Exopolysaccharides of lactic acid bacteria isolated from honeybee gut and effects of their antibiofilm activity against Streptococcus mutans.

Streptococcus mutans is one of the main factors in formation of cariogenic biofilms. New strategies need to be developed to reduce the formation of cariogenic biofilms. For this purpose, bacterial exopolysaccharides (EPS) could be considered as new agents against biofilms. Therefore, cell-bound (b-EPS) and released exopolysaccharides (r-EPS) were extracted from the strains Apilactobacillus kunkeei K1.10 and Latilactobacillus curvatus Kar.9b isolated from the microbiota of honeybees, and their antibiofilm effects on S. mutans biofilm formation were determined. The highest reduction in biofilm formation was achieved by r-EPS of L. curvatus Kar.9b and A. kunkeei K1.10. Scanning electron micrographs (SEM) showed that r-EPS inhibited biofilm formation by reducing adhesion of S. mutans. To increase the production of r-EPS from A. kunkeei K1.10, the effects of different incubation conditions were also analyzed. The highest EPS production was obtained during 48 h-incubation at 37ºC in a medium containing 1% fructose. r-EPS can be used as a raw material to inhibit cariogenic biofilms. Further studies revealing the detailed structural analysis of r-EPS and the mechanism of action of its antibiofilm effect could be beneficial. Finally, b-EPS and r-EPS from lactic acid bacteria were found to have very different properties in terms of their antibiofilm properties.

The Effects of Antiperspirant Aluminum Chlorohydrate on the Development of Antibiotic Resistance in Staphylococcus epidermidis.

This study investigates the effects of the antiperspirant aluminum chlorohydrate on the development of antibiotic resistance in commensal Staphylococcus epidermidis isolates. The isolates were exposed to aluminum chlorohydrate for 30 days. The bacteria that developed resistance to oxacillin and ciprofloxacin were isolated, and the expression levels of some antibiotic resistance genes were determined using quantitative reverse transcriptase PCR. Before and after exposure, the minimum inhibitory concentration (MIC) values of the bacteria were determined using the microdilution method. A time-dependent increase was observed in the number of bacteria that developed resistance and increased MIC values. Consistent with the ciprofloxacin resistance observed after exposure, an increase in norA, norB/C, gyrA, gyrB, parC, and parE gene expression was observed. In addition to aluminum chlorohydrate exposure, oxacillin resistance was observed in all test bacteria in the group only subcultured in the medium, suggesting that phenotypic resistance cannot be correlated with chemical exposure in light of these data. The increase in mecA gene expression in selected test bacteria that acquired resistance to oxacillin after exposure compared with control groups suggests that the observed resistance may have been related to aluminum chlorohydrate exposure. To our knowledge, this is the first time in the literature that the effects of aluminum chlorohydrate as an antiperspirant on the development of antibiotic resistance in Staphylococcus epidermidis have been reported.

Analysis of Chemical Structure and Antibiofilm Properties of Exopolysaccharides from Lactiplantibacillus plantarum EIR/IF-1 Postbiotics.

Previous studies have indicated that the exopolysaccharides of lactic acid bacteria exhibit antibiofilm activity against non-oral bacteria by preventing their initial adhesion to surfaces and by downregulating the expression of genes responsible for their biofilm formation. The aims of this study were to (1) characterize the exopolysaccharides (EPSs) of Lactobacillus plantarum EIR/IF-1 postbiotics, (2) test their antibiofilm effect on dual biofilms, and (3) evaluate their bacterial auto-aggregation, co-aggregation, and hydrocarbon-binding inhibitory activity. The EPSs were characterized by FTIR, HPLC, and thermogravimetric analysis. Bacterial auto- and co-aggregation were tested by Kolenbrander's method and hydrocarbon binding was tested by Rosenberg's method. Dual biofilms were formed by culturing Fusobacterium nucleatum ATCC 25586 with one of the following bacteria: Prevotella denticola ATCC 33185, P. denticola AHN 33266, Porphyromonas gingivalis ATCC 33277, P. gingivalis AHN 24155, and Filifactor alocis ATCC 35896. The EPSs contained fractions with different molecular weights (51 and 841 kDa) and monosaccharides of glucose, galactose, and fructose. The EPSs showed antibiofilm activity in all the biofilm models tested. The EPSs may have inhibited bacterial aggregation and binding to hydrocarbons by reducing bacterial hydrophobicity. In conclusion, the EPSs of L. plantarum EIR/IF-1, which consists of two major fractions, exhibited antibiofilm activity against oral bacteria, which can be explained by the inhibitory effect of EPSs on the auto-aggregation and co-aggregation of bacteria and their binding to hydrocarbons.

