were collected for villi morphology measurements. At d 22 and 28 (1- and 7-days postinoculation), 24 ileal tissue samples per treatment were collected for relative gene expression analysis. At d 42, 24 cecal content samples per treatment were collected for CJ enumeration. Finally, on d 44, 96 birds per treatment were processed to determine carcass yield and 16 carcass rinses per treatment were collected to determine CJ prevalence after processing. Diet or inoculation did not impact broiler performance (P > 0.05). Limited differences were observed in intestinal morphology, and villus height and crypt depth were different only in the ileum at d 42 (P=0.0280 and P=0.0162, respectively). At d 1 postinoculation, differences between treatments inoculated with CJ and PBS were observed in the expression of avian beta defensin 10 (AvBD10), interleukin 1ß (IL-1ß), and interleukin 10 (IL-10) (P < 0.05). At d 7 postinoculation, expression of AvBD10, IL-1ß, and IL-10 was similar among all treatments (P > 0.05). At d 42, all birds, regardless the inoculation, had similar levels of CJ recovered from cecal contents (P > 0.05). After processing, carcass yield and CJ prevalence postchilling was similar in all treatments (P > 0.05). Overall, under the conditions of this study, the addition of YCW during a CJ challenge did not have an impact in growth performance, innate immune response, cecal colonization, carcass yield, or CJ prevalence after processing.

Distinct roles of carbohydrate-binding modules in multidomain β-1,3-1,4-glucanase on polysaccharide degradation.

Lam16A is a novel GH16 β-1,3-1,4-lichenase isolated from the genus Caldicellulosiruptor which can utilize untreated carbohydrate components of plant cell walls. Its catalytic module has been characterized that the six carbohydrate-binding modules (CBMs) were queued in the C-terminus, but their roles were still unclear. Here, full-length and CBM-truncated mutants of Lam16A were purified and characterized through heterologous expression in Escherichia coli. The profiles of these proteins, including the enzyme activity, degrading efficiency, substrate-binding affinity, and thermostability, were explored. Full-length Lam16A with six CBMs showed excellent thermostability and the highest activity against barley β-glucan and laminarin with optimum pH of 6.5. The CBMs stimulated degrading ability of the catalytic module, especially against β-1,3(4)-glucan-based polysaccharides. The released products from β-1,3-1,4-glucan by Lam16A or its truncated mutants revealed an endo-type glycoside hydrolase. Lam16As exhibited strong binding affinities to the insoluble polysaccharides, especially Lam16A-1CBM. The degradation of yeast cell walls by Lam16A enzyme solution relative to the control reduced the absorbance values at OD800 by ~ 85% ± 1.2, enabling the release of up to ~ 0.057 ± 0.0039µg/mL of the cytoplasmic protein into the supernatant, lowering the viability of the cells by ~ 70.3% ± 6.9, thus causing significant damage in the cell wall structure. Taken together, CBMs could influence the substrate specificity, thermal stability, and binding affinity of β-1,3-1,4-glucanase. These results demonstrate the great potential of these enzymes to promote the bioavailability of β-1,3-glucan oligosaccharides for health benefits. KEY POINTS: • Carbohydrate-binding modules strongly influenced the enzyme activity and binding affinity, and further impacted glycoside hydrolase activity. • Lam16A enzymes have sufficient ability to hydrolyze β-1,3-1,4-glucan-based polysaccharides. • Lam16As provide a powerful tool to promote the bioavailability of β-1,3-glucan oligosaccharides.

Optimal Extraction and Deproteinization Method for Mannoprotein Purification from Kluyveromyces marxianus.

