Extracellular Polysaccharide Production Research Articles

Effect of LongZhang Gargle on Dual-Species Biofilm of Candida albicans and Streptococcus mutans.

Bioactive natural products have become a hot spot for oral disease treatments. At the present study, LongZhang Gargle was investigated for its effects on single-species biofilms of Candida albicans and dual-species biofilms of Candida albicans and Streptococcus mutans. Two different models of single and dual-species biofilms were grown in YNBB medium under appropriate conditions. Biofilm biomass, biofilm architecture, and cell activity in biofilms were assessed using Crystal Violet Staining, MTT, scanning electron microscopy (SEM), and confocal laser scanning microscopy (CLSM). Significant reductions of biofilm biomass and fungus activity were obtained when treated with LongZhang Gargle at 2% (P < 0.05), 4% (P < 0.05), and 8% (P < 0.05) in single-species biofilms of C. albicans, and at 4% (P < 0.05) and 8% (P < 0.05) in double-species biofilms. Suppression of density, thickness, and the proportion of hyphae and fungal spores were obtained under SEM and CLSM. In conclusion, LongZhang Gargle affects single and dual-species biofilms by inhibiting biofilm biomass, cell activity, and formation of hyphae, but it does not affect the production of Extracellular polysaccharides (EPS). We speculate that LongZhang Gargle would be a promising natural drug, which can be used in treatment against C. albicans and S. mutans in oral diseases.

Shrimp lectin–conjugated copper sulfide nanoparticles enhance immune response and gene expression in Etroplus suratensis infected with Aeromonas hydrophila

Lectins are hemagglutinating glycoproteins which act as pathogen recognition receptors that can bind to the pathogen-associated molecular patterns. Shrimp lectin (Md-Lec) is a lectin that is reported to have antimicrobial property. In this study, the CuS nanoparticles (CuS NPs) which were previously reported to have wide range of antimicrobial activity were conjugated with Md-Lec. Thus, their minimum inhibitory concentration (MIC) against aquatic pathogens was reduced four times than that of their individual activity. Moreover, antibacterial and antibiofilm assay was performed with Md-Lec, pCuS NPs, and Md-Lec conjugated with pCuS NPs (Md-Lec-pCuS NPs) against Aeromonas hydrophila at 25, 50, and 100 μg/mL. The hydrophobicity index and EPS (extracellular polysaccharide) production of A. hydrophila were decreased after treating with Md-Lec-pCuS NPs. The non-specific immune parameters—lysozyme assay, peroxidase assay, and respiratory burst activity—were elevated considerably in the infected fishes that are treated with Md-Lec-pCuS NPs at their MIC. Further, immune gene (COX2, MyD88, and LEC) expression analysis at different time intervals was also done in the infected Etroplus suratensis that are treated with Md-Lec-pCuS NPs at 3.13 μM concentration. The results reveal that the immune gene expression was upregulated at the 6th hour of infection in the infected fish that are treated with Md-Lec-pCuS NPs (3.13 μM) and thus it can be used as a therapeutic agent which can act as an immunostimulant for the production of disease-free fishes in aquaculture industry.

Antioxidant potential and optimization of production of extracellular polysaccharide by Acinetobacter indicus M6

