Colony-forming Unit Counts Research Articles

Effects of Hydrogen-rich Water on Cariogenic Bacteria.

Some kinds of electrolysed water have been reported to exhibit antioxidant and bactericidal activity. However, studies on the effect of electrolysed hydrogen-rich water (EHW) with a neutral pH on cariogenic bacteria are limited. This study aimed to evaluate the feasibility of using EHW as a mouthwash by examining its various effects on cariogenic bacteria. To test the bactericidal and anti-biofilm formation effects of EHW on Streptococcus mutans and Streptococcus sobrinus, bacterial growth curves, colony-forming unit (CFU) counts, and crystal violet staining of biofilms were examined after exposing the bacterial pellets to EHW or tap water as a control for one minute. In addition, the expressions of glucosyltransferase and glucan-binding proteins encoding genes were examined using real-time PCR. Bacterial growth and biofilm formation were inhibited, and the number of CFUs was significantly reduced in the EHW group compared to the control group. The expression of genes encoding glucosyltransferases (gtfB, gtfC, and gtfI) and glucan-binding proteins (gbpC and dblB) were also decreased in the EHW group compared to the control. Exposing cariogenic bacteria to EHW at neutral pH for one minute can effectively inhibit bacterial growth and biofilm formation in vitro, suggesting that EHW is a promising mouthwash.

Análise antifúngica da incorporação do óleo essencial de Cymbopogon citratus (d.c.) Stapf em polimetacrilato de metila

ABSTRACT Objective: Evaluated the antifungal effect of the incorporation of different concentrations of the essential oil Cymbopogon citratus (capim santo), into polymethylmethacrylate (PMMA) against Candida albicans. Methods: Fifty specimens were fabricated and divided into five groups: Group 1, PMMA + 10% essential oil (n=10); Group 2, PMMA + 15% essential oil (n=10); Group 3, PMMA + 20% essential oil (n=10); Group 4, PMMA + 25% essential oil (n=10); Group 5, PMMA (n=10). PMMA powder was mixed with the monomer and the mixture was placed in disc-shaped cavities measuring 15 mm in diameter, 2 mm thick. To evaluate the antifungal activity of the experimental specimens, the standard strain of Candida albicans was tested. After incubation, the colony count of each plate was performed using a digital colony counter, obtaining the number of colony forming units (CFU) and the Kruskal-Wallis test was applied. Results: There was statistically significant difference in the CFU count of Candida albicans as a consequence of the addition of Cymbopogon citratus essential oil to PMMA (p < 0.001) and values were significantly higher in comparison with those of all the other groups, when the essential oil was incorporated as incorporated into the PMMA in the concentration of 20%. In the other concentrations, no difference in values was observed in comparison with the Control Group without essential oil of Cymbopogon citratus. Conclusion: The acrylic resin with the essential oil incorporated into it in different concentrations provided no effect against development of the genus Candida.

Antimicrobial Photodynamic Therapy as a Technique for Decontamination of Acrylic Resin Devices Provided by Different Dental Laboratories

Introduction: Dentures, occlusal splints, surgical guides and orthodontic appliances are examples of acrylic resin devices made in dental laboratories, which must be disinfected and even sterilized before insertion into the oral cavity. This study evaluated the antimicrobial effect of photodynamic therapy (PDT) applied to acrylic resin specimens received from different laboratories. Methods: Three hundred standardized specimens were ordered from six randomly selected laboratories registered in the Council of Dentistry of Ceará (n=50). The PDT consisted in the association of 22 µM erythrosine, as a photosensitizer (P), and a 520-nm LED at 38 J/cm2 (L). The specimens of each laboratory were randomly distributed into five groups: positive control, sterilized with ethylene oxide; negative control, untreated (P-L-); erythrosine control, only stained (P+L-); LED control, only irradiated (P-L+); PDT (P+L+). Then, the specimens were individually sonicated in saline solution; the suspension was diluted, plated on culture mediums (blood agar, sabouraud dextrose agar and a non-selective chromogenic agar), and incubated for 48 hours at 37°C. Colony-forming-unit (CFU) counts were done and statistical tests of Kruskal-Wallis/Dunn were carried out. Results: The specimens from all laboratories were contaminated with bacteria and yeasts. Staphylococcus aureus, Staphylococcus saprophyticus, Escherichia coli, Enterococcus spp., Klebsiella and Pseudomonas spp. were identified. The PDT significantly reduced CFU counts (P<0.0001), compared to P-L-. Conclusion: PDT was able to effectively decontaminate the acrylic resin specimens provided from dental laboratories.

