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The procedure of ultra-rapid extraction (PURE) and loop-mediated isothermal amplification for tuberculosis (LAMP-TB) is a simple and rapid manual tuberculosis diagnostic with medium-throughput capability. Because of its simplicity, this method could be useful in resource-limited conditions such as microscopy centers in developing countries. This study was conducted to evaluate the clinical performance of this method in a point-of-care setting. The performance was compared to that of smear microscopy and liquid culture in a hospital laboratory in Haiti, which is considered a representative facility for the implementation of this method. The sensitivity, based on culture-positivity, was 86% (95% confidence interval: 81.3-90.3%) and that based on the smear-negative and culture-positive results was 51% (38.7-63.5%). The specificity based on sample negativity for both smear and culture was 98.4% (96.8-99.2). These results are nearly equivalent to those of a clinical study performed in Japan and are comparable with those of other nucleic acid amplification methods. Thus, approximately 18% more tuberculosis patients could be identified by adding the LAMP-TB method to routine smear microscopy in field settings in Haiti. In addition, it is suggested that local technicians could perform LAMP-TB after only short-term training.

Electrical engineering staff (reorganized to Electrical and Computer engineering in 2004) has taught electrical engineering students for mathematics classes in Wakayama National College of Technology (WNCT) in Japan from 2007. Effects of the mathematics classes by the electrical engineering staff were evaluated by using the results of the mathematics placement tests. The results of the tests are compared with averages of other National Colleges of Technology in Japan and other divisions of WNCT. As a result, an ability of electrical engineering students for mathematics improved dramatically with increasing the ratio of mathematics classes by the electrical engineering staff. It should seem that these good results were achieved by eagerness and strong motivation of the staff with electrical specialty for improving the ability of their division's students. The reasons for the improvement are discussed and the unique class of mathematics for college of technology is also introduced.

Diagnosis and treatment of tuberculosis (TB) will likely navigate a historical turning point in the 2010s with a new management paradigm emerging. However, global control of TB remains a formidable challenge for the decades to come. The estimated case detection rate of TB globally was 66%, and there were 310 000 estimated multidrug-resistant TB (MDR-TB) cases among the 6.2 million TB patients notified in 2011. Although new tools are being introduced for the diagnosis of MDR-TB, there are operational and cost issues related to their use that require urgent attention, so that the poor and vulnerable can benefit. World Health Organization (WHO) estimated that globally, 3.7% of new cases and 20% of previously treated cases have MDR-TB. However, the scale-up of programmatic management of drug-resistant TB is slow, with only 60 000 MDR-TB cases notified to WHO in 2011. The overall proportion of treatment success of MDR-TB notified globally in 2009 was 48%, far below the global target of 75% success rate. Although new tools and drugs have the potential to significantly improve both case detection and treatment outcome, adequate health systems and human resources are needed for rapid uptake and proper implementation to have the impact required to eliminate TB. Hence, the global TB community should broaden its scope, seek intersectoral collaboration and advocate for cost reduction of new tools, while ensuring that the basics of TB control are implemented to reduce the TB burden through the current ‘prevention through case management’ paradigm.

Performance of interferon-γ (IFN-γ) release assays still needs to be improved. The data on the performance of QuantiFERON-TB Gold Plus (QFT-Plus), a new-generation of QFT assay are limited. This study evaluated the diagnostic performance of QFT-Plus, and compared to that of QuantiFERON-TB Gold In-Tube (QFT-GIT). Blood samples were collected from 162 bacteriologically confirmed tuberculosis (TB) patients and 212 Mycobacterium tuberculosis-uninfected volunteers; these samples were then tested with QFT-GIT and QFT-Plus. The IFN-γ concentration of QFT-Plus was lower than that of QFT-GIT in TB patients (p

PURPOSE The BCG immunization has long been performed in Japan. Although the BCG immunization service is the responsibility of the municipality, the manner in which the BCG immunization is delivered differs from municipality to municipality. The purpose of this study was to clarify how the different manner of the BCG immunization delivery systems influenced the BCG immunization coverage. METHODS The study of BCG immunization coverage was conducted in the Tama area located in the western suburbs of Tokyo in 2004. The birth data and the immunization history by the age of 3 years were collected in the three-year-old health check-up from a total of 2,341 children residing in the Tama area. Based on the age at immunization for each child, the BCG immunization coverage was calculated according to the types of the BCG immunization delivery system. The immunization types were defined as follows; the BCG immunization given on the occasion of the mass health check-up (Group 1); the exclusive mass BCG immunization in a monthly service (Group 2); the exclusive mass BCG immunization in a bimonthly service (Group 3); the exclusive mass BCG immunization in services of fewer than every two months (Group 4); and the immunization given on an individual basis by a general practitioner (Group 5). A univariate analysis was performed to examine the relationship between the BCG immunization coverage by the age of 6 months and the difference among the BCG immunization delivery systems, followed by a multivariate regression analysis to adjust for the factors related to the demography, health care services and the socio-economic status of the municipalities. RESULTS Unadjusted odds ratios and adjusted odds ratios for BCG unimmunized children under the age of 6 months by the BCG immunization delivery manner groups were OR 1 reference, adj. OR 1 reference in Group 1; OR 1.42 CI 0.87-2.29, adj. OR 4.01 CI 2.24-7.11 in Group 2; OR 4.96 CI 3.66-6.82, adj. OR 15.59 CI 10.10-24.49 in Group 3;OR 18.60 CI 13.77-25.49, adj. OR 48.17 CI 29.62-79.75 in Group 4; and OR 4.24 CI 2.86-6.31, adj. OR 15.61 CI 9.05-27.26 in Group 5. The univariate analysis and multivariate regression analysis revealed an influence of the BCG immunization delivery manner on the BCG immunization coverage. CONCLUSION The choice of BCG immunization delivery manner is very important to raise the BCG immunization coverage. The BCG immunization given on the occasion of the mass health check-up and the high-frequent immunization service are thought to improve the BCG immunization coverage.

