HIV Diagnostic Tests
April 1997
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HIV Diagnostic Tests April 1997 View PDF
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Laboratory tests can give important clues about the health status of people with HIV. Some of these tests—specifically complete blood counts (CBC), chemistry screens, T-Cell counts and viral load tests—should be done shortly after someone finds out they are HIV positive to establish a "baseline" measure of immune status and viral activity. Establishing this baseline helps people and their health care providers monitor disease progression as well as the effects of treatments. Age, sex, stress, current therapies, active infections and other factors can affect the results of these tests, and test results should be interpreted with these other factors in mind. Understanding your laboratory test results can be a daunting task, but the tests can help you take charge of your health and understand why your physician may be prescribing particular tests and medications. Most common test reports make interpretation of results simpler by including a "normal" range (high and low values). Those results which fall outside or at the edges of these normal ranges are likely to be the most significant. You should keep track of your test results to look for overall trends. You may simply keep copies of your test results, but it may be easier to make a chart or table to help keep track of all your test results and easily note trends or changes. There are some basic guidelines to consider when interpreting laboratory results:
Common
Laboratory Tests Complete Blood
Count (CBC) Generally, even people without symptoms of HIV disease should have a CBC test performed every 6-12 months. People whose bloodwork trends are changing may want to have CBC tests performed every 3 months (or more frequently). People with symptoms of HIV disease should have a CBC every 3-6 months. Testing is done more frequently in people with symptoms of anemia (low red blood cell count), leukopenia (low white blood cells) and thrombocytopenia (low platelets). In all cases, if a change occurs that causes you or your physician any concern, the tests should be repeated a few weeks later to confirm the first result. Keep in mind that of the following tests, the most important markers are the red blood cell, white blood cell and platelet counts. Red Blood Cell (RBC) Count - Hemoglobin - Hematocrit - Mean Corpuscular Volume (MCV) - Mean Corpuscular Hemoglobin (MHC) and Platelet Count - White Blood Cell (WBC) Count - White Cell Differential - |
| Chemistry
Screen ("Chem 25" or SMA-25) A chemistry panel examines the levels of 25 chemicals in the blood and can help determine if the body is functioning properly. (Other versions of the chemistry test monitor 12, 14, or 20 chemicals and are called SMA12, SMA14, and SMA20 respectively.) A chemistry test should be done once a year in people not taking medications and more often in people who are. Some of the important values are: Cholesterol and Triglycerides - Amylase - Liver Function Tests - Kidney Function Tests - Glucose - Proteins - CD4+ cell counts were long considered the best predictor of disease stage and risk of developing AIDS-related complications. It is now generally agreed that CD4+ cell counts alone are inadequate for measuring disease progression and response to therapy because CD4+ counts are subject to variability. They tell us how many cells are present, but do not tell us about their function. The combined use of CD4+ counts and viral load testing will provide a more complete picture of an individual’s status and response to therapy. However, CD4+ cell counts, not viral load, are a better indicator for when to start preventative therapy for opportunistic infections. Lymphocyte Subsets -
Because the absolute T cell counts can be variable, it is helpful to look at relative percentages of CD4+ and CD8+ cells to get a better picture of immune status. The CD4+ percentage is the percentage of CD4+ cells in the total number of lymphocytes; this may be a better indicator of disease progression than CD4+ cell counts alone because of the variability of absolute numbers. Normal CD4+% is 28-58%. The CD4+/CD8+ ratio is another helpful indicator. This ratio is normally about 2.0 (that is, for every 2 CD4+ cells there is 1 CD8+ cell). In HIV disease, this ratio is inverted; as the CD8+ cell level rises, the CD4+ cells decrease. Although T cell counts are useful as a rough guide for starting treatments or medications and monitoring immune function, they are subject to variation and cannot be an absolute indicator of health or illness. Tests can vary according to time of day, current infections, lack of sleep, stress and other biological factors. Variation in the laboratory used, as well as how quickly the test is performed after the blood is drawn, can also affect test results. Therefore, it is very important to look at overall trends and not be alarmed by any one individual test result. Viral
Load Tests What are
the tests?
