Spatio-Temporal Tracking and Phylodynamics of an Urban Dengue 3 Outbreak in Sao Paulo, Brazil

The dengue virus has a single-stranded positive-sense RNA genome of similar to 10.700 nucleotides with a single open reading frame that encodes three structural (C, prM, and E) and seven nonstructural (NS1, NS2A, NS2B, NS3, NS4A, NS4B, and NS5) proteins. It possesses four antigenically distinct serotypes (DENV 1-4). Many phylogenetic studies address particularities of the different serotypes using convenience samples that are not conducive to a spatio-temporal analysis in a single urban setting. We describe the pattern of spread of distinct lineages of DENV-3 circulating in Sao Jose do Rio Preto, Brazil, during 2006. Blood samples from patients presenting dengue-like symptoms were collected for DENV testing. We performed M-N-PCR using primers based on NS5 for virus detection and identification. The fragments were purified from PCR mixtures and sequenced. The positive dengue cases were geo-coded. To type the sequenced samples, 52 reference sequences were aligned. The dataset generated was used for iterative phylogenetic reconstruction with the maximum likelihood criterion. The best demographic model, the rate of growth, rate of evolutionary change, and Time to Most Recent Common Ancestor (TMRCA) were estimated. The basic reproductive rate during the epidemics was estimated. We obtained sequences from 82 patients among 174 blood samples. We were able to geo-code 46 sequences. The alignment generated a 399-nucleotide-long dataset with 134 taxa. The phylogenetic analysis indicated that all samples were of DENV-3 and related to strains circulating on the isle of Martinique in 2000-2001. Sixty DENV-3 from Sao Jose do Rio Preto formed a monophyletic group (lineage 1), closely related to the remaining 22 isolates (lineage 2). We assumed that these lineages appeared before 2006 in different occasions. By transforming the inferred exponential growth rates into the basic reproductive rate, we obtained values for lineage 1 of R(0) = 1.53 and values for lineage 2 of R(0) = 1.13. Under the exponential model, TMRCA of lineage 1 dated 1 year and lineage 2 dated 3.4 years before the last sampling. The possibility of inferring the spatio-temporal dynamics from genetic data has been generally little explored, and it may shed light on DENV circulation. The use of both geographic and temporally structured phylogenetic data provided a detailed view on the spread of at least two dengue viral strains in a populated urban area.

Disparity between Multilocus Enzyme Electrophoresis, Microsatellite Markers and Pulsed-Field Gel Electrophoresis in epidemiological tracking of Candida albicans

Various molecular systems are available for epidemiological, genetic, evolutionary, taxonomic and systematic studies of innumerable fungal infections, especially those caused by the opportunistic pathogen C. albicans. A total of 75 independent oral isolates were selected in order to compare Multilocus Enzyme Electrophoresis (MLEE), Electrophoretic Karyotyping (EK) and Microsatellite Markers (Simple Sequence Repeats - SSRs), in their abilities to differentiate and group C. albicans isolates (discriminatory power), and also, to evaluate the concordance and similarity of the groups of strains determined by cluster analysis for each fingerprinting method. Isoenzyme typing was performed using eleven enzyme systems: Adh, Sdh, M1p, Mdh, Idh, Gdh, G6pdh, Asd, Cat, Po, and Lap (data previously published). The EK method consisted of chromosomal DNA separation by pulsed-field gel electrophoresis using a CHEF system. The microsatellite markers were investigated by PCR using three polymorphic loci: EF3, CDC3, and HIS3. Dendrograms were generated by the SAHN method and UPGMA algorithm based on similarity matrices (S(SM)). The discriminatory power of the three methods was over 95%, however a paired analysis among them showed a parity of 19.7-22.4% in the identification of strains. Weak correlation was also observed among the genetic similarity matrices (S(SM)(MLEE) x S(SM)(EK) x S(SM)(SSRs)). Clustering analyses showed a mean of 9 +/- 12.4 isolates per cluster (3.8 +/- 8 isolates/taxon) for MLEE, 6.2 +/- 4.9 isolates per cluster (4 +/- 4.5 isolates/taxon) for SSRs, and 4.1 +/- 2.3 isolates per cluster (2.6 +/- 2.3 isolates/taxon) for EK. A total of 45 (13%), 39(11.2%), 5 (1.4%) and 3 (0.9%) clusters pairs from 347 showed similarity (Si) of 0.1-10%, 10.1-20%, 20.1-30% and 30.1-40%, respectively. Clinical and molecular epidemiological correlation involving the opportunistic pathogen C. albicans may be attributed dependently of each method of genotyping (i.e., MLEE, EK, and SSRs) supplemented with similarity and grouping analysis. Therefore, the use of genotyping systems that give results which offer minimum disparity, or the combination of the results of these systems, can provide greater security and consistency in the determination of strains and their genetic relationships. (C) 2010 Elsevier B.V. All rights reserved.

