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Mantoux Test – Procedure, Normal Range and Result Interpretation

What is Mantoux Test?

  • The Mantoux test, often referred to as a skin test, is a method used to determine if someone has been exposed to Mycobacterium tuberculosis, the bacteria responsible for tuberculosis (TB). This exposure could be due to an active TB infection or a past encounter with the bacteria. The test is particularly useful for monitoring the effectiveness of the BCG vaccination, which is administered to protect against TB.
  • This test is commonly conducted among certain groups of people. These include children, healthcare workers, and individuals who either have a high risk of TB exposure or show symptoms suggesting a TB infection.
  • It’s important to note that the Mantoux test doesn’t differentiate between an active TB infection and a latent (inactive) one. Also, a positive result from this test alone isn’t enough to confirm an active TB infection. In cases where the Mantoux test shows a positive reaction, additional tests are necessary. These may include a sputum smear test, culture tests, and chest X-rays, all of which help in accurately diagnosing an active TB infection.

Principle of Mantoux Test

The Mantoux test operates on the principle of delayed-type hypersensitivity, specifically Type IV hypersensitivity. This test is designed to assess a person’s cell-mediated immunity against the bacterium Mycobacterium tuberculosis. To conduct this test, a substance known as Purified Protein Derivative (PPD) is used, which is essentially a mycobacterial antigen.

During the test, a small amount of PPD, typically 5 units in 0.1 ml, is injected intradermally using a needle of 26, 27, or 30 gauge. The test area is then observed for a reaction over the following 48 to 72 hours. What the medical professionals look for is an area of induration, which means a raised, hardened area at the injection site. It’s important to understand that redness, or erythema, around the area is not considered a significant indicator in this test.

The reaction that occurs at the site of the injection is due to the body’s immune response. When PPD is injected, it triggers memory T-helper 1 (Th1) cells to release cytokines. These cytokines attract macrophages and granulocytes to the area, which is what causes the induration and, to a lesser extent, erythema. This hypersensitivity reaction starts about 5-6 hours after the injection and typically reaches its peak between 48 and 74 hours.

Requirements for Mantoux test

For conducting the Mantoux test, a few specific items are essential:

  1. Graduated 1ml Syringe: This is a crucial tool for administering the test. The syringe, marked with accurate measurements, ensures that the correct dose of the test substance is injected.
  2. PPD or Tuberculin: PPD stands for Purified Protein Derivative, which is the substance used in the test. It is derived from the bacteria that cause tuberculosis and is used to check the body’s reaction to these bacteria.
  3. Spirit Swab: Before the injection, the skin where the PPD will be injected needs to be cleaned to prevent infection. A spirit swab, usually soaked in alcohol, is used for this purpose. It sterilizes the area, ensuring that the test is conducted in a safe and hygienic manner.

Procedure of Mantoux test

  1. Prepare the PPD Reagent: Before starting, make sure the PPD (Purified Protein Derivative) reagent is at room temperature. This is important for the accuracy of the test.
  2. Choose the Injection Site: The test is usually performed on the dorsal (back) surface of the forearm, approximately 4cm below the elbow. It’s important to select an area of skin that is clear of any obstructions like scars or sores, as these can affect the results.
  3. Clean the Injection Site: Use a spirit swab to disinfect the area where you’ll inject the PPD. It’s crucial to let the skin dry after disinfecting to prevent any irritation or infection.
  4. Prepare the Syringe: Draw a little more than 0.1 ml of PPD into a 1 ml syringe. Then, adjust the amount to exactly 0.1 ml and ensure there are no air bubbles in the syringe, as these can interfere with the test.
  5. Inject the PPD: Use a 27 gauge needle to inject the PPD into the skin intradermally (just under the skin). The goal is to create a small raised area on the skin, often called a ‘bleb’ or ‘wheal’, about 6 to 8 mm in diameter.
  6. Mark the Injection Site: After the injection, mark the area so it’s easy to find when reading the results.
  7. Read the Results: The results of the Mantoux test are read 48 to 72 hours after the injection. The key thing to look for is induration (a raised, hardened area) at the injection site. The size of the induration is what determines if the test is positive or negative.

