Introduction

Tuberculosis (TB) is a potentially fatal bacterial infection caused by the bacterium Mycobacterium tuberculosis (M. tuberculosis). It is a highly contagious droplet infection that primarily affects the lungs. However, it can affect numerous organ systems. Treatment for TB is available and effective, yet it remains a significant public health concern and a leading cause of morbidity and mortality worldwide, especially in developing countries. It is estimated that more than 1.7 billion people are infected with M. tuberculosis. Although the overall incidence and prevalence of tuberculosis have declined, the incidence of multidrug-resistant tuberculosis remains significant.[1]

Active and Latent TB Infection (LTBI)

A person with an active infection usually presents with constitutional symptoms, including unexplained weight loss, fever, fatigue, loss of appetite, and night sweats. LTBI is asymptomatic and non-infectious.

Early diagnosis of active TB is crucial to managing and preventing its spread. LTBI has a worldwide prevalence of 33%. Those with immunocompromised states are at an increased risk of reactivation and progression to active TB disease, which is symptomatic and highly contagious. The risk of LTBI progressing to active disease is greatest within the first two years of exposure. Therefore, detection and treatment of latent TB are critical to controlling the spread of TB and reducing the disease burden.[2][3]

The risk of LTBI progression to active disease declines with age, secondary to increased immunity. The risk of progression in infants is 50%, and this decreases to 1% to 2% by age ten.[4]

TB Screening Tests

Two screening tests are available to detect TB infection in the United States:

• Tuberculin skin (TST) or purified-protein derivative (PPD) skin test via the Mantoux technique:[5][6]
• Interferon-gamma release assay blood test (IGRA)[7]

Both tests measure delayed-type hypersensitivity reaction or type IV cell-mediated immunity involving T-lymphocytes, which are activated after exposure to mycobacteria. Notably, a positive test does not distinguish between latent and active TB. Therefore, symptom assessment and further testing with chest radiography, sputum test for acid-fast bacilli, or chest computed tomography scans are essential to diagnosing active infection.

There is no definitive test to diagnose LTBI. Instead, it is a clinical diagnosis based on a history of prior TB infection and ruling out active disease.

PPD Skin Test (TST)

Robert Koch described the tuberculin reaction in 1890, and Felix Mandel developed the test in 1908. Charles Mantoux was credited with creating the technique of injecting material intradermally on the inner surface of the forearm. In 1934, Florence Seibert published her method of obtaining the purified-protein derivative, effectively creating the PPD test.[8]

The tuberculin protein used in the test is extracted from Mycobacterium tuberculosis cultures and is used as a purified-protein derivative. A standardized PPD-S utilizes a tuberculous mycobacterium. Non-tuberculous mycobacteria are identified by a letter other than S. The test results are interpreted by measuring the delayed-type hypersensitivity reaction. The peak of the induration reaction occurs 24 hours after the test and is secondary to cell infiltration.

It takes approximately six to eight weeks after exposure to bacteria for the PPD test to become positive. The PPD test should be read between 48 and 72 hours after administration.[9]

Specimen Collection

The PPD TST is performed on the surface of the forearm; no specimen collection is required.[10]

Procedures

Administration of PPD Skin Test

In the United States, two US Food and Drug Administration (FDA)-approved PPD tuberculin antigens are available: Tubersol and Aplisol. Other tuberculin antigens, such as RT-23, are sometimes used outside the United States. The dosage of PPD-S and RT-23 formulations differ. The dosage for PPD-S formulation is five units in 0.1 ml. No dose adjustments are required in hepatic and renal failure patients. 

According to the Centers for Disease Control (CDC), this test is performed using the Mantoux technique, which involves injecting 0.1 mL of a solution containing five units of tuberculin-purified protein derivative into the inner surface of the forearm via the intradermal route. It should be administered two or more inches from the elbow, wrist, or other injection sites. According to the 2005 CDC guidelines, it can be administered to the back of the shoulder if neither arm can be used.

This test is performed using a one mL tuberculin syringe and a needle less than 0.5 inches, with the needle bevel facing upward. The needle should be inserted slowly at an angle of five to fifteen degrees, and the bevel should be visible below the skin. A wheal or elevation of the skin, six to ten mm in diameter, is formed when a small amount of the PPD solution is injected intradermally, ensuring the correct administration. If a wheal is not created, the test must be repeated immediately on another site at least five cm from the original administration site.