The reported effect of anti-tuberculosis chemotherapy on interferon-gamma (IFN-γ) release in response to specific Mycobacterium tuberculosis antigens has been inconsistent. The objective of this study was therefore to determine the effect of anti-tuberculosis chemotherapy on IFN-γ response to ESAT-6 and CFP-10. QuantiFERON®-TB Gold (QFT-G) was performed, and the IFN-γ response to ESAT-6 and CFP-10 were measured, for 50 people with culture-confirmed tuberculosis prior to initiating treatment and periodically for up to 120 weeks following initiation of said treatment. IFN-γ responses and bacteriological response were compared. The average IFN-γ response to ESAT-6 and CFP-10 and the proportion of QFT-G results that were positive decreased during chemotherapy and for several weeks thereafter, reaching lows at weeks 48 to 56. Furthermore, these measures were lower at 48 weeks for those with bacteriological reversion prior to the second monthly visit than for those with slower reversion. Although it was shown that anti-tuberculosis treatment generally reduced the specific release of IFN-γ, the effect is so variable that it could be used as a monitor of progress of chemotherapy with great care and reservation.

Isoniazid (INH) is a key agent in the treatment of tuberculosis. In Mycobacterium tuberculosis, INH is converted to its active form by KatG, a catalase-peroxidase, and attacks InhA, which is essential for the synthesis of mycolic acids. We sequenced furA-katG and fabG1-inhA in 108 INH-resistant (INH(r) ) and 51 INH-susceptible (INH(s) ) isolates, and found three mutations in the furA-katG intergenic region (Int(g-7a) , Int(a-10c) and Int(g-12a) ) in four of 108 INH(r) isolates (4%), and the furA(c41t) mutation with an amino acid substitution in 18 INH(r) isolates (17%). These mutations were not found in any of 51 INH(s) isolates tested. We reconstructed these mutations in isogenic strains to determine whether they conferred INH resistance. We found that the Int(g-7a) , Int(a-10c) and Int(g-12a) single mutations in the furA-katG intergenic region decreased katG expression and conferred INH resistance. In contrast, the furA(c41t) mutation was not sufficient to confer INH resistance. These results suggested that downregulation of katG is a mechanism of INH resistance in M. tuberculosis and that mutations in the furA-katG intergenic region play a role in this resistance mechanism.

The aim of this study was to establish the importance of detecting fluoroquinolone (FQ) resistance in multidrug resistant (MDR) Mycobacterium tuberculosis, and to show the usefulness of a hybridization-based line probe assay (LiPA) for detecting gyrA mutations. Thirty-three MDR M. tuberculosis isolates were collected from a total of sixty MDR isolates identified in Japan over 6 months during a national surveillance study in 2002. Seventeen MDR isolates were collected by the National Center for Global Health and Medicine in Japan over 6 years from 2003 to 2008. These 50 isolates were examined for FQ susceptibility, and analysed by LiPA and gyrA sequencing. Among them, 22 (44 %) showed FQ resistance. All FQ-resistant isolates had at least one mutation in gyrA. The results of the LiPA were fully consistent with the DNA sequencing results. Given that on the basis of our results almost half of the MDR M. tuberculosis isolates in Japan might have resistance to FQ, it is important to monitor FQ resistance in patients with MDR tuberculosis (TB), as well as with drug-susceptible TB, prior to commencing treatment. For the detection of FQ resistance, LiPA is useful and can rapidly and efficiently assess FQ resistance.

Isoniazid (INH) is an effective first-line antituberculosis drug. KatG, a catalase-peroxidase, converts INH to an active form in Mycobacterium tuberculosis , and katG mutations are major causes of INH resistance. In the present study, we sequenced katG of 108 INH-resistant M. tuberculosis clinical isolates. Consequently, 9 novel KatG mutants with a single-amino-acid substitution were found. All of these mutants had significantly lower INH oxidase activities than the wild type, and each mutant showed various levels of activity. Isolates having mutations with relatively low activities showed high-level INH resistance. On the basis of our results and known mutations associated with INH resistance, we developed a new hybridization-based line probe assay for rapid detection of INH-resistant M. tuberculosis isolates.

