Volume 31 Issue 7
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CAO Shu Hui, CHEN Yan Qing, SUN Yong, LIU Yang, ZHENG Su Hua, ZHANG Zhi Guo, LI Chuan You. Screening of Serum Biomarkers for Distinguishing between Latent and Active Tuberculosis Using Proteome Microarray[J]. Biomedical and Environmental Sciences, 2018, 31(7): 515-526. doi: 10.3967/bes2018.069
Citation: CAO Shu Hui, CHEN Yan Qing, SUN Yong, LIU Yang, ZHENG Su Hua, ZHANG Zhi Guo, LI Chuan You. Screening of Serum Biomarkers for Distinguishing between Latent and Active Tuberculosis Using Proteome Microarray[J]. Biomedical and Environmental Sciences, 2018, 31(7): 515-526. doi: 10.3967/bes2018.069

Screening of Serum Biomarkers for Distinguishing between Latent and Active Tuberculosis Using Proteome Microarray

doi: 10.3967/bes2018.069
Funds:

the Natural Science Foundation of China 81470091

Beijing Municipal Administration of Hospitals Ascent Plan DFL20151501

More Information
  • Author Bio:

    CAO Shu Hui, male, born in 1977, PhD, majoring in bacteriology and immunology

  • Corresponding author: LI Chuan You, PhD, Professor, Tel:86-10-69545354;Fax:86-10-69546819, E-mail:lichuanyou6688@hotmail.com
  • Received Date: 2018-02-24
  • Accepted Date: 2018-06-11
  • Objective To identify potential serum biomarkers for distinguishing between latent tuberculosis infection (LTBI) and active tuberculosis (TB). Methods A proteome microarray containing 4, 262 antigens was used for screening serum biomarkers of 40 serum samples from patients with LTBI and active TB at the systems level. The interaction network and functional classification of differentially expressed antigens were analyzed using STRING 10.0 and the TB database, respectively. Enzyme-linked immunosorbent assays (ELISA) were used to validate candidate antigens further using 279 samples. The diagnostic performances of candidate antigens were evaluated by receiver operating characteristic curve (ROC) analysis. Both antigen combination and logistic regression analysis were used to improve diagnostic ability. Results Microarray results showed that levels of 152 Mycobacterium tuberculosis (Mtb)-antigen-specific IgG were significantly higher in active TB patients than in LTBI patients (P < 0.05), and these differentially expressed antigens showed stronger associations with each other and were involved in various biological processes. Eleven candidate antigens were further validated using ELISA and showed consistent results in microarray analysis. ROC analysis showed that antigens Rv2031c, Rv1408, and Rv2421c had higher areas under the curve (AUCs) of 0.8520, 0.8152, and 0.7970, respectively. In addition, both antigen combination and logistic regression analysis improved the diagnostic ability. Conclusion Several antigens have the potential to serve as serum biomarkers for discrimination between LTBI and active TB.
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Screening of Serum Biomarkers for Distinguishing between Latent and Active Tuberculosis Using Proteome Microarray

doi: 10.3967/bes2018.069
Funds:

the Natural Science Foundation of China 81470091

Beijing Municipal Administration of Hospitals Ascent Plan DFL20151501

  • Author Bio:

  • Corresponding author: LI Chuan You, PhD, Professor, Tel:86-10-69545354;Fax:86-10-69546819, E-mail:lichuanyou6688@hotmail.com

Abstract:  Objective To identify potential serum biomarkers for distinguishing between latent tuberculosis infection (LTBI) and active tuberculosis (TB). Methods A proteome microarray containing 4, 262 antigens was used for screening serum biomarkers of 40 serum samples from patients with LTBI and active TB at the systems level. The interaction network and functional classification of differentially expressed antigens were analyzed using STRING 10.0 and the TB database, respectively. Enzyme-linked immunosorbent assays (ELISA) were used to validate candidate antigens further using 279 samples. The diagnostic performances of candidate antigens were evaluated by receiver operating characteristic curve (ROC) analysis. Both antigen combination and logistic regression analysis were used to improve diagnostic ability. Results Microarray results showed that levels of 152 Mycobacterium tuberculosis (Mtb)-antigen-specific IgG were significantly higher in active TB patients than in LTBI patients (P < 0.05), and these differentially expressed antigens showed stronger associations with each other and were involved in various biological processes. Eleven candidate antigens were further validated using ELISA and showed consistent results in microarray analysis. ROC analysis showed that antigens Rv2031c, Rv1408, and Rv2421c had higher areas under the curve (AUCs) of 0.8520, 0.8152, and 0.7970, respectively. In addition, both antigen combination and logistic regression analysis improved the diagnostic ability. Conclusion Several antigens have the potential to serve as serum biomarkers for discrimination between LTBI and active TB.

CAO Shu Hui, CHEN Yan Qing, SUN Yong, LIU Yang, ZHENG Su Hua, ZHANG Zhi Guo, LI Chuan You. Screening of Serum Biomarkers for Distinguishing between Latent and Active Tuberculosis Using Proteome Microarray[J]. Biomedical and Environmental Sciences, 2018, 31(7): 515-526. doi: 10.3967/bes2018.069
Citation: CAO Shu Hui, CHEN Yan Qing, SUN Yong, LIU Yang, ZHENG Su Hua, ZHANG Zhi Guo, LI Chuan You. Screening of Serum Biomarkers for Distinguishing between Latent and Active Tuberculosis Using Proteome Microarray[J]. Biomedical and Environmental Sciences, 2018, 31(7): 515-526. doi: 10.3967/bes2018.069
  • Tuberculosis (TB) is a major cause of morbidity and mortality due to infectious diseases worldwide. According to data from the World Health Organization, approximately 10.4 million new cases of TB occurred, and 1.8 million people died from TB worldwide in 2015[1].Among individuals infected with Mycobacterium tuberculosis (Mtb), approximately 5% develop active TB over 2-5 years, whereas the remaining 95% develop latent TB infection (LTBI) without clinical, radiological, or bacteriological evidence of active TB, but show immune responses to Mtb infection[2-4]. Currently, nearly one-third of the world's population have LTBI. However, these individuals have a 10% chance of reactivation of the latent infection, which may ultimately progress to active TB during their lifetimes. So, individuals with LTBI are potential reservoirs of active TB[1, 5]. Therefore, identification of LTBI is urgent for decreasing the risk of developing active TB, especially in certain high-risk populations. At the same time, early diagnosis followed by treatment of active TB is the most effective method for controlling TB epidemics.

    Currently, early detection of individuals with Mtb infection mainly relies on the host's cell-mediated immune response to pathogen-specific antigens. Examples of such tests are the in vivo tuberculin skin test (TST) and in vitro interferon gamma release assays (IGRAs)[6]. The TST is based on the use of a purified protein derivative (PPD), which is isolated from culture filtrates of the tubercle bacilli. Specific antigens in the PPD can elicit a cross-reaction with other mycobacterium species, including the BCG vaccine, thereby showing poor specificity[7, 8]. IGRAs are based on the measurement of interferon gamma (IFN-γ) released from T-lymphocytes exposed to Mtb antigens, which is easier, faster, and more sensitive than the TST. Hence, IGRAs are commonly used by clinical physicians[9].In particular, IGRAs can differentiate between Mtb infection and BCG vaccination and exclude interference of most non-tuberculosis mycobacterium. However, neither of these tests distinguish between active TB and LTBI[10, 11]. Thus, there is an urgent requirement for rapid and simple tests that indicate disease status.

    Serological assays have a long history and have been widely used for the diagnosis of several infectious diseases because of convenient sampling, low cost, easy procedure, and short turnaround time. Currently, several serological commercial kits based on antibody detection for TB identification have been developed, such as Linonex TB kits, InBios Active TbDetect IgG ELISA, IBL M. tuberculosis IgG ELISA, and Anda Biologicals TB ELISA[12, 13]; however, these kits do not yield accurate results, which limits their clinical use. Since serological detection in a simple dipstick format can incorporate various antigens, investigation of serum biomarkers for both diagnosing TB and determining disease status is a worthwhile endeavor.

    The present study aimed to screen potential serum biomarkers for distinguishing between LTBI and active TB at the system level using the Mtb proteome microarray containing 4, 262 antigens. We observed that the levels of 152 Mtb antigen-specific IgG antibodies were higher in the active TB group than in the LTBI group, and these antigens showed both stronger associations with each other and involvement in various biological processes. We further validated 11 candidate antigens with ELISA and used ROC analysis to evaluate the diagnostic performance of these antigens. In addition, both antigen combination and the logistic regression model exhibited better ability than any single antigen for distinguishing between LTBI and active TB patients, which provides the necessary groundwork required for establishing a new method for determining TB disease status.

  • This study was approved by the ethics committee of the Beijing Chest Hospital, Capital Medical University (Beijing, China) in accordance with the Declaration of Helsinki. Written informed consent for the use of samples was obtained from all participants.

    From June 2014 to December 2015, 319 subjects were included in this study and divided into three groups: active TB patients group (active TB group), LTBI subjects group (LTBI group), and healthy control group (HC group). Active TB patients were recruited from the Beijing Chest Hospital; LTBI subjects and healthy controls were recruited from the Institute of Tuberculosis Prevention and Control of Changping district (Beijing, China). Active TB patients were diagnosed according to the guidelines of 'Pulmonary TB Diagnosis and Therapy' authored by the Tuberculosis Branch Association of the Chinese Medical Association, which included abnormal chest radiography, positive Ziehl-Neelsen-stained sputum smear and/or sputum culture, and the presence of TB clinical symptoms. All active TB patients had been treated for no longer than 2 weeks at the time of their blood sampling. Every LTBI subject showed a positive response to both the TST (> 10 mm) and the T-spot TB assay, whereas healthy controls showed negative responses to both the TST (< 5 mm) and the T-spot TB assay. Both LTBI subjects and healthy controls were free of all clinical symptoms or abnormal chest radiographic findings, which are indicative of active TB. In addition, all subjects with HIV and those treated with immunosuppressive medications were excluded. Assays for TST, anti-HIV, and T-spot were performed using TB-PPD (Sanroadbio, Beijing, China), ELISA reagent kit (Livzon Diagnostic Inc., Zhuhai, China), and T-spot TB reagent kit (Oxford Immunotec, Abingdon, UK), respectively, according to manufacturers' instructions.

  • Three milliliters peripheral venous blood was collected. Sera were obtained by centrifugation at 1, 509 × g for 10 min. Then, 400 μL aliquots of sera were aliquoted into 0.5 mL clean eppendorf tubes and stored at -80 ℃ until further use.

  • The Mtb proteome microarrays used in our study were purchased from BCBIO (Guangzhou, China). Microarrays comprise 3, 829 proteins encoded by genes of H37Rv (Mtb standard strain) and 433 proteins encoded by genes of CDC1551 (pathogenic strain), which were spotted in duplicates on polymer slides (polymer-slide H, CapitalBio)[14]. Expression of these GST-tagged recombinant proteins was verified in a Saccharomyces cerevisiae expression system. In addition, positive controls of human IgG and IgM, and negative controls of bovine serum albumin (BSA) were also printed in duplicates on the microarrays.

    First, microarrays were blocked for 1 h at room temperature with agitation in blocking buffer [3% BSA in 1× TBST (Tris-buffered saline plus 0.1% Tween 20 detergent, pH 7.4)]. Three milliliters of serum samples (1:50 dilution in TBST) were overlaid onto protein microarrays and then incubated at room temperature for 3 h. After washing three times for 5 min each with TBST, microarrays were probed with goat anti-human IgG conjugated with Cy3 (Jackson Laboratory, PA, USA) diluted 1:1, 000 in TBST and incubated in a dark room at room temperature for 45 min. Next, microarrays were washed three times with TBST and then twice with double-distilled water in the dark. Finally, microarrays were dried in a SlideWasher (CapitalBio, Beijing, China) at room temperature, and scanned at 532 nm in a GenePix 4200A (Molecular Devices, CA, USA). Fluorescence data were analyzed using GenePix Pro 6.0 software (Molecular Devices, CA, USA).

  • Mtb antigens were purchased from BCBIO (Guangzhou, China). Each well of the 96-well flat-bottom plates (Thermo, Copenhagen, Denmark) was coated with 100 μL of 5 μg/mL individual antigens in coating buffer (0.1 mol/L carbonate/bicarbonate, pH 9.6) and stored at 4 ℃ overnight. Plates were washed three times with phosphate-buffered saline containing 0.05% Tween 20 (PBST) for 5 min each, and then blocked with 200 μL/well PBST containing 1% bovine serum albumin (PBST-B) at room temperature for 2 h in a humidified chamber. After washing three times with PBST, 100 μL of serum samples diluted 1:400 in PBST-B were added to antigen-coated wells and incubated at room temperature for 1 h. Subsequently, plates were washed five times, 100 μL/well of horseradish peroxidase-conjugated anti-human IgG antibody (CWBiotech, Beijing, China) diluted 1:30, 000 in PBST-B was added, and incubated at room temperature for 1 h. Next, plates were washed five times, and developed using 100 μL/well of TMB substrate (BD, NJ, USA) in a dark room for 10 to 15 min until visible color was apparent; the reaction was stopped using 50 μL/well of 2 mol/L sulfuric acid. Finally, optical density was determined at 450 nm using an automatic microplate reader (Perlong, Beijing, China).

  • Protein microarray data were obtained using GenePix Pro 6.0. The background signal of the raw data was corrected to eliminate variations between arrays, followed by normalization using the limma package of the R programming language (http://www.protein-microarray.com). This pre-processed data was used for further analysis. Differences in microarray data between active TB and LTBI were analyzed with a t-test for two independent samples. Fold change was defined as the log-transformed ratio of active TB to LTBI. Hierarchical cluster, obtained using R statistical software, was represented by log-transformed values. Protein-protein interaction networks were analyzed using STRING 10.0 (http://string-db.org/) and visualized with a confidence level of 0.25 as the parameter setting.

  • Differences in age and gender among the three groups were analyzed using the Kruskal-Wallis test and Chi-square test, respectively. Differences in optical density values among the three groups were analyzed using a one-way analysis of variance with Bonferroni comparisons. Differences between paired proportions were analyzed with McNemar's test. P values less than 0.05 were considered statistically significant.

    For each antigen, diagnostic performance was examined using ROC curve analysis to determine AUCs and their 95% confidence intervals (CI). The cutoff level of each antigen was determined at the maximum Youden's index (YI = Sensitivity + Specificity − 1).

    For analyzing antigen combinations, we selected the top three antigens based on AUCs and assigned a score of 0 or 1 to each antigen result depending on whether it was below or above the cutoff level for the antigen; a positive responder was determined if any two antigen tests were positive with total score ≥ 2.

    For logistic regression analysis, we selected 11 antigens as factors to construct the logistic regression model, including 64 LTBI individuals, and 62 active TB patients as the training set. We used a stepwise forward selection procedure to determine candidate biomarkers that contributed maximally to distinguish between LTBI and active TB. Stepwise procedures were guided by an F value probability of 0.05 for inclusion, and 0.10 for exclusion. The leave-one-out method was used to evaluate the model's generalization ability. Coefficients for antigens included in the final step were calculated. The accuracy of the established regression model was evaluated by ROC analysis. We selected independently 29 LTBI, and 30 active TB patients as the validation set.

    Statistical analyses were performed using GraphPad Prism V5.0 (GraphPad Software, CA, USA) and SPSS version 17.0 (IBM, NY, USA).

