Colony-forming Unit Counts Research Articles

Strong Activity and No Resistance Induction Exerted by Cell-Free Supernatants from Lacticaseibacillus rhamnosus against Mono-Species and Dual-Species Biofilms of Wound Pathogens in In Vivo-like Conditions.

It is widely agreed that microbial biofilms play a major role in promoting infection and delaying healing of chronic wounds. In the era of microbial resistance, probiotic strains or their metabolic products are emerging as an innovative approach for the treatment of hard-to-heal (chronic) wounds due to their antimicrobial, healing, and host immune-modulatory effects. In this study, we aimed to investigate the potential of cell-free supernatants (CFS) from Lacticaseibacillus rhamnosus GG against mono- and dual-species biofilms of wound pathogens in a 3D in vitro infection model. Mature biofilms of Pseudomonas aeruginosa and Staphylococcus aureus were obtained on collagen scaffolds in the presence of a simulant wound fluid (SWF) and treated with CFS at different doses and time intervals. At 1:4 dilution in SWF, CFS caused a marked reduction in the colony forming-unit (CFU) numbers of bacteria embedded in mono-species biofilms as well as bacteria released by the biofilms in the supernatant. CFU count and electron microscopy imaging also demonstrated a marked antibiofilm effect against dual-species biofilms starting from 8 h of incubation. Furthermore, CFS exhibited acceptable levels of cytotoxicity at 24 h of incubation against HaCaT cells and, differently from ciprofloxacin, failed to induce resistance after 15 passages at sub-inhibitory concentrations. Overall, the results obtained point to L. rhamnosus GG postbiotics as a promising strategy for the treatment of wound biofilms.

Development and validation of quantitative PCR assays for HIV-associated cryptococcal meningitis in sub-Saharan Africa: a diagnostic accuracy study

HIV-associated cryptococcal meningitis is the second leading cause of AIDS-related deaths, with a 10-week mortality rate of 25-30%. Fungal load assessed by colony-forming unit (CFU) counts is used as a prognostic marker andto monitor response to treatment in research studies. PCR-based assessment of fungal load could be quicker andless labour-intensive. We sought to design, optimise, and validate quantitative PCR (qPCR) assays for the detection, identification, and quantification of Cryptococcus infections in patients with cryptococcal meningitis in sub-Saharan Africa. We developed and validated species-specific qPCR assays based on DNA amplification of QSP1 (QSP1A specific to Cryptococcus neoformans, QSP1B/C specific to Cryptococcus deneoformans, and QSP1D specific to Cryptococcus gattii species) and a pan-Cryptococcus assay based on a multicopy 28S rRNA gene. This was a longitudinal study that validated the designed assays on cerebrospinal fluid (CSF) of 209 patients with cryptococcal meningitis at baseline (day 0) and during anti-fungal therapy (day 7 and day 14), from the AMBITION-cm trial in Botswana and Malawi (2018-21). Eligible patients were aged 18 years or older and presenting with a first case of cryptococcal meningitis. When compared with quantitative cryptococcal culture as the reference, the sensitivity of the 28S rRNA was 98·2% (95% CI 95·1-99·5) and of the QSP1 assay was 90·4% (85·2-94·0) in CSF at day 0. Quantification of the fungal load with QSP1 and 28S rRNA qPCR correlated with quantitative cryptococcal culture (R2=0·73 and R2=0·78, respectively). Both Botswana and Malawi had a predominant C neoformans prevalence of 67% (95% CI 55-75) and 68% (57-73), respectively, and lower C gattii rates of 21% (14-31) and 8% (4-14), respectively. We identified ten patients that, after 14 days of treatment, harboured viable but non-culturable yeasts based on QSP1 RNA detection (without any positive CFU in CSF culture). QSP1 and 28S rRNA assays are useful in identifying Cryptococcus species. qPCR results correlate well with baseline quantitative cryptococcal culture and show a similar decline in fungal load during induction therapy. These assays could be a faster alternative to quantitative cryptococcal culture to determine fungal load clearance. The clinical implications of the possible detection of viable but non-culturable cells in CSF during induction therapy remain unclear. European and Developing Countries Clinical Trials Partnership; Swedish International Development Cooperation Agency; Wellcome Trust/UK Medical Research Council/UKAID Joint Global Health Trials; and UKNational Institute for Health Research.