The steadily growing epidemic of diabetes mellitus poses a threat for global tuberculosis (TB) control. Previous studies have identified an important association between diabetes mellitus and TB. However, these studies have limitations: very few were carried out in low-income countries, with none in Africa, raising uncertainty about the strength of the diabetes mellitus-TB association in these settings, and many critical questions remain unanswered. An expert meeting was held in November 2009 to discuss where there was sufficient evidence to make firm recommendations about joint management of both diseases, to address research gaps and to develop a research agenda. Ten key research questions were identified, of which 4 were selected as high priority: (i) whether, when and how to screen for TB in patients with diabetes mellitus and vice versa; (ii) the impact of diabetes mellitus and non-diabetes mellitus hyperglycaemia on TB treatment outcomes and deaths, and the development of strategies to improve outcomes; (iii) implementation and evaluation of the tuberculosis 'DOTS' model for diabetes mellitus management; and (iv) the development and evaluation of better point-of-care diagnostic and monitoring tests, including measurements of blood glucose and glycated haemoglobin A(1c) (HbA(1c)) for patients with diabetes mellitus. Implementation of this research agenda will benefit the control of both diseases.

The apathy of bystanders often prevails when instances of bullying, hidden crime and extortion occur in communities such as schools, business work areas, underclass ghettos, prisons and the military. The present study models apathetic behavior as a non-cooperative game and attempts to verify this theory through experiments. Furthermore, our research suggests that the apathy of bystanders generally decrease as the number of citizens in a community decrease. In our experimental cases, if the number of members in a group decreases from 40 members to 20 members, the concerned and helpful behavior of bystanders increases by 21%.

BACKGROUND: To develop a more accurate methodology for diagnosing active tuberculous pleurisy, as well as peritonitis and pericardits of tuberculous origin, we established an antigen-specific interferon gamma (IFN-gamma)-based assay that uses cavity fluid specimens. METHODS: Over a 19-month period, 155 consecutive, nonselected patients with any cavity effusion were evaluated. Study subjects were 28 patients with bacteriologically confirmed active tuberculous serositis and 47 patients with definitive nontuberculous etiology. Culture was performed for 18 h with fluid mononuclear cells in the supernatant of the effusion together with saline or Mycobacterium tuberculosis-specific antigenic peptides, early secretory antigenic target 6 and culture filtrate protein 10. IFN-gamma concentrations in the culture supernatants were measured. RESULTS: In patients with active tuberculous serositis, antigen-specific IFN-gamma responses of cavity fluid samples were significantly higher than those of nontuberculous effusion samples. Area under the receiver operating characteristic (AUROC) curve was significantly greater for cavity fluid IFN-gamma response (AUROC curve, 0.996) than for cavity fluid adenosine deaminase and whole-blood IFN-gamma responses (AUROC curve, 0.882 and 0.719, respectively; P = .037 and P < .001, respectively). Although the AUROC curve was greater for cavity fluid IFN-gamma response than for background cavity fluid IFN-gamma level (AUROC curve, 0.975), the AUROC curves were not statistically significantly different (P = .74). However, multivariate logistic regression analysis revealed that cavity fluid IFN-gamma responses were significantly associated with the diagnosis, even after adjustment for background IFN-gamma level (adjusted odds ratio, 1.21; 95% confidence interval, 1.03-1.42; P < .001). CONCLUSIONS: The cavity fluid IFN-gamma assay could be a method for accurately and promptly diagnosing active tuberculous serositis.

Resistance of Mycobacterium tuberculosis to pyrazinamide (PZA) derives mainly from mutations in the pncA gene. We developed a reverse hybridization-based line probe assay with oligonucleotide probes designed to detect mutations in pncA . The detection of PZA resistance was evaluated in 258 clinical isolates of M. tuberculosis . The sensitivity and specificity of PZA resistance obtained by this new assay were both 100%, consistent with the results of conventional PZA susceptibility testing. This assay can be used with sputa from tuberculosis patients. It appears to be reliable and widely applicable and, given its simplicity and rapid performance, will be a valuable tool for diagnostic use.