Targeting Nrf2 with Probiotics and Postbiotics in the Treatment of Periodontitis.

Periodontitis is a destructive disease of the tooth-surrounding tissues. Infection is the etiological cause of the disease, but its extent and severity depend on the immune-inflammatory response of the host. Immune cells use reactive oxygen species to suppress infections, and there is homeostasis between oxidative and antioxidant mechanisms during periodontal health. During periodontitis, however, increased oxidative stress triggers tissue damage, either directly by activating apoptosis and DNA damage or indirectly by activating proteolytic cascades. Periodontal treatment aims to maintain an infection and inflammation-free zone and, in some cases, regenerate lost tissues. Although mechanical disruption of the oral biofilm is an indispensable part of periodontal treatment, adjunctive measures, such as antibiotics or anti-inflammatory medications, are also frequently used, especially in patients with suppressed immune responses. Recent studies have shown that probiotics activate antioxidant mechanisms and can suppress extensive oxidative stress via their ability to activate nuclear factor erythroid 2-related factor 2 (Nrf2). The aim of this narrative review is to describe the essential role of Nrf2 in the maintenance of periodontal health and to propose possible mechanisms to restore the impaired Nrf2 response in periodontitis, with the aid of probiotic and postbiotics.

The role of microbiota-derived postbiotic mediators on biofilm formation and quorum sensing-mediated virulence of Streptococcus mutans: A perspective on preventing dental caries.

Dental caries is not only one of the most prevalent diseases worldwide, but also a public health problem, undoubtedly. Among the various species of cariogenic bacteria, Streptococcus mutans is considered to be the major etiological pathogen of dental caries. The present study aimed to assess the influence of microbiota-derived postbiotic mediators (PMs) on the pathogenesis of dental caries. Within this aim, the antibacterial (agar diffusion method) and antibiofilm (crystal violet assay) characteristics of PMs derived from Lactiplantibacillus plantarum EIR/IF-1, Lactiplantibacillus curvatus EIR/DG-1, and Lactiplantibacillus curvatus EIR/BG-2 were analyzed against S. mutans (ATCC 25175). According to the results, PMs of the strain EIR/IF-1, isolated from infant feces showed the highest inhibitory effect (pH-dependent). Besides, sub-MIC doses of all PMs eliminated the biofilm formation following the co-treatment and pre-treatment assays. The reduction of cell viability and notable changes in biofilm formation were also confirmed both on glass coverslips and ex vivo human tooth surfaces by confocal laser scanning microscopy and scanning electron microscopy. Moreover, sub-MIC values of PMs down-regulated the expression of gtfC, comA, and comX, without any significant growth inhibition. Organic acids, fatty acids, and vitamins in PMs were also reported. Overall, these findings indicated the possible preventive role of microbiota-derived PMs in the pathogenesis of dental caries.

Investigation of antibacterial properties of polyacrylonitrile fibers modified by new functional groups and silver nanoparticles.

This work reports the surface modification of polyacrylonitrile (PAN) fibers by graft copolymerization to ensure the decoration of homogenous and dense Ag nanoparticles. Two facile and subsequent modification processes resulted in a PAN fiber composite with an intact fibrous structure, sufficiently conductive for antistatic application and antibacterial activity. In the first step, some chemically attractive monomers and monomer mixtures, such as acrylic acid (AA), AA-itaconic acid (AA-IA), AA-acrylamide (AA-AAm), were introduced to the fiber surface by grafting. The grafting process was evidenced by FTIR, 1H-NMR, and SEM techniques. The second step aimed to form a chelate structure by Ag+ ions with the coordination centers imparted to the PAN structure, and then, Ag nanoparticles (AgNPs) were decorated on the copolymer fiber surfaces by reducing with the NaBH4. The presence, distribution, and changes that occurred after the AgNPs decoration were also monitored by the SEM technique. It was obtained that the AgNPs could not be easily removed from the composites, which presented an appearance as if they were dyed with Ag. It was determined that the composite fibers gained a certain degree of conductivity with the surface resistivity value of 109-102 Ω/cm2. The antibacterial activity of the composites against E. coli and S. aureus was examined by the zone of inhibition test compared to their detergent-washed samples.