Transient and Steady-State Analysis of the Affine Combination of Two Adaptive Filters

In this paper, we propose an approach to the transient and steady-state analysis of the affine combination of one fast and one slow adaptive filters. The theoretical models are based on expressions for the excess mean-square error (EMSE) and cross-EMSE of the component filters, which allows their application to different combinations of algorithms, such as least mean-squares (LMS), normalized LMS (NLMS), and constant modulus algorithm (CMA), considering white or colored inputs and stationary or nonstationary environments. Since the desired universal behavior of the combination depends on the correct estimation of the mixing parameter at every instant, its adaptation is also taken into account in the transient analysis. Furthermore, we propose normalized algorithms for the adaptation of the mixing parameter that exhibit good performance. Good agreement between analysis and simulation results is always observed.

Stochastic stability analysis for the constant-modulus algorithm

We derive an easy-to-compute approximate bound for the range of step-sizes for which the constant-modulus algorithm (CMA) will remain stable if initialized close to a minimum of the CM cost function. Our model highlights the influence, of the signal constellation used in the transmission system: for smaller variation in the modulus of the transmitted symbols, the algorithm will be more robust, and the steady-state misadjustment will be smaller. The theoretical results are validated through several simulations, for long and short filters and channels.

Quantitative ferromagnetic resonance analysis of CD133 stem cells labeled with iron oxide nanoparticles

The aim of this work is to provide a quantitative method for analysis of the concentration of superparamagnetic iron oxide nanoparticles (SPION), determined by means of ferromagnetic resonance (FMR), with the nanoparticles coupled to a specific antibody (AC133), and thus to express the antigenic labeling evidence for the stem cells C D133(+). The FMR efficiency and sensitivity were proven adequate for detecting and quantifying the low amounts of iron content in the C D133(+) cells (similar to 6.16 x 10(5) pg in the volume of 2 mu l containing 4.5 x 1011 SPION). The quantitative method led to the result of 1.70 x 10(-13) mol of Fe (9.5 pg), or 7.0 x 10(6) nanoparticles per cell. For the quantification analysis via the FMR technique it was necessary to carry out a preliminary quantitative visualization of iron oxide-labeled cells in order to ensure that the nanoparticles coupled to the antibodies are indeed tied to the antigen at the stem cell surface and that the cellular morphology was conserved, as proof of the validity of this method. The quantitative analysis by means of FMR is necessary for determining the signal intensity for the study of molecular imaging by means of magnetic resonance imaging (MRI).

Digital subtraction radiographic analysis of the combination of bioabsorbable membrane and bovine morphogenetic protein pool in human periodontal infrabony defects

OBJECTIVES: This study assessed the bone density gain and its relationship with the periodontal clinical parameters in a case series of a regenerative therapy procedure. MATERIAL AND METHODS: Using a split-mouth study design, 10 pairs of infrabony defects from 15 patients were treated with a pool of bovine bone morphogenetic proteins associated with collagen membrane (test sites) or collagen membrane only (control sites). The periodontal healing was clinically and radiographically monitored for six months. Standardized pre-surgical and 6-month postoperative radiographs were digitized for digital subtraction analysis, which showed relative bone density gain in both groups of 0.034 ± 0.423 and 0.105 ± 0.423 in the test and control group, respectively (p>0.05). RESULTS: As regards the area size of bone density change, the influence of the therapy was detected in 2.5 mm² in the test group and 2 mm² in the control group (p>0.05). Additionally, no correlation was observed between the favorable clinical results and the bone density gain measured by digital subtraction radiography (p>0.05). CONCLUSIONS: The findings of this study suggest that the clinical benefit of the regenerative therapy observed did not come with significant bone density gains. Long-term evaluation may lead to a different conclusions.

