Shotgun metagenomic analysis of saliva microbiome suggests Mogibacterium as a factor associated with chronic bacterial osteomyelitis.

Osteomyelitis of the jaw is a severe inflammatory disorder that affects bones, and it is categorized into two main types: chronic bacterial and nonbacterial osteomyelitis. Although previous studies have investigated the association between these diseases and the oral microbiome, the specific taxa associated with each disease remain unknown. In this study, we conducted shotgun metagenome sequencing (≥10 Gb from ≥66,395,670 reads per sample) of bulk DNA extracted from saliva obtained from patients with chronic bacterial osteomyelitis (N = 5) and chronic nonbacterial osteomyelitis (N = 10). We then compared the taxonomic composition of the metagenome in terms of both taxonomic and sequence abundances with that of healthy controls (N = 5). Taxonomic profiling revealed a statistically significant increase in both the taxonomic and sequence abundance of Mogibacterium in cases of chronic bacterial osteomyelitis; however, such enrichment was not observed in chronic nonbacterial osteomyelitis. We also compared a previously reported core saliva microbiome (59 genera) with our data and found that out of the 74 genera detected in this study, 47 (including Mogibacterium) were not included in the previous meta-analysis. Additionally, we analyzed a core-genome tree of Mogibacterium from chronic bacterial osteomyelitis and healthy control samples along with a reference complete genome and found that Mogibacterium from both groups was indistinguishable at the core-genome and pan-genome levels. Although limited by the small sample size, our study provides novel evidence of a significant increase in Mogibacterium abundance in the chronic bacterial osteomyelitis group. Moreover, our study presents a comparative analysis of the taxonomic and sequence abundances of all genera detected using deep salivary shotgun metagenome data. The distinct enrichment of Mogibacterium suggests its potential as a marker to distinguish between patients with chronic nonbacterial osteomyelitis and chronic bacterial osteomyelitis, particularly at the early stages when differences are unclear.

Exploring the genetic diversity of the Japanese population: Insights from a large-scale whole genome sequencing analysis.

The Japanese archipelago is a terminal location for human migration, and the contemporary Japanese people represent a unique population whose genomic diversity has been shaped by multiple migrations from Eurasia. We analyzed the genomic characteristics that define the genetic makeup of the modern Japanese population from a population genetics perspective from the genomic data of 9,287 samples obtained by high-coverage whole-genome sequencing (WGS) by the National Center Biobank Network. The dataset comprised populations from the Ryukyu Islands and other parts of the Japanese archipelago (Hondo). The Hondo population underwent two episodes of population decline during the Jomon period, corresponding to the Late Neolithic, and the Edo period, corresponding to the Early Modern era, while the Ryukyu population experienced a population decline during the shell midden period of the Late Neolithic in this region. Haplotype analysis suggested increased allele frequencies for genes related to alcohol and fatty acid metabolism, which were reported as loci that had experienced positive natural selection. Two genes related to alcohol metabolism were found to be 12,500 years out of phase with the time when they began to increase in the allele frequency; this finding indicates that the genomic diversity of Japanese people has been shaped by events closely related to agriculture and food production.

Role of Macrophage Colony-Stimulating Factor for Staphylococcal Infection in the Oral Cavity.

OBJECTIVE: There are few valid indicators of oral infection owing to the complexity of pathogenic factors in oral diseases. Salivary markers are very useful for scrutinizing the symptoms of disease. To provide a reliable and useful predictive indicator of infection for opportunistic pathogens in individuals with compromised immune systems, such as those with periodontal diseases and Human Immunodeficiency Virus (HIV), this study examines opportunistic pathogens such as C. albicans and staphylococci and macrophage colony-stimulating factor (M-CSF) and CA125/MUC16 in saliva. The aim was to explore the correlations investigated among these factors. METHODS: Samples were divided into two groups (based on patient sex, the absence and presence of dentures in elderly, or HIV-positive patients and healthy subjects), and the correlation was analyzed in two groups of elderly patients with periodontal disease (64.5 ± 11.2 years old) and HIV-infected patients (41.9 ± 8.4 years old). Healthy subjects (33.8 ± 9.1 years old) were also analyzed as a control. Levels of C. albicans, staphylococci, and M-CSF, which is an immunological factor for the differentiation of macrophage, and CA125/MUC16, which provides a protective lubricating barrier against infection, were investigated. RESULTS: A significant and positive correlation between the levels of M-CSF and staphylococci was found in elderly individuals and HIV-positive patients treated with antiretroviral therapy. A significant and positive correlation between the levels of M-CSF and CD125/MUC16 was also found in both patients. These correlations were enhanced in both patients as compared with healthy subjects. CONCLUSION: Salivary M-CSF might be useful as a new indicator of opportunistic infection caused by staphylococci and a defense against infection in immunocompromised hosts.

