Oral Microbiota Composition Predicts Early Childhood Caries Onset.

As the most common chronic disease in preschool children in the United States, early childhood caries (ECC) has a profound impact on a child's quality of life, represents a tremendous human and economic burden to society, and disproportionately affects those living in poverty. Caries risk assessment (CRA) is a critical component of ECC management, yet the accuracy, consistency, reproducibility, and longitudinal validation of the available risk assessment techniques are lacking. Molecular and microbial biomarkers represent a potential source for accurate and reliable dental caries risk and onset. Next-generation nucleotide-sequencing technology has made it feasible to profile the composition of the oral microbiota. In the present study, 16S ribosomal RNA (rRNA) gene sequencing was applied to saliva samples that were collected at 6-mo intervals for 24 mo from a subset of 56 initially caries-free children from an ongoing cohort of 189 children, aged 1 to 3 y, over the 2-y study period; 36 children developed ECC and 20 remained caries free. Analyses from machine learning models of microbiota composition, across the study period, distinguished between affected and nonaffected groups at the time of their initial study visits with an area under the receiver operating characteristic curve (AUC) of 0.71 and discriminated ECC-converted from healthy controls at the visit immediately preceding ECC diagnosis with an AUC of 0.89, as assessed by nested cross-validation. Rothia mucilaginosa, Streptococcus sp., and Veillonella parvula were selected as important discriminatory features in all models and represent biomarkers of risk for ECC onset. These findings indicate that oral microbiota as profiled by high-throughput 16S rRNA gene sequencing is predictive of ECC onset.

Our Essential and Endangered Dentist-Scientist Workforce.

Future advances in dental medicine rely on a robust and stable pipeline of dentist-scientists who are dedicated to research inspired by the patients' condition. The biomedical research community faces external and internal pressures that have been building over years. This is now threatening the current and future status of basic, translational and patient-oriented research by dentist-scientists who study dental, oral and craniofacial diseases, population sciences, and prevention. The dental academic, research and practicing communities can no longer ignore the warning signs of a system that is under considerable stress. Here, the authors report findings of the Physician-Scientist Workforce Working Group, charged by the National Institutes of Health (NIH) Director, to perform quantitative and qualitative analyses on dentist-scientists by addressing the size, composition and activities of the group, relative to other health professions. From 1999 to 2012, trends in the numbers of grant applications and awards to dentist-scientists point to an overall decline. Disturbing are the low numbers of new investigators who apply for Early Career NIH Programs. While more seasoned dentist researchers enjoy greater success, the average age of first-time funded dentists is 52.7 y for females and 54.6 y for males, with a relatively low number of applications submitted and funded. These new data led the panel to stress the need to expand the capacity of the dentist-scientist workforce to leverage technologies and research opportunities that benefit the profession at-large. Suggestions were made to invest in developing clinical research faculty, including those with foreign degrees, through new training mechanisms. The creation of new alliances between national organizations like the American Association for Dental Research, the American Dental Education Association and the American Dental Association will undoubtedly lead to bold and concerted actions that must be pursued with a sense of urgency. A more supportive culture within dental schools and universities for dentist-scientists is needed, as their success is critical to the future career choices of their mentees. Knowledge Transfer Statement: Advances in dental medicine rely on a pipeline of dentist-scientists who are dedicated to research inspired by the patients' condition. Despite the recent advancement in technology and innovation, the dental community can no longer ignore the various pressures that threaten the future of the dentist-scientist profession. Here, the authors report findings of the Physician-Scientist Workforce Working Group of NIH that were published in 2014, and draw attention to the key issues threatening the NIH-funded pool of dentist-scientists.

The use of absorbable monofilament polydioxanone suture in pediatric cardiovascular operations.

Growth of suture lines and anastomoses is required for long-term success after the repair of congenital cardiovascular anomalies. Polydioxanone, an absorbable monofilament suture material, has been used in a variety of operations since April, 1983. Twenty-two of the 46 procedures were coarctation repairs. Complete repairs for anomalous pulmonary veins and transposition of the great arteries, as well as Fontan procedures and systemic-pulmonary shunts, have been performed. Angiographic, gross, and microscopic examination showed good healing. There was no anastomotic disruption or aneurysm formation. The results with this absorbable vascular suture have been uniformly encouraging in a follow-up of up to 30 months.

Immunodetection of Parkin protein in vertebrate and invertebrate brains: a comparative study using specific antibodies.

