Approval status of essential therapeutic drugs for systemic sclerosis versus that of drugs for rheumatoid arthritis.

Objective: Systemic sclerosis, a rare disease characterized by chronic multisystem fibrosis, requires lifelong management, necessitating enough insurance coverage for the patient. Official drug approval is the first step to ensuring that the drug is covered by insurance. In this study, we investigated the approval status of essential therapeutic drugs for systemic sclerosis across eight countries and compared it with that of drugs for rheumatoid arthritis. Methods: The essential therapeutic drug lists for systemic sclerosis and rheumatoid arthritis were taken from the guidelines of the American College of Rheumatology and the European Alliance of Associations for Rheumatology. Official drug approval status for the selected drugs was confirmed by searching representative Internet databases from eight countries: the United States, the United Kingdom, Germany, France, Italy, Switzerland, Japan, and the Republic of Korea. Results: A total of 21 and 16 drugs were selected for systemic sclerosis and rheumatoid arthritis, respectively. The drug approval rates of the 21 drugs for systemic sclerosis varied among countries. Most drugs used to treat pulmonary arterial hypertension, which were developed recently and are expensive, are approved by most countries; however, most older drugs-which are still essential for management of Raynaud's phenomenon, digital ulcers, interstitial lung disease, and skin fibrosis-are not approved by most countries. By contrast, almost all of the 16 drugs used to treat rheumatoid arthritis, whether old or new, are approved by most countries. Conclusion: Approval rates for drugs used to treat systemic sclerosis, a rare disease, are much lower than those for drugs used to treat rheumatoid arthritis. Thus, approval rates of essential therapeutic drugs for systemic sclerosis need to improve, which will benefit patients by increasing the number of drugs covered by insurance.

An epidemic of cataract surgery in Korea: the effects of private health insurance on the National Health Insurance Service.

OBJECTIVES: In Korea, the National Health Insurance Service (NHIS) covers essential healthcare expenses, including cataract surgery. To address concerns that private health insurance (PHI) might have inflated the need for such procedures, we investigated the extent of the PHI-attributable increase in cataract surgery and its impact on NHIS-reimbursed expenses. METHODS: This retrospective, observational study uses nationwide claims data for cataract surgery from 2016 to 2020. We examined trends in utilization and cost, and we estimated the excess numbers of (1) cataract operations attributable to PHI and (2) types of intraocular lenses used for cataract surgery in 2020. RESULTS: Between 2016 and 2020, a 36.8% increase occurred in the number of cataract operations, with increases of 63.5% and 731.8% in the total healthcare costs reimbursed by NHIS and PHI, respectively. Over a 5-year period, the surgical rate per 100,000 people doubled for patients aged <65 years (from 328 in 2016 to 664 in 2020). Among the 619,771 cases in 2020 of cataract surgery reimbursed by the Korean diagnosis-related group system, more non-NHIS-covered intraocular lenses were used for patients aged <65 years than ≥65 years (68.1 vs. 14.2%). In 2020 alone, an estimated 129,311 excess operations occurred, accounting for an excess cost of US$115 million. CONCLUSIONS: A dramatic increase in the number and cost of cataract operations has occurred over the last 5 years. The PHI-related increase in operations resulted in increased costs to NHIS. Measures to curtail the non-indicated use of cataract surgery should be implemented regarding PHI.

Health Inequities in Cancer Incidence According to Economic Status and Regions Are Still Existed Even under Universal Health Coverage System in Korea: A Nationwide Population Based Study Using the National Health Insurance Database.

