Environmental risks and sphingolipid signatures in adult asthma and its phenotypic clusters: a multicentre study.

BACKGROUND: Adult asthma is phenotypically heterogeneous with unclear aetiology. We aimed to evaluate the potential contribution of environmental exposure and its ensuing response to asthma and its heterogeneity. METHODS: Environmental risk was evaluated by assessing the records of National Health Insurance Research Database (NHIRD) and residence-based air pollution (particulate matter with diameter less than 2.5 micrometers (PM2.5) and PM2.5-bound polycyclic aromatic hydrocarbons (PAHs)), integrating biomonitoring analysis of environmental pollutants, inflammatory markers and sphingolipid metabolites in case-control populations with mass spectrometry and ELISA. Phenotypic clustering was evaluated by t-distributed stochastic neighbor embedding (t-SNE) integrating 18 clinical and demographic variables. FINDINGS: In the NHIRD dataset, modest increase in the relative risk with time-lag effect for emergency (N=209 837) and outpatient visits (N=638 538) was observed with increasing levels of PM2.5 and PAHs. Biomonitoring analysis revealed a panel of metals and organic pollutants, particularly metal Ni and PAH, posing a significant risk for current asthma (ORs=1.28-3.48) and its severity, correlating with the level of oxidative stress markers, notably Nε-(hexanoyl)-lysine (r=0.108-0.311, p<0.05), but not with the accumulated levels of PM2.5 exposure. Further, levels of circulating sphingosine-1-phosphate and ceramide-1-phosphate were found to discriminate asthma (p<0.001 and p<0.05, respectively), correlating with the levels of PAH (r=0.196, p<0.01) and metal exposure (r=0.202-0.323, p<0.05), respectively, and both correlating with circulating inflammatory markers (r=0.186-0.427, p<0.01). Analysis of six phenotypic clusters and those cases with comorbid type 2 diabetes mellitus (T2DM) revealed cluster-selective environmental risks and biosignatures. INTERPRETATION: These results suggest the potential contribution of environmental factors from multiple sources, their ensuing oxidative stress and sphingolipid remodeling to adult asthma and its phenotypic heterogeneity.

Prevention and management of tuberculosis in solid organ transplantation: A consensus statement of the transplantation society of Taiwan.

Solid organ transplant recipients have an increased risk of tuberculosis (TB). Due to the use of immunosuppressants, the incidence of TB among solid organ transplant recipients has been consistently reported to be higher than that among the general population. TB frequently develops within the first year after transplantation when a high level of immunosuppression is maintained. Extrapulmonary TB and disseminated TB account for a substantial proportion of TB among solid organ transplant recipients. Treatment of TB among recipients is complicated by the drug-drug interactions between anti-TB drugs and immunosuppressants. TB is associated with an increased risk of graft rejection, graft failure and mortality. Detection and management of latent TB infection among solid organ transplant candidates and recipients have been recommended. However, strategy to mitigate the risk of TB among solid organ transplant recipients has not yet been established in Taiwan. To address the challenges of TB among solid organ transplant recipients, a working group of the Transplantation Society of Taiwan was established. The working group searched literatures on TB among solid organ transplant recipients as well as guidelines and recommendations, and proposed interventions to strengthen TB prevention and care among solid organ transplant recipients.

Comprehensive phylogenetic analyses of Orchidaceae using nuclear genes and evolutionary insights into epiphytism.

Orchidaceae (with >28,000 orchid species) are one of the two largest plant families, with economically and ecologically important species, and occupy global and diverse niches with primary distribution in rainforests. Among orchids, 70% grow on other plants as epiphytes; epiphytes contribute up to ~50% of the plant diversity in rainforests and provide food and shelter for diverse animals and microbes, thereby contributing to the health of these ecosystems. Orchids account for over two-thirds of vascular epiphytes and provide an excellent model for studying evolution of epiphytism. Extensive phylogenetic studies of Orchidaceae and subgroups have ;been crucial for understanding relationships among many orchid lineages, although some uncertainties remain. For example, in the largest subfamily Epidendroideae with nearly all epiphytic orchids, relationships among some tribes and many subtribes are still controversial, hampering evolutionary analyses of epiphytism. Here we obtained 1,450 low-copy nuclear genes from 610 orchid species, including 431 with newly generated transcriptomes, and used them for the reconstruction of robust Orchidaceae phylogenetic trees with highly supported placements of tribes and subtribes. We also provide generally well-supported phylogenetic placements of 131 genera and 437 species that were not sampled by previous plastid and nuclear phylogenomic studies. Molecular clock analyses estimated the Orchidaceae origin at ~132 million years ago (Ma) and divergences of most subtribes from 52 to 29 Ma. Character reconstruction supports at least 14 parallel origins of epiphytism; one such origin was placed at the most recent common ancestor of ~95% of epiphytic orchids and linked to modern rainforests. Ten occurrences of rapid increase in the diversification rate were detected within Epidendroideae near and after the K-Pg boundary, contributing to ~80% of the Orchidaceae diversity. This study provides a robust and the largest family-wide Orchidaceae nuclear phylogenetic tree thus far and new insights into the evolution of epiphytism in vascular plants.