We developed a DNA sequencing-based method to detect mutations in the genome of drug-resistant Mycobacterium tuberculosis . Drug resistance in M. tuberculosis is caused by mutations in restricted regions of the genome. Eight genome regions associated with drug resistance, including rpoB for rifampin (RIF), katG and the mabA ( fabG1 )- inhA promoter for isoniazid (INH), embB for ethambutol (EMB), pncA for pyrazinamide (PZA), rpsL and rrs for streptomycin (STR), and gyrA for levofloxacin, were amplified simultaneously by PCR, and the DNA sequences were determined. It took 6.5 h to complete all procedures. Among the 138 clinical isolates tested, 55 were resistant to at least one drug. Thirty-four of 38 INH-resistant isolates (89.5%), 28 of 28 RIF-resistant isolates (100%), 15 of 18 EMB-resistant isolates (83.3%), 18 of 30 STR-resistant isolates (60%), and 17 of 17 PZA-resistant isolates (100%) had mutations related to specific drug resistance. Eighteen of these mutations had not been reported previously. These novel mutations include one in rpoB , eight in katG , one in the mabA-inhA regulatory region, two in embB , five in pncA , and one in rrs. Escherichia coli isolates expressing individually five of the eight katG mutations showed loss of catalase and INH oxidation activities, and isolates carrying any of the five pncA mutations showed no pyrazinamidase activity, indicating that these mutations are associated with INH and PZA resistance, respectively. Our sequencing-based method was also useful for testing sputa from tuberculosis patients and for screening of mutations in Mycobacterium bovis . In conclusion, our new method is useful for rapid detection of multiple-drug-resistant M. tuberculosis and for identifying novel mutations in drug-resistant M. tuberculosis .

Novel lophine dimer (hexaarylbiimidazole, HABI) containing a hydrophilic oligo-ethylene glycol group was prepared as a radical initiator. Absorption of a lophyl radical generated by photo dissociation of the lophine dimer was observed around 600nm by photo irradiation at 254nm. Electron beam (EB) pulse radiolysis of the lophine dimer suggested generation of lophyl radical by EB irradiation similarly, showing that the material is also applicable to electron beam lithography. The lophine dimmer showed excellent stability in developing solution and high resolution due to introduction of hydrophilic substituent.

Objective.To examine the hypothesis that results of the QuantiFERON-TB Gold assay (QFT-G), a whole-blood test for detection of tuberculosis infection, are more significantly related to known risk factors for tuberculosis infection in healthcare workers (HCWs) who have received bacille Calmette-Guerin vaccine than are results of the Mantoux tuberculin skin test (TST).Design.All HCWs (approximately 510) from a 370-bed general hospital in Tokyo where patients with and patients without tuberculosis are treated were invited to participate in the study. All study participants completed a questionnaire about their Mycobacterium tuberculosis infection risk factors as HCWs at the general hospital. They were then tested for LTBI by means of the QFT-G, followed by the TST. Statistical analyses were performed to compare results of each test with M. tuberculosis infection risk factors (age, length of employment in the healthcare industry, history of working with tuberculosis-positive patients in a tuberculosis ward or in the outpatient department of the hospital's tuberculosis clinic for more than 1 year, chest radiograph evidence of healed tuberculosis, history of performing bronchoscope procedures, and job classification), and for TST-positive HCWs, to compare the QFT-G result with the TST induration diameter.Results.A total of 332 HCWs (95% of whom had been vaccinated with BCG) participated in the study, and 33 had positive QFT-G results, suggesting a prevalence of LTBI of 9.9%. Of 304 HCWs who underwent TST, 283 (93.1%) had an induration diameter of 10 mm or more. Multiple logistic regression analysis revealed that positive QFT-G results were significantly associated with age and with a history of working in a tuberculosis ward or an outpatient department of a tuberculosis clinic. TST results were not correlated with any of the tuberculosis infection risk factors we evaluated.Conclusions.Positive QFT-G results were closely associated with the presence of risk factors for LTBI in a hospital setting, suggesting that the QFT-G can detect LTBI in a population composed predominately of BCG vaccinees. Because most HCWs worldwide have been vaccinated with BCG, the QFT-G offers a significant improvement over the TST in tuberculosis screening programs and minimizes un-warranted use of tuberculosis prophylaxis.