PURPOSE: The purpose of this study is to determine the level of health equity in relation to cancer incidence. MATERIALS AND METHODS: We used the National Health Insurance claims data of the National Health Insurance Service between 2005 and 2022 and annual health insurance and medical aid beneficiaries between 2011 and 2021 to investigate the disparities of cancer incidence. We calculated age-sex standardized cancer incidence rates by cancer and year according to the type of insurance and the trend over time using the annual percentage change. We also compared the hospital type of the first diagnosis by cancer type and year and cancer incidence rates by cancer type and region in 2021 according to the type of insurance. RESULTS: The total cancer incidence increased from 255,971 in 2011 to 325,772 cases in 2021. The absolute difference of total cancer incidence rate between the NHI beneficiaries and the medical aid (MA) recipients increased from 510.1 cases per 100,000 population to 536.9 cases per 100,000 population. The odds ratio of total cancer incidence for the MA recipients increased from 1.79 (95% confidence interval [CI], 1.77 to 1.82) to 1.90 (95% CI, 1.88 to 1.93). Disparities in access to hospitals and regional cancer incidence were profound. CONCLUSION: This study examined health inequities in relation to cancer incidence over the last decade. Cancer incidence was higher in the MA recipients, and the gap was widening. We also found that regional differences in cancer incidence still exist and are getting worse. Investigating these disparities between the NHI beneficiaries and the MA recipients is crucial for implementing of public health policies to reduce health inequities.

Incidence, Severity, and Mortality of Influenza During 2010-2020 in Korea: A Nationwide Study Based on the Population-Based National Health Insurance Service Database.

BACKGROUND: The epidemiology of influenza is commonly used to understand and establish relevant health policies for emerging respiratory infections, including coronavirus disease 2019 (COVID-19). However, Korea has no confirmed nationwide data on influenza incidence, severity, and mortality rate. METHODS: We conducted a cross-sectional study to obtain epidemic data on influenza at the national level using National Health Insurance claims data during 2010 to 2020. Influenza cases were defined as 90-day timeframe episodes based on all inpatient and outpatient claims data with disease code J09, J10, and J11. Influenza incidence, severity, and mortality rate were calculated, and logistic regressions were performed to assess the associations of demographic characteristics and comorbidity with influenza-related hospitalization, severe illness, and death. RESULTS: There were 0.4-5.9% influenza cases in the population from 2010 to 2020, with 9.7-18.9%, 0.2-0.9%, and 0.03-0.08% hospitalized, used in the intensive care unit, and dead, respectively. Age-standardized incidence and mortality rates were 424.3-6847.4 and 0.2-1.9 per 100,000 population, respectively. While more than half of the influenza cases occurred in populations aged younger than 20 years, deaths in older than 60 years accounted for more than two-thirds of all deaths. CONCLUSION: This study provided the simplest but most important statistics regarding Korean influenza epidemics as a reference. These can be used to understand and manage other new acute respiratory diseases, including COVID-19, and establish influenza-related policies.

The impact of the COVID-19 pandemic in the healthcare utilization in Korea: Analysis of a nationwide survey.

BACKGROUND: COVID-19 has brought changes in daily life and increased the medical burden. This study aims to evaluate the delays in healthcare services and related factors in the general population during the COVID-19 pandemic. METHODS: We took a nationally representative sample and conducted a mobile phone-based survey. The study was conducted anonymously. Of the 3377 subjects who consented to participate, a total of 2097 finished the survey. The primary outcome was respondents' experiences with delayed (1) health screenings, (2) non-urgent medical visits, (3) medical visits for chronic disease, and (4) emergency visits during the COVID-19 pandemic. RESULTS: Of 2097 respondents, females, residents of the Seoul metropolitan area, those with private insurance, those without chronic diseases, smokers, and drinkers had higher risk of delays in health screening and non-urgent medical visits after adjustment. Among chronic disease patients, those who were over 60 years old (adjusted odds ratio 0.36, 95% CI 0.14-0.92) showed lower risk of delayed medical visit. Residents of the Seoul metropolitan area, those with private insurance, smokers, and drinkers were all associated with experiencing delayed health screening and non-urgent medical visits had higher risk of delays in chronic disease visits and emergent medical visits. CONCLUSIONS: Delayed access to healthcare services is associated with poor outcomes and may cause different complications. Efforts are needed to prevent delays in medical use due to infectious diseases such as COVID-19. Considering the possibility of the emergence of infectious diseases, various countermeasures are needed to prevent delays in medical visit.

Interim outcomes of delamanid for the treatment of MDR- and XDR-TB in South Korea.

