Machine-learning from Pseudomonas putida KT2440 transcriptomes reveals its transcriptional regulatory network.

Bacterial gene expression is orchestrated by numerous transcription factors (TFs). Elucidating how gene expression is regulated is fundamental to understanding bacterial physiology and engineering it for practical use. In this study, a machine-learning approach was applied to uncover the genome-scale transcriptional regulatory network (TRN) in Pseudomonas putida KT2440, an important organism for bioproduction. We performed independent component analysis of a compendium of 321 high-quality gene expression profiles, which were previously published or newly generated in this study. We identified 84 groups of independently modulated genes (iModulons) that explain 75.7% of the total variance in the compendium. With these iModulons, we (i) expand our understanding of the regulatory functions of 39 iModulon associated TFs (e.g., HexR, Zur) by systematic comparison with 1993 previously reported TF-gene interactions; (ii) outline transcriptional changes after the transition from the exponential growth to stationary phases; (iii) capture group of genes required for utilizing diverse carbon sources and increased stationary response with slower growth rates; (iv) unveil multiple evolutionary strategies of transcriptome reallocation to achieve fast growth rates; and (v) define an osmotic stimulon, which includes the Type VI secretion system, as coordination of multiple iModulon activity changes. Taken together, this study provides the first quantitative genome-scale TRN for P. putida KT2440 and a basis for a comprehensive understanding of its complex transcriptome changes in a variety of physiological states.

Interdisciplinary Management of a Patient with Dentinogenesis Imperfecta Type II Using a Combination of CAD-CAM and Analog Techniques: A Clinical Report.

Type II dentinogenesis imperfecta is an autosomal dominant condition that affects dentin which increases the complexity of the predictability of restorative treatment. Computer-aided design and computer-aided manufacturing (CAD-CAM) technologies permit the creation of highly accurate devices and dental prostheses that simplify the planning and execution of advanced implant surgery and full-mouth rehabilitation. This clinical report presents the interdisciplinary management of a 20-year-old male with dentinogenesis imperfecta type II. In this article, a combination of analog and CAD-CAM technologies were used to fabricate devices that aided planning, assisted intermaxillary fixation and implant placement, served as interim prostheses, and permitted the accurate establishment of esthetics and occlusion of the definitive full-arch prostheses.

The lichen symbiosis re-viewed through the genomes of Cladonia grayi and its algal partner Asterochloris glomerata.

BACKGROUND: Lichens, encompassing 20,000 known species, are symbioses between specialized fungi (mycobionts), mostly ascomycetes, and unicellular green algae or cyanobacteria (photobionts). Here we describe the first parallel genomic analysis of the mycobiont Cladonia grayi and of its green algal photobiont Asterochloris glomerata. We focus on genes/predicted proteins of potential symbiotic significance, sought by surveying proteins differentially activated during early stages of mycobiont and photobiont interaction in coculture, expanded or contracted protein families, and proteins with differential rates of evolution. RESULTS: A) In coculture, the fungus upregulated small secreted proteins, membrane transport proteins, signal transduction components, extracellular hydrolases and, notably, a ribitol transporter and an ammonium transporter, and the alga activated DNA metabolism, signal transduction, and expression of flagellar components. B) Expanded fungal protein families include heterokaryon incompatibility proteins, polyketide synthases, and a unique set of G-protein α subunit paralogs. Expanded algal protein families include carbohydrate active enzymes and a specific subclass of cytoplasmic carbonic anhydrases. The alga also appears to have acquired by horizontal gene transfer from prokaryotes novel archaeal ATPases and Desiccation-Related Proteins. Expanded in both symbionts are signal transduction components, ankyrin domain proteins and transcription factors involved in chromatin remodeling and stress responses. The fungal transportome is contracted, as are algal nitrate assimilation genes. C) In the mycobiont, slow-evolving proteins were enriched for components involved in protein translation, translocation and sorting. CONCLUSIONS: The surveyed genes affect stress resistance, signaling, genome reprogramming, nutritional and structural interactions. The alga carries many genes likely transferred horizontally through viruses, yet we found no evidence of inter-symbiont gene transfer. The presence in the photobiont of meiosis-specific genes supports the notion that sexual reproduction occurs in Asterochloris while they are free-living, a phenomenon with implications for the adaptability of lichens and the persistent autonomy of the symbionts. The diversity of the genes affecting the symbiosis suggests that lichens evolved by accretion of many scattered regulatory and structural changes rather than through introduction of a few key innovations. This predicts that paths to lichenization were variable in different phyla, which is consistent with the emerging consensus that ascolichens could have had a few independent origins.

