Gabriel Alejandro Martínez-Castañón

Anti-biofilm activity of chitosan gels formulated with silver nanoparticles and their cytotoxic effect on human fibroblasts.

The development of multi-species biofilms in chronic wounds is a serious health problem that primarily generates strong resistance mechanisms to antimicrobial therapy. The use of silver nanoparticles (AgNPs) as a broad-spectrum antimicrobial agent has been studied previously. However, their cytotoxic effects limit its use within the medical area. The purpose of this study was to evaluate the anti-biofilm capacity of chitosan gel formulations loaded with AgNPs, using silver sulfadiazine (SSD) as a standard treatment, on strains of clinical isolates, as well as their cytotoxic effect on human primary fibroblasts. Multi-species biofilm of Staphylococcus aureus oxacillin resistant (MRSA) and Pseudomonas aeruginosa obtained from a patient with chronic wound infection were carried out using a standard Drip Flow Reactor (DFR) under conditions that mimic the flow of nutrients in the human skin. Anti-biofilm activity of chitosan gels and SSD showed a log-reduction of 6.0 for MRSA when chitosan gel with AgNPs at a concentration of 100 ppm was used, however it was necessary to increase the concentration of the chitosan gel with AgNPs to 1000 ppm to get a log-reduction of 3.3, while the SSD showed a total reduction of both bacteria in comparison with the negative control. The biocompatibility evaluation on primary fibroblasts showed better results when the chitosan gels with AgNPs were tested even in the high concentration, in contrast with SSD, which killed all the primary fibroblasts. In conclusion, chitosan gel formulations loaded with AgNPs effectively prevent the formation of biofilm and kill bacteria in established biofilm, which suggest that chitosan gels with AgNPs could be used for prevention and treatment of infections in chronic wounds. The statistic significance of the biocompatibility of chitosan gel formulations loaded with AgNPs represents an advance; however further research and development are necessary to translate this technology into therapeutic and preventive strategies.

Anti-biofilm and cytotoxicity activity of impregnated dressings with silver nanoparticles

Infections arising from bacterial adhesion and colonization on chronic wounds are a significant healthcare problem. Silver nanoparticles (AgNPs) impregnated in dressing have attracted a great deal of attention as a potential solution. The goal of the present study was to evaluate the anti-biofilm activities of AgNPs impregnated in commercial dressings against Pseudomonas aeruginosa, bacteria isolated of chronic wounds from a hospital patient. The antimicrobial activity of AgNPs was tested within biofilms generated under slow fluid shear conditions using a standard bioreactor. A 2-log reduction in the number of colony-forming units of P. aeruginosa was recorded in the reactor on exposure to dressing impregnated with 250ppm of AgNPs, diameter 9.3±1.1nm, and also showed compatibility to mammalian cells (human fibroblasts). Our study suggests that the use of dressings with AgNPs may either prevent or reduce microbial growth in the wound environment, and reducing wound bioburden may improve wound-healing outcomes.

Silver nanoparticles with antimicrobial activities against Streptococcus mutans and their cytotoxic effect

Microbial resistance represents a challenge for the scientific community to develop new bioactive compounds. The goal of this research was to evaluate the antimicrobial activity of silver nanoparticles (AgNPs) against a clinical isolate of Streptococcus mutans, antibiofilm activity against mature S. mutans biofilms and the compatibility with human fibroblasts. The antimicrobial activity of AgNPs against the planktonic clinical isolate was size and concentration dependent, with smaller AgNPs having a lower minimum inhibitory concentration. A reduction of 2.3 log in the number of colony-forming units of S. mutans was observed when biofilms grown in a CDC reactor were exposed to 100 ppm of AgNPs of 9.5±1.1 nm. However, AgNPs at high concentrations (>10 ppm) showed a cytotoxic effect upon human dermal fibroblasts. AgNPs effectively inhibited the growth of a planktonic S. mutans clinical isolate and killed established S. mutans biofilms, which suggests that AgNPs could be used for prevention and treatment of dental caries. Further research and development are necessary to translate this technology into therapeutic and preventive strategies.