Association between an Increased Serum CCL5 Level and Pathophysiology of Degenerative Joint Disease in the Temporomandibular Joint in Females.

Degenerative joint disease of the temporomandibular joints (DJD-TMJ) clinically manifests with symptoms such as orofacial pain, joint sounds and limited jaw movements. Our research group previously reported the functional necessity of a chemokine-chemokine receptor axis of CCL5-CCR5 in osteoclasts. Accumulated studies reported that this axis was involved in the pathogenesis of bone and joint destructive diseases, suggesting CCL5 as a potent biomarker. This study investigated whether or not the serum level of CCL5 can be a biomarker of DJD-TMJ and concomitantly analyzed changes in the serum and urine levels of bone markers to see whether or not changes in the rate of bone metabolism were predisposing. We enrolled 17 female subjects with diagnosed DJD-TMJ and sexually and age-matched 17 controls. The serum CCL5 level in DJD-TMJ subjects was significantly higher than that in the control subjects. Multivariate analyses indicated an association between an augmented CCL5 level and the rate of bone metabolism, especially in relatively young DJD-TMJ subjects without other systemic symptoms. A principal component analysis of serum markers and our pharmacological experiment using a postmenopausal model of ovariectomized rats suggested that an augmented serum CCL5 level specifically reflected DJD-TMJ and that covert changes in the rate of bone metabolism predisposed individuals to DJD-TMJ.

Effects of Perioperative Oral Management in Patients with Cancer.

Perioperative oral management (POM) is used to prevent pneumonia in patients with cancer. However, the factors that expose hospitalized patients to increased risk of developing pneumonia remain unclear. For example, no study to date has compared the incidence of pneumonia in hospitalized patients by cancer primary lesion, or POM implementation, or not. We determined which patients were most likely to benefit from POM and examined the effects of POM on pneumonia prevention and mortality. In a total of 9441 patients with cancer who underwent surgery during hospitalization, there were 8208 patients in the No POM group, and 1233 in the POM group. We examined between-group differences in the incidence of pneumonia and associated outcomes during hospitalization. There was no significant between-group difference in the incidence of pneumonitis, however, patients with lung, or head and neck cancers, demonstrated a lower incidence of postoperative pneumonia. Among patients with lung and pancreatic cancers, mortality was significantly lower in the POM group. POM appears effective at reducing the risk of postoperative pneumonia in patients with certain cancers. Further, mortality was significantly lower in patients with lung and pancreatic cancers who received POM; hence, POM may be an effective adjuvant therapy for patients with cancer.

Whole blood transcriptome profiling identifies gene expression subnetworks and a key gene characteristic of the rare type of osteomyelitis.

Chronic non-bacterial osteomyelitis (CNO) is a rare and severe inflammatory bone disorder that can occur in the jaw. It is often associated with systemic conditions including autoimmune deficiency. Medical management of patients and establishment of a correct diagnosis are difficult as the etiology of the disease remains unknown. Therefore, little is known about the disease characteristics at the gene expression level. Here, we explored aspects of CNO based on whole blood RNA sequencing (>6 Gb per sample) of 11 patients and 9 healthy controls in Japan and on a recently developed method that is applicable to small datasets, can estimate a directed gene network, and extract a subnetwork of genes underlying patient characteristics. We identified nine subnetworks, comprising 26 differentially regulated edges and 36 genes, with the gene encoding glycophorin C (GYPC) presenting the highest discrimination ability. The expression of the gene was mostly lower in patients with CNO than in the healthy controls, suggesting an abnormal status of red cells in patients with CNO. This study enhances our understanding of CNO at the transcriptome level and further provides a framework for whole blood RNA sequencing and analysis of data obtained for a better diagnosis of the disease.

Physical oral care prevents ventilator-associated pneumonia in Vietnam: A prospective interventional study.

