TB tests for diagnosis and drug resistance
There are several tests available to help with the diagnosis of TB. There are also TB tests to find out whether the TB bacteria that someone is infected with are susceptible to drug treatment or are drug resistant. TB tests to find out if someone has drug resistant TB, are known as drug susceptibility tests.
TB diagnostic tests
There are TB tests which can be used to determine is someone has latent TB, which means that they are infected with TB bacteria. There are also TB tests, which when considered alongside other factors, such as whether someone has symptoms of TB, can confirm a diagnosis of active TB or TB disease.
Evidence of TB bacteria
Mycobacterium tuberculosis, the bacteria that cause TB © NIAID
The development of TB disease is a two stage process. In the first stage, known as latent TB, a person is infected with TB bacteria. In the second stage, known as active TB or TB disease, the bacteria have reproduced sufficiently to usually cause the person to have become sick.
A diagnosis of active TB can only be confirmed when there is definite evidence of TB bacteria in the person's body. Some of the diagnostic tools look directly for TB bacteria. Others such as the chest X-ray look for the effect of the bacteria on the person suspected of having TB.
Current TB tests - some problems
Some of the current TB tests take a long time to obtain a result, and some are not very accurate. The tests either have low sensitivity (the ability to correctly detect people with TB) and/or low specificity (the ability to correctly detect people who haven't got TB).
If a test has low sensitivity, it means that there will be a significant number of "false negatives", meaning that the test result is suggesting that a person has not got TB when they actually have. Similarly, a low specificity means that there will be a significant number of "false positives" suggesting that a person has TB when they actually haven't.
Acute pulmonary TB can be easily seen on an X-ray. However, the picture it presents is not specific and a normal chest X-ray cannot exclude extra pulmonary TB. Also, in countries where resources are more limited, there is often a lack of X-ray facilities.
The TB skin test
The TB skin test is a widely used diagnostic tool, and in countries with low rates of TB it is often used to test for latent TB infection. The problem with using it in countries with high rates of TB infection is that the majority of people may have latent TB.
The TB skin test involves injecting a small amount of fluid (called tuberculin) into the skin in the lower part of the arm. If the skin reacts by swelling after 48 to 72 hours, the person is probably infected with Tuberculosis bacteria. The Mantoux test is the type of test most often used, although the Heaf and Tine tests are still used in some countries. None of these tests though will guarantee a correct result.
False positive results happen with the TB skin test because the person has been infected with a different type of bacteria, rather than the one that causes TB. It can also happen because the person has been vaccinated with the BCG vaccine, and this vaccine is widely used in countries with high rates of TB infection. False negative results particularly happen with children, older people and people with HIV.
TB Interferon gamma release assays (IGRAs)
The Interferon Gamma Release Assays (IGRAs), are a new type of more accurate TB test. In this context referring to an assay is simply a way of referring to a test or procedure.
T-SPOT®.TB Test © Oxford Immunotec
IGRAs are blood tests that measure a person's immune response to the bacteria that cause TB. The immune system mounts a complex response to TB bacteria, and produces some special molecules called cytokines. These assays work by detecting a cytokine called the interferon gamma cytokine. They are performed in practice by taking a blood sample and mixing it with special substances to identify if the cytokine is present.
Two IGRAs that have been approved by the U.S. Food and Drug Administration (FDA), and are commercially available in the U.S., are the QuantiFERON®-TB Gold test, and the T-SPOT®.TB TB test.
The advantages of an IGRA test includes the fact that it only requires a single patient visit to conduct the TB test, results can be available within 24 hours, and prior BCG vaccination does not cause a false positive result. Disadvantages include the fact that the blood sample must be processed fairly quickly, laboratory facilities are required, and the test is for latent TB. It is also thought that the IGRAs may not be as accurate in people who have HIV.1 In low prevalence resource rich settings, IGRAs are beginning to be used in place of the TB skin test.2
Serological tests for TB
Serological tests for TB are tests carried out on samples of blood, and they claim to be able to diagnose TB by detecting antibodies in the blood. However, testing for TB by looking for antibodies in the blood is very difficult.
As a result serological tests, sometimes called serodiagnostic tests, for TB are inaccurate and unreliable, and the World Health Organisation has warned that these tests should not be used to try and diagnose active TB. Some countries have banned the use of serological or serodiagnostic tests for TB.
Serological tests for TB are very different from the IGRA tests described above.
Sputum smear microscopy
A sputum smear stained using fluorescent acid fast stain © CDC/R W Smithwick
Smear microscopy of sputum is often the first TB test to be used in countries with a high rate of TB infection. Sputum is a thick fluid that is produced in the lungs and the airways leading to the lungs, and a sample of sputum is usually collected by the person coughing.
For the diagnosis of TB several samples of sputum will normally be collected.3 Historically it has been recommended that three sputum specimens are collected on two consecutive days, but in 2007 the World Health Organisation (WHO) recommended that just two specimens could be examined from consecutive days. Now it has been suggested that two specimens can be collected on the same day without any loss of accuracy.45
To do the test a very thin layer of the sample is placed on a glass slide, and this is called a smear. A series of special stains are then applied to the sample, and the stained slide is examined under a microscope for signs of the TB bacteria.6
Sputum smear microscopy is inexpensive and simple, and people can be trained to do it relatively quickly and easily. In addition the results are available within hours. The sensitivity though is only about 50-60%.7 In countries with a high prevalence of both pulmonary TB and HIV infection, the detection rate can be even lower, as many people with TB and HIV co-infection have very low levels of TB bacteria in their sputum, and are therefore recorded as sputum negative.