Documentation of the injection site, date and time of test administration, the name of the person performing the test, product lot number, and the manufacturer should be done. The patient should avoid touching the area and keep the site uncovered and clean. Since an allergic reaction to tuberculin is possible, epinephrine should be available.

Reading of the PPD Skin Test

A type IV delayed hypersensitivity reaction to the injected tuberculin PPD antigen is induced. It is a time-sensitive test. The reaction begins five to six hours after intradermal test placement but should be measured 48 to 72 hours after test placement when the peak effect occurs. After 48 to 72 hours, the reaction begins to subside. Erythema and induration are seen at the injection site. The diameter of induration should be demarcated and measured in mm perpendicularly to the long axis of the forearm. Erythema has no diagnostic value and should not be measured. Induration is palpable and should be measured by inspection and palpation.[11][2]

Tests that are read after 72 hours are inaccurate as they tend to underestimate the size of the skin reaction, and therefore, the reliability of the test is compromised. Repeat testing is recommended if the reaction is not read within 72 hours. The repeat test should be administered as soon as possible, preferably within seven days of the initial test, to avoid a boosting effect. The second test site should be in a different location, like the other arm.[2]

Proper administration and interpretation of the PPD skin test require standardizing this test procedure, adequate staff training, and supervision. Although the PPD skin test is commonly used worldwide, its interpretation is challenging.[5] If such assurances cannot be made, IGRA testing should be considered.

Indications

Screening for infection with M. tuberculosis is indicated as a part of baseline TB risk assessment or for evaluating symptoms consistent with TB. Screening tests include the TST and IGRA, as described above. Screening tests are not recommended as a part of routine healthcare and should be performed in high-risk populations to detect latent or active TB disease.

According to 2017 guidelines for the diagnosis of active or latent TB, published by the CDC, American Thoracic Society (ATS), and Infectious Diseases Society of America (IDSA), TB screening should be performed in patients with an increased risk of new TB infection and those with an increased risk of reactivation disease.

Patients with an increased risk of new TB infection are those with known exposure to M. tuberculosis, those with close and casual contact with a patient with untreated, active respiratory TB, individuals with LTBI and an increased risk of disease progression, employees and residents of homeless shelters and correctional facilities, and some healthcare workers. Certain healthcare workers, including pulmonologists and respiratory therapists, require serial testing, given their high risk of exposure.[12]

The risk of reactivation is six times higher in individuals with severe immunocompromise, such as chemotherapy or organ transplant, HIV infection, lymphoma, leukemia, head or neck cancer, those with evidence of healed TB on chest radiograph, silicosis, renal failure requiring hemodialysis, and those receiving treatment with tumor necrosis factor-alpha (TNF) inhibitors. Therefore, all individuals meeting these criteria should have a single test to rule out TB infection.

The risk of reactivation is three to six times higher in individuals with diabetes mellitus and those on systemic glucocorticoid therapy (greater than 15 mg/day for more than one month). Testing should be limited to those with an increased prevalence of TB, including those living homeless, those who use injection drugs, contacts of those with active TB, and those born outside the US who emigrated from countries with an incidence of TB greater than 100/100,000.

The risk of reactivation is 1.5 to three times higher in individuals weighing less than 85% of ideal body weight, those who smoke cigarettes, and those with evidence of small granulomas on chest radiography. Testing should be individualized for this group, with particular consideration for those living homeless, those who use injection drugs, contacts of those with active TB, and those born outside the US who emigrated from countries with an incidence of TB greater than 100/100,000.[13]

Screening test selection should be based on CDC, ATS, and IDSA guidelines. The PPD skin test is preferred over the IGRA test for serial testing of individuals who should be tested regularly. This is because repeat IGRA test results are difficult to interpret. An IGRA is preferred over the PPD skin test when there is a low to intermediate risk of progression from latent to active disease. An IGRA is also the preferred screening test when the patient is unlikely to return to have the PPD test read, has a history of prior BCG vaccination, or has a previous history of non-tuberculous infection.

The PPD test may be an alternative when IGRA is unavailable or too expensive. In addition, either test may be used if there is a high risk of progression to active disease.

Dual testing with IGRA and PPD (in any order) is performed in low-risk individuals if the initial test is positive. The law requires latent TB infection testing; however, it is not indicated medically. A positive PPD test is followed by IGRA testing to confirm the presence of infection. IGRA testing should be done within three days of initial test placement. LTBI treatment is offered to individuals who are positive for both tests.

