Many scientists and physicians agree that there are important issues concerning present Lyme disease (LD) testing. In this post I will address how the commercial testing cleared by the Food and Drug Administration and approved by the Centers for Disease Control and Prevention (CDC) results in under-identification of the number of individuals, especially children, suffering from LD.
First let’s review the recommended testing to confirm exposure to Borrelia Burgdorferi (BB), the bacteria that causes LD. Testing consists of two parts or “tiers.” Results of these two tests depend upon the ability to detect the antibodies that our bodies make when exposed to the BB bacteria. The first is an enzyme-linked immunosorbent assay (ELISA). The ELISA is considered a screening test. According to the Center for Disease Control and Prevention recommendations, if the ELISA result is negative, the search is over and no further testing is needed. If the result is equivocal or positive, the second part of the test, or second tier called a Western Blot (WB), should be done. The WB looks for the presence of antibodies to certain proteins (each numbered according their different weights) associated with BB. These proteins are depicted on special testing strips as lines called bands. The bands are numbered 18, 22, 23-25, 28, 30, 31, 34, 39, 41, 45, 58, 66, 73, 88, and 93, reflecting their weight in units called kilodaltons. Testing consists of exposing the patient’s serum (blood without the clotting factors or blood cells) to the antigen bands to see if a reaction occurs. A positive band is supposed to represent an antibody response to a protein found on the Lyme spirochete. I say “supposed to” because there is the potential that some of the bands can become positive from other infections.
The initial goal of this two-tiered approach, using the ELISA as an initial screen followed by the more specific WB, was to create a test that was highly sensitive and specific in identifying the studied infection. By eliminating the unlikely cases during the first part of the testing and then using more specific identifiers in the second part, the test should be very good, or specific, in identifying most real or true cases of LD. This, in a nutshell, is the rationale for using the two-tiered testing.
The problems with this type of testing are many. One major difficulty in using the ELISA and WB is that the bacterial organism responsible for Lyme disease, BB, is in itself immunosuppressive. So how could a test that depends upon the person’s immune response (the production of antibodies) be much good if we know the organism suppresses the immune system? In other words, if what you are measuring (antibodies) can be affected by the cause of the illness itself (BB), then depending on this test to make the LD diagnosis is questionable. This major flaw in the recommended testing diminishes the value of the test. Unfortunately, relying on a test that is so subject to error is the the weak foundation that has provided support for much of LD research. This testing is additionally problematic in many ways.
1.The ELISA is a poor screening test.
The concept of two-tier testing is not unique to LD and has been used effectively in diagnosing other illnesses such as Acquired Immunodeficiency Syndrome (AIDS). But in LD, the first step, the ELISA, is very limited, on average detecting only 56% of cases, meaning that it is so very insensitive, it actually misses 44% of cases of people who do have LD. This is in stark contrast to the 99.5% effective rate found when used in testing for AIDS.1
Compounding this insult the rule that only those whose results are either indeterminate or positive on the initial ELISA test should go on to the second part of the test. Basically, present recommended use of the ELISA to screen for LD actually eliminates close to half the LD cases from further testing. According to the official guidelines, testing is halted with a negative ELISA and the individual is told that he/she does not have LD. Clearly, this easily leads to an underdiagnosis of LD infections. Going on to use the WB in those who were indeterminate or positive on the ELISA, results in a 99% specificity (accuracy), but if you’ve already missed half the infected group, then the testing is quite problematic.
2. The Western Blot has its own issues.
The WB looks for the presence of an immune response to BB proteins, focusing on two types of antibodies: immunoglobulin M (IgM) and immunoglobulin G (IgG). When a person comes in contact with an infectious agent, the body makes IgM antibodies as its first line of defense. In general, it takes two to four weeks to be at a consistently detectable level, with production peaking at around four weeks and becoming undetectable after 6 months. Persistent ongoing detectable IgM levels beyond the one-month period are subject to controversy in their meaning. Some scientists consider these false positives, while others view it as evidence of persistent infection and associated with chronic illness.
The second antibody produced in the body is IgG. It develops over four to eight weeks after exposure to BB, peaks at approximately six weeks and is gone in less than one year. IgG antibodies are produced to target specific threats like viruses, bacteria and other potentially harmful microorganisms, and forms the basis of long-term protection against microorganisms. Typically, with infectious illness exposure, the IgM antibody response decreases after a while and one is left only with the IgG response, but this transition is not so clear with exposure to the Lyme bacteria.
In the WB test, the laboratory compares the patient’s blood with blots representing a pattern of numbered bands to those seen in previously well documented CDC LD cases. To be considered a positive WB, the tested blot must show a match by having the required minimum number of reactive bands.
As noted, the bands are numbered by weight, with the following bands previously identified as: 18, 22, 23-25, 28, 30, 31, 34, 39, 41,45, 58, 66, 73, 88, and 93. All of the bands written in black are specific indicators that are seen only in BB infections. Three of these bands have been considered highly specific for LD and are given names based on their outer surface proteins (OSP). These bands are known as OSP A (Band 31), OSP B (Band 34) and OSP C (Band 23). Some people believe that a positive result on even only one of these highly specific bands is good evidence of exposure to the BB bacteria. It’s important to keep in mind that one can be exposed to an infectious agent (e.g. virus, bacteria, fungus, etc.) but not necessarily become ill. Therefore exposure and actual illness are different
In the listing, the color blue has been used for bands 28, 45, 58, 66 which are considered nonspecific to LD. This means they can appear positive because the person can have other infections, not only LD . Band 41 is somewhat controversial regarding specificity and that is why it is green.
