As we know Tuberculosis is a major killer among infectious diseases globally and The only reservoir of Tuberculosis is humans. It become very important to diagnose tuberculosis early to stop the transmission of disease.

India is among those 30 high TB burden countries that contribute to around 87% of all cases in the world³. Therefore early diagnosis of Tuberculosis in India becomes even more important.

Diagnosis of Pulmonary Tuberculosis

Of all the systems, tuberculosis primarily affects the lungs. Tuberculosis of the lungs is called Pulmonary tuberculosis.

An algorithm for the diagnosis of Pulmonary TB is given below:-

Suptum Smear Microscopy

sputum sample of the patient is taken and spread over the glass slide to see the bacilli causing tuberculosis under the microscope.

• Only those presenting with symptoms suggestive of Pulmonary tuberculosis are offered Sputum Smear Microscopy. Sputum smear microscopy is also done for the follow-up of pulmonary TB patients.
• 2 samples of sputum are taken. One is taken on the spot (spot sample) under supervision during the interaction with the patient. Another sample is taken in the early morning in the container provided to the patient.
• Both samples collected from patients are examined under the microscope after preparing the smear on the slides.
• Two techniques are used to prepare the slides:-
  • ZN (zeihl-Neelson) staining and
  • Fluorescence staining
• sputum smear microscopy though cheap and widely available, suffers from a number of limitations also as low sensitivity, operator-dependent results, quality of sputum sample, etc.

Molecular test

1. CBNAAT

CBNAAT (Cartridge Based Nucleic Acid Amplification Test) is a molecular test that can detect a very small number of Tuberculosis bacilli in two hours. It has the additional advantage of detecting Rifampicin resistance. 

Molecular testing is done in the:-

• All the smear-positive tuberculosis patients to know the status of Rifampicin resistance.
• Upfront CBNAAT (directly CBNAAT, without sputum smear) is done in diabetics, persons living with HIV, and children. since in children the sample collection is difficult, therefore sputum smear is skipped and direct CBNAAT is done.
• It is not done in patients who have completed their TB treatment, as it can detect even dead bacteria.
• It takes around 2 hours to give results.

2. Truenat

It is a simpler chip-based version of CBNAAT which can be decentralized for better penetration into the periphery for quicker molecular tests.

3. LPA (Line Probe Assay)

LPA is done in all the smear or CBNAAT-positive cases to know to decide on the treatment modifications if needed. It detects the Isoniazid resistance in addition to Rifampicin resistance. LPA require 2 sputum samples (Spot and Early Morning). It takes around 72 hours for test results to come but requires a sophisticated lab setup.

Culture test

In the culture test the sample is processed and cultured on various media to grow the colonies of mycobacterium. Culture tests are highly sensitive and specific and are considered the gold standard for the diagnosis of TB. Since they take around 2-8 weeks time for the results, they have a limited role in the early diagnosis of TB. however, they are used for the follow-up of drug-resistant TB cases to ensure a relapse-free cure. They include:-

• Solid culture media (LJ Media)
• Liquid culture systems eg- BACTEC MGIT 960

Liquid culture is done for those samples which came out Rifampicin resistant on CBNAAT or LPA. it can detect the resistance for almost all anti-tubercular drugs.

Imaging test

Though not so specific, may be used to support the diagnosis when confirmatory test cant be used. they include:-

Chest X-ray

Chest X-ray is also done in infections involving the respiratory system. 

• It is not a specific test for Tuberculosis.
• It can narrow the scope but cannot confirm the diagnosis of Tuberculosis 
• similar findings on X-ray can be found in other diseases like silicosis, bronchiectasis or pneumonia.
• It also can not differentiate active tuberculosis from healed tuberculosis.

Therefore X-ray is a supportive investigation, not a diagnostic.

CT scan

Although a CT scan is not done routinely to diagnose TB, some characteristic findings may be seen on the CT chest even when the chest X-ray is normal. 