Objectives: Delamanid is a new anti-TB drug, but few data exist on its use outside clinical trials. The purpose of this study was to evaluate the efficacy as well as the safety and tolerability of a delamanid-containing regimen for 24 weeks in the treatment of MDR- and XDR-TB. Methods: We performed a retrospective cohort study among patients with MDR/XDR-TB who were treated with a delamanid-containing regimen in seven hospitals in South Korea. Results: A total of 32 patients with MDR-TB, of which 6 (18.8%) were XDR-TB, were included and all completed 24 weeks of delamanid treatment. Of 19 patients (59.4%) who had positive culture sputum at the initiation of delamanid treatment, the proportion of culture conversion at 8 weeks was 72.2% (13 of 18) in solid medium and 50.0% (7 of 14) in liquid medium. The proportion of culture conversion at 24 weeks was 94.4% (17 of 18) in solid medium and 92.9% (13 of 14) in liquid medium. The median time to culture conversion was 33 days (range = 5-81) using solid medium and 57 days (range = 8-96) using liquid medium. Of the 32 patients, there was no serious adverse event or death. Three patients developed a transient QTcF of > 500 ms. Conclusions: The use of delamanid combined with optimized background regimens has the potential to achieve high culture conversion rates at 24 weeks with an acceptable safety and tolerability profile in patients with MDR/XDR-TB.

Trends in the prescription of benzodiazepines for the elderly in Korea.

BACKGROUND: This study examined trends in the prescription of benzodiazepines for the elderly (age over 65 years) in Korea, a country with a higher level of spending on pharmaceuticals compared to that in other Organization for Economic Cooperation and Development (OECD) countries, and identified factors related to the inappropriate use of such drugs. METHODS: We used the National Health Insurance Claims Data (NHICD) for the period 2009-2013, including all reimbursed drug-prescribing information. Following the OECD's prescribing quality indicators (PQIs), we looked at the prevalence, quantities, durations, and inappropriate (long-term or high-quantity) use of benzodiazepines, some of the most widely prescribed, but potentially inappropriate, drugs for the elderly. We also performed multivariate logistic regression analyses to identify factors related to the inappropriate use of these drugs. RESULTS: The annual prevalence of benzodiazepine prescribing for elderly subjects decreased slightly over time but remained high (37.9% in 2009 and 35.1% in 2013). There were also small decreases in the inappropriate long-term use of benzodiazepines over the five years, with a 0.6 decrease in the Defined Daily Dose and a 4.1 per 1,000 decreases in elderly user-days. The proportion of subjects using long-acting benzodiazepines also fell from 263.6 to 220.4 per 1,000 elderly patients. The regression analyses found that the inappropriate long-term use of benzodiazepines in the elderly was significantly related to the patients visiting several institutions and physicians prescribing more than 30 days' worth of medication. CONCLUSIONS: The prevalence of prescribing potentially inappropriate drugs, such as benzodiazepines, remains high in Korea. Policy efforts, such as a periodic assessment of prescribing, restricting prescribing days, and more practical guidelines, are needed to improve the quality of prescribing.

Multidrug-Resistant Tuberculosis Treatment Outcomes in Relation to Treatment and Initial Versus Acquired Second-Line Drug Resistance.