BACKGROUND: Ventilator-associated pneumonia (VAP) has emerged as a critical issue in the intensive care unit (ICU) because of its high burden on patients and medical staff. Here, we examined the potential for reducing VAP incidence through physical oral care interventions without any medication. METHODS: This prospective interventional study compared VAP incidence during an 8-month baseline period (usual oral care) and a 9-month intervention period (physical oral care with sponge brush) among patients who received mechanical ventilation for >48 h in a tertiary care hospital in Vietnam from 2017 to 2019. Physical oral care was provided by general ICU nurses who had been trained by dentists and infection control nurses. VAP was diagnosed using the Clinical Pulmonary Infection Score. RESULTS: In total, 423 patients were enrolled in the baseline group and 454 patients were enrolled in the intervention group; 303 and 300 patients, respectively, were included in the analysis. Two hundred thirty-eight VAP episodes were identified: 135 (44.6%) during the baseline period and 103 (34.3%) during the intervention period. Univariate analysis revealed significant reduction of VAP occurrence in the intervention period (odds ratio = 0.65; 95% confidence interval = 0.47-0.90; P = 0.010). The incidences of VAP per 1000 ventilator-days were 63.4 (135/2128) during the baseline period and 48.4 (103/2128) during the intervention period (P = 0.038). CONCLUSIONS: Physical oral care without any medication (e.g., chlorhexidine) reduced VAP incidence in the ICU. This method could be used to reduce VAP incidence, particularly in countries with limited medical resources.

Mental foramen in panoramic radiography can be a reference for discrimination of punched-out lesions in the mandible in patients with symptomatic multiple myeloma: A cross-sectional study.

Multiple myeloma (MM) is a hematopoietic malignancy characterized by monoclonal proliferation of plasma cells. MM features bony radiolucencies called punched-out lesions (POLs), which require appropriate diagnosis due to increased risk of surgically-related adverse events. Although dental surgeons can identify dental focal infections (DFIs) in MM patients, the prevalence and characteristics of POLs in the jawbone of MM patients have not been investigated. We examined the prevalence of POLs in the mandible of MM patients, evaluated its relationship with MM International Staging System progression, and examined panoramic radiographs as a diagnostic reference for POLs in a single center in Japan. We identified 98 patients (55 men, 43 women) with a median age of 63 (range, 34 to 91) years. Of these, 18 patients (18.4%) had POLs in the mandible, including two patients in stage I (2/37; 5.4%), six in stage II (6/43; 14.0%), and ten in stage III (10/18; 55.6%). The prevalence of POLs significantly increased with MM stage progression (p < 0.0001). POLs confirmed on computed tomography (CT) were also detected on panoramic radiographs. The Hounsfield unit value at the site of POLs was nearly the same or lower than that of the mental foramen. Although the prevalence of POLs in the mandible is low, dental surgeons need to differentiate POLs as radiological findings when examining DFIs in MM patients. Confirmation of POLs in the mandible is possible by CT and panoramic radiography, and the mental foramen is likely to be a reference for discrimination.

What does oral care mean to society?

Oral care is defined in a narrow sense as cleaning of the teeth, oral cavity, and dentures, and in a broad sense as the maintenance of oral functions (feeding, swallowing, chewing, speech, aesthetics, etc.), dental treatment, feeding and swallowing training, and articulation training. In the past, it was recognized as simply cleaning the mouth, but the concept of oral care has gradually expanded, and many studies and surveys have been conducted in cooperation with various other professions. As a result, oral health care is involved not only in the prevention of pneumonia, but also in the onset and suppression for severity of diabetes, cardiovascular diseases, some malignant tumors, cerebrovascular diseases, rheumatoid arthritis, dementia, etc. It is also a powerful supportive therapy in cancer treatment. In the terminal stages of life, oral health care can help people to maintain their dignity by continuing to consume food orally until the end of their lives, and in times of disaster, oral health care has been found to be as important as attention to deep vein thrombosis. It has also been found to be effective in preventing severe diseases such as COVID-19. And, although it has not been discussed much, it has been found to have medical and economic benefits such as reducing the duration of hospitalization and treatment costs. This article reviews the results of research to date.

Dispersion of Aerosols Generated during Dental Therapy.