  • In total, 319 subjects were enrolled in the final analysis, which consisted of 94 healthy controls, 113 LTBI subjects, and 112 active TB patients. Both demographic and clinical characteristics of all subjects in this study are shown in Table 1. In the healthy control, LTBI, and active TB groups, median ages were 40.5 years (range, 18-65 years), 45 years (range, 20-69 years), and 42 years (range, 18- 65 years), respectively; the male to female ratios were 44/50, 46/67, and 53/59, respectively, and there were no statistical differences among them with respect to age (P = 0.268) or gender (P = 0.600). For the T-spot TB assay, results for healthy controls and LTBI subjects were negative and positive, respectively. Furthermore, the T-spot TB assay detected 89 out of 93 patients with active TB, indicating a sensitivity of 95.7%. In addition, none of the subjects was infected with HIV or was treated with immunosuppressive medications.

    Items Health Controls LTBI Subjects Active TB Patients
    Total number 94 113 112
    Median age (range), years 40.5 (18-65) 45 (20-69) 42 (18-65)
    Gender, male/female 44/50 46/67 53/59
    Abnormal chest radiograph, n (%) 0 (0) 0 (0) 112 (100.0)
    Bacteria positive, n (%) nd nd 112 (100.0)
    TST results
      Induration < 5 mm, n (%) 94 (100.0) 0 (0) nd
      Induration 5-10 mm, n (%) 0 (0.0) 0 (0) nd
      Induration > 10 mm, n (%) 0 (0.0) 113 (100.0) nd
    T-SPOT results
      Positive, n (%) 0 (0) 113 (100.0) 89 (79.5)
      Negative, n (%) 94 (100.0) 0 (0) 4 (3.6)
      Unknown, n (%) 0 (0) 0 (0) 19 (16.9)
    No immunosuppression, n (%) 94 (100.0) 113 (100.0) 112 (100.0)
    HIV-negative, n (%) 94 (100.0) 113 (100.0) 112 (100.0)
    Note. n, number of subjects; nd, not done.

    Table 1.  Demographic Characteristics and Clinical Details of Study Subjects

    Among these subjects, we randomly selected 20 each of LTBI and active TB patients as the screening population (screening using microarray), and 94 healthy controls, 93 LTBI individuals, and 92 active TB patients as the validation population (validation using indirect ELISA).

  • After normalizing and excluding the outlier microarrays, 18 and 19 microarray results were obtained from LTBI subjects and active TB patients, respectively. We further revealed that the concentrations of 152 Mtb antigen-specific IgG antibodies were significantly higher in active TB patients than in LTBI subjects based on a P value of < 0.05 and a fold-change (Log2) of > 0.18 (Supplementary Table S1, available in www.besjournal.com). Moreover, the differences in IgG reactivity against 152 antigens in each individual serum from LTBI individuals and active TB patients were subjected to hierarchical clustering, which distinguished between LTBI and active TB patients (Figure 1).