Evaluation of Biofilm Formation in Clear Aligners with Different Materials and The Effect of Lugol on the Formed Biofilm by Microbiological and SEM Analysis

Bu çalışmanın amacı; günümüzde kullanımı yaygınlaşan şeffaf plaklardaki biyofilm tutulumunu ve %5 Lugol’un hem biyofilm oluşumuna (inhibisyon) hem de oluşmuş biyofilmin eradikasyonuna etkisini araştırmaktır. Üç farklı marka şeffaf plak 5mm’lik yuvarlaklar halinde kesilip (n=10) üzerlerine S. mutans biyofilm oluşumu hem koloni sayma yöntemi hem de SEM ile incelendi. %5 lugolun etkisi MIC olarak tespit edildi. MIC altı değerde inhibisyon ve MIC -2 MIC değerlerde deneyleri yapıldı. Verilerin değerlendirilmesinde Kruskal Wallis varyans analizi (ANOVA) ve tekrarlayan ölçümlerde varyans analizi kullanıldı. Koloni oluşturan birim sayımı (cfu) ve SEM görüntüleri ile elde edilen veriler karşılaştırıldığında her üç plakda da 3. Saatin sonunda biyofilm oluşumu izlenmiş ve 48 saatin sonunda maximuma ulaşmıştır. %5 Lugol her üç plakta da tam inhibisyon sağlamış ancak eradikasyonda etkili olmakla birlikte tam bir uzaklaştırma yapamamıştır. %5 Lugol biyofilm oluşmadan uygulandığında biyofilm oluşumunu şeffaf plak markası farketmeksizin engellemiş, ancak oluşmuş S. mutans biyofilmlerini kısmen uzaklaştırabilmiştir.

DIVERSE STRESS TREATMENTS AND ACINETOBACTER BAUMANNII PERSISTER FORMATION

This study was aimed to investigate diverse stress treatments on Imipenem susceptible A. baumannii for persister cells formation. Isolates were assessed for persister cells presence by colony forming unit counting (CFU/ml) after exposure to Imipenem (10 µg/ml) in presence and absence of acid stress treatment (pH 6) for 0-7 hr. In- vitro activity of curcumin to synergize Imipenem to kill persister cells was investigated. The frequency of hicAB and relEB of type II toxin- antitoxin (TA) loci was evaluated by polymerase chain reaction. The transcription level of hicA and relE toxin genes was assessed by quantitative reverse transcriptase polymerase chain reaction. The level of persistence varied between isolates and across the stress conditions. Imipenem treated isolates gave persister fraction of 0.07% to 0.4% whereas Imipenem + pH6 treated isolates gave 0.3% to 0.7% of the initial untreated population. No significant reduction in persistence level was observed after combination with curcumin (P value &gt; 0.05). The hicAB and relEB loci were detected in 100% and 61.5% of Imipenem susceptible isolates, respectively. Imipenem triggered hicA toxin gene transcription while acid stress triggered hicA and relE toxin genes transcription in persister cells.

Biofilm Formation, Virulence Factors and Antifungal Susceptibility of Candida spp. Isolated From the Oral Cavity of Diabetes Mellitus Patients

Background: Candida species colonize human microbiota and some conditions, such as immunosuppression or chronic illness, predispose the individual to fungal infections; among them, diabetes mellitus, a metabolic disorder frequently associated with higher rates of yeast infections. Material and Methods: The prevalence of Candida species in the oral cavity of patients with diabetes mellitus was evaluated and the carriage was compared between type 1 and type 2 diabetic groups. In addition, in vitro susceptibility to antifungals, biofilm formation, cell surface hydrophobicity, and the production of hydrolytic enzymes were tested. Results: The results demonstrated the presence of different Candida species in the oral cavity of diabetic patients; and, also showed that type 1 diabetic patients are more susceptible to Candida colonization. Almost all isolates produce virulence factors such as proteases, phospholipases, or form biofilm; and they are sensitive to fluconazole and nystatin. Conclusion: Colonization of Candida spp. oral isolates from type 1 and type 2 diabetic patients were similar; however, type 1 presented a higher colony-forming unit counting. Overall, Candida isolates from the oral cavity of diabetic patients are potential pathogens of candidiasis.