Antimicrobial and optical properties of a new biogenic silica-coated silver nanoparticles incorporated into experimental resin.

Evaluate the effects of incorporating silica-coated silver nanoparticles (Ag@SiO2 NPs) into odontological clinic resin materials. Silver nanoparticles coated with silicon dioxide were added to the experimental resin matrix at 1, 3, and 5wt%. Degree of conversion (DC), optical properties (total transmittance and color change), and microstructural analysis were evaluated. Materials were tested for silver ion release, cytotoxicity in dental pulp fibroblasts, Streptococcus mutans biofilm growth by Colony-Forming Unit (CFU) and confocal laser scanning microscopy (CLSM). Groups had a similar DC, despite significant differences observed in transmittance and color change analysis for all groups with NPs. Silver ion release values were below the detection limit after 72h for all groups, and NPs incorporation did not show a statistical difference from the control on pulp fibroblasts assay. After 72h, the CFU count was significantly reduced by 74% from 3wt% of Ag@SiO2NPs. CLSM evaluation on S. mutans colonies showed a dose-dependent decrease in the emitted fluorescence. The application of Ag@SiO2 NPs in a resinous matrix, demonstrates a significant reduction of S. mutans CFU in oral biofilm, at concentrations from 3wt%, without an increase in cytotoxicity. The reduced transmittance values did not affect the DC, although a significant color change was perceived in all concentrations. Key words:Nanoparticles, Silver Compounds, Composite Dental Resin, Anti-Bacterial Agent, Optical Imaging.

Evaluation of the Effectiveness of Disinfectants on Impression Materials.

Background The disinfection of dental impression materials is a cornerstone of infection control in dental practice. This study aimed to evaluate the effectiveness of two disinfectants, sodium hypochlorite and iodophor, on alginate and silicone impression materials, which are prone to microbial contamination. Methods The study was structured into two main groups based on the impression material: Group I (alginate) and Group II (addition silicone), each further subdivided into two subgroups for disinfection with sodium hypochlorite and iodophor. For each subgroup, initial microbial swabs were taken before any treatment, followed by a second swab after rinsing and a final swab after disinfection. The mean colony-forming unit (CFU) counts, standard deviations, and standard errors of the mean were calculated for each stage of treatment. Results Prior to disinfection, Group I had a mean CFU count of 2,529.40, while Group II had a lower mean CFU of 1,417.40. After rinsing, there was a significant decrease in CFUs in both groups, with Group I at 1,337.10 and Group II at 415.10. Post-disinfection, Group I showed a mean CFU count of 73.00 for sodium hypochlorite and 0.00 for iodophor. Similarly, Group II achieved a CFU reduction of 99.00 with sodium hypochlorite and 0.00 with iodophor, demonstrating a marked reduction in microbial presence. Conclusion Iodophor was exceptionally effective in disinfecting both alginate and silicone impression materials, eliminating all detectable CFUs. Sodium hypochlorite also significantly reduced microbial counts but was not as effective as iodophor. Rinsing prior to disinfection was instrumental in reducing the microbial load, underscoring its importance in the disinfection protocol.

Cryptocarya moschata extract decreases single and mixed biofilms on acrylic resins.

This study proposed to assess the effect of Cryptocarya moschata extract on single and mixed biofilms formed on denture base and reline acrylic resin. Single and mixed biofilms of Candida albicans and Streptococcus mutans were formed on the samples and treated with C. moschata extract; Nystatin solution at 100,000 IU/mL or Penicillin antibiotic solution at 100,000 IU/mL; or PBS solution. Antimicrobial activity was analyzed by counting colony-forming units, metabolism assay, assessment of protein components of the biofilm matrix, and of cell viability using confocal laser scanning microscopy (CLSM). Data were submitted to ANOVA and Tukey's post-test (α = 0.05). Cryptocarya moschata extract reduced cell viability of C. albicans and S. mutans single and mixed biofilms formed on samples. For all types of biofilms in the C. moschata group, there was a log reduction of the biofilm, proven by the Alamar Blue assay. Analyzing the extracellular matrix protein components, groups treated with the extract exhibited a lower level of fluorescence compared to the PBS groups. Reduction in thickness biofilm and viable cells was perceptible in the C. moschata group when assessing through CLSM. Cryptocarya moschata extract reduced the single and mixed biofilms of C. albicans and S. mutans on acrylic resins.

