Metagenomic analysis of the viral flora of pine marten and European badger feces.

A thorough understanding of the diversity of viruses in wildlife provides epidemiological baseline information about potential pathogens. Metagenomic analysis of the enteric viral flora revealed a new anellovirus and bocavirus species in pine martens and a new circovirus-like virus and geminivirus-related DNA virus in European badgers. In addition, sequences with homology to viruses from the families Paramyxo- and Picornaviridae were detected.

Genogroup I and II picobirnaviruses in respiratory tracts of pigs.

Sequence-independent amplification and specific reverse transcription PCRs identified genogroup I and II picobirnaviruses in respiratory tracts of pigs. These data expand knowledge of picobirnavirus diversity and tropism. Genetic relationships between porcine respiratory and human enteric picobirnaviruses suggest cross-species transmission of picobirnaviruses between pigs and humans.

Pulmonary pathology of pandemic influenza A/H1N1 virus (2009)-infected ferrets upon longitudinal evaluation by computed tomography.

We investigated the development of pulmonary lesions in ferrets by means of computed tomography (CT) following infection with the 2009 pandemic A/H1N1 influenza virus and compared the scans with gross pathology, histopathology and immunohistochemistry. Ground-glass opacities observed by CT scanning in all infected lungs corresponded to areas of alveolar oedema at necropsy. These areas were most pronounced on day 3 and gradually decreased from days 4 to 7 post-infection. This pilot study shows that the non-invasive imaging procedure allows quantification and characterization of influenza-induced pulmonary lesions in living animals under biosafety level 3 conditions and can thus be used in pre-clinical pharmaceutical efficacy studies.

Efficacy of the enamel matrix derivative to induce cementogenesis in vital and endodontically treated teeth with osseous dehiscence defects.

This experiment assessed the efficacy of the enamel matrix derivative (EMD) to regenerate cementum in vital and endodontically treated teeth with osseous dehiscence defects. Five adult female beagle dogs were used. Thirty maxillary teeth (bilateral maxillary canines and second and fourth premolars) were randomly divided into two experimental groups (groups A and B, containing 12 teeth each) and one control group (group C). Endodontic treatment was only performed on teeth in group A compared with teeth in groups B and C. Buccal osseous dehiscence defects were surgically created in teeth from all groups. Teeth in the experimental group were treated with the EMD, whereas the controls were not. After 5 months, the animals were sacrificed and block sections of the teeth in experimental and control groups were processed for histological analysis. Newly regenerated cementum was observed in all teeth in groups A and B. No cementum regeneration was observed in group C. There was a significant difference in cementum generation between the experimental and control groups (P < 0.001). EMD therapy induces cementogenesis in vital and endodontically treated teeth with osseous dehiscence defects.

Identification and characterization of deer astroviruses.

The threat of emerging infectious viruses in humans requires a more effective approach regarding virus surveillance. A thorough understanding of virus diversity in wildlife provides epidemiological baseline information about pathogens and may lead to the identification of newly emerging pathogens in the future. In this study, diarrhoea samples from an outbreak of gastrointestinal illness in a Danish population of European roe deer were gathered for which no aetiological agent could be identified. Large-scale molecular RNA virus screening, based on host nucleic acid depletion, sequence-independent amplification and sequencing of partially purified viral RNA, revealed the presence of novel astroviruses, CcAstV-1 and CcAstV-2, in two of ten diarrhoea samples. Whether these viruses were responsible for causing diarrhoea remains to be determined. Phylogenetic analyses on amplified sequences showed that these viruses were most closely related to each other, were a novel species in the genus Mamastrovirus and may represent two different serotypes.

Human picobirnaviruses identified by molecular screening of diarrhea samples.

