Exopolysaccharide Biosynthesis Research Articles

Phenotypic features and analysis of genes supporting probiotic action unravel underlying perspectives of Bacillus velezensis VTX9 as a potential feed additive for swine

PurposeTo date, a total of 13 probiotic Bacillus species are considered as a Generally Recognized as Safe organism (GRAS) approved by the US Federal Food, Drug, and Cosmetic Act (FDCA), which are used for food and feed additives. However, Bacillus velezensis is not considered as a probiotic candidate in swine farming due to a lack of genetic basis of probiotic action-related traits. Therefore, the present study was undertaken to exploit the genetic basis underlying the probiotic traits of B. velezensis VTX9.MethodsThe genome sequencing of B. velezensis VTX9 was performed on a PacBio Sequel platform. The probiotic properties including biosafety, antioxidative capacity, and riboflavin and exopolysaccharide production were evaluated by using genotypic and phenotypic analysis. The secondary metabolite potentials were also predicted.ResultsStrain VTX9 isolated from swine feces proved some probiotic properties including resistance to 3 mM H2O2, 0.6 mM bile salt, low pH, and antipathogenic activity. The complete genome of B. velezensis VTX9 consists of a 3,985,800 bp chromosome that housed 3736 protein-coding genes and 5 plasmids with the size ranging from 7261 to 20,007 bp. Genome analysis revealed no functional genes encoding enterotoxins and transferable antibiotic resistance, which confirmed the safety of VTX9. A total of 82 genes involved in gastrointestinal stress tolerance were predicted, which has not been reported previously. The maximum production of riboflavin reached 769 ± 7.5 ng/ml in LB medium after 72 h, which was in agreement with the complete de novo riboflavin biosynthetic pathway exploited for the first time in the B. velezensis genome. Antagonistic activity against pathogenic bacteria was attributed to 10 secondary metabolites clusters. The presence of a large gene cluster involved in biosynthesis of exopolysaccharides underscored further the adhesion and biofilm-forming capabilities of VTX9 in swine intestines.ConclusionOur results revealed for the first time that B. velezensis VTX9 has the potential to be a probiotic candidate. The information provided here on the genome of B. velezensis VTX9 opens new opportunities for using B. velezensis as a feed additive for swine farming in the future.

Genomic and molecular evidence reveals novel pathways associated with cell surface polysaccharides in bacteria.

Amino acid (acyl carrier protein) ligases (AALs) are a relatively new family of bacterial amino acid adenylating enzymes with unknown function(s). Here, genomic enzymology tools that employ sequence similarity networks and genome context analyses were used to hypothesize the metabolic function(s) of AALs. In over 50% of species, aal and its cognate acyl carrier protein (acp) genes, along with three more genes, formed a highly conserved AAL cassette. AAL cassettes were strongly associated with surface polysaccharide gene clusters in Proteobacteria and Actinobacteria, yet were prevalent among soil and rhizosphere-associated α- and β-Proteobacteria, including symbiotic α- and β-rhizobia and some Mycolata. Based on these associations, AAL cassettes were proposed to encode a noncanonical Acp-dependent polysaccharide modification route. Genomic-inferred predictions were substantiated by published experimental evidence, revealing a role for AAL cassettes in biosynthesis of biofilm-forming exopolysaccharide in pathogenic Burkholderia and expression of aal and acp genes in nitrogen-fixing Rhizobium bacteroids. Aal and acp genes were associated with dltBD-like homologs that modify cell wall teichoic acids with d-alanine, including in Paenibacillus and certain other bacteria. Characterization of pathways that involve AAL and Acp may lead to developing new plant and human disease-controlling agents as well as strains with improved nitrogen fixation capacity.

Isolation, Phylogenetic and Gephyromycin Metabolites Characterization of New Exopolysaccharides-Bearing Antarctic Actinobacterium from Feces of Emperor Penguin.