Pseudomonas fluorescens increases mycorrhization and modulates expression of antifungal defense response genes in roots of aspen seedlings.

BACKGROUND: Plants, fungi, and bacteria form complex, mutually-beneficial communities within the soil environment. In return for photosynthetically derived sugars in the form of exudates from plant roots, the microbial symbionts in these rhizosphere communities provide their host plants access to otherwise inaccessible nutrients in soils and help defend the plant against biotic and abiotic stresses. One role that bacteria may play in these communities is that of Mycorrhizal Helper Bacteria (MHB). MHB are bacteria that facilitate the interactions between plant roots and symbiotic mycorrhizal fungi and, while the effects of MHB on the formation of plant-fungal symbiosis and on plant health have been well documented, the specific molecular mechanisms by which MHB drive gene regulation in plant roots leading to these benefits remain largely uncharacterized. RESULTS: Here, we investigate the effects of the bacterium Pseudomonas fluorescens SBW25 (SBW25) on aspen root transcriptome using a tripartite laboratory community comprised of Populus tremuloides (aspen) seedlings and the ectomycorrhizal fungus Laccaria bicolor (Laccaria). We show that SBW25 has MHB activity and promotes mycorrhization of aspen roots by Laccaria. Using transcriptomic analysis of aspen roots under multiple community compositions, we identify clusters of co-regulated genes associated with mycorrhization, the presence of SBW25, and MHB-associated functions, and we generate a combinatorial logic network that links causal relationships in observed patterns of gene expression in aspen seedling roots in a single Boolean circuit diagram. The predicted regulatory circuit is used to infer regulatory mechanisms associated with MHB activity. CONCLUSIONS: In our laboratory conditions, SBW25 increases the ability of Laccaria to form ectomycorrhizal interactions with aspen seedling roots through the suppression of aspen root antifungal defense responses. Analysis of transcriptomic data identifies that potential molecular mechanisms in aspen roots that respond to MHB activity are proteins with homology to pollen recognition sensors. Pollen recognition sensors integrate multiple environmental signals to down-regulate pollenization-associated gene clusters, making proteins with homology to this system an excellent fit for a predicted mechanism that integrates information from the rhizosphere to down-regulate antifungal defense response genes in the root. These results provide a deeper understanding of aspen gene regulation in response to MHB and suggest additional, hypothesis-driven biological experiments to validate putative molecular mechanisms of MHB activity in the aspen-Laccaria ectomycorrhizal symbiosis.

Treatment of edentulous mandible with metal-resin fixed complete dentures: A 15- to 20-year retrospective study.

OBJECTIVES: The purpose of this retrospective study was to report the implant and prosthetic complications of mandibular metal-resin fixed complete dentures (MRFCDs) opposing a maxillary complete removable dental prosthesis (CRDP) in a 15- to 20-year post-placement follow-up period. MATERIAL AND METHODS: Dental records of 24 edentulous patients treated by a mandibular MRFCD and a maxillary CRDP were reviewed. Complications for the implants, MRFCDs, and CRDPs were recorded in four different recall periods: 0-5 years, 5-10 years, 10-15 years, and more than 15 years. The survival and failure times based on Kaplan-Meier statistics were analyzed using Lifetest procedures. Product-limit survival estimates were used for cumulative survival rates (CSRs). RESULTS: The mean service time was 18.5 years. The CSR for the implants and MRFCDs was 91.8% at 16.9 years (confidence intervals: 85.2% and 95.5%) and 80% at 19.6 years (confidence intervals: 44.1% and 94.1%), respectively. The implant failures after 15 years occurred because of a tumor resection. Acrylic resin tooth fracture (45.8% of patients) and wear (75% of patients) were the most common complications with the MRFCD. Retaining screw complications [loosening (8.1% of retaining screws) and fracture (11.3% of retaining screws)] were also common. CONCLUSIONS: The outcomes seen with MRFCD over the long term were favorable. After placement of prostheses, 1 implant loss was observed potentially due to prosthetic/hygiene/periodontal factors. However, potential maintenance complications such as acrylic resin tooth fracture and/or wear and retaining screw loosening and fracture may occur in the long term when a similar design is used for the mandible.

Ultrasensitive Microstring Resonators for Solid State Thermomechanical Analysis of Small and Large Molecules.