The Remarkable Antibiofilm Activity of the Sweet Tooth Mushroom, Hydnum repandum (Agaricomycetes), Displaying Synergetic Interactions with Antibiotics.

This study examines the antioxidant, anticancer, antimicrobial, and antibiofilm activities of Hydnum repandum, an edible and medicinal mushroom known as sweet tooth or wood hedgehog. H. repandum ethanolic extract had a high amount of myricetin and apigenin and displayed antiproliferative effects against the MCF-7 and HT-29 cell lines. Moreover, the extract displayed antimicrobial and antibiofilm activities against methicillin-resistant Staphylococcus aureus ATCC 43300, S. epidermidis ATCC 35984, Escherichia coli ATCC 25922, and Pseudomonas aeruginosa ATCC 27853. Synergetic interactions have been observed when antibiotics such as kanamycin and ampicillin are used together with mushroom extract. The minimum biofilm eradication concentration values were lower for H. repandum extract than for antibiotics. This study demonstrates that H. repandum has antibiofilm potential against biofilms and confronts antibiotic resistance.

Comparison of the chemical composition and biological effects of Clitocybe nebularis and Infundibulicybe geotropa.

Clitocybe mushrooms have long been recognized for their various therapeutic potential and medicinal properties. A few members of the genus are considered edible and many others are poisonous. This study investigated the ethanolic extracts obtained from C. nebularis (CN) and I. geotropa (IG) mushrooms for phenolic content and antioxidant, antiproliferative, antimicrobial, and antibiofilm activities. The data from ultra-performance liquid chromatography and Fourier transform infrared spectroscopy analysis of the mushrooms were presented for the first time. According to the results, both ethanolic extracts contain high levels of phenolic (catechin, myricetin, quercetin, rutin, gallic acid, vanillic acid) compounds. Fourier transform infrared spectroscopy results may suggest the presence of clitopycin in CN extract. The ethanol extract of CN scavenged about 79% and the IG 78% of the free 2,2-diphenyl-1-picrylhydrazyl radicals. Additionally, the CN and IG extracts inhibited glutathione-S-transferase by 10%-18% at all concentrations. The CN extract effectively inhibited aldose reductase by 30%-80% at all concentrations. Besides, the CN extract showed promising antiproliferative activity on HT-29 and MCF-7 cell lines. On the other hand, CN and IG extracts displayed inhibitory effects on some multidrug-resistant Gram-positive bacteria and effectively inhibited biofilm production. The obtained results showed that C. nebularis and I. geotropa extracts presented inhibition of biofilm production. Therefore, C. nebularis was demonstrated to be a potential source of natural medicine.

Mathematical Models for the Biofilm Formation of Geobacillus and Anoxybacillus on Stainless Steel Surface in Whole Milk.

Biofilm formation of Geobacillus thermodenitrificans, Geobacillus thermoglucosidans and Anoxybacillus flavithermus in milk on stainless steel were monitored at 55°C, 60°C, and 65°C for various incubation times. Although species of Geobacillus showed a rapid response and produced biofilm within 4 h on stainless steel, a delay (lag time) was observed for Anoxybacillus. A hyperbolic equation and a hyperbolic equation with lag could be used to describe the biofilm formation of Geobacillus and Anoxybacillus, respectively. The highest biofilm formation amount was obtained at 60°C for both Geobacillus and Anoxybacillus. However, the biofilm formation rates indicated that the lowest rates of formation were obtained at 60°C for Geobacillus. Moreover, biofilm formation rates of G. thermodenitrificans (1.2-1.6 Log10CFU/mL·h) were higher than G. thermoglucosidans (0.4-0.7 Log10CFU/mL·h). Although A. flavithermus had the highest formation rate values (2.7-3.6 Log10CFU/mL·h), this was attained after the lag period (4 or 5 h). This study revealed that modeling could be used to describe the biofilm formation of thermophilic bacilli in milk.

Weissella cibaria EIR/P2-derived exopolysaccharide: A novel alternative to conventional biomaterials targeting periodontal regeneration.