The novel coronavirus pandemic has resulted in an urgent need to study the risk of infection from aerosols generated during dental care and to conduct a review of infection controls. However, existing studies on aerosol particles related to dental treatment have mainly evaluated only the scattering range. Few studies have been conducted on the specifics of the generation of aerosol particles in clinical settings, their mechanisms and patterns of distribution throughout open or enclosed spaces, the duration that they remain suspended in air, and the amount and size of particles present. To minimize the influence of background particles, laser lights, a high-sensitivity camera, and particle counters were used in a large super clean laboratory to investigate the dynamics of aerosols generated during the operation of dental micromotors. The results indicate that aerosols tend to scatter upward immediately after generation and then gradually disperse into the surroundings. Most of the particles are less than 5 µm in size (only a few are larger), and all particles are widely distributed over the long term. Our research clearly elucidates that aerosols produced in dental care are distributed over a wide area and remain suspended for a considerable time in dental clinics before settling.

Preventive Effects of Sustainable and Developmental Perioperative Oral Management Using the "Oral Triage" System on Postoperative Pneumonia after Cancer Surgery.

Perioperative oral management is widely recognized in the healthcare system of Japan. Conventionally, the surgeon refers patients with oral problems to a dental or oral surgery clinic in the hospital. However, frequent in-house referrals were found to increase the number of incoming patients resulting in unsustainable situations due to an insufficient workforce. In 2011, the Center for Perioperative Medicine was established at our hospital to function as a management gateway for patients scheduled to undergo surgery under general anesthesia. The "oral triage" system, wherein a dental hygienist conducts an oral screening to select patients who need preoperative oral hygiene and functional management, was established in 2012. A total of 37,557 patients who underwent surgery at our hospital from April 2010 to March 2019 (two years before and seven years after introducing the system) were evaluated in this study. The sustainability and effectiveness of introducing the system were examined in 7715 cancer surgery patients. An oral management intervention rate of 20% and a significant decrease in the incidence of postoperative pneumonia (aOR = 0.50, p = 0.03) indicated that this system could be useful as a sustainable and developmental oral management strategy to manage surgical patients with minimal human resources.

Cost-Effectiveness Analysis of Perioperative Oral Management after Cancer Surgery and an Examination of the Reduction in Medical Costs Thereafter: A Multicenter Study.

In April 2012, perioperative oral management (POM) was approved for inclusion in the national health insurance system of Japan to prevent the occurrence of pneumonia, a major complication in cancer patients. The subsequent decrease in the incidence of postoperative pneumonia indicated the prophylactic effect of POM. The constant increase in health expenditure necessitates a cost-effectiveness analysis. In addition, the effect of reducing healthcare costs owing to health technologies must be evaluated. In the present multi-institutional study, the cost-effectiveness analysis of POM was conducted by comparing the incidence of postoperative pneumonia and the healthcare costs between patients who received surgery for malignant tumors before (n = 11,886) and after (n = 13,668) the introduction of POM. Additionally, the effect of reducing healthcare costs was evaluated. Reductions in the number of patients who developed pneumonia, duration of hospitalization, and number of deaths were observed after the introduction of POM. The incremental cost-effectiveness ratio was 111,927 yen, hence the prevention of postoperative pneumonia needs 111,927 yen per patient in healthcare costs. Consequently, a maximum reduction of 250,368,129 yen in healthcare costs was observed between the incremental costs for pneumonia treatment and the cost of POM. These findings indicate that improvements in cost-effectiveness can be expected in the future through the development of procedure and system for POM.

Risk Factors for Post-Tooth Extraction Complications in HIV-Infected Patients: a Retrospective Study.

We aimed to assess the rate and risk factors of postoperative complications following tooth extraction in HIV-infected patients by CD4 count. The study participants were 231 HIV-infected patients who underwent tooth extraction at our institution between January 2007 and December 2011. Blood test results, underlying diseases, surgical site, extraction method, and postoperative complication data were obtained from medical records. Potential risk factors of postoperative complications were analyzed using multivariate logistic regression. Patients were divided into two groups: 61 (26%) patients with a CD4 count < 200/µL, and 170 (74%) with a CD4 count ≥ 200/µL. Of the 231 patients, 12 (5.2%) developed postoperative complications (alveolar osteitis, n = 10; surgical site infection, n = 2). The rate of complications did not differ between the CD4 < 200/µL group (1.6%) and the CD4 ≥200/µL group (6.5%) (adjusted odds ratio [aOR]: 9.328, 95% confidence interval [CI]: 0.470, 185.229; P = 0.1431). Surgical extraction with bone excavation, but without CD4 count, was identified as a risk factor for post-extraction complications (aOR: 22.037, 95% CI: 1.519, 319.617; P = 0.0234). A low CD4 count is not a risk factor for post-extraction complications in patients with HIV infection. We conclude that tooth extraction should be performed based on dental/oral conditions, and not delayed until CD4 count improvement.

