Characterization of the unique oral microbiome of children harboring Helicobacter pylori in the oral cavity.

Objective: Helicobacter pylori infection is acquired in childhood via the oral cavity, although its relationship with the characteristics of the oral microbiome has not been elucidated. In this study, we performed comprehensive analysis of the oral microbiome in children and adults with or without H. pylori in the oral cavity. Methods: Bacterial DNA was extracted from 41 adult and 21 child saliva specimens, and H. pylori was detected using PCR. 16S rRNA gene amplification was performed for next-generation sequencing. Bioinformatic analyses were conducted using Quantitative Insights into Microbial Ecology 2 (QIIME 2). Results: Faith's phylogenetic diversity analysis showed a significant difference between H. pylori-negative adult and child specimens in terms of α-diversity (p < 0.05), while no significant difference was observed between H. pylori-positive adult and child specimens. There was also a significant difference in ß-diversity between H. pylori-positive and negative child specimens (p < 0.05). Taxonomic analysis at the genus level revealed that Porphyromonas was the only bacterium that was significantly more abundant in both H. pylori-positive adults and children than in corresponding negative specimens (p < 0.01 and p < 0.05, respectively). Conclusion: These results suggest unique oral microbiome characteristics in children with H. pylori infection in the oral cavity.

Sublingual Dermoid Cyst in Young Child.

A dermoid cyst is a benign congenital lesion of ectodermal origin that can arise in any region of the body, though occurrence is rare. A young girl aged 2 years 4 months was referred to our hospital because of a painless mass in the floor of the mouth. Intraoral examination findings revealed a painless movable elastic soft mass on the floor of the mouth measuring approximately 15 mm in diameter. Magnetic resonance imaging indicated a cystic lesion, with low signal intensity shown in T1-weighted and extremely high signal intensity in T2-weighted images. These clinical findings indicated the presence of a dermoid cyst and removal was planned. Under general anesthesia with nasal intubation, surgical removal was performed through an incision on the floor of the mouth. Blunt dissection exposed the integrity of the cyst capsule, which was weakly attached to adjacent tissue. The excised mass was 19 mm × 14 mm × 11 mm in size. Histological examination findings confirmed a diagnosis of dermoid cyst. The operation was successfully completed without any complications and the postoperative course was good. It is important to properly evaluate cysts in children and provide proper treatment with appropriate timing.

Large-Scale Survey of Missing Deciduous Anterior Teeth on Medical Examination at the Age of 3.5 Years.

Tooth anomalies in childhood may negatively affect the healthy development of the dentition and occlusion; hence, it is important to examine the actual oral condition at an early stage. The present study was performed to understand the state of missing deciduous anterior teeth in children aged 3.5 years who underwent dental checkups in Matsubara City. In total, 3508 children received oral examinations, and items such as erupted deciduous teeth and teeth anomalies were recorded. Among these children, those with missing anterior deciduous teeth were selected, and their details were analyzed. In the 216 children, there were 266 missing anterior deciduous teeth. Congenitally missing anterior deciduous teeth were observed in 80 children, and fused teeth were observed in 128 children. The missing teeth were predominantly located in the mandible and occurred more frequently on the right side. The most common reason for acquired missing teeth was trauma, and no cases of spontaneous loss due to systemic disease were found in this study. Screening for various tooth anomalies is expected to play an important role in cultivating a better understanding of the oral cavity of children, developing healthy dentitions, and contributing to the early detection of some systemic diseases.

Detection of Helicobacter pylori from Extracted Teeth of a Patient with Idiopathic Thrombocytopenic Purpura.