Parkin is an intracellular protein that plays a significant role in the etiopathogenesis of autosomal recessive juvenile parkinsonism. Using immunoblot methods, we found Parkin isoforms varying from 54 to 58 kDa in rat, mouse, bird, frog and fruit-fly brains. Immunocytochemical studies carried out in rats, mice and birds demonstrated multiple cell types bearing the phenotype for Parkin throughout telencephalic, diencephalic, mesencephalic and metencephalic brain structures. While in some instances Parkin-containing neurons tended to be grouped into clusters, the majority of these labeled nerve cells were widely scattered throughout the neuraxis. The topographical distribution and organizational pattern of Parkin within major functional brain circuits was comparable in both rats and mice. However, the subcellular localization of Parkin was found to vary significantly as a function of antibody reactivity. A consistent cytoplasmic labeling for Parkin was observed in rodent tissue incubated with a polyclonal antibody raised against the human Parkin protein and having an identical amino-acid sequence with that of the rat. In contrast, rodent tissue alternately incubated with a polyclonal antibody raised against a different region of the same human Parkin protein but having 10 mismatched amino-acid sequence changes with those of the rat and mouse, resulted in nuclear labeling for Parkin in rat but not mouse neurons. This difference in epitope recognition, however, was reversed when mouse brain tissue was heated at 80 degrees C, apparently unmasking target epitopes against which the antisera were directed. Collectively, these results show a high degree of conservation in the cellular identity of Parkin in animals as different as drosophilids and mammals and points to the possibility that the biochemical specificities of Parkin, including analogous functional roles, may have been conserved during the course of evolution.

The storage stability of 3-hydroxybutyrate in serum, plasma, and whole blood.

The storage stability of 3-hydroxybutyrate in whole blood, serum, and plasma was evaluated. The levels of 3-hydroxybutyrate were measured using an enzymatic rate reaction of the specific 3-hydroxybutyrate dehydrogenase with the NAD-NADH coupled reaction. 3-Hydroxybutyrate was found to be a stable analyte in whole blood, plasma, and serum. The long-term stability of beta-hydroxybutyrate allows ample time for separation of blood components and offers storage of samples to meet quality control needs as well as the possibility of mailing specimens. Further studies indicate that NaF plasma, heparin plasma, and serum are the preferred specimens, because both EDTA and oxalate showed the most significant interference with the determination of 3-hydroxybutyrate.

Approaches to the artificial heart. Invited speaker.

Over the last two decades, the implantable artificial heart has evolved from an idea to a device capable of completely supporting the circulation for periods now exceeding 5 months. Although initial animal studies were limited by thromboembolism and device breakage, the usual causes of death in experimental animals are now infection, atrioventricular valve obstruction, elastomer bladder calcification, or inadequate cardiac output because of the relatively rapid growth of the young calves. As a result of the bulky nature of the energy converter and the substantial risk of infection with large diameter percutaneous tubes, clinical use of their air-powered artificial hearts will be limited to patients who are awaiting or being prepared for heart transplantation. Artificial hearts with implanted energy converters are being developed for permanent heart replacement. These devices require well-designed, durable mechanical components and sophisticated control systems. Although initial designs centered around thermal engines powered by a completely implantable nuclear energy source, the excessive cost and potential dangers have shifted the focus away from the nuclear system. Several electrically driven artificial hearts, based on samarium-cobalt magnet brushless direct-current motors, are now undergoing bench testing and will be ready for long-term animal studies within 2 years. This research will culminate with the availability of an "off-the-shelf" electrically powered artificial heart for use in patients with a wide range of nonrepairable forms of end-stage heart disease.

End-to-end repair of aortic coarctation using absorbable polydioxanone suture.

Based on previous laboratory work, we have used polydioxanone absorbable suture in a variety of vascular and cardiac repairs in pediatric patients. However, some investigators have expressed concern about the potential for aneurysm formation at the anastomotic site. Between March 1983 and June 1989, 15 patients (7 male, 8 female) aged 2.5 months to 9.2 years (mean, 3.7 years) had resection of coarctation of the aorta and end-to-end anastomosis with polydioxanone absorbable suture. Thirteen patients have returned for routine postoperative evaluation, the follow-up time ranging from 11 to 49 months (mean, 23 months). Noninvasive two-dimensional, pulsed-wave Doppler and color echocardiography and magnetic resonance imaging studies demonstrated good anatomical repair and no anastomotic aneurysm formation or residual coarctation of the aorta in any patient after end-to-end anastomosis with polydioxanone. In summary, this intermediate follow-up study has revealed no vascular complications related to the repair of coarctation with absorbable polydioxanone suture.