Survival case of rhinocerebral and pulmonary mucormycosis due to Cunninghamella bertholletiae during chemotherapy for acute myeloid leukemia: a case report.

A 42-year-old man diagnosed with acute myeloid leukemia complained of progressive swelling of the right side of his face with pain 11 days after the third cycle of consolidation therapy with high-dose arabinosylcytosine-cytarabine. Head and neck magnetic resonance imaging showed a mass lesion in his right maxillary sinus with parapharyngeal involvement, which included the right masseter muscle, intraorbital involvement, and an abscess in his brain. Chest computed tomography revealed peribronchial small nodules in his right upper lobe and a necrotic tumor in his right lower lobe. Molds identified as Cunninghamella bertholletiae were isolated from the necrotic ulcer. According to these results, chemotherapy for leukemia was discontinued. High-dose liposomal amphotericin (10 mg/kg/day) was initiated. Because renal dysfunction occurred, the dosage was decreased to 6 mg/kg and combined with 150 mg/day micafungin. Debridement of necrotic tissue in the right maxillary sinus and establishment of the fenestration between the sinus and oral cavity were performed. Subsequently, brain and lung lesions were surgically removed. Rhinocerebral mucormycosis was successfully treated without relapse over 3 years by a 112-day course of intravenous anti-fungal therapy and 223-day course of terbinafine and partial surgical removal, respectively, to maintain masticatory and ocular functions. To our knowledge, there has been no other report of a long-term survival case of rhinocerebral mucormycosis due to C. bertholletiae.

Screening for dental focal infections in febrile patients with hematologic malignancies who received chemotherapy: a retrospective cohort study.

Source of fever in chemotherapy patients is often unknown. Fever can also be fatal. No observational studies have determined the incidence of dental focal infection (DFI)-associated fever, despite oral cavity being a potential source of infection. We report the incidence of fever after chemotherapy in patients with hematological malignancies and their association with DFIs in 441 patients visiting our institution during a 6-year period. Dental treatments, including tooth extraction, were performed, and their oral and hematological profiles were monitored after chemotherapy. Fever was evident in 87 (38.5%) of 226 patients (≥ 38˚C) after the first cycle of chemotherapy. Sepsis due to DFIs (n = 4; 4.6%) was evaluated. Chemotherapy was delayed due to DFI in one case. Fever after chemotherapy should be differentiated from oral infections. Our study emphasizes the significance of DFI in patients with fever after chemotherapy and can help in improving the prognosis of patients.

Shotgun metagenome sequencing identification of a set of genes encoded by Actinomyces associated with medication-related osteonecrosis of the jaw.

Medication-related osteonecrosis of the jaw (MRONJ) is intractable and severely affects a patient's quality of life. Although many cases of MRONJ have been reported in the past decade, the disease pathophysiology is unclear and there are no evidence-based therapeutic strategies. MRONJ usually features bone inflammation and infection. Prior studies that explored the association between MRONJ and microbial infection used the culture-based approach, which is not applicable to hundreds of unculturable taxa in the human oral microbiome, or 16S ribosomal RNA gene sequencing, which does not provide quantitative information of the abundance of specific taxa, and information of the presence, abundance, and function of specific genes in the microbiome. Here, deep shotgun metagenome sequencing (>10 Gb per sample) of bulk DNA extracted from saliva of MRONJ patients and healthy controls was performed to overcome these limitations. Comparative quantitative analyses of taxonomic and functional composition of these deep metagenomes (initially of 5 patients and 5 healthy controls) revealed an average 10.1% increase of genus Actinomyces and a 33.2% decrease in genus Streptococcus normally predominant in the human oral microbiota. Pan-genome analysis identified genes present exclusively in the MRONJ samples. Further analysis of the reads mapping to the genes in the extended dataset comprising five additional MRONJ samples and publicly available dataset of nine healthy controls resulted in the identification of 31 genes significantly associated with MRONJ. All these genes were encoded by Actinomyces genomic regions. Of these, the top two abundant genes were almost exclusively encoded by Actinomyces among usual taxa in the human oral microbiota. The potential relationships of these key genes with the disease are discussed at molecular level based on the literature. Although the sample size was small, this study will aid future studies to verify the data and characterize these genes in vitro and in vivo to understand the disease mechanisms, develop molecular targeted drugs, and for early stage screening and prognosis prediction.