Phylogenetic analysis and character evolution of tribe Arethuseae (Orchidaceae) reveal a new genus Mengzia.

Delimitation of the tribe Arethuseae has varied considerably since it was first defined. The relationships within Arethuseae, particularly within the subtribe Arethusinae, remain poorly elucidated. In this study, we reconstructed the phylogeny of Arethuseae, using six plastid markers (matK, ycf1, rbcL rpoc1, rpl32-trnL and trnL-F) from 83 taxa. The ancestral state reconstruction of 11 selected morphological characters was also conducted to identify synapomorphies and assess potential evolutionary transitions. Morphological character comparision between the distinct species Bletilla foliosa and other species are conducted. Our results unequivocally supported the monophyly of Arethuseae, which included highly supported clades and a clear synapomorphy of non-trichome-like lamellae. Furthermore, B. foliosa formed a separate clade in the subtribe Arethusinae, instead of clustering with the other Bletilla species in the subtribe Coelogyninae. The morphological characters comparision further showed that the B. foliosa clade could be distinguished from other genera in Arethuseae by multiple characters, including presence of lateral inflorescence, three lamellae with trichome-like apex and four pollinia. In light of these molecular and morphological evidences, we propose Mengzia as a new genus to accommodate B. foliosa and accordingly provide descriptions of this new genus and combination.

The prevalence and clinical features of chronic obstructive pulmonary disease patients with traits of asthma in Taiwan.

BACKGROUND/PURPOSE: The application of the checkbox for identifying patients with traits of both chronic obstructive pulmonary disease (COPD) and asthma proposed by the 2015 Global Initiative for Asthma (GINA)/Global Initiative for Chronic Obstructive Lung Disease (GOLD) recommendations has not been well studied although such identification is important in clinical practice. Thus, we aimed to investigate the prevalence and features of COPD coexistent with asthma traits diagnosed based on the 2015 GINA/GOLD strategies, and explore the gap between guidelines and routine practice in the diagnosis and pharmacological management of such condition in a COPD cohort. METHODS: COPD subjects were enrolled retrospectively throughout Taiwan. A patient record form was completed for each participant and the data were analyzed. RESULTS: Of 340 participants, the prevalence of COPD coexistent with traits of asthma was 39.4% and 30.3% based on guidelines and physician's judgment, respectively. Coexistent patients were characterized by blood eosinophilia, higher total immunoglobulin E (IgE) levels, preserved lung function, and the presence of gastro-esophageal reflux disease and atopic disease while total IgE level > 100 kU/L and the presence of atopic disease were predictors for coexistent patients. Gaps existed in the diagnosis (a weak agreement with kappa = 0.53) and treatment (non-adherence to the preferred therapy in 18.4% of physician-judged coexistent patients) in COPD patients with asthma traits. The exacerbation history was similar between coexistent and non-coexistent patients. CONCLUSION: We found that measuring circulatory eosinophil and total IgE levels may raise clinicians' awareness of the presence of traits of asthma in the management of COPD.

Completion Rate and Safety of Programmatic Screening and Treatment for Latent Tuberculosis Infection in Elderly Patients With Poorly Controlled Diabetic Mellitus: A Prospective Multicenter Study.