Thermal analysis plays an important role in both industrial and fundamental research and is widely used to study thermal characteristics of a variety of materials. However, despite considerable effort using different techniques, research struggles to resolve the physicochemical nature of many thermal transitions such as amorphous relaxations or structural changes in proteins. To overcome the limitations in sensitivity of conventional techniques and to gain new insight into the thermal and mechanical properties of small- and large-molecule samples, we have developed an instrumental analysis technique using resonating low-stress silicon nitride microstrings. With a simple sample deposition method and postprocess data analysis, we are able to perform rapid thermal analysis of direct instrumental triplicate samples with only pico- to nanograms of material. Utilizing this method, we present the first measurement of amorphous alpha and beta relaxation, as well as liquid crystalline transitions and decomposition of small-molecule samples deposited onto a microstring resonator. Furthermore, sensitive measurements of the glass transition of polymers and yet unresolved thermal responses of proteins below their apparent denaturation temperature, which seem to include the true solid state glass transition of pure protein, are reported. Where applicable, thermal events detected with the setup were in good agreement with conventional techniques such as differential scanning calorimetry and dynamic mechanical analysis. The sensitive detection of even subtle thermal transitions highlights further possibilities and applications of resonating microstrings in instrumental physicochemical analysis.

Early implant placement for a patient with ectodermal dysplasia: Thirteen years of clinical care.

Patients with ectodermal dysplasia have abnormalities of 2 or more structures that originate from the ectoderm. The oral manifestations often include the congenital absence of teeth and malformed teeth. This clinical report describes the interdisciplinary care from childhood through the definitive dental rehabilitation completed at skeletal maturation to replace the missing teeth in a patient with ectodermal dysplasia. Treatment began at 9 years of age with an implant-assisted mandibular overdenture to improve function and replace the missing mandibular teeth. Orthodontic treatment for the consolidation of space, composite resin restorations, and interim removable dental prostheses were provided to improve esthetics and replace the missing maxillary teeth. Skeletal growth was monitored, and orthognathic surgery was performed at the cessation of growth. The definitive rehabilitation consisted of a mandibular fixed dental prosthesis supported by dental implants and a maxillary removable dental prosthesis to restore the patient to esthetics and function.

Nanomechanical Infrared Spectroscopy with Vibrating Filters for Pharmaceutical Analysis.

Standard infrared spectroscopy techniques are well-developed and widely used. However, they typically require milligrams of sample and can involve time-consuming sample preparation. A promising alternative is represented by nanomechanical infrared spectroscopy (NAM-IR) based on the photothermal response of a nanomechanical resonator, which enables the chemical analysis of picograms of analyte directly from a liquid solution in only a few minutes. Herein, we present NAM-IR using perforated membranes (filters). The method was tested with the pharmaceutical compound indomethacin to successfully perform a chemical and morphological analysis on roughly 100 pg of sample. With an absolute estimated sensitivity of 109±15 fg, the presented method is suitable for ultrasensitive vibrational spectroscopy.

Inactivation of the microRNA-183/96/182 cluster results in syndromic retinal degeneration.

The microRNA-183/96/182 cluster is highly expressed in the retina and other sensory organs. To uncover its in vivo functions in the retina, we generated a knockout mouse model, designated "miR-183C(GT/GT)," using a gene-trap embryonic stem cell clone. We provide evidence that inactivation of the cluster results in early-onset and progressive synaptic defects of the photoreceptors, leading to abnormalities of scotopic and photopic electroretinograms with decreased b-wave amplitude as the primary defect and progressive retinal degeneration. In addition, inactivation of the miR-183/96/182 cluster resulted in global changes in retinal gene expression, with enrichment of genes important for synaptogenesis, synaptic transmission, photoreceptor morphogenesis, and phototransduction, suggesting that the miR-183/96/182 cluster plays important roles in postnatal functional differentiation and synaptic connectivity of photoreceptors.

Nanomechanical Pyrolytic Carbon Resonators: Novel Fabrication Method and Characterization of Mechanical Properties.

Micro- and nanomechanical string resonators, which essentially are highly stressed bridges, are of particular interest for micro- and nanomechanical sensing because they exhibit resonant behavior with exceptionally high quality factors. Here, we fabricated and characterized nanomechanical pyrolytic carbon resonators (strings and cantilevers) obtained through pyrolysis of photoresist precursors. The developed fabrication process consists of only three processing steps: photolithography, dry etching and pyrolysis. Two different fabrication strategies with two different photoresists, namely SU-8 2005 (negative) and AZ 5214e (positive), were compared. The resonant behavior of the pyrolytic resonators was characterized at room temperature and in high vacuum using a laser Doppler vibrometer. The experimental data was used to estimate the Young's modulus of pyrolytic carbon and the tensile stress in the string resonators. The Young's moduli were calculated to be 74 ± 8 GPa with SU-8 and 115 ± 8 GPa with AZ 5214e as the precursor. The tensile stress in the string resonators was 33 ± 7 MPa with AZ 5214e as the precursor. The string resonators displayed maximal quality factor values of up to 3000 for 525-µm-long structures.