The global threat of (re)emerging infectious viruses requires a more effective approach regarding virus surveillance and diagnostic assays, as current diagnostics are often virus species specific and not able to detect highly divergent or unknown viruses. A systematic exploration of viruses that infect humans is the key to effectively counter the potential public health threat caused by new and emerging infectious diseases. The human gut is a known reservoir of a wide variety of microorganisms, including viruses. In this study, Dutch clinical diarrhea samples for which no etiological agent could be identified by available cell culture, serological, or nucleic acid-based tests were gathered. Large-scale molecular RNA virus screening based on host nucleic acid depletion, sequence-independent amplification, and sequencing of partially purified viral RNA from a limited number of clinical diarrhea samples revealed four eukaryotic virus species. Among the detected viruses were a rhinovirus and a new picobirnavirus variant. In total, approximately 20% of clinical diarrhea samples contained human picobirnavirus sequences. The Dutch picobirnaviruses belonged to different phylogenetic clades and did not group with other picobirnaviruses according to year of isolation or host species. Interestingly, the average age of patients infected with picobirnavirus was significantly higher than that of uninfected patients. Our data show that sequence-independent amplification of partially purified viral RNA is an efficient procedure for identification of known and highly divergent new RNA viruses in clinical diarrhea samples.

Human astrovirus infection in a patient with new-onset celiac disease.

Many diseases with unknown etiology may be caused by unidentified viruses. Sequence-independent amplification revealed a new astrovirus, similar to VA1, in a 4-year-old male diagnosed with celiac disease. This expands the geographic range of this virus to include Europe and may associate astrovirus infection with the onset of celiac disease.

Healing of apical periodontitis after endodontic treatment with and without obturation in dogs.

The principle of obturation of the root canal space is generally accepted. This belief has dominated the field of endodontics despite the absence of credible scientific evidence. Little information is available on the possible relationships between endodontic success and obturation of the root canal system. The present study compares the healing of instrumented and obturated versus instrumented and nonobturated root canal systems with apical periodontitis. Fifty-six root canals in 28 third and fourth bilateral lower premolar teeth with completely formed apices in seven mixed German Shepherd dogs were used. Apical lesions were created by accessing the canals, removing the pulp and leaving them open to the oral environment for 42 days. The teeth in the control group were instrumented, irrigated, and then obturated using a lateral condensation technique with gutta-percha cones and AH26 Plus as the endodontic sealer. The teeth in the experimental group were instrumented and irrigated without obturation. All teeth in both groups were sealed coronally. After 190 days, the animals were euthanized. Vital perfusion with 10% formalin through common carotid artery was performed. Thirty to 40 serial 5-microm thick sections were obtained from each root and stained with hematoxylin and eosin for histological evaluation by observer blinded to the treatment allocation. The control group showed less cementum and dentin resorption in comparison with test group (p < 0.5). No statistically significant difference was found in other parameters between the control and test groups (p > 0.05). The noteworthy finding of this study was that there was no difference in healing of apical periodontitis between the instrumented and obturated and instrumented and nonobturated root canal system. The success of endodontic treatment ultimately depends on the elimination of the microorganism, host response and mechanical closure (coronal seal) of treated root canals that may provide a potential for future bacterial contamination.

Comparison of antifungal activity of white-colored and gray-colored mineral trioxide aggregate (MTA) at similar concentrations against Candida albicans.

The killing effect of similar concentrations of white-colored MTA and gray-colored MTA against Candida albicans was assessed in vitro using the tube dilution test. A direct correlation was found between MTA concentration and its killing effect on C. albicans. At time period zero, both white-colored and gray-colored MTA samples showed Candida growth regardless of the concentration. Tubes containing gray-colored MTA in concentrations of 50, 25, 12.5, 6.25, and 3.125 mg/ml and tubes containing white-colored MTA in concentrations of 50 and 25 mg/ml did not show C. albicans growth at any of the other time periods tested. Tubes containing white-colored MTA in concentrations of 12.5 mg/ml or less showed Candida growth at all the time periods tested. Statistically, a significant difference was found between tubes containing either 50 mg/ml or 25 mg/ml and tubes containing lower concentrations of white-colored MTA (p < 0.001). A significant difference was also found between tubes containing gray-colored MTA in concentrations of 12.5 mg/ml or less and tubes containing similar concentrations of white-colored MTA (p < 0.001). It appears that both gray-colored and white-colored MTA in concentrations of 50 mg/ml and 25 mg/ml are effective in killing C. albicans for periods of up to 1 wk. Lower concentrations of gray-colored MTA may still be effective while lower concentrations of white-colored MTA may not.