BackgroundExtracellular polysaccharides (ECPs) produced by biofilm-producing marine bacterium have great applications in biotechnology, pharmaceutical, food engineering, bioremediation, and bio-hydrometallurgy industries. The ECP-producing strain was identified as Acinetobacter indicus M6 species by 16S rDNA analysis. The polymer produced by the isolate was quantified and purified and chemically analyzed, and antioxidant activities have been studied. The face-centered central composite design (FCCCD) was used to design the model. ResultsThe results have clearly shown that the ECP was found to be endowed with significant antioxidative activities. The ECP showed 59% of hydroxyl radical scavenging activity at a concentration of 500 μg/mL, superoxide radical scavenging activity (72.4%) at a concentration of 300 μg/mL, and DPPH˙ radical scavenging activity (72.2%) at a concentration of 500 μg/mL, respectively. Further, HPLC and GC-MS results showed that the isolated ECP was a heteropolymer composed of glucose as a major monomer, and mannose and glucosamine were minor monomers. Furthermore, the production of ECP by Acinetobacter indicus M6 was increased through optimization of nutritional variables, namely, glucose, yeast extract, and MgSO4 by “Response Surface Methodology”. Moreover the production of ECP reached to 2.21 g/L after the optimization of nutritional variables. The designed model is statistically significant and is indicated by the R2 value of 0.99. The optimized medium improved the production of ECP and is two folds higher in comparison with the basal medium. ConclusionsAcinetobacter indicus M6 bacterium produces a novel and unique extracellular heteropolysaccharide with highly efficient antioxidant activity. GC-MS analyses elucidated the presence of quite uncommon (1→4)-linked glucose, (1→4)-linked mannose, and (→4)-GlcN-(1→) glycosidic linkages in the backbone. The optimized medium improved the production of ECP and is two folds higher in comparison with the basal medium. The newly optimized medium could be used as a promising alternative for the overproduction of ECP.

Effects of extracellular DNA on dual-species biofilm formed by Streptococcus mutans and Candida albicans

Streptococcus mutans is the most important acid-producing pathogen that causes dental caries, while Candida albicans is an opportunistic fungal pathogen that is frequently detected in conjunction with heavy infection by S. mutans. Their interactions in dental plaque biofilms remain unclear. Extracellular DNA (eDNA) is found in oral biofilms, but its effects have not been thoroughly defined. In this study, the role of eDNA in dual-species biofilms formed by S. mutans and C. albicans was investigated. With eDNA removal, the growth of both strains was not affected, but the formation of dual-species biofilms obviously decreased. In addition, the removal of eDNA spatially disrupted the structure of the dual-species biofilm. It was also shown that eDNA mainly affected the initial attachment and development stages of the dual-species biofilms but not the well-developed biofilms. A similar phenomenon was also observed in the cell viability of dual-species biofilms after DNase I treatment. To further exploration, we analyzed the expression of genes associated with biofilm formation in both S. mutans and C. albicans. We determined that the co-cultivation of S. mutans and C. albicans promotes the expression of genes related to extracellular polysaccharide production (e.g., gtfC), adhesion (e.g., spaP, epa1), mycelial transformation (e.g., hwp1), and drug resistance (e.g., cdr2). However, these genes were significantly downregulated when the eDNA of the dual-species biofilm was removed by adding DNase I compared to those untreated groups. Altogether, eDNA removal, such as that by DNase I treatment, could be considered a promising strategy to control oral biofilms and biofilm-associated oral diseases.

Staphylococcus aureus induces an itaconate-dominated immunometabolic response that drives biofilm formation

Staphylococcus aureus is a prominent human pathogen that readily adapts to host immune defenses. Here, we show that, in contrast to Gram-negative pathogens, S. aureus induces a distinct airway immunometabolic response dominated by the release of the electrophilic metabolite, itaconate. The itaconate synthetic enzyme, IRG1, is activated by host mitochondrial stress, which is induced by staphylococcal glycolysis. Itaconate inhibits S. aureus glycolysis and selects for strains that re-direct carbon flux to fuel extracellular polysaccharide (EPS) synthesis and biofilm formation. Itaconate-adapted strains, as illustrated by S. aureus isolates from chronic airway infection, exhibit decreased glycolytic activity, high EPS production, and proficient biofilm formation even before itaconate stimulation. S. aureus thus adapts to the itaconate-dominated immunometabolic response by producing biofilms, which are associated with chronic infection of the human airway.