Immune thrombocytopenic purpura (ITP) is an autoimmune disease characterized by isolated cryptogenic thrombocytopenia due to a transient or persistent reduction in platelet count. Many patients with ITP have shown improved platelet count after Helicobacter pylori eradication therapy. However, there have been no studies regarding H. pylori in the oral cavity of patients with ITP. Here, we describe a patient with ITP whose oral samples exhibited H. pylori. A 64-year-old woman with ITP came to our hospital with chief complaints that required oral surgery, including tooth extraction and cystectomy. Bacterial DNA from H. pylori was confirmed on the extracted tooth, but was not detected in the saliva taken at the time. Bacterial DNA from H. pylori was detected on the suture around the extraction socket, which was removed at 10 days post-operation. However, H. pylori DNA was not detected in other oral samples at 10 or 30 days post-operation. A urea breath test was carried out in the gastrointestinal clinic at 60 days post-operation, which revealed no presence of H. pylori in the gastrointestinal tract. These results suggest that teeth with severe bacterial infections may be a potential reservoir of H. pylori for patients with ITP.

Effects of Enzyme Replacement Therapy for Primary Teeth in a Patient with Infantile Hypophosphatasia.

Hypophosphatasia (HPP) is a skeletal disorder characterized by hypomineralization of bone, with early exfoliation of primary teeth. Alkaline phosphatase enzyme replacement therapy (ERT) has been shown to improve bone hypomineralization for patients with HPP, although its dental effects are unknown. A 20-month-old Japanese boy diagnosed with infantile HPP was referred to our clinic because of early exfoliation of primary teeth. The patient had been followed by a pediatrician since the age of 3 months, due to slow weight gain. At the age of 12 months, primary incisors showed sudden exfoliation; at the age of 19 months, a diagnosis of HPP was made based on bone and dental manifestations. ERT was initiated at the age of 21 months. The patient demonstrated stable periodontal conditions of primary molars that erupted after initiation of ERT, due to improved alveolar bone and tooth mineralization. Thus, ERT may improve both dental and systemic conditions.

Contribution of Streptococcus mutans to Helicobacter pylori colonisation in oral cavity and gastric tissue.

Helicobacter pylori is presumed to infect gastric tissue via the oral cavity in childhood, whereas risk factors for H. pylori infection in the oral cavity are unknown. In this study, we analysed the effects of Streptococcus mutans, a major cariogenic bacterial species, on H. pylori colonisation in the oral cavity, as well as gastric tissue. Rats in the weaning period were infected with S. mutans in the oral cavity, then fed a caries-inducing diet to facilitate S. mutans colonisation. One month after S. mutans infection, rats were infected with H. pylori in the oral cavity; rats were then euthanised at 1 month after H. pylori infection. H. pylori was detected in the oral cavities of rats infected with both S. mutans and H. pylori, but not in rats infected with H. pylori alone. In addition, H. pylori colonisation in the gastric tissue and typical gastrointestinal damage were observed in rats infected with both S. mutans and H. pylori. When H. pylori was co-cultured with in vitro biofilm formed by S. mutans, a large number of H. pylori bacteria invaded the biofilm formed by S. mutans. Our results suggest that S. mutans is involved in the establishment of H. pylori infection.

Distribution of Helicobacter pylori and Periodontopathic Bacterial Species in the Oral Cavity.

The oral cavity may serve as a reservoir of Helicobacter pylori. However, the factors required for H. pylori colonization are unknown. Here, we analyzed the relationship between the presence of H. pylori in the oral cavity and that of major periodontopathic bacterial species. Nested PCR was performed to detect H. pylori and these bacterial species in specimens of saliva, dental plaque, and dental pulp of 39 subjects. H. pylori was detected in seven dental plaque samples (17.9%), two saliva specimens (5.1%), and one dental pulp (2.6%) specimen. The periodontal pockets around the teeth, from which dental plaque specimens were collected, were significantly deeper in H. pylori-positive than H. pylori-negative subjects (p < 0.05). Furthermore, Porphyromonas gingivalis, a major periodontopathic pathogen, was detected at a significantly higher frequency in H. pylori-positive than in H. pylori-negative dental plaque specimens (p < 0.05). The distribution of genes encoding fimbriae (fimA), involved in the periodontal pathogenicity of P. gingivalis, differed between H. pylori-positive and H. pylori-negative subjects. We conclude that H. pylori can be present in the oral cavity along with specific periodontopathic bacterial species, although its interaction with these bacteria is not clear.