Predicting bacterial community assemblages using an artificial neural network approach.

Understanding the interactions between the Earth's microbiome and the physical, chemical and biological environment is a fundamental goal of microbial ecology. We describe a bioclimatic modeling approach that leverages artificial neural networks to predict microbial community structure as a function of environmental parameters and microbial interactions. This method was better at predicting observed community structure than were any of several single-species models that do not incorporate biotic interactions. The model was used to interpolate and extrapolate community structure over time with an average Bray-Curtis similarity of 89.7. Additionally, community structure was extrapolated geographically to create the first microbial map derived from single-point observations. This method can be generalized to the many microbial ecosystems for which detailed taxonomic data are currently being generated, providing an observation-based modeling technique for predicting microbial taxonomic structure in ecological studies.

Modeling forest ecosystem responses to elevated carbon dioxide and ozone using artificial neural networks.

Rising atmospheric levels of carbon dioxide and ozone will impact productivity and carbon sequestration in forest ecosystems. The scale of this process and the potential economic consequences provide an incentive for the development of models to predict the types and rates of ecosystem responses and feedbacks that result from and influence of climate change. In this paper, we use phenotypic and molecular data derived from the Aspen Free Air CO2 Enrichment site (Aspen-FACE) to evaluate modeling approaches for ecosystem responses to changing conditions. At FACE, it was observed that different aspen clones exhibit clone-specific responses to elevated atmospheric levels of carbon dioxide and ozone. To identify the molecular basis for these observations, we used artificial neural networks (ANN) to examine above and below-ground community phenotype responses to elevated carbon dioxide, elevated ozone and gene expression profiles. The aspen community models generated using this approach identified specific genes and subnetworks of genes associated with variable sensitivities for aspen clones. The ANN model also predicts specific co-regulated gene clusters associated with differential sensitivity to elevated carbon dioxide and ozone in aspen species. The results suggest ANN is an effective approach to predict relevant gene expression changes resulting from environmental perturbation and provides useful information for the rational design of future biological experiments.

A multi-omic survey of black cottonwood tissues highlights coordinated transcriptomic and metabolomic mechanisms for plant adaptation to phosphorus deficiency.

Introduction: Phosphorus (P) deficiency in plants creates a variety of metabolic perturbations that decrease photosynthesis and growth. Phosphorus deficiency is especially challenging for the production of bioenergy feedstock plantation species, such as poplars (Populus spp.), where fertilization may not be practically or economically feasible. While the phenotypic effects of P deficiency are well known, the molecular mechanisms underlying whole-plant and tissue-specific responses to P deficiency, and in particular the responses of commercially valuable hardwoods, are less studied. Methods: We used a multi-tissue and multi-omics approach using transcriptomic, proteomic, and metabolomic analyses of the leaves and roots of black cottonwood (Populus trichocarpa) seedlings grown under P-deficient (5 µM P) and replete (100 µM P) conditions to assess this knowledge gap and to identify potential gene targets for selection for P efficiency. Results: In comparison to seedlings grown at 100 µM P, P-deficient seedlings exhibited reduced dry biomass, altered chlorophyll fluorescence, and reduced tissue P concentrations. In line with these observations, growth, C metabolism, and photosynthesis pathways were downregulated in the transcriptome of the P-deficient plants. Additionally, we found evidence of strong lipid remodeling in the leaves. Metabolomic data showed that the roots of P-deficient plants had a greater relative abundance of phosphate ion, which may reflect extensive degradation of P-rich metabolites in plants exposed to long-term P-deficiency. With the notable exception of the KEGG pathway for Starch and Sucrose Metabolism (map00500), the responses of the transcriptome and the metabolome to P deficiency were consistent with one another. No significant changes in the proteome were detected in response to P deficiency. Discussion and conclusion: Collectively, our multi-omic and multi-tissue approach enabled the identification of important metabolic and regulatory pathways regulated across tissues at the molecular level that will be important avenues to further evaluate for P efficiency. These included stress-mediating systems associated with reactive oxygen species maintenance, lipid remodeling within tissues, and systems involved in P scavenging from the rhizosphere.