Effects of nitrogen from different sources on mycelial biomass and polysaccharide production and pellet morphology in submerged cultures of Grifola frondosa

The effects of nitrogen in the medium on the production of mycelial biomass, extracellular polysaccharides (EPS), and intracellular polysaccharides (IPS) was investigated in submerged cultures of Grifola frondosa. In addition, the effects on pellet morphology were examined. The maximum production levels of mycelial biomass (2.32 g/L), EPS (1.58 g/L), and IPS (29.1 mg/L) were obtained when the nitrogen sources in the medium were yeast extract, malt extract, and peptone, respectively. Using yeast extract as the nitrogen source yielded the maximum mycelial biomass, and morphological characterization revealed a composition of 47% large pellets (fraction L), 20% small pellets (fraction S), and 33% adhesive mycelia (fraction A). The maximum circularity value and the minimum roughness value of the pellets were observed using yeast extract cultures. Both the compactness (0.53) and circularity (0.15) of the pellets were the lowest among the seven types of nitrogen sources, but the roughness (2.86) was the highest in malt extract, which was the nitrogen source that resulted in maximum polysaccharide production. The results revealed that the production levels of mycelial biomass, EPS, and IPS of G. frondosa were associated with changes in pellet morphology due to the source of nitrogen in the medium.

Contamination and Biofilm Formation of Foodborne and Opportunistic Pathogens in Yellow-Feathered Chicken Carcass.

This study aims to analyze the contamination and biofilm formation of foodborne and opportunistic pathogens in yellow-feathered chicken carcasses sampled in different seasons and to prove the relationship between biofilm-forming ability and bacterial extracellular polysaccharide (EPS) production. A total of 78 strains were isolated from chicken samples. The strains consisted of 30.8% Escherichia coli, 14.1% Pseudomonas aeruginosa, 12.8% Salmonella enteritidis, 12.8% Klebsiella pnenmoniae, 10.2% Enterobacter cloacae, 8.9% Proteus mirabilis, 5.1% Klebsiella oxytoca, 1.3% Staphylococcus aureus, and 1.3% Citrobacter braakii. Crystal violet staining assay revealed six strains with strong biofilm-forming ability, namely, E. coli S7, K. oxytoca B12, K. pnenmoniae B6, S. enteritidis H4, P. aeruginosa M5, and S. aureus G1, which showed had high abilities of cell motility and EPS production. Confocal laser scanning microscopy and scanning electron microscopy showed that all six strains can form mature biofilm architectures after 5 d of cultivation. This study may serve as a reference to control the contamination of foodborne pathogens in yellow-feathered chicken and enhance the quality and shelf life of these chicken products.

In vitro Antibacterial Activity of an FDA-Approved H+-ATPase Inhibitor, Bedaquiline, Against Streptococcus mutans in Acidic Milieus.

BackgroundDental caries is an acid-related disease. Current anti-caries agents mainly focus on the bacteriostatic effect in a neutral environment and do not target acid-resistant microorganisms related to caries in acidic milieus.ObjectivesTo assess the in vitro antibacterial activities of bedaquiline against oral pathogens in acidic milieus.MethodsStreptococcus mutans, Streptococcus sanguinis, and Streptococcus salivarius were used to prepare the mono-/multiple suspension and biofilm. The MIC and IC50 of bedaquiline against S. mutans were determined by the broth microdilution method. Bedaquiline was compared regarding (i) the inhibitory activity in pH 4–7 and at different time points against planktonic and biofilm; (ii) the effect on the production of lactic acid, extracellular polysaccharide, and pH of S. mutans biofilm; (iii) the cytotoxicity effects; and (iv) the activity on H+-ATPase enzyme of S. mutans.ResultsIn pH 5 BHI, 2.5 mg/L (IC50) and 4 mg/L (MIC) of bedaquiline inhibited the proliferation and biofilm generation of S. mutans and Mix in a dose-dependent and time-dependent manner, but it was invalid in a neutral environment. The lactic acid production, polysaccharide production, and pH drop range reduced with the incorporation of bedaquiline in a pH 5 environment. Its inhibitory effect (>56 mg/L) against H+-ATPase enzyme in S. mutans and its non-toxic effect (<10 mg/L) on periodontal ligament stem cells were also confirmed.ConclusionBedaquiline is efficient in inhibiting the proliferation and biofilm generation of S. mutans and other oral pathogens in an acidic environment. Its high targeting property and non-cytotoxicity also promote its clinical application potential in preventing caries. Further investigation of its specific action sites and drug modification are warranted.