Orthodontic adhesive loaded with different proportions of ZrO2 silver-doped nanoparticles: An in vitro μTBS, SEM, EDX, FTIR, and antimicrobial analysis.

Zirconium dioxide silver-doped nanoparticles (ZrO2AgDNPs) impacts the adhesive material in terms of its physical characteristics, antimicrobial properties, degree of conversion (DC), and micro-tensile bond strength (μTBS) of orthodontic brackets to the enamel surface. A comprehensive methodological analysis utilizing a range of analytical techniques, including scanning electron microscopy coupled with energy-dispersive x-ray spectroscopy (EDX), Fourier-transform infrared (FTIR) spectroscopy, DC analysis, and μTBS testing. A light-curable orthodontic adhesive, specifically Transbond XT, was combined with ZrO2AgDNPs at 2.5% and 5%. As a control, an adhesive with no incorporation of ZrO2AgDNPs was also prepared. The tooth samples were divided into three groups based on the weightage of NPs: group 1: 0% ZrO2AgDNPs (control), group 2: 2.5 wt% ZrO2AgDNPs, and group 3: 5 wt% ZrO2AgDNPs. EDX graph demonstrated silver (Ag), Zirconium (Zr), and Oxygen (O2), The antibacterial efficacy of adhesives with different concentrations of NPs (0%, 2.5%, and 5%) was assessed using the pour plate method. The FTIR spectra were analyzed to identify peaks at 1607 cm-1 corresponding to aromatic CC bonds and the peaks at 1638 cm-1 indicating the presence of aliphatic CC bonds. The μTBS was assessed using universal testing machine (UTM) and bond failure of orthodontic brackets was seen using adhesive remanent index (ARI) analysis. Kruskal-Wallis test assessed the disparities in survival rates of Streptococcus mutans. Analysis of variance (ANOVA) and post hoc Tukey multiple comparisons test calculated μTBS values. The lowest μTBS was observed in group 1 adhesive loaded with 0% ZrO2AgDNPs (21.25 ± 1.22 MPa). Whereas, the highest μTBS was found in group 3 (26.19 ± 1.07 MPa) adhesive loaded with 5% ZrO2AgDNPs. ZrO2AgDNPs in orthodontic adhesive improved μTBS and has acceptable antibacterial activity against S mutans. ZrO2AgDNPs at 5% by weight can be used in orthodontic adhesive alternative to the conventional method of orthodontic adhesive for bracket bonding. RESEARCH HIGHLIGHTS: The highest μTBS was found in orthodontic adhesive loaded with 5% ZrO2AgDNPs. ARI analysis indicates that the majority of the failures fell between 0 and 1 among all investigated groups. The colony-forming unit count of S. mutans was significantly less in orthodontic adhesive loaded with nanoparticles compared with control. The 0% ZrO2AgDNPs adhesive showed the highest DC.

Electromagnetic fields effects on microbial growth in cocoa fermentation: A controlled experimental approach using established growth models

Understanding the effects of electromagnetic fields is crucial in the fermentation of cocoa beans, since through precise control of fermentation conditions the sensory and nutritional properties of cocoa beans could be improved. This study aimed to evaluate the effect of oscillating magnetic fields (OMF) on the kinetic growth of the core microbial communities of the Collections Castro Naranjal (CCN 51) cocoa bean. The data was obtained by three different models: Gompertz, Baranyi, and Logistic. The cocoa beans were subjected to different OMF strengths ranging from 0 mT to 80 mT for 1 h using the Helmholtz coil electromagnetic device. The viable microbial populations of lactic acid bacteria (LAB), acetic acid bacteria (AAB), and yeast (Y) were quantified using the colony-forming unit (CFU) counting method. The logistic model appropriately described the growth of LAB and Y under magnetic field exposure. Whereas the Baranyi model was suitable for describing AAB growth. The microbial populations in cocoa beans exposed to magnetic fields showed lower (maximum specific growth rate (μmax), values than untreated controls, with AAB exhibiting the highest average growth rate value at 5 mT and Y having the lowest average maximum growth rate value at 80 mT. The lower maximum specific growth rates and longer lag phases when exposed to magnetic fields compared to controls demonstrate the influence of magnetic fields on microbial growth kinetics.