Antibacterial effect of two mineral trioxide aggregate (MTA) preparations against Enterococcus faecalis and Streptococcus sanguis in vitro.

The antibacterial effects of gray-colored MTA (GMTA) and white-colored MTA (WMTA) against Enterococcus faecalis and Streptococcus sanguis were assessed in vitro using the tube dilution test. Broth tubes were prepared and divided into experimental and control groups. Aliquots of each of the tested microorganisms were taken from a stock culture and added to each experimental and positive control group. All groups were incubated at 37 degrees C and evaluated for turbidity at 0, 1, 24, 48, and 72-hour time periods. A direct correlation was found between GMTA and WMTA concentrations and their antibacterial effect. Tubes containing GMTA in concentrations of 50, 25, and 12.5 mg/ml did not show E. faecalis growth at any of the time periods tested whereas tubes containing WMTA showed E. faecalis growth at all concentrations and time periods tested. Statistically significant differences were found between tubes containing GMTA in concentrations of 50, 25 and 12.5 mg/ml and tubes containing similar concentrations of WMTA (p < 0.001). Tubes containing GMTA in concentrations of 50, 25, 12.5, 6.25, and 3.12 mg/ml and tubes containing WMTA in concentrations of 50, 25, and 12.5 mg/ml did not show S. sanguis growth at any of the time periods tested. Statistically significant differences were found between tubes containing GMTA in concentrations 6.25 and 3.12 mg/ml and tubes containing similar concentrations of WMTA (p < 0.001). It appears that the susceptibility of E. faecalis and S. sanguis to MTA differed and that GMTA requires lower concentrations than WMTA to exert the same antibacterial effect against each of the microorganisms tested.

Differential diagnosis of large periapical lesions using cone-beam computed tomography measurements and biopsy.

This study compared the differential diagnosis of large periapical lesions (granuloma versus cyst) to traditional biopsy using Cone Beam Computed Tomography (CBCT), the NewTom 3G. Large lesions were scanned and a preoperative diagnosis based on gray value measurements of the imaged lesion area was made. After surgery a biopsy report was obtained and compared to the CBCT diagnosis. In 13 out of 17 cases, the diagnosis coincided. In 4 out of 17 cases, the CBCT read cyst with the oral pathologist's diagnosis being granuloma. Thus, the CBCT may provide a more accurate diagnosis than biopsy and histology providing a diagnosis without invasive surgery and/or waiting a year to see if nonsurgical therapy is effective.

Anatomical redesign for the treatment of dens invaginatus type III with open apexes: a literature review and case presentation.

BACKGROUND: Dens invaginatus is a rare dental anomaly that may give rise to many complex anatomical forms. The complexity of the internal anatomy may create challenges for the complete removal of the diseased pulpal tissue and the subsequent sealing of the canal system. CASE DESCRIPTION: The authors discuss the modification of the internal anatomy under the operating microscope, allowing the clinician better access to treat predictably the canal system with conventional or alternative techniques. CONCLUSION AND CLINICAL IMPLICATIONS: Considering the anatomical variations and the challenges that a dens invaginatus may present, a practitioner may consider a modification of the internal anatomy of the canal system to gain better access for proper instrumentation, disinfection and sealing of the root canal system using conventional or contemporary techniques.

Human saliva penetration of root canals obturated with two types of mineral trioxide aggregate cements.

The sealing ability of orthograde mineral trioxide aggregate (MTA) root canal filling against human saliva was assessed in vitro. Leakage of gray-colored MTA, white-colored MTA and vertically condensed gutta-percha and sealer were compared. Forty-three extracted single-rooted human teeth were serially instrumented to a file size 40/0.06 at the apex and obturated with either gray-colored MTA (group A), white-colored MTA (group B), or gutta-percha and Kerr Canal Sealer EWT (group C). The teeth were then mounted in a model to test for saliva leakage. After 42 days, one root in group A (9.1%), four roots in group B (36.4%), and nine roots in group C (81.8%) showed saliva leakage. Statistically, a significant difference was found between group A and group C (p < 0.001). No statistical difference was found between group A and group B. Out of the root samples that leaked, the one sample in group A leaked after 36 days. Of the four samples in group B, one sample leaked after 32 days, one sample after 33 days, and two samples after 39 days. All nine samples in group C had leaked after 19 days. It appears that under the conditions of this study both MTA preparations may be more resistant to human saliva leakage than vertically condensed gutta-percha and sealer.