Healing and regeneration of periodontium are considered as a complex physiological process. Therefore, treatments need to be addressed with highly effective components modulating the multiple pathways. In this study, exopolysaccharide (EPS) produced by Weissella cibaria EIR/P2, was partially purified from the culture supernatant and subjected to characterization within the aim of evaluating its potential for periodontal regeneration. High-Performance Liquid Chromatography analysis revealed a single-peak corresponding to the glucose which identified the EPS as dextran. Fourier transform-infrared spectra were also displayed characteristic peaks for polysaccharides. According to the results of gel permeation/size exclusion-chromatography, the molecular mass was determined to be 8 × 106 Da. To clarify its anti-bacterial activity on Streptococcus mutans, effects on viability and biofilm formation was evaluated. At 50 mg/mL, dextran exhibited a bactericidal effect with 70% inhibition on biofilm formation. Besides, dose-dependent antioxidant effects were also detected. The efficacy of dextran in enhancing the viability of human periodontal ligament fibroblast cells (hPDLFCs) was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium-bromide (MTT) assay, and an increase was observed in the viability of hPDLFCs. In conclusion, dextran derived from W. cibaria can be potentially used as a multi-functional bioactive polymer in the design of new therapeutic strategies to promote healing and regeneration of periodontium.

Antioxidant, Anticancer, Antimicrobial, and Antibiofilm Properties of the Culinary-Medicinal Fairy Ring Mushroom, Marasmius oreades (Agaricomycetes).

This study is based on the phenolic composition and the antioxidant, anticancer, antimicrobial, and antibiofilm activities of the edible mushroom Marasmius oreades from Turkey. The phenolic composition of an M. oreades ethanol extract was measured by using the Folin-Ciocalteu method, aluminium chloride colorimetry, and ultraperformance liquid chromatography. The antioxidant activity was evaluated on the basis of DPPH radical scavenging activity. The effect of the M. oreades ethanol extract was also screened in order to determine glutathione-S-transferase, glutathione peroxidase, catalase, and superoxide dismutase enzyme activities. The antimicrobial activity of the mushroom extract was evaluated by using well diffusion and was based on the minimum inhibitory concentration. In addition, the antibiofilm potential of M. oreades was analyzed against Gram-positive and -negative bacteria. Finally, the anticancer effects of the mushroom extract were tested on colon (HT-29) and breast (MCF-7 and MDA-MB-231) cancer cell lines by using the MTT assay. The results revealed that the total amount of phenolics in the ethanol extract of M. oreades was 10.990 ± 0.0007 mg gallic acid equivalent/100 g, and the total amount of flavonoids was 1.139 ± 0.0052 mg quercetin equivalent/100 g. The ultraperformance liquid chromatography results indicated that the M. oreades ethanol extract contained various phenolic compounds: catechin, ferulic, gallic acid, and vanillic acid. The M. oreades ethanol extract scavenged about 80% of DPPH free radicals. It did not show any effect on the glutathione-S-transferase, glutathione peroxidase, and catalase enzyme activities, but its maximal concentration (10 mg/mL) increased superoxide dismutase activity (8%). The ethanol extract of M. oreades showed a moderate anticancer effect on the HT-29, MCF-7, and MDA-MB-231 cell lines. Although the ethanolic extract of the mushroom did not show sufficient antibacterial activity, it presented a strong antibiofilm effect against all studied pathogenic strains at the tested concentrations.

Anoxybacillus and Geobacillus biofilms in the dairy industry: effects of surface material, incubation temperature and milk type.

Anoxybacillus (A. flavithermus, A. kamchatkensis subsp. asachharedens, A. caldiproteolyticus and A. tepidamans) and Geobacillus (two strains of G. thermodenitrificans, G. thermoglucosidans and G. vulcanii) isolates and reference strains in whole milk were evaluated for their biofilm production on six different abiotic surfaces. G. thermodenitrificans DSM 465T had the highest cell counts (>4 log10 CFU cm-2) on glass and stainless steel (SS) at 55 and 65 °C, respectively. G. thermodenitrificans D195 had the highest counts on SS at 55 °C (>5 log10 CFU cm-2) and polyvinyl chloride (PVC) at 65 °C (>4 log10 CFU cm-2), indicating the existence of strain variation. The ideal surfaces for all strains were SS and glass at 55 °C, but their preferences were polystyrene and SS at 65 °C. Moreover, Anoxybacillus members were more prone to form biofilms in skim milk than in semi-skim and whole milk, whereas the results were the opposite for Geobacillus. Both the attachment and sporulation of Geobacillus in whole milk was higher than in semi-skim or skim milk. This study proposes that the surface material, temperature and milk type had a cumulative effect on biofilm formation.