Trp-Containing Antibacterial Peptides Impair Quorum Sensing and Biofilm Development in Multidrug-Resistant Pseudomonas aeruginosa and Exhibit Synergistic Effects With Antibiotics.

Pseudomonas aeruginosa uses quorum sensing (QS) to control virulence, biofilm formation and antibiotic efflux pump expression. The development of effective small molecules targeting the QS system and biofilm formation represents a novel attractive strategy. In this present study, the effects of a series of Trp-containing peptides on the QS-regulated virulence and biofilm development of multidrug-resistant P. aeruginosa, as well as their synergistic antibacterial activity with three classes of traditional chemical antibiotics were investigated. The results showed that Trp-containing peptides at low concentrations reduced the production of QS-regulated virulence factors by downregulating the gene expression of both the las and rhl systems in the strain MRPA0108. Biofilm formation was inhibited in a concentration-dependent manner, which was associated with extracellular polysaccharide production inhibition by downregulating pelA, algD, and pslA transcription. These changes correlated with alterations in the extracellular production of pseudomonal virulence factors and swarming motility. In addition, the combination of Trp-containing peptides at low concentration with the antibiotics ceftazidime and piperacillin provided synergistic effects. Notably, L11W and L12W showed the highest synergy with ceftazidime and piperacillin. A mechanistic study demonstrated that the Trp-containing peptides, especially L12W, significantly decreased β-lactamase activity and expression of efflux pump genes OprM, MexX, and MexA, resulting in a reduction in antibiotic efflux from MRPA0108 cells and thus increasing the antibacterial activity of these antibiotics against MRPA0108.

Evaluation of physio-biochemical potentials of alkaliphilic bacterial diversity in bauxite processing residues of diverse restoration history

Bauxite residue, the waste by-product of bauxite ore processing in alumina industry is mostly deposited around the areas of alumina plants and often leads to a number of environmental and human health problems. Here we report the physico-chemical and microbiological analysis of bauxite residue samples of different restoration history (0–40 years). The samples were characterized by high alkalinity (pH 8.5–12.5) and metal content along with poor organic matter. While the presence of Sodalite, Hematite, Boehmite, Gibbsite, Anatase and Quartz was confirmed by X-ray powder diffraction (XRD) analysis, heavy metal contamination was evaluated by applying contamination factor and pollution load index. The aerobic microbial density as well as activity of the samples were in general very low and ranged from 1.2 to 8.96 × 102 cfu/g and 0.32–0.96 µg fluorescein/g/h. A total of 12 phenotypically distinguishable bacteria were isolated in pure form which showed significant tolerance to nickel (Ni), chromium (Cr), aluminium (Al), iron (Fe), manganese (Mn), magnesium (Mg) and lead (Pb), high pH and NaCl and were resistant to a number of antibiotics. Further, tolerance to alkalinity, transformation of waste to neutral one, production of diverse enzymes, extracellular polysaccharide and formation of biofilm, the unique features of these bacterial isolates could be of immense importance for potential applications in bioremediation and bioleaching.

Role of c-di-GMP in improving stress resistance of alginate-chitosan microencapsulated Bacillus subtilis cells in simulated digestive fluids

Probiotics (Bacillus subtilis 04178) were entrapped in alginate-chitosan microcapsules by high-voltage electrostatic process. The encapsulation pattern was established as entrapped low density cells with culture (ELDCwc). The performance of ELDCwc cells was investigated against stress environments of simulated digestive fluids. After incubation in simulated gastric (pH 2.5) and intestinal fluids (4% bile salt) for 2h, the survival rate of ELDCwc cells (18.19% and 27.54%) was significantly higher than that of the free cells (0.0000009% and 0.0005%). The reason why B. subtilis embedded in microcapsules can resist the stress environments was that the mass production of extracellular proteins and polysaccharides prompted B. subtilis to form cell aggregates. The production of extracellular proteins and polysaccharides were regulated by the concentration of c-di-GMP and the expression of ydaJKLMN operon, abbA, sinI, slrA, slrB, abrR and sinR. c-di-GMP is important for the production of extracellular polymer substance to enhance probiotic viability in stress environments.