Biofilm formation by Staphylococcus pseudintermedius on titanium implants

Osteomyelitis often involves Staphylococcus spp. as the isolated genus in domestic animal cases. Implant-related infections, frequently associated with biofilm-forming microorganisms like staphylococci species, necessitate careful material selection. This study assessed biofilm formation by Staphylococcus pseudintermedius on titanium nuts used in veterinary orthopaedic surgery. Biofilm quantification employed safranin staining and spectrophotometric measurement, while bacterial counts were determined in colony-forming units (CFU). Scanning Electron Microscopy (SEM) evaluated the biofilm morphology on the surface of titanium nuts. All samples had CFU counts. Absorbance values that evidence biofilm formation were observed in seven of the eight samples tested. SEM images revealed robust bacterial colonization, and significant extracellular polymeric substance production, and the negative control displayed surface irregularities on the nut. Whole genome sequencing revealed accessory Gene Regulator (agr) type III in six samples, agr IV and agr II in two each. Genes encoding hlb, luk-S, luk-F, siet, se_int, and the icaADCB operon were identified in all sequenced samples. Other exfoliative toxins were absent. Biofilm formation by S. pseudintermedius was detected in all samples, indicating the susceptibility of orthopaedic titanium alloys to adhesion and biofilm formation by veterinary species. The biofilm formation capacity raises concerns about potential post-surgical complications and associated costs.

Microbiome-friendly PS/PVP electrospun fibrous membrane with antibiofilm properties for dental engineering.

Dental caries is one of the most prevalent and biofilm-associated oral diseases in humans. Streptococcus mutans, with a high ability to form biofilms by adhering to hard surfaces, has been established as an important etiological agent for dental caries. Therefore, it is crucial to find a way to prevent the formation of cariogenic biofilm. Here, we report an electrospun fibrous membrane that could inhibit the adhesion and biofilm formation of S. mutans. Also, the polystyrene (PS)/polyvinyl pyrrolidone (PVP) electrospun fibrous membrane altered the 3D biofilm architecture and decreased water-insoluble extracellular polysaccharide production. Notably, the anti-adhesion mechanism which laid in Coulomb repulsion between the negatively charged PS/PVP electrospun fibrous membrane and S. mutans was detected by zeta potential. Furthermore, metagenomics sequencing analysis and CCK-8 assay indicated that PS/PVP electrospun fibrous membrane was microbiome-friendly and displayed no influence on the cell viability of human gingival epithelial cells and human oral keratinocytes. Moreover, an in vitro simulation experiment demonstrated that PS/PVP electrospun fibrous membrane could decrease colony-forming unit counts of S. mutans effectively, and PS/PVP electrospun fibrous membrane carrying calcium fluoride displayed better anti-adhesion ability than that of PS/PVP electrospun fibrous membrane alone. Collectively, this research showed that the PS/PVP electrospun fibrous membrane has potential applications in controlling and preventing dental caries.

Comparative Evaluation of Antimicrobial Activity of 0.2% Lemongrass Oil and 0.2% Chlorhexidine as a Preprocedural Mouth Rinse: A Randomised Controlled, Parallel Design, Split-mouth Clinical Trial

Introduction: The aerosols produced by an ultrasonic scaler contain bacteria that may enter the respiratory systems of both dental surgeons and patients. Lemongrass oil is one of the essential oils with enormous antibacterial, antifungal, antioxidant, anti-inflammatory, and antipyretic effects. Aim: To evaluate and compare the efficacy of commercially available preprocedural mouth rinses, containing 0.2% lemongrass oil mouthwash and a 0.2% chlorhexidine gluconate solution, in reducing the levels of viable bacteria in aerosols. Materials and Methods: This single-centre double-blinded randomised controlled parallel design split-mouth clinical trial was planned among patients in the Department of Periodontics attending Vishnu Dental College and Hospital in Bhimavaram, Andhra Pradesh, India. The study was conducted for 40 days, from August 2022 to October 2022. Two distinct types of preprocedural mouth rinses (lemongrass oil and chlorhexidine) were used on 56 participants with Stage II or III and Grade B periodontitis. A total of 56 subjects were randomly assigned to two groups: Group A (0.2% lemongrass oil) and Group B (0.2% chlorhexidine), using a simple coin toss method. Colonyforming units (CFUs) with and without rinsing in each group were assessed and studied. CFUs collected on blood agar plates were counted during ultrasonic scaling both with and without rinsing. International Business Machine (IBM) Statistical Package for Social Sciences (SPSS) version 20.0 was used for data analysis. Paired t-tests and independent t-tests were used for intragroup and intergroup comparisons for both groups. Results: The mean CFU count in the chlorhexidine group without rinsing was 259.6±26.6, and with rinsing, it was 140.7±22.6, which was statistically significant (p-value < 0.001). In the lemongrass oil group, the CFU count without rinsing was 263.5±26.9, and with rinsing, it was 147.18±33.82, which was also statistically significant (p-value<0.001). In the intergroup comparison, the p-values for both groups-using preprocedural rinse (Group A vs. Group B for quadrants 2 and 3) and without preprocedural rinse (Group A vs. Group B for quadrants 1 and 4)-were 0.40 and 0.58, respectively, which were not significant. Conclusion: Both preprocedural mouth rinses were equally effective in reducing microbial counts in aerosols during ultrasonic scaling.