Bone ingrowth and initial stability of titanium and porous tantalum dental implants: a pilot canine study.

PURPOSE: To investigate if a dental implant system with a midsection covered by 3-dimensionally porous tantalum material would exhibit stability comparable with a traditional threaded titanium alloy implant system and whether bone would grow into the porous section. METHODS: Three experimental and 3 control implants were placed in the individual mandibles of 8 dogs. Resonance frequency analysis assessed implant stability at 0, 2, 4, 8, and 12 weeks of healing. Histomorphometric and backscattered scanning electron microscopic analyses examined the presence of bone ingrowth into the experimental implant's porous section and bone-to-implant contact along the titanium surfaces of both implants. RESULTS: Implant stability did not significantly differ during 0 to 12 weeks of healing. Progressive tissue mineralization developed inside porous sections from weeks 2 to 12. Porous implants exhibited a combination of progressive osseointegration along their titanium surfaces and bone ingrowth inside their porous tantalum sections. CONCLUSIONS: Cortical and apical implant threads, combined with the porous section, were able to stabilize the experimental implant to the same degree as the fully threaded control implant.

Outcomes of a contemporary credentialing and privileging program in a dental school.

The current credentialing and privileging (C&P) climate has evolved due to a risk reduction/management awareness of increased institutional legal liability. This recognition affects dental colleges and has caused the implementation of C&P processes. Contemporary best practices for methods, processes, and structure are reported here. Data reported from the process show how and what clinicians' red flags were discovered during the process. Conclusions include the following: C&P is a significant process to introduce in terms of institutional resources and commitment. This process includes increased clinician and administrative burden that needs to have a governor. Attention to experiences of other institutions can reduce but not eliminate challenges from the clinician and some administrators. A primary data-based verification process administered by a credentialing specialist can make the process valid and workable.

Evaluation of a mucoadhesive fenretinide patch for local intraoral delivery: a strategy to reintroduce fenretinide for oral cancer chemoprevention.

Systemic delivery of fenretinide in oral cancer chemoprevention trials has been largely unsuccessful due to dose-limiting toxicities and subtherapeutic intraoral drug levels. Local drug delivery, however, provides site-specific therapeutically relevant levels while minimizing systemic exposure. These studies evaluated the pharmacokinetic and growth-modulatory parameters of fenretinide mucoadhesive patch application on rabbit buccal mucosa. Fenretinide and blank-control patches were placed on right/left buccal mucosa, respectively, in eight rabbits (30 min, q.d., 10 days). No clinical or histological deleterious effects occurred. LC-MS/MS analyses of post-treatment samples revealed a delivery gradient with highest fenretinide levels achieved at the patch-mucosal interface (no metabolites), pharmacologically active levels in fenretinide-treated oral mucosa (mean: 5.65 µM; trace amounts of 4-oxo-4-HPR) and undetectable sera levels. Epithelial markers for cell proliferation (Ki-67), terminal differentiation (transglutaminase 1-TGase1) and glucuronidation (UDP-glucuronosyltransferase1A1-UGT1A1) exhibited fenretinide concentration-specific relationships (elevated TGase1 and UGT1A1 levels <5 µM, reduced Ki-67 indices >5 µM) relative to blank-treated epithelium. All fenretinide-treated tissues showed significantly increased intraepithelial apoptosis (TUNEL) positivity, implying activation of intersecting apoptotic and differentiation pathways. Human oral mucosal correlative studies showed substantial interdonor variations in levels of the enzyme (cytochrome P450 3A4-CYP3A4) responsible for conversion of fenretinide to its highly active metabolite, 4-oxo-4-HPR. Complementary in vitro assays in human oral keratinocytes revealed fenretinide and 4-oxo-4-HPR's preferential suppression of DNA synthesis in dysplastic as opposed to normal oral keratinocytes. Collectively, these data showed that mucoadhesive patch-mediated fenretinide delivery is a viable strategy to reintroduce a compound known to induce keratinocyte differentiation to human oral cancer chemoprevention trials.

Modeling interaction networks between host, diet, and bacteria predicts obesogenesis in a mouse model.