Effect of S. mutans combinations with bifidobacteria/lactobacilli on biofilm and enamel demineralization.

The present study evaluated the ability of Bifidobacterium and Lactobacillus species associated with streptococci to increase insoluble extracellular polysaccharide (EPS) production and initial caries lesion progression. Bovine enamel blocks (n = 190; 4 mm x 4 mm) were prepared, selected according to initial surface hardness (SH), and divided into two groups: a) double combinations: S. mutans with Bifidobacterium or Lactobacillus, and b) triple combinations: S. mutans and S. sobrinus with Bifidobacterium or Lactobacillus species. The blocks were exposed to the bacterial associations for 7 days. Subsequently, quantity of EPS from biofilms and caries lesion depth were determined by means of colorimetric and cross-sectional enamel hardness (ΔKHN) analysis. The data were submitted to one-way analysis of variance, followed by the Bonferroni test (p < 0.05). S. mutans with B. animalis or B. dentium produced a higher quantity of EPS; S. mutans + B. animalis led to the highest ∆KHN. S. mutans + S. sobrinus + B. longum induced greater EPS and ∆KHN values. In conclusion, associations of B. animalis and B. longum with streptococci promoted EPS production and caries lesion progression.

Effect of collagen and EPS components on the viscoelasticity of Pseudomonas aeruginosa biofilms.

Pseudomonas aeruginosa is an opportunistic pathogen that causes thousands of deaths every year in part due to its ability to form biofilms composed of bacteria embedded in a matrix of self-secreted extracellular polysaccharides (EPS), e-DNA, and proteins. In chronic wounds, biofilms are exposed to the host extracellular matrix, of which collagen is a major component. How bacterial EPS interacts with host collagen and whether this interaction affects biofilm viscoelasticity is not well understood. Since physical disruption of biofilms is often used in their removal, knowledge of collagen's effects on biofilm viscoelasticity may enable new treatment strategies that are better tuned to biofilms growing in host environments. In this work, biofilms are grown in the presence of different concentrations of collagen that mimic in vivo conditions. In order to explore collagen's interaction with EPS, nine strains of P. aeruginosa with different patterns of EPS production were used to grow biofilms. Particle tracking microrheology was used to characterize the mechanical development of biofilms over two days. Collagen is found to decrease biofilm compliance and increase relative elasticity regardless of the EPS present in the system. However, this effect is minimized when biofilms overproduce EPS. Collagen appears to become a de facto component of the EPS, through binding to bacteria or physical entanglement.

Production and Characterization of Extracellular Polymeric Substances by marine Halomonas sp. NASH isolated from Wadi El-Natroun

Halophilic micro-organisms often synthesize and produce extracellular polysaccharides (EPS), whose physical, chemical properties and material properties vary greatly from each other. The extracellular polysaccharide (EPS) development of Halomonas sp. MN795630 strain type halophilic bacterium (NASH) was investigated and whether biotechnological applications were feasible. After 168 hours of incubation, 4 g/L of EPS was produced and all elements from the medium were completely used during the growth. Sucrose has been identified as the most favorable carbon source for production of EPS and maximum production (6 g/l). Beef extract level was shown to be the best for EPS production among different nitrogen sources. Optimum production of EPS (10 g/L) were achieved by supplementing the medium with 4M NaCl, pH adjusted at 9 and the medium was inoculated with 7% initial inoculum. The purified EPS were characterized chemically. Fourier transform infrared (FTIR) spectrophotometer was observed in several functional groups. EPS also demonstrated an significant inhibitor of Candida albicans ATCC 10231 and Pseudomonas aeruginosa ATCC 9027 (20.4 and 14.7 mm), respectively. EPS show satisfactory results when applied as anti-oxidant, anti-inflammatory and emulsifier.