Peripheral Arterial Disease Associated With Caries and Periodontal Disease

BACKGROUND Peripheral arterial disease (PAD) is an important cardiovascular disorder of the peripheral arteries. Chronic infections, such as periodontitis, may play an important role in the etiology and pathophysiology of PAD and other cardiovascular conditions. Recently, Streptococcus mutans has been found with high frequency in atheromatous plaques. The aim of this study is to evaluate the possible clinical and microbiologic association between PAD and periodontitis and dental caries. METHODS Thirty patients with PAD and 30 control individuals were selected. PAD and its severity were established by the use of the ankle-brachial index (ABI). Clinical attachment loss (AL); probing depth; decayed, missing, and filled teeth (DMFT) index; and C-reactive protein (CRP) levels were evaluated. The presence of bacterial DNA from Streptococcus mutans, Porphyromonas gingivalis, Tannerella forsythia, Prevotella intermedia, Treponema denticola, and Aggregatibacter actinomycetemcomitans was identified by polymerase chain reaction in subgingival biofilm and serum. RESULTS Patients with ≥30% AL ≥ 4 mm had six-fold increased risk of having PAD (odds ratio = 8.18; 95% confidence interval = 1.21 to 35.23; P = 0.031). There was statistical difference in the CRP (P = 0.0413) and DMFT index (P = 0.0002), with elevated number of missing teeth (P = 0.0459) in the PAD group compared with the control group. There were no significant differences in the frequency of bacteria in serum and subgingival plaque. CONCLUSION There was a positive relationship between periodontitis based on AL and PAD determined by the ABI (odds ratio = 8.18).

Enamel roughness and depth profile after phosphoric acid etching of healthy and fluorotic enamel.

BACKGROUND Dental fluorosis requires aesthetic treatment to improve appearance and etching of enamel surfaces with phosphoric acid is a key step for adhesive restorations. The aim of this study was to evaluate surface roughness and a depth profile in healthy and fluorotic enamel before and after phosphoric acid etching at 15, 30 and 60 seconds. METHODS One hundred and sixty enamel samples from third molars with no fluorosis to severe fluorosis were evaluated by atomic force microscopy. RESULTS Healthy enamel showed a statistically significant difference (p < 0.05) between mean surface roughness at 15 seconds (180.3 nm), 30 seconds (260.9 nm) and 60 seconds (346.5 nm); depth profiles revealed a significant difference for the 60 second treatment (4240.2 nm). For mild fluorosis, there was a statistically significant difference (p < 0.05) between mean surface roughness for 30 second (307.8 nm) and 60 second (346.6 nm) treatments; differences in depth profiles were statistically significant at 15 seconds (2546.7 nm), 30 seconds (3884.2 nm) and 60 seconds (3612.1 nm). For moderate fluorosis, a statistically significant difference (p < 0.05) was observed for surface roughness for 30 second (324.5 nm) and 60 second (396.6 nm) treatments. CONCLUSIONS Surface roughness and depth profile analyses revealed that the best etching results were obtained at 15 seconds for the no fluorosis and mild fluorosis groups, and at 30 seconds for the moderate fluorosis group. Increasing the etching time for severe fluorosis decreased surface roughness and the depth profile, which suggests less micromechanical enamel retention for adhesive bonding applications.

Antibacterial and antibiofilm activities of the photothermal therapy using gold nanorods against seven different bacterial strains

The objective of this work was to determine the bactericidal and antibiofilm activities of gold nanorods (AuNRs) using plasmonic photothermal therapy (PPTT) against oral microorganisms. AuNRs were synthesized by the seed and growth solution method and the gold nanoclusters were characterized with a size of 33.2 nm ± 2.23 length and 7.33 nm ± 1.60 width. The efficacy of PPTT related to its temperature was done reaching 67°C. Minimum inhibitory concentration (MIC) and minimum bactericide concentration (MBC) of AuNRs and AuNRs PPTT were determined against Enterococcus faecalis, Staphylococcus aureus, Streptococcus mutans, Streptococcus sobrinus, Streptococcus oralis, Streptococcus salivarius, and Escherichia coli growth. The antibiofilm activity of AuNRs was explored by fluorescence microscopy. After experimental analyses, AuNRs PPTT shows better results in MICs and MBCs, when it was compared with AuNRs alone. The laser employed to activate the AuNRs had no antibacterial effect against oral microbes. The MICs and MBCs values were higher for S. aureus and E. coli and lower against S. oralis. Surprisingly, the AuNRs alone presented a high antibiofilm activity, inhibiting the biofilm formation of S. mutans. Altogether, these results strongly suggest that AuNRs could be an interesting option to control oral biofilms.