BACKGROUND: Resistance to second-line drugs develops during treatment of multidrug-resistant (MDR) tuberculosis, but the impact on treatment outcome has not been determined. METHODS: Patients with MDR tuberculosis starting second-line drug treatment were enrolled in a prospective cohort study. Sputum cultures were analyzed at a central reference laboratory. We compared subjects with successful and poor treatment outcomes in terms of (1) initial and acquired resistance to fluoroquinolones and second-line injectable drugs (SLIs) and (2) treatment regimens. RESULTS: Of 1244 patients with MDR tuberculosis, 973 (78.2%) had known outcomes and 232 (18.6%) were lost to follow-up. Among those with known outcomes, treatment succeeded in 85.8% with plain MDR tuberculosis, 69.7% with initial resistance to either a fluoroquinolone or an SLI, 37.5% with acquired resistance to a fluoroquinolone or SLI, 29.3% with initial and 13.0% with acquired extensively drug-resistant tuberculosis (P < .001 for trend). In contrast, among those with known outcomes, treatment success increased stepwise from 41.6% to 92.3% as the number of drugs proven effective increased from ≤1 to ≥5 (P < .001 for trend), while acquired drug resistance decreased from 12% to 16% range, depending on the drug, down to 0%-2% (P < .001 for trend). In multivariable analysis, the adjusted odds of treatment success decreased 0.62-fold (95% confidence interval, .56-.69) for each increment in drug resistance and increased 2.1-fold (1.40-3.18) for each additional effective drug, controlling for differences between programs and patients. Specific treatment, patient, and program variables were also associated with treatment outcome. CONCLUSIONS: Increasing drug resistance was associated in a logical stepwise manner with poor treatment outcomes. Acquired resistance was worse than initial resistance to the same drugs. Increasing numbers of effective drugs, specific drugs, and specific program characteristics were associated with better outcomes and less acquired resistance.

Association of cardiovascular health screening with mortality, clinical outcomes, and health care cost: a nationwide cohort study.

OBJECTIVE: To determine whether a cardiovascular disease (CVD) health screening program is associated with CVD-related health conditions, incidence of cardiovascular events, mortality, healthcare utilization, and costs. METHODS: Cohort study of a 3% random sample of all Korea National Health Insurance members 40years of age or older and free of CVD or CVD-related health conditions was conducted. A total 443,337 study participants were followed-up from January 1, 2005 through December 31, 2010. RESULTS: In primary analysis, the hazard ratios for CVD mortality, all-cause mortality, incident composite CVD events, myocardial infarction, cerebral infarction, and cerebral hemorrhage comparing participants who attended a screening exam during 2003-2004 compared to those who did not were 0.58 (95% CI: 0.53-0.63), 0.62 (95% CI: 0.60-0.64), 0.82 (95% CI: 0.78-0.85), 0.84 (95% CI: 0.75-0.93), 0.84 (95% CI: 0.79-0.89), and 0.73 (95% CI: 0.67-0.80), respectively. Screening attenders had higher rates of newly diagnosed hypertension, diabetes mellitus, and dyslipidemia, lower inpatient days of stay and cost, and lower outpatient cost compared to non-attenders. CONCLUSIONS: Participation in CVD health screening was associated with lower rates of CVD, all-cause mortality, and CVD events, higher detection of CVD-related health conditions, and lower healthcare utilization and costs.

Variations in the hospital standardized mortality ratios in Korea.

OBJECTIVES: The hospital standardized mortality ratio (HSMR) has been widely used because it allows for robust risk adjustment using administrative data and is important for improving the quality of patient care. METHODS: All inpatients discharged from hospitals with more than 700 beds (66 hospitals) in 2008 were eligible for inclusion. Using the claims data, 29 most responsible diagnosis (MRDx), accounting for 80% of all inpatient deaths among these hospitals, were identified, and inpatients with those MRDx were selected. The final study population included 703 571 inpatients including 27 718 (3.9% of all inpatients) in-hospital deaths. Using logistic regression, risk-adjusted models for predicting in-hospital mortality were created for each MRDx. The HSMR of individual hospitals was calculated for each MRDx using the model coefficients. The models included age, gender, income level, urgency of admission, diagnosis codes, disease-specific risk factors, and comorbidities. The Elixhauser comorbidity index was used to adjust for comorbidities. RESULTS: For 26 out of 29 MRDx, the c-statistics of these mortality prediction models were higher than 0.8 indicating excellent discriminative power. The HSMR greatly varied across hospitals and disease groups. The academic status of the hospital was the only factor significantly associated with the HSMR. CONCLUSIONS: We found a large variation in HSMR among hospitals; therefore, efforts to reduce these variations including continuous monitoring and regular disclosure of the HSMR are required. OBJECTIVES: The hospital standardized mortality ratio (HSMR) has been widely used because it allows for robust risk adjustment using administrative data and is important for improving the quality of patient care. METHODS: All inpatients discharged from hospitals with more than 700 beds (66 hospitals) in 2008 were eligible for inclusion. Using the claims data, 29 most responsible diagnosis (MRDx), accounting for 80% of all inpatient deaths among these hospitals, were identified, and inpatients with those MRDx were selected. The final study population included 703 571 inpatients including 27 718 (3.9% of all inpatients) in-hospital deaths. Using logistic regression, risk-adjusted models for predicting in-hospital mortality were created for each MRDx. The HSMR of individual hospitals was calculated for each MRDx using the model coefficients. The models included age, gender, income level, urgency of admission, diagnosis codes, disease-specific risk factors, and comorbidities. The Elixhauser comorbidity index was used to adjust for comorbidities. RESULTS: For 26 out of 29 MRDx, the c-statistics of these mortality prediction models were higher than 0.8 indicating excellent discriminative power. The HSMR greatly varied across hospitals and disease groups. The academic status of the hospital was the only factor significantly associated with the HSMR. CONCLUSIONS: We found a large variation in HSMR among hospitals; therefore, efforts to reduce these variations including continuous monitoring and regular disclosure of the HSMR are required.