Determination of the biofilm production capacities and characteristics of members belonging to Bacillaceae family.

The biofilm characteristics of many endospore-forming bacilli, especially the thermophiles are still unclear. In this study, a detailed identification and description of biofilm production characteristics of totally 145 isolates and reference strains belonging to Bacillaceae family, displaying thermophilic (n = 115), facultative thermophilic (n = 24) and mesophilic (n = 6) growth from genera Anoxybacillus, Geobacillus, Thermolongibacillus, Aeribacillus, Brevibacillus, Paenibacillus and Bacillus were presented. The incubation temperatures were adjusted to 37, 45 and 55-65 °C for mesophiles, facultative thermophiles, and thermophiles, respectively. The bacilli were evaluated based on their colony morphotypes on Congo red (CR) agar, their complex exopolysaccharide production on calcofluor supplemented tryptic soy agar, and as well as their pellicle formation at the liquid-air surface in tryptic soy broth cultures. Their biofilm production capabilities were also tested on abiotic surfaces of both polystyrene and stainless steel by crystal violet binding assay and viable biofilm cell enumerations, respectively. As a result, the biofilm production capacities of Bacillaceae members from genera to species level, the effects of osmolarity, temperature, incubation time and abiotic surfaces on biofilm formation as well as the CR morphotypes associated with the biofilm production were able to reveal in a wide group of bacilli. Besides, general enrichment-inoculation approaches and methodologies were also offered, which allow and facilitate the screening and determining the biofilm producing endospore forming bacilli.

Biofilm characteristics and evaluation of the sanitation procedures of thermophilic Aeribacillus pallidus E334 biofilms.

The ability of Aeribacillus pallidus E334 to produce pellicle and form a biofilm was studied. Optimal biofilm formation occurred at 60 °C, pH 7.5 and 1.5% NaCl. Extra polymeric substances (EPS) were composed of proteins and eDNA (21.4 kb). E334 formed biofilm on many surfaces, but mostly preferred polypropylene and glass. Using CLSM analysis, the network-like structure of the EPS was observed. The A. pallidus biofilm had a novel eDNA content. DNaseI susceptibility (86.8% removal) of eDNA revealed its importance in mature biofilms, but the purified eDNA was resistant to DNaseI, probably due to its extended folding outside the matrix. Among 15 cleaning agents, biofilms could be removed with alkaline protease and sodium dodecyl sulphate (SDS). The removal of cells from polypropylene and biomass on glass was achieved with combined SDS/alkaline protease treatment. Strong A. pallidus biofilms could cause risks for industrial processes and abiotic surfaces must be taken into consideration in terms of sanitation procedures.

A Comparative Study: Taxonomic Grouping of Alkaline Protease Producing Bacilli.

Alkaline proteases have biotechnological importance due to their activity and stability at alkaline pH. 56 bacteria, capable of growing under alkaline conditions were isolated and their alkaline protease activities were carried out at different parameters to determine their optimum alkaline protease production conditions. Seven isolates were showed higher alkaline protease production capacity than the reference strains. The highest alkaline protease producing isolates (103125 U/g), E114 and C265, were identified as Bacillus licheniformis with 99.4% and Bacillus mojavensis 99.8% based on 16S rRNA gene sequence similarities, respectively. Interestingly, the isolates identified as Bacillus safensis were also found to be high alkaline protease producing strains. Genotypic characterizations of the isolates were also determined by using a wide range of molecular techniques (ARDRA, ITS-PCR, (GTG)5-PCR, BOX-PCR). These different techniques allowed us to differentiate the alkaliphilic isolates and the results were in concurrence with phylogenetic analyses of the 16S rRNA genes. While ITS-PCR provided the highest correlation with 16S rRNA groups, (GTG)5-PCR showed the highest differentiation at species and intra-species level. In this study, each of the biotechnologically valuable alkaline protease producing isolates was grouped into their taxonomic positions with multi-genotypic analyses.