The 1951 Tuberculosis Control Law of Japan was amended extensively and has been in effect since April, 2005. The revision of the National Tuberculosis Program (NTP) is to respond to the tremendous changes that have occurred during the last 50 years in tuberculosis epidemiology and in the environment in tuberculosis control implementation. In this review, the main points and framework of the revisions were summarized and the perspective of the development of new technical innovations relevant to each area of the revised TB control legislation is discussed. Also, challenges of Japan's NTP in the recent future are discussed, including the controversies over the proposed abolishment of the Tuberculosis Control Law. 1. IMMUNIZATION: In the revision of NTP, the BCG vaccination of elementary school and junior-high school entrants was discontinued. In order to strengthen the early primary vaccination for infants, the new Law has adopted the direct vaccination scheme omitting tuberculin testing prior to immunization. This program is implemented to young babies, i.e., less than six months old, as defined by the decree. It is a heavy responsibility for the municipalities to ensure the high coverage of immunization when the period of legal vaccination is rather strictly limited practically to the fourth to sixth months after birth. The safe direct vaccination is another new challenge where appropriate management of the Koch's phenomenon or similar reactions should be warranted. 2. CHEMOPROPHYLAXIS: Though unfortunately suspended for some legal reason currently, the expansion and improvement of chemoprophylaxis, or treatment of latent tuberculosis infection, to cover anyone with higher risk of clinical development of TB would have a tremendous effect in Japan, especially since 90% of patients who developed TB were infected tens of years ago. The technical innovations in diagnosis of TB infection such as QuantiFERON will be very helpful. Development of new drugs or drug regimens for this purpose is also expected. 3. CASE DETECTION: The "indiscriminate" screening scheme in the periodic mass health examination has been replaced with a selective one. Only subjects aged 65 or older are eligible for the screening, supplemented with selected occupational groups who are considered to become source of infection, should they develop tuberculosis, such as health-care providers and school teachers. Local autonomies are also responsible for offering screening to the socio-economic high-risk populations, such as homeless people, slum residents, day laborers, and/or workers in small businesses, as decided by the autonomies at their disposal. Another important mode of active case-finding, i.e., contact investigation has been legally enforced so that anybody cannot refuse to be examined by the Health Center. This investigation service will be greatly enhanced by such new technologies as DNA fingerprinting of TB bacilli and a new diagnostic of TB infection. Regarding the clinical service of the symptomatic patients that detect 75% of new cases currently will be improved in its quality by introducing an external quality assurance system of commercial bacteriological laboratory services. 4. TREATMENT AND PATIENT SUPPORT: The revised NTP clearly states the government's responsibility for treating TB patients in close cooperation with a doctor, in the framework of the DOTS Japan version. While the development of new anti-tuberculosis drugs will be realized in the near future, Japan still has to overcome the issues of improper practice of treatment, as well as the government's slow process for approving drugs to be used for multi-drug resistant TB and non-tuberculous mycobacterioses, such as quinolones, macrolides and others. 5. PREFECTURAL TB CONTROL PLAN: In order to respond to the specific issues of tuberculosis problem in the respective prefectures in terms of epidemiology or in available resources, the Law requests every prefecture to develop and implement its own TB control plan. The rather abruptly proposed argument of abolishing Tuberculosis Control Law and merging the NTP to Infectious Disease Control Law threatens us with the government's premature departure from active commitment in the tuberculosis control. This reminds us of the United States' remarkable experience of the tuberculosis resurgence in the late 1980's and early 1990's, after 15 years of abandonment of the governmental efforts during 1970s.

To identify the diseases that correlate with suspended particle concentration in the ambient air, a cross-sectional epidemiological study was conducted using the annual vital statistics and air pollution estimates of 1881 points throughout Japan. The concentration of suspended particulate matters (SPMs) 10 microm or less in diameter were hypothetically converted to PM(2.5) values (converted PM(2.5) or cPM(2.5)) by using a conversion factor obtained from 25 estimates in Japan. Among various causes of death, a significant correlation was observed between both the SPM and cPM(2.5) (SPM/cPM(2.5)) levels and the age-adjusted death rates of ischemic heart disease or hypertensive heart disease in both genders. Correlation was noted with pneumonia, asthma, chronic bronchitis/emphysema, or lung cancer only in females. Unexpectedly, breast, endometrial, and ovarian cancer also showed significant increases in mortality rates related to the SPM/cPM(2.5) level, suggesting a role for suspended particles in the ambient air with or without gaseous component as a possible endocrine-disrupting, estrogenic agent. Multivariate regression analysis of confounding factors, smoking rate, population density, and hormone-related factors revealed consistent significance of SPM/cPM(2.5) in these diseases.

症例は72歳,女性.平成14年5月20日ころから胸部不快感を自覚していた.27日夜中より胸部不快感に加え呼吸困難も出現したため,救急受診された.来院時,血圧60mmHg台のショック状態であり,CPKの上昇(1,751IU/l)および心臓超音波検査にて4/4度の僧帽弁閉鎖不全を認めた.気管内挿管および大動脈内バルーンパンピング駆動下に冠状動脈造影(CAG)を施行し,回旋枝の完全閉塞を認めたため,手術準備を行いながら経皮的冠状動脈形成術(PCI)を行った.ひき続き胸骨正中切開,右側左房切開でアプローチ,僧帽弁は前外側乳頭筋が完全断裂しており,St. Jude Medical®弁(27mm)で僧帽弁置換術(後尖温存)を施行した.術後経過は良好で,術後27日目に退院となり,現在外来にて経過観察中である.乳頭筋断裂の中でも前外側乳頭筋断裂は発生が希であるが,左冠状動脈系の心筋梗塞が原因であることから術前にCAGを行い,できる限り早期の僧帽弁置換術の施行と必要に応じて術前PCIまたは同時冠状動脈バイパス術(CABG)を考慮すべきであると思われた.