FiltekTM Silorane and FiltekTM Supreme XT resins: tissue reaction after subcutaneous implantation in isogenic mice

The aim of this study was to evaluate the tissue compatibility of a silorane-based resin system (FiltekTM Silorane) and a methacrylate-based nanoparticle resin (FiltekTM Supreme XT) after implantation in the subcutaneous connective tissue of isogenic mice. One hundred and thirty five male isogenic BALB/c mice were randomly assigned to 12 experimental and 3 control groups, according to the implanted material and the experimental period of 7, 21 and 63 days. At the end of each period, the animals were killed and the tubes with the surrounding tissues were removed and processed for microscopic analysis. Samples were subjected to a descriptive and a semi-quantitative analyses using a 4-point scoring system (0-3) to evaluate the collagen fiber formation and inflammatory infiltrate. Data were statistically analyzed using the Kruskal Wallis test (?=0.05). The results showed that there was no significant difference between the experimental and control groups considering the three evaluation periods (p>0.05). The silorane-based and the methacrylate-based nanoparticle resins presented similar tissue response to that of the empty tube (control group) after subcutaneous implantation in isogenic mice.

Isolation of Pseudomonas aeruginosa strains from dental office environments and units in Barretos, state of São Paulo, Brazil, and analysis of their susceptibility to antimicrobial drugs

A wide variety of opportunistic pathogens has been detected in the tubing supplying water to odontological equipment, in special in the biofilm lining of these tubes. Among these pathogens, Pseudomonas aeruginosa, one of the leading causes of nosocomial infections, is frequently found in water lines supplying dental units. In the present work, 160 samples of water, and 200 fomite samples from forty dental units were collected in the city of Barretos, State of São Paulo, Brazil and evaluated between January and July, 2005. Seventy-six P. aeruginosa strains, isolated from the dental environment (5 strains) and water system (71 strains), were tested for susceptibility to six antimicrobial drugs most frequently used against P. aeruginosa infections. Susceptibility to ciprofloxacin, followed by meropenem was the predominant profile. The need for effective means of reducing the microbial burden within dental unit water lines is emphasized, and the risk of exposure and cross-infection in dental practice, in special when caused by opportunistic pathogens like P. aeruginosa, are highlighted.

Analysis of TMJ vibration sounds before and after use of two types of occlusal splints

Temporomandibular joint (TMJ) sounds are important and common physical signs of temporomandibular disorders (TMD). The aim of this study was to evaluate the influence of the effect of the use of occlusal bite splints (stabilizing and repositioning) on the sounds produced in the TMJ, by means of the electrovibratography (EVG). Thirty-one patients with TMD from the Dental School of Ribeirão Preto, University of São Paulo, Brazil were selected for this study. Group 1 (n=23) wore stabilizing bite splints and Group 2 (n=8) used anterior repositioning splints. Before and after treatment with occlusal splints both groups were analyzed using the SonoPAK Q/S recording system (BioResearch System, Inc.). The treatments with stabilizing bite splints were satisfactory, since they reduced the total amount of the sound energies (p<0.05), but the use of anterior repositioning splints for no more than 4 weeks produced significantly better results (p<0.01). The total amount of vibration energy involved in the vibrating movements of the TMJ showed significant improvement using anterior repositioning splints.