Effect of chlorhexidine mouthrinse on prevention of microbial contamination during EBUS-TBNA: a randomized controlled trial

BackgroundAlthough endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) is a minimally invasive procedure, fatal infectious complications have been reported. However, adequate preventive strategies have not been determined. We aimed to investigate the effect of chlorhexidine mouthrinse on the prevention of microbial contamination during EBUS-TBNA.MethodsIn this single-center, assessor-blinded, parallel-group randomized controlled trial, we randomly assigned adult participants undergoing EBUS-TBNA using a convex probe to gargle for 1 minute with 100 mL of 0.12% chlorhexidine gluconate before EBUS-TBNA or to receive usual care (no chlorhexidine mouthrinse). Aspiration needle wash samples were collected immediately after completion of EBUS-TBNA by instilling sterile saline into the used needle. The primary outcome was colony forming unit (CFU) counts per mL of needle wash samples in aerobic cultures. Secondary outcomes were CFU counts per mL of needle wash samples in anaerobic cultures, fever within 24 hours after EBUS-TBNA, and infectious complications within 4 weeks after EBUS-TBNA.ResultsFrom January 2021 to June 2021, 106 patients received either chlorhexidine mouthrinse (n = 51) or usual care (n = 55). The median CFU counts of needle wash samples in aerobic cultures were not significantly different in the two groups (10 CFU/mL vs 20 CFU/mL; P = 0.70). There were no significant differences between the groups regarding secondary outcomes, including median CFU counts in anaerobic cultures (P = 0.41) and fever within 24 hours after EBUS-TBNA (11.8% vs 5.6%, P = 0.31). There were no infectious complications within 4 weeks in both groups.ConclusionsChlorhexidine mouthrinse did not reduce CFU counts in needle wash samples of EBUS-TBNA.Trial registrationClinicalTrials.gov, NCT04718922. Registered on 22/01/2021.

Salivary Streptococcus mutans colony-forming unit count in patients with and without orthodontic appliances.

The aim of this study was to determine the S. mutans colony-forming unit count (CFU/mL) in participants with and without fixed orthodontic appliances. It was an observational cross-sectional study on 21 participants, all over 18 years of age, non-smokers, without removable oral appliances, who had not been under antibiotic treatment within the previous three months. Sociodemographic variables, oral hygiene habits, S. mutans CFU/mL count, and salivary pH were assessed. Saliva samples were collected, and the data was analyzed using Fisher's exact and Kruskal Wallis tests. A p-value <0.05 was considered statistically significant. Fourteen (66.7%) of the participants were female; average age was 20.4 ± 2.2 years. The group without fixed orthodontic appliances had the highest salivary S. mutans CFU/mL count (Me: 56.0×103, IQR: 9.2×103 - 75.5×103), but there was no statistically significant difference between groups (p=0.7459). There was a statistically significant difference in salivary pH, with the metal orthodontic appliance group having the lowest pH (p=0.0478). No statistically significant difference in salivary S. mutans CFU/mL count was found between groups. Salivary pH was lower in the group with metal appliances than in the groups with non-metal appliances and without appliances.

Development and Characterization of Novel Orthodontic Adhesive Containing PCL-Gelatin-AgNPs Fibers.

Enamel demineralization around brackets is a relatively common complication of fixed orthodontic treatment, which seriously affects the aesthetics of teeth. In this study, a novel orthodontic adhesive containing polycaprolactone−gelatin−silver nanoparticles (PCL−gelatin−AgNPs) composite fibers was prepared to prevent enamel demineralization of orthodontic treatment. First, PCL−gelatin−AgNPs fibers film prepared by electrospinning was made into short fibers and added to traditional orthodontic adhesives (Transbond XT, 3M Unitek) in three different ratios to design a series of composite adhesives containing antibacterial materials. The antimicrobial performance of the control product and the three samples were then evaluated by bacterial live/dead staining, colony-forming unit (CFU) counts, tensile bond strength (TBS), and adhesive residue index (ARI) scores. The composite adhesives’ antimicrobial properties increased with the increasing content of PCL−gelatin−AgNPs short fibers. The addition of complex antimicrobial fibers to 3M Transbond XT adhesive can significantly reduce the CFU of bacterial biofilms (p < 0.05). The bacterial survival rate on the surface of the specimen decreased with the increase of PCL−gelatin−AgNPs short fibers (p < 0.05). The TBS and ARI values (n = 10) indicated that adding PCL−gelatin−AgNPs short fibers had no significant adverse effect on adhesion. Therefore, adding PCL−gelatin−AgNPs short fibers makes it possible to fabricate orthodontic adhesives with strong antibacterial properties without compromising the bonding ability, which is essential for preventing enamel demineralization around the brackets.