The various tests are appropriate for different stages of HIV disease. Q-PCR is the most sensitive and can detect very low levels of virus in the blood, but the bDNA test has been shown to be the most accurate in quantifying high levels of virus. Every test has a certain error level, and some tests may have a degree of error as high as 20%. In addition, the tests require different amounts of blood—bDNA uses 2ml (several tablespoons) while NASBA and Q-PCR require only 100 and 200 microliters respectively (much less). This may be an issue for people receiving other tests that require large amounts of blood. New versions of these tests are being studied, and each "generation" of test generally provides greater sensitivity and accuracy of the HIV RNA level. Roche Molecular Systems is developing a new ultra-sensitive test that will be able to detect between 20-50 copies of HIV RNA, allowing us to know earlier whether a therapy is succeeding. Similarly, Chiron is developing second and third generation bDNA tests which will be able to detect 500 and 50 copies of HIV RNA respectively and Organon Teknika has a second generation test which will be able to detect 400 copies of HIV RNA. Viral load, disease progression, and antiretroviral therapyHigher HIV RNA levels correlate to lower CD4+ counts, more rapid declines in CD4+ cell counts and more rapid disease progression. HIV RNA levels seem to be a good predictor of long-term outcomes with regard to disease progression and death. People with HIV RNA levels greater than 100,000 have been shown to be ten times more likely to progress to AIDS over the following five years compared to those with levels less than 100,000. People with constant viral load levels less than 10,000 appeared to have a decreased risk of disease progression. Some exciting potential uses of viral load tests include using the tests for evaluating responses to anti-HIV therapy and for deciding the appropriate time to begin, change or add therapies. Viral load tests may quickly tell researchers, physicians and patients whether or not a drug is effective and when its effectiveness begins to wane. This information may make treatment decision-making a much more rational process than before. Effective antiviral therapy has been shown to significantly reduce HIV RNA levels in the blood within one week of the start of treatment. Whether it is starting, switching or adding an antiviral therapy, HIV RNA levels have been shown to drop in response to a new attack against the virus. No significant change in viral load levels indicates that the particular drug regimen is probably not working. With the use of these new tests, people can now make quicker decisions on the effect of their treatments. (Note: people with more advanced-stage disease (CD4+ cell counts less than 50) may take more time to see decreases in HIV RNA after initiating a new regimen.) Preliminary results from a recent study showed that viral load can predict the duration of viral suppression during protease inhibitor therapy. Duration of response to therapy was defined as the time from starting therapy until a 0.3 log increase in viral load was seen. Interestingly, duration of response to therapy was predicted only by the lowest viral load levels reached on that therapy. How long a treatment was likely to last was not predicted by starting viral load, initial CD4+ counts, magnitude of viral load drop or magnitude of CD4+ cell increase. Thus it is very important to get viral load below the limits of detection so that the therapy will last as long as possible. In addition, another study showed that the benefit of therapy in reducing risk of disease progression depends on the amount of reduction, regardless of baseline HIV RNA levels. For example, a one log reduction from 150,000 copies HIV RNA to 15,000 copies reduces the relative risk of disease progression the same amount as a one log reduction from 50,000 copies to 5,000 copies. Increasing HIV RNA levels may indicate the development of drug resistance. Researchers hope that changes in HIV RNA may give an earlier warning of impending drug failure, signals that can be read before a person suffers serious decline of CD4+ counts, disease progression or death. Recent studies suggest that a rapid return to pre-therapy HIV RNA levels (within 6 months of initiation of therapy) may be associated with a higher risk of disease progression and potential drug failure. These factors taken together may provide a better way to know when to start or change therapies than with the earlier method of using only CD4+ levels. Viral load
in plasma and other HIV reservoirs
Results of HIV RNA tests can be difficult to understand. Many physicians have been using HIV RNA tests since the first of these tests received FDA approval in June of 1996. However, clear guidelines are needed to assist people in understanding when to initiate or alter antiviral therapy. A few general guidelines are provided here, but they are likely to change as more research is done and new tests become available. As with CD4+ counts, what is most important is the trend of HIV RNA levels, not just any individual test result. Both the size and the duration of the drop are likely to be important in evaluating the success of a given therapy. People with undetectable viral load should remember that "undetectable" does not mean that the virus is gone, but that it is simply at such low levels that it is below the sensitivity level of the test. Viral load levels in the plasma may be undetectable, but as of today we are not certain that virus is not present in other parts of the body which are harder to access, such as the lymph tissue and the brain. However, recent data do indicate that a correlation may exist between viral levels seen in plasma and other parts of the body. These guidelines are being clarified and modified as physicians and researchers learn more about the predictive value of viral load tests.