Mannoproteins, mannose-glycosylated proteins, play an important role in biological processes and have various applications in industries. Several methods have been already used for the extraction of mannoproteins from yeast cell-wall. The aim of this study was to evaluate the extraction and deproteinization of mannan oligosaccharide from the K. marxianus mannoprotein. To acquire crude mannan oligosaccharides, K. marxianus mannoproteins were deproteinized by the Sevage, TCA, and HCL methods. Total nitrogen, crude protein content, fat, carbohydrate and ash content were measured according to the monograph prepared by the meeting of the Joint FAO/WHO Expert Committee and standard. Mannan oligosaccharide loss, percentage of deproteinization, and chemical composition of the product were assessed to check the proficiency of different methods. Highly purified (95.4%) mannan oligosaccharide with the highest deproteinization (97.33 ± 0.4%) and mannan oligosaccharide loss (25.1 ± 0.6%) were obtained following HCl method. HCl, was the most appropriate deproteinization method for the removal of impurities. This preliminary data will support future studies to design scale-up procedures.

Yeast Cell Wall Compounds on The Formation of Fermentation Products and Fecal Microbiota in Cats: An In Vivo and In Vitro Approach.

The effects of yeast cell wall compounds (YCWs) being added to cat food on hindgut fermentation metabolites and fecal microbiota were assessed in in vivo Experiment 1 (Exp. 1) and in vitro Experiments 2 and 3 (Exp. 2 and 3). In Exp. 1, the cats' diets were supplemented with two dietary concentrations (46.2 and 92.4 ppm) of YCWs (YCW-15 and YCW-30, respectively), and a negative control diet with no compound in three groups (six cats per group) was used to assess the fecal score, pH, digestibility, fermentation products, and microbiota. In Exp. 2, feces from the cats that were not supplemented with YCWs (control) were used as an inoculum. A blend of pectin, amino acids, and cellulose was used as a substrate, and the YCW compound was added at two levels (5 and 10 mg). In Exp. 3, feces from cats fed YCWs were used as an inoculum to test three different substrates (pectin, amino acids, and cellulose). In Exp. 2 and 3, the gas production, pH, and fermentation products (ammonia, SCFAs, and BCFAs) were assessed. YCW-30 resulted in a higher digestibility coefficient of the crude protein, organic matter (OM) (p < 0.05), and energy of the diet (p < 0.10). Regarding the fermentation products, YCW-15 showed a trend toward higher concentrations of propionate, acetate, lactate, ammonia, isobutyrate, and valerate, while YCW-30 showed a trend (p < 0.10) toward higher levels of butyrate and pH values. The bacteroidia class and the genus Prevotella were increased by using YCW-30 and the control. At the gender level, decreased (p < 0.01) Megasphaera was observed with YCW inclusion. The microbiota differed (p < 0.01) among the groups in their Shannon indexes. For beta diversity, YCW-30 showed higher indexes (p = 0.008) than the control. The microbiota metabolic profile differed in the pathway CENTFERM-PWY; it was more expressed in YCW-30 compared to the control. In Exp. 2, the YCWs showed a higher ratio (p = 0.006) of the fermentation products in the treatments with additives with a trend towards a high dose of the additive (10 mg). In Exp. 3, the effects of the substrates (p < 0.001), but not of the YCWs, on the fermentation products were observed, perhaps due to the low dietary concentrations we used. However, the marked responses of the fermentation products to the substrates validated the methodology. We could conclude that the YCWs, even at low dietary concentrations, affected fecal SCFA production, reduced the fecal pH, and modulated the fecal microbiota in the cats. These responses were more pronounced under in vitro conditions.

Titrating Avidity of Yeast-Displayed Proteins Using a Transcriptional Regulator.

Yeast surface display is a valuable tool for protein engineering and directed evolution; however, significant variability in the copy number (i.e., avidity) of displayed variants on the yeast cell wall complicates screening and selection campaigns. Here, we report an engineered titratable display platform that modulates the avidity of Aga2-fusion proteins on the yeast cell wall dependent on the concentration of the anhydrotetracycline (aTc) inducer. Our design is based on a genomic Aga1 gene copy and an episomal Aga2-fusion construct both under the control of an aTc-dependent transcriptional regulator that enables stoichiometric and titratable expression, secretion, and display of Aga2-fusion proteins. We demonstrate tunable display levels over 2-3 orders of magnitude for various model proteins, including glucose oxidase enzyme variants, mechanostable dockerin-binding domains, and anti-PDL1 affibody domains. By regulating the copy number of displayed proteins, we demonstrate the effects of titratable avidity levels on several specific phenotypic activities, including enzyme activity and cell adhesion to surfaces under shear flow. Finally, we show that titrating down the display level allows yeast-based binding affinity measurements to be performed in a regime that avoids ligand depletion effects while maintaining small sample volumes, avoiding a well-known artifact in yeast-based binding assays. The ability to titrate the multivalency of proteins on the yeast cell wall through simple inducer control will benefit protein engineering and directed evolution methodology relying on yeast display for broad classes of therapeutic and diagnostic proteins of interest.