BACKGROUND: Poor control of diabetes mellitus (DM) increases active tuberculosis (TB) risk. Understanding risk factors for latent TB infection (LTBI) in this population and intervention completion rates is crucial for policy making. METHODS: Under a collaborative multidisciplinary team consisting of public health professionals, endocrinologists, and pulmonologists, patients aged >45 years with poorly controlled DM (pDM), defined as having a glycated hemoglobin level of ≥9% within the preceding year, were enrolled by endocrinologists from 2 hospitals; these patients underwent LTBI screening by using QuantiFERON (QFT). Once-weekly isoniazid and rifapentine for 12 weeks (3HP) or daily isoniazid for 9 months (9H) was administered by pulmonologists. QFT-positivity predictors were evaluated using logistic regression. Completion rates and safety were also investigated. RESULTS: Among 980 patients with pDM (age: 64.2 ±â€…9.7 years), 261 (26.6%) were QFT-positive. Age, DM duration, chronic kidney disease stage ≥3, and dipeptidyl peptidase-4 inhibitor use, not using metformin, were associated with QFT-positivity. Preventive therapy (3HP: 138; 9H: 62) was administered in 200 (76.6%) QFT-positive patients. The completion rates of 3HP and 9H were 84.1% and 79.0%, respectively (P = .494). Nine (6.5%) and zero patients in the 3HP and 9H groups, respectively, developed systemic drug reactions (P = .059); 78.3% and 45.2% had ≥1 adverse drug reactions (P < .001); and post-treatment QFT conversion rates were 32% and 20%, respectively (P = .228). CONCLUSIONS: LTBI prevalence exceeds 25% in elderly patients with pDM. Under care from a collaborative multidisciplinary team, the completion rate of preventive therapy, regardless of regimen could approach, or even exceed 80% in this population.

Cross-phytogroup assessment of foliar epiphytic mycobiomes.

The foliar surface forms one of the largest aboveground habitats on Earth and maintains plant-fungus relationships that greatly affect ecosystem functioning. Despite many studies with particular plant species, the foliar epiphytic mycobiome has not been studied across a large number of plant species from different taxa. Using high-throughput sequencing, we assessed epiphytic mycobiomes on leaf surfaces of 592 plant species in a botanical garden. Plants of angiosperms, gymnosperms, and pteridophytes were involved. Plant taxonomy, leaf side, growing environment, and evolutionary relationships were considered. We found that pteridophytes showed the higher fungal species diversity, stronger mutualistic fungal interactions, and a greater percentage of putative pathogens than gymnosperms and angiosperms. Plant taxonomic group, leaf side, and growing environment were significantly associated with the foliar epiphytic mycobiome, but the similarity of the mycobiomes among plants was not directly related to the distance of the host evolutionary tree. Our results provide a general understanding of the foliar fungal mycobiomes from pteridophytes to angiosperms. These findings will facilitate our understanding of foliar fungal epiphytes and their roles in plant communities and ecosystems.

Update on guidelines for the treatment of COPD in Taiwan using evidence and GRADE system-based recommendations.

Chronic obstructive pulmonary disease (COPD) has significant contributions to morbidity and mortality world-wide. Early symptoms of COPD are not readily distinguishable, resulting in a low rate of diagnosis and intervention. Different guidelines and recommendatations for the diagnosis and treatment of COPD exist globally. The first edition of clinical practice guidelines for COPD was published in 2016 by the Ministry of Health and Welfare in Taiwan in collaboration with the Taiwan evidence-based medicine association and Cochrane Taiwan, and was revised in 2019 in order to update recent diagnostic and therapeutic modalities for COPD and its acute exacerbation. This revised guideline covered a range of topics highlighted in the Global Initiative for Chronic Obstructive Lung Disease (GOLD) report, including strategies for the diagnosis, assessment, monitoring, and management of stable COPD and exacerbations, with particular focus on evidence from Taiwan. The recommendations included in the revised guideline were formed based on a comprehensive systematic review or meta-analysis of specific clinical issues identified by an expert panel that surveyed relevant scientific evidence in the literature and guidelines published by the clinical communities and organizations nationally and internationally. The guidelines and recommendations are applicable to the clinical settings in Taiwan. We expect this revised guideline to facilitate the diagnosis, treatment and management of patients with COPD by physicians and health care professionals in Taiwan. Adaptations of the materials included herein for educational and training purposes is encouraged.

Identification and drug susceptibility testing for nontuberculous mycobacteria.