A low-shrinkage-stress and anti-bacterial adherent dental resin composite: physicochemical properties and biocompatibility.

Polymerisation shrinkage and biofilm accumulation are the two main problems associated with dental resin composites (DRCs) that induce secondary caries, which can cause restoration failure. Polymerisation shrinkage can lead to microleakage gaps between the tooth and the DRCs, causing the aggregation of bacteria and development of secondary caries. Reducing the shrinkage stress (SS) and improving the resistance to bacterial adhesion have always been the focus of this field in modifying DRCs. A thiol-ene resin system can effectively reduce the polymerisation SS via its step-growth mechanism for delaying the gel point. Fluorinated compounds can reduce the surface free energies, thereby reducing bacterial adhesion. Thus, in this study, a range of mass fractions (0, 10, 20, 30, and 40 wt%) of a fluorinated thiol-ene resin system were added to a fluorinated dimethacrylate resin system/tricyclo decanedimethanol diacrylate to create a fluorinated methacrylate-thiol-ene ternary resin matrix. DRCs were prepared using the obtained ternary resin matrix, and their physical and chemical properties, effect on bacterial adhesion, and biocompatibility were investigated. The results demonstrated that the volumetric shrinkage and SS of the DRCs were reduced with no reduction in conversion degree even after the thiol-ene resin system was added. All DRC-based fluorinated resin systems exhibited an excellent anti-bacterial adhesion effect, as evidenced by the colony-forming unit counts, live/dead bacterial staining, and crystal violet staining tests against Streptococcus mutans (S. mutans). The genetic expressions associated with the bacterial adhesion of S. mutans were substantially affected after being cultured with fluorinated DRCs. All fluorinated DRCs demonstrated good biocompatibility through the in vitro cytotoxicity test and live/dead staining images of the L-929 cells. The above results illustrate that the DRCs based on the fluorinated methacrylate-thiol-ene resin matrix can be potentially applied in clinical practice due to their low SS and anti-bacterial adhesion effect.

The antimicrobial activity of an antiseptic soap against Candida Albicans and Streptococcus Mutans single and dual-species biofilms on denture base and reline acrylic resins.

To evaluate the effect of antiseptic soap on single and dual-species biofilms of Candida albicans and Streptococcus mutans on denture base and reline resins. Samples of the resins were distributed into groups (n = 9) according to the prevention or disinfection protocols. In the prevention protocol, samples were immersed in the solutions (Lifebuoy, 0.5% sodium hypochlorite solution and PBS) for 7, 14 and 28 days before the single and dual-species biofilms formation. Overnight denture disinfection was simulated. In the disinfection protocol, samples were immersed in the same solutions during 8 hours after the single and dual-species biofilms formation. Antimicrobial activity was analyzed by counting colony-forming units (CFU/mL) and evaluating cell metabolism. Cell viability and protein components of the biofilm matrix were evaluated using confocal laser scanning microscopy (CLSM). Data were submitted to ANOVA, followed by Tukey's post-test (α = 0.05) or Dunnett's T3 multiple comparisons test. In the prevention protocol, Lifebuoy solution effectively reduced the number of CFU/mL of both species. In addition, the solution decreased the cell metabolism of the microorganisms. Regarding disinfection protocol, the Lifebuoy solution was able of reduce approximately of 2-3 logs for all the biofilms on the denture base and reline resin. Cellular metabolism was also reduced. The images obtained with CLSM corroborate these results. Lifebuoy solution was effective in reducing single and dual-species biofilms on denture base and reline resins.