Effects of dietary nucleotide and yeast cell wall on growth performance, feed utilization, anti-oxidative and immune response of grass carp (Ctenopharyngodon idella)

A 70-day feeding trial was conducted to study the effects of dietary nucleotide, yeast cell wall (containing 20% β -glucan) and their combination on growth performance, feed utilization and immune response of grass carp (Ctenopharyngodon idella) with 69.97 ± 0.05 g of initial body weight. Four isonitrogenous (about 38% crude protein) and isolipidic (about 5% crude lipid) diets were established. Based on the control diet (CD), the other three experimental diets were prepared by adding 0.01% of nucleotide (NT), 0.1% of yeast cell wall (YCW) and NT (0.01%) +YCW (0.1%), respectively. Results showed that no significant difference was found in survival of grass carp ranging from 94.44% to 97.78% among all the groups (P > 0.05). Compared with the control group, weight gain rate, muscle crude protein content, serum protein, trypsin and chymotrypsin activities in midgut, lysozyme and immunoglobulin M in serum significantly increased in fish fed the YCW diet (P < 0.05). The significantly highest weight gain rate, villus height and digestive enzyme activities in midgut and innate immune parameters in serum were found in fish fed the NT + YCW diet (P < 0.05). The gene expressions of β-defensin, hepcidin, il-10 and tgf-β1 in the midgut, and tor and s6k1 in liver significantly increased in fish fed the NT + YCW diet. Meanwhile, the gene expressions of il-1β and tnf-α in the midgut decreased significantly (P < 0.05). The liver histology showed the better development in dietary NT and/or YCW supplemented groups than those in the control group. In conclusion, combination of dietary NT and YCW had significantly synergetic improvements on the growth, feed utilization, digestive enzymes, innate immunity and histology of midgut and liver of grass carp.

Comparative Assessment of Different Yeast Cell Wall-Based Mycotoxin Adsorbents Using a Model- and Bioassay-Based In Vitro Approach.

Frequently reported occurrences of deoxynivalenol (DON), beauvericin (BEA), and, to a lesser extent, ochratoxin A (OTA) and citrinin (CIT) in ruminant feed or feedstuff could represent a significant concern regarding feed safety, animal health, and productivity. Inclusion of yeast cell wall-based mycotoxin adsorbents in animal feeds has been a common strategy to mitigate adverse effects of mycotoxins. In the present study, an in vitro approach combining adsorption isotherm models and bioassays was designed to assess the efficacy of yeast cell wall (YCW), yeast cell wall extract (YCWE), and a postbiotic yeast cell wall-based blend (PYCW) products at the inclusion rate of 0.5% (w/v) (ratio of adsorbent mass to buffer solution volume). The Hill's adsorption isotherm model was found to best describe the adsorption processes of DON, BEA, and CIT. Calculated binding potential for YCW and YCWE using the Hill's model exhibited the same ranking for mycotoxin adsorption, indicating that BEA had the highest adsorption rate, followed by DON and CIT, which was the least adsorbed. PYCW had the highest binding potential for BEA compared with YCW and YCWE. In contrast, the Freundlich isotherm model presented a good fit for OTA adsorption by all adsorbents and CIT adsorption by PYCW. Results indicated that YCW was the most efficacious for sequestering OTA, whereas YCWE was the least efficacious. PYCW showed greater efficacy at adsorbing OTA than CIT. All adsorbents exhibited high adsorption efficacy for BEA, with an overall percentage average of bound mycotoxin exceeding 60%, whereas moderate efficacies for the other mycotoxins were observed (up to 37%). Differences in adsorbent efficacy of each adsorbent significantly varied according to experimental concentrations tested for each given mycotoxin (p < 0.05). The cell viability results from the bioassay using a bovine mammary epithelial cell line (MAC-T) indicated that all tested adsorbents could potentially mitigate mycotoxin-related damage to bovine mammary epithelium. Results from our studies suggested that all tested adsorbents had the capacity to adsorb selected mycotoxins in vitro, which could support their use to mitigate their effects in vivo.