Health behaviors of people with retinitis pigmentosa in the republic of Korea.

PURPOSE: To investigate the health behaviors of people with retinitis pigmentosa (RP) in the Republic of Korea. METHODS: An online questionnaire was used to survey members of the Korean Retinitis Pigmentosa Society (KRPS), over 3 months from December 2010 to February 2011. Controls were selected from participants of the 4th Korean National Health and Nutrition Examination Survey and were matched with people with RP using the propensity score method to optimize comparative analysis. A semi-structured interview was also conducted with five members of the KRPS to identify potential reasons for physical inactivity and unhealthy diet and to develop effective interventions. RESULTS: A total of 194 eligible people with RP responded to the online survey, and 187 individuals were matched with the control population by propensity score matching. RP subjects reported a lower rate of current smoking and greater use of preventive healthcare services. However, people with RP were more physically inactive (50.8% vs 27.3%, p < 0.001) and consumed more fast food (69.5% vs 58.3%, p = 0.024) than the control population. CONCLUSION: Physical inactivity and fast food consumption were more frequently reported in people with RP than the general population. Efforts to understand the possible reasons and develop interventions to improve these health behaviors are warranted.

The predictability of claim-data-based comorbidity-adjusted models could be improved by using medication data.

BACKGROUND: Recently, claim-data-based comorbidity-adjusted methods such as the Charlson index and the Elixhauser comorbidity measures have been widely used among researchers. At the same time, there have been an increasing number of attempts to improve the predictability of comorbidity-adjusted models. We tried to improve the predictability of models using the Charlson and Elixhauser indices by using medication data; specifically, we used medication data to estimate omitted comorbidities in the claim data. METHODS: We selected twelve major diseases (other than malignancies) that caused large numbers of in-hospital mortalities during 2008 in hospitals with 700 or more beds in South Korea. Then, we constructed prediction models for in-hospital mortality using the Charlson index and Elixhauser comorbidity measures, respectively. Inferring missed comorbidities using medication data, we built enhanced Charlson and Elixhauser comorbidity-measures-based prediction models, which included comorbidities inferred from medication data. We then compared the c-statistics of each model. RESULTS: 247,712 admission cases were enrolled. 55 generic drugs were used to infer 8 out of 17 Charlson comorbidities, and 106 generic drugs were used to infer 14 out of 31 Elixhauser comorbidities. Before the inclusion of comorbidities inferred from medication data, the c-statistics of models using the Charlson index were 0.633-0.882 and those of the Elixhauser index were 0.699-0.917. After the inclusion of comorbidities inferred from medication data, 9 of 12 models using the Charlson index and all of the models using the Elixhauser comorbidity measures were improved in predictability but, the differences were relatively small. CONCLUSION: Prediction models using Charlson index or Elixhauser comorbidity measures might be improved by including comorbidities inferred from medication data.