Carvacrol Targets SarA and CrtM of Methicillin-Resistant Staphylococcus aureus to Mitigate Biofilm Formation and Staphyloxanthin Synthesis: An In Vitro and In Vivo Approach.

Carvacrol is an essential oil traditionally used in culinary processes as spice due to its aromatic nature and also known for various biological activities. In the present study, the antivirulence efficacy of carvacrol against methicillin-resistant Staphylococcus aureus (MRSA) is explored. MRSA is an opportunistic pathogen capable of causing various superficial and systemic infections in humans. Biofilm formation and virulence factors of MRSA are responsible for its pathogenesis and resistance. Hence, the aim of this study was to explore the antibiofilm and antivirulence efficacy of carvacrol against MRSA. Carvacrol at 75 μg/mL inhibited MRSA biofilm by 93%, and it also decreased the biofilm formation on polystyrene and glass surfaces. Further, microscopic analyses revealed the reduction in microcolony formation and collapsed structure of biofilm upon carvacrol treatment. The growth curve analysis and the Alamar blue assay showed the nonfatal effect of carvacrol on MRSA. Further, carvacrol significantly reduced the production of MRSA biofilm-associated slime and extracellular polysaccharide. In addition, carvacrol strongly inhibited the antioxidant pigment staphyloxanthin and its intermediates’ synthesis in MRSA. Inhibition of biofilm and staphyloxanthin by carvacrol enhanced the susceptibility of MRSA to oxidants and healthy human blood. Quantitative polymerase chain reaction (qPCR) analysis unveiled the downregulation of sarA-mediated biofilm gene expression and staphyloxanthin-associated crtM gene expression. The sarA-dependent antibiofilm potential of carvacrol was validated using S. aureus Newman wild-type and isogenic ΔsarA strains. In silico molecular docking analysis showed the high binding efficacy of carvacrol with staphylococcal accessory regulator A (SarA) and 4,4′-diapophytoene synthase (CrtM) when compared to positive controls. Furthermore, the in vivo efficacy of carvacrol against MRSA infection was demonstrated using the model organism Galleria mellonella. The results revealed the nontoxic nature of carvacrol to the larvae and the rescuing potential of carvacrol against MRSA infection. Finally, the current study reveals the potential of carvacrol in inhibiting the biofilm formation and staphyloxanthin synthesis of MRSA by targeting the global regulator SarA and a novel antivirulence target CrtM.

Fusobacteriumnucleatum Adheres to Clostridioides difficile via the RadD Adhesin to Enhance Biofilm Formation in Intestinal Mucus

Although Clostridioides difficile infection (CDI) is known to involve the disruption of the gut microbiota, little is understood regarding how mucus-associated microbes interact with C difficile. We hypothesized that select mucus-associated bacteria would promote C difficile colonization and biofilm formation. To create a model of the human intestinal mucus layer and gut microbiota, we used bioreactors inoculated with healthy human feces, treated with clindamycin and infected with C difficile with the addition of human MUC2-coated coverslips. C difficile was found to colonize and form biofilms on MUC2-coated coverslips, and 16S rRNA sequencing showed a unique biofilm profile with substantial cocolonization with Fusobacterium species. Consistent with our bioreactor data, publicly available data sets and patient stool samples showed that a subset of patients with C difficile infection harbored high levels of Fusobacterium species. We observed colocalization of C difficile and F nucleatum in an aggregation assay using adult patients and stool of pediatric patients with inflammatory bowel disease and in tissue sections of patients with CDI. C difficile strains were found to coaggregate with F nucleatum subspecies invitro; an effect that was inhibited by blocking or mutating the adhesin RadD on Fusobacterium and removal of flagella on C difficile. Aggregation was shown to be unique between F nucleatum and C difficile, because other gut commensals did not aggregate with C difficile. Addition of F nucleatum also enhanced C difficile biofilm formation and extracellular polysaccharide production. Collectively, these data show a unique interaction of between pathogenic C difficile and F nucleatum in the intestinal mucus layer.