Adhesion of an endodontic sealer to dentin and gutta-percha: shear and push-out bond strength measurements and SEM analysis

The use of an adequate method for evaluation of the adhesion of root canal filling materials provides more reliable results to allow comparison of the materials and substantiate their clinical choice. The aims of this study were to compare the shear bond strength (SBS) test and push-out test for evaluation of the adhesion of an epoxy-based endodontic sealer (AH Plus) to dentin and gutta-percha, and to assess the failure modes on the debonded surfaces by means of scanning electron microscopy (SEM). Three groups were established (n=7): in group 1, root cylinders obtained from human canines were embedded in acrylic resin and had their canals prepared and filled with sealer; in group 2, longitudinal sections of dentin cylinders were embedded in resin with the canal surface smoothed and turned upwards; in group 3, gutta-percha cylinders were embedded in resin. Polyethylene tubes filled with sealer were positioned on the polished surface of the specimens (groups 2 and 3). The push-out test (group 1) and the SBS test (groups 2 and 3) were performed in an Instron universal testing machine running at crosshead speed of 1 mm/min. Means (±SD) in MPa were: G1 (8.8±1.13), G2 (5.9±1.05) and G3 (3.8±0.55). Statistical analysis by ANOVA and Student's t-test (a=0.05) revealed statistically significant differences (p<0.01) among the groups. SEM analysis showed a predominance of adhesive and mixed failures of AH Plus sealer. The tested surface affected significantly the results with the sealer reaching higher bond strength to dentin than to gutta-percha with the SBS test. The comparison of the employed methodologies showed that the SBS test produced significantly lower bond strength values than the push-out test, was skilful in determining the adhesion of AH Plus sealer to dentin and gutta-percha, and required specimens that could be easily prepared for SEM, presenting as a viable alternative for further experiments.

Morphological analysis of glass, carbon and glass/carbon fiber posts and bonding to self or dual-cured resin luting agents

OBJECTIVE: The aim of this study was to evaluate the morphology of glass (GF), carbon (CF) and glass/carbon (G/CF) fiber posts and their bond strength to self or dual-cured resin luting agents. MATERIAL AND METHODS: Morphological analysis of each post type was conducted under scanning electron microscopy (SEM). Bond strength was evaluated by microtensile test after bisecting the posts and re-bonding the two halves with the luting agents. Data were subjected to two-way ANOVA and Tukey's test (α=0.05). Failure modes were evaluated under optical microscopy and SEM. RESULTS: GF presented wider fibers and higher amount of matrix than CF, and G/CF presented carbon fibers surrounded by glass fibers, and both involved by matrix. For CF and GF, the dual-cured material presented significantly higher (p<0.05) bond strength than the self-cured agent. For the dual agent, CF presented similar bond strength to GF (p>0.05), but higher than that of G/CF (p<0.05). For the self-cured agent, no significant differences (p>0.05) were detected, irrespective of the post type. For GF and G/CF, all failures were considered mixed, while a predominance of adhesive failures was detected for CF. CONCLUSION: The bonding between fiber posts and luting agents was affected by the type of fibers and polymerization mode of the cement. When no surface treatment of the post is performed, the bonding between glass fiber post and dual-cured agent seems to be more reliable.

Influence of cervical preflaring on determinationof apical file size in mandibular molars: SEM analysis