Because the tests measure viral activity, anything that affects the production of virus can influence test results. In the normal progression of HIV disease, viral levels tend to rise slowly, and a sudden, sustained rise can indicate that the disease is likely to progress more rapidly. However, a sudden rise can also be caused by other active infections, such as the flu or an herpes outbreak, because previously HIV-infected, but inactive, CD4+ cells suddenly ‘wake up’ (are activated by the flu or herpes virus) and start producing HIV. Additionally, certain vaccines, such as the flu vaccine, can cause short-term increases in HIV RNA levels. This has caused some controversy around whether or not HIV-positive people should get routine flu shots. In general, the guidance is that the increase in viral levels resulting from vaccination are much lower than the rises caused by the actual flu infection, and these increased viral levels are not sustained for more than a few weeks. Viral levels can also be affected by antiviral therapy, making viral load tests an important tool in making treatment decisions. |
| How
often should viral load be measured? Initially two HIV RNA measurements should be taken approximately 2-4 weeks apart to establish a ‘baseline’ level. Afterwards, tests should be repeated every 3-4 four months along with CD4+ cell counts. Because other infections, such as a common flu, can cause a temporary increase in viral levels, any sudden rise in virus should be checked with another test 2-4 weeks later. People should generally avoid having viral load tests done during an active infection (e.g. active cold or flu), after routine immunizations (flu, tetanus, and pneumococcus) or during flare-ups of genital herpes or other opportunistic infections. These factors can all cause a 10 to 100 fold increase in viral levels that usually returns to baseline within a few weeks after the resolve of the infection or post vaccination. Tests should be performed more frequently when critical decisions are being made regarding treatment, and a test should be performed 3-4 weeks after starting or changing therapies. In addition, because the tests can vary slightly in their results, people using a particular test should continue to use that test to get reliable results and correct trends. Getting viral load testsBoth Roche and Chiron, manufacturers of Q-PCR and bDNA respectively, offer Patient Assistance Programs to supply their tests free-of-charge to people who have no other means to pay for them ($150-250 each). The availability of these programs may be limited geographically, but you can call the following numbers for more information:
Roche’s Q-PCR is FDA-approved, and most insurance providers and Medicaid will pay for tests once they are approved by the FDA. The bDNA test is still awaiting approval by the FDA. Check with your personal provider for specific coverage details. CommentaryOngoing research will help us understand the best use of HIV RNA tests in regular health monitoring. As more research is done, we will obtain clearer and stricter guidelines as to the use of viral load tests for monitoring responses to antiviral therapy and for predicting disease progression. The combination of viral load tests, CD4+ cell counts and other health markers help provide a clearer picture of the risk of disease progression, immune status and the body’s fight against HIV. The increasingly widespread use of viral load tests warrants that clearer guidelines be developed to assist people in understanding viral activity. Because viral load tests are only performed on blood plasma, the next stage of research needs to examine whether these tests are an accurate predictor of viral activity in other parts of the body that have high viral activity, such as the lymph nodes, brain, bone marrow and genital secretions. If a correlation exists, then as viral load levels are reduced in the blood, we will know that levels are also being reduced in other inaccessible tissues. Recent data show that this is probably true of the lymphoid tissues. There has been recent talk that it may be possible to eradicate the virus altogether in people with sustained undetectable viral load levels (undetectable on the 20 copy-sensitive assay) and who are taking combination antiviral therapies, but this idea is still in the very early theoretical stages (see PI Perspective #19, "Eradication of HIV - Hope or Hype?"). People should be warned that stopping or reducing antiviral therapy because the viral load is perceived as "undetectable" may result in the resurgence of the virus because HIV may be hiding out in other reservoirs of the body not detected by viral load tests. In fact, this has been seen in people in clinical studies. Reducing antiviral therapy may also lead to drug resistance. Researchers are looking into the development of another type of HIV test that would detect HIV inside a cell, before it has had a chance to replicate and before it would be detected by the new viral load test. If virus is present in cells despite undetectable HIV RNA plasma levels, it is more likely that that viral levels will rise again at some point. If eradication is a true possibility, a highly sensitive viral load test needs to be developed to make certain that no virus is present in the body. HIV viral load is only the first area where these new testing technologies (Polymerase Chain Reaction, or PCR, and other amplification techniques) have been applied. Other tests based on these techniques are similarly used in research to measure other viral diseases, such as CMV, and bacterial diseases, such as tuberculosis (TB), both of which are of great concern for people living with HIV. As these tests become standardized, understood and accepted they will help us better understand the relationship between HIV, opportunistic infections (OIs) and the immune system. The best predictor of disease progression is likely to be not one single test but a combination of tests, including viral load tests, CD4+ counts and other diagnostic tests, that each tell us about different elements of the immune system’s status and fight against HIV. Whereas CD4+ cell counts tell us what has already happened in the body, viral load levels allow us to predict what might happen in the future. Used together, these tests will allow physicians and patients to make earlier and more definitive decisions about treatment and health status. |
| Resistance
