This document summarizes various methods for diagnosing tuberculosis (TB) including: 1. Radiographic examination of the chest to identify features of primary or reactivated TB. 2. Laboratory tests like smear microscopy, culture and nucleic acid amplification to detect Mycobacterium tuberculosis from respiratory or other specimens. 3. Immunological tests like tuberculin skin test, interferon-gamma release assays and serological tests to detect TB infection.

25. Tuberculin skin test ▪ TST has been used to identify patients actively infected with TB, ▪ to measure the prevalence of infection in a community, and to select susceptible or high-risk patients for BCG vaccination . How to do TST : ▪ TST works by intradermally injecting 0.1 mL of 5 TU PPD on the forearm. (Mantoux's method) On examination, after 48-72 hours, a positive reaction is indicated by erythema and induration of > 10 mm in size. Erythema (redness) alone is not taken as a positive reaction . ▪ To detect persons at high risk for LTBI or at high risk to progress to disease

27. ☻ A positive TST result is a manifestation of type IV delayed hypersensitivity. □ False-positive test findings can occur in ; ● cross-reactions caused by non-tuberculous (atypical) mycobacterial infection ● bacille Calmette-Gu´ erin (BCG) vaccination.

28. Causes of False-Negative Tuberculin Skin Test 1: Factors Releated to the Person Being Tested ● Concurrent infections ● Human immunodeficiency virus ● Other viral infections (measles, mumps, chickenpox) ● Bacterial diseases (typhoid fever, brucellosis, typhus, leprosy, pertussis, overwheiming tuberculosis) ● virus vaccination (measles, mumps, polio) ● Diseases affecting lymphoid organs (Hodgkin's disease, lymphoma, chronic lymphocytic leukemia, sarcoidosis) ● Age (newborns, elderly) ● Metabolic conditions (advanced renal failure, malnutrition) ● Recent surgery ● Certain medications (e.g.,TNF- α antagonists, corticosteroids and other immunosuppressive drugs), will suppress the IV response and T-lymphocyte function

29. 2 : Factors Related to the Test ● Loss of antigen potency (exposure to heat and/or light) ● Incorrect administration (too little antigen, deep injection) ● Incorrect reading (inexperienced reader, reader bias, recording error

32. Enzyme-linked immunospot for interferon-gamma The ELISPOT assay for diagnosis of M. tuberculosis infection is based on the rapid detection of T cells specific for M. tuberculosis antigens. IFN-γ released in vivo from these cells can be detected by the extremely sensitive ELISPOT In a preliminary trial, this test was positive in 96 % of 47 TB patients and in 85 % of 26 persons presumed to have latent TB. The ELISPOT test was negative in 26 BCG-vaccinated control subjects, and this specificity implies a major advantage over TST ELISPOT offers a more accurate approach than TST for the identification of individuals who have latent TB infection . Each such T cell gives rise to a dark spot and the readout is the number of spots. The T cells enumerated by the ELISPOT assay are effector cells that have recently encountered antigen in vivo and can rapidly release IFN-γ when re- exposed to the antigen

33. Quantiferon-TB test The IFN-γ secreted by T-cells into the plasma is measured by ELISA to indicate the likelihood of TB infection . Different studies demonstrated that the QuantiFERON-TB test was comparable to TST in its ability to detect latent TB infection. also showed that the QuantiFERON-TB test was less affected by BCG vaccination QuantiFERON-TB was approved by the Food and Drug Administration (FDA) of the United States (US) in 2001. In 2003, the US Centers for Disease Control and Prevention released guidelines for using the QuantiFERON®-TB Test in the diag- nosis of latent M. tuberculosis infection ,

34. QuantiFERON-TB Gold , This in vitro test detects the release of interferon-gamma (IFN-γ) from lymphocytes of sensitized persons when their blood is incubated with peptide mixtures simulating two M. tuberculosis proteins called ESAT-6 and CFP-10 . the TB-specific antigens ESAT-6 and CFP- 10 that are only present in M. tuberculosis and are absent from all strains of M.bovis (BCG) and most environmental mycobacteria ,

35. LIMITATIONS IFN γ assays are expensive tests and their higher cost appears to limit their wider applicability, especially in resource-limited settings and developing countries,. The ELISPOT test is not yet suitable for widespread use, because it is costly and requires isolation of mononuclear cells, a procedure that is not performed in clinical laboratories .