This study investigated the influence of cervical preflaring with different rotary instruments on determination of the initial apical file (IAF) in mesiobuccal roots of mandibular molars. Fifty human mandibular molars whose mesial roots presented two clearly separated apical foramens (mesiobuccal and mesiolingual) were used. After standard access opening and removal of pulp tissue, the working length (WL) was determined at 1 mm short of the root apex. Five groups (n=10) were formed at random, according to the type of instrument used for cervical preflaring. In group 1, the size of the IAF was determined without preflaring of the cervical and middle root canal thirds. In groups 2 to 5, preflaring was performed with Gates-Glidden drills, ProTaper instruments, EndoFlare instruments and LA Axxes burs, respectively. Canals were sized manually with K-files, starting with size 08 K-files, inserted passively up to the WL. File sizes were increased until a binding sensation was felt at the WL and the size of the file was recorded. The instrument corresponding to the IAF was fixed into the canal at the WL with methylcyanoacrylate. The teeth were then sectioned transversally 1 mm short of the apex, with the IAF in position. Cross-sections of the WL region were examined under scanning electron microscopy and the discrepancies between canal diameter and the diameter of IAF were calculated using the tool "rule" (FEG) of the microscope's proprietary software. The measurements (µm) were analyzed statistically by Kruskal-Wallis and Dunn's tests at 5% significance level. There were statistically significant differences among the groups (p<0.05). The non-flared group had the greatest discrepancy (125.30 ± 51.54) and differed significantly from all flared groups (p<0.05). Cervical preflaring with LA Axxess burs produced the least discrepancies (55.10 ± 48.31), followed by EndoFlare instruments (68.20 ± 42.44), Gattes Glidden drills (68.90 ± 42.46) and ProTaper files (77.40 ± 73.19). However, no significant differences (p>0.05) were found among the rotary instruments. In conclusion, cervical preflaring improved IAF fitting to the canals at the WL in mesiobuccal roots of maxillary first molars. The rotary instruments evaluated in this study did not differ from each other regarding the discrepancies produced between the IAF size and canal diameter at the WL.

Ex vivo analysis of root canal cleaning using Endo-PTC associated to NaOCl and different irrigant solutions

The aim of this study was to assess qualitatively, by means of SEM images, the cleaning of the dentin walls of root canals after chemical-surgical preparation using Endo-PTC cream with 0.5% and 1% sodium hypochlorite and different final irrigating solutions. Seventy-two single-rooted human teeth were divided into eight groups and prepared using Endo-PTC cream with sodium hypochlorite (NaOCl) at different concentrations, and irrigated with NaOCl at different concentrations. Final irrigation was performed with either EDTA-T or EDTA-C. The best results were obtained with Group 1, followed by Groups 5, 2, 7, 8, 3, 6 and 4. We can conclude that the use of 0.5% NaOCl during instrumentation and final flush of the root canals was more efficient in cleaning than was 1% sodium hypochlorite. EDTA-T was more efficient in removing smear layer than EDTA-C, and the cervical third presented better cleaning of the root canal walls than did the middle third, which showed cleaner dentin walls than the apical third.

Coronal and apical leakage analysis of two different root canal obturation systems

This study compared the coronal and apical leakage of AH Plus with gutta-percha to that of Epiphany with Resilon. Twenty-four single rooted teeth were instrumented and divided into 2 groups according to the solutions for smear layer removal and the obturation materials employed: Group A - 17% EDTA-T and AH Plus with gutta-percha; Group B - primer and Epiphany with Resilon. The Group B specimens were light-cured in the coronal area for 20 s. The external root surfaces were covered with a double layer of ethyl cyanoacrylate, except for the apical foramen and the cavity access. The teeth were immersed in 0.5% methylene blue for 48 h. The specimens were rinsed, dried and axially split for dye penetration measurement with the ImageLab 2.3 software. The t-test showed no significant differences for coronal leakage between the groups, but there were significant differences for apical leakage between the groups (P < 0.05). AH Plus with gutta-percha and Epiphany with Resilon provided the same coronal seal, whereas Epiphany with Resilon provided the best apical seal.

Analysis of contamination of endodontic absorbent paper points

PURPOSE: The aim of this study was to assess the contamination status of endodontic absorbent paper points from sterilized or not sterilized commercial packs, as well as paper points exposed to the dental office environment. METHODS: Twenty absorbent paper points were evaluated for contamination status packed under different conditions: commercial/sterilized pack, commercial/non-sterilized pack, exposed to the clinical environment, and intentionally contaminated (positive control). Contamination was determined qualitatively and quantitatively by aerobiosis, capnophilic growth, and pour plate. The Petri dishes were analyzed with a colony counter, and the results were expressed as colony-forming units. The data were analyzed by Kruskal-Wallis test (α=0.05). RESULTS: No difference in colony-forming units was found among the groups of endodontic absorbent paper points. All groups were contaminated by fungi and bacteria. CONCLUSION: It can be concluded that the sterilization of absorbent endodontic paper points before clinical use should be recommended regardless of commercial presentation