    Name L1 L2 L3 L5 L6 L7 L8 L9 L10 L11 L12 L13 L14 L15 L16 L18 L19 L20 A1 A2 A3 A4 A5 A6 A7 A8 A9 A10 A11 A12 A13 A14 A15 A16 A18 A19 A20 Fold change (Log2) P value
    Rv0379 8.3491782 6.2903204 5.8762276 6.4405914 8.0420408 8.0936946 7.2329981 8.2203904 6.3279363 6.1181267 6.0957997 8.1158571 8.3239144 7.2287831 7.8648127 6.7331704 8.9338898 6.6794144 7.2813882 9.6584977 9.926081 8.978122 7.2564087 8.4931519 8.1520579 9.4238362 10.673998 8.0820386 10.70095 10.954123 11.99734 8.8251697 8.4082266 12.650203 13.758344 10.723222 8.730426 0.417770064 7.77E-06
    Rv3841 10.242245 7.5800733 7.3498213 7.9971314 5.769736 5.7702772 6.5681823 8.5151497 7.9835148 7.5627985 7.1648398 7.7810613 6.2780138 6.0762377 7.4515165 7.3758343 9.9470853 7.7492584 8.6136389 7.8690714 12.044714 8.8265217 7.277246 10.129875 10.998924 11.232083 9.7009306 11.393826 7.3199922 9.9914507 10.082166 7.4819171 11.212392 13.575217 10.417595 9.4470854 12.577226 0.414755188 1.19E-05
    Rv3190c 7.3733508 10.413173 8.2969942 9.3483153 7.211019 7.0578386 9.8147715 10.394686 8.8763918 6.5270723 8.0658541 7.6952589 8.5519846 8.7706159 9.9645737 8.847094 9.2181983 7.0162382 10.460602 12.296939 12.081969 11.391606 8.7558325 11.669257 10.506195 10.830471 12.162605 11.327933 11.515915 10.767905 10.758721 12.162605 10.431843 11.937504 12.879088 11.866002 11.619375 0.411458656 2.56E-09
    Rv1483 7.5987427 7.8879505 8.2757547 8.3816482 8.5845724 8.9781705 8.2235729 8.5734425 8.6782129 8.9804309 9.2148661 7.7636504 9.0467124 8.1103122 8.8035498 8.0204223 8.2460404 7.7236564 8.4237559 13.38782 9.5768292 12.223228 8.0706537 12.774646 11.669257 12.223228 9.7485006 11.717555 12.78021 10.183723 9.5593928 13.758344 13.055373 8.8211599 10.932973 9.0805367 12.78021 0.402224709 2.14E-06
    Rv2421c 7.6369405 6.8505005 6.7417458 6.6739014 7.4019211 8.1896234 8.3399418 6.1467534 6.5496445 7.3007739 6.4734575 7.7981635 8.1674519 8.2403259 5.8872593 6.7898572 7.4638011 6.3679218 7.3717442 10.810575 7.8538797 10.761906 6.9394494 11.900448 11.818093 9.4977798 8.2811393 10.249528 10.617704 7.9095422 7.7042669 11.397897 11.964498 8.0913551 7.6245142 8.1119663 10.257515 0.39568212 1.29E-05
    Rv2928 7.9087819 6.2613301 6.4972082 6.3740435 7.89223 6.0244463 6.2438668 6.1479758 6.1387066 5.5851077 5.8009375 7.2937137 6.1143934 5.9447971 8.93555 7.2742685 7.8035326 6.3854338 8.200557 9.8401148 7.7653934 8.7575554 9.1251269 9.612247 9.0481595 8.7989169 7.7661579 8.8489326 8.7617273 7.4317456 8.9788697 9.1823419 8.6317904 7.9098749 8.2401753 7.8592369 9.797653 0.370036401 1.75E-08
    Rv2031c 6.8208515 6.0546301 6.6727487 6.2191481 7.613001 5.9213299 5.929078 6.8054547 6.1683855 5.8917976 5.8289113 6.8902455 6.0886513 6.3874929 6.3016622 6.7289734 6.8315949 6.6077193 7.8176054 6.9309494 9.8984299 7.2827637 7.9252977 7.2249701 6.9253877 7.1038611 8.0433612 8.6644635 7.3034878 8.5580843 8.0124233 8.7103341 6.2055835 9.753778 9.3805067 8.0939956 13.38782 0.363653005 4.22E-05
    Rv1474c 8.3250528 6.1444028 6.0719942 6.7559549 7.1513953 6.9446525 8.5620913 8.147383 6.2801309 6.2463967 6.0483955 8.2758866 6.8428462 7.011195 9.5252871 6.5430932 8.23404 6.6791142 8.5625781 12.398564 8.8342358 8.5431174 6.6355925 12.414698 12.340987 9.3325009 7.9963771 9.7844711 8.4760195 8.1349251 9.1904823 9.0789581 11.158235 7.7818309 9.0035605 7.6839698 8.4922127 0.360116396 4.62E-05
    Rv2906c 5.670648 6.327279 6.3668663 6.7221993 6.6263945 7.0177027 5.7891352 8.131681 6.2747419 6.4872235 6.8955154 5.6799853 8.3513581 8.9370752 7.7356504 6.5935761 6.5656697 6.4914207 8.6584187 8.0306532 8.0415102 10.803165 7.2749102 10.505461 8.357535 9.4230551 9.1826597 8.5538826 8.7365247 7.788431 8.5843132 8.7832237 7.6389762 9.4046104 9.2268572 9.3663243 7.646153 0.358525308 1.49E-07
    Rv1498A 7.7376025 7.0180888 6.8674103 7.7690789 8.6246687 8.6905627 7.1659917 7.8546747 7.57535 7.0832234 6.9580158 7.5880341 8.7283271 9.1932939 7.1026245 6.5574967 8.5936079 7.0613432 8.0317736 10.494522 9.4296979 10.767905 8.0355265 9.5422647 9.1108183 10.512899 10.311709 12.024334 8.4527651 9.8887725 9.6757966 10.835817 9.0368968 11.063385 9.8706611 9.8981928 9.8207884 0.357088806 1.43E-08
    Rv1337 8.5782554 6.2727234 5.5816801 6.389916 7.5152048 7.206287 7.8048974 8.5521464 6.2358644 6.1544031 6.1610989 8.1035621 7.6450087 7.0034073 8.6174032 6.00422 8.9259271 5.9270756 8.203378 9.7051384 8.9904078 9.6792579 8.3792888 8.3175188 8.6804047 9.7072596 9.021254 9.0869422 10.434739 9.7104059 9.0896238 10.136697 7.8915744 9.8101172 8.9776426 9.1188848 8.3546608 0.351451617 2.35E-07
    Rv3324c 7.4727605 7.0726695 6.7347435 7.494265 7.0622932 7.6004602 6.4318588 7.9676188 7.4415174 7.1103562 7.6438013 7.0271232 7.8107834 9.1175295 6.60658 6.7754203 9.1467135 6.6200129 7.5476951 9.57369 11.669257 10.319612 6.5313228 9.8938929 9.9814129 9.5191524 9.8728204 9.0152162 8.4757591 8.6581222 9.3206183 9.5126366 10.166692 9.6528761 9.3873205 9.7354009 8.8982876 0.338792973 1.73E-07
    Rv3899c 10.878719 8.9422482 9.3892025 8.8869599 8.6031849 6.534539 10.665886 10.60589 8.9458764 9.4684363 10.408274 9.5851356 6.2129768 5.9799666 8.9782375 8.7376968 9.661131 10.605242 10.944664 11.287995 10.814208 10.351259 8.9345342 11.98143 14.476539 10.024501 12.879614 10.756596 10.58048 11.693095 10.863784 11.182093 14.476539 11.211205 11.533673 11.353532 10.744906 0.327969314 1.04E-05
    Rv3297 7.7813994 5.6182535 6.1052333 5.4971678 6.9399519 7.9138471 7.4582257 7.483583 5.8763718 5.8471977 5.8221673 7.8600523 8.1197832 7.8561676 7.4640846 5.5979532 8.2422007 6.0720134 8.0403314 8.8512019 8.3494275 8.2443216 9.5103595 8.6813129 8.6925627 8.5906839 8.5278079 8.4594752 8.0076713 9.2347291 8.2846653 8.784727 8.6614981 8.456184 8.3027635 8.3821577 9.1430675 0.323521261 5.32E-07
    Rv2097c 7.1916979 8.0109329 7.1209251 8.2490101 7.7300946 7.8408914 6.6585778 9.1217991 7.7163833 6.6072822 7.1135338 7.5074509 8.0719083 6.470887 8.5686616 7.3563087 8.8263803 6.9107803 9.584024 9.8869939 8.6073539 9.4928411 7.6556056 10.170391 10.137253 9.3796335 9.376369 12.780565 9.0023389 7.4385149 9.2407789 10.19675 10.422212 8.7537607 8.2546144 8.990879 9.8349583 0.30866537 1.64E-06
    Rv0494 7.9871333 8.2065344 7.7069909 8.1708896 6.8158618 7.3528225 7.8903556 8.4978053 8.1364095 7.6044947 8.0957332 7.6724274 7.8731396 8.2413668 8.0611209 8.2219168 7.9183471 8.4545126 8.2987061 10.240039 12.847623 8.4619199 9.1671807 9.6784715 9.5325174 9.6693619 10.235642 9.524654 11.665861 9.0663128 9.7529722 10.269072 9.4578679 8.8331852 9.3381832 9.4858602 9.9577551 0.298200616 1.64E-07
    Rv0187 8.8232853 7.4450685 6.2496276 8.3954547 7.4291489 9.9407169 7.0852199 9.9531627 7.0619347 7.1590189 6.6333529 8.1892439 8.6919428 7.6648694 8.046311 7.8490522 8.8566171 6.8448879 8.4758665 10.68926 9.8569648 9.4281083 8.4161572 9.7596431 10.22339 8.9195598 9.0665249 9.9529185 10.366093 10.262419 9.6431775 9.8977508 10.822424 9.0129456 9.7000577 9.4570823 9.9841484 0.292062114 7.31E-07
    Rv0597c 11.471435 9.4059438 9.3330073 8.8470712 7.6300437 6.0939683 12.497314 10.228972 9.1380965 8.9671398 8.2183149 11.583256 6.7136558 8.1024876 9.0254082 8.8890505 10.972541 10.752485 11.052981 12.10235 11.008002 11.282732 9.0388019 11.567553 13.38782 10.274864 11.764286 10.562024 10.681328 12.49461 11.068791 11.237638 12.5251 11.764286 11.852338 11.371153 11.035079 0.286161815 6.11E-05
    Rv1815 8.4231501 6.0931732 6.4466628 6.4020118 7.5803447 8.083283 8.0265984 7.7413293 6.2309725 5.8337724 6.0218168 7.6145292 7.7323708 9.2008373 7.9677197 7.0065187 9.4536961 6.6348314 9.1040157 10.081743 8.2111263 9.4701028 7.7411365 9.3235513 9.4654895 9.0096934 9.5749866 8.9370249 8.5939845 8.1308778 8.3638136 9.0004057 8.9712766 9.2722326 8.7338513 8.937261 9.5098547 0.285274474 3.81E-06
    Rv2632c 7.2609725 6.2786847 6.1796287 6.7815145 7.2408685 7.409093 6.0971735 7.7032662 6.3827642 5.4731893 5.9219602 6.4990705 7.2025417 7.7612409 8.0567929 6.8574841 6.8046469 6.5785487 8.2931632 8.335863 11.453677 7.9710465 7.4257194 8.904729 8.6462559 8.2870977 8.4169069 7.6898417 7.7675221 7.1668065 8.143752 8.3588517 8.9237831 7.9515688 7.113221 8.0718429 8.2820636 0.281975391 2.08E-06
    Rv3743c 6.6502233 6.024088 6.722078 5.9443692 7.9789893 7.7669555 6.8206562 7.5828474 5.9850755 5.9744258 6.1048917 7.0058925 7.0826412 6.8321064 6.1532483 6.5699699 6.4516148 6.8053779 7.0507215 8.1494925 8.1564109 7.8132082 8.0921705 8.5095688 8.4167333 8.2885085 7.683132 8.3211327 8.2282789 8.9239248 7.897236 8.4134169 8.3943236 7.5448296 7.7250497 7.4792636 8.9421417 0.276704901 4.67E-09
    Rv0062 6.7147166 5.9921393 6.394957 5.9986691 7.8129309 7.9533142 6.8964188 6.3557954 6.0632925 6.1147847 6.1080683 6.7793326 8.3340954 7.8081895 6.7701384 6.2036511 6.7620538 5.9735531 7.6008308 9.4143069 6.6935892 8.1818485 9.1837559 9.2126708 8.9741816 8.4629102 7.2104237 8.5431751 8.5173887 6.7672277 6.9082414 9.0497581 8.3784739 7.1979036 7.6183087 7.0567969 9.6835929 0.275617138 1.08E-05
    Rv2499c 8.2967935 6.8018474 6.7430886 6.9928632 8.179409 8.0111114 6.7178617 7.693924 6.9521883 6.4610722 6.9063655 7.7802629 8.0578754 8.5629183 7.3442559 6.8127903 8.573419 6.8663219 8.1003876 9.4414948 8.8943539 9.3702005 7.8098731 9.2888405 8.9578941 9.0445844 9.3833262 8.9377117 9.1010013 9.1895958 9.1482214 8.3040482 8.9831078 9.2442711 9.2939733 9.2481935 9.1338934 0.275352585 4.98E-09
    Rv3625c 7.0110514 7.9440728 6.4800295 6.5230113 7.478578 8.2572199 7.7152385 8.0681928 7.6652661 7.1307919 8.2034697 7.0628218 8.2629301 8.8386944 8.1335389 7.057611 7.8035121 6.9790941 7.9582615 10.077463 9.162567 8.9320983 9.7114333 8.9946485 9.723673 9.5021459 10.174634 8.6894632 7.3940609 8.7478567 9.4020881 10.413173 8.0684958 9.6342418 9.4954779 9.14411 8.6571969 0.269997903 7.37E-08
    Rv2716 6.5895907 6.2082826 6.0436056 5.8271377 7.6110127 7.889531 6.0411936 8.1574845 6.2535656 5.8487676 5.9923778 6.2681868 7.8734922 8.0346158 7.9293128 6.7061748 7.0891621 6.8624596 8.5502686 9.6439194 7.1902022 8.0879192 7.4434023 8.7783181 8.4874932 8.608162 7.3825463 8.0779459 8.6618287 7.1893454 7.9558537 9.3019822 8.7243654 7.9429808 8.1339035 7.4556365 9.1993011 0.269758356 2.70E-06
    Rv2860c 8.1169528 8.0432543 7.1598019 7.6600741 6.6091469 6.1371048 7.2269755 8.4850046 7.1185467 7.381858 7.2022153 7.9737581 6.6040277 6.2728577 7.8288071 7.1824611 7.9868769 6.4052158 8.549173 9.4045547 8.8484038 8.7673414 6.9505866 9.2501628 8.3369416 8.9117892 9.088763 8.3585195 8.4555793 8.601775 9.3887881 9.2586659 8.9288596 8.8172791 9.2050378 8.6228253 9.3185055 0.268484786 1.80E-08
    Rv2026c 9.7893241 6.5708104 7.0704452 6.2262416 8.8888402 9.3426826 8.5995939 9.7759013 6.5333667 6.1847363 6.1353623 8.1832736 6.4721581 6.5966895 9.1808346 7.0119125 9.6262775 7.1293906 10.637107 10.755129 8.2382338 8.8604758 8.0007254 10.567584 12.296939 8.7390976 8.0145894 10.470983 9.112491 8.2285784 8.1214479 9.9312656 11.732927 7.9466698 8.4184455 8.3935577 8.3845358 0.266150071 0.00076452
    Rv2903c 6.4740465 7.5415281 7.8384415 7.6180486 7.695041 5.8078409 8.0008196 7.8196254 7.4201731 6.8487366 7.064432 6.4790112 6.1526033 5.9442504 6.964646 7.6029363 7.0539874 7.854384 8.2074165 7.3605666 12.223228 7.4224495 8.57938 9.5238404 9.5327777 10.482553 7.955414 7.7854233 6.8567086 7.7370164 8.8872102 7.601863 9.9645737 8.5866768 9.0159117 7.572625 7.3540316 0.26558796 0.00014869
    Rv0861c 7.6315587 6.4010965 6.3577009 7.2350981 8.4332142 6.4675745 7.6684342 8.3030484 6.5684655 6.2587724 6.5173718 7.775154 6.8966653 6.4369037 8.4418728 7.0757974 7.8848781 6.0674394 9.520667 9.2285455 8.4472062 9.1260901 9.315057 7.9824904 8.1315759 7.2711375 9.2470647 8.1281469 8.3540141 7.9686188 9.2823379 7.8387866 8.157792 8.6382553 9.6073351 8.2044902 8.1450955 0.262396123 7.95E-07
    Rv0214 7.4968379 8.3124023 7.0341622 7.4474096 8.2937944 7.8774911 6.4686333 7.7419802 7.4595382 6.9648769 7.5619527 8.276585 7.1494417 8.9108522 7.4084538 8.207783 9.9799655 6.9732524 8.7581651 9.5484941 8.7086059 8.6210244 7.4967762 10.102111 9.3296245 9.095828 9.7605316 7.8081049 9.4653562 7.7522881 10.020711 11.149624 9.2800702 10.555126 11.337877 9.7919245 8.016326 0.261529684 1.38E-05
    Rv3601c 6.2786075 6.4866139 6.8435146 6.181057 7.4461907 6.0263364 6.508403 7.0120404 6.4298393 6.1699266 6.2128521 6.3353354 5.9976149 6.1597763 6.6822579 6.5863422 6.1734633 5.9370139 7.2831032 7.9948957 6.7163674 7.9177776 8.1484875 8.3650539 8.3482776 8.2202079 7.2062897 7.9046657 7.8975491 6.1059504 6.9390399 8.0021816 8.0554329 7.5051446 6.7247675 7.7839664 8.4773062 0.256489902 4.33E-08
    Rv2198c 9.2065065 8.5064888 6.4155027 8.296374 7.5092836 7.2528871 10.77607 9.4275278 9.2789421 8.1517437 6.5672193 7.9058539 6.6935122 6.2754589 10.122887 6.3599007 8.9464497 6.18597 8.5010914 11.165662 8.9154972 9.2031566 9.6294888 7.6001058 9.4856845 11.393826 8.2382769 9.3291162 10.080478 11.274624 8.8713933 9.8037384 8.7281088 11.648793 8.9742668 8.7506757 9.4095968 0.253163925 0.00051299
    Rv0242c 8.0613881 10.388796 9.6181397 9.8321169 11.813108 11.524138 7.2911073 9.8830863 10.906549 9.5971805 9.6958842 10.141244 9.0774726 6.5972797 9.6711266 10.069176 7.949697 10.349247 9.8401479 12.51965 10.998729 11.944063 7.8868472 10.86947 10.66911 11.399324 12.650203 11.25335 11.746973 12.78021 11.136211 12.78021 10.812603 12.10235 11.781031 11.909289 11.764286 0.252337277 5.58E-05
    Rv2572c 7.8929393 5.8389698 6.145203 5.7955342 7.3372949 8.0671714 7.4400066 6.7497931 6.0701611 5.660125 5.5252968 7.8921711 8.4545508 8.3395317 7.0804854 6.1421819 7.5919402 5.9604826 6.9428703 8.2239401 8.0051793 7.7760407 8.2794656 8.7749214 8.3602048 8.3528435 8.2516087 8.1777718 8.1315436 7.9067117 7.9295084 8.7200978 8.4554076 8.1696396 7.7241847 8.2801359 9.2164675 0.250415518 2.14E-05
    Rv2804c 5.8782379 5.9547963 6.4388239 6.3029923 6.740037 7.3794842 5.8292382 6.2594265 5.6567906 6.6151909 6.8764638 5.7222571 7.7870073 7.9384992 6.2951146 6.8319253 5.9362176 6.8520924 6.9389168 7.5282056 7.6874411 8.8007754 7.1162751 8.1864103 7.7343335 8.4330276 7.7251245 8.2021053 7.4289663 7.0097031 7.5459822 7.7878762 7.9252644 7.9391026 7.528989 8.1201146 7.5272584 0.249304099 2.58E-07
    Rv1725c 7.0409249 7.9761689 6.8949163 7.818675 5.5469045 5.8108319 7.1472265 8.1944582 7.5521226 7.7824014 7.333382 7.0504715 5.5809189 5.7629281 8.4615178 6.1596178 7.1249958 6.0518097 8.6380468 8.3346972 7.6564465 7.9061137 7.6406669 7.9986877 8.2908016 8.1782208 9.4998947 7.7105706 7.8452473 7.6647155 8.3882946 8.1225319 8.1645966 9.553497 8.9079402 8.4077187 8.235236 0.248809581 1.02E-05
    Rv3144c 6.3632227 7.3090935 6.8442563 7.1840239 7.5121578 7.7141969 6.8609231 7.0334865 7.6140158 7.8331864 7.1562257 6.3494377 7.5515983 7.383046 7.3087103 6.7740345 6.280101 7.2742547 7.7503567 9.6930826 6.7143771 9.66256 7.5001688 9.5445153 10.101356 9.1508354 7.0519284 9.5781238 10.125742 6.4087592 7.0337635 9.041696 9.8601341 7.1305415 7.1821691 7.1900395 9.6388434 0.243264737 0.00030121
    Rv1270c 7.0697623 7.8814206 7.6422186 7.9700832 8.1474708 9.0516616 5.7445755 8.8505438 8.4062138 7.8887198 8.1622807 6.2623874 7.9980592 7.6305363 7.8740503 8.1309311 8.5769588 8.1764463 8.7414841 9.6863173 8.1896506 12.10235 10.251861 9.5634663 9.7269728 8.7882175 7.8506617 9.6357306 10.401416 8.1426207 7.8911749 10.047963 10.329515 8.1289363 8.3359699 8.5747547 10.320549 0.242939773 4.69E-05