In this paper we use a laboratory experiment toexamine the Pivotal mechanism when applied to a binarydecision on a public project of a fixed size. Wefirst point out that the well-known incentivecompatibility of the Pivotal mechanism is true only in aweak sense; There are always strategies otherthan truth-telling that do no worse for a subject thantruth-telling, in any particular set of circumstances.This weakness of the incentive compatibility ofthe Pivotal mechanism makes it difficult for subjects tounderstand that truth-telling is the unique dominantstrategy for the mechanism, unless they havecomprehensive understanding of the payoff structure,with the result that subjects often do not play thedominant strategy. We suggest that this tendency todepart from the dominant strategy can be overcome byproviding subjects with more information about the payoffstructure. We controlled the level of informationprovision in the laboratory and verified that thestrategies used are closer to the dominant strategywhen more information is provided. Under noinformation provision conditions were outcomes poor,but under ``wide'' provision conditions, in which eachsubject experienced a variety of true personalvaluations of the project, departure from thedominant strategy was smaller in magnitude, and under``deep'' provision conditions, in which detailed payofftables were available to each subject, the rate of useof the dominant strategy increased significantly.

A white rot fungus, Coriolus hirsutus, exhibited a strong ability to decolorize melanoidin in cultures not supplemented with nitrogenous nutrients. Addition of peptone to the cultures lowered the ability of the fungus to decolorize melanoidin, but that of inorganic nitrogens (Ns), ammonium and nitrate did not bring about any marked reduction in the ability. These results suggest an inhibitory effect of organic N on melanoidin decolorization. Therefore, for enhancing the decolorization of melanoidin in wastewaters by the fungus, activated sludge pretreatment of the wastewaters was expected to be effective, i.e., activated sludge is capable of converting available organic N into inorganic N. To confirm this, waste sludge heat treatment liquor (HTL), wastewater from a sewage treatment plant, was pretreated with activated sludge. In practice, pretreatment of HTL under appropriate conditions accelerated the fungal decolorization of HTL. In the pretreated HTL, the fungus was shown to produce a high level of manganese-independent peroxidase (MIP). Addition of Mn(II) to the pretreated HTL caused a further increase in the decolorization efficiency of the fungus and a marked increase in the manganese peroxidase (MnP) activity. Consequently, the increases in MIP and MnP activities were considered to play an important role in the enhanced ability of C. hirsutus to decolorize HTL.

Thyroid functions were analyzed before, during and after interferon (IFN) therapy in patients with chronic hepatitis C. According to the results of routine thyroid function tests and measurements of the levels of anti-thyroid autoantibody prior to the therapy, patients were divided into 2 groups; Group A (19 patients) had at least one abnormal finding related to the thyroid, and Group B (40 patients) did not show any abnormality. Five patients (26% ) in Group A and 4 (10%) in Group B showed thyroid dysfunctions which were very clearly reflected by thyrotropin (TSH) measurements. Interestingly, the time of peak TSH elevation in Group A (mean±SD, 4.3±0.8 months) was significantly earlier than that in Group B (6.8±0.8). Most patients in Group B were diagnosed as having destructive thyroiditis. These findings may suggest that the pathogenesis of IFN-induced thyroid dysfunction consists not only of exacerbation of pre-existing thyroid autoimmunity but also of de novo destructive changes even in the intact thyroid before IFN therapy.(Internal Medicine 37: 246-252, 1998)

An opinion survey concerning the management of Graves' hyperthyroidism was conducted among the council members of the Japan Thyroid Association. The selection of 3 major treatments by 90 respondents for their patients was 98.6±4.2% for antithyroid drug (ATD), 7.8±12.6% for partial thyroidectomy and 5.2±8.1% for radioiodide. They expressed a movement away from the past trend of surgery because of postoperative complications and unsatisfactory therapeutic results, and they assumed a further reduction in the future. On the other hand, the frequency of radioiodide treatment was not considered to have decreased greatly, and they expected a slight increase in the future. Of the respondents, 65% suggested that hyperthyroidism should be completely cured even if the patient would fall into hypothyroidism. The major reascns for choosing surgery or radioiodide after ATD were the adverse effects of ATD and the age and social backgrounds of the patients. Large goiter size was the 3rd reason for surgery but was a minimal indicator for radioiodide. As for ATD treatment, none of the respondents reported the routine application of any uniform fixed-time therapy protocol. Japanese Graves' patients were shown to be less responsive to ATD than Caucasian patients. This was assumed to result at least from high iodide intake, and half of them had ordered their patients to restrict iodide intake. Furthermore, 78% of them had treated with a combined therapy of ATD and thyroid hormone. Most of them apply this for selected patients mainly to lower TSH receptor antibody activity, to better control their patients and to reduce the goiter size. All but 8 (9%) did not give T4 (or T3) after the cessation of ATD, and they felt this to be unnecessary, doubtful about the effect, unsuitable or even possible to induce recurrence. The excellent findings reported by Hashizume et al. (N Engl J Med 324: 947-953, 1991) are well known among them. However, most of them did not agree with the efficacy of the protocol to reduce TRAb or to improve the remission rate, and 90% of the respondents did not intend to apply the protocol immediately. In conclusion, the Japanese thyroidologists were shown to highly prefer ATD, and they intended to treat their patients for longer periods of time only by ATD until clinical remission is achieved. The combination therapy is widely used, but most of them do not consider it effective. The therapeutic protocol reported by Hashizume et al. was not accepted widely in Japan.