Quorum Sensing Inhibition Attenuates the Virulence of the Plant Pathogen Ralstonia solanacearum Species Complex.

Strains of Ralstonia solanacearum species complex (RSSC) cause "bacterial wilt" on a wide range of plant species and thus lead to marked economic losses in agriculture. Quorum sensing (QS), a bacterial cell-cell communication mechanism, controls the virulence of RSSC strains by regulating the production of extracellular polysaccharide (EPS) and secondary metabolites, biofilm formation, and cellular motility. R. solanacearum strain OE1-1 employs (R)-methyl 3-hydroxymyristate (3-OH MAME) as a QS signal, which is synthesized by the PhcB methyltransferase and sensed by the PhcS/PhcRQ two-component system. We describe the design, synthesis, and biological evaluation of inhibitors of the phc QS system. Initial screening of a small set of QS signal analogues revealed that methyl 3-hydroxy-8-phenyloctanoate, named, PQI-1 (phc quorum sensing inhibitor-1), inhibited biofilm formation by strain OE1-1. To improve its inhibitory activity, the derivatives of PQI-1 were synthesized, and their QS inhibition activities were evaluated. PQIs-2-5 evolved from PQI-1 more strongly inhibited not only biofilm formation but also the production of ralfuranone and EPS. Furthermore, RNA-Seq analysis revealed that the PQIs effectively inhibited QS-dependent gene expression and repression in strain OE1-1. On the other hand, the PQIs did not affect the canonical QS systems of the representative reporter bacteria. These antagonists, especially PQI-5, reduced wilting symptoms of the tomato plants infected with strain OE1-1. Taken together, we suggest that targeting the phc QS system has potential for the development of chemicals that protect agricultural crops from bacterial wilt disease.

Elucidation of a Novel Role of YebC in Surface Polysaccharides Regulation of Escherichia coli bipA-Deletion.

The BipA (BPI-inducible protein A) protein is ubiquitously conserved in various bacterial species and belongs to the translational GTPase family. Interestingly, the function of Escherichia coli BipA is not essential for cell growth under normal growth conditions. However, cultivation of bipA-deleted cells at 20°C leads to cold-sensitive growth defect and several phenotypic changes in ribosome assembly, capsule production, and motility, suggesting its global regulatory roles. Previously, our genomic library screening revealed that the overexpressed ribosomal protein (r-protein) L20 partially suppressed cold-sensitive growth defect by resolving the ribosomal abnormality in bipA-deleted cells at low temperature. Here, we explored another genomic library clone containing yebC, which encodes a predicted transcriptional factor that is not directly associated with ribosome biogenesis. Interestingly, overexpression of yebC in bipA-deleted cells diminished capsule synthesis and partially restored lipopolysaccharide (LPS) core maturation at a low temperature without resolving defects in ribosome assembly or motility, indicating that YebC may be specifically involved in the regulation of exopolysaccharide and LPS core synthesis. In this study, we collectively investigated the impacts of bipA-deletion on E. coli capsule, LPS, biofilm formation, and motility and revealed novel roles of YebC in extracellular polysaccharide production and LPS core synthesis at low temperature using this mutant strain. Furthermore, our findings suggest that ribosomal defects as well as increased capsule synthesis, and changes in LPS composition may contribute independently to the cold-sensitivity of bipA-deleted cells, implying multiple regulatory roles of BipA.

An Evaluation of the Antibacterial Properties of Tormentic Acid Congener and Extracts From Callistemon viminalis on Selected ESKAPE Pathogens and Effects on Biofilm Formation.