Tests HIV resistance has recently become a major concern for people with HIV. Resistance usually occurs when the drugs being used are not potent enough to completely stop HIV replication. If HIV can reproduce at all in the presence of drugs, it has the opportunity to make changes in its structure (mutations) until it finds one that allows it to reproduce in spite of the drugs. Once such a mutation occurs, it then grows unchecked and soon is the dominant strain of HIV in the individual. The drug becomes progressively weaker against the ‘new’ strain of HIV (though it may still have some value). Most of the research on HIV resistance has been done in laboratory studies. It is unclear how much information from these studies is relevant to humans. However, increasing research on HIV drug resistance is being incorporated into studies. There are two general ways in which researchers attempt to determine whether resistance is occurring. One approach, called genotypic testing, seeks to determine any changes to a part of HIV’s genetic structure, which change the way the virus makes key proteins (like the protease or reverse transcriptase enzyme). Such changes are referred to as mutations. It is known that certain mutations result in the loss of a given drug’s effectiveness, so researchers primarily look for these known mutations. The other approach, called phenotypic testing, is a more direct measure of resistance. It examines the amount of drug needed to inhibit the growth of HIV in a laboratory setting. In its natural state, when HIV is not resistant to a particular drug, known levels of the drug completely suppress viral replication. Resistant HIV requires higher levels of the same drug to get an equal level of suppression. However, it is not feasible to increase the dose of a drug indefinitely as this leads to increased toxicities. Most drugs are already given at near their maximum tolerated dose. Thus, if a person’s resistant virus requires 10 times more drug before it will stop replicating, the person can not simply take a larger dose of the drug to overcome resistance. Generally, any time viral mutations make it necessary to use a dose 4 times higher than standard to suppress replication, the virus is considered to have high-level resistance. The only choice is to use a different drug. Not all genotypic changes lead to phenotypic changes, and a lot of work is being done to better understand these findings. Generally, it takes a few genotypic changes before you see a phenotypic change. However, there are some drugs, such as 3TC (Epivir), which require only a single mutation to induce high level phenotypic resistance. Both genotypic and phenotypic resistance testing have their drawbacks. While phenotypic resistance is probably more relevant, it does not necessarily provide information about which drugs a person might still benefit from. Genotypic resistance gives specific information about changes in the enzyme and scientists know, to some degree, which changes affect which drugs. But those changes do not always mean that the drug has completely lost effectiveness. Ideally, both genotypic and phenotypic information would be obtained to make the best treatment decisions. Unfortunately, measuring genotypic or phenotypic drug resistance is difficult, laborious and expensive. Only a few university laboratories can perform these tests. Work is proceeding on increasing availability of these technologies. Several companies are working on new technologies to detect resistance rapidly and these technologies are currently in clinical studies. One test which is being offered by Specialty Labs in Santa Monica, California detects genotypic changes affecting the nucleoside analogue drugs but not changes which affect non-nucleoside reverse transcriptase inhibitors or the protease inhibitors. This test costs about $300 and currently some insurance companies are covering these costs. Due to start-up problems, there have been some delays in getting results from this test. A group in Belgium is developing a process called the RT-antivirogram which provides phenotypic resistance testing of the nucleoside analogue drugs and the non-nucleoside reverse transcriptase inhibitors. Work is being done to extend this to include phenotypic testing of protease inhibitors. Additionally, since this test is still for research and has not been approved by the FDA, most third party payers will not reimburse the $600 cost for the test. Affymetrix of Santa Clara California has developed a high technology GeneChip system which detects genotypic changes in both the protease and reverse transcriptase enzymes. This system should be able to determine the presence of resistant mutations for all of the currently approved antiretroviral drugs. Conducting this test can be fairly simple because it can use the same blood sample used to test viral load by PCR (polymerase chain reaction). Interpretation of the test results is somewhat more difficult. Unfortunately, this test is not yet available commercially although we expect that there will be limited access to this technology soon. Project Inform has made further development and availability of this test a high level advocacy goal for the coming year. Less Frequent Routine TestsThe following tests are considered routine in HIV-positive individuals, but do not need to be performed as frequently. PPD Skin Test and Chest X-rays - Pap Smears - The following tests are not considered routine for HIV-infected people without symptoms, but may be necessary in later stages of HIV disease. As CD4+ counts decline, the possibility of opportunistic infections increases, and thus various tests should be performed to monitor for these infections. Hepatitis Serology - Toxoplasmosis Serology (IgG) - |
Table
of Common Tests and Ranges
Interpreting Your Viral Load Numbers Users of viral load tests have had to learn or relearn a few words not heard since math class. Because changes in viral load can be very large, researchers use a "LOGarithmic" scale to express the test results. The log scale is simply a shorthand way to express large numbers. The following is a method to help you figure out your log change based on your viral load numbers. The graph below gives you the log values on the horizontal axis and their respective copy numbers on the vertical axis. Here are some simple directions to figure out your very own viral load log change.
One last thing that may be helpful for conceptualizing logs is that a 1 log change is the same as a 90% change. For example, a change from 100,000 down to 10,000 is a 1 log drop or a 90% decrease. A 2 log change is equivalent to a 99% change. And, finally, a 0.5 log change is the same as a 70% change. |
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