Effect of Adding an Organic Binder on Health of Cows Fed with Mycotoxins Contaminated Diet

Mycotoxins contamination occurring in dairy cow's diet is responsible for devastating effects on livestock health. Among different strategies, using organic adsorbents is a promising approach to reduce the toxicity of mycotoxins. This study investigated the effects of an organic adsorbent containing Lactobacillus brevis TD4, Lactobacillus paracasei TD3, and Saccharomyces cerevisiae cell wall on milk production, somatic cell count, blood parameters (white blood cell [WBC], lymphocyte [LYM], neutrophil, basophil, monocyte, eosinophil, red blood cell, haemoglobin, haematocrit, mean corpuscular volume, mean corpuscular haemoglobin) and liver function (alanine transaminase [ALT], gamma-glutamyl transferase [GGT], aspartate transaminase [AST], alkaline phosphatase [ALP], urea, total protein, albumin) in dairy cows fed a naturally contaminated diet with Zearalenone and Deoxynivalenol. The amounts of mycotoxins Zearalenone (ZEA) and Deoxynivalenol (DON) in feed ingredients were measured using the HPLC method. Ten Holstein dairy cows received organic adsorbents daily in their diets for four weeks. Milk and blood samples were taken from cows before the start of feeding adsorbent (CTRL), during the feeding period (FP), and one week after removal of adsorbent from the diet (RP). Totally, the amount of measured ZEA and DON toxins in the diet were 389 and 1254.6 ppb, respectively. Feeding of organic adsorbent significantly increased milk production, total serum protein, and albumin compared to CTRL (P&lt;0.01). Also, numerically lower somatic cell count in their milk and a significant decrease (P&lt;0.01) in serum urea were resulted. Among examined blood parameters, the number of WBC and LYM significantly decreased (P&lt;0.01) after feeding with the organic binder in comparison to the control period. Furthermore, except for a significant increase in the level of AST (P&lt;0.05), the other liver function examined parameters were not affected. The consumption of feed containing low-cost organic adsorbent including Lactobacillus bacteria and yeast cell wall can improve the physical condition and health of dairy cows and reduce economic losses in livestock production.

Cecropin D-derived synthetic peptides in the fight against Candida albicans cell filamentation and biofilm formation.

The recent progressive increase in the incidence of invasive fungal infections, especially in immunocompromised patients, makes the search for new therapies crucial in the face of the growing drug resistance of prevalent nosocomial yeast strains. The latest research focuses on the active compounds of natural origin, inhibiting fungal growth, and preventing the formation of fungal biofilms. Antimicrobial peptides are currently the subject of numerous studies concerning effective antifungal therapy. In the present study, the antifungal properties of two synthetic peptides (ΔM3, ΔM4) derived from an insect antimicrobial peptide - cecropin D - were investigated. The fungicidal activity of both compounds was demonstrated against the yeast forms of Candida albicans, Candida tropicalis, and Candida parapsilosis, reaching a MFC99.9 in the micromolar range, while Candida glabrata showed greater resistance to these peptides. The scanning electron microscopy revealed a destabilization of the yeast cell walls upon treatment with both peptides; however, their effectiveness was strongly modified by the presence of salt or plasma in the yeast environment. The transition of C. albicans cells from yeast to filamentous form, as well as the formation of biofilms, was effectively reduced by ΔM4. Mature biofilm viability was inhibited by a higher concentration of this peptide and was accompanied by increased ROS production, activation of the GPX3 and SOD5 genes, and finally, increased membrane permeability. Furthermore, both peptides showed a synergistic effect with caspofungin in inhibiting the metabolic activity of C. albicans cells, and an additive effect was also observed for the mixtures of peptides with amphotericin B. The results indicate the possible potential of the tested peptides in the prevention and treatment of candidiasis.