Fluoride-resistant Streptococcus mutans within cross-kingdom biofilms support Candida albicans growth under fluoride and attenuate the in vitro anti-caries effect of fluorine.

Fluoride-resistant Streptococcus mutans (S. mutans) might affect the ecological balance of biofilms in the presence of fluoride. We used a S. mutans and Candida albicans (C. albicans) cross-kingdom biofilm model to investigate whether fluoride-resistant S. mutans in biofilms would support C. albicans growth under fluoride stress and attenuate the in vitro anti-caries effect of fluorine. The impact of fluoride-resistant S. mutans on formation of cross-kingdom biofilms by S. mutans and C. albicans in the presence of fluoride was investigated in vitro using the crystal violet staining assay. Biofilm constitution was determined using colony-forming unit (CFU) counts and fluorescent in situ hybridization (FISH). Extracellular polysaccharide (EPS) generation in biofilms was determined by EPS/bacterial dying and water-insoluble polysaccharide detection. Acid production and demineralization were monitored using pH, lactic acid content, and transversal microradiography (TMR). The gene expression of microorganisms in the cross-kingdom biofilm was measured using qRT-PCR. Our results showed that both C. albicans and fluoride-resistant S. mutans grew vigorously, forming robust cross-kingdom biofilms, even in the presence of sodium fluoride (NaF). Moreover, fluoride-resistant S. mutans-containing cross-kingdom biofilms had considerable cariogenic potential for EPS synthesis, acid production, and demineralization ability in the presence of NaF than fluoride-sensitive S. mutans-containing biofilms. Furthermore, the gene expression of microorganisms in the two cross-kingdom biofilms changed dissimilarly in the presence of NaF. In summary, fluoride-resistant S. mutans in cross-kingdom biofilms supported C. albicans growth under fluoride and might attenuate the anti-caries potential of fluorine by maintaining robust cross-kingdom biofilm formation and cariogenic virulence expression in vitro in the presence of NaF.

Evaluation of aloe vera gel as a local drug delivery agent in the treatment of chronic periodontitis: A clinical and microbiological study

Abstract Background: Local delivery of antibacterial agents into periodontal pockets provides a multitude of benefits over systemic antimicrobials resulting in the increased preference for such use among periodontal health-care providers. Aim: The study aims to determine the antimicrobial efficacy of adjunctive use of the herbal extract “aloe vera gel” (Aloe vera (L.) Burm. f.) along with scaling and root planing (SRP) in patients with chronic periodontitis. Materials and methods: This prospective, single-blind randomized split-mouth clinical trial was done among 22 subjects diagnosed with chronic periodontitis. Periodontal parameters and mean log counts of porphyromonas gingivalis (P. gingivalis) evaluated using real-time polymerase chain reaction were recorded. SRP was performed both at control and test sites. The evaluation of clinical and microbiological parameters was done at baseline and 3 months’ post-intervention. Data were analyzed using Statistical Package for Social Sciences (SPSS) V.20 software. Results: While significant improvement in both clinical and microbiological parameters from baseline to three months was observed in both the test and control sites, the mean reduction in plaque index, periodontal pocket depth, and mean log colony-forming unit counts of P. gingivalis were significantly higher in the test sites compared to the control sites. Conclusion: The results of this study reject the null hypothesis in favor of the alternative hypothesis that adjunctive application of locally delivered aloe vera gel for the treatment of chronic periodontitis enhances the beneficial effects of mechanical therapy.

Comparative Evaluation of Antibacterial Activity of Three Different Universal Bonding Agents Against Streptococcus mutans and Enterococcus faecalis: An In Vitro Study