Effect of white-colored mineral trioxide aggregate in different concentrations on Candida albicans in vitro.

The antifungal action of different concentrations of white-colored mineral trioxide aggregate (MTA) against Candida albicans was assessed in vitro. Fresh mix of MTA was prepared at concentrations varying from 0.78 mg/ml to 50 mg/ml by dilution with 10 ml molten agar at 45 degrees C. The MTA-agar compound was thoroughly mixed and the uniform mix was then poured into sterile Petri dishes and allowed to set. A total of 348 agar plates were prepared and divided into experimental groups of 11 plates each and control groups of 5 plates each. Plates of agar without MTA served as positive control and plates without C. albicans served as negative control. Fresh inoculate of C. albicans was prepared by growing an overnight culture from a stock culture. Aliquots of C. albicans were then taken from the stock culture and plated on the agar compound of the experimental and positive control groups. All plates were incubated at 37 degrees C for 1, 24, 48, and 72-h periods. At each time period, the presence of C. albicans colonies was assessed and recorded. A direct correlation was found between MTA concentration and its inhibition effect on C. albicans growth. Plates containing MTA in concentration of 50 mg/ml showed significantly better killing action against C. albicans in all of the time periods tested (p < 0.001). Plates containing MTA in concentration of 25 mg/ml showed antifungal activity only at 1 and 24-h time periods. Plates containing lower concentrations of MTA did not show any antifungal activity. It appears that under the conditions of this study, white-colored MTA in concentration of 50 mg/ml is effective in killing C. albicans for periods of up to 3 days. Lower MTA concentrations may not be effective.

Locoregional control in infants with neuroblastoma: role of radiation therapy and late toxicity.

PURPOSE: To review patterns of failure in infants with neuroblastoma and determine late toxicity and efficacy of radiotherapy (RT) on locoregional control. MATERIALS AND METHODS: From 1955 to 1998, 53 children (35 males and 18 females) <1 year old with neuroblastoma were seen at our institution. Twenty-one (40%) were 1 month), and primary site were not found to impact on survival or progression. None of the Stage 1, 2A, or 2B patients recurred. One of 15 Stage 3 and 5 of 6 Stage 4 children recurred (6 distant metastases, 4 local failure). Four of 6 (67%) LN+ patients treated with locoregional RT and 8 of 10 (80%) LN+ patients treated without RT were locally controlled. There was no isolated locoregional relapse. Two Stage 4S patients died of respiratory compromise secondary to hepatomegaly. RT toxicity: For the 20 infants who received RT, 13 are alive with long-term follow-up ranging from 9.3 to 41 years, median 23 years. The 10 and 15-year musculoskeletal toxicity rates were 38.5% and 47.3% for those receiving RT and 3.3% for no RT (p = 0.02, log-rank test). Five of 6 infants <6 months of age and 1 of 7 >/=6 months developed musculoskeletal toxicity. Musculoskeletal effects were seen in 6 RT patients and included bony hypoplasia in 6, scoliosis in 5, soft tissue hypoplasia in 3, slipped capital femoral epiphysis in 2, kyphosis in 1, and osteochondroma in 1. Three required orthopedic intervention, all receiving >/=20 Gy. One child developed bowel obstruction at 21 months and another developed a leiomyosarcoma in the treatment field 34 years after RT. CONCLUSION: Our study shows that most LN+ infants achieve locoregional control without RT. Infants <6 months receiving RT were the most susceptible to musculoskeletal abnormalities. Further studies are needed to determine if cardiovascular anomalies are more frequently seen in children with neuroblastoma.

In vivo determination of extra-target doses received from serial tomotherapy.