Ankylosed Primary Molar in a Japanese Child with Hypophosphatasia.

Hypophosphatasia (HPP) is a rare genetic disorder; affected patients may experience early exfoliation of primary teeth, especially anterior teeth. However, there have been few reports regarding longitudinal follow-up for primary teeth, especially posterior teeth, until their replacement with permanent teeth. Here, we describe a patient with HPP who underwent follow-up from 1 to 9 years of age. A 14-month-old boy was referred to our hospital with the chief complaint of early loss of primary anterior teeth. He was diagnosed with odonto-type HPP by his pediatrician, due to low serum alkaline phosphatase concentration and early exfoliation of primary teeth with bone hypomineralization. The patient experienced exfoliation of three additional primary anterior teeth by 4 years and 1 month of age. Partial dentures were applied for space maintenance; there were no problems regarding subsequent replacement with permanent teeth in the anterior region. However, the primary mandibular right first molar appeared to be submerged when the patient was 8 years and 3 months of age; the severity of submergence was greater when the patient was 9 years of age. The affected primary molar was considered to be ankylosed; it was extracted when the patient was 9 years and 4 months of age. Histopathological analysis of the tooth revealed disturbed cementum formation, which is a typical characteristic of teeth in patients with HPP. On the basis of these findings, we hypothesize that the disturbed cementum formation could lead to susceptibility to early exfoliation of anterior teeth, as well as occurrence of ankylosis involving posterior teeth.

Association between Helicobacter pylori infection and dental pulp reservoirs in Japanese adults.

BACKGROUND: Helicobacter pylori (H. pylori) colonize the stomach and are considered an etiological agent of gastric cancer. The oral cavity is a transmission route to the stomach, but the exact site of colonization has not yet been explicated. Our study investigated the association between H. pylori infection and presence in oral samples. METHODS: Dental pulp, supragingival plaque, and saliva from 192 patients visiting the Dentistry's outpatient clinic were collected for testing. The H. pylori ureA gene was identified via Nested PCR. Urine anti-H. pylori antibody test was utilized to detect infection. RESULTS: Twenty-five subjects were found to be antibody-positive. PCR analysis of dental pulp revealed that 23 subjects possessed the ureA gene. Twenty-one subjects were positive for both antibodies and genes in dental pulp. PCR testing revealed that 2 subjects were positive in dental plaque but negative for saliva. The subjects positive for H. pylori in dental pulp expressed clinical signs of severe dental caries. CONCLUSIONS: H. pylori infected subjects expressed H. pylori in samples from the oral cavity. The main reservoir for infection within the oral cavity was determined to be dental pulp. Moreover, H. pylori are likely transmitted from dental caries to the root canal.

Potential involvement of Helicobacter pylori from oral specimens in overweight body-mass index.

The bacterium Helicobacter pylori was originally classified in the Campylobacter genus, which contains major periodontopathic bacterial species, and H. pylori DNA has been found in the oral cavity. Although many studies show an association between the presence of periodontal bacteria and an overweight body-mass index (BMI; >25 kg/m2), the relationship between body weight and the presence of H. pylori in the oral cavity has not been demonstrated. Herein, we analysed the relationship between H. pylori in the oral cavity and systemic conditions, including the overweight BMI. Saliva specimens and extracted teeth were obtained from 87 subjects; the distribution of H. pylori among these specimens was analysed with the polymerase chain reaction. Subjects with an overweight BMI exhibited significantly higher detection rates of H. pylori in saliva, compared with non-overweight subjects (BMI <25 kg/m2) (P < 0.05). A clinical history of digestive diseases was not associated with the presence of H. pylori in overweight subjects, whereas subjects with both severe dental caries and an overweight BMI showed a higher detection rate of H. pylori in saliva specimens, compared with other groups. These results suggest that the detection of H. pylori in the oral cavity could be associated with the overweight BMI, which was predominant among subjects with severe dental caries.