A new versatile actinobacterium designated as strain NJES-13 was isolated from the feces of the Antarctic emperor penguin. This new isolate was found to produce two active gephyromycin analogues and bioflocculanting exopolysaccharides (EPS) metabolites. Phylogenetic analysis based on pairwise comparison of 16S rRNA gene sequences showed that strain NJES-13 was closely related to Mobilicoccus pelagius Aji5-31T with a gene similarity of 95.9%, which was lower than the threshold value (98.65%) for novel species delineation. Additional phylogenomic calculations of the average nucleotide identity (ANI, 75.9–79.1%), average amino acid identity (AAI, 52.4–66.9%) and digital DNA–DNA hybridization (dDDH, 18.6–21.9%), along with the constructed phylogenomic tree based on the up-to-date bacterial core gene (UBCG) set from the bacterial genomes, unequivocally separated strain NJES-13 from its close relatives within the family Dermatophilaceae. Hence, it clearly indicated that strain NJES-13 represented a putative new actinobacterial species isolated from the gut microbiota of mammals inhabiting the Antarctic. The obtained complete genome of strain NJES-13 consisted of a circular 3.45 Mb chromosome with a DNA G+C content of 67.0 mol%. Furthering genome mining of strain NJES-13 showed the presence of five biosynthetic gene clusters (BGCs) including one type III PKS responsible for the biosynthesis of the core of gephyromycins, and a series of genes encoding for bacterial EPS biosynthesis. Thus, based on the combined phylogenetic and active metabolites characterization presented in this study, we confidently conclude that strain NJES-13 is a novel, fresh actinobacterial candidate to produce active gephyromycins and microbial bioflocculanting EPS, with potential pharmaceutical, environmental and biotechnological implications.

OsaR (PA0056) Functions as a Repressor of the Gene fleQ Encoding an Important Motility Regulator in Pseudomonas aeruginosa.

FleQ plays a crucial role in motility and biofilm formation by regulating flagellar and exopolysaccharide biosynthesis in Pseudomonas aeruginosa. It has been reported that the expression of FleQ is transcriptionally downregulated by the virulence factor regulator Vfr. Here, we demonstrated that a LysR-type transcriptional regulator, OsaR, is also capable of binding to the promoter region of fleQ and repressing its transcription. Through gel shift and DNase I footprinting assays, the OsaR binding site was identified and characterized as a dual LysR-type transcriptional regulator box (AT-N11-AT-N7-A-N11-T). Mutation of the A-T palindromic base pairs in the fleQ promoter not only reduced the binding affinity of OsaR in vitro but also derepressed fleQ transcription in vivo. The OsaR binding site was found to cover the Vfr binding site; knockout of osaR or vfr separately exhibited no effect on the transcriptional level of fleQ; however, fleQ expression was repressed by overexpression of osaR or vfr. Furthermore, simultaneously deleting both osaR and vfr resulted in an upregulation of fleQ, but it could be complemented by the expression of either of the two repressors. In summary, our work revealed that OsaR and Vfr function as two transcriptional repressors of fleQ that bind to the same region of fleQ but work separately. IMPORTANCE Pseudomonas aeruginosa is a widespread human pathogen, which accounts for serious infections in the hospital, especially for lung infection in cystic fibrosis and chronic obstructive pulmonary disease patients. P. aeruginosa infection is closely associated with its motility and biofilm formation, which are both under the regulation of the important transcription factor FleQ. However, the upstream regulatory mechanisms of fleQ have not been fully elucidated. Therefore, our research identifying a novel regulator of fleQ as well as new regulatory mechanisms controlling its expression will be significant for better understanding the intricate gene regulatory mechanisms related to P. aeruginosa virulence and infection.

Genome Mining Associated with Analysis of Structure, Antioxidant Activity Reveals the Potential Production of Levan-Rich Exopolysaccharides by Food-Derived Bacillus velezensis VTX20

Exopolysaccharides (EPSs) produced by Bacillus species have recently emerged as promising commercial antioxidants in various industries, such as pharmaceutics and biomedicine. However, little is known about EPS production and function from Bacillus velezensis so far. In the present study, the effect of sugar sources on EPS production by B. velezensis VTX20 and the genetic biosynthesis, characteristics, and antioxidant activity of the resulting EPS were evaluated. The strain VTX20 produced the maximum EPS yield of 75.5 ± 4.8 g/L from an initial 200 g/L of sucrose after a 48-h cultivation. Through genomic analysis, ls-levB operon was found, for the first time, to be responsible for the levan-type EPS production in B. velezensis. Biochemical and structural characterization further confirmed the majority of levan, followed by an extremely low level of dextran biopolymer. The water solubility index and water holding capacity of the EPSs were 81.9 ± 3.4% and 100.2 ± 3.4%, respectively. In vitro antioxidant activity analyses showed strong scavenging activity for 1,1-diphenyl-2-picrylhydrazyl and hydroxyl radical values of 40.1–64.0% and 16.0–40%, respectively. These findings shed light on the EPS biosynthesis of B. velezensis at both structural and genetic levels and the potential application of EPS as a natural antioxidant for pharmaceutical and biomedical industries.

