Methods for Diagnosis of Tuberculosis

Abstract

Methods for diagnosis of tuberculosis are disclosed. In particular, the invention relates to the use of a panel of biomarkers for aiding diagnosis, prognosis, and treatment of tuberculosis. The identified biomarkers can be used to detect active tuberculosis as well as to distinguish active tuberculosis from latent tuberculosis and other pulmonary and infectious diseases, and for monitoring responses to treatment.

1 - 34 . (canceled) 
     
     
         35 . A method of treating a patient for tuberculosis, comprising:
 (a) identifying the patient as having tuberculosis based on the expression levels of biomarkers for GBP5, DUSP3, and a normalization gene in a biological sample; and   (b) administering an effective amount of at least one antibiotic to the patient.   
     
     
         36 . The method of  claim 35 , wherein step (a) further comprises identifying the patient as having tuberculosis based on the expression levels of biomarkers for GBP5, DUSP3, KLF2, and the normalization gene. 
     
     
         37 . The method of  claim 35 , wherein step (a) comprises distinguishing active tuberculosis from latent tuberculosis by analyzing the expression levels of each biomarker in conjunction with respective reference value ranges for each biomarker,
 wherein similarity of the expression levels of biomarkers for GBP5, DUSP3, and the normalization gene to reference value ranges for a patient with active tuberculosis indicates that the patient has active tuberculosis, and similarity of the expression levels of biomarkers for GBP5, DUSP3, and the normalization gene to reference value ranges for a patient with latent tuberculosis indicates that the patient has latent tuberculosis.   
     
     
         38 . The method of  claim 37 , wherein step (b) comprises a different treatment regimen for the patient identified as having latent tuberculosis than the patient identified as having active tuberculosis. 
     
     
         39 . The method of  claim 37 ,
 wherein the patient identified as having active tuberculosis is administered more than one antibiotic for a period of greater than one month, to treat active tuberculosis, and   wherein the patient identified as having latent tuberculosis is administered at least one antibiotic for a period of less than 12 months, to treat latent tuberculosis.   
     
     
         40 . The method of  claim 35 , wherein the at least one antibiotic is selected from the group consisting of rifampicin, isoniazid, pyrazinamide, ethambutol, rifapentine, ethionamide, moxifloxacin, and streptomycin. 
     
     
         41 . The method of  claim 37 , further comprising administering an effective amount of a corticosteroid to the patient with active tuberculosis. 
     
     
         42 . The method of  claim 37 , further comprising
 monitoring the patient's response to treatment or relapse after treatment to determine treatment length or antibiotic resistance, or prevent latent tuberculosis progressing to active tuberculosis in the patient.   
     
     
         43 . The method of  claim 35 , further comprising evaluating disease severity in the patient, by calculating a TB score for the patient based on the levels of expression of the biomarkers, wherein an increasing TB score indicates worsening tuberculosis infection and a decreasing TB score indicates recovery from active tuberculosis, or wherein a decreasing TB score indicates worsening tuberculosis infection and a decreasing TB score indicates recovery from active tuberculosis. 
     
     
         44 . The method of  claim 35 , wherein the biological sample comprises whole blood, sputum, monocytes, or macrophages. 
     
     
         45 . The method of  claim 35 , wherein the biomarkers are RNA biomarkers quantified by PCR. 
     
     
         46 . The method of  claim 45 , comprising:
 a) contacting the biological sample from the patient with sets of primers that detect biomarkers for GBP5, DUSP3, and the normalization gene in the biological sample;   b) conducting one or more polymerase chain reaction (PCR);   c) generating amplicons that is produced by the PCR for GBP5, DUSP3, and the normalization gene;   d) contacting the amplicons with at least one probe for each of said biomarkers, wherein each probe comprises a detectable label.   
     
     
         47 . A method of diagnosing latent tuberculosis in a patient, comprising:
 a) obtaining a first biological sample from the patient;   b) detecting and analyzing expression levels of biomarkers for GBP5, DUSP3, and KLF2 in the first biological sample, in conjunction with respective reference value ranges for said biomarkers, wherein similarity of the levels of expression of biomarkers for GBP5, DUSP3, and KLF2 to reference value ranges for a patient with latent tuberculosis indicate that the patient has latent tuberculosis; and   c) administering a treatment regimen comprising an antibiotic or a combination of antibiotics for treating latent tuberculosis, to the patient.   
     
     
         48 . The method of  claim 47 , wherein step b) further comprises detecting and analyzing expression levels of a normalization gene. 
     
     
         49 . The method of  claim 47 , further comprising monitoring the patient's response to treatment by
 e) measuring levels of expression of biomarkers for GBP5, DUSP3, and KLF2 in a second biological sample from the patient, wherein the second biological sample is obtained from the patient at a second time point;   f) comparing the levels of expression of the biomarkers in the first biological sample to the levels of expression of the biomarkers in the second biological sample, or calculating TB scores based on the levels of expression of the biomarkers in the first biological sample and the second biological sample, to determine if the tuberculosis infection in the patient is improving or worsening; and   e) optionally administering a second treatment regimen that results in improved health conditions of the patient.   
     
     
         50 . The method of  claim 47 , wherein the subject suspected or at risk of developing tuberculosis displays one or more characteristic symptoms of tuberculosis, or have a genetic predisposition or presence of one or more developmental, environmental, or behavioral risk factors for tuberculosis. 
     
     
         51 . A kit comprising primers and probes for detecting and/or measuring expression levels of biomarkers for GBP5, DUSP3, and a normalization gene,
 wherein the primers comprise a first polymerase chain reaction primer pair for detecting a biomarker for GBP5, a second polymerase chain reaction primer pair for detecting a biomarker for DUSP3, and a third polymerase chain reaction primer pair for detecting a biomarker for the normalization gene; and   wherein the probes comprise at least one polymerase chain reaction probe for detecting a biomarker for GBP5, at least one polymerase chain reaction probe for detecting a biomarker for DUSP3, and at least one polymerase chain reaction probe for detecting a biomarker for the normalization gene.   
     
     
         52 . The kit of  claim 51 , wherein the kit further comprises,
 a fourth polymerase chain reaction primer pair for detecting a biomarker for KLF2; and   at least one polymerase chain reaction probe for detecting a biomarker for KLF2.   
     
     
         53 . The kit of  claim 51 , wherein each probe comprises a detectable label. 
     
     
         54 . The kit of  claim 53 , wherein each probe comprises a fluorescent dye and a quencher molecule.