1201. Reduction of Methicillin-Resistant Staphylococcus aureus Surface Microbial Burden and related Healthcare Associated Infections with the implementation of an Advanced Photocatalytic Oxidation Technology in a Medical-Surgical Intensive Care Unit

Abstract Background Methicillin Resistant Staphylococcus aureus (MRSA) bacteria have been long established as a major cause of infections. MRSA infections occurring in blood or other sterile sites are associated with poorer health outcomes and an increased risk of mortality, especially when acquired in a healthcare setting. The ongoing COVID-19 pandemic has not only stymied reduction efforts but has precipitated increases in invasive HO-MRSA infections, with increases seen of 12-34% in national MRSA standardized infection ratios in 2020 compared to 2019. The facility had seen similar increases of HO-MRSA infections, defined as a positive culture on or after the 3rd hospital day, despite no change in clinical practice. Methods A prospective study was conducted in a 22 bed Medical-Surgical Intensive Care Unit from November 2021 to March 2022. 50 surface samples were collected from surfaces throughout the unit including two nurses’ stations, physician charting area, and 5 areas in 7 patient rooms. The advanced photocatalytic oxidation (aPCO) equipment was then installed in the HVAC ductwork throughout the ICU and activated. Sampling was then repeated every four weeks during the study period. The facility’s normal disinfection protocols were unchanged. HO-MRSA infections attributed to the unit were also tracked during the study period. Changes in MRSA surface burden were calculated using a repeated methods ANOVA with post hoc analyses as appropriate. Rates of HO-MRSA infections per 1,000 patient days were compared using the chi-square test. Results There was a 98% statistically significant decrease in MRSA surface burden from the baseline to final post-activation test. The average colony forming unit count (CFU) went from 427 to 3 CFU/100cm2 during the same time. HO-MRSA infections also had a statistically significant decrease when compared to the same time frame a year prior and the immediate 6 months prior to the study. Figure 1Average MRSA Surface Reduction Conclusion The aPCO technology resulted in a reduction of MRSA on frequently touched surfaces in a high-traffic ICU. Corresponding decreases in HO-MRSA cases were also seen. This study highlights a novel aPCO technology and its efficacy at reducing microbial burden and healthcare-onset MRSA infections despite no change in practice and through the continued COVID-19 pandemic. Disclosures Caitlin Stowe, MPH, CPH, CIC, CPHQ, VA-BC, ActivePure Medical: Employee|PDI Healthcare: Employee.

Effect of teriparatide on drug treatment of tuberculous spondylitis: an experimental study

Tuberculous spondylitis often develops catastrophic bone destruction with uncontrolled inflammation. Because anti-tuberculous drugs do not have a role in bone formation, a combination drug therapy with a bone anabolic agent could help in fracture prevention and promote bone reconstruction. This study aimed to investigate the influence of teriparatide on the effect of anti-tuberculous drugs in tuberculous spondylitis treatment. We used the virulent Mycobacterium tuberculosis (Mtb) H37Rv strain. First, we investigated the interaction between teriparatide and anti-tuberculosis drugs (isoniazid and rifampin) by measuring the minimal inhibitory concentration (MIC) against H37Rv. Second, we evaluated the therapeutic effect of anti-tuberculosis drugs and teriparatide on our previously developed in vitro tuberculous spondylitis model of an Mtb-infected MG-63 osteoblastic cell line using acid-fast bacilli staining and colony-forming unit counts. Selected chemokines (interleukin [IL]-8, interferon γ-induced protein 10 kDa [IP-10], monocyte chemoattractant protein [MCP]-1, and regulated upon activation, normal T cell expressed and presumably secreted [RANTES]) and osteoblast proliferation (alkaline phosphatase [ALP] and alizarin red S [ARS] staining) were measured. Teriparatide did not affect the MIC of isoniazid and rifampin. In the Mtb-infected MG-63 spondylitis model, isoniazid and rifampin treatment significantly reduced Mtb growth, and cotreatment with teriparatide did not change the anti-tuberculosis effect of isoniazid (INH) and rifampin (RFP). IP-10 and RANTES levels were significantly increased by Mtb infection, whereas teriparatide did not affect all chemokine levels as inflammatory markers. ALP and ARS staining indicated that teriparatide promoted osteoblastic function even with Mtb infection. Cotreatment with teriparatide and the anti-tuberculosis drugs activated bone formation (ALP-positive area increased by 705%, P = 0.0031). Teriparatide was effective against Mtb-infected MG63 cells without the anti-tuberculosis drugs (ARS-positive area increased by 326%, P = 0.0037). Teriparatide had no effect on the efficacy of anti-tuberculosis drugs and no adverse effect on the activity of Mtb infection in osteoblasts. Furthermore, regulation of representative osteoblastic inflammatory chemokines was not changed by teriparatide treatment. In the in vitro Mtb-infected MG-63 cell model of tuberculous spondylitis, cotreatment with the anti-tuberculosis drugs and teriparatide increased osteoblastic function.