Correlation between exopolysaccharide biosynthesis and gastrointestinal tolerance of Lactiplantibacillus plantarum.

This study aimed to investigate the correlation between the level of exopolysaccharide (EPS) biosynthesis and gastrointestinal tolerance of 12 Lactiplantibacillus plantarum strains. In this study, the EPS production and survival rate of 12strains of L.plantarum under gastrointestinal stress were determined. Results showed that the EPS biosynthesis level of L.plantarum in semi-defined medium ranged from 9.84 to 26.05mg/L. The survival rates of all strains in simulated gastric juice at pH 3.0 ranged from 43.52% to 112.73%. Among them, eight strains were higher than 90%, while only one strain was lower than 50%. The survival rates of all strains in simulated intestinal juice ranged from 50.36% to 125.39%, among which eight strains were higher than 80%. The survival rates of all strains under 0.1% bile salt stress ranged from 3.39% to 109.34%, among which four strains were higher than 80% and three strains were lower than 60%. Besides, the survival rates of all strains under 0.5% bile salt stress ranged from 0.42% to 95.34%. The results indicated that the 12 L.plantarum strains had good tolerance to simulated gastric juice at pH 3.0, simulated intestinal juice and 0.1% bile salt. Notably, it was observed that the survival rates of L.plantarum strains under simulated gastric juice at pH 3.0 and simulated intestinal juice were significantly positively correlated with EPS biosynthesis (p<0.01). The yield of EPS of L.plantarum was related to simulated gastric juice and simulated intestinal juice environment. It was speculated that the production of EPS may be one of the strategies for L.plantarum to adapt to the part of gastrointestinal environment. In the future, we could analyse the protection mechanism of EPS from the gene level.

Expression of genes involved in exopolysaccharide synthesis in Lactiplantibacillus plantarum VAL6 under environmental stresses.

Environmental factors can alter exopolysaccharide biosynthesis in lactic acid bacteria (LAB). To further clarify this potential relationship, the mRNA expression of genes involved in exopolysaccharide synthesis such as glmU, pgmB1, cps4E, cps4F, cps4J, and cps4H in Lactiplantibacillus plantarum VAL6 under different conditions including temperature, pH, sodium chloride (NaCl), and carbon dioxide (CO2) intensification culture was studied. The transcriptomic data revealed that the exposure of L. plantarum VAL6 at pH 3 increased the expression level of cps4H but decreased the expression levels of pgmB1 and cps4E. Under pH 8, cps4F and cps4E were significantly upregulated, whereas pgmB1 was downregulated. Similarly, the expression levels of cps4F, cps4E, and cps4J increased sharply under stresses at 42 or 47°C. In the case of NaCl stress, glmU, pgmB1, cps4J, and cps4H were downregulated in exposure to NaCl at 7 and 10% concentrations while cps4E and cps4F were upregulated at 1h of 10%-NaCl treatment and at 5h of 4%-NaCl treatment. Remarkably, CO2 intensification culture stimulated the expression of all tested genes. In addition, simultaneous changes in expression of cps4E and cps4F under environmental challenges may elicit the possibility of an association between the two genes. These findings indicated that the expression level of eps genes is responsible for changes in the yield and monosaccharide composition of exopolysaccharides under environmental stresses.

Study of exopolysaccharides from lactic acid bacteria and their industrial applications: a review

SummaryThis review describes and discusses the structure, biosynthesis and applications of exopolysaccharides from lactic acid bacteria. These substances are classified as homopolysaccharides, which are synthesised from sucrose through the action of extracellular glycosyltransferases or heteropolysaccharides, which are synthesised from repeating unit precursors formed in the cytoplasm and assembled extracellularly by the sequential addition of nucleotide sugars. The industrial application of exopolysaccharides is linked to enhancing the texture and rheological properties of certain fermented products and their production in situ being of particular interest. The chemical characteristics of exopolysaccharides influence interactions with milk proteins in fermented dairy products. These compounds reduce gel syneresis and increase the viscosity, water retention capacity and firmness of the gel, all of which are desirable characteristics for the development of low‐fat dairy products. Similarly, they have applications in the production of gluten‐free bakery products and fermented meat products.