Diagnosis of Extra-Pulmonary Tuberculosis

• Tuberculosis other than that of the Lungs is called extra-pulmonary tuberculosis. It is usually difficult to demonstrate the TB bacteria, hence diagnosis is made primarily on clinical suspicion.
• some specific tests may be used depending upon the organ involved like:- FNAC of lymph nodes (in the TB of lymph nodes) and CBNAAT.

• Attempts should be made to establish the microbiological confirmation of presumptive extrapulmonary TB by collecting the sample and performing CBNAAT.
  • Samples like CSF (cerebrospinal fluid), pleural fluid, synovial fluid, and ascitic fluid can be subjected to CBNAAT, though the sensitivity varies.
  • Negative CBNAAT in these samples does not rule out tuberculosis, whereas positive CBNAAT confirms the disease.

• If CBNAAT is negative or can not be performed, Other investigations like chest X-ray, Ultrasound imaging, CT Scan, and Magnetic resonance imaging (MRI) can be used to support the clinical diagnosis.

Diagnosis of Latent Tuberculosis

As we know bacilli casing TB sometimes remains dormant in the body and doesn’t cause the disease. This is called latent TB. latent TB can become Active TB at any time during the lifetime. Therefore it becomes important to diagnose and treat Latent TB before it flourishes into a disease.

The test offered to diagnose latent TB:-

1. TST (Tuberculin Skin Test) also known as Montoux Tuberculin Skin Test
2. IGRA (Interferon Gamma Releasing Assay)

At present These tests are offered to:-

• All the non- symptomatic Household contacts of pulmonary tuberculosis patients (symptomatic household contacts of Pulmonary TB are screened for active TB as per the algorithm of diagnosis)
• Population with weak immune systems like cancer patients, those on immuno-suppressive treatment, organ transplant recipients, and persons on interferon-gamma etc.

Tuberculin Skin Test 

Also known as the Montoux test. 

Principal: 

• delayed hypersensitivity reaction (type IV hypersensitivity reaction) is the basis of TST. When a small amount (0.1ml) of antigen (PPD^* RT23 or PPD-S) similar to TB bacteria is injected into the body, The cell-mediated immunity of the body having mycobacterium reacts to the antigen and induration appears with or without redness.
  • *PPD is a Purified Protein Derivative, the mycobacterial antigen.

How is it done:-

• 0.1 ml tuberculin antigen (2 TU of PPD RT23 or 5 TU of PPD-S) is injected intra-dermal at the forearm on the volar aspect. Bleb (wheal) formation is ensured for successful injection. If bleb (wheal) is not formed or is less than 6mm, injection is repeated 2 inches away from the previous site or on the other arm.

Reading the test:-

• The patient has to visit the doctor again after 48 hours of the injection (48-72 hours of injection) for assessment of the test. Induration is noted by following a method as shown in the video. If this induration comes out to be more than 10mm it is taken as reactive and the patient is offered TPT.

The Tuberculin test is not specific and may be found positive in the individuals vaccinated with the BCG vaccine. Since TB is quite common in India we put individuals on the TPT based on TST results.

IGRA (Interferon-Gamma Release Assay)

The blood sample of the patient is mixed with the antigen of Mycobacterium. WBC of an individual infected with Mycobacterium releases the Interferon-gamma on interaction with the antigen of Mycobacterium tuberculosis.

It is a relatively more specific test. Blood samples of non-symptomatic household contacts of pulmonary TB patients are tested. after ruling out active TB, when a person is found positive for IGRA is put on the TPT (TB Preventive Treatment).

• IGRA is a bit expensive, in Himachal Pradesh it is offered free of cost.
• prior BCG vaccination has no effect on it.
• A blood sample has to be processed within 8-30 hours. 
• It doesn’t differentiate between active and latent TB.
• There is limited data on IGRA in children less than 5 years therefore they are offered TST for now.

Diagnosis of Tuberculosis in Children

All attempts should be made to establish the microbiological diagnosis. CBNAAT is the preferred investigation. If CBNAAT is possible due to some reason, sputum smear microscopy can be performed. If the Sample could not be retrieved or is not possible, a Chest X-ray and TST (Mantoux test) with 2 TU of PPD RT 23 is done.