Results of aortic anastomoses made under tension using polydioxanone suture.

After early repair of congenital cardiovascular defects, such as coarctation of the aorta, late stenosis may become a problem. Use of absorbable sutures has been shown to be superior to use of nonabsorbable sutures in allowing growth of an anastomotic site along with the individual. Some concern has been raised, however, about the potential for aneurysm formation at the site of anastomosis when absorbable sutures are used. This study was undertaken to observe the effects of longitudinal tension on anastomoses made with absorbable polydioxanone suture in growing animals. Six piglets (aged 3 to 4 weeks) underwent a 1-cm resection of the infrarenal aorta and reanastomosis with polydioxanone suture. One animal died prematurely of respiratory illness. The 5 remaining animals were killed after 6 months. The excised aortas demonstrated no stenosis, no dilatation, and no burst-test failure to 250 mm Hg. Histological examination showed disrupted elastic laminae without thinning of aortic wall in all samples of aorta. We conclude that polydioxanone suture is a suitable suture material for vascular anastomoses made under tension where growth of the anastomotic site is expected.

Vascular anastomoses in growing vessels: the use of absorbable sutures.

Primary end-to-end infrarenal aortic anastomoses were performed in 36 piglets using two synthetic absorbable suture materials: polydioxanone and coated polyglactin. Animals were killed at 1, 4, and 11 weeks and 6 months following operation. Each aorta was removed, burst-tested to 300 mm Hg, radiographed, and examined histologically. All anastomoses were patent, and no burst-test failures occurred. Stenosis occurred in 14 of 17 animals at 1 and 4 weeks, respectively. One of 5 animals exhibited stenosis at 11 weeks, and none of the 14 animals had stenosis 6 months postoperatively. Histological examination revealed fibrosis replacing areas of disrupted elastica at 6 months in both suture groups. This study suggests that absorbable suture material, in particular polydioxanone, because of its excellent handling characteristics and prolonged tensile strength retention, will be useful for the repair of vascular and cardiac anomalies where growth of the suture line is required.

A low-cost fiber-optic instrument to colorimetrically detect patients with Barrett's esophagus for early detection of esophageal adenocarcinoma.

In Barrett's esophagus, the precursor to esophageal adenocarcinoma, the squamocolumnar junction (SCJ) between the normal esophagus and the stomach like mucosa is proximally displaced. Currently it can be detected only by an expensive upper GI endoscopic procedure. We have developed a minimally invasive and easy to operate colorimetric instrument for the low-cost detection of Barrett's esophagus. The instrument is based on a flexible, narrow diameter, fiber-optic probe that performs a colorimetric scan of the esophageal lumen. The instrument was clinically evaluated in 50 subjects. The instrument could identify both symmetric and asymmetric SCJ's. The SCJ locations determined by the colorimetric instrument correlated strongly (R2 = 0.89) with those determined by endoscopy. The instrument identified the SCJ locations accurately (Mean of difference +/- SEM: 0.97 +/- 1.72 cm) and reproducibly (Mean of absolute difference +/- SEM: 1.33 +/- 1.40 cm). The instrument has a 90% sensitivity of identifying patients with Barrett's esophagus, based on the clinical algorithm that if the SCJ is located at a distance less than 37 cm from the teeth, then the subject has Barrett's esophagus, otherwise the subject does not have Barrett's esophagus. In conclusion, the colorimetric instrument has the potential of being a cost-effective way of determining patients likely to have Barrett's esophagus in the population.

Observations on the use of a cation exchange resin for the preparation of metal-depleted media for lymphocyte culture.

A chelating resin specific for divalent cations (Chelex) was used to prepare metal-depleted media for lymphocyte culture. A batch procedure (resin in pH 7.4 phosphate buffer/specimen, 1:1) removed 70-80% of iron, 77-87% of copper and 88-98% of zinc, calcium and magnesium. At variance with other reports, when a resin/specimen ratio of 1:4 was used, iron chelation decreased to 40%, whereas other cation chelation remained unchanged. Best chelation for iron and calcium was obtained at pH 5-6.4; for copper, zinc and magnesium, at pH 7.4-8.0. During the procedure protein content decreased by 8-10%; arginine and lysine by 80%; asparagine, cystine, tyrosine and phenylalanine by 60%, other amino acids by 35%. These new data suggest that cation-depleted media prepared with Chelex may be used to study the effects of cations on lymphocytes in culture, provided that the most appropriate pH and resin/specimen ratio are selected and adequate amino acid replacement is performed. Results on normal human lymphocytes are reported.