Great progress has recently been made in methodologies for identifying nontuberculous mycobacteria (NTM). Recommendations for drug susceptibility testing (DST) of NTM have been expanded and updated by the Clinical and Laboratory Standards Institute and are crucial in the management of NTM infections. This article summarizes the clinically relevant molecular methods used to discriminate NTM species and updates the information on DST. Furthermore, recent progress on new antimicrobials against NTM infections is reviewed.

Phylogenomics of Orchidaceae based on plastid and mitochondrial genomes.

To advance our knowledge of orchid relationships and timing of their relative divergence, we used 76 protein-coding genes from plastomes (ptCDS) and 38 protein-coding genes from mitochondrial genomes (mtCDS) of 74 orchids representing the five subfamilies and 18 tribes of Orchidaceae, to reconstruct the phylogeny and temporal evolution of the Orchidaceae. In our results, the backbone of orchid tree well supported with both datasets, but there are conflicts between these trees. The phylogenetic positions of two subfamilies (Vanilloideae and Cypripedioideae) are reversed in these two analyses. The phylogenetic positions of several tribes and subtribes, such as Epipogiinae, Gastrodieae, Nerviliinae, and Tropidieae, are well resolved in mtCDS tree. Thaieae have a different position among higher Epidendroideae, instead of sister to the higher Epidendroideae. Interrelationships of several recently radiated tribes within Epidendroideae, including Vandeae, Collabieae, Cymbidieae, Epidendreae, Podochileae, and Vandeae, have good support in the ptCDS tree, but most are not resolved in the mtCDS tree. Conflicts between the two datasets may be attributed to the different substitution rates in these two genomes and heterogeneity of substitution rate of plastome. Molecular dating indicated that the first three subfamilies, Apostasioideae, Cypripedioideae and Vanilloideae, diverged relatively quickly, and then there was a longer period before the last two subfamilies, Orchidoideae and Epidendroideae, began to radiate. Most mycoheterotrophic clades of Orchidaceae evolved in the last 30 million years with the exception of Gastrodieae.

Effects of Analyzer Detection Method, Ethnicity, and Reference Individuals on Reference Interval of Immature Platelet Fraction.

BACKGROUND: Immature platelet fraction (IPF) is a new biomarker for thrombopoiesis and inflammation. However, the reference interval (RI) is wildly discrepant among published reports. This study aimed to establish the RI of IPF for a population in Taiwan and evaluate the effects the detection method of the analyzer, ethnicity, and reference individuals have on the RI of IPF. METHODS: The RI of absolute IPF (A-IPF) and IPF% were established with healthy subjects from the outpatient services of the Health Management Department of Taichung Veterans General Hospital between January 1, 2015 and March 1, 2016. These values were used along with published reports for meta-analysis. RESULTS: A-IPF (109/L) and IPF% of Taiwanese were 6.9 - 7.6 and 3.1 - 3.4, respectively. Significant differences were found when performing paired comparisons of the RI of A-IPF and IPF% published in reports. For A-IPF, there was only one paired comparison with a significant difference (Z > 1.96) across 6 reports. Thus, the contribution of the factors examined on the RI of IPF cannot be determined. For IPF%, there were 8 paired comparisons with significant differences across 10 reports. The discrepancy rates of RI for IPF% were 41.2%, 50.0%, and 25.0% with the difference of reference individuals, the analyzer method, and ethnicity, respectively. CONCLUSIONS: The RIs of Taiwanese for A-IPF and IPF% were established. Furthermore, the analyzer detection method and the reference individuals contribute to the discrepancy of the RI for IPF% and should be considered cautiously when the value of IPF is interpreted.

A phylogenetic analysis of molecular and morphological characters of Herminium (Orchidaceae, Orchideae): evolutionary relationships, taxonomy, and patterns of character evolution.

The first comprehensive phylogenetic study of the orchid genus Herminium and its allies is presented, based on seven molecular markers (nuclear internal transcribed spacer, Xdh, chloroplast matK, psaB, psbA-trnH, rbcL and trnL-F) and 37 morphological characters. Phylogenetic analyses indicate that Herminium as currently delimited is paraphyletic and that several genera are deeply nested within it. Based on parsimony analysis of total evidence, the generic circumscription of Herminium is expanded to include Androcorys, Bhutanthera, Frigidorchis and Porolabium. Apomorphic and plesiomorphic character states are identified for various clades recovered in this study. A few species currently wrongly assigned to Peristylus and Platanthera are here included in Herminium. All necessary new combinations are made.