603. Comparing assays for Clinical Phage Microbiology in biofilm

Abstract Background Non-resolving infections are commonly associated with bacterial biofilm formation. One promising solution is the use of lytic bacterial bacteriophages, as an adjunctive approach with antibiotics to penetrate biofilms and cure infections. A major challenge in the use of phages for therapy is the need for accurate personal matching of the most effective phage-antibiotic combination against the target bactera. This matching, termed “Clinical Phage Microbiology” (CPM) is even more problematic regarding biofilm and there are no standard methods for that. Here we compared several approaches for CPM in biofilms. Methods The efficacy of phages on the biofilm was tested in 8 methods using 5 phages at two concentrations. To this end five Pseudomonas aeruginosa (PA) specific phages were grown for biofilm establishment. Next, we followed the metabolic activity of the cultures using: (1)Agilent Seahorse XF Analyzers., and (2) Symcel’s calScreener calorimeter. In addition, we measured the levels of extracellular DNA in the biofilm supernatant using (3) rtPCR and (4) online by SYBR Green without amplification. The 5th method used GFP PA14 with an allure-red dye that allows the assessment of the adherent bacteria only. Last, we used the commonly used (6) Colony Forming Units (CFU) counts, (7) Crystal Violet (CV), and (8) Live/dead stain as our gold standard controls. Results Out of these methods, the calScreener (Figure 1A-F), and Seahorse were able to differentiate between phages real-time. The calSreener also showed a clear difference between biofilm and planktonic growth curves. The Real-Time PCR and the Live dead stain methods have shown a significant differentiation at the treatment endpoint. The other methods, CFU, CV, Syber Green, and Allura red did not show a significant difference between the phages. It should be noted that phages biofilm ranking did not correlate with the planktonic one. Figure 1 heat flow was measured using calScreener over 21 hours, comparing 5 different phages at a PFU of 108 and 106 on Pseudomonas aeruginosa planktonic (A) or biofilm (B-F). (A) planktonic bacteria and PASA16 phage treated, (B) KLEIN 3 phage, (C) PASA16 phage, (D) PB2F phage, (E) D9 phage, (F) PB2C phage. Figure 2 Comparison, using rtPCR, of 5 different phages at a PFU of 10^8 and 10^6 treated for 24 hours, and then their supernatant was filtered. Primers for housekeeping genes (A) ldhD, (B) rpoD. Conclusion We have compared 8 different methods as assays for phage screening. In this model of PA biofilm and phages, we found that calScreener and Seahorse are superior while CFU, CV, Syber Green, Allura red are not as useful. Disclosures Ran Nir-Paz, MD, BiomX: Advisor/Consultant|Technophage: Advisor/Consultant.

Effectiveness Evaluation of a UV-C-Photoinactivator against Selected ESKAPE-E Pathogens.

