Manual Blood Score

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Johnnie Johnson. Jon Brion. Joni Mitchell. Jonny Greenwood. Joshua Redman. Joshua Rifkin. Jule Styne. Julius Hemphill. Ken Burns. Kenny Garrett. Kevin Hays. Kremerata Baltica. Kronos Quartet. The whiskers above and below represent the range of values. Circles indicate outliers. B, Receiver operating characteristics ROC curves for distinguishing progressors and nonprogressors prior to sputum conversion at different intervals. Solid line indicates receiver operating characteristic curve distinguishing patients with active tuberculosis from patients without tuberculosis in the Brazil Active Screen Study cohort.

AUC indicates area under the curve. Each dot indicates a patient. Black line indicates linear regression. Each dot represents a patient. Color scale reflects log 10 day Total Glycolytic Activity Index score. D, Patients with ATB with 3-gene TB scores above the median had significantly higher likelihood of prolonged lung pathology as measured by positron emission tomography—computed tomography. Conflicts of interest comprise financial interests, activities, and relationships within the past 3 years including but not limited to employment, affiliation, grants or funding, consultancies, honoraria or payment, speaker's bureaus, stock ownership or options, expert testimony, royalties, donation of medical equipment, or patents planned, pending, or issued.

If you have no conflicts of interest, check "No potential conflicts of interest" in the box below. The information will be posted with your response. Not all submitted comments are published. Please see our commenting policy for details. Published online October 26, 6 :e The CTRC patients also had positron emission tomography—computed tomography scans at diagnosis, week 4, and week Analyses were performed from September to June This gene expression diagnostic approach should be considered for further validation and future implementation.

There are currently no commercially available tests, to our knowledge, that satisfy these criteria. Solid culture results take weeks to provide a diagnosis and liquid culture grown in the mycobacteria growth indicator tube MGIT still requires 7 to 10 days for a positive diagnosis and 42 days before negativity can be declared. Several host response-based transcriptional signatures have been described for diagnosis of ATB 13 - 20 and for anticipating progression from latent to active disease. Further, none of these host response-based signatures have been shown to be associated with the lung pathology during ATB to date.

We previously described a 3-gene signature derived and validated in a multicohort analysis of whole-blood transcriptome profiles in 14 independent cohorts from 11 countries that addresses many of these challenges. The 3-gene TB score was calculated for every sample described in this article and used for all statistical comparisons relevant to that sample.

The ACS cohort has been previously described by other groups. However, phenotypic information was only available for individuals. Therefore, 9 adolescents were excluded from further analysis. Of the individuals, 43 developed ATB during the study called progressors ; the remaining individuals are referred to as nonprogressors. Written informed consent was obtained from participants.

For adolescents, consent was obtained from parents or legal guardians of adolescents and written informed assent from each adolescent. Every patient with ATB identified during the study was notified and underwent treatment. Prisoners provided written informed consent in a private room without involvement of prison staff. Participation decisions were not reported to prison staff unless an individual was diagnosed with TB, in which case the diagnosis was reported, as mandated by Brazilian law, and free treatment was provided.

Between January and February , all inmates aged 18 years or older at these 2 prisons were recruited, of whom declined consent; the remaining inmates consented to participate and were enrolled. Individuals were not excluded from the study for any potential comorbidities. Participants demographic eTable 1 in the Supplement and clinical information was ascertained by a standard questionnaire eAppendix in the Supplement.

Two sputum samples were collected, including 1 spot sample after the interview and another the next morning, consistent with the WHO recommendations at the time. Smear microscopy Ziehl-Neelsen and solid culture were used to test for Mtb. Samples were decontaminated using the Petroff method and culturing was performed using modified Ogawa medium 23 - 25 and maintained for 60 days until it was considered negative.

Radiography was not available in the prisons, and a TB case was therefore defined as the presence of at least 1 positive culture test. In these prisons, previous studies found that 1. Their demographic eTable 1 in the Supplement and clinical information was ascertained by a standard questionnaire, and 2 sputum samples and whole-blood samples in PAXgene RNA tubes were collected. All ATB cases, defined as a positive sputum culture for Mtb, and consecutive controls that were sputum culture negative were included in gene expression analysis on a case to control ratio of 1 to 1.

The CTRC has been previously described by other groups. These adults include MGIT culture-positive patients with ATB, 21 healthy controls, and 17 patients with other lung diseases pneumonia or asthma.

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Biometrics were collected for these patients at the time of diagnosis prior to treatment initiation baseline , 1 week, 4 weeks, and 24 weeks study-defined end of treatment [EOT] for all patients eAppendix in the Supplement. The complete list of biometrics has been previously described. Patient clinical outcomes were not known at the time of reading. Sputum cultures were performed throughout the course of the study. Patients who completed therapy and had at least the last 2 consecutive sputum cultures at the EOT as negative were considered cured.

Seven patients with positive sputum test at the EOT were considered to have failed treatment. Two patients with EOT contaminated sputum cultures were considered unevaluable and were removed from the analysis. For the ACS, 17 our primary outcome was diagnosis of ATB, and the secondary outcome was association with progression from latent to active disease. Overall sensitivity and specificity values for diagnosis of ATB across the 3 cohorts were calculated by pooling true-positives, false-positives, true-negatives, and false-negatives from the 3 data sets.