Confirmed False Positive Proteinuria in Patients with Systemic Lupus Erythematosus Taking Hydroxychloroquine: a Spot Sample Measurement.

BACKGROUND: A false-positive screening result is associated with harmful treatment or follow-up costs. This study aimed to estimate the rate of false positive proteinuria with the dipstick in patients with systemic lupus erythematosus (SLE) taking hydroxychloroquine. METHODS: A total of 334 patients with a positive dipstick and confirmed by total urine protein with quantification assay were enrolled. The experimental group included those with SLE taking hydroxychloroquine, and the rest was the control group. The difference of the rate of false positive in the dipstick was analyzed using the chi-square test and odds ratio (OR) between groups. Qualitative tracking of potential interference in the dipstick was performed. RESULTS: The results revealed that the rate of false positive with a dipstick for the experimental and control groups were 29.5% and 5.0% (p = 0.000), respectively. The OR with 95% confidence interval (CI) of the rate of false positive for the experimental group with respect to the control group was 5.95 (95% CI: 2.80 - 12.65). Qualitative tracking showed that the dipstick was influenced to become false-positive when hydroxychloroquine concentration was ≥ 30 mg/dL. CONCLUSIONS: Hydroxychloroquines like plaquenil or geniquin may lead to a high rate of false positive with the dipstick method. A quantification assay is recommended for proteinuria measurement in patients with SLE taking hydroxychloroquines.

Molecular systematics of subtribe Orchidinae and Asian taxa of Habenariinae (Orchideae, Orchidaceae) based on plastid matK, rbcL and nuclear ITS.

The subtribe Orchidinae, distributed predominantly in Eastern Asia and the Mediterranean, presents some of the most intricate taxonomic problems in the family Orchidaceae with respect to generic delimitation. Based on three DNA markers (plastid matK, rbcL, and nuclear ITS), morphological characters, and a broad sampling of Orchidinae and selected Habenariinae mainly from Asia (a total of 153 accessions of 145 species in 31 genera), generic delimitation and phylogenetic relationships within the subtribe Orchidinae and Habenariinae from Asia were assessed. Orchidinae and Asian Habenariinae are monophyletic, and Orchidinae is divided into distinct superclades. Many genera, such as Amitostigma, Habenaria, Hemipilia, Herminium, Platanthera, Peristylus and Ponerorchis, are not monophyletic. Habenaria is subdivided into two distantly related groups, while Platanthera is subdivided into three even more disparate groups. Many previously undetected phylogenetic relationships, such as clades formed by the Amitostigma-Neottianthe-Ponerorchis complex, Platanthera latilabris group, Ponerorchis chrysea, Sirindhornia, and Tsaiorchis, are well supported by both molecular and morphological evidence. We propose to combine Hemipiliopsis with Hemipilia, Amitostigma and Neottianthe with Ponerorchis, Smithorchis with Platanthera, and Aceratorchis and Neolindleya with Galearis, and to establish a new genus to accommodate Ponerorchis chrysea. Tsaiorchis and Sirindhornia are two distinctive genera supported by both molecular data and morphological characters. A new genus, Hsenhsua, and 41 new combinations are proposed based on these findings.

Sputum bacterial microbiota signature as a surrogate for predicting disease progression of nontuberculous mycobacterial lung disease.

OBJECTIVES: Predicting progression of nontuberculous mycobacterial lung disease (NTM-LD) remains challenging. This study evaluated whether sputum bacterial microbiome diversity can be the biomarker and provide novel insights into related phenotypes and treatment timing. METHODS: We analyzed 126 sputum microbiomes of 126 patients with newly diagnosed NTM-LD due to Mycobacterium avium complex, M. abscessus complex, and M. kansasii between May 2020 and December 2021. Patients were followed for 2 years to determine their disease progression status. We identified consistently representative genera that differentiated the progressor and nonprogressor by using six methodologies. These genera were used to construct a prediction model using random forest with 5-fold cross validation. RESULTS: Disease progression occurred in 49 (38.6%) patients. Compared with nonprogressors, α-diversity was lower in the progressors. Significant compositional differences existed in the ß-diversity between groups (p=0.001). The prediction model for NTM-LD progression constructed using seven genera (Burkholderia, Pseudomonas, Sphingomonas, Candidatus Saccharibacteria, Phocaeicola, Pelomonas, and Phascolarctobacterium) with significantly differential abundance achieved an area under curve of 0.871. CONCLUSIONS: Identification of the composition of sputum bacterial microbiome facilitates prediction of the course of NTM-LD, and maybe used to develop precision treatment involving modulating the respiratory microbiome composition to ameliorate NTM-LD.