A positive WB assay is based on having two of the following three bands 23, 39, 41 being positive to be considered an IgM (Immunoglobulin M) antibody positive test. A positive WB IgG (Immunoglobulin G) antibody test requires the presence of 5 of 10 bands: 18, 23-25, 28, 30, 39, 41, 45, 58, 66, or 93.
According to present CDC guidelines, an IgM antibody test can be considered a positive indicator of early exposure to the infection only during the first 30 days after onset of illness. A positive IgM antibody test is generally considered a marker of an acute (recent onset) illness. The official recommendation is that positive IgM antibody results should be disregarded if the patient has been sick for more than 30 days (i.e. 30 days after the bite.) After that time the “gospel” according to the CDC and Infectious Disease Society of America (IDSA) proclaim that a positive IgM antibody test is a false positive. A false positive means that the test is read as positive but isn’t due to LD but caused by another infection or problem.
As noted previously, the typical immunologic progression with infections is a transition from initial production of IgM antibodies to the subsequent production of IgG antibodies. But the situation in LD isn’t typical . As previously discussed, BB bacteria are capable of immunosuppression. This means that the organism itself can interfere with the immune system’s response to infections. The normal transition from making IgM antibodies to IgG antibodies can be hindered. In part this is because of problems that occur in what’s called germinal centers, the part of the lymphoid tissue where the antibodies are made.2 “The rule” that all IgM antibodies present after 30 days from the initial infection must be considered false positives, effectively serves to dismiss and minimize the number of real cases of LD. A negative IgM WB test with a positive IgG WB are considered to indicate either late-stage LD or are a residual result left over from a past infection that is no longer present.
The result of this two-tiered testing system is meant to be 99% specific - meaning these are real cases of LD and not false positives. This present testing approach is so flawed that a statistical analysis by Cook and Puri found that the LD two-tiered testing resulted in 500 times more false-negative (read as non-BB but really is) outcomes than similar two-tiered tests used in the diagnosis of AIDS.3
The 1994 Second National Conference on Serologic Diagnosis of
Lyme Disease was held in Dearborn Michigan. It was attended by representatives of the Association of State and Territorial Public Health Laboratory Directors, CDC, the Food and Drug Administration (FDA), the National Institutes of Health (NIH), the Council of State and Territorial
Epidemiologists, and the National Committee for Clinical Laboratory
Standards. The goal was to established a set of nationwide standards
for LD testing for the purpose of creating consistency in reporting WB results. Unfortunately, the standards selected at that meeting are still in use. In addition to selecting the two-tier testing, the decision was made to eliminate two of the highly specific bands, 31, and 34, from the required testing.
These bands were removed as a vaccine against LD using these proteins was in the planning stage. Investigators knew that the use of these two specific proteins in the vaccine could create false positives for these bands in vaccinated individuals. Put another way, because these proteins were to be used in the vaccine, the vaccinated individuals could test positive for these bands but not have LD.
Participants at the Dearborn meeting seemed to doubt the ability of doctors to remember to ask if the individual had already received the vaccine when the person was getting new testing for LD. Could it be that such a simple step, of asking a question, was all that would have been needed to retain these two highly specific bands as possibilities for optimal testing? In 1998, the Food and Drug Administration (FDA) approved a Lyme vaccine, LYMErix™, which absolutely did have the potential to make these two very specific bands become positive in vaccinated individuals. Although for a variety of reasons that are complicated and will not be addressed here, by February 26, 2002, SmithKline Beecham withdrew the vaccine from the market.
What was accomplished by removing these two highly specific bands? It’s important to keep in mind that these bands were so significant that they were used to make the vaccine. Eliminating them from the diagnostic testing detracted from the WB test’s diagnostic sensitivity. Band 31 is also special because it is not seen until at least months after initial infection. Its presence could potentially serve as an indication of an ongoing infection (chronic rather than acute infection at time of testing.) In addition, bands 31 and 34 have been associated with the presence of neuropsychiatric illness in LD, which could have a crucial impact in the approach to treatment in similarly affected patients. By the elimination of these bands, potentially critical diagnostic, and therefore possible helpful treatment, information was removed.
Why so many of the test’s bands required to make the diagnosis are non-specific, while the two specific bands continue to be omitted, lacks medical, as well as common, logic.
Dr. Paul Fawcett and colleagues reported results of an important study at a Rheumatology Symposia Conference in Texas in 1995. The research was designed to look at the effect of eliminating the two specific bands from the WB criteria.4 The authors compared the diagnostic utility of applying the older criteria (including Bands 31 and 34) vs. the newer WB criteria (as decided at the Dearborn meeting) to 66 child patients with known histories of a tick bite, an erythema migrans rash and symptoms of the illness. Inclusion of Bands 31 and 34 resulted in the identification of 100% of the youth as positive for LD. Using the newer criteria where OSP A and OSP B were omitted resulted in only 31% of the youth being identified as positive for LD. From their analysis, the investigators concluded, “The proposed Western Blot Reporting Criteria are grossly inadequate, because it excluded 69% of the infected children. Utilizing the newer criteria did ensure all cases that tested positive truly showed LD but it served as another