Mental health of people with retinitis pigmentosa.

PURPOSE: The purpose of this study was to evaluate and compare the mental health of patients with retinitis pigmentosa (RP) with that of the general population of Korea. METHODS: Online surveys were completed by patients registered with the KRPS (Korean Retinitis Pigmentosa Society), an online organization that promotes research on RP and provides advocacy and online and offline support and information for patients with RP. Control population was selected from the fourth round of the KNHANES (Korean National Health and Nutrition Examination Survey). One hundred eighty-seven patients with RP were matched with the control population using the propensity-score method to optimize comparative analysis. RESULTS: Stress was reported in 51.9% of RP patients and 29.4% of controls (p < 0.001). Depressive mood of at least 2 weeks' duration in the previous year was reported by 34.8 and 17.1% of patients and controls, respectively (p < 0.001). Suicidal thoughts were reported by 38.5 and 12.9% of patients and controls, respectively (p < 0.001), although there was no significant difference in the number of suicide attempts between the groups (2.1 vs. 1.6%, p = 0.703). In multivariate analysis, disability rating was significantly associated with stress (adjusted odds ratio, 0.46; 95% confidence interval, 0.24 to 0.88). CONCLUSIONS: People with RP had poorer mental health than the general population. Further investigations are warranted on the mental health of RP patients, and appropriate welfare services are needed to decrease the impact of mental illness in this population.

Alcohol consumption and the risk of type 2 diabetes mellitus: effect modification by hypercholesterolemia: the Third Korea National Health and Nutrition Examination Survey (2005).

OBJECTIVE: While the protective nature of moderate alcohol consumption against diabetes mellitus is well known, inconsistent findings continue to be reported. The possibility of different mixes of effect modifiers has been raised as a reason for those inconsistent findings. Our study aim was to examine potential effect modifiers that can change the effect of alcohol consumption on type 2 diabetes. METHODS: From data in the third Korea National Health and Nutrition Examination Survey, 3,982 individuals over the age of 30 years who had not been diagnosed with diabetes were selected for inclusion in the study population. Breslow and Day's test and the Wald test between hypercholesterolemia and alcohol consumption in a multiple logistic regression model were used to assess effect modification. RESULTS: Odds ratios for diabetes stratified by alcohol consumption strata and assessed using Breslow and Day's tests for homogeneity indicated that hypercholesterolemia was not a significant confounding factor (p=0.01). However, the Wald test for interaction terms, which is a conservative method of effect modification, was significant (p=0.03). CONCLUSIONS: The results indicate that moderate alcohol consumption is not necessarily protective for type 2 diabetes mellitus, if a person has hypercholesterolemia. People who have hypercholesterolemia should be aware of the risk associated with alcohol consumption, a risk that contrasts with the reported protective effect of moderate alcohol consumption on diabetes.

Rapid detection of Mycobacterium tuberculosis biomarkers in a sandwich immunoassay format using a waveguide-based optical biosensor.

Early diagnosis of active tuberculosis (TB) remains an elusive challenge, especially in individuals with disseminated TB and HIV co-infection. Recent studies have shown a promise for the direct detection of pathogen-specific biomarkers such as lipoarabinomannan (LAM) for the diagnosis of TB in HIV-positive individuals. Currently, traditional immunoassay platforms that suffer from poor sensitivity and high non-specific interactions are used for the detection of such biomarkers. In this manuscript, we demonstrate the development of sandwich immunoassays for the direct detection of three TB-specific biomarkers, namely LAM, early secretory antigenic target 6 (ESAT6) and antigen 85 complex (Ag85), using a waveguide-based optical biosensor platform. Combining detection within the evanescent field of a planar optical waveguide with functional surfaces that reduce non-specific interactions allows for the ultra-sensitive and quantitative detection of biomarkers (an order of magnitude enhanced sensitivity, as compared to plate-based ELISA) in complex patient samples (urine, serum) within a short time. We also demonstrate the detection of LAM in urine from a small sample of subjects being treated for TB using this approach with excellent sensitivity and 100% corroboration with disease status. These results suggest that pathogen-specific biomarkers can be applied for the rapid and effective diagnosis of disease. It is likely that detection of a combination of biomarkers offers greater reliability of diagnosis, rather than detection of any single pathogen biomarker. NCT00341601.