BACKGROUND AND PURPOSE: The purpose of this study was to perform in-vivo measurements of extracranial doses received by patients undergoing serial tomotherapy of the head and neck. MATERIAL AND METHODS: Intensity modulated radiotherapy treatment (IMRT) plans were designed for nine patients using the CORVUS treatment planning system (NOMOS Corp.). These plans were delivered using a tertiary collimator dedicated for serial tomotherapy attached to a 10-MV linear accelerator. For each patient, one optically stimulated luminescence dosimeter (OSLD) was placed on the sternum and one on the lower abdomen. The OSLDs were then processed, thereby estimating the in vivo absorbed doses to the sternum and gonads as a function of distance from the treatment site. RESULTS: The OSLDs were shown to measure known doses to within 5%, thereby validating their accuracy for this dose and energy range. In the patient studies, the dose received by the OSLDs varied in direct proportion to the number of monitor units delivered and inversely with the distance from the target volume; the patient dose at a distance of 15 cm from the target is approximately 0.4% of the total monitor units delivered, and drops to below 0.1% of the total MUs at approximately 40 cm from the center of the target. The average sternal dose was 1353 mSv and the average abdominal dose was 327 mSv for an average prescribed dose of 60.1 Gy. This can be attributed, at least partially, to the inefficient treatment delivery that on average required 9.9 MU/0.01 Gy. CONCLUSIONS: While IMRT reduces the normal tissue volume in the high-dose region, it also increases the overall monitor units delivered, and hence the whole-body dose, when compared with conventional treatment delivery. As has been noted in existing literature, these increases in whole-body dose from radiotherapy delivery may increase the likelihood of a radiation-induced secondary malignancy. Therefore, it is important to assess the risk of secondary malignancies from IMRT delivery, and compare this relative risk against the potential benefits of decreased normal tissue complication probabilities.

Herpesviruses in periapical pathosis: an etiopathogenic relationship?

BACKGROUND: Much remains to be learned about the etiopathogenesis of periapical pathosis, especially about the molecular events preceding and causing disease onset. Human cytomegalovirus and Epstein-Barr virus, 2 herpesviruses, are discussed in this review as they relate to apical periodontitis in humans. RESULTS: Cytomegalovirus or Epstein-Barr virus active infections are detected in more than 90% of granulomas of symptomatic and large periapical lesions. Dual infection with cytomegalovirus and Epstein-Barr virus is closely associated with symptomatic lesions. Herpes simplex virus active infection has no apparent relationship to periapical disease. DISCUSSION: The available evidence suggests the involvement of active cytomegalovirus and Epstein-Barr infections in the etiopathogenesis of apical periodontitis. In periapical pathosis, herpesviruses may cause the release of tissue-destructive cytokines, the overgrowth of pathogenic bacteria, and the initiation of cytotoxic or immunopathologic events. Immune impairment resulting from herpesvirus infection may aid bacteria at several stages of the pathogenesis of periapical lesions, including growth in the periapical environment, possible invasion of tissue, and direct damage to tissue. Unraveling the etiology and pathogenesis of periapical pathosis may require a broadening of our experimental approaches to include studies on interactions among herpesviruses, bacteria, and host immune reactions. Understanding the significance of herpesviruses in the development of periapical lesions may aid in the diagnosis, prevention, and treatment of the diseases.

Evaluation of the reliability and accuracy of using cone-beam computed tomography for diagnosing periapical cysts from granulomas.

INTRODUCTION: The purpose of this study was to evaluate the reliability and accuracy of cone-beam computed tomographic (CBCT) imaging against the histopathologic diagnosis for the differential diagnosis of periapical cysts (cavitated lesions) from (solid) granulomas. METHODS: Thirty-six periapical lesions were imaged using CBCT scans. Apicoectomy surgeries were conducted for histopathological examination. Evaluator 1 examined each CBCT scan for the presence of 6 radiologic characteristics of a cyst (ie, location, periphery, shape, internal structure, effects on surrounding structure, and perforation of the cortical plate). Not every cyst showed all radiologic features (eg, not all cysts perforate the cortical plate). For the purpose of finding the minimum number of diagnostic criteria present in a scan to diagnose a lesion as a cyst, we conducted 6 receiver operating characteristic curve analyses comparing CBCT diagnoses with the histopathologic diagnosis. Two other independent evaluators examined the CBCT lesions. Statistical tests were conducted to examine the accuracy, inter-rater reliability, and intrarater reliability of CBCT images. RESULTS: Findings showed that a score of ≥4 positive findings was the optimal scoring system. The accuracies of differential diagnoses of 3 evaluators were moderate (area under the curve = 0.76, 0.70, and 0.69 for evaluators 1, 2, and 3, respectively). The inter-rater agreement of the 3 evaluators was excellent (α = 0.87). The intrarater agreement was good to excellent (κ = 0.71, 0.76, and 0.77). CONCLUSIONS: CBCT images can provide a moderately accurate diagnosis between cysts and granulomas.