Comparison of oral flora before and after triple therapy for Helicobacter pylori eradication in patient with gastric disease.

The oral cavity is recognized as a major route for infection by Helicobacter pylori, which colonizes the gastric mucosa. Therapeutic options for elimination in patients with digestive disease have been established, though whether antibiotics are effective for H. pylori harbored in the oral cavity as well as oral commensal bacteria remain unknown. A 29-year-old woman visited a gastrointestinal clinic with a chief complaint of stomach ache. Gastroscopy and urine test findings for H. pylori IgG led to a definitive diagnosis of goose bumps gastritis induced by bacteria in the stomach, and triple therapy for H. pylori eradication was administered. Oral samples were obtained 10 times at a dental hospital clinic from 1 week before until 6 months after starting triple therapy. Nested PCR amplifying the ureA gene and PCR assays using species-specific primer sets were performed to detect H. pylori and major oral commensal bacterial species, respectively, in those samples. Bacterial DNA encoding the ureA gene of H. pylori in oral specimens was detected prior to starting therapy, which was then reduced during and not detected after finishing treatment. Although the populations of major oral pathogenic bacteria, such as periodontopathic and oral streptococcal species, were drastically reduced during triple therapy, most had recovered within approximately 1 week after ending treatment. These results suggest that a conventional triple therapy approach for eradication of H. pylori in cases of digestive disease is effective for elimination of the bacterium from the oral cavity. On the other hand, after finishing such therapy, the oral flora composition is not drastically changed.

Molecular and clinical analyses of Helicobacter pylori colonization in inflamed dental pulp.

BACKGROUND: Recently, dental pulp has been considered a possible source of infection of Helicobacter pylori (H. pylori) in children. We previously developed a novel PCR system for H. pylori detection with high specificity and sensitivity using primer sets constructed based on the complete genome information for 48 H. pylori strains. This PCR system showed high sensitivity with a detection limit of 1-10 cells when serial dilutions of H. pylori genomic DNA were used as templates. However, the detection limit was lower (102-103 cells) when H. pylori bacterial DNA was detected from inflamed pulp specimens. Thus, we further refined the system using a nested PCR method, which was much more sensitive than the previous single PCR method. In addition, we examined the distribution and virulence of H. pylori in inflamed pulp tissue. METHODS: Nested PCR system was constructed using primer sets designed from the complete genome information of 48 H. pylori strains. The detection limit of the nested PCR system was 1-10 cells using both H. pylori genomic DNA and bacterial DNA isolated from inflamed pulp specimens. Next, distribution of H. pylori was examined using 131 inflamed pulp specimens with the nested PCR system. In addition, association between the detection of H. pylori and clinical information regarding endodontic-infected teeth were investigated. Furthermore, adhesion property of H. pylori strains to human dental fibroblast cells was examined. RESULTS: H. pylori was present in 38.9% of inflamed pulp specimens using the nested PCR system. H. pylori was shown to be predominantly detected in primary teeth rather than permanent teeth. In addition, samplings of the inflamed pulp were performed twice from the same teeth at 1- or 2-week intervals, which revealed that H. pylori was detected in most specimens in both samplings. Furthermore, H. pylori strains showed adhesion property to human dental fibroblast cells. CONCLUSION: Our results suggest that H. pylori colonizes inflamed pulp in approximately 40% of all cases through adhesion to human dental fibroblast cells.

The in vivo Inhibition of Oral Biofilm Accumulation and Streptococcus mutans by Ceramic Water.