Healthcare-associated infections (HAI) worldwide includes infections by ESKAPE-E pathogens. Environmental surfaces and fomites are important components in HAI transmission dynamics, and shoe soles are vectors of HAI. Ultraviolet (UV) disinfection is an effective method to inactivate pathogenic microorganisms. In this study, we investigated whether the SANITECH UV-C shoe sole decontaminator equipment that provides germicidal UV-C radiation could effectively reduce this risk of different pathogens. Six standard strains and four clinical MDR strains in liquid and solid medium were exposed to a UV-C System at specific concentrations at other times. Bacterial inactivation (growth and cultivability) was investigated using colony counts and resazurin as metabolic indicators. SEM was performed to assess the membrane damage. Statistically significant reduction in cell viability for all ATCCs strains occurred after 10 s of exposure to the UV-C system, except for S. enterica, which only occurred at 20 s. The cell viability of P. aeruginosa (90.9%), E. faecalis and A. baumannii (85.3%), S. enterica (82.9%), E. coli (79.2%) and S. aureus (71.9%) was reduced considerably at 20 s. In colony count, after 12 s of UV-C exposure, all ATCC strains showed a 100% reduction in CFU counts, except for A. baumannii, which reduced by 97.7%. A substantial reduction of colonies above 3 log10 was observed at 12 and 20 s in all bacterial strains tested, except for A. baumannii ATCC 19606 (12 s). The exposure of ATCCs bacterial strains to the UV-C system for only 2 s was able to reduce 100% in the colony forming units (CFU) count in all ATCCs strains, S. aureus, P. aeruginosa, E. coli, A. baumannii, E. faecalis, except the S. enterica strain which had a statistically significant reduction of 99.7%. In ATCC strains, there was a substantial decrease in colonies after 4 s (sec) of exposure to the UV-C system, with a reduction ranging from 3.78-4.15 log10 CFU/mL. This reduction was observed in MDR/ESKAPE-E strains within 10 s, showing that UV-C could eliminate above 3.84 log10 CFU/mL. SEM showed a reduction of pili-like appendages after UV-C treatment in all strains except for E. coli (ATCC 25922). The Sanitech UV-C shoe sole decontaminator equipment from Astech Serv. and Fabrication Ltd. (Petrópolis, Brazil), effectively killed in vitro a series of ATCCs and MDR/ESKAPE-E bacteria of sanitary interest, commonly found in the hospital environment.

Effect of gastric acid on the surface roughness and bacterial adhesion of bulk-fill composite resins.

The purpose of this in vitro study was to evaluate the effect of gastric acid on the surface roughness and biofilm formation of bulk-fill composite resins. Twenty-seven samples of each composite resin were obtained: G1: Filtek Z250 XT (Z250), G2: Filtek Bulk Fill (FTK), G3: Tetric N-Ceram Bulk Fill (TTC), and G4: Aura Bulk Fill (AUR). The samples were quantitatively analyzed for surface roughness (Ra) using a roughness tester (n=15) and for biofilm formation (Cn) by the counting of colony-forming units (CFUs/mL) (n=9) in three different moments: after polishing (Ra0 and Cn0), after gastric acid immersion (Ra1 and Cn1), and after gastric acid and simulated tooth brushing (Ra2 and Cn2). Qualitative analysis through surface topography (n=3) was evaluated by scanning electron microscopy (SEM). Ra values were subjected to two-way repeated measures ANOVA, followed by Tukey's test. Cn values were subjected to Kruskal-Wallis analysis, followed by multiple comparisons analysis (α=0.05). Z250 and FTK showed significant increases in surface roughness at Ra1. There were fewer CFUs/mL on TTC and AUR in relation to those of Z250 and FTK for Cn0, Cn1 and Cn2. The SEM images showed that gastric acid increased the formation of cracks, exposure of fillers and micro cavities for all composite resins. After tooth brushing, the topographical changes were more evident but did not influence biofilm formation. The gastric acid promoted both degradation of the surfaces and bacterial adhesion for all composite resins.

The Quinone-Derived Small Molecule M5N32 Is an Effective Anti-Helicobacter pylori Agent Both In Vivo and In Vitro.

Helicobacter pylori has become increasingly resistant to all commonly used clinical antibiotics. Therefore, new anti-H. pylori drugs need to be identified. Recently, quinones were found to inhibit growth of H. pylori with quinone-derived small-molecule compounds identified as having antitumor effects. The minimum inhibitory concentrations of the compounds against H. pylori were measured by agar plate dilution method. The inhibition of biofilm formation by the compounds was assessed by SYTO9-PI double staining. The reactive oxygen species induced by the compounds were detected by DCFH-DA stain. The clearance effects of the compounds for H. pylori in mouse were evaluated by counting colony-forming units and hematoxylin and eosin staining. Our results revealed strong inhibition of M5N32 in vitro against H. pylori in both the planktonic and biofilm-forming states. Resistance to M5N32 was not developed in successive generations of the bacteria. In vivo, the combination of M5N32 and omeprazole showed enhanced effects in comparison to the standard triple therapy. M5N32 was nontoxic to normal tissues. M5N32 is effective in the treatment of H. pylori infections, providing potential development of anti-H. pylori medicines in the treatment of H. pylori infections.

Real-time in-situ electrochemical monitoring of Pseudomonas aeruginosa biofilms grown on air-liquid interface and its antibiotic susceptibility using a novel dual-chamber microfluidic device.