In 2019, the CDC updated guidelines for PPD screening and testing recommendations indicated in healthcare personnel. In the setting of a previously negative TB test and no prior history of TB disease, testing with TST (PPD) or IGRA should occur. If IGRA testing is positive or the TST is greater than five mm, the patient should be considered as having a new TB infection. If the initial test is negative, retesting should occur eight to ten weeks after known exposure to TB. The repeat TST is positive if induration is greater than ten mm and the measurement has increased from the first test by greater than six mm. Serial testing with IGRA in healthcare personnel is undefined, mainly secondary to increased rates of false conversion.

A documented prior positive TST does not require retesting but rather a TB symptom screen. However, if symptoms are present, further evaluation should occur. Screening should include individual baseline TB risk assessment. Routine annual screening should not be done without occupational risk or exposure.

In 2020, the World Health Organization released its guidelines for diagnosing and treating TB infection. The PPD skin test or the IGRA may be used to diagnose TB, without preference for either. 

In 2021, the National Tuberculosis Controllers Association (NTCA) released its guidance for screening, diagnosing, and treating TB infection. IGRA testing is preferred for most patients born outside the US who were vaccinated with or may have been vaccinated with BCG. For others, either IGRA testing or a PPD TST may be used. Routine use of testing with both the PPD TST and IGRA is not advised but may be considered for those who may have a decreased immune response to the tests, those at risk for severe disease, and those in whom there is a high index of suspicion of infection. In addition, the PPD TST or IGRA may be used for low-risk patients requiring testing. Confirmatory retesting with either method should be performed if the test is positive.

Potential Diagnosis

A positive skin reaction may occur in the following situations:

• Active TB infection
• Latent TB infection
• History of BCG vaccination with live attenuated mycobacterial strain
• Infection with non-tuberculosis mycobacteria

Normal and Critical Findings

PPD Skin Test Interpretation Based on CDC Guidelines

The result of the PPD test is either positive or negative. However, the size of the induration diameter cutoff (5, 10, or 15 mm) for the test to be considered positive is based on certain risk factors. As the diameter cutoff increases, the sensitivity of this test declines, and the specificity increases. For instance, the sensitivity of this test is highest (98%) for a positivity cutoff of 5 mm of induration.[2]

Induration of less than 5 mm is considered positive in the following cases:

• Individuals infected with HIV who have had close contact with an active contagious case

Induration of 5 mm or greater is considered positive in the following cases:

• Immunosuppressed individuals, including those receiving long-term steroids (the equivalent of prednisone ≥15 mg/day for ≥1 month), chemotherapy, or tumor necrosis factor-alpha inhibitors
• Individuals infected with HIV
• Close contact with patients with active TB 
• Prior TB signs on chest radiography, such as fibronodular changes
• Organ transplant recipients

Induration of 10 mm or more is considered positive in the following cases:

• Emigrants from countries where TB is considered endemic, or the incidence of infection is greater than 25/100,000
• Employees and residents in high-risk settings, such as prisons, mycobacteriology labs, healthcare settings, and homeless shelters
• Injection drug users
• Mycobacteriology laboratory professionals
• Children less than four years of age
• Those with chronic medical conditions that increase the risk of reactivation TB, including diabetes mellitus, chronic renal failure on dialysis, silicosis, malignancy (leukemia, lymphoma, or cancer of the head, neck, or lung), a history of jejunoileal bypass
• Individuals weighing less than 90% of the ideal body weight
• Infants, children, and adolescents exposed to those in high-risk categories

Induration of 15 mm or more is considered positive in the following cases:

• Healthy individuals older than four years of age with a low likelihood of TB infection[14]

Positive PPD Skin Test 

If the risk of TB infection is very high, the PPD test need not be repeated. The greater the risk due to risk factors, the higher the positive predictive value of the PPD TST. A TB symptom assessment, physical examination, and chest radiograph usually follow the positive PPD test. 

If there are no symptoms of TB and no evidence of active tuberculosis infection on the physical examination and chest radiograph, the patient most likely has latent TB (LTBI), and treatment should be initiated. However, if the clinical evaluation or chest radiograph is positive, additional tests, including a sputum test, urine test, or tissue biopsy, may be performed to confirm the diagnosis of active disease. 