Rapid isolation of exopolysaccharide-producing Streptococcus thermophilus based on molecular marker screening.

As a natural food additive, exopolysaccharide (EPS) produced by Streptococcus thermophilus can improve product viscosity and texture. The protein EpsA is a putative pathway-specific transcriptional regulator for EPS biosynthesis in S.thermophilus. According to comparative analysis of EPS biosynthetic gene clusters, a conserved region of epsA (609 bp) was employed to design primer pair epsA-F/R as a molecular marker for the isolation of EPS-producing (EPS+ ) S. thermophilus. Two EPS+ S. thermophiles strains, AR333 and S-3, were band-positive, whereas Lactococcus lactis NZ9000 (non-EPS-producing, EPS- ), Lactobacillus casei LC2W (EPS+ ) and L. plantarum AR113 (EPS+ ) were negative by polymerase chain reaction (PCR) amplicon bands using the epsA probe. This indicated good specificity of the epsA probe to EPS+ S. thermophilus. Moreover, based on PCR screening with the epsA probe, 23 positive strains were isolated and identified as S. thermophilus from our microbial library and natural fermented milk with 141.3-309.2mg L-1 of EPS production, demonstrating the validity of our molecular marker screening method. The designed molecular marker of epsA can rapidly screen EPS+ S. thermophilus, which has potential application in the dairy and other food industries. © 2021 Society of Chemical Industry.

Transcription activation of two clusters for exopolysaccharide biosynthesis by phosphorylated DctD in Vibrio vulnificus.

NtrC-mediated production of exopolysaccharides (EPS), essential components for Vibrio vulnificus biofilms, is highly increased in the presence of dicarboxylic or tricarboxylic acids. Gel-shift assays showed that regulation of the EPS-gene cluster I (EPS-I cluster) by NtrC was direct via binding of phosphorylated NtrC (p-NtrC) to the regulatory region of the EPS-I cluster. In contrast, p-NtrC did not bind to the EPS-II and EPS-III clusters, suggesting that NtrC regulation was not direct and another transcription factor belonging to an NtrC-regulon might play a role in activating their transcription. A candidate transcription factor, DctD, of which expression was induced by NtrC, activated the expression of the EPS-II and EPS-III clusters via direct binding to their upstream regions. Under growth conditions with either dicarboxylic or tricarboxylic acids, the expression of NtrC was induced and the transcription of dctD was activated. Furthermore, DctD exhibited higher transcriptional activity under the conditions with dicarboxylic acids than with tricarboxylic acids. Therefore, this study demonstrates that under dicarboxylate-rich conditions, both the abundance and activity of DctD were markedly induced, which activates the expression of two EPS clusters to maximize biosynthesis of EPS facilitating biofilm maturation in V. vulnificus.

Biosynthesis of a sulfated exopolysaccharide, synechan, and bloom formation in the model cyanobacterium Synechocystis sp. strain PCC 6803.

Extracellularpolysaccharides of bacteria contribute to biofilm formation, stress tolerance, and infectivity. Cyanobacteria, the oxygenic photoautotrophic bacteria, uniquely produce sulfated extracellular polysaccharides among bacteria to support phototrophic biofilms. In addition, sulfated polysaccharides of cyanobacteria and other organisms have been focused as beneficial biomaterial. However, very little is known about their biosynthesis machinery and function in cyanobacteria. Here, we found that the model cyanobacterium, Synechocystis sp. strain PCC 6803, formed bloom-like cell aggregates embedded in sulfated extracellular polysaccharides (designated as synechan) and identified whole set of genes responsible for synechan biosynthesis and its transcriptional regulation, thereby suggesting a model for the synechan biosynthesis apparatus. Because similar genes are found in many cyanobacterial genomes with wide variation, our findings may lead elucidation of various sulfated polysaccharides, their functions, and their potential application in biotechnology.