Biofilms are communities of bacterial cells encased in a self-produced polymeric matrix that exhibit high tolerance toward environmental stress. Despite the plethora of research on biofilms, most P. aeruginosa biofilm models are cultured on a solid-liquid interface, and the longitudinal growth characteristics of P. aeruginosa biofilm are unclear. This study demonstrates the real-time and noninvasive monitoring of biofilm growth using a novel dual-chamber microfluidic device integrated with electrochemical detection capabilities to monitor pyocyanin (PYO). The growth of P. aeruginosa biofilms on the air-liquid interface (ALI) was monitored over 48 h, and its antibiotic susceptibility to 6 h exposure of 50, 400, and 1600 µg/ml of ciprofloxacin solutions was analyzed. The biofilm was treated directly on its surface and indirectly from the substratum by delivering the CIP solution to the top or bottom chamber of the microfluidic device. Results showed that P. aeruginosa biofilm developed on ALI produces PYO continuously, with the PYO production rate varying longitudinally and peak production observed between 24and 30 h. In addition, this current study shows that the amount of PYO produced by the ALI biofilm is proportional to its viable cell numbers, which has not been previously demonstrated. Biofilm treated with ciprofloxacin solution above 400 µg/ml showed significant PYO reduction, with biofilms being killed more effectively when treatment was applied to their surfaces. The electrochemical measurement results have been verified with colony-forming unit count results, and the strong correlation between the PYO electrical signal and the viable cell number highlights the usefulness of this approach for fast and low-cost ALI biofilm study and antimicrobial tests.

Efficacy of organo-selenium-incorporated urinary catheter tubing for in vitro growth inhibition of E. coli, K. pneumoniae, P. aeruginosa, and H. influenzae.

Catheter-associated urinary tract infections are of significant medical burden in cost, morbidity, and mortality. Experimental selenium-coated medical devices have demonstrated non-toxic in vitro and in vivo antimicrobial activity. While antimicrobial-coated catheters have shown efficacy in preventing CAUTIs, selenium has not been tested in this context. The purpose of this in vitro study is to evaluate selenium-incorporated urinary catheters for inhibition of uropathogenic bacterial growth and biofilm formation. Urinary catheters incorporated with 1% organo-selenium and standard (uncoated) catheters were incubated in vitro with E. coli, K. pneumoniae, P. aeruginosa, H. influenzae, and combinations of these bacteria. Growth was evaluated by colony-forming unit count and visualized with confocal laser and scanning electron microscopy. Organo-selenium catheter material integrity was also tested by soaking the tubing in phosphate-buffered saline for 12weeks at 37°C. Organo-selenium-incorporated catheters demonstrated total reduction (100%) of in vitro bacterial growth and biofilm formation for E. coli, K. pneumoniae, H. influenzae, and a combination of these species when compared to control. P. aeruginosa growth was inhibited by approximately 4 logs (99.99%). Complete inhibition of E. coli growth was maintained after long-term phosphate-buffered saline soaking. The results demonstrate that organo-selenium was stably incorporated into catheter tubing and inhibited bacterial attachment, growth, and biofilm formation for multiple uropathogenic organisms. Furthermore, long-term soaking of organo-selenium tubing in phosphate-buffered saline did not show any decline in bacterial growth inhibition or biofilm formation. These findings suggest that organo-selenium-incorporated catheters may be advantageous in preventing catheter-associated urinary tract infections and warrant further in vivo and clinical evaluation.

In vitro antibacterial effects of photodynamic therapy against Enterococcus faecalis in root canals of deciduous teeth

ObjectiveThis study aimed at evaluating the in vitro antibacterial efficacy of photodynamic therapy (PDT) on planktonic E. faecalis and its biofilm in the root canal of infected deciduous teeth.MethodsForty root canals of maxillary deciduous anterior teeth were enlarged up to #35 K-file and inoculated with E. faecalis for 21 days. The root canals were randomly assigned into four groups (n = 10): The normal saline group (control), 1% NaClO group, PDT group, and the 1% NaClO + PDT group. Paper point samples were obtained at baseline (S1) and after treatment (S2). The colony-forming units (CFU) were counted, and the bacterial growth rate calculated. From each subgroup, 5 samples were randomly selected after treatment and a scanning laser confocal microscope (CLSM) used to determine the distribution of dead / living bacteria on the biofilm surface of each subgroup. A scanning electron microscope (SEM) was used to observe bacterial morphologies in the root canal walls of the remaining 5 samples in each subgroup. The Kruskal–Wallis test and Dunn test with boferroni adjustment were used to analyze the effect of the different treatment techniques on the E. faecalis in root canals.ResultsCompared to the saline group, PDT significantly reduced bacterial counts in the root canal (p < 0.05). The CFU counts were lowest (p < 0.05) in the 1% NaClO and in 1% NaClO + PDT groups. The rate of bacterial death on the surface of the biofilm in the PDT group was significantly increased after treatment (p < 0.05), and the rate of bacterial death was highest in 1%NaClO group and 1%NaClO + PDT group (p < 0.05).ConclusionPDT has an antibacterial activity against E. faecalis in the root canal of deciduous teeth. Its activity against planktonic E. faecalis is better than the activity on the intact biofilm. The antibacterial activity of PDT on E. faecalis in root canals of deciduous teeth is lower compared to that of 1% NaClO.