Identification of the main proteins secreted byKluyveromyces marxianusand their possible roles in antagonistic activity against fungi

Lytic enzymes secreted by Kluyveromyces marxianus can lyse Saccharomyces cerevisiae cells. Their ability to hydrolyze yeast cell walls can be used in biotechnological applications, such as the production of glucans and protoplasts, as well as a biological control agent against plant pathogenic fungi. Herein, 27 proteins secreted by K. marxianus were identified by mass spectrometry analyses. Importantly, 14 out of the 27 proteins were classified as hydrolases. Indeed, the enzyme extract secreted by K. marxianus caused damage to S. cerevisiae cells and reduced yeast cell viability. Moreover, K marxianus inhibited spore germination and mycelial growth of the phytopathogenic fungus Botrytis cinerea in simultaneous cocultivation assays. We suggest that this inhibition may be partially related to the yeast's ability to secrete lytic enzymes. Consistent with the in vitro antagonistic tests, K. marxianus was able to protect strawberry fruits inoculated with B. cinerea. Therefore, these findings suggest that K. marxianus possesses potential as a biocontrol agent against strawberry gray mold during the postharvest stage and may also have potential against other phytopathogenic fungi by means of its lytic enzymatic arsenal.

Evaluation of long-term supplementation of a direct-fed microbial and enzymatically hydrolyzed yeast cell culture product on feedlot growth performance, efficiency of dietary net energy utilization, heat stress measures, and carcass characteristics in beef steers.

The objective of this research was to determine the influence of long-term supplementation (258 d) of a direct-fed microbial (DFM) and yeast cell wall (YCW) product used alone or in combination on growth performance, dietary net energy utilization, and carcass characteristics in beef steers finished under climatic conditions in the Northern Plains (NP). Single-sourced Charolais × Red Angus steers [n = 256; body weight = 246 ± 1.68 kg] were blocked by pen location in a 2 × 2 factorial arrangement of DFM and YCW. Steers were administered a series of diets common to the NP and administered ractopamine hydrochloride (RH; 300 mg/kg) during the last 28 d of the finishing phase. Steers were vaccinated and poured at processing and individually weighed on days 1, 14, 42, 77, 105, 133, 161, 182, 230, and 258. Temperature-humidity index (THI) was calculated during RH supplementation. For 98% of the experiment, the THI was lower than 72 and thus cattle were not under high-ambient temperature. On days 1, 2, 21, and 22 of RH supplementation, respiration rates (RR), and panting scores (PS) were determined before and after AM and PM feedings (0700 h, 1100 h, 1400 h, and 1700 h). A DFM + YCW interaction was noted for the proportion of steers categorized as PS 2.0 at 1100 h on day 21 (P = 0.03) and RR on day 21 at 1400 h (P = 0.02). Control steers had a greater proportion of PS 2.0 compared to DFM or YCW steers (P ≤ 0.05), while DFM + YCW steers did not differ from others (P ≥ 0.05); DFM + YCW steers had greater (P < 0.05) RR compared to DFM steers, while control and YCW steers did not differ from others (P ≥ 0.05). No DFM + YCW interactions or main effects (P ≥ 0.05) were observed for cumulative growth performance measures. However, YCW steers had 2% lower (P = 0.04) dry matter intakes compared to steers not fed YCW. No DFM + YCW interactions or main effects (P ≥ 0.05) were observed for carcass traits or liver abscess severity. However, a DFM + YCW interaction (P < 0.05) was noted for the distribution of USDA yield grade (YG) 1 and Prime carcasses. Control steers had a greater proportion (P < 0.05) of YG 1 carcasses compared to other treatments. DFM+YCW steers had a greater proportion (P < 0.05) of USDA Prime carcasses compared to DFM or YCW but were similar to control steers, which were also similar to DFM or YCW. Overall, the use of DFM and YCW alone or in combination had minimal effects on growth performance, carcass traits, and heat stress measures in steers finished in NP climatic conditions.