Relationship between total cell counts and exopolysaccharide production of Streptococcus thermophilus T9 in reconstituted skim milk

The main objective of this study was to establish a correlation between total bacterial count (TBC) and exopolysaccharide (EPS) production in a specific Streptococcus thermophilus strain. TBC was determined based on real-time quantitative polymerase chain reaction (qPCR) standard curves of S. thermophilus T9 cultured in reconstituted skim milk. EPS yields were measured following isolation and purification steps using a dextran standard curve. The relationship between TBC of and EPS production by S. thermophilus T9 was analyzed alongside these standard curves. The results showed that EPS yield reached 36.6 ± 0.8 mg L−1 following 6 h of incubation. Over the same period, the streptococcal counts measured by qPCR increased from 4.71 ± 0.15 to 7.52 ± 0.09 log10 TBC mL−1. Overall, EPS biosynthesis has been shown to be a growth-related trait in S. thermophilus T9 (R2 = 0.991). In conclusion, the monitoring of TBC may provide an insight into the mechanism of EPS production and, therefore, its use can be recommended for this purpose and for EPS quantification. To our knowledge, this is the first research that has associated qPCR-based TBC with EPS yield in S. thermophilus.

Artocarpus integrifolia seed lectin enhances membrane damage, oxidative stress and biofilm inhibition activity of silver nanoparticles against Staphylococcus aureus

Functionalizing the nanoparticles to target the bacteria surface is an effective approach to curb drug resistance pathogens. The present study describes the synthesis of N-myristoyltaurine capped silver nanoparticles (AgNPs) and functionalization of AgNPs with jacalin (JAgNPs), a lectin from the seeds of Artocarpus integrifolia . JAgNPs has fourfold lower minimum inhibitory concentration than AgNPs and kills the S. aureus in less than 30 min through inducing oxidative stress and membrane damage. JAgNPs (31.25 µM) was found to have excellent biofilm inhibitory activity at lower concentration than AgNPs (62.5 µM). This was judged by crystal violet assay, light microscopy imaging and confocal laser scanning microscopy (CLSM) method. CLSM analysis reveals that JAgNPs is highly effective as compared to AgNPs in reducing the bio-volume and average biofilm thickness to 18.3 ± 2.5 µm 3 /µm 2 and 14.0 ± 2.6 µm, respectively. Scanning electron microscopy (SEM) study reveals that JAgNPs prevents the colony formation and inhibited exopolysaccharide synthesis, thereby reducing the formation of biofilm. At minimum biofilm inhibitory concentrations, SEM and double staining technique using CLSM provides evidence that cells are intact and live, suggesting that NPs exhibit true biofilm activity. The findings indicate that jacalin functionalization may offer a unique way to use nanosilver at lower concentrations for treating infection associated with S. aureus biofilm. • Jacalin functionalized AgNPs kills S. aureus in less than 30 min. • NPs exert antibacterial activity through inducing oxidative stress and membrane damage. • Jacalin-AgNPs exhibit antibiofilm activity at 0.5X MIC. • NPs inhibits exopolysaccharide biosynthesis.

Lyophilized alginate-based microspheres containing Lactobacillus fermentum D12, an exopolysaccharides producer, contribute to the strain\u2019s functionality in vitro

Lactobacillus (Limosilactobacillus) fermentum D12 is an exopolysaccharide (EPS) producing strain whose genome contains a putative eps operon. Whole-genome analysis of D12 was performed to disclose the essential genes correlated with activation of precursor molecules, elongation and export of the polysaccharide chain, and regulation of EPS synthesis. These included the genes required for EPS biosynthesis such as epsA, B, C, D and E, also gt, wzx, and wzy and those involved in the activation of the precursor molecules galE, galT and galU. Both the biosynthesis and export mechanism of EPS were proposed based on functional annotation. When grown on MRS broth with an additional 2% w/v glucose, L. fermentum D12 secreted up to 200 mg/L of a mixture of EPSs, whose porous structure was visualized by scanning electron microscopy (SEM). Structural information obtained by 1HNMR spectroscopy together with composition and linkage analyses, suggested the presence of at least two different EPSs, a branched heteropolysaccharide containing t-Glcp and 2,6-linked Galf, and glycogen. Since recent reports showed that polysaccharides facilitate the probiotic-host interactions, we at first sought to evaluate the functional potential of L. fermentum D12. Strain D12 survived simulated gastrointestinal tract (GIT) conditions, exhibited antibacterial activity against enteropathogenic bacteria, adhered to Caco-2 cells in vitro, and as such showed potential for in vivo functionality. The EPS crude extract positively influenced D12 strain capacity to survive during freeze-drying and to adhere to extracellular matrix (ECM) proteins but did not interfere Caco-2 and mucin adherence when added at concentrations of 0.2, 0.5, and 1.0 mg/mL. Since the viable bacterial count of free D12 cells was 3 logarithmic units lower after the exposure to simulated GIT conditions than the initial count, the bacterial cells had been loaded into alginate for viability improvement. Microspheres of D12 cells, which were previously analyzed at SEM, significantly influenced their survival during freeze-drying and in simulated GIT conditions. Furthermore, the addition of the prebiotic substrates mannitol and lactulose improved the viability of L. fermentum D12 in freeze-dried alginate microspheres during 1-year storage at 4 °C compared to the control.