    Rv1967 6.2571671 6.4378532 7.053584 6.6634502 6.8193829 7.8092476 6.9003438 6.4746268 6.5728419 6.778461 6.3503337 6.5946644 7.9053585 8.2884229 6.8262677 7.8586084 6.373915 7.359882 7.5127151 10.784951 6.4849105 9.6270699 7.745514 10.579391 10.004553 8.0383499 6.5751828 9.6814159 8.4866573 6.6003761 6.5324675 8.2826244 11.353532 6.6644487 6.9851304 6.7587376 7.8315858 0.24276565 0.0020226
    Rv0632c 7.1066734 6.7686946 7.7963558 6.4125428 5.4546043 5.8456335 7.6208857 8.2657181 6.7596457 6.8204727 6.6992958 7.4116117 5.9825135 5.9034512 8.0022893 7.6514666 7.8924446 7.9264868 8.5610289 7.6312812 8.6925299 7.3653139 7.1895547 7.4604483 8.8853992 9.7311726 8.9811038 9.3392879 8.3571267 7.4194177 8.0217954 7.3549242 8.2932516 8.8558222 8.2512082 8.3389519 8.8994341 0.241438894 1.35E-05
    Rv2288 6.3063629 5.6927008 6.1785901 5.7468104 6.5914302 6.8967151 6.4209232 7.3458053 6.0417776 5.9006815 5.6210108 6.0312756 7.9060875 7.6955791 6.4659841 5.7817036 6.3234094 5.9841482 7.3092618 7.354068 7.7803556 7.0778728 7.8725231 7.5403747 7.7324864 7.2957317 7.822537 7.2330322 7.5330212 7.4266551 7.5649077 7.5346677 7.32894 8.1110979 7.3897636 7.9862537 7.489098 0.241100005 4.00E-07
    Rv1408 6.8750429 5.7245474 6.0470636 5.8365417 5.3588573 5.6900694 7.660512 7.028151 5.7429685 6.0098609 5.7916968 7.1811317 7.1622516 6.8021386 7.8961083 6.2357122 7.2470574 6.1138438 7.8051818 7.6838363 8.6183361 7.3837387 6.6812631 7.1197009 7.1054823 6.8123472 8.6099114 7.037562 7.5789452 6.7763778 8.2698809 7.3268532 7.6350972 8.5590638 7.5798462 8.7747009 7.7376592 0.23986797 1.01E-05
    Rv0258c 6.9044212 7.827023 7.7213055 7.3690015 7.2237715 7.4009334 6.6636694 7.6728244 7.3095352 8.0434103 8.2564432 6.725985 7.3965594 7.3055574 7.2257561 7.8119739 8.0758024 7.4896869 8.5599548 8.4634707 9.6204915 8.3558118 7.6845613 8.3730098 9.2147701 7.9385529 10.349821 8.3690403 8.1780105 9.8411773 9.907153 7.8714662 8.0520758 9.1802554 9.5302119 9.801818 8.1546434 0.238908859 4.96E-07
    Rv2114 6.8912133 6.470406 6.5448766 6.3578113 6.2182355 6.3953288 5.7894724 6.6440762 6.7079913 7.3612603 7.0379018 6.1912108 6.5113194 7.3687467 6.0715258 6.0298457 6.7066605 6.3854338 6.0099505 7.9583024 5.9912935 9.9258553 6.1642456 9.8071096 8.7490157 12.414698 5.7832629 8.8462824 7.4270104 6.4610904 5.8850359 9.6854959 9.5225551 6.210356 6.2054874 6.1017604 7.4132365 0.23859921 0.00814781
    Rv2342 6.2762676 5.9536134 6.3066479 5.8864575 8.0382644 7.4195171 6.0616795 6.2336894 5.8946729 5.931524 6.0060943 5.8295573 7.3579661 6.7585818 6.3001438 6.1423144 6.2059449 5.7727378 7.3345927 8.0782925 6.4932644 7.1454735 8.5914496 8.0740318 7.8720885 7.6838579 6.7986442 7.4263937 7.4730218 6.5084051 6.6244128 8.2361661 8.5011514 7.1977095 6.8647453 7.1115069 8.381152 0.238129506 3.58E-06
    Rv3322c 6.9285338 5.3998335 5.953778 5.4883265 7.3025144 7.7185932 7.1523609 6.2568614 5.4165457 6.1022746 5.5736896 6.9336512 7.2233319 7.3004057 7.4956529 5.8532156 7.091898 6.0641846 7.4285085 7.4008146 7.6043762 7.8021601 6.6801375 7.3670689 8.0958309 6.9701142 8.6439765 8.4397289 7.8489021 7.5630312 7.7304756 7.1777765 6.9630168 8.509695 8.4488358 7.9681932 7.3242899 0.237981604 8.04E-06
    Rv2728c 7.2035017 6.7470556 6.9559499 6.8189848 8.4362631 9.0578691 8.4255377 9.6756886 7.2828907 8.7200845 8.2486142 7.9407075 8.6696246 9.3289095 9.4601348 6.6986545 9.796205 7.0502718 9.5713348 10.878154 9.1740312 10.242055 7.4647172 9.2050912 9.2788164 9.1609332 9.9424713 9.9986264 9.8815712 9.9279234 9.6494495 9.747845 9.3106413 9.9011273 9.506251 9.556456 9.5757175 0.23465603 2.15E-05
    Rv2023c 7.773004 5.7548473 5.9209752 5.7792318 6.3669487 6.6424857 6.8414671 7.3315974 5.8065235 6.1684558 5.9231349 7.0526744 6.0539818 6.1879354 6.6230361 6.5342748 7.1008832 6.713816 7.3985806 7.5334065 7.3555415 8.358874 7.5070843 6.7443458 8.0841752 6.821488 8.1442276 7.8809362 7.848377 7.2730267 7.8683239 6.7880731 7.6191637 7.908537 8.5243064 7.5779403 7.3918783 0.233085416 1.73E-07
    Rv2202c 7.7467191 5.6749106 5.8282051 6.0083829 8.2237317 8.3805102 6.441398 7.3966221 5.8151337 5.8647554 5.8653171 7.5851024 7.1320012 7.500278 7.1497927 5.9951267 7.349761 6.0920345 7.5587022 7.2718954 7.5592095 9.7779677 7.8810825 7.7186204 7.1920041 8.2510025 9.5383449 7.0069742 7.6287723 7.2067381 8.5177139 7.5120003 6.3748097 8.5741149 7.5161515 8.5916105 9.4784789 0.230567859 0.00021243
    Rv3259 6.4874586 6.6881167 6.4317907 6.7850137 7.2448451 6.0926026 6.8483356 6.073359 6.4661041 6.2778493 6.626848 6.1601438 6.1195093 6.2512173 5.7894378 6.6167299 6.1231056 6.0693744 6.8451348 7.8741744 6.32266 7.8879098 8.0882092 8.5170598 8.257601 8.0414183 6.5931666 7.948405 7.9428955 6.223808 6.6151744 7.7821793 8.0428723 7.0422585 7.1377602 7.0719366 8.351643 0.230295036 1.49E-06
    Rv3277 6.565491 5.9109382 6.1156688 5.8021268 6.1864726 5.8242219 6.8603708 6.8035645 5.9788219 5.8507106 5.7462976 6.8528253 5.8699689 5.8879911 6.6445046 6.6633318 6.6641508 6.2473414 6.6675918 7.5900234 6.629311 7.5878572 6.7874733 7.6709738 7.9715126 7.168811 7.2223368 7.8109867 8.1508356 6.6795637 6.7387685 7.7043372 7.3420646 7.0974536 7.6736815 6.8550007 7.8821701 0.229873611 1.19E-08
    Rv1109c 7.4124215 6.2377275 5.893762 6.0820689 7.1213229 7.4456527 7.95564 7.0658626 6.0117818 6.0588593 6.1135249 7.2445214 7.9359539 7.8734885 7.5752604 6.7122465 7.3821315 6.956575 7.3160226 8.0279817 7.8539234 9.9859649 7.0553523 9.0433923 7.7477618 9.8026672 7.4336025 8.7514009 6.8946018 7.4226899 7.5993594 10.731152 8.6761799 7.4798217 7.3503301 7.8589891 7.7571003 0.229456556 0.00017061
    Rv3906c 7.9678877 6.3528925 5.4998588 6.620618 7.08646 6.6615051 7.6781266 8.3194884 6.1866632 6.2180439 6.1972031 8.0135552 7.7060655 6.6402676 8.3399459 6.3227046 7.8896093 5.9902953 7.9896649 9.4549406 8.5477773 8.1144711 6.633428 7.69688 7.7606274 7.8779376 8.0611264 8.0422606 10.479252 8.6453502 8.2186095 8.8741386 7.3358626 7.6951632 7.6351696 8.4147977 8.0542116 0.22932232 7.68E-05
    Rv0440 7.5673823 8.6743843 7.6580359 8.8865878 9.0089736 7.3754723 7.9926208 7.7978768 10.328124 8.6241366 8.4741541 7.2604935 7.271521 7.6602653 8.3027289 9.2714979 10.501241 13.342173 6.8800517 7.3812957 10.567433 11.382286 7.2975502 11.488053 10.556999 13.18007 11.302883 12.978491 7.7911063 7.6124335 11.746973 9.5115063 6.6999525 9.5958504 11.82753 14.476539 10.649406 0.22852367 0.01361936
    Rv2559c 7.239709 7.5869735 7.5073464 8.194183 5.7289291 6.1203456 7.0439919 8.5627114 7.7776555 7.9921982 7.7849546 7.2749867 6.1125243 6.2998395 7.8461504 7.5831344 7.8466294 7.7009701 8.1980549 9.3793072 7.5986751 8.2507077 7.7441791 9.0666482 9.2551489 8.7896892 7.9539767 11.731149 8.4186419 7.5274125 8.4040548 8.8297759 9.3585426 7.8559688 8.6210241 7.6907673 8.8252969 0.228522025 5.72E-05
    Rv2484c 7.6660144 5.7439734 6.1067656 6.0295787 8.5718641 8.3316712 7.8107213 8.1613189 5.7696025 8.0170173 7.2764813 7.6868504 6.3408081 5.992306 7.2735954 6.5848616 7.827557 6.7519321 7.7700594 7.9973045 8.0932772 7.9564456 8.034999 8.0093225 8.7036819 8.3084166 9.1821368 8.3481139 7.9942898 8.2181693 8.0245848 8.1246567 8.2284238 8.9462169 8.758951 8.9182766 8.4299625 0.22685347 2.44E-05
    Rv1124 6.9558531 6.3102044 6.3081809 6.2886097 7.8395272 8.8149415 7.2422745 7.6233969 6.2644089 7.4874717 7.4510128 7.1521062 8.229352 8.0768932 7.2245687 6.5141789 8.333554 6.2653352 7.9639532 7.7590273 8.2902486 9.3639893 8.0975471 8.3803353 7.5180449 9.0458489 9.7608291 8.2515586 7.1261386 7.9303016 8.9743994 9.5475135 8.0274053 9.4446038 8.7289846 9.1454186 7.5323481 0.225315387 2.00E-05
    Rv1959c 6.821238 6.2903204 6.3253885 6.1909011 8.3839251 7.6095259 6.6158235 6.4993124 6.0399731 6.3072321 6.5126355 6.570944 7.039596 6.5989266 6.1901912 6.2079022 6.6416095 5.7133296 6.9811194 8.4277643 6.4539594 7.9085257 8.585406 8.6567315 8.2672413 8.1028736 6.7419361 8.1196989 7.9038065 6.997583 6.6468395 8.5121088 8.0107757 7.162816 6.9419535 6.8608476 8.9941468 0.225089443 1.77E-05
    Rv0793 6.9806847 6.3023261 6.3907187 6.6425674 8.0259658 7.542901 6.3984887 6.7456206 6.136946 6.240348 6.2204565 6.5567923 7.3376776 6.7418468 6.8982798 6.4914388 6.6416095 5.8248163 7.2642999 9.0750238 6.4427159 7.6899892 8.5881255 8.9131052 8.3780127 8.3867375 6.6442048 8.2017725 8.1326452 6.7175966 6.5950795 8.9489619 8.3767992 6.9253297 6.9517999 6.7451874 9.2125063 0.22497435 6.69E-05
    Rv3709c 12.577226 8.6251929 7.4640201 8.8134993 8.3319017 8.8691409 12.78021 9.0382472 7.5868681 8.6195127 9.2137321 11.864104 7.3149742 7.8366624 8.3268878 7.6562996 11.849452 6.6113503 8.9382679 11.302883 9.8937158 10.701305 7.4731978 11.732927 13.758344 10.137217 10.674293 10.446156 11.151468 10.012029 10.088233 10.822444 13.758344 9.5209739 10.588138 9.6781786 10.872467 0.224914393 0.00483437
    Rv3501c 7.3025193 6.0785573 6.4860374 6.207792 7.1322838 6.0893854 6.9865755 7.4005331 6.1130354 5.7630725 5.8301007 6.8132232 6.4380643 6.0923255 7.9954463 6.7887678 7.5737467 6.6088019 7.6010697 8.118781 6.6315905 7.3542018 8.3195489 8.3380222 8.3147379 8.1438587 7.2672795 7.8340003 8.0154609 6.2321639 7.5267766 8.6340781 8.7188696 7.6579661 6.96174 7.316532 8.6229036 0.224357475 6.06E-06
    Rv1197 6.8066986 6.6154225 6.4460732 6.4915294 6.3967442 6.4924839 6.506232 6.8197332 6.947426 7.0175901 7.4521271 6.3558369 6.5429676 7.9426726 6.2810488 5.8103317 6.9146684 6.3864488 6.1386005 9.4616536 7.0165866 7.3318254 6.4468087 8.6327173 8.3260297 8.621831 6.8214879 7.9417377 8.1815325 7.4119574 6.9161648 9.4084364 8.9419769 7.2675337 6.939388 7.1140454 9.0926696 0.221971869 0.00011999
    Rv3107c 7.7055981 6.8916997 7.1020261 7.1761312 6.6868556 6.8777786 8.1172497 5.6049054 7.0315305 7.1353599 6.9876775 7.7077833 6.7498553 6.891253 7.1144163 7.0453747 7.6138518 7.1833195 6.0942248 7.5918756 7.7730074 8.0310907 7.5138255 10.049626 9.6015717 10.929899 7.2904428 9.4798733 7.2631445 7.5159836 7.2442736 8.3653101 9.4618509 7.0275344 6.8202598 7.3636709 11.504475 0.220159981 0.00187163
    Rv0217c 7.3246171 9.6773835 6.9065112 8.7721368 6.8240777 6.1393095 7.8460607 8.216947 8.5017126 8.3268902 8.0654962 7.6244168 6.5575802 6.2856195 7.4608033 7.1280339 8.8710844 6.8392017 8.2431011 7.6882163 8.3385789 9.5991638 10.198216 10.167905 9.0309925 9.4818029 8.8185982 9.4532838 7.2177421 8.9771232 8.3506985 10.855894 9.2441805 8.8660898 8.5352157 8.3254909 7.3657012 0.218908609 0.0001941
    Rv3533c 6.7320874 6.0531088 6.2687261 5.9192689 7.335569 8.179593 6.8231819 6.4400704 6.1786013 7.7850116 7.4984349 6.860454 7.4754978 7.2881156 6.4721087 6.1655988 6.5478741 5.7031999 7.5122673 8.6095349 6.811463 7.9262722 8.4873796 8.5605623 8.5664461 8.2643043 7.3414944 8.2390331 8.245884 6.5848846 6.8198414 8.6465184 8.1639389 7.086435 7.364474 7.01978 9.1321408 0.217366821 2.81E-05
    Rv0270 5.5090119 6.6551386 7.0820409 7.3806853 7.6812624 7.4408594 6.2242423 7.4719108 6.7209856 7.9606031 8.2955927 6.2292272 7.5914757 7.1526388 6.9660392 7.1231381 6.4070268 7.0191146 7.6485062 7.7051153 8.1291913 8.4769483 7.4620291 7.9298979 7.8425985 7.506507 8.4660323 8.0917313 7.6649964 7.1924947 9.3103449 7.4743415 7.8371924 9.5393228 8.8521384 11.025151 7.512667 0.21664549 8.21E-05
    Rv1951c 6.8523757 6.2018889 6.2202067 6.0731394 8.3017077 7.2089287 6.9140553 6.6393395 6.2524245 6.3621353 6.9812191 6.9321475 6.7650371 6.7856576 7.0082766 5.9026138 7.1503999 6.116901 7.0952164 9.1117052 6.5532164 7.7372325 8.3801253 8.8635779 8.3121065 8.3378597 6.5186098 8.1805343 8.3282102 6.8786463 6.5767725 9.1040069 8.3056865 6.6528353 7.0381474 6.6665622 9.3193861 0.216160387 0.00014913
    Rv2723 6.9790277 6.2758688 6.4494359 6.5158535 8.0270041 8.0198752 6.7236277 7.2403333 6.0637449 6.5493605 7.1220471 6.8370637 8.0173751 7.997772 7.8591547 6.8998295 6.4833353 6.9196762 7.4196234 8.6307328 8.1901431 7.9014526 8.2769857 8.4484322 8.4542382 8.2353896 8.0550545 8.3240352 8.0900482 8.3320456 8.199009 8.2317593 8.100161 7.9499581 7.9599931 8.0910766 8.7261697 0.215385085 3.75E-07
    Rv2678c 6.6821195 7.5851832 6.5648805 6.7354322 7.800481 7.9140522 7.1457141 6.0687364 6.8348676 6.3234473 6.2091615 7.059859 7.9510768 7.6098814 6.981629 6.1303579 6.856083 5.8955583 6.3265319 9.3071558 6.9383348 7.9354149 9.1481698 8.8284296 8.726136 8.3059596 7.3676298 7.9847492 8.079643 7.082084 6.997367 9.0944489 8.3623549 7.2809545 7.8106132 7.1693144 9.4776067 0.213793533 9.29E-05
    Rv1144 7.7739312 6.2010055 6.8260497 6.4094525 6.3798057 6.8434963 7.729919 7.6406254 6.6304983 7.108401 7.2919002 8.4603435 7.0693195 7.3523245 7.4854461 6.5249425 8.1825928 6.9500851 7.8588403 9.4954952 7.2270991 8.5076259 8.7033967 9.0647919 8.7005546 8.4780218 7.6584627 8.1304885 8.5358202 7.4790978 8.2844838 9.18828 8.6593715 7.3834964 7.8490851 7.0374635 9.490021 0.213665858 7.26E-06
    Rv0557 7.4232165 6.6396665 6.6452994 6.4678558 7.5401141 7.6529702 7.6464489 7.8138992 6.6439464 6.7711182 8.4479356 7.3767584 6.88782 7.4144135 7.5610945 6.9451794 7.7143988 6.4927422 8.0625802 9.8975847 7.0929256 7.8594351 8.1003492 8.9612526 9.1781684 8.8908818 6.9409302 8.5141728 9.388498 7.6696785 7.7742823 9.7355075 8.884881 7.385297 8.0711101 6.9012127 9.7986298 0.212544963 6.11E-05
    Rv2369c 7.0077245 6.0908213 6.0464733 6.4518682 7.3310872 7.7084059 6.9869167 7.8945637 6.1490596 6.0142353 5.9852792 7.2389614 8.0794674 7.7709783 7.9887818 6.5015305 7.0513316 6.669743 7.7253853 8.3175308 8.0470797 7.8656795 8.3147509 8.4352932 8.5094335 7.4614731 7.5414379 8.1571149 7.8345152 9.0381746 7.6986439 8.4551877 8.1160601 7.3775716 7.5600105 7.4924062 8.8643584 0.212206523 5.06E-06
    Rv0589 6.6073621 6.0801922 6.3621178 5.8515355 7.0652498 7.8681009 7.4353335 6.4453921 6.1002174 6.0871698 6.0012332 6.686885 7.6984529 7.4579719 6.0465475 6.5984017 6.1332291 6.2920639 7.7258695 8.3927338 6.4836078 7.9609366 6.7135417 8.1916373 8.629699 8.3181211 7.0110537 8.2146258 8.1962264 6.071746 6.6648849 8.249152 8.2697624 7.1934516 7.1948706 7.1009547 8.7038036 0.212153107 5.11E-05
    Rv2849c 6.7229311 7.8814206 8.0516639 8.022471 7.6959633 8.1182713 6.6904118 6.6591018 6.9025007 6.7999532 7.7083846 6.2242399 8.0656025 8.6092886 8.9704751 8.2643425 7.0745779 8.4991078 8.2222998 8.2749898 8.5373507 9.8635073 7.9155019 9.2649109 8.9640556 8.5217984 9.9305641 8.8789559 8.2169454 8.0382119 9.6219698 8.5028891 8.9856195 10.068752 9.0884428 8.3402512 8.1678615 0.211576988 1.13E-05
    Rv0400c 6.5526715 6.0061401 6.4003581 5.9555066 7.6793416 8.0976254 6.958109 6.9597292 6.2012241 5.9457194 6.0368393 6.9438399 8.5024358 8.4972036 7.0477205 6.2088541 7.044861 5.9966169 7.9354595 9.0546048 6.7156071 7.7944404 6.474108 9.0686928 8.6854999 8.4209965 7.2684743 8.2577628 8.3101426 6.4493076 7.0669846 8.9850539 8.5677802 7.2630102 7.4614544 7.1458651 9.3693139 0.210723878 0.00041924
    Rv2808 6.1377146 6.8867832 7.3126971 6.2241144 6.5497823 7.0899921 5.8823172 6.4870827 6.559138 6.5873188 6.5652987 6.0783838 7.4405197 7.5524653 6.6088656 6.8723812 6.0381488 6.7081118 7.0905427 8.5505219 6.7784603 8.0141337 7.0419995 8.6669825 8.144468 8.3971141 7.5553327 7.8053636 7.9540568 6.1995012 7.1240701 8.6393625 8.0423131 7.2886474 6.0996431 7.6663864 8.9879913 0.210439561 2.57E-05