Results and Interpretation

  1. Measurement of Reaction: After 48-72 hours of administering the PPD, the area of induration (the raised, hardened area) is measured in millimeters. It’s important to note that redness, or erythema, around the area is not considered significant for this test and is therefore not measured.
  2. Interpreting the Results: The Centers for Disease Control (CDC) provides guidelines on how to interpret these measurements, taking into account the person’s risk of TB infection and progression to disease. There are three main thresholds for a positive result, based on the size of the induration and the individual’s risk factors:
    • Induration of ≥5 mm: This is considered positive in high-risk groups such as:
      • People infected with HIV.
      • Those who have recently been in close contact with TB patients.
      • Individuals with chest X-ray results showing changes consistent with past TB.
      • Patients who have received organ transplants or are otherwise immunosuppressed.
    • Induration of ≥10 mm: This is considered positive in groups like:
      • Recent immigrants (within the last 5 years) from areas with high TB prevalence.
      • Injection drug users.
      • Residents and employees in high-risk environments like prisons, nursing homes, healthcare facilities, residential facilities for AIDS patients, and homeless shelters.
      • Mycobacteriology laboratory personnel.
      • Individuals with certain health conditions such as silicosis, diabetes, chronic renal failure, certain blood disorders, and specific cancers.
      • Children and adolescents exposed to high-risk adults.
    • Induration of ≥15 mm: This threshold is for people who don’t have any known risk factors for TB.

Normal range of Mantoux test

  • Positive Reaction:
    • Typically, an induration of 15 millimeters or more is considered positive in individuals with no known risk factors for TB.
    • In individuals with certain risk factors (e.g., close contact with active TB cases, immunosuppression), a smaller induration (e.g., 5 mm or more) may be considered positive.
  • Negative Reaction:
    • An induration of less than 5 millimeters is usually considered negative.
  • Indeterminate Reaction:
    • In some cases, the reaction may be neither clearly positive nor negative, and the result is considered indeterminate.

Classification of tuberculin reaction

The classification of tuberculin reactions is a critical aspect of interpreting Mantoux test results, and it involves considering specific increments of induration. The person’s medical risk factors determine the threshold at which the induration is considered positive. The following classification elucidates these thresholds and their associated medical risk factors:

  1. 5 mm or More: Positive Result in Specific Cases
    • Medical Risk Factors:
      • An HIV-positive person.
      • Persons with recent contacts with a TB patient.
      • Individuals with nodular or fibrotic changes on chest X-ray consistent with old healed TB.
      • Patients with organ transplants and other immunosuppressed individuals.
    • Function: This threshold signifies a positive result in individuals with heightened susceptibility to tuberculosis due to specific medical conditions or recent exposure.
  2. 10 mm or More: Positive Result in Various Groups
    • Medical Risk Factors:
      • Recent arrivals (less than five years) from high-prevalence countries.
      • Injection drug users.
      • Residents and employees of high-risk congregate settings (e.g., prisons, nursing homes, hospitals, homeless shelters, etc.).
      • Mycobacteriology lab personnel.
      • Persons with clinical conditions placing them at high risk (e.g., diabetes, prolonged corticosteroid therapy, leukemia, end-stage renal disease, chronic malabsorption syndromes, low body weight, etc.).
      • Children less than four years of age or children and adolescents exposed to adults in high-risk categories.
    • Function: This threshold indicates positive results in individuals with diverse risk factors, including recent arrivals, specific occupational exposures, and those with underlying health conditions.
  3. 15 mm or More: Positive Result in Individuals with No Known Risk Factors
    • Medical Risk Factors:
      • Persons with no known risk factors for TB.
    • Function: This threshold denotes a positive result in individuals without identified risk factors, suggesting exposure to Mycobacterium tuberculosis.
  4. Tuberculin Test Conversion: Increase of 10 mm or More Within a Two-Year Period
    • Alternative Criteria:
      • Increases of 6, 12, 15, or 18 mm are also considered.
    • Function: Tuberculin test conversion is defined as a substantial increase in induration within a specified timeframe, irrespective of age. Alternative criteria provide flexibility in assessing changes over time.