Whole genome and acid stress comparative transcriptome analysis of Lactiplantibacillus plantarum ZDY2013.

Previous study has reported that Lactiplantibacillus plantarum ZDY2013 which was screened from traditional Chinese fermented soybeans has a strong acid resistance. The purpose of this study was to uncover the genes potentially related to its genetic adaptation and probiotic profiles, based on comparative genomic and comparative transcriptome analysis. We got the basic information about L. plantarum ZDY2013 and identified genes which are related to genetic adaptation and probiotic profiles, including carbohydrate transport and metabolism, cell wall/membrane/envelope biogenesis, proteolytic enzyme systems and amino acid biosynthesis, CRISPR adaptive immunity, stress responses, ability to adhere to the host intestinal wall, exopolysaccharide (EPS) biosynthesis, and bacteriocin biosynthesis. Comparative transcriptome showed CK group (normal MRS culture L. plantarum ZDY2013) and SCL group (pH 3.0 MRS culture L. plantarum ZDY2013) had 652 significant differentially expressed genes including 310 up-regulated genes and 342 down-regulated genes. Besides that, these genes had been classified through KEGG and GO functional annotation. In addition, we also found top 20 KEGG pathways adjusted to acid stress. Then, some genes were selected to verify the transcriptome analysis and explore the mechanism of how L. plantarum ZDY2013 tolerate acid stress. We found that some genes of ABC transporter, phosphotransferase system, oxidation reduction process, membrane transporter and phosphorylation metabolism process had a significant change. These results suggested that comparative characterization of the L. plantarum ZDY2013 genome and transcriptome provided the genetic basis for further elucidating the functional mechanisms of it.

An cost-effective production of bacterial exopolysaccharide emulsifier for oil pollution bioremediation

The thermotolerant bacterium Geobacillus stearothermophilus DG1 was cultured with agroindustrial by-product molasses supplemented with NaNO3 and (NH4)2HPO3 to produce exopolysaccharides (EPSs) for the bioremediation of oil contamination. The EPSs produced under the optimal conditions contained a relatively high content of carbohydrates (60.32%), proteins (11.82%) and uronic acids (1.32%) as the main constituents. The EPSs successfully emulsified a range of hydrocarbons and exhibited excellent stability by thermal treatmanet and over a broad range of pH as well as a strong tolerance to different salinity levels. Additionally, hazardous constituents of the crude oil such as heteroaromatics, alkanes and alkyl cyclohexanes were readily degraded. A gene cluster containing 26 open reading frames (ORFs) responsible for EPS biosynthesis was isolated and characterised. Metabolic pathways related to EPS production and hydrocarbon degradation were reconstructed, and the analysis of the relationship between bioemulsion and biodegradation revealed that the EPSs enhanced petroleum hydrocarbon solubility and bioavailability, thereby promoting its uptake and utilization by microorganisms. This may also provide a carbon resource for sustainable bioremediation to generate sugar precursors for the synthesis of EPSs. Our results promoted the emulsification efficiency of crude oil and the findings illuminate the potential for EPS application in the bioremediation of hydrocarbons in different environments.

Yeast exopolysaccharides and their physiological functions.

Mounting evidence indicated the capability of various microorganisms in biosynthesis of exopolysaccharides (EPSs). A wide range of evidence extensively investigated the ability of bacterial species for EPS synthesis and their favorable effects, so little is known regarding yeast species. Many factors like composition of growth media and fermentation conditions are related to the structural and physical properties of EPSs. The EPS protects the producer yeast strain against extreme environment. Researchers proposed that yeast EPSs have priority over bacterial EPSs because of high yields of EPS biosynthesis and easy separation methods from growth media. Besides, they have drawn increasing attention due to their interesting biological activities, food, pharmaceutical, and cosmetics applications. Although a limited number of studies exist, this review aims to highlight the EPS structure and various applications of known yeast species in detail.