    Rv2291 8.2514994 8.5106938 7.8285736 8.5598151 5.8665251 6.3580971 6.5045559 9.3134377 8.0435098 7.3152234 8.3136075 6.8536015 6.2507609 7.3516935 9.1400934 8.0593663 8.7649666 8.8792955 9.7084236 8.3838325 7.7383799 9.6989748 7.9163761 9.6187625 8.6279216 10.142103 8.7473845 9.1004311 7.3394505 7.6917855 8.9005889 10.170755 9.3842665 10.010876 9.0141668 10.427616 8.5439008 0.21026421 0.00034801
    Rv2021c 6.6241424 6.2761505 6.4055435 6.2806502 8.1864037 7.7126629 6.2659644 7.1492878 6.2364174 6.1519553 6.3037457 6.6342802 6.9687203 6.594982 6.6186473 6.3138826 6.5889112 5.6377654 7.1972204 8.2499793 6.6103126 8.4831603 8.7372581 8.3792075 7.9050787 8.159748 6.9626559 7.7858705 7.9875904 6.6818042 6.9536453 8.1386675 7.5708294 6.9180924 6.2671534 7.3316256 8.9331482 0.210209284 3.08E-05
    Rv3495c 7.8435094 5.8750463 6.1842431 6.0255898 7.9948073 7.870374 7.7377245 7.8189482 6.0566491 5.9103246 5.773126 8.1013372 8.5585983 8.6856552 7.6509705 6.0269774 7.9027522 6.1497345 7.8330625 8.4484672 7.8917437 8.0534911 8.6766436 8.8590425 8.611569 8.2875355 8.20736 8.3373561 8.3448545 7.4786826 7.8379638 8.3044036 8.1535137 8.1794727 7.5768043 8.4942966 8.8762212 0.209704323 0.00020609
    Rv3184 7.310353 6.1037186 5.9505954 6.2832533 7.0081257 7.3964607 6.7703983 7.4876925 6.0389934 7.2895024 7.3099342 7.4788339 6.8523303 6.8362025 7.6407727 6.4723548 7.2652274 6.6736797 7.926423 8.6757686 8.5110091 8.4810685 7.8441503 7.8352504 7.3211894 7.6877605 7.9917674 7.4664938 8.9927142 7.789708 8.1827607 7.9766929 7.3103974 7.3171977 7.4919156 8.6232862 8.1445203 0.209684024 1.84E-07
    Rv2520c 8.0858191 7.0776914 7.7536492 7.2832387 6.5157584 6.7723351 7.0659698 7.9532704 7.678732 7.7062745 7.4754035 7.8882967 7.2790008 8.4009064 7.9069639 7.4408386 8.3750335 7.6242148 7.7328139 10.100819 10.98035 9.4454931 6.9845271 8.5265724 7.8752709 7.9478375 8.5398775 8.8163161 9.70575 8.3079888 8.8688855 8.6898396 8.5980361 8.6986632 8.6695632 9.4883672 8.3809463 0.209678209 1.60E-05
    Rv0389 7.1827528 6.039459 6.2567975 6.2964642 8.0238976 7.3330554 6.7649891 7.6565178 6.0688405 6.1010412 6.0029379 6.4520016 7.3331729 7.2855242 8.0269634 6.5834381 7.97059 6.7628961 7.8145651 8.0087504 9.5313067 7.5506519 7.7848986 7.7613674 8.6575046 7.2803936 7.4990177 8.1724523 8.7440675 7.4011623 7.6596607 7.7484965 8.1316337 7.9315159 7.9378274 7.9566517 7.9473553 0.209515248 6.69E-06
    Rv1247c 6.5372622 6.1434676 6.3338963 6.175442 6.4051965 6.859346 6.1384929 6.9086335 6.2176705 5.7492681 5.8997663 6.3162488 7.0589977 7.2891689 7.1721836 6.5338719 6.2957651 6.2305737 7.4650275 7.7487784 7.1226477 7.5788627 6.8692732 8.0019658 7.6739637 7.5681857 7.8385254 7.1124302 7.3817818 6.5904344 7.4891916 7.6492842 8.0159378 7.5372963 6.2297934 7.9412168 8.087316 0.20947154 6.59E-08
    Rv1602 6.8933109 6.984798 6.8586612 6.5624497 5.5091416 5.7856398 7.2173905 6.5633073 7.4856603 8.1805464 7.8352841 7.0893116 5.6566439 5.7917653 6.416599 5.9827144 7.580814 6.1776003 8.0048621 7.5572277 7.2020984 8.35444 6.8354347 7.5545587 7.9060027 6.6937525 9.0727502 8.3071083 7.8012694 6.9934344 7.4758054 7.2980654 7.5958159 8.7395412 8.1110384 8.0386769 7.5892598 0.209209545 4.48E-05
    Rv3210c 6.0495126 7.0542621 7.2796517 7.2124076 7.052712 7.2337552 5.8524324 6.6164854 7.5532098 7.2947793 7.3940059 6.1462024 7.2245151 7.2051316 7.9066519 7.6562996 6.397931 7.3309911 7.9843926 7.3817169 7.2406169 7.9701904 6.6837678 7.9823152 8.3333702 11.81162 7.5187507 9.5993248 10.048692 6.2574588 7.1914347 8.0571455 9.1251887 8.1099447 7.2202265 8.3108196 7.482761 0.20913478 0.00122852
    Rv2413c 7.7927753 5.632358 6.1949909 6.9373653 7.8653023 8.0675073 6.3648613 7.7760367 5.8751502 8.224576 7.4907322 7.2322127 7.4519428 7.5434787 7.7732441 5.7322244 7.5418754 6.1384282 8.1746324 7.8967814 9.2736163 7.3661308 8.6125338 8.2628435 8.4542998 7.9165002 8.4276216 7.8025688 7.9118223 7.9176378 8.6367752 7.7236052 8.0009774 9.0887316 8.7500234 7.4466181 8.0703017 0.209056938 3.33E-05
    Rv2070c 7.6121924 6.2778469 6.860321 7.2404894 6.9678326 6.8989394 6.8786038 7.9190285 6.204255 6.9318469 6.60962 7.7923678 6.6866395 5.6959895 8.3211839 6.8380473 7.8876101 6.6075568 7.0398585 9.1861229 8.5400877 8.0440491 6.5767788 7.8743538 7.829331 7.9409012 8.0004047 7.9038442 10.239563 8.4508538 8.1013471 8.7201881 7.5249527 7.7245824 7.7502758 8.4428282 8.0293887 0.208116427 3.07E-05
    Rv0476 7.3012574 6.1238809 6.0462895 5.8066225 6.7771832 6.9264524 8.0711596 5.6272462 5.9967883 5.9013788 5.8370383 7.350135 7.6878999 8.0060631 7.6448384 6.6547572 7.2184155 6.822633 6.5958381 9.7585084 6.6084572 8.2445756 6.7948787 8.586389 8.777771 8.2336227 6.6744595 8.7229933 8.4551213 6.8341539 6.7283855 9.131293 8.0255233 6.8737797 7.2366646 6.6008234 9.6209059 0.207986081 0.00114508
    Rv2503c 9.0080002 6.8192997 8.1324031 6.5670397 7.7224157 8.0169043 7.7399466 7.3373318 6.5471683 8.158015 7.5112365 6.9989299 8.1545266 8.4359353 7.9376135 8.6330564 9.3014272 9.4890971 8.4199022 9.9953819 8.8496294 9.5985132 6.9156757 9.2910756 9.8058259 8.8882672 8.4432755 8.7400203 8.8746987 9.7485006 9.4118774 10.160112 10.114597 9.5294212 10.07773 7.9670787 8.8832977 0.207652154 5.49E-05
    Rv1702c 6.8219227 8.3383996 6.6682918 7.9232596 7.9756591 7.1866295 6.6907732 7.5606315 6.8962324 7.2767953 7.4687206 7.0396146 6.9959514 6.3352607 7.1391375 7.1628517 6.8808878 6.8476541 7.5242569 7.9311105 9.4245386 7.2323855 7.1275461 7.9425242 7.5655956 8.1026752 10.590645 7.3224096 7.3495133 8.5082636 9.6457595 7.4707449 7.5038915 9.300779 9.0535719 9.4995843 8.3419047 0.207075619 0.0001213
    Rv2502c 6.6394946 7.9741535 8.5684659 6.4271362 8.603303 7.8957422 7.1572246 8.2239646 6.9265714 6.5752103 6.4756633 7.0194347 6.641754 6.7222088 7.6566851 8.1146112 6.9693454 7.3118289 8.0949572 9.7717453 7.5859646 8.6180678 9.1937469 9.1358269 9.0126314 8.2558217 7.68982 8.6871074 8.9434613 7.2928264 8.1323001 8.9447129 8.1202634 7.7482604 8.1648497 7.4097437 9.8460676 0.206426911 2.78E-05
    Rv2530c 6.803303 6.5181546 6.691543 6.5085851 5.8369768 6.1583006 6.8552683 6.8614745 6.5132482 6.8508916 6.2358028 6.8565487 6.2334394 6.9794465 7.4686964 6.6706876 6.7186447 5.8000858 7.8454911 7.9512188 6.5833916 7.8807165 8.1973684 8.0507616 8.1650066 8.1656284 7.1463746 7.8165181 7.9095485 6.4243363 7.1815881 7.7828179 7.9615162 7.057693 6.9665777 7.0500941 8.2529906 0.206333994 3.19E-07
    Rv1070c 5.9227339 7.0613109 6.7495955 7.2502628 7.3727996 7.4690799 5.9396228 8.0995398 7.4721445 7.5144518 7.789461 5.8955462 7.8976933 10.02142 8.2877294 6.7618599 8.6659961 6.8064524 8.0623736 8.5242849 8.7786769 8.9806087 6.8325549 9.6027611 8.338448 8.03819 7.907883 7.9782873 7.9721722 7.614648 8.9416991 7.7277305 8.8814256 8.8564061 8.974967 12.01772 7.9115157 0.206293509 0.00112971
    Rv2140c 10.14901 7.5468231 8.2529551 7.662908 7.3797155 7.6895017 7.6248906 7.5232831 8.4805358 7.6349783 7.405246 8.6336 7.6729795 8.5913969 6.8376002 7.8069222 9.8958854 8.0136371 7.0212743 9.4660814 9.6075652 9.9275889 6.5250235 10.217164 9.6190411 9.8725436 9.761231 10.134919 7.8558903 10.180142 9.3202381 10.34067 10.357409 9.4207719 8.6223555 10.244544 7.8217928 0.206082669 0.00038544
    Rv0145 9.3490334 8.5899679 8.551127 8.7234645 12.78021 8.6277625 8.2345496 9.1269735 9.1202829 8.8170382 8.0825387 11.393826 7.9942483 9.1960078 7.9544692 8.2855034 10.018765 9.252856 9.7257166 11.004181 10.686571 10.864367 8.5744993 12.223228 12.060624 9.727716 10.146924 10.175428 10.509893 9.9973328 9.6329155 11.780836 12.099688 9.6168164 10.05322 10.407366 10.335503 0.204711052 0.00030779
    Rv3750c 7.4899051 7.2637923 7.0751387 7.0997668 6.1330236 6.7296596 6.6788147 7.9491717 8.1900017 7.6415773 7.9272228 7.7185115 6.2404093 7.0639707 7.2465689 7.4362556 7.7545926 7.3891702 7.1678948 8.2860935 9.2209557 7.8029906 6.9010287 8.1382122 7.8498969 7.907256 9.4857915 7.5530067 7.8072533 9.0089117 9.0900578 7.8776024 8.0365893 9.183623 9.6304607 10.062473 8.2861086 0.203839172 3.64E-05
    Rv0690c 6.0086155 8.3955252 7.8936759 6.1463139 8.4087737 7.9229728 6.4438813 7.8517916 6.9601318 7.5380702 7.8846593 6.3742178 7.5387431 7.1298418 6.5692189 8.0085228 7.2504229 7.4353871 7.6070965 7.3145407 7.3892228 9.9449279 6.5102292 9.5257993 7.9956503 9.1272398 8.5021043 8.8574891 7.1597933 6.9754059 8.2506255 9.0347196 8.4401791 9.2601061 8.5897868 7.6510707 12.024334 0.203593418 0.00150578
    Rv2284 5.9355483 5.9384983 6.6551616 5.8828116 6.2793237 6.4580334 6.8342352 7.8290946 5.8231709 6.6594922 6.5640394 6.2373589 6.1672343 7.0806362 7.3272384 7.0787083 5.8703496 8.042163 7.7253853 7.5637669 7.2847841 8.2707496 7.6422848 7.79934 7.9834114 7.9396649 7.9249942 8.0452691 7.8346184 5.8317319 7.2063569 7.3803607 6.731166 8.1462577 7.1527939 8.0835608 7.692565 0.203588049 1.40E-05
    Rv0390 9.2166313 7.5720656 7.2501698 7.5241636 8.7006926 9.0869527 6.4891407 8.893402 7.5746134 7.523643 7.8322599 7.829491 7.0328334 5.9610797 7.6060711 7.7162896 9.3000923 7.9548183 8.6090993 8.5377371 9.3664931 8.6868565 7.9649358 10.471046 8.1411529 8.8862255 9.3542855 8.2552914 9.1289975 9.0671318 9.4915758 8.4646233 9.5956343 8.6725548 10.737078 9.6391911 8.156911 0.201552144 8.47E-05
    Rv3590c 7.5029275 6.8327779 6.0076329 6.4592501 7.9520505 6.4973049 7.6618806 7.1839449 6.7692938 6.4118277 6.4378192 7.376255 6.6819402 6.5214247 7.5773869 5.7891415 7.2983112 5.6848985 7.5168204 8.5863005 7.4419595 8.1667319 8.3191717 8.387454 7.8657405 7.7313973 7.0559036 8.1393513 8.5623423 7.3423609 7.3429924 8.3838236 8.0553405 6.8471608 7.128146 7.0083508 8.9424413 0.201101665 1.34E-05
    Rv0914c 11.423935 9.7639059 9.1056313 8.8767191 8.6652465 7.9593116 12.223228 10.332727 9.2553006 11.919654 10.756042 11.422873 7.2295942 8.4806405 9.0723955 8.8614531 10.560292 10.452591 11.114803 11.980408 10.861416 10.835388 9.0688921 11.469255 12.978491 9.8893153 11.781748 10.612988 10.094566 12.060624 11.106452 11.357078 12.236683 11.648277 11.951014 11.669257 11.145582 0.200143278 0.00044359
    Rv2002 7.6093934 5.968065 6.4463687 6.0691473 7.0073975 8.0702808 7.1783868 6.8509008 6.2580066 7.4601467 7.7303542 7.450223 8.1028796 7.7144838 6.8388302 6.6777969 7.6516381 6.4896885 8.0701848 8.2210749 7.748586 8.473949 7.5104612 8.1117657 8.6790832 8.3236452 8.6282542 8.1661492 8.0626191 7.6849342 7.6613867 7.9043646 8.283638 8.4694257 8.0778769 8.2370669 8.3395636 0.199707709 1.65E-06
    RV1237 7.9886353 6.3369427 5.9189492 6.1720482 7.7477773 5.649325 7.2539905 7.6791349 6.2244398 6.0877635 6.0452313 7.9096433 6.3293409 5.4974229 7.1714753 6.5317928 7.7052069 6.5642113 7.298946 7.8939629 7.9521156 7.2975463 7.2104833 7.5643304 7.7230688 7.7924901 7.3861684 7.6382577 9.2656874 8.2094424 7.4453678 7.7775872 7.8189657 7.4928477 7.4547835 7.5758564 7.6194535 0.199304461 5.92E-05
    Rv1898 7.3702816 7.487157 7.730418 7.8827366 6.452257 6.190058 6.7541547 7.7103417 6.7678424 6.3093723 7.1207523 7.4895247 6.4351216 6.126241 7.3264955 7.4888118 7.4371771 7.1267332 7.1482044 8.3016559 8.3358058 8.0556482 8.547838 8.5715701 8.1168521 8.1210216 7.9414304 7.701452 7.9649143 8.9805038 7.8256343 8.0251047 8.0638738 7.8230168 8.160436 7.7667331 8.6519853 0.198738024 2.28E-07
    Rv3446c 6.8174165 7.1924478 6.2657614 7.4289715 7.3734869 7.3754723 6.9023312 7.1253758 7.1519337 6.9039287 6.5702748 6.4476922 7.0327962 6.6236377 6.4150466 6.3713123 6.1410544 6.267368 8.5167596 8.1872525 6.9764624 7.6091639 6.7037475 8.5329869 8.4032823 8.3383623 7.5227708 8.0915532 8.0337761 5.8564541 7.5056213 8.031542 8.4502428 7.7910515 7.4692351 7.5440585 8.7083193 0.19857201 7.03E-06
    Rv1700 7.9738665 6.415962 5.8866802 6.3882546 6.8075512 6.5549692 6.5811774 7.8661586 6.2040086 6.0946808 6.0991305 7.9276833 7.5276268 7.6802713 7.8271271 7.1312019 8.0059301 6.5713997 7.3780546 8.8345096 8.2458186 8.0575619 6.8023641 7.975979 8.0506612 8.1442294 7.5137768 8.1228391 9.4864208 8.3846931 8.0123167 8.9229937 7.6443808 7.3955437 7.629732 7.4559607 7.941213 0.197870382 4.32E-05
    Rv1411c 6.9657694 7.7324855 7.6756093 7.3021978 5.5149034 5.8593874 7.7941155 6.7028267 7.6251164 8.1797192 7.4820103 7.1566713 5.7114676 6.0183913 6.3364151 7.5668776 7.2180919 7.6947702 8.0981024 7.6397657 7.5374311 7.3334206 7.1652164 7.3770757 7.8999267 9.9603712 9.0810552 8.1895094 7.2722094 7.4268579 7.5870859 7.5955193 7.8840687 10.116895 8.4900113 8.1196 8.3398222 0.197045847 0.00030435
    Rv0308 7.0944744 7.0960519 6.2281986 7.0402268 7.1257445 7.547564 6.8005411 8.1036829 6.6484665 5.4634314 5.6571117 6.9511723 7.8740493 8.0086323 7.578158 6.630977 7.0362349 6.7353941 6.9797223 9.4520358 6.8490415 7.8259005 6.3806118 9.1114191 8.9359151 8.5005652 7.0587254 8.3321498 8.354149 7.3273147 7.6066867 9.6909271 9.0402422 6.9966187 6.9419535 6.766054 9.8057463 0.196581572 0.00110637
    Rv2184c 7.2130181 8.8944824 7.9657205 6.5514625 8.4707393 7.7465212 7.0010727 7.1127062 7.1821749 6.2989294 6.9091673 8.1611414 6.2024712 6.2693332 7.3686112 8.222302 7.4983017 7.4932933 8.5011358 9.5124254 7.2721223 8.0688375 9.3291114 8.9798941 9.3687333 8.4566389 7.6807916 8.4645517 8.2150168 7.4969356 7.673051 9.5808268 8.6710904 8.0152227 8.2584441 7.347124 9.4499125 0.196487387 7.85E-05
    Rv3686c 7.2444754 7.7383271 7.9139309 8.0428161 5.6875915 5.5447296 7.7884609 7.7019533 8.6661711 8.1418603 7.2225951 7.510436 5.9552304 5.7434059 7.8020139 7.7399865 7.8896093 7.8976166 8.0097499 8.9252195 8.2730496 9.110572 7.4661967 7.8699812 9.1445944 7.8830426 8.6655391 9.2048242 9.392441 8.5909492 8.1225102 7.7862665 8.1509746 8.747632 8.5138476 8.3091812 7.6723266 0.195553302 0.0001513
    Rv3035 6.5089905 7.5031261 6.942918 7.2391122 7.547953 7.9029398 6.9043015 7.3330682 7.466791 7.1221136 7.4611669 6.5250468 7.98155 7.5117512 6.9970593 6.997375 6.4096827 6.9696561 7.3975423 8.9713021 6.9411332 9.0182768 7.6773907 9.8600978 9.7524194 8.817673 7.1692939 8.9315469 8.5185575 6.7845094 6.8631566 8.1059471 9.3601712 7.2138333 7.3880696 7.3903534 10.110413 0.195057161 0.00045928
    Rv3911 6.6440424 7.0267467 7.0047582 6.600185 7.2907968 7.7012556 7.5644633 6.7377614 6.7334506 7.111795 6.8056381 6.956223 7.57886 7.1951712 6.7845861 6.7020497 6.4586422 6.5940413 7.4617794 8.253133 7.9705864 7.5360836 7.7313732 8.2765536 8.4566995 7.9792044 8.1450182 7.9600012 8.0034138 7.9279435 7.6489592 8.0987651 7.8584169 8.0137726 7.7513481 8.0591239 8.4412121 0.194436199 7.03E-11
    Rv1654 8.2969143 7.8866405 7.3784948 8.1808785 8.5481748 6.5700661 7.2845041 9.1213218 7.7261084 6.1390722 6.1449562 9.1621556 8.1508772 5.9169854 8.6893576 7.3890156 8.4152595 7.0943634 8.9588164 7.9148343 9.5243485 7.7890825 8.3550261 9.7180689 7.5603949 9.0117337 8.4870212 8.3107562 8.4641766 9.4824766 9.9095602 9.5973472 8.6555689 8.6194882 9.1474512 9.919209 7.3043796 0.193846344 0.00041621
    Rv1400c 10.146766 6.4305655 6.5901582 6.5957288 8.2716844 7.5749844 7.3145511 8.0323673 6.2404135 5.4311921 5.7679218 7.4107949 6.85726 6.6103753 7.8643651 6.9598925 11.829657 6.871523 8.4158953 7.2700474 8.9491673 8.0950876 8.7133993 8.2587613 11.764286 7.651914 7.6177896 8.6852109 6.577285 7.3922086 8.8261033 6.8784871 8.8631728 8.9290434 8.524532 7.3712375 11.397864 0.192447625 0.01642773