Culture media for Mycobacterium Tuberculosis

Mycobacterium tuberculosis, the causative agent of tuberculosis, requires specialized culture media for optimal growth and isolation. The commonly employed medium for cultivating these bacteria is the Lowenstein-Jensen media. The composition and characteristics of this media contribute to the selective growth of Mycobacterium tuberculosis while inhibiting the proliferation of contaminants.

Composition of Lowenstein-Jensen Media:

  1. Whole Eggs: The media incorporates whole eggs, providing a rich source of nutrients essential for the growth of Mycobacterium tuberculosis. The eggs contribute proteins, lipids, and other vital components required for bacterial metabolism.
  2. Asparagine: Asparagine, an amino acid, serves as a nitrogen source in the media. Nitrogen is a critical element for the synthesis of proteins and nucleic acids, supporting the bacterial growth process.
  3. Glycerol: Glycerol is included in the media formulation, serving as a carbon source. Bacteria utilize glycerol as an energy and carbon substrate, facilitating metabolic processes.
  4. Malachite Green: Malachite green is an important component with inhibitory properties. It helps suppress the growth of contaminants, ensuring the specificity of the culture for Mycobacterium tuberculosis.

Cultivation Process:

  1. Incubation Temperature: The culture process involves incubating the specimen at a temperature of 37 °C. This temperature is conducive to the optimal growth of Mycobacterium tuberculosis.
  2. Duration of Incubation: The incubation period typically spans 2 to 3 weeks, but the cultures are maintained for an extended duration, ranging from 6 to 8 weeks. This extended period allows for the detection of slow-growing mycobacterial species.
  3. Microscopic Identification: Modern diagnostic techniques involve the identification of Mycobacterium tuberculosis using fluorescent microscopy. This method enhances the speed and accuracy of detection.

Classification Based on Growth Rate:

  1. Rapid Growers: This group comprises mycobacterial species that form colonies within a short period, usually 2 to 3 days. Their swift growth distinguishes them from slow growers.
  2. Slow Growers: Mycobacteria classified as slow growers exhibit a more extended growth period, typically taking 2 to 3 weeks to form colonies.

Medically Important Mycobacteria:

  1. M. tuberculosis: The primary causative agent of tuberculosis, characterized by a slowly progressive chronic infection, primarily affecting the lungs.
  2. M. bovis: Another medically significant species causing tuberculosis, with the capacity to infect various organs.
  3. M. leprae: Responsible for leprosy, a chronic infectious disease affecting the skin and peripheral nerves.
  4. Atypical Mycobacteria: This group includes mycobacterial species other than M. tuberculosis, M. bovis, and M. leprae. They can cause opportunistic infections in individuals with compromised immune systems.

False positive result

A false positive result in the Mantoux test means that the test indicates a person has been exposed to the tuberculosis bacteria when, in reality, they haven’t. This can happen for several reasons, especially in individuals who are at low risk for tuberculosis:

  • Infection with Non-Tuberculous Mycobacteria (NTM): There are types of mycobacteria other than the one that causes tuberculosis. Infection with these NTM can lead to a positive reaction in the Mantoux test, even though the person does not have tuberculosis.
  • BCG Vaccination: The BCG vaccine is used to protect against tuberculosis, but it can cause a false positive result in the Mantoux test. This is because the vaccine contains a strain of mycobacteria similar to the one that causes tuberculosis.
  • Incorrect Antigen Administration: If the wrong antigen is used during the test, or if it is administered incorrectly, this can lead to a misleading positive result.
  • Incorrect Interpretation of Results: Sometimes, the way the results are read can lead to a false positive. It’s important that the test is interpreted correctly, taking into account factors like the size of the induration and the individual’s risk factors for TB.

False negative result

In some cases, individuals may have a negative response to the Tuberculin Skin Test (TST), even if they are infected with M. tuberculosis. This is known as a false-negative result. There are several factors that can contribute to this:

  • Concurrent Infections: Certain viral infections like measles, mumps, chickenpox, and HIV, bacterial infections such as typhoid fever, brucellosis, typhus, leprosy, and pertussis, or fungal infections can interfere with the TST, leading to a false-negative result.
  • Chronic Renal Failure: This condition can impact the immune response, potentially leading to a false-negative TST.
  • Low Protein States: Severe protein depletion or conditions like afibrinogenemia can affect the body’s ability to mount an immune response to the test.
  • Diseases Affecting Lymphoid Organs: Diseases such as Hodgkin’s disease, lymphoma, chronic leukemia, and sarcoidosis, which impact lymphoid organs, can alter the immune response, leading to false-negative results.
  • Immunosuppressive Drugs: Medications like medical steroids, which suppress the immune system, can result in a lack of response to the TST.
  • Age-Related Immune Response: Very young children (under 6 months) and elderly patients may have an immature or weakened immune system, respectively, affecting the test results.
  • Stress Factors: Various stressors, including surgery, burns, mental illness, and graft-versus-host reactions, can influence the immune system’s response.
  • Improper Handling: Incorrect storage or handling of the antigen, or inaccuracies in measuring or interpreting the results, can lead to a false-negative.
  • Recent Vaccinations: Live virus vaccinations can interfere with the TST response.
  • Recent TB Infection: In cases of a very recent TB infection, the body may not have had enough time to develop a response detectable by the TST.

Uses of Mantoux Test

  • Screening for Tuberculosis:
    • The Mantoux test is widely employed as a screening tool to identify individuals who may have been exposed to Mycobacterium tuberculosis.
    • It is particularly valuable in populations with a higher risk of TB, such as healthcare workers, immigrants from high-prevalence regions, and individuals in close contact with TB patients.
  • Tuberculosis Diagnosis:
    • The Mantoux test is an essential component in the diagnosis of tuberculosis.
    • A positive test indicates that an individual has mounted an immune response to Mycobacterium tuberculosis, suggesting current or past infection.
  • Identification of Latent Tuberculosis Infection (LTBI):
    • The test is crucial for detecting latent tuberculosis infection, where individuals harbor the bacteria but do not exhibit active disease symptoms.
    • LTBI individuals have the potential to develop active TB in the future, and identifying them allows for preventive measures.
  • Epidemiological Studies:
    • The Mantoux test is utilized in epidemiological studies to assess the prevalence of tuberculous infection within specific populations.
    • This data aids public health authorities in understanding the extent of TB transmission and implementing targeted interventions.
  • Contact Tracing:
    • In situations where an individual has been in close contact with someone diagnosed with active tuberculosis, the Mantoux test is used for contact tracing.
    • It helps identify individuals who may have been exposed to the bacteria and guides further diagnostic and preventive measures.
  • Assessment of TB Control Programs:
    • Public health agencies use the Mantoux test results to evaluate the effectiveness of TB control programs.
    • Monitoring the prevalence of tuberculous infection helps assess the impact of interventions and guide future public health strategies.
  • Pre-employment and Occupational Health Screening:
    • Some industries, especially healthcare, may require pre-employment or periodic Mantoux testing to ensure a safe working environment.
    • This is particularly relevant for occupations where individuals may be at an increased risk of TB exposure.
  • Evaluation of BCG Vaccination Status:
    • The Mantoux test is used to assess the response to Bacillus Calmette–Guérin (BCG) vaccination, a vaccine administered in some countries to protect against severe forms of TB, such as tuberculous meningitis.
    • A positive result in an individual who has received the BCG vaccine is interpreted as latent TB infection rather than vaccine effectiveness.
  • Monitoring Treatment Response:
    • In cases where TB treatment is initiated, the Mantoux test may be used to monitor the effectiveness of treatment.
    • A decrease in the size of induration over time may indicate a positive response to therapy.
  • Research and Surveillance:
    • The Mantoux test is utilized in research settings to study the immune response to Mycobacterium tuberculosis and to gather data for surveillance purposes.
    • Research studies may explore variations in test performance, the impact of new diagnostics, and trends in TB prevalence.

Advantages of Mantoux Test

  • Sensitivity:
    • The Mantoux test is highly sensitive in detecting individuals who have been exposed to Mycobacterium tuberculosis.
    • It can identify both active tuberculosis cases and individuals with latent tuberculosis infection (LTBI), providing a comprehensive view of TB prevalence.
  • Early Detection of Infection:
    • The test can detect early stages of infection, even before symptoms of active tuberculosis manifest.
    • Early detection allows for timely intervention and preventive measures to reduce the risk of progression to active disease.
  • Cost-Effective:
    • The Mantoux test is relatively inexpensive compared to some other diagnostic methods for tuberculosis.
    • Its cost-effectiveness makes it suitable for large-scale screening programs and resource-limited settings.
  • Ease of Administration:
    • The test is straightforward to administer, requiring only a small injection of tuberculin under the skin.
    • It can be performed by trained healthcare professionals, and the injection is minimally invasive.
  • **Readily Available Tuberculin:
    • Purified protein derivative (PPD), the tuberculin used in the Mantoux test, is commercially available and easily accessible.
    • This availability contributes to the widespread implementation of the test in various healthcare settings.
  • Wide Applicability:
    • The Mantoux test is applicable across diverse populations, including adults, children, and individuals with varying health statuses.
    • Its versatility makes it valuable for screening in different demographic groups.
  • Standardized Interpretation Criteria:
    • Clear and standardized criteria exist for interpreting the test results, helping to minimize subjectivity in result interpretation.
    • This standardization enhances the reproducibility of the test across different healthcare providers.
  • Useful in Contact Tracing:
    • The Mantoux test is instrumental in contact tracing scenarios, where individuals exposed to active tuberculosis cases need to be identified.
    • It aids in assessing the extent of potential transmission within a specific community or setting.
  • Integration into Public Health Programs:
    • The test has been integrated into many national and international public health programs focused on tuberculosis control.
    • Its inclusion in these programs facilitates systematic screening and monitoring efforts.
  • Historical Data and Comparisons:
    • The Mantoux test has a long history of use, and historical data using this test are available for many populations.
    • Longitudinal data and comparisons over time contribute to a better understanding of trends in TB prevalence.
  • Research Utility:
    • The Mantoux test is commonly used in research studies to assess the immune response to Mycobacterium tuberculosis and evaluate new diagnostic tools.
    • Its use in research contributes to the development of improved strategies for tuberculosis control.

Limitations of Mantoux Test

  • Non-Specificity:
    • The Mantoux test lacks specificity, meaning it can yield positive results in individuals who have been vaccinated with Bacillus Calmette–Guérin (BCG) or those exposed to non-tuberculous mycobacteria.
    • BCG vaccination, which is common in many countries, can cause a false-positive reaction and complicate result interpretation.
  • False Positives:
    • The test may produce false-positive results in individuals with a history of BCG vaccination, even if they have not been exposed to Mycobacterium tuberculosis.
    • Cross-reactivity with environmental mycobacteria can also lead to false positives.
  • False Negatives:
    • False-negative results may occur in individuals with compromised immune systems, such as those with HIV or other immunosuppressive conditions.
    • Recent TB infection (within 8-10 weeks) may not induce a detectable immune response, leading to false negatives.
  • Variability in Immune Response:
    • Individual variations in immune responses can affect the test results.
    • Some individuals may not mount a significant immune response, leading to false negatives, while others may exhibit exaggerated responses, potentially resulting in false positives.
  • Interfering Conditions:
    • Certain medical conditions, such as sarcoidosis, infectious mononucleosis, and live virus vaccinations, can suppress the reaction to the tuberculin, leading to false negatives.
    • Corticosteroid therapy or other immunosuppressive medications can also influence test results.
  • Subjectivity in Measurement:
    • The test result interpretation involves measuring the induration size, and this process introduces a degree of subjectivity.
    • The precision of the measurement depends on the healthcare provider’s skill and consistency.
  • Boosted Reaction:
    • Some individuals who have been previously exposed to tuberculin may experience a “boosted” reaction upon subsequent testing, leading to an apparent conversion.
    • This phenomenon may not necessarily indicate recent infection but rather a recall of the immune response from the initial exposure.
  • Inappropriate Timing:
    • The timing of the test reading is critical, and deviations from the recommended 48-72 hours may affect result accuracy.
    • Reading the test too early or too late may lead to misinterpretation.
  • Inadequate Sensitivity in Children:
    • In very young children, the Mantoux test may have reduced sensitivity, potentially resulting in false-negative results.
    • Children under the age of 5 may not reliably produce a measurable reaction.
  • Variability in Tuberculin Products:
    • The source and composition of tuberculin products may vary, leading to differences in test performance.
    • Variability in the quality and potency of tuberculin products may impact the reliability of the test.
  • Not Suitable for Recent Infections:
    • The Mantoux test may not be suitable for detecting very recent TB infections, as it takes time for an immune response to develop.

Important Notes on Mantoux test

  • Preference in Children Under 5 Years:
    • The skin test is the preferred method for tuberculosis screening in children under the age of 5.
    • This preference is likely due to the practicality and reliability of the Mantoux test in this age group.
  • Basis of the Tuberculin Skin Test:
    • The test relies on the delayed-type hypersensitivity skin reaction produced by prior infection with the M. tuberculosis bacterium.
    • Specialized immune cells, known as T cells, sensitized by previous infection, play a pivotal role in initiating the immune response.
  • Mechanism of Skin Reaction:
    • T cells, attracted to the skin site, release lymphokines, chemical messengers that induce induration through local vasodilation.
    • This process leads to fluid deposition, edema, fibrin deposition, and the attraction of various inflammatory cells to the site.
  • Incubation Period for Positivity:
    • An incubation period of two to 12 weeks is typically required after exposure to M. tuberculosis for the Mantoux test to yield a positive result.
    • This emphasizes the importance of considering the exposure timeline for accurate interpretation.
  • Contraindications and Safety:
    • The Mantoux test is generally safe for infants, pregnant women, and HIV-infected individuals.
    • Contraindications exist only for those who have had a severe reaction to a previous tuberculin skin test.
  • Repeat Testing and Site Selection:
    • If the initial test is improperly administered, a repeat test can be given immediately, selecting a site several centimeters away from the original injection site.
    • This ensures accurate results and minimizes the impact of potential administration errors.
  • False-Negative Results:
    • A negative test does not necessarily imply freedom from tuberculosis.
    • Immunocompromised individuals, those undergoing cancer chemotherapy, or those with AIDS may exhibit false-negative results.
  • BCG Vaccination and False-Positive Results:
    • Individuals who received the BCG vaccine may show a positive skin reaction, leading to a false-positive result.
    • Persistence of this positive reaction is more likely in individuals vaccinated after the first year of life or those receiving multiple doses.
  • Other Mycobacteria and False Positives:
    • Infections with mycobacteria other than M. tuberculosis may lead to false-positive TB skin test results.
    • This highlights the importance of considering potential sources of mycobacterial exposure.
  • Safety and Lack of Side Effects:
    • The Mantoux test typically does not produce side effects.
    • Allergic reactions are rare, and since the test does not involve live bacteria, there is no risk of developing tuberculosis from the test.

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