Comparative Proteomic Analyses Between Biofilm-Forming and Non-biofilm-Forming Strains of Corynebacterium pseudotuberculosis Isolated From Goats.

Caseous lymphadenitis (CLA) is a chronic disease that affects small ruminants and causes economic losses in the associated breeding system. The causative agent of CLA is Corynebacterium pseudotuberculosis, a Gram-positive bacterium that exhibits tropism for external and internal lymph nodes and induces abscess formation in the host. Bacterial communities often produce a biofilm matrix that serves various functions, including protection against hostile environmental conditions, antibiotics, and the host immune response. Although biofilm formation has been reported for C. pseudotuberculosis, not all strains demonstrate this property in culture. In this work, we report the first comparative proteomic analysis of one biofilm-forming (CAPJ4) and one biofilm-non-forming strain (CAP3W) of C. pseudotuberculosis isolated from goats. Bacterial whole cell protein extracts were obtained for mass spectrometry analyses. Using LC-MS/MS, our studies reveal three and four proteins exclusively found in the CAPJ4 and CAP3W proteome, respectively. In addition, label-free quantitative analysis identified 40 proteins showing at-least 2-fold higher values in CAPJ4 compared CAP3W proteome Notably, CAPJ4 differentially synthesized the penicillin-binding protein, which participates in the formation of peptidoglycans. CAPJ4 also exhibited upregulation of N-acetylmuramoyl-L-alanine amidase and galactose-1-phosphate uridylyltransferase, which are involved in biofilm formation and exopolysaccharide biosynthesis. Here, we demonstrate that biofilm formation in C. pseudotuberculosis is likely associated with specific proteins, some of which were previously shown to be associated with virulence and biofilm formation in other organisms. Our findings may drive studies related to the bacterial mechanisms involved in the biofilm formation, in addition to providing targets for the treatment of CLA.

Exopolysaccharide Characterization of Rhizobium favelukesii LPU83 and Its Role in the Symbiosis With Alfalfa.

One of the greatest inputs of available nitrogen into the biosphere occurs through the biological N2-fixation to ammonium as result of the symbiosis between rhizobia and leguminous plants. These interactions allow increased crop yields on nitrogen-poor soils. Exopolysaccharides (EPS) are key components for the establishment of an effective symbiosis between alfalfa and Ensifer meliloti, as bacteria that lack EPS are unable to infect the host plants. Rhizobium favelukesii LPU83 is an acid-tolerant rhizobia strain capable of nodulating alfalfa but inefficient to fix nitrogen. Aiming to identify the molecular determinants that allow R. favelukesii to infect plants, we studied its EPS biosynthesis. LPU83 produces an EPS I identical to the one present in E. meliloti, but the organization of the genes involved in its synthesis is different. The main gene cluster needed for the synthesis of EPS I in E. meliloti, is split into three different sections in R. favelukesii, which probably arose by a recent event of horizontal gene transfer. A R. favelukesii strain devoided of all the genes needed for the synthesis of EPS I is still able to infect and nodulate alfalfa, suggesting that attention should be directed to other molecules involved in the development of the symbiosis.

Effects of an EPS Biosynthesis Gene Cluster of Paenibacillus polymyxa WLY78 on Biofilm Formation and Nitrogen Fixation under Aerobic Conditions

Exopolysaccharides (EPS) are of high significance in bacterial biofilm formation. However, the effects of EPS cluster(s) on biofilm formation in Paenibacillus species are little known. In this study, we have shown that Paenibacillus polymyxa WLY78, a N2-fixing bacterium, can form biofilm. EPS is the major component of the extracellular matrix. The genome of P. polymyxa WLY78 contains two putative gene clusters (designated pep-1 cluster and pep-2 cluster). The pep-1 cluster is composed of 12 putative genes (pepO-lytR) co-located in a 13 kb region. The pep-2 cluster contains 17 putative genes (pepA-pepN) organized as an operon in a 20 kb region. Mutation analysis reveals that the pep-2 cluster is involved in EPS biosynthesis and biofilm formation. Disruption of the pep-2 cluster also leads to the enhancement of motility and change of the colony morphology. In contrast, disruption of the pep-1 cluster does not affect EPS synthesis or biofilm formation. More importantly, the biofilm allowed P. polymyxa WLY78 to fix nitrogen in aerobic conditions, suggesting that biofilm may provide a microaerobic environment for nitrogenase synthesis and activity.