    Rv2359 7.9750726 7.2821299 6.5601153 7.7110603 7.2781814 7.5562506 5.8029322 7.9373361 9.0709028 7.8330149 7.7246025 6.8577024 7.8731396 8.0738561 7.0061656 6.7412791 8.6764922 6.8466835 6.7026314 8.615154 9.4011459 8.5232997 6.6146708 8.5225021 7.9762555 8.6816635 10.233456 7.8206927 7.6614751 8.4478265 9.3623236 8.1734984 8.7450307 9.4790314 9.6953704 9.6743588 8.215707 0.191951846 0.00038405
    Rv3012c 7.4226038 6.0917631 5.9630888 5.6875561 8.2140118 7.4652267 8.1445239 6.0469063 5.8806035 8.3863326 7.3806437 7.4994964 6.4757722 6.6330261 6.4903178 5.9164398 7.2570939 6.000779 6.8092356 8.5351079 6.6739644 7.8236521 8.1698588 8.4227511 8.5769563 8.3310555 6.6332129 8.6743689 8.123337 7.2340114 6.6335259 9.3258389 8.3883678 6.6282431 7.2583917 6.6671359 9.3028349 0.191513973 0.00143351
    Rv0758 6.5017296 6.6687954 6.807707 6.5078441 6.2878192 6.54865 6.4222228 7.6969925 6.9791661 7.3952741 6.9843102 6.5118352 6.5189077 7.2147676 7.1036254 6.3297089 6.2844779 6.7449883 7.6429793 7.7227897 7.741813 7.4158983 5.9350039 7.7325715 8.1269282 7.9565999 8.215855 7.8346568 7.6872584 6.8432506 7.7381327 7.7170684 7.9142945 8.5630904 7.4995286 8.2423733 7.8679187 0.190832383 5.22E-07
    Rv1942c 6.3296368 7.5366619 6.924461 7.5378606 6.5579485 6.0771115 6.0030564 7.4245309 7.2918038 6.3685667 6.8828042 6.2621399 6.3630163 6.2260401 6.3895378 6.7808498 6.3120382 6.9282907 7.6038998 7.757172 7.5845983 8.1276717 7.8524978 8.1003301 7.7129917 7.9951683 7.1220475 7.3248118 7.671494 8.0583477 7.1945314 7.7291453 7.2694915 7.174939 7.1798957 7.3443282 7.9935616 0.190635289 1.36E-07
    RV0687 7.8145991 6.5972826 6.5424419 6.5548456 6.2430185 6.5825015 7.6697943 6.5865492 6.8104527 7.2524563 7.5217494 7.3919817 6.3998957 7.4797054 6.2068362 6.173173 7.5109425 6.4277074 6.0351859 7.327698 8.3258775 6.9821293 7.9893592 7.5478573 9.1919025 8.4523362 8.5705647 8.0463706 6.9864734 7.9729764 8.5284092 8.028304 7.8846609 7.8827304 8.4804161 7.4105411 7.4523044 0.190625718 2.99E-05
    Rv3249c 6.9390555 5.8280145 6.1512342 5.9306674 7.2781896 7.6962823 7.5726127 6.869517 6.1936669 6.2343155 6.1197347 7.0084131 7.9687628 7.2981781 6.8859878 6.1101422 6.7090049 5.6974341 7.0554349 7.0837534 6.8726188 9.456611 7.1391708 7.821196 8.6033342 7.7424181 7.6483094 8.5202338 7.6533143 6.4594562 6.89897 8.0425396 8.9707356 7.3827811 7.2302958 7.4144785 7.1418318 0.190489705 0.00020419
    Rv3611 5.9335619 7.4012283 6.6798831 6.8680712 6.4179844 5.6580251 7.0720513 7.2672247 7.129884 7.8307867 7.5416803 6.367948 5.6356131 5.7445302 6.6276813 6.5515563 5.9109041 6.603076 7.238061 7.6419707 7.2084592 7.8035156 7.9332274 7.6794511 8.1627782 7.3172913 7.5736847 7.951029 7.9507107 6.3967676 7.1647959 7.1276149 7.6397201 7.7906448 7.4309968 7.9820354 7.5990922 0.190082591 1.17E-05
    Rv0970 7.1873069 6.7005646 6.620289 6.9939104 7.178671 8.0293812 7.8246298 6.6312802 6.4773022 7.7191711 6.4498698 7.1102482 8.1049665 7.9981566 6.6800035 7.5413039 6.9414706 7.7732267 7.2261918 7.2763562 7.2952219 10.136689 7.2019401 9.7631317 9.3707576 9.3657927 7.1497176 9.5944407 7.0797092 7.1744849 7.3530066 10.893172 10.040862 7.2308154 7.128836 7.6763689 7.542186 0.190070097 0.00277466
    Rv1524 6.2450402 8.2611324 8.0345541 7.3758207 6.6489026 6.9874765 6.4396632 7.6186707 7.3106482 8.1805541 7.7030031 6.6114282 7.0440372 7.6523515 7.6838306 7.9472129 6.9475588 7.5678214 8.2323649 8.6114072 9.1454289 8.9734703 6.5935055 8.3542224 9.010671 8.2310371 8.5491681 8.7222005 8.4356721 8.1800116 8.0961322 8.236428 8.1488787 8.7586069 8.7085006 7.7894839 8.4809388 0.189990895 2.59E-06
    Rv3001c 7.2820895 6.927794 6.359556 6.6296366 6.9626099 7.0937395 7.5359774 8.1753179 6.612606 7.0844745 8.1232853 7.2782848 6.8462939 7.0726625 7.8168248 6.8046326 7.3505105 6.5209292 7.7419348 9.6307629 6.8910386 7.6661088 8.0357731 8.8461385 9.0686616 8.6323279 6.7262053 8.5893199 9.4676151 7.369182 6.9832754 9.2854703 8.7291094 6.9761219 7.6554017 6.6867493 9.6718664 0.18952026 0.00043669
    Rv3029c 6.5987855 6.2518317 6.4858187 6.4941068 8.4467973 8.1177286 6.5829114 7.1376889 6.1970163 6.1476282 6.1407309 6.6438469 7.7323708 6.9510746 7.2319904 6.1091069 6.459959 6.0733283 7.9213927 7.7912645 7.5902119 7.767296 8.337876 7.7537005 7.7733678 7.6705335 7.8643919 7.5944721 7.7220717 6.9766666 7.9866615 7.6610896 7.9336955 7.4570762 7.5221431 7.4946075 7.7175421 0.188706861 1.36E-05
    Rv2216 6.7724007 6.4622067 6.2816542 6.3506234 8.3031877 8.5538918 6.0090444 7.690532 6.6109622 6.4274585 7.5205449 6.4751674 8.4205662 8.1307291 7.1565269 6.5451897 7.3419755 6.9246231 8.1003973 8.2716652 7.5827609 9.0869328 7.5217826 8.0572308 8.0287291 8.4328353 7.8595337 7.768282 8.0374811 7.6926128 7.9695685 8.5090489 8.0168539 8.7959717 8.4914126 7.9062648 7.8095395 0.188467607 4.87E-05
    Rv1575 6.235218 5.9161435 5.9554571 5.7923113 7.8714761 8.112205 6.9186303 6.9654249 5.9163579 5.932806 5.8541991 6.4322328 7.0493538 6.8425476 6.2375832 6.2630481 6.1119384 6.7063397 6.9440896 7.8562328 6.5680606 8.0730673 7.7330675 7.5920575 8.2343186 6.9888878 7.0523475 8.0805298 8.0677176 6.3878916 6.8109087 7.2303025 7.5247513 7.1721191 7.5383667 7.2551719 7.7429291 0.188281323 4.46E-05
    Rv0427c 7.8034508 7.4753507 6.9625555 7.1874269 7.5448877 6.7168153 6.6849388 8.2254914 7.796858 7.4698717 7.6459681 7.9710084 7.3736697 6.0009993 7.443675 7.2064919 8.0818588 7.5343688 8.1508547 9.2294559 8.7880146 9.0103721 6.4567378 7.7999275 7.9935884 8.6000892 8.0271281 8.1739007 10.192115 8.7632113 8.1533596 9.0988584 8.6991362 8.1176199 8.3141501 8.5245337 7.9751714 0.187895463 1.71E-05
    Rv2398c 7.9286589 6.19848 5.7495334 6.0537104 7.228984 6.8364065 7.1366392 8.1349099 6.1444155 6.1621662 6.1034635 7.9464258 7.6372561 6.9242464 8.1552997 6.2743092 7.9674539 6.1467704 7.6475863 8.2549473 8.4075597 7.6194372 7.0929769 7.6477964 7.3456974 7.4707974 7.705917 7.6550067 9.655172 8.5050948 7.8258532 8.1186997 7.3676494 7.8109693 7.5682842 8.2811248 7.9254889 0.187257753 0.00019205
    Rv0251c 6.932854 7.5664272 7.6147823 7.2527043 5.6840208 6.1516955 6.8956948 9.9524656 7.484848 7.7959513 7.8653417 6.3832432 6.2874967 6.2851834 6.1850003 7.5786267 7.2306368 7.2881894 9.1126023 8.2146071 7.2272828 8.171504 6.6351294 8.7176277 8.6311926 8.1697357 7.7144372 8.3811147 8.1018273 7.1751586 8.1500971 7.9802547 8.5766963 8.2933204 9.0852322 7.9375352 8.0600134 0.187025983 0.00047263
    Rv3647c 6.5276886 6.0468959 6.0566373 6.1366547 7.5678286 7.6711301 6.429446 6.5258681 6.2214782 5.7622411 6.0195446 6.6190016 7.3949918 7.7086701 7.3755013 6.3935803 6.775 6.2328926 6.9178697 7.8979876 6.8830066 7.5834913 8.3967313 7.9749245 7.5379749 8.0308167 7.2310568 7.242654 7.5866843 7.4709954 7.1289474 7.8394729 7.5552001 7.7943358 7.4740524 7.1789307 7.8291665 0.187005395 6.44E-06
    Rv1846c 9.7652492 5.7331082 5.92929 6.160345 7.4254517 7.83975 8.2885924 7.0530073 5.6547229 5.9508739 6.0075822 8.3020367 7.7477241 7.8429746 7.0942823 6.1894034 8.7129599 6.1607666 7.4770261 7.7000141 8.6711107 7.1281938 6.762879 7.9791326 8.0628242 7.9900971 9.4065673 8.7250383 7.5619778 7.9955333 9.7686612 7.6979436 7.6766324 8.0360252 8.7876016 8.3686023 7.8119916 0.186705662 0.00302066
    Rv0394c 6.9827983 8.0674431 8.1503646 7.8647011 7.6068548 7.9095046 8.1168082 8.3049337 7.8215017 7.9625353 7.7189051 7.0208062 5.9535751 5.7741947 8.364003 8.1435012 7.0764308 7.9029239 7.8985231 9.9424951 8.4478713 9.2048127 7.2945181 8.6356042 8.8302634 8.8220948 8.4360172 9.2595523 8.5586939 9.3426822 8.1138059 8.6990168 8.2836686 8.284822 9.0114843 7.9242678 9.2330889 0.186202001 3.01E-05
    Rv2788 6.853321 7.767368 7.7512895 8.1332104 8.370278 7.1293195 6.5143772 8.5440416 6.8251709 8.3391945 8.1411559 6.8235683 7.1171868 6.7975288 7.2454465 8.2388495 8.0950417 8.0183876 8.6451792 7.6887482 7.5909122 9.6263866 9.0523041 9.2148298 9.813036 9.1853777 8.097633 8.8496379 7.4546289 7.7412511 8.7226818 10.16894 8.8993464 8.7254382 8.9621823 8.3567313 7.3125957 0.185578433 7.85E-05
    Rv0474 7.4680449 7.0775898 6.0217093 7.0580418 7.7012003 6.7109951 7.2263579 7.1341829 6.76695 6.584382 6.465298 7.4381425 6.7382495 6.4484884 6.9852935 6.2468305 7.3935956 6.0827909 7.0278195 8.357403 7.8455287 7.7300291 7.9472021 8.3042792 7.8210926 7.9720296 7.4729231 7.6999309 7.9191624 7.5733751 7.8904176 8.1840773 8.1227051 7.0397812 7.4733407 7.5028612 8.4242163 0.185522282 1.65E-07
    Rv1842c 7.4945815 7.8199206 7.7413619 8.2389459 6.7814737 6.065093 7.4041245 8.380552 8.1652258 8.4947879 8.4336327 7.0514101 6.3630163 6.5277243 9.3508526 8.3213279 7.3372815 7.9396167 9.3079454 10.074176 7.6477894 8.9345342 7.3656994 9.8567621 9.7050146 9.3764679 7.1102196 9.391001 9.9445197 7.4729379 7.5595072 9.6353915 9.7687028 7.3919792 7.6081317 7.1462639 10.231692 0.185342391 0.00195944
    Rv0625c 7.2468973 7.3687697 7.1195149 7.4765619 6.2426194 6.6976093 7.5316606 8.0890591 7.2915739 8.8583593 8.742244 7.3234414 7.7464891 7.4765642 8.2429159 7.0683014 7.2878864 6.9659211 7.7304834 9.5570422 7.6386802 8.8651539 7.749955 8.9014746 9.276946 8.532363 7.6987162 9.4318712 9.2612184 8.2723944 7.5857514 9.39926 8.7466408 7.5781328 8.1292801 7.5621662 9.8464958 0.185320515 7.15E-05
    Rv2422 7.7727951 6.0884742 5.9210165 5.9259597 5.5858468 5.9419619 8.2021924 6.3550303 5.8177601 6.0624084 5.5927339 7.6225849 6.0995522 5.9614442 8.5780146 6.4906165 7.691769 5.9180417 6.912833 7.2780891 7.5059226 8.0701306 6.4393691 7.2064208 8.192907 6.7722778 7.8145178 8.1665219 8.1200264 6.9312998 7.4659131 7.416583 6.6259192 7.5558504 7.9649351 7.3968653 7.3184987 0.185042499 0.00091311
    Rv1472 6.335069 5.9938163 5.7924539 6.0698255 7.2673293 7.7941524 5.7054962 7.1693735 5.5572915 5.6888474 6.0761174 6.0772163 7.8898921 8.0826334 7.2735155 6.4335262 6.4952511 6.1530157 7.7919495 7.557498 6.8560977 8.586288 8.7471308 7.4708342 7.0833376 7.6259939 7.1277725 7.3861256 7.3506741 6.8019742 7.0325916 7.2107526 7.2025618 7.4333539 7.2938591 7.3767122 7.3900974 0.184009563 0.00022483
    Rv2908c 7.1023195 6.7163086 6.1920917 6.7883789 7.3259649 5.7949685 8.0658185 7.7299376 6.5948387 7.3289516 6.6299739 7.3172417 6.5756457 6.5256319 7.0396878 6.1953721 6.8316145 5.9235673 7.2220737 7.9576894 7.8417217 8.4885072 7.6561443 7.3609098 7.9255873 6.6819198 8.2284185 8.0937967 8.1602814 7.3827301 7.7528457 7.4117737 6.9197126 8.1449249 7.9021557 8.1981972 7.7567547 0.183778603 5.70E-06
    Rv2146c 7.3642645 6.3418363 6.1949909 6.0373594 7.4367817 7.1231383 7.8631726 5.6632841 6.3332439 6.2930149 7.8963988 7.4169425 6.7005994 6.6748641 5.9083298 5.9878095 7.3853629 5.9755836 5.8425859 8.3422314 6.6857605 7.6847837 6.4199789 8.3535139 8.2753207 8.5046292 6.6894504 8.2472334 8.0990367 7.5404969 6.5737801 9.3797403 8.0732623 6.4491535 7.4136572 6.6004347 9.4022174 0.18364447 0.00190786
    Rv3039c 6.8209556 7.1084086 7.3270572 7.0931082 7.7877039 7.4156983 5.7890413 6.8510811 7.3543974 7.7152746 7.8131946 6.8403366 7.6727626 7.6865003 7.0115211 7.5471375 7.5070389 7.5068619 7.2810939 9.2253075 7.5609144 7.8858366 7.0289646 8.6818643 8.7140322 8.749468 7.760091 8.4350922 8.3331039 6.734612 7.94025 9.3833084 8.9868433 8.3518467 8.4268214 8.2913909 9.0903275 0.183592989 1.95E-05
    Rv0842 7.338301 5.9053542 6.1274228 7.3851872 7.721419 7.3761463 7.783319 7.9468806 5.8031112 7.0904241 6.7046822 7.711804 6.7138446 6.8333149 7.6740521 6.5122302 7.2621888 7.1845097 7.2025968 8.77415 8.4360194 8.3940343 7.3320067 7.8569968 7.666508 7.6585857 7.8824902 7.9739625 9.108959 8.3779399 7.9857155 7.8490368 7.5291815 8.0410711 7.9653704 8.2560355 8.0029086 0.183181452 8.67E-06
    Rv1420 6.7367907 5.9671502 6.2377126 5.9228406 8.0767656 7.9815524 7.2176221 6.9165769 6.3523032 6.122467 6.0260759 6.909765 6.9209057 6.6714029 7.1031438 5.4311921 6.6282144 5.4539337 7.5769886 8.0520673 6.5931101 8.1307672 8.5184107 7.7734098 8.390093 7.2092282 6.9339465 7.8424493 8.1859655 6.282461 6.7680205 7.6500749 7.4523621 6.9553858 6.9497381 6.9154867 8.0351084 0.183038629 0.00023287
    Rv0248c 7.2581429 6.2638673 6.580919 5.9890519 7.2663051 8.2038034 6.7787055 6.7094222 6.3102936 6.9142826 7.732166 6.8618629 8.4148614 8.269184 8.6793546 6.6259068 6.7158914 6.0640317 6.6564862 8.7413155 8.2808036 8.0696482 8.5665309 8.6120846 8.636834 8.3383623 7.3954046 8.3302799 8.4871785 6.6141796 7.6672645 8.5860813 8.3285673 7.3621237 7.5843119 7.1519759 9.4914159 0.182535402 0.00044521
    Rv1241 7.6906717 6.1250641 6.0505745 6.3376288 7.0016506 5.6319396 8.2883431 7.1589178 6.2085455 6.2140016 6.0442238 7.8935086 6.0855481 5.6287823 7.9735286 6.2365735 7.5050185 5.647792 6.6296102 7.5856792 7.6402625 7.7247163 7.4857689 7.4378206 8.0499916 6.9269456 8.018951 8.2070058 8.6272207 6.7053508 7.5431087 7.4989721 6.5677593 7.7430906 7.4576139 7.6725751 7.8627741 0.182201726 0.00046616
    Rv0483 8.6439519 8.620336 8.560155 8.6207955 9.019563 8.779989 7.8830684 10.149821 9.6986563 7.6594887 8.0496865 9.7558819 8.9886402 5.6268556 8.2686447 9.0105494 9.1689225 8.1351502 8.3330794 9.3193597 9.1872016 10.763502 6.7235882 9.7233399 9.8037384 10.136767 10.587545 10.062226 10.989464 9.3462093 9.7065864 10.799596 9.7192027 10.913467 9.2988818 9.5813402 10.188437 0.182038296 0.00061134
    Rv2671 7.1868022 8.3846313 7.8603937 8.4081658 6.104603 6.7560885 6.5920919 8.2372002 8.2931609 7.8662429 8.2320447 7.2450422 6.7460367 7.6233947 7.3747548 7.8949412 8.4891431 7.9282551 7.716173 8.4433931 9.3027924 8.318875 6.7602421 9.0335384 7.8434474 8.1820972 10.235694 7.6829779 7.4911153 8.5295966 9.8242331 8.2910866 9.1227026 9.8623239 9.6219023 9.8148678 8.1062905 0.180783073 0.00042852
    RV1404 7.2124449 8.51104 8.5009532 8.2708053 7.966444 5.7125327 6.7500923 8.2431262 8.6328294 8.5413618 8.5605687 6.9204591 5.8321579 5.9206979 9.5096396 8.4309241 6.6806942 9.068908 8.3993044 10.295139 7.8871798 8.0746148 7.0202068 10.560727 10.453794 10.745343 7.3694483 13.758344 7.2494356 8.0822011 7.3802296 8.0783171 11.308075 7.5045814 7.4372925 7.6405769 7.3675657 0.180653289 0.02527516
    Rv3379c 7.5731754 5.8154319 6.0412005 5.7354843 6.8345847 7.7952554 7.4610169 7.2633988 5.7986362 7.2102479 6.6245199 7.817155 7.7480628 8.0608195 7.2245412 6.4067156 7.4019495 5.9530621 7.5850028 8.6650473 6.7379303 7.8649024 8.4202364 8.9059539 8.6801087 8.4653828 7.1608841 8.1110409 8.1979766 6.4780409 6.7501541 8.6917748 8.2208849 7.0790127 7.080964 7.0583247 9.1052954 0.180598367 0.00073036
    Rv1526c 6.5458638 7.9168452 7.6136389 6.8455065 7.8982413 7.6755877 7.7940877 6.9507891 7.1785048 7.0599914 7.3332323 7.3907052 6.043083 6.2775123 6.6338998 7.8537993 7.0670318 7.6325561 7.7069259 7.7631482 6.647541 10.24271 7.0484252 8.1768213 11.533374 8.408003 7.6066596 9.0742752 7.9575168 6.5772474 6.9399736 6.87253 8.0036544 7.8393825 7.4741401 8.6997973 10.572797 0.180314515 0.00463433
    Rv3042c 6.317055 7.4277001 6.5043224 8.502226 7.7672138 7.2809906 6.2935788 9.722582 6.7430463 6.9479511 8.1109014 7.0717699 7.0820401 7.7868858 8.8436067 7.0901844 9.0478147 7.0210602 9.1611764 8.9577551 7.7500185 8.1944056 6.9164608 9.1633502 8.9544334 7.9553631 7.793821 8.4683934 9.7977963 7.8695199 8.9757197 8.5646953 8.9169426 8.2489764 9.5825955 7.4593788 9.4057417 0.180265349 0.00079713
    Note. Fold change was defined as the log2-transformed ratio of active TB to LTBI. P values < 0.05 showed statistical significance; L, LTBI; A, active TB.