A constitutively activating mutation encoding Asp578--Gly in transmembrane helix 6 of the lutropin/choriogonadotropin receptor (LHR) is the most common cause of gonadotropin-independent, male-limited precocious puberty. This mutant LHR produces a 4.5-fold increase in basal cAMP when expressed in COS-7 cells. To better understand the normal role of Asp578 in the LHR we studied the effect of seven other amino acid substitutions at this position. No agonist binding or response was detected with the Asp578--Pro mutant. Agonist binding affinity was unaffected by the other substitutions and estimated receptor concentrations ranged from 11 to 184% of wild type. Substitution of Asp578 with Asn, a similarly sized, uncharged residue, did not produce agonist-independent activation. In contrast, replacement with Glu, Ser, or Leu caused 4. 9-5.6-fold stimulation of basal cAMP. Substitution with Tyr (8.5-fold) or Phe (7.5-fold) had a greater activating effect. Only the Tyr, Phe, and Leu mutants showed constitutive activation of the inositol phosphate pathway. Our data suggest that it is the ability of the Asp578 side chain to serve as a properly positioned hydrogen bond acceptor, rather than its negative charge, that is important for stabilizing the inactive state of the LHR. A bulky aromatic side chain at position 578 may further destabilize the inactive receptor conformation.

Although being one of the most thoroughly studied diseases in epidemiology, tuberculosis (TB) is now re-emerging with quite new epidemiological characteristics, such as TB combined with HIV infection, and multiple-drug resistance TB. These new aspects can add very serious impact on the existing health burden of TB, especially its serious inequality between developed and developing parts of the world in demographic, social and economic terms. These epidemiological aspects of TB problem will be illustrated in a stepwise way, using several mathematical models. The application of epidemetric model of TB is also shown for evaluation of some TB control measures. J Epidemiol, 1996 ; 6 : S57-S63.

To evaluate the efficacy of high-dose intravenous steroid pulse followed by oral steroids in the treatment of thyroid-associated ophthalmopathy, we performed clinical assessment and measurement of retroorbital muscle enlargement in 27 patients before and after the therapy, and followed them up longitudinally. The mean duration of follow up is (mean +/- SD) 29.8 +/- 23.8 months (range 4-92). Diplopia disappeared in 10 patients and ameliorated in 11 patients. The degree of proptosis decreased in 15 patients and the fall in visual acuity improved in a third of the patients. The total ophthalmopathy index (OI) decreased from 7.0 +/- 1.9 to 3.0 +/- 1.5. The extraocular muscle enlargement (EME), expressed as the maximal ratio of extraocular muscle thickness to the diameter of the optic nerve, decreased from 2.33 +/- 0.56 to 1.27 +/- 0.26. No major side effects were found in any patient. The improvement in the eye disease was found immediately after the pulse therapy, prior to the start of the following therapy by oral steroids and/or orbital irradiation. Both of OI and EME decreased with time after the therapy and did not get worse after withdrawing oral steroids. The efficacy of the therapy evaluated by degrees of improvement in OI and in EME was significantly greater in females than in males. Although there was a significant positive correlation between initial OI and EME values and initial TBII and TSAb activities, a significant correlation was seen only between the degrees of improvement in EME and changes in TBII activity due to the therapy. The duration of eye disease, thyroid status, treatment with anti-thyroid drug, smoking and experience of previous treatment did not affect the efficacy of the present therapy. We conclude that high-dose intravenous steroid pulse therapy is effective and safe for thyroid-associated ophthalmopathy.

In order to determine what and how many test items are necessary for the initial tests when examining patients with thyroid disorders, a questionnaire was sent to the council members of the Japan Thyroid Association. Thyroid disorders were divided into 4 categories; hyperthyroidism, hypothyroidism, diffuse goiter, and nodular goiter. The questionnaire was designed to assess: 1) tests routinely ordered at the initial examination; 2) minimum test battery necessary; 3) the 5 most important tests in order; and 4) selection of only 2 tests at most. The data on 112 completed questionnaires were collected and analyzed. When the choice was limited to only 2 tests, TSH and fT4 were commonly selected for hyperthyroidism hypothyroidism and diffuse goiter, but these 2 tests would be insufficient for the differentiation of disorders. In the case of nodular goiter, ultrasonogram and fine needle aspiration biopsy (FNA) were selected. The mean number of routinely ordered tests was 6.62 +/- 2.00 (S.D.) for hypothyroidism and 8.06 +/- 2.48 for nodular goiter, but many fewer tests were cited as absolutely necessary. In hyperthyroidism, 4.56 +/- 1.92 tests were required, in hypothyroidism 4.39 +/- 1.92, in diffuse goiter 4.93 +/- 1.75, and in nodular goiter 5.15 +/- 2.13. Most of the function-related tests, ultrasonogram and autoantibodies were commonly selected, while TBII and thyroid 123I or 99mTcO4 uptake in hyperthyroidism, and FNA, Tg and scintigram in nodular goiter were considered to be for specific purposes. Summarizing all these, the authors propose 5 or 6 tests to be necessary as the initial tests when examining patients with 4 individual thyroid disorders.