Molecular systematics of Dendrobium (Orchidaceae, Dendrobieae) from mainland Asia based on plastid and nuclear sequences.

Dendrobium is one of the three largest genera and presents some of the most intricate taxonomic problems in the family Orchidaceae. Based on five DNA markers and a broad sampling of Dendrobium and its relatives from mainland Asia (109 species), our results indicate that mainland Asia Dendrobium is divided into eight clades (with two unplaced species) that form polytomies along the spine of the cladogram. Both Dendrobium and Epigeneium are well supported as monophyletic, whereas sect. Dendrobium, sect. Densiflora, sect. Breviflores, sect. Holochrysa, are paraphyletic/polyphyletic. Many ignored phylogenetic relationships, such as the one of major clades formed by D. jenkinsii and D. lindleyi (two members of sect. Densiflora), the Aphyllum group, the Devonianum group, the Catenatum group, the Crepidatum group, and the Dendrobium moniliforme complex are well supported by both molecular and morphological evidence. Based on our data, we propose to broaden sect. Dendrobium to include sect. Stuposa, sect. Breviflores, and sect. Holochrysa and to establish a new section to accommodate D. jenkinsii and D. lindleyi. Our results indicated that it is preferable to use a broad generic concept of Dendrobium and to pursue an improved infrageneric classification at sectional level, taking into account both morphology and current molecular findings.

Usefulness of offline fractional exhaled nitric oxide measurements in the elderly asthmatic patients.

Fractional exhaled nitric oxide (FeNO) measurement is an important noninvasive tool for evaluating eosinophilic inflammation of the airways. However, measurement of online FeNO in elderly people may not be possible if they are unable to perform the expiratory maneuver. The offline FeNO test is an alternative method that obviates the need for the expiratory maneuver. Adult asthma patients were enrolled and assigned to one of two age groups: group 1, aged >65 years old (n = 29), and group 2, control group, aged 20-65 years old (n = 27). Online and offline FeNO measurements were performed for all patients. There were no significant differences in pulmonary function parameters (forced expiratory volume in 1 second [FEV1], percentage of predicted FEV1, and FEV1/forced vital capacity [FVC]) between the two groups but group 1 had significantly lower Asthma Control Test scores. There was a high correlation between online and offline FeNO levels (r = 0.908; p < 0.001). There was significantly higher mean FeNO level, regardless of online or offline measurements, in patients with poorly controlled asthma. Offline FeNO measurement was found to be a simple and convenient method of measuring FeNO in elderly patients and can be used as an objective guide for further management of eosinophilic inflammatory airway disease in this population.

Incremental yield of serial sputum examinations in the diagnosis of pulmonary tuberculosis in Taiwan: Findings of a pragmatic trial.

BACKGROUND: Presumptive tuberculosis (TB) cases commonly had two to three sputum examinations in Taiwan. The incremental yield of serial sputum examinations has not been assessed before. METHODS: In a pragmatic trial, presumptive TB patients with a frontline nucleic acid amplification test (NAAT) were classified as group A. Those without a frontline NAAT were randomized into group B frontline NAAT as intervention, and group C usual care. We investigated expected incremental yields and the number of examinations required for detection of one additional TB case from each serial sputum smear and culture. RESULTS: Of 6835 presumptive TB cases, 395 (5.8%) were smear positive for acid-fast bacilli, and 195 (2.8%) culture positive for M tuberculosis. The expected incremental yield from a third smear was 3.5% and examination of 1712 (95% credibility interval 586-4706) third smears was required to detected one additional TB case. Sensitivity of one smear with an NAAT in group B was 46.8% (95% confidence interval 32.1%-61.9%), and that of two smears in Group C 40.0% (95% confidence interval 25.7%-55.7%). The expected incremental yield from a third culture was 8.4%, and the number of third cultures required to detect one additional TB case was 394 (95% credibility interval 231-670). CONCLUSIONS: The incremental yield of the third sputum smear was negligible. It may be reasonable to perform an NAAT, smear and culture on the first specimen and culture alone on the second. The utility of the third serial culture for the detection of additional TB case is debatable.