The use of fluorescent microscopy is a way of making sputum tests more accurate. With a fluorescent microscope the smear is illuminated with a quartz halogen or high pressure mercury vapour lamp, allowing a much larger area of the smear to be seen and resulting in more rapid examination of the specimen.
One disadvantage though is that a mercury vapour lamp is expensive and lasts a very short time. Such lamps also take a while to warm up, they burn significant amounts of electricity, and electricity supply problems can significantly shorten their life span. One way of overcoming these problems is the use of light emitting diodes (LEDs). These switch on extremely quickly, have an extremely long life, and they don't explode.8
In 2011 the World Health Organisation issued a policy statement recommending that conventional fluorescence microscopy should be replaced by LED microscopy. It also recommended that in a phased way, that LED microscopy should replace conventional Ziehl-Neelsen light microscopy.9
Colonies of Mycobacterium tuberculosis growth on a culture plate © CDC/Dr George Kubica
Culturing is a method of studying bacteria by growing them on media containing nutrients. Media can be either solid media on culture plates or bottles of liquid media (culture broths). Different media are used to make it as easy as possible for the suspected microorganisms to grow.
To isolate a single bacterial species from a mixture of different bacteria, solid media are normally used. Individual cells dividing on the surface do not move away from each other as they would do in liquid, and after many replications they form visible colonies composed of tens of millions of cells all derived from a single cell.
Culturing and identification of M. tuberculosis provides a definitive diagnosis of TB and can significantly increase the number of cases found. Culture can also provide drug susceptibility testing, showing which TB drugs a person's bacteria is resistant to. I.e. Has the person got MDR or XDR TB. However, culture is much more complex and expensive than microscopy to perform as it requires specific equipment and enhanced laboratory facilities.
Diagnosing TB using culture can also take weeks because of the slow growth of TB bacilli.10 It averages 4 weeks to get a conclusive result using the most common methods of solid media, with another 4-6 weeks to produce drug susceptibility results. Efforts have been made to reduce this time lag, the most common approach being to use improved liquid culture paired with solid media.
Clinical and sub clinical active TB
If a person has clinical active TB it usually means that they have symptoms or a chest x-ray which shows that they have TB, in addition to usually having a positive sputum test or culture. Sub clinical active TB means that although the person has had a positive sputum test or culture result, they have no symptoms and a chest x-ray does not show that they have TB.11
It is mostly just a very few people who have both HIV and TB who have sub clinical active TB. The great majority of people with active TB have symptoms.
TB drug susceptibility tests
Drug susceptibility testing means testing to find out which drugs the TB bacteria in a patient are susceptible to, and can therefore determine whether the person has got drug resistant TB. Some drug susceptibility tests, such as the Xpert TB test can be used to diagnose TB, as well as testing for some types of drug resistance.
TB tests summary
Diagnosing TB, and in particular diagnosing drug resistant TB, promptly and accurately, remains a significant challenge, particularly in resource poor settings. Although new TB tests are becoming available, they are generally too expensive for developing countries and also require significant laboratory facilities, including the availability of highly trained staff. This results in delays in providing patients with the appropriate drug treatment, and contributes to the on going global TB epidemic. It has been said regarding TB in India, and this applies to many other countries as well:
“What is sorely needed is a simple, cheap, point of care diagnostic test, and an .. economic, molecular test for drug resistance.”
- "Guidelines for intensified case finding and isoniazid preventative therapy for people living with HIV in resource constrained settings", Geneva, WHO, 2011 9 http:/www.who.int/tb/publications/2011/
- "Fact Sheets Interferon-Gamma Release Assays -Blood Tests for TB Infection", CDC www.cdc.gov/tb/publicationsfactsheets/
- "Sputum Culture", WebMD www.webmd.com/lung/sputum-culture
- Davis, J Lucian "Diagnostic accuracy of same-day microscopy versus standard microscopy for pulmonary tuberculosis: a systematic review and meta-analysis", The Lancet Infectious Diseases 23rd October 2012 www.thelancet.com/
- Kirwan, Daniela E "Same-day diagnosis and treatment of tuberculosis", The Lancet Infectious Diseases 23rd October 2012 www.thelancet.com/
- "Sputum Gram stain - Overview", University of Maryland Medical Center www.umm.edu/ency/article/
- Siddiqi, Kamran "Clinical diagnosis of smear-negative pulmonary tuberculosis in low-income countries: the current evidence", The Lancet Infectious Diseases, Vol 3, May 2003, 288 www.thelancet.com/journals/
- "TB diagnosis: Improving the yield with fluorescence microscopy", 2007 www.aidsmap.com/TB-diagnosis-Improving-the-yield-with-fluorescence-microscopy/
- "Fluorescent light-emitting diode (LED) microscopy for diagnosis of tuberculosis", WHO, 2011 www.who.int/tb/laboratory/policy_statements/en/index.html
- "New Laboratory Diagnostic Tools for Tuberculosis Control", Stop TB Partnership, 2009 http://apps.who.int/tdr/svc/publications/non-tdr-publications/
- Lillian M, "High Rates of Clinical and Subclinical Tuberculosis among HIV Infected Ambulatory Subjects in Tanzania", Clinical Infectious Diseases, 2005;40:1500-7 www.cid.oxfordjournals.org/content/40/10/1500.full.pdf
- Kuldeep Sing Sachdeva in Kelly Morris, "The new face of tuberculosis", The Lancet Infectious Diseases, Vol 11, October 2011, 736 www.thelancet.com/journals/
- Kuldeep Sing Sachdeva, "TB in India: burden, progress, and needs", TB diagnostics in India conference August 2011, tbevidence.org/2011/11/conference-on-tb-diagnostics-in-india-from-importation-and-imitation-to-innovation/