Rapid, high-throughput detection of rifampin resistance and heteroresistance in Mycobacterium tuberculosis by use of sloppy molecular beacon melting temperature coding.

Rifampin resistance in Mycobacterium tuberculosis is largely determined by mutations in an 80-bp rifampin resistance determining region (RRDR) of the rpoB gene. We developed a rapid single-well PCR assay to identify RRDR mutations. The assay uses sloppy molecular beacons to probe an asymmetric PCR of the M. tuberculosis RRDR by melting temperature (T(m)) analysis. A three-point T(m) code is generated which distinguishes wild-type from mutant RRDR DNA sequences in approximately 2 h. The assay was validated on synthetic oligonucleotide targets containing the 44 most common RRDR mutations. It was then tested on a panel of DNA extracted from 589 geographically diverse clinical M. tuberculosis cultures, including isolates with wild-type RRDR sequences and 25 different RRDR mutations. The assay detected 236/236 RRDR mutant sequences as mutant (sensitivity, 100%; 95% confidence interval [CI], 98 to 100%) and 353/353 RRDR wild-type sequences as wild type (specificity, 100%; 95% CI, 98.7 to 100%). The assay identified 222/225 rifampin-resistant isolates as rifampin resistant (sensitivity, 98.7%; 95% CI, 95.8 to 99.6%) and 335/336 rifampin-susceptible isolates as rifampin susceptible (specificity, 99.7%; 95% CI, 95.8 to 99.6%). All mutations were either individually identified or clustered into small mutation groups using the triple T(m) code. The assay accurately identified mixed (heteroresistant) samples and was shown analytically to detect RRDR mutations when present in at least 40% of the total M. tuberculosis DNA. This was at least as accurate as Sanger DNA sequencing. The assay was easy to use and well suited for high-throughput applications. This new sloppy molecular beacon assay should greatly simplify rifampin resistance testing in clinical laboratories.

Mycolic acids as diagnostic markers for tuberculosis case detection in humans and drug efficacy in mice.

Mycolic acids are attractive diagnostic markers for tuberculosis (TB) infection because they are bacteria-derived, contain information about bacterial species, modulate host-pathogen interactions and are chemically inert. Here, we present a novel approach based on mass spectrometry. Quantification of specific precursor → fragment transitions of approximately 2000 individual mycolic acids (MAs) resulted in high analytical sensitivity and specificity. We next used this tool in a retrospective case-control study of patients with pulmonary TB with varying disease burdens from South Korea, Vietnam, Uganda and South Africa. MAs were extracted from small volume sputum (200 µl) and analysed without the requirement for derivatization. Infected patients (70, 19 of whom were HIV+) could be separated from controls (40, 20 of whom were HIV+) with a sensitivity and specificity of 94 and 93%, respectively. Furthermore, we quantified MA species in lung tissue of TB-infected mice and demonstrated effective clearance of MA levels following curative rifampicin treatment. Thus, our results demonstrate for the first time the feasibility and clinical relevance of direct detection of mycobacterial lipids as biomarkers of TB infection.

Association of antigen-stimulated release of tumor necrosis factor-alpha in whole blood with response to chemotherapy in patients with pulmonary multidrug-resistant tuberculosis.