Abstract Aims: The purpose of this work is to assess the antibacterial activity of three distinct bonding systems, Single Bond Universal, Tetric N-Bond Universal, and One Coat 7.0 on Streptococcus mutans and Enterococcus faecalis. Materials and Methods: The antibacterial efficacy of dentin bonding systems against S. mutans and E. faecalis was investigated in this study. Standardized inoculums and selective media were used in this investigation, and the components that were analyzed were immediately used on the cultures. Dentin bonding system (4 mm × 4 mm) discs were created, cured, cleaned, and incubated with bacterial inoculums for 48 h. A colony counter was used to measure the colony-forming unit count. The dentin bonding agents evaluated in this study revealed different inhibitory effects. Statistical Analysis Used: Submitted to the analysis of variance. Results: Given the constraints of the current in vitro investigation, it was determined that the One Coat 7.0 Universal (Coltene) bonding agent showed the highest antibacterial activity against both S. mutans and E. faecalis. Conclusion: The dentin bonding agents taken in this study revealed different inhibitory effects on bacterial growth, in which One Coat 7.0 Universal (Coltene) bonding agent performed the highest antibacterial effect against both the bacteria taken in the study, that is S. mutans and E. faecalis. among all the three bonding agents taken.

Chip-Based Molecular Evaluation of a DNA Extraction Protocol for Candida Species from Positive Blood Cultures.

The diagnosis of Candida bloodstream infection (BSI) may rely on a PCR-based analysis of a positive blood culture (PBC) obtained from the patient at the time of BSI. In this study, a yeast DNA extraction protocol for use on PBCs was developed and evaluated with the molecular mouse (MM) yeast blood (YBL) chip-based PCR assay, which allowed us to detect nine medically relevant Candida species. We studied 125 simulated or clinical PBCs for Candida species. A positive correlation between the DNA concentration and colony-forming unit count was found for simulated (Spearman's ρ = 0.58; p < 0.0001) and clinical (Spearman's ρ = 0.23, p = 0.09) PBCs. The extracted DNA yielded positive results with the MM YBL chip assay that agreed with the Candida species-level identification results for 63 (100%) of 63 isolates from simulated PBCs and 66 (99.5%) of 67 isolates from clinical PBCs. The false-negative result was for one C. tropicalis isolate that grew together with C. albicans in PBC. None of the 30 (Candida)-negative clinical BCs included as negative controls yielded a positive result with the MM YBL chip assay. Our DNA extraction protocol for the Candida species couples efficiency and simplicity together. Nevertheless, further studies are needed before it can be adopted for use with the MM YBL chip assay.

Analysis of the quantitative composition of the dental biofilm depending on the state of the hard dental tissues

The microflora of the oral cavity affects not only the development and course of the pathology of the hard dental tissues but also the effectiveness of its treatment and prevention. Therefore, it is equally important for a dentist to determine the species composition of microorganisms and their quantitative characteristics. The aim of the work was to study the quantitative composition of the planktonic form of the dental biofilm of the vestibular surface of the cervical teeth in young patients; its analysis in patients with a wedge-shaped defect, cervical caries and clinically intact teeth. A microbiological study of the dental biofilm of intact premolars of 33 caries-resistant patients (average age [23.90±5.34] years) was conducted. All patients were divided into three groups depending on the condition of the hard dental tissues. The dental biofilm removed from the lesion was additionally examined in patients with wedge-shaped defects. Microorganisms were identified by morphological, tinctorial and biochemical characteristics. A quantitative study of the population level was carried out by counting colony-forming units (CFU) in one gram of biomaterial which was expressed as a decimal logarithm (lg CFU/g). An imbalance between representatives of pathological and normal microflora was determined. Patients with cervical caries had a significantly higher chance of having Str. mutans and Lactobacterium in the dental biofilm, and a lower chance Str. γ-haemolyticus (p&lt;0.05). It confirms the role of Str. mutans and Lactobacterium in the etiology of cervical dental caries. There were no significant differences between the quantitative composition of the dental biofilm of patients of different groups that is probably related to the study of its planktonic component, the criteria for forming groups and the high level of caries resistance of the examined enamel. The obtained results can be useful in prescribing treatment and preventive measures for patients with cervical pathology of the teeth as well as in evaluating their effectiveness. In our opinion, it will contribute to a significant reduction in its prevalence and intensity among young people. Keywords: microorganisms, оral cavity, wedge-shaped defect, cervical caries.