    Table Supplementary Table S1.  Raw data of the serum reactivity against 152 Mtb antigens

    Figure 1.  Hierarchical clustering of differentially expressed serum biomarkers. Each column indicates one serum sample and each row indicates one Mtbantigen. Data were normalized by Z score estimation, and the difference in serum reactivity was visualized as a color spectrum. A, active TB; L, latent TB infection.

  • STRING analysis was performed to investigate the interactions of 152 candidate antigens (Figure 2). Among these antigens, 13 pairs, namely, Rv2499c vs. Rv2503c, Rv2502c vs. Rv1070c, Rv0400c vs. Rv0914c, Rv2908c vs. Rv2906c, Rv0400c vs. Rv1472c, Rv0632c vs. Rv0400c, Rv0914c vs. Rv2503c, Rv0400c vs. Rv1070c, Rv0914c vs. Rv1472c, Rv0914c vs. Rv0632c, Rv0914c vs. Rv1070c, Rv3501c vs. Rv0589c, and Rv2502c vs. Rv2499c, exhibited strong associations with the highest confidence (combined score > 0.9), implying that multiple Mtb antigens may collaborate to participate in resistance to host immune responses.

    Figure 2.  STRING analyses of candidate Mtb antigens. Interaction networks were visualized with a confidence level of 0.25. Stronger associations are represented by thicker lines.