Significance of human cytomegalovirus and Epstein-Barr virus in inducing cytokine expression in periapical lesions.

INTRODUCTION: Because herpesviruses might be etiologically involved in periapical pathosis of endodontic origin, this study aimed to determine the occurrence of human cytomegalovirus (HCMV), Epstein-Barr virus (EBV), and the expression of mRNA transcripts of tumor necrosis factor (TNF)-α, γ-interferon (IFN), interleukin (IL)-1ß, IL-6, IL-12, and IL-10 in periapical granulomatous lesions collected in conjunction with apicoectomy. METHODS: A total of 9 symptomatic and 6 asymptomatic teeth with periapical lesions were studied. Periapical samples were collected in conjunction with apicoectomy, which was being performed because of radiographic evidence of incomplete periapical healing after conventional root canal therapy. By using established polymerase chain reaction primers and procedures, polymerase chain reaction assays were used to identify herpesvirus and cytokine gene expression. RESULTS: The difference in occurrence of HCMV, EBV, and cytokines between symptomatic and asymptomatic periapical lesions was statistically significant: HCMV (P = .048), EBV (P = .002), IFN (P = .001), IL-1 (P = .012), IL-6 (P = .026), IL-10 (P = .026), IL-12 (P = .012), and TNF (P < .001) (Mann-Whitney U test). There was a significant correlation between EBV, HCMV, and TNF, γ-IFN, IL-1, and IL-12 in symptomatic periapical lesions (Spearman test). CONCLUSIONS: The present findings provide evidence of a putative role of HCMV and EBV in the pathogenesis of symptomatic periapical pathosis. The release of tissue-destructive cytokines might be of pathogenetic significance.

Bacterial flora of dental periradicular lesions analyzed by the 454-pyrosequencing technology.

INTRODUCTION: Symptomatic teeth with periradicular lesions of infectious origin remain a significant challenge in dentistry, and the reason for the acute perturbation is incompletely understood. The present study used pyrosequencing of bacterial 16S ribosomal RNA (rRNA) genes to characterize the microbiota of periradicular lesions. METHODS: Thirteen periradicular lesions from 11 symptomatic and 2 asymptomatic teeth were sampled during apical surgery. Samples were subjected to DNA extraction and 16S rRNA polymerase chain reaction (PCR) amplification. PCR amplicons were then sequenced by using the Roche 454 GS FLX platform. Data were analyzed with the Quantitative Insights into Microbial Ecology (QIIME) software package. RESULTS: Seven of the 13 periradicular lesions (53.8%) yielded PCR amplicons, which generated 35,731 high-quality DNA sequences belonging to 10 bacterial phyla and 73 bacterial genera. All 7 lesions were associated with symptoms. The phyla with most bacterial taxa were Proteobacteria (proportion of total bacterial taxa, 33.3%), Firmicutes (30.9%), Actinobacteria (12.2%), and Bacteroidetes (11.4%). The most abundant genera were Fusobacterium (average of total sequences, 21.0%), Streptococcus (8.0%), Prevotella (7.5%), Corynebacterium (7.2%), Porphyromonas (6.0%). and Actinomyces (5.8%). CONCLUSIONS: This study demonstrated that the microbiota of symptomatic periapical lesions is predominated by anaerobic bacteria but also contains substantial levels of streptococci, actinomyces, and bacteria not previously identified in the oral cavity. The etiopathogenic role and therapeutic implication of periradicular bacteria need to be determined.