The search for a healthy sugar substitute in aid to lower the incidence of Early Childhood Caries: a comparison of sucrose, xylitol, erythritol and stevia

A pursuit to find a healthy alternative to sucrose with less cariogenic potential, which can potentially lower the incidence of Early Childhood Caries (ECC), by means of comparison. Primary tooth enamel blocks (n=32) were randomly divided into four groups and exposed to 5% concentrations of the respective test groups (sucrose, xylitol, erythritol and stevia). All samples were inoculated with S. mutans standard strain (ATCC 25175) at room temperature. Analysis of Colony Forming Units (CFUs), acidity measurements (pH) and Scanning Electron Microscopy (SEM) observations were done after 6, 12, 18 and 24 h and compared. After 6 h, the marginal mean CFU count indicated equal S. mutans growth in all groups. Stevia showed lower CFU counts compared to other groups at 12, 18 and 24 h. The pH levels for all non-fermentable sugar substitutes (NSS) initially decreased but never below the critical pH=5.5 and stabilized from 12 to 18 h. The pH levels of sucrose dropped and remained below pH=5.5 at all time intervals. The SEM analysis of S. mutans supported the CFU results indicating growth in the presence of sucrose and reduction in the presence of the NSS.Compared to sucrose, xylitol, erythritol and stevia have less cariogenic potential with reduced growth of S. mutans and subsequent acidity levels. Stevia had the least cariogenic potential of all the NSS tested, followed by erythritol and then xylitol.

Sterilization Procedures for Titanium Alloy Surfaces Leads to Higher Expression of Biofilm-Related Staphylococcus aureus Genes.

Background: Around 1-2% of all implantation surgeries lead to implant-related infections, incurring costs of $40,000-$160,000 per total hip PJI. The 5-year mortality rate of prosthetic joint infections is up to 21%. To prevent infections during surgery, sterile surgery rooms and procedures have been developed and certified standards have been established. To guarantee the sterility, implants can be acquired already sterile from manufacturers. Some titanium implants can be delivered unsterilized with a manual for sterilization procedure in compliance with ISO 17664. The aim of this study is to evaluate if the most used sterilization methods (steam sterilization in an autoclave and UV light sterilization) of titanium alloys, can influence the biofilm forming capacity of Staphylococcus aureus. In this study, we examined the influence of sterilization methods on the gene expression of biofilm-associated genes and regulators. Methods: We compared gene expression of icaADBC, SarA, SigB, and SodA on titanium CP4 and Ti6Al4V alloys sterilized by UV-light and pressurized saturated steam sterilization. We performed RT-qPCR after RNA extraction of Staphylococcus aureus ATCC 29213. In addition, bacterial cell growth on the sterilized titanium surfaces was examined by colony forming unit counting on agar plates after 24 h of incubation. Results: Colony forming units of S. aureus on titanium CP4 samples showed a higher tendency in colony counts when sterilized with UV light than with pressurized saturated steam (autoclaved). Similarly, colony forming unit counts on Ti6Al4V samples showed tendencies of higher numbers on UV light sterilized samples than on autoclaved samples. Gene expression of icaADBC, SarA and SodA between steamed samples and UV light sterilized samples showed no difference on titanium CP4 samples, whereas SigB showed higher gene expression on titanium CP4 samples when sterilized with UV light than in an autoclave. On autoclaved Ti6Al4V samples, all examined genes showed 4 to 9 times higher fold changes in gene expression than on UV light sterilized samples. Conclusions: This study indicates that steam sterilization of Ti6Al4V can increase biofilm formation of S. aureus on its surface. The significantly increased gene expression of biofilm responsible genes may indicate a modification of titanium surfaces on alloy components. This may promote biofilm formation that can lead to implant-infections in vivo.