BACKGROUND: We have previously reported that TNF-α levels correlate to total mycobacterial burden in tuberculosis (TB) patients. OBJECTIVE: To characterize the dynamics of cytokine responses in TB patients during chemotherapy to identify potential surrogate markers for effective treatment. METHODS: Following induction by culture filtrate proteins in whole blood, production patterns of TNF-α, IL-10, IFN-γ and IL-12 were measured in 23 non-multidrug-resistant (MDR)-TB and 16 MDR-TB patients and in 31 healthy controls. Rates of mycobacterial clearance from the sputum were then measured and compared. RESULTS: Prior to the initiation of chemotherapy, TNF-α and IL-10 levels were significantly higher in TB patients than in healthy controls while IFN-γ and IL-12 levels were similar. During chemotherapy, the levels of all 4 cytokines increased. We evaluated these responses separately in patients that did and did not clear their sputum culture at 2 and 6 months. At 2 months, decreases in both IFN-γ and IL-12 correlated strongly with a successful early response, while after 6 months of therapy, when half (7/14) of MDR-TB patients were still sputum culture positive, downregulation of TNF-α was uniquely correlated with sputum conversion between the groups. CONCLUSION: Our findings suggest the possibility that the regulation of TNF-α production in whole blood may be a more specific indicator of sputum conversion at 6 months than IFN-γ, IL-12 or IL-10 in MDR-TB patients.

Mutations in extensively drug-resistant Mycobacterium tuberculosis that do not code for known drug-resistance mechanisms.

BACKGROUND: Highly lethal outbreaks of multidrug-resistant (MDR) and extensively drug-resistant (XDR) tuberculosis are increasing. Whole-genome sequencing of KwaZulu-Natal MDR and XDR outbreak strains prevalent in human immunodeficiency virus (HIV)-infected patients by the Broad Institute identified 22 novel mutations which were unique to the XDR genome or shared only by the MDR and XDR genomes and not already known to be associated with drug resistance. METHODS: We studied the 12 novel mutations which were not located in highly-repetitive genes to identify mutations that were truly associated with drug resistance or were likely to confer a specific fitness advantage. RESULTS: None of these mutations could be found in a phylogenetically and geographically diverse set of drug-resistant and drug-susceptible Mycobacterium tuberculosis isolates, suggesting that these mutations are unique to the KZN clone. Examination of the 600-basepair region flanking each mutation revealed 26 new mutations. We searched for a convergent evolutionary signal in the new mutations for evidence that they emerged under selective pressure, consistent with increased fitness. However, all but 1 rare mutation were monophyletic, indicating that the mutations were markers of strain phylogeny rather than fitness or drug resistance. CONCLUSIONS: Our results suggest that virulent XDR tuberculosis in immunocompromised HIV-infected patients can evolve without generalizable fitness changes or other XDR-specific mutations.

Neutrophils are the predominant infected phagocytic cells in the airways of patients with active pulmonary TB.

BACKGROUND: The exact role of neutrophils in the pathogenesis of TB is poorly understood. Recent evidence suggests that neutrophils are not simply scavenging phagocytes in Mycobacterium tuberculosis (Mtb) infection. METHODS: Three different types of clinical specimens from patients with active pulmonary TB who underwent lung surgery were examined: sputum, BAL fluid, and cavity contents. Differential cell separation and quantification were performed for intracellular and extracellular bacteria, and bacterial length was measured using microscopy. RESULTS: Neutrophils were more abundant than macrophages in sputum (86.6% +/- 2.2% vs 8.4% +/- 1.3%) and in BAL fluid (78.8% +/- 5.8% vs 11.8% +/- 4.1%). Inside the cavity, lymphocytes (41.3% +/- 11.2%) were the most abundant cell type, followed by neutrophils (38.8% +/- 9.4%) and macrophages (19.5% +/- 7.5%). More intracellular bacilli were found in neutrophils than macrophages in sputum (67.6% +/- 5.6% vs 25.2% +/- 6.5%), in BAL fluid (65.1% +/- 14.4% vs 28.3% +/- 11.6%), and in cavities (61.8% +/- 13.3% vs 23.9% +/- 9.3%). The lengths of Mtb were shortest in cavities (1.9+/- 0.1 microm), followed by in sputum (2.9 +/- 0.1 microm) and in BAL fluid (3.6 +/- 0.2 microm). CONCLUSIONS: Our results show that neutrophils are the predominant cell types infected with Mtb in patients with TB and that these intracellular bacteria appear to replicate rapidly. These results are consistent with a role for neutrophils in providing a permissive site for a final burst of active replication of the bacilli prior to transmission.