ESKAPE pathogens, namely, Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species, are responsible for a majority of all healthcare-acquired infections (HAI). The bacteria cause nosocomial infections in immunocompromised patients. Extracts from Callistemon viminalis have been shown to have antibacterial, antifungal, and anti-inflammatory activities. Tormentic acid congener, a pentacyclic triterpene saponin, was isolated from C. viminalis leaves. This study aimed to investigate the antibacterial effects of tormentic acid congener and leaf extracts on biofilm formation by A. baumannii, S. aureus, S. pyogenes, and P. aeruginosa. The antibacterial effects were determined by the microbroth dilution method, and ciprofloxacin was used as the standard antibacterial drug. Biofilm formation and detachment assays were performed using crystal violet staining. Production of extracellular polymeric DNA and polysaccharides from biofilms was also determined. Tormentic acid congener showed time-dependent antibacterial activity against P. aeruginosa with a MIC of 100 µg/ml and caused significant protein leakage. Antibacterial activity was found when tormentic acid congener was tested against both S. aureus and P. aeruginosa. The MICs were found to be 25 µg/ml and 12.5 µg/ml for P. aeruginosa and S. aureus cells, respectively. S. pyogenes was found to be susceptible to tormentic acid congener and the hydroethanolic extract with an MIC of 100 µg/ml and 25 µg/ml, respectively. A. baumannii was found not to be susceptible to the compound or the extracts. The compound and the extracts caused a significant decrease in the biofilm extracellular polysaccharide content of S. pyogenes. The extracts and tormentic acid congener caused detachment of biofilms and decreased the release of extracellular DNA and capsular polysaccharides from biofilms of P. aeruginosa and S. aureus. Tormentic acid congener and extracts, thus, have significant antibacterial and antibiofilm activities on these selected ESKAPE bacteria and can act as source lead compounds for the development of antibacterial triterpenoids.

Ergosterol Pathway of Candida albicans Promotes the Growth and Cariogenic Virulence of Streptococcus mutans

To investigate the effect of Candida albicans ( C. albicans) on proliferation and virulence of Streptococcus mutans ( S. mutans), and to verify the role of the ergosterol pathway in it. After single- and co-cultivation of C. albicans wild-type 5314 and S. mutansUA159, the absorbance value of OD 600and colony-forming units (CFU) were detected to reflect the influence of C. albicans on the growth of S. mutans. To reflect the influence of C. albicanson the virulence of S. mutans, the production of extracellular polysaccharides was detected by anthrone-vitriol method, and the acid production capacity was detected by a pH meter. After single- and co-cultivation of C. albicans mutant strains and S. mutans, the growth of S. mutans was evaluated by CFU. After inhibiting the ergosterol pathway by 0 mg/L, 0.012 5 mg/L, and 0.025 mg/L fluconazole for 24 h, the CFU of single- and co-cultivated C. albicans wild-type and S. mutans were detected. The OD 600absorbance value and CFU in theco-cultivation of C. albicans wild-type and S. mutans was higher than that in the single culture ( P<0.05), and the production of extracellular polysaccharides in S. mutans was increased when S. mutans was co-cultured with C. albicans ( P<0.05), accompanied with a more obvious decrease of pH ( P<0.05). Fourteen strains in whole 42 C. albicans mutant strains lost the growth-promoting effect on S. mutans, including 6 ergosterol synthesis-related mutant strains. After co-cultivation of the 6 ergosterol synthesis-related mutant strains and S. mutans, the CFU of S. mutans remained unchanged or decreased. After inhibiting the ergosterol pathway by 0.012 5 mg/L and 0.025 mg/L fluconazole, the CFU of S. mutans in the co-cultivation of C. albicans wild-type and S. mutans was lower than that without fluconazole treatment ( P<0.05), while the CFU of C. albicans and S. mutans single-cultivations did not change significantly ( P>0.05) and the CFU of C. albicans in the co-cultivation of C. albicans wild-type and S. mutans did not change significantly ( P>0.05). C.albicans can enhance the growth ability and virulence of S. mutans through the ergosterol-related pathway. This process can be inhibited by fluconazole, which is expected to become a novel strategy to prevent and treat dental caries.