A Randomized Double-blind Placebo-controlled Study on the Efficacy of Oregano (Origanum Vulgare) Leaf Ointment on the Reduction of 24-hour Colony Forming Units of Staphylococcus Aureus in Infected Open Wounds of Children Aged 8-12 Years Old

The practice of traditional medicine has been a vital part of Philippine medical practice, and, as a consequence, has led to innumerable researches conducted by the Department of Health (DOH) as to their role in the Filipino well-being. Out of the 10 named herbal plants studied by the department, oregano (Origanum vulgare) was chosen for its uses on the disorders of the skin, gastrointestinal and respiratory tract in the communities; and was therefore used in this study for the evaluation of its antimicrobial property, formulated as a leaf ointment against Staphylococcus aureus. OBJECTIVES: Specifically, this research aimed to determine a significant difference between the colony forming units (CFUs) on infected wounds treated with the placebo and oregano (Origanum vulgare) ointments. METHODOLOGY: A randomized double-blind placebo-controlled research study was conducted to the 60 infected wounds of the students of Dona Juana Actub Lluch Memorial Central School and Ubaldo Laya Memorial Central School with respondents aging 8-12 years old. Pre-treatment wound swabbing was done for 24-hour microbial culture, from which CFUs were counted. Post-treatment swabbing was performed after 5 days of treatment and subsequent CFUs were obtained. Using 0.05 level of significance, Mann-Whitney Pairwise Comparison Test was used to determine the significant difference in CFU count reduction between the two treatments. Findings showed reduction of CFUs in both ointments with greater reduction of CFUs in wounds treated Oregano leaf ointment as compared to those with the placebo. Statistical analysis of the study revealed that there was a significant difference between Placebo and Oregano (Origanum vulgare) on the reduction of CFUs in a 24-hour bacterial culture of infected open wounds.

Synthesis, Characterization, and Antibacterial Efficacy of Borosilicate Compound against Escherichia coli

In this study, a glassy borosilicate compound was synthesized using recycled glass and natural clays. Even though glass recycling is the generally accepted standard practice for managing glass waste, fine fractions of container soda-lime glass or cullet of other compositions are still disposed of in landfills. Thus, advanced upcycled products that offer greater economic motivation for implementation in industry may be the key to success, but these are frequently linked to alternative methods of product synthesis. Here, a simple and facile route of borosilicate compound production has been synthesized and characterized. The physicochemical characterization of the compounds was carried out to determine their properties and the antibacterial efficacy of the synthesized compound against Escherichia coli (E. coli) was investigated. The structural and spectroscopic characteristics were identified as a compound that conformed to quartz, cristobalite, and silicon hexaboride (SiB6). For the antibacterial activity, two test types were typically performed; in the first one, the dilutions of the grind were combined with chloramphenicol at a concentration of 20 µg/mL to perform a synergistic action against the bacteria and in the second one, only the amorphous borosilicate compound was tested against E. coli ATCC 25922 strains. The treatments applied considered the dilutions from 8 to 40 µg/mL. The minimum inhibitory concentration (MIC) sensitivity tests began with incubation at 37 °C in the tubes and subsequent seeding in Petri dishes for colony-forming unit (CFU) counting. The results obtained indicated that the samples possessed a productive antibacterial effect, which support their use in various biomedical applications.

Organotypic model of the gingiva for studying bacterial and viral pathogens implicated in periodontitis

ABSTRACT Background Three-dimensional (3D) tissue models bridge the gap between conventional two-dimensional cell cultures and animal models. The aim of this study was to develop an organotypic 3D gingival (OTG) model to provide a tool to investigate bacterial and viral pathogens in periodontitis. Methods The OTG model composed of gingival fibroblasts (GFs) and telomerase-immortalized gingival keratinocytes (TIGKs) was constructed and applied to study infections by Porphyromonas gingivalis and herpes simplex virus 1 (HSV-1). Immunohistochemical staining, confocal microscopy, qPCR, titration techniques, and colony-forming unit counts were applied to interrogate epithelial markers expression, monitor P. gingivalis and HSV-1 presence, and evaluate the immune response along with the efficiency of antimicrobial drugs. Results The OTG model resembled the morphology of the human gingiva. During infection, both pathogens penetrated deep into the tissue and persisted for a few days with P. gingivalis also forming a biofilm on the cell surface. The infection triggered the expression of inflammatory mediators in cells and both pathogens were efficiently eliminated by specific antimicrobials. Conclusions Presented OTG model constitutes a simple and convenient tool to study the interaction between bacterial and viral pathogens within the gingival tissue, including penetration, persistence and biofilm formation. It is also suitable to examine the efficiency of antimicrobial drugs.