Negative PPD Skin Test

If the patient is at a high risk of developing an active TB infection, a repeat test is recommended if the initial test is negative. Repeat testing should also be performed if the initial test was done within eight weeks of known exposure. In very high-risk patients, the repeat test may also be negative secondary to a lack of immune response. In this situation, treatment may be considered. 

PPD Skin Test Conversion 

According to the CDC, a repeat PPD TST is considered positive if induration is greater than ten mm and has increased by at least six mm compared with the previous test. This test interpretation is less sensitive but more specific.

PPD Skin Test Reversion

The conversion of a previously positive test to a negative one is called Mantoux test reversion and is seen in fewer than 10% of individuals. This phenomenon may be seen in those with waning natural immunity, such as in older adults, where the previous positive PPD result was the result of a boosted response on two-step testing.

Interfering Factors

False-positive Reaction

Falsely positive test results occur when the test result is positive in the absence of M. tuberculosis infection. Since the PPD test has low specificity, low-risk individuals with a positive test may be false positives. A false-positive reaction may be seen in the following cases:[2]

• Previous vaccination with Bacillus Calmette-Guerin (BCG)
• Infections with non-tuberculous mycobacteria
• Improper administration of the test
• Incorrect reading or misinterpretation of the test

The IGRA test may be considered in individuals with prior BCG vaccination since childhood BCG immunization will not affect IGRA results. 

False-negative Reaction

Falsely negative test results occur when there is an inadequate or absent response to tuberculin protein in the presence of M. tuberculosis infection. A false-negative reaction may be seen in the following cases:[2]

• Inadequate T-cell response or cutaneous anergy secondary to immunosuppression or waning natural immunity
• Immunosuppressive medications
• Comorbid conditions (chronic renal disease, recent surgery, severe burns, severe malnutrition, lymphoma, sarcoidosis)
• M. tuberculosis infection within eight weeks of testing
• Old mycobacteria infection, i.e., many years prior, may not be detected 
• Children less than six months old
• Older adults
• Recent viral, bacterial, or fungal infection
• HIV infection 
• Recent live-virus vaccination (measles, mumps, polio) within 4 to 6 weeks of the test
• Improper administration of the test
• Incorrect reading or misinterpretation of the test
• Defective tuberculin material

Two-step Testing and Boosted Reaction

Two-step testing involves administering a subsequent test after an initial false-negative test. The second test should be administered between one and four weeks after the first test.[5]

In some individuals, the reaction to tuberculin antigen wanes over time, which results in a false-negative reaction. In individuals with a prior TB infection from years earlier, sensitization to tuberculin may be weak, and there may be a false-negative PPD test. However, if a subsequent test is administered, the tuberculin PPD may stimulate the immune system. A boosted reaction is seen on subsequent testing due to "recalling" of the immune response.

Complications

The PPD TST may cause the following side effects with unknown frequency.

A severe hypersensitivity reaction characterized by urticaria, dyspnea, or angioedema may occur within 15 minutes of tuberculin administration or six to twelve hours after testing. Local side effects may occur at the injection site, including pain, discomfort, hematoma, scarring, pruritus, necrosis, ulceration, blistering, or swelling. In addition, fever, presyncope, or syncope may occur.

Patient Safety and Education

Any person with a prior severe reaction to the PPD TST should not be tested again using this method; severe reactions include necrosis, blistering, ulcerations, or anaphylactic shock.

PPD administration is not contraindicated in the following groups: those having received prior BCG vaccination, pregnant patients, those with HIV infection, infants, and those having previously received a PPD test.

Clinical Significance

The PPD test is simple, inexpensive, and rapidly identifies TB infection. In cases of latent TB, treatment should be initiated to prevent progression to active disease. Shorter treatment regimens are more likely to be completed than the six to nine months isoniazid regimen. Shorter duration regimens are recommended by the CDC and NTCA and include three months of weekly isoniazid and rifapentine, four months of daily rifampin, or three months of daily isoniazid and rifampin.

The detection and treatment of LTBI decrease the burden of active TB.[15][16]

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Disclosure: Parul Pahal declares no relevant financial relationships with ineligible companies.

Disclosure: Elizabeth Pollard declares no relevant financial relationships with ineligible companies.

Disclosure: Sandeep Sharma declares no relevant financial relationships with ineligible companies.