Characterization and biocompatibility of chitosan gels with silver and gold nanoparticles

The presence of bacterial resistance to antibiotics is a very important issue and the search of new alternatives is necessary. In this work, a combination of chitosan gel with silver or gold nanoparticles was prepared and characterized using thermal, rheology, bactericide, and biocompatibility analyses. ESEM images were also taken to visualize the incorporation of the nanoparticles into the gel matrix. Thermal analysis showed a better thermal stability in the chitosan-gold nanoparticles gels compared to the chitosan-silver nanoparticles gels. Rheology analyses showed that the viscosity of the gels decreased when velocity increased and there were differences in viscosity when silver and gold nanoparticles concentrations change. ESEM images showed the presence of agglomerates of silver and gold nanoparticles into the gel matrix with a good distribution; in some cases the formation of microstructures was found. Bactericide results show that the sematerials present an antibacterial activity against S. aureus, S. mutans, and E. coli. The biocompatibility test showed neither negative reaction nor wound healing delay after the application of the gels in an in vivo test. The gels with silver and gold nanoparticles could be used to treat wound infections in oral or skin applications.

Molecular identification and antibiotic resistant bacteria isolated from primary dentition infections

BACKGROUND Bacterial resistance to antibiotics is a health problem in many parts of the world. The aim of this study was to identify bacteria from dental infections and determine bacterial resistance to antibiotics used in dental care in the primary dentition. METHODS This cross-sectional study comprised 60 children who presented for dental treatment for active dental infections in the primary dentition. Samples from dental infections were collected and bacteria were identified by polymerase chain reaction (PCR) assay. Bacterial resistance to antibiotics was determined by colony forming units on agar plates containing amoxicillin, clindamycin and amoxillicin-clavulanic acid (A-CA) tested at 8 μg/ml or 16 μg/ml. RESULTS Clindamycin in both concentrations tested (8 μg/ml and 16 μg/ml) showed the highest bacterial resistance (85.9%), followed by amoxicillin (43.7%) and A-CA (12.0%). All comparisons among the three antibiotics used in the study exhibited statistical significance (p = <0.05) in both concentrations tested (8 μg/ml and 16 μg/ml), and under aerobic and anaerobic conditions. The most prevalent resistant species identified by PCR in primary dentition infections were: Streptococcus oralis and Prevotella intermedia (75.0%); Treponema denticola and Porphyromonas gingivalis (48.3%); Streptococcus mutans (45.0%); Campylobacter rectus; and Streptococcus salivarius (40%). CONCLUSIONS This study demonstrated that A-CA exhibited the lowest bacterial resistance for clinical isolates in primary dentition infections.

Evaluation of vascular tone and cardiac contractility in response to silver nanoparticles, using Langendorff rat heart preparation

Silver nanoparticles (AgNPs) have been widely used because of their antimicrobial properties. However, several reports suggest that AgNPs exposure promote cardiac effects that involve nitric oxide (NO) and oxidative stress (OS). Nevertheless, there are no studies related to AgNPs-induced effects in cardiac physiology. The aim of this study was to evaluate the AgNPs direct actions on coronary vascular tone and cardiac contractility using Langendorff rat heart preparation. Low concentrations of AgNPs (0.1 and 1 μg/mL) increased NO derived from inducible NO-synthase (iNOS), without modifying cardiac parameters. Meanwhile, high concentrations (10 and 100 μg/mL) promoted a sustained vasoconstriction and increased cardiac contractility related to OS, leading to rhabdomyolysis. Furthermore, AgNPs were internalized in the cardiac muscle, hindering classic actions induced by phenylephrine (Phe) and acetylcholine (ACh). These data suggest that AgNPs affect cardiac physiology in function of the concentration and in part of the NO generation, NOS expression and OS.

Bovine serum albumin and chitosan coated silver nanoparticles and its antimicrobial activity against oral and nonoral bacteria

Antimicrobial agents have been developed for drug-resistance infections, which have been rapidly increasing; however, the control of involved microorganisms is still a challenge. In this work, SNP with bovine serum albumin (BSA) and chitosan (CS) coatings were prepared with an aqueous reduction method, characterized using dispersion light scattering, transmission electron microscopy, and thermal analysis. Antibacterial activity was tested on seven oral and nonoral bacteria by microdilution test and scanning electron microscopy. Six different sizes and shapes of coated SNP were prepared and used. Characterization revealed narrow size and good distribution of particles, spherical and pseudospherical shapes, and the presence of coatings on the SNP surfaces. All samples showed antimicrobial activity, although smaller sizes and CS samples had the best inhibition effects. The highest microbial resistance was shown by Gram-positive bacteria. Although coated SNP action depends on particular bacterium, BSA and CS coated SNP could be used for drug-resistance infections.