Cloning of TSHR gene and some subsequent studies using the gene were described. Enormous numbers of studies have been performed since the cloning of TSHR gene. Recent molecular studies on TSH receptor and TSHRAb gave various impacts on thyroidology and are resolving past problems. We mainly focused on regulation, processing and glycosylation, TSH- and TSHRAb binding sites, T cell epitopes, and signal transduction of TSHR. Furthermore, we isolated and characterized TSHRAb genes using lymphocytes producing monoclonal TSHRAb obtained from patients with Graves' disease and primary hypothyroidism. Thus, both antigen and antibody genes are cloned. Combined use of these genes will help to investigate the interactions between TSHR and TSHRAb, and may be expected to contribute to the understanding of molecular mechanisms underlying the pathogenesis of autoimmune thyroid disease as well as the physiology of the thyroid gland.

We treated a patient who was hyperthyroid due to Graves' disease and strongly resistant to methimazole (MMI): in spite of good compliance, she needed 150 mg of MMI daily to control her hyperthyroidism. To elucidate the reasons of resistance to MMI, her serum and intrathyroidal MMI concentrations were determined by high pressure liquid chromatography (HPLC). After taking a 30 mg dose of MMI, she had a similar serum MMI concentration-time curve to that of a normal subject: drug malabsorption and rapid drug metabolism were not evident when studied after surgical treatment. After her serum containing MMI was incubated with Protein G, the MMI concentration of the fraction not bound to Protein G did not change significantly from that of untreated serum: the possibility of anti-MMI IgG antibody production was considered unlikely. Furthermore, the intrathyroidal concentration of MMI in a surgically obtained tissue specimen was 3 micrograms/g wet tissue and appeared to be comparable with those of other Graves' tissues reported. Considering that the patient had been taking 150 mg per day of MMI by the time of thyroidectomy, her intrathyroidal MMI concentration was relatively low, suggesting possible impairment of intrathyroidal MMI accumulation. The possibilities of impaired intrathyroidal actions and the severity of hyperthyroidism, especially high T3 levels, also remained as possible causes. In conclusion, here was a severely hyperthyroid patient who was poorly responsive to conventional doses of MMI, and impairment of thyroid uptake of MMI or of pathways after uptake were considered as possible mechanisms.

We have examined the role of the 2nd cytoplasmic loop of the TSH receptor (TSHR) in TSH- and TSHR autoantibody-stimulated cAMP and inositol phosphate formation using mutants created by substituting sequences from the alpha 1- or beta 2-adrenergic receptors (AR). Unlike similar substitution mutants involving the 3rd cytoplasmic loop that lose agonist-induced inositol phosphate but not cAMP increase after transfection into Cos-7 cells, mutants involving the 2nd loop showed significant change in generating both signals. Mutant B525, which substitutes residues 525-527 with a comparable beta 2-AR sequence, exhibited a complete loss in TSH- or Graves' immunoglobulin G-increased cAMP signaling and a lesser loss in phosphoinositide signaling. This is a unique mutant in which cAMP response was completely lost in all those involving the 2nd or 3rd cytoplasmic loop. On the other hand, mutant B528, in which residues 528-532 are substituted with a comparable beta 2-AR sequence, exhibited the most profound loss in phosphoinositide signaling. Mutants involving portions surrounding residues 528-532 in the 2nd cytoplasmic loop had milder losses in agonist-increased phosphoinositide signaling and much lesser losses in agonist-increased cAMP generation. The transfection efficiency of all transfectants was the same. All transfectants with mutant or wild type TSHR had a similar amount and identical profile of TSHR mRNA in Northern blots and TSHR forms on Western blots. Thus, the 2nd cytoplasmic loop is important for agonist-induced cAMP as well as for phosphoinositide signal generation, whereas the 3rd loop appears to be important only for the latter. The most important determinant for agonist-increased cAMP signal generation is in the middle of the 2nd loop, around residues 525-527. In contrast, the determinants most critical for agonist-induced phosphoinositide signaling are also located in the middle of the 2nd loop, around residues 528-532, and those with less importance are broadly distributed.

We investigated the role of the 1st cytoplasmic loop of the thyrotropin receptor (TSHR) on signal transduction using mutants as we did the 2nd and 3rd cytoplasmic loops [Kosugi S. et al. (1994) Mol. Endocrinol., in press; and 7, 1009-1020]. Five substitution mutants involving the first cytoplasmic loop showed a TSH- or Graves' IgG-stimulated cAMP response despite the low TSH binding Bmax. All the mutants completely lost or markedly decreased the TSH- or Graves' IgG-stimulated inositol phosphate increase. These findings suggest that the l st cytoplasmic loop of the TSHR does not play a crucial role in agonist-induced adenylate cyclase activation but that it is important for phosphoinositide signaling.