Host-microbiome interactions are known to have substantial effects on human health, but the diversity of the human microbiome makes it difficult to definitively attribute specific microbiome features to a host phenotype. One approach to overcoming this challenge is to use animal models of host-microbiome interaction, but it must be determined that relevant aspects of host-microbiome interactions are reflected in the animal model. One such experimental validation is an experiment by Ridura et al. In that experiment, transplanting a microbiome from a human into a mouse also conferred the human donor's obesity phenotype. We have aggregated a collection of previously published host-microbiome mouse-model experiments and combined it with thousands of sequenced and annotated bacterial genomes and metametabolomic pathways. Three computational models were generated, each model reflecting an aspect of host-microbiome interactions: 1) Predict the change in microbiome community structure in response to host diet using a community interaction network, 2) Predict metagenomic data from microbiome community structure, and 3) Predict host obesogenesis from modeled microbiome metagenomic data. These computationally validated models were combined into an integrated model of host-microbiome-diet interactions and used to replicate the Ridura experiment in silico. The results of the computational models indicate that network-based models are significantly more predictive than similar but non-network-based models. Network-based models also provide additional insight into the molecular mechanisms of host-microbiome interaction by highlighting metabolites and metabolic pathways proposed to be associated with microbiome-based obesogenesis. While the models generated in this study are likely too specific to the animal models and experimental conditions used to train our models to be of general utility in a broader understanding of obesogenesis, the approach detailed here is expected to be a powerful tool of investigating multiple types of host-microbiome interactions.

Effects of a topically applied bioadhesive berry gel on loss of heterozygosity indices in premalignant oral lesions.

PURPOSE: The aim of this study was to assess the effects of topical application of a 10% (w/w) freeze-dried black raspberry (FBR) gel on oral intraepithelial neoplasia (IEN) variables that included histologic diagnoses and loss of heterozygosity (LOH) indices. Microsatellite instability and/or LOH at tumor suppressor gene-associated chromosomal loci have been associated with a higher risk for oral IEN progression to oral squamous cell carcinoma. Previously, our laboratories have shown that FBRs are well tolerated and possess potent antioxidant, apoptotic, and differentiation-inducing properties. EXPERIMENTAL DESIGN: Each participant with IEN served as their own internal control. Before treatment, all lesions were photographed, and lesional tissue was hemisected to obtain a pretreatment diagnosis and baseline biochemical and molecular variables. Gel dosing (0.5 g applied four times daily for 6 weeks) was initiated 1 week after the initial biopsy. Genomic DNA was isolated from laser-captured basilar and suprabasilar surface epithelial cells followed by PCR amplification using primer sets that targeted known and presumed tumor suppressor gene loci associated with INK4a/ARF, p53, and FHIT. Allelic imbalance was determined by sequence analysis using normal participant tissues to establish microsatellite marker peak patterns and allele sizes. RESULTS: Confirming earlier phase I data, none of the 27 participants developed FBR gel-associated toxicities. Furthermore, our results show histologic regression in a subset of patients as well as statistically significant reduction in LOH at tumor suppressor gene-associated loci. CONCLUSIONS: These preliminary data suggest that further evaluation of berry gels for oral IEN chemoprevention is warranted.

Effects of implant design and surface roughness on crestal bone and soft tissue levels in the esthetic zone.

PURPOSE: To evaluate the effects of implant geometry and collar macrostructure and microstructure on crestal bone height, mobility values, and attachment levels using two different types of radiographic analysis. MATERIALS AND METHODS: Subjects included in this study were those who had single-tooth implants restored in the maxillary esthetic zone. Bone levels were measured from the implant-abutment (IA) interface on standardized radiographs at baseline and after 5 years using digital and standard methods. The digital method converted the radiographs into three-dimensional models based on radiodensity so that bone levels could be measured. Clinical attachment levels and Periotest values were recorded and all data were analyzed statistically. RESULTS: The bone position from the IA interface for the rough-collar implant group was a mean of -0.61 +/- 0.08 mm and it was -1.55 +/- 0.10 mm for the smooth-collar implant group (-1.96 +/- 0.16 mm for implants with a stepped collar and -0.85 +/- 0.19 mm for implants with a straight collar). The mean bone level change over the 5-year period for the rough implants was -0.19 +/- 0.09 mm and it was -0.36 +/- 0.06 mm for the smooth implants. The crestal bone position relative to the implant at the time of surgery influenced mean bone level changes significantly. There were statistically significant differences in clinical attachment levels depending on implant location and on implant group and length. CONCLUSIONS: Implants with straight collars had less bone loss at the 5-year interval than implants with stepped collars. Bone level changes were greater when the implant was placed further subcrestal. The digitally converted radiographic method was significantly more precise for evaluating changes in bone levels. These findings suggest that implant geometry and implant surface roughness may play a significant role in bone level changes in the esthetic zone.