Pneumonia prevention effects of perioperative oral management in approximately 25,000 patients following cancer surgery.

AIM: We conducted a multicenter study to explore the risk factors of developing pneumonia and the effectiveness of perioperative oral management (POM) for the prevention of pneumonia in postsurgical patients. METHODS AND RESULTS: A survey covering eight regional hospitals was conducted over 4 years, from April 2010 to March 2014. Using the Diagnosis Procedure Combination database, a target group of 25,554 patients with cancer who underwent surgery was selected and assessed from a population of 346,563 patients without pneumonia on admission (sample population). The study compared the incidence of pneumonia and attempted to identify the significant predictive factors for its occurrence in these patients using multiple logistic regression analysis. Comparative assessment for the occurrence of pneumonia before and after POM implementation showed a significant incidence decrease after POM introduction in the target group, with no such change observed in the sample population. Multiple logistic regression analysis showed that the odds ratio for pneumonia occurrence after POM introduction was 0.44, indicating a reduced risk of pneumonia. CONCLUSION: POM in cancer patients was indeed effective in reducing the incidence of pneumonia in hospitals and thereby helped in preventing pneumonia during hospitalization.

Retrospective Study on Laser Treatment of Oral Vascular Lesions Using the "Leopard Technique": The Multiple Spot Irradiation Technique with a Single-Pulsed Wave.

OBJECTIVE: This study aimed to retrospectively evaluate the efficacy and safety of laser treatment of oral vascular lesions using the multiple spot irradiation technique with a single-pulsed wave. BACKGROUND DATA: In laser therapy for vascular lesions, heat accumulation induced by excessive irradiation can cause adverse events postoperatively, including ulcer formation, resultant scarring, and severe pain. To prevent heat accumulation and side effects, we have applied a multiple pulsed spot irradiation technique, the so-called "leopard technique" (LT) to oral vascular lesions. This approach was originally proposed for laser treatment of nevi. It can avoid thermal concentration at the same spot and spare the epithelium, which promotes smooth healing. The goal of the study was to evaluate this procedure and treatment outcomes. PATIENTS AND METHODS: The subjects were 46 patients with 47 oral vascular lesions treated with the LT using a Nd:YAG laser (1064 nm), including 24 thick lesions treated using a combination of the LT and intralesional photocoagulation. RESULTS: All treatment outcomes were satisfactory without serious complications such as deep ulcer formation, scarring, bleeding, or severe swelling. CONCLUSIONS: Laser therapy with the LT is a promising less-invasive treatment for oral vascular lesions.

Non-Syndromic Familial Keratocystic Odontogenic Tumour: A Rare Case Report in Japanese Identical Twins.

Keratocystic Odontogenic Tumour (KCOT) is unicystic or multicystic intraosseous benign tumour of odontogenic origin that recurs due to locally destructive behaviour. KCOTs are usually the first manifestation of Nevoid Basal Cell Carcinoma Syndrome (NBCCS), an autosomal dominant disorder also known as Gorlin's syndrome and they are most frequently observed familial symptom regardless of patients' nationality. In addition, the recurrence rate and multiplicity of KCOTs is relatively high as compared to that of other sporadic carcinomas. KCOT has been considered as a non-hereditary lesion and its familial onset is an extremely rare event in non-NBCCS cases. Here, we describe previously unreported non-syndromic multiple KCOT cases in identical twins in a Japanese family. The subjects were female Japanese identical twins who were 26 and 27 years old, respectively, at the time of diagnosis for KCOT. They had no major or minor features of NBCCS other than KCOT. Although there were lesions that were likely to be dentigerous cysts based on radiographic findings, one of them was KCOT. This case report highlights the importance of precise diagnosis, choice of surgical method and careful observation for multiplicity or familial onset in sporadic KCOT cases without NBCCS.