  • The functional classification of 152 antigens was further analyzed using the TB database (Figure 3; Supplementary Table S2, available in www.besjournal.com). These Mtb antigens were involved in various biological processes. The top three of these processes with known functions were intermediary metabolism and respiration (27.63%), cell wall and cell processes (19.74%), and lipid metabolism (11.84%). In addition, large proportions (12.50%) of the antigens were conserved hypothetical proteins whose particular functions remain unknown.

    Figure 3.  Functional category of candidate Mtb antigens. Analysis is based on the Sanger Institute database. N, number of antigens; PE/PPE, Pro-Glu/Pro-Pro-Glu.

    Annotation Rv Number
    1 intermediary metabolism and respiration Rv3841 Rv2421c Rv3324c Rv2097c Rv0187 Rv1815 Rv0062 Rv2499c Rv2716
    Rv2860c Rv3601c Rv2198c Rv1408 Rv3322c Rv2728c Rv2202c Rv0793 Rv3709c
    Rv0217c Rv2678c Rv1144 Rv2849c Rv2291 Rv0389 Rv1602 Rv2070c Rv2140c
    Rv0690c Rv2284 Rv1898 Rv1654 Rv1400c RV0687 Rv1524 Rv3001c Rv3029c Rv2216
    Rv0248c Rv2671 Rv3379c Rv1526c Rv3042c
    2 cell wall and cell processes Rv0379 Rv1337 Rv3743c Rv3625c Rv2903c Rv2804c Rv1270c Rv2288 Rv0258c
    Rv3277 Rv3906c Rv2559c Rv1197 Rv3533c Rv2723 Rv3495c Rv2520c Rv0476
    RV1237 Rv1411c Rv0308 Rv3035 Rv0970 Rv2398c Rv0394c Rv1842c Rv0625c
    Rv2146c Rv0842 Rv0483
    3 conserved hypotheticals Rv3190c Rv1498A Rv3899C Rv2632c Rv1725c Rv2114 Rv2342 Rv2023c Rv3259
    Rv1951c Rv2369c Rv2808 Rv2413c Rv0390 Rv3446c Rv2184c Rv3686c Rv3611
    Rv2422
    4 lipid metabolism Rv1483 Rv2928 Rv0214 Rv0242c Rv0632c Rv2484c Rv3107c Rv0270 Rv0557
    Rv0400c Rv2503c Rv2502c Rv1070c Rv0145 Rv0914c Rv2002 Rv1472 Rv3039c
    5 virulence, detoxification, adaptation Rv2031c Rv0597c RV2026C Rv1967 Rv0440 Rv1124 Rv1959c Rv3501c Rv0589
    Rv1247c Rv2530c Rv1942c Rv0251c Rv1241
    6 information pathways Rv2906c Rv3297 Rv0861c Rv2572c Rv1109c Rv3210c Rv1700 Rv3911 Rv3012c
    Rv0427c Rv3647c Rv2908c Rv1420
    7 regulatory proteins Rv1474c Rv0494 Rv2021c Rv2359 Rv0758 Rv3249c Rv1846c Rv2788 Rv0474
    RV1404
    8 insertion seqs and phages Rv3184 Rv1702c Rv3750c Rv1575
    9 PE/PPE Rv3144c Rv3590c
    Note. Annotations are according to Sanger Institute database (http://tuberculist.epfl.ch/).

    Table Supplementary Table S2.  Functional Classification of 152 Mtb Antigens

  • To validate the potential capability of candidate antigens for discriminating LTBI from active TB, 11 antigens that had high fold-change values and could easily be expressed and purified were selected and further assessed by ELISA using 279 serum samples including 94 healthy controls, 93 LTBI patients, and 92 active TB patients. As expected, the 11 antigen-specific IgG levels were significantly higher in active TB patients than in LTBI individuals, which is consistent with microarray results (Figure 4).

    Figure 4.  Scatter plots of IgG levels against 11 Mtb antigens in patients with active TB, LTBI, and healthy controls. Bars indicate means, and error bars indicate SEM. *P < 0.05; ***P < 0.001; ns, not significant.

  • The performance of 11 antigens in distinguishing between LTBI and active TB was evaluated using ROC curves, and their AUCs, specificities, and sensitivities were calculated (Table 2). Results showed that Rv2031c, Rv1408, and Rv2421c had higher AUC of 0.8520 (95% CI: 0.7954-0.9086), 0.8152 (95% CI: 0.7494-0.8809) and 0.7970 (95% CI: 0.7271-0.8668), respectively. Several other antigens, including Rv2716, Rv2002, Rv2097c, Rv0248c, Rv2026c, Rv0389, Rv2906c, and Rv2928, also distinguished between LTBI and active TB, and the results were statistically significant (P < 0.05).

    Rv Number Gene Name AUC (95% CI) PValue Sensitivity (%, 95% CI) Specificity (%, 95% CI)
    Rv2031c hspX 0.8520 (0.7954-0.9086) < 0.0001 75.00 (64.89-83.45) 84.95 (76.03-91.52)
    Rv1408 rpe 0.8152 (0.7494-0.8809) < 0.0001 75.00 (64.89-83.45) 87.10 (78.55-93.15)
    Rv2421c nadD 0.7970 (0.7270-0.8668) < 0.0001 77.17 (67.25-85.28) 82.80 (73.57-89.83)
    Rv2716 Rv2716 0.7931 (0.7212-0.8650) < 0.0001 66.30 (55.70-75.83) 97.85 (92.45-99.74)
    Rv2002 fabG3 0.7765 (0.7047-0.8484) < 0.0001 73.91 (63.71-82.52) 83.87 (74.80-90.68)
    Rv2097c Rv2097c 0.7184 (0.6415-0.7954) < 0.0001 60.87 (50.14-70.88) 80.65 (71.15-88.11)
    Rv0248c Rv0248c 0.7176 (0.6395-0.7956) < 0.0001 69.57 (59.10-78.73) 73.12 (62.92-81.79)
    Rv2026c Rv2026c 0.7010 (0.6237-0.7782) < 0.0001 67.39 (56.82-76.80) 69.89 (59.50-78.97)
    Rv0389 purT 0.7021 (0.6254-0.7789) < 0.0001 58.70 (47.95-68.87) 82.80 (73.57-89.83)
    Rv2906c trmD 0.6605 (0.5781-0.7430) 0.0002 52.17 (41.50-62.70) 89.25 (81.11-94.72)
    Rv2928 tesA 0.6584 (0.5760-0.7409) 0.0002 42.39 (32.15-53.14) 92.47 (85.11-96.92)
    Note. 95% CI: 95% confidence interval.

    Table 2.  AUCs for Discriminating between LTBI and Active TB Patients

  • According to AUC values, we selected the top three antigens, namely Rv2031c, Rv1408, and Rv2421c to perform antigen combination analysis. Our results showed that the positive rate of antigen combination in the LTBI group (namely, specificity) was increased to 93.6%, which is higher than that obtained for any single antigen (Figure 5A). There was a statistically significant difference in the detection rate of Rv2421c vs. combination (P = 0.002), and Rv2031c vs. combination (P = 0.039). The positive rate of antigen combination in the active TB group (namely, sensitivity) was increased to 81.5%, which was slightly higher than that observed for each antigen, although it was not statistically significant (Figure 5B). In addition, the positive rate of each antigen did not show any statistical difference with respect to the LTBI or the active TB group.

    Figure 5.  Distribution of positive and negative responders to Rv2031c, Rv1408, Rv2421c antigens, and a combination of these antigens. (A) Latent TB infection group. (B) Active tuberculosis group. For antigen combinations, a positive responder was determined if any two antigens yielded a positive result. Detection rates were compared using McNemar's test.

  • Logistic regression analysis was used to enhance diagnostic ability. Our results showed that Rv1408, R0248, Rv2026c, Rv2716, Rv2031c, Rv2928, and Rv2121c were the major predictive antigens for differentiating between LTBI and active TB patients. Therefore, a regression model Logit P = -6.28 + 16.92 × Rv1408 - 11.26 × R0248 + 19.82 × Rv2026c + 25.49 × Rv2716 + 11.35 × Rv2031c - 23.07 × Rv2928 + 11.55 × Rv2121c was established. In the training set, the AUC of the model was 0.9844 (95% CI: 96.62%- 1.003%), with sensitivity and specificity of 96.77% (95% CI: 88.83%-99.61%) and 93.75% (95% CI: 84.76%-98.27%), respectively (Figure 6A). In the validation set, the AUC of the model was 0.9632 (95% CI: 91.40%-1.012%), with sensitivity and specificity of 93.33% (95% CI: 77.93%-99.18%) and 93.1% (95% CI: 77.23%-99.15%), respectively (Figure 6B).

    Figure 6.  ROC curve analysis of logistic regression model. (A) Training set. (B) Validation set. AUC, sensitivity, and specificity are shown.

  • Since Mtb is an intracellular pathogen, cell-mediated immunity has always been considered as the predominant defense mechanism against Mtb infection. However, accumulating evidence suggests that B cell-mediated humoral immune response may play an important role in Mtb infection and is related to the progression of latent infection to active TB disease[15-18]. Previous studies showed several Mtb antigens such as ESAT-6 (Rv3875), CFP-10 (Rv3874), Ag85B (Rv1886c), Hsp16.3 (16-kD, Rv2031c), 38 kD (Rv0934), LAM, Rv2029c, Rv2628, and Rv1813c have previously been reported to be able to discriminate between LTBI and active TB[19-21]. Recently, Lu et al.[22] demonstrated that antibody glycosylation patterns could also distinguish between LTBI and active TB. In our study, we aimed to screen novel serum biomarkers for discrimination between LTBI and active TB at the systemic level using an Mtb proteome microarray containing 4, 262 antigens. We found that the concentrations of 152 Mtb antigen-specific IgG antibodies were higher in the active TB group than in the LTBI group. This result provided potential serum biomarkers and data that may facilitate a better understanding of differences in the host humoral immune response between LTBI and active TB. Among previously reported and commonly used antigens, only Rv2031c was included in our results. This may be because the protein antigens used in previous studies were all produced using an Escherichia coli-based cell-free transcription system and without further purification. However, in our study, the antigens spotted on the Mtb proteome microarray were all produced using a eukaryotic yeast expression system. The use of a eukaryotic system results in the expressed protein antigens having many more posttranslational modifications than those produced using a prokaryotic system. In the present study, the antigens were obtained by gently using affinity purification to maintain the activity that is necessary for functional studies[16]. This is important since specific features of these antigens may have a great influence on the antibody response. In addition, the enrollment of subjects with a different infection time or status and with or without anti-TB treatment may impact the results. These reasons may underlie the inconsistency of current approaches for the serological diagnosis of TB.

    In our validation experiments using the 11 candidate antigens and ELISA, we used healthy controls to ensure the accuracy of the test, i.e., the levels of antigen-specific IgG antibodies must be higher in both the LTBI and active TB groups than in the healthy control group. It is noteworthy that Rv0389, Rv2002, and Rv2026c showed significant differences among the healthy control, LTBI, and active TB groups, suggesting that these antigens may elicit stronger humoral immune responses despite low bacterial load in LTBI individuals. Therefore, Rv0389, Rv2002, and Rv2026c antigens could be potential biomarkers for diagnosing Mtb infection and determining disease status. However, further studies in this regard are warranted.

    Antigens assessed in this study (Rv2031c, Rv1408, and Rv2421c) could not discriminate between LTBI subjects and healthy controls. However, they could distinguish between LTBI and active TB with sufficient accuracy, indicating that these antigens may have potential value by offering improved predictive accuracy when used in combination with IGRAs. Rv2031c is involved in virulence, detoxification, and adaptation to biological processes and is predominantly expressed by Mtb during latency[23, 24]. In contrast, both Rv1408 and Rv2421c are involved in intermediary metabolism and respiration, which are required for the long-term survival of Mtb. Specifically, Rv1408 is ribulose-phosphate 3-epimerase (rpe), which catalyzes the reversible epimerization of D-ribulose 5-phosphate to D-xylulose 5-phosphate. Previous studies showed that Rv1408 expression was downregulated during nitrogen starvation, suggesting that it was involved in nitrate/nitrite metabolism[25]. Rv2421c is nicotinate-nucleotide adenylyltransferase, which transfers phosphate groups during NAD biosynthesis and salvage. Cloete et al. reported that Rv2421c was upregulated during starvation or dormancy by hypoxia[26].This is the first study to demonstrate that Rv1408 and Rv2421c antigens might be used for distinguishing LTBI from active TB patients.

    Since significant heterogeneity exists among individuals and various regions, a single antigen can never achieve satisfactory performance for the diagnosis of TB. Deng et al.[14] have demonstrated that the combination of several serum biomarkers showed better performance in distinguishing between active TB patients and recovered individuals, with 78.4% diagnostic accuracy, 80.2% sensitivity, and 79.0% specificity.He et al.[27] reported that the combination of Rv3618 with the 38 kD 38E6 could improve the sensitivity of TB diagnosis. In this study, we combined several antigens to improve diagnostic ability. We selected the top three antigens with high AUC values to conduct antigen combination analysis. Our results showed that the specificity of the antigen combination was significantly higher than those of single antigens. We also used logistic regression analysis to develop a regression model containing seven antigens and cross-validated the results using another independent sample. Once again, our results showed that both specificity and sensitivity were significantly better than those observed with any single antigen. Although logistic regression analysis is rarely used in studies on TB, it has been applied in several other studies[28, 29]. However, it is noteworthy that the seven antigens included in the regression model were not the top seven out of the 11 antigens according to AUC values. Some of them had relatively lower AUCs but higher specificity than other antigens, which when combined with other antigens of higher sensitivity, may result in a mutual complementation pattern for distinguishing LTBI from active TB.

    However, this study still has certain limitations. The primary limitation is that the sputum smear or bacterial culture of active TB patients were Mtb-positive and bacteria-negative TB patients were not considered. In addition, the sample size in the logistic regression model construction and validation phases was small. Therefore, further studies for verifying the diagnostic performance of the regression model with larger sample size, including bacteria-negative TB patients, are required in the future.

    In conclusion, we have revealed that a variety of Mtb antigens could elicit stronger serum IgG responses in active TB patients when compared with LTBI subjects using proteome microarrays, which may be useful for understanding host humoral immune responses in individuals with different Mtb infection statuses. Furthermore, several antigens were further validated. Each of these antigens exhibited a moderate diagnostic performance in discriminating between active TB and LTBI while using the top three antigens in combination or using a logistic regression model established with seven antigens could improve the diagnostic sensitivity and specificity. Of these two approaches, the logistic regression model with seven antigens had the best performance, suggesting that it may serve as an index for distinguishing between active TB and LTBI.

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