Demonstration of a possible source of error with an electric pulp tester.

The purpose of this study was to determine whether electrical current can travel between adjacent teeth through contacting interproximal amalgam restorations. Twenty-two extracted human premolars were restored with class II amalgam restorations. They were then mounted in pairs in self-curing resin, simulating adjacent teeth. Only their restorations were allowed to contact. An electrical multimeter was used to supply electrical current and to measure any current passing through the mounted teeth. One current measurement was made from the buccal enamel of a tooth to the occlusal surface of the filling in that tooth. A second measurement was made from the buccal enamel of the same tooth to the occlusal surface of the adjacent filling. These two measurements were made for all 11 trials. Statistically, all current that entered the first tooth passed to the adjacent tooth. This finding suggests a potential source of error when using an electric pulp tester.

Testing of household products and materials for emission of toluene diisocyanate.

Polyurethane products were subjected to chamber testing to determine their emission rates of 2,4- and 2,6-toluene diisocyanate (TDI). The polyurethane (PU) products included carpet padding, furniture cushions, sheet foam, varnishes, and sealants, as well as a commercially-applied water sealant product for concrete that contained up to 4 percent TDI by weight. The PU products were screened in a 9-L glass chamber, under elevated temperature and chamber loading conditions, using both a time-integrated sampling and analysis method specific for TDI and a continuous but non-specific real-time monitor for isocyanates. None of the products normally found in residences showed a positive response in the screening tests, indicating that TDI emissions and consequently toxic effects from such products are negligible. However, the commercially-applied water sealant gave a positive response in the screening test. Further testing of that product at realistic temperatures showed initial TDI emission rates of about 300,000 micrograms/m2/hr, with emissions lasting only one hour or less. At 21 and 27 degrees C, about 1 percent and 5 percent, respectively, of the TDI content of the product was released to the air. The emitted TDI was predominantly the 2,6-isomer, although the TDI originally present in the product was predominantly the 2,4-isomer.

Evaluation of a simple weight-loss method for determining the permeation of organic liquids through rubber films.

The standard ASTM method for determining permeation of liquid penetrants through polymeric films (F739-85) was compared with a simple weight-loss method. The weight-loss permeation cell was constructed of off-the-shelf components, a conventional analytical balance was used to measure the weight loss, and a chamber with continuous ventilation was used to hold the cell at constant temperature and evacuate the permeating penetrant. The advantages and limitations of the weight-loss method were illustrated by using data obtained from the permeation of acetonitrile, n-hexane, and methanol through films of four acrylonitrile-butadiene copolymers. The steady-state flux obtained by using the weight-loss method gave results statistically equivalent to the more analytically complex ASTM standard method. The weight-loss method required the experimenter to monitor the weight of the cell and its contents over time to obtain the steady-state flux whereas the ASTM method required the experimenter to chemically analyze for the penetrant concentration in the effluent gas stream as a function of time. The ASTM method required more analytical skill and training and more costly analytical equipment than did the simple weight-loss method. The weight-loss method needs further improvement and validation but shows promise even in its present form. By using the weight-loss method, the potential exists for far more chemical protective clothing users to conduct their own low-cost permeation testing as an initial screening to determine the relative permeation performance of candidate protective clothing materials. The weight-loss method is not meant to replace the standard ASTM method but to supplement it as a screening test.

Three-dimensional (3-D) imaging in post-mortem examinations: elucidation and identification of cranial and facial fractures in victims of homicide utilizing 3-D computerized imaging reconstruction techniques.

The analysis of cranial and facial fractures in skeletal remains of homicidal victims can prove challenging for forensic anthropologists and forensic pathologists in postmortem examination. In such cases, the use of 3-D computerized imaging to elucidate the fractures and patterns of injuries can provide strong medical evidence that is very useful during litigation and at trial. The authors describe 3-D reconstructions of the skull performed as part of forensic postmortem examination in a recent victim of homicide.