Combustion-synthesized titanium carbide ceramics uniformly disperse silver, producing silver ions and hydroxyl radicals in water. This generates antimicrobial activity against various bacteria. One such bacterium is Streptococcus mutans, a gram-positive anaerobic bacterium known as a major pathogen of dental caries. In this study, we analyzed the inhibition of oral biofilms and S. mutans by ceramic water in in vitro and human studies. S. mutans strains showed significantly lower antimicrobial and sucrose-dependent adhesion activity in the presence of ceramic powder compared with untreated culture medium. Confocal microscopy revealed that S. mutans biofilm structures with ceramic powder were thin and coarse. Twenty-seven volunteers (13 males, 14 females; 18-37 years old, mean 25.2 years) were enrolled for subsequent studies. After each meal, one group was asked to rinse with ceramic water while the other rinsed with untreated water for 1 week. After 1 week, the rinsing contents were switched between the groups and the same protocol was followed for an additional week. After rinsing with ceramic water, the average plaque score was 43.0 ± 3.7, which was significantly lower than the baseline value (74.1 ± 5.7, p < 0.001). However, no significant difference was observed when rinsing with untreated water. In addition, the total number of S. mutans in saliva was significantly reduced after rinsing with ceramic water compared with untreated water (p < 0.05). These results suggest that ceramic water possesses antimicrobial activity against S. mutans and inhibits biofilm formation. Rinsing with ceramic water can also inhibit dental plaque formation and S. mutans colonization in humans.

Contribution of the Collagen-Binding Proteins of Streptococcus mutans to Bacterial Colonization of Inflamed Dental Pulp.

Streptococcus mutans is a major pathogen of dental caries. Collagen-binding proteins (CBPs) (approximately 120 kDa), termed Cnm and Cbm, are regarded as important cell surface antigens related to the adherence of S. mutans to collagenous tissue. Furthermore, CBP-positive S. mutans strains are associated with various systemic diseases involving bacteremia, such as infective endocarditis. Endodontic infection is considered to be an important cause of bacteremia, but little is known regarding the presence of S. mutans in dental pulp tissue. In the present study, the distribution and virulence of S. mutans in dental pulp tissues were investigated by focusing on CBPs. Adhesion and invasion properties of various S. mutans strains were analyzed using human dental pulp fibroblasts (HDPFs). CBP-positive strains had a significantly higher rate of adhesion to HDPFs compared with CBP-defective isogenic mutant strains (P<0.001). In addition, CBP-positive strains induced HDPF proliferation, which is a possible mechanism related to development of hyperplastic pulpitis. The distribution of S. mutans strains isolated from infected root canal specimens was then analyzed by PCR. We found that approximately 50% of the root canal specimens were positive for S. mutans. Approximately 20% of these strains were Cnm-positive, while no Cbm-positive strains were isolated. The Cnm-positive strains isolated from the specimens showed adhesion to HDPFs. Our results suggest that CBP-positive S. mutans strains exhibit high colonization in dental pulp. This could be a possible virulence factor for various systemic diseases.

Detection of Helicobacter pylori DNA in inflamed dental pulp specimens from Japanese children and adolescents.

The oral cavity has been implicated as a source of Helicobacter pylori infection in childhood. Various PCR methods have been used to detect H. pylori DNA in oral specimens with various detection rates reported. Such disparity in detection rates complicates the estimation of the true infection rate of H. pylori in the oral cavity. In the present study, we constructed a novel PCR system for H. pylori detection and used it to analyse oral specimens. Firstly, the nucleotide alignments of genes commonly used for H. pylori detection were compared using the complete genome information for 48 strains registered in the GenBank database. Candidate primer sets with an estimated amplification size of approximately 300-400 bp were selected, and the specificity and sensitivity of the detection system using each primer set were evaluated. Five sets of primers targeting ureA were considered appropriate, of which a single primer set was chosen for inclusion in the PCR system. The sensitivity of the system was considered appropriate and its detection limit established as one to ten cells per reaction. The novel PCR system was used to examine H. pylori distribution in oral specimens (40 inflamed pulp tissues, 40 saliva samples) collected from Japanese children, adolescents and young adults. PCR analysis revealed that the detection rate of H. pylori in inflamed pulp was 15 %, whereas no positive reaction was found in any of the saliva specimens. Taken together, our novel PCR system was found to be reliable for detecting H. pylori. The results obtained showed that H. pylori was detected in inflamed pulp but not saliva specimens, indicating that an infected root canal may be a reservoir for H. pylori.