The pooled values were then used to calculate sensitivity and specificity to give total sensitivity and total specificity. At The 3-gene TB score did not distinguish progressors from nonprogressors in samples collected more than 6 months prior Figure 1. Of the remaining 98 patients with ATB, 7 failed treatment. We compared the 3-gene TB score of the patients with ATB with that of healthy controls and patients with other lung diseases at the time of diagnosis.

Further, the 3-gene TB score at EOT continued to be higher in those with persistent lung inflammation compared with those who were radiologically clear at EOT eFigure 2 in the Supplement. We chose median instead of mean as it is robust to outliers. Collectively, across these 3 cohorts the 3-gene TB score had The score was significantly associated with the progression of individuals from LTBI to ATB 6 months prior to positive sputum test results.

Finally, the 3-gene TB score at baseline correlated with the severity of lung inflammation in patients with ATB. The patients with a higher 3-gene TB score at the time of diagnosis had persistent lung inflammation at the EOT. The 3-gene TB score satisfied the WHO TPP for a non—sputum-based triage test, 7 demonstrating its potential for systemic screening to identify those who should be tested further for confirming diagnosis of ATB.

There is an unmet need for noninvasive or minimally invasive biomarkers that can be used in clinical trials and during treatment for ATB as a proxy of lung pathology. The studies further observed that the 3-gene TB score at the time of diagnosis was significantly associated with 6-month radiological outcome. Patients with a 3-gene TB score higher than the median at time of diagnosis were 3 times more likely to have persistent lung inflammation.

The 3-gene TB score measured at the EOT was higher in those with ongoing lung inflammation, and accurately identified those who failed treatment as defined by a positive culture at the EOT.

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These results indicate that the 3-gene TB score may be useful in measuring lung pathology and monitoring treatment response because patients often cannot generate sputum after 6 months of treatment, and can be easily lost to follow-up when culture results take 40 to 60 days to declare negativity. Since its publication in , 18 the 3-gene TB score is now shown to distinguish patients with ATB from those with LTBI, other diseases, and healthy controls in 19 independent cohorts consisting of over individuals from 14 countries across all age groups children, adolescents, and adults with or without HIV coinfection.

To our knowledge, the 3-gene TB score is the first signature demonstrated to work in an active screen study, and to demonstrate that peripheral immune response can be a reasonable proxy for ongoing lung pathology during ATB. Collectively, these results show a substantial potential for the 3-gene TB score to monitor an individual with a latent Mtb infection through progression to and diagnosis of ATB and evaluate response to treatment. There are several limitations of our analysis. Because there is not yet a commercially available version of the test, we do not have a universal 3-gene TB score threshold for diagnosing ATB at the time.

Such a threshold can only be determined once the 3-gene TB score is ported to a point-of-care device. Second, in the BASS, use of solid media instead of liquid culture and lack of radiography would arguably misclassify a number of patients. However, such a misclassification would likely bias performance of the 3-gene TB score toward the null hypothesis, leading to underestimate of its specificity.

Third, only 7 of the individuals in the CTRC had failed treatment. Hence, further validation is required to better understand how closely the 3-gene TB score tracks with response to treatment. Fourth, the 3-gene TB score reliably identified progressors only up to 6 months prior to diagnosis, but not in samples beyond 6 months prior to diagnosis. Therefore, although the 3-gene TB score moderately correlated with lung pathology, it provides additional clinically useful information that no other biomarker to date provides.

Hence, an active screen study with larger sample size is required to better estimate specificity. Despite these limitations, the results of our analyses are consistent with previously published results in more than samples. The 3-gene TB score is associated with disease progression, performs well in active case finding, and tracks with treatment response and lung pathology. These performance characteristics make it a potential test for ruling out ATB and a strong candidate for monitoring the status of an individual over the course of infection, disease, and treatment.

Published: October 26, Author Contributions: Drs Warsinske and Khatri had full access to all of the data in the study and take responsibility for the integrity of the data and the accuracy of the data analysis. Drs Andrews and Khatri were co—senior authors. Drs Khatri and Sweeney are coinventors on the 3-gene TB score pending patent owned by Stanford University, which has been licensed for commercialization. Dr Malherbe reported receiving grants from the South African Medical Research Council during the conduct of the study and grants from the European and Developing Countries Clinical Trials Partnership outside the submitted work.

Dr Walzl reported receiving grants from the Bill and Melinda Gates Foundation during the conduct of the study; in addition, Dr Walzl has a patent to the United Kingdom patent office pending that is not related to the 3-gene TB signature. Dr Winter reported receiving grants from the Bill and Melinda Gates Foundation during the conduct of the study and outside the submitted work. Dr Sweeney is an employee and stockholder for Inflammatix, Inc; in addition, Dr Sweeney had a patent to methods of tuberculosis diagnosis pending and licensed.

Dr Andrews reported receiving grants from the National Institutes of Health, grants from the Bill and Melinda Gates Foundation, and grants from the Stanford Center for Clinical and Translational Education during the conduct of the study. Dr Khatri reported receiving grants from the Bill and Melinda Gates Foundation, personal fees from Vir Bio and grants from the National Institute of Allergy and Infectious Diseases, National Institutes of Health during the conduct of the study; and personal fees from Vir Bio and Inflammatix, Inc outside the submitted work; in addition, Dr Khatri had a patent to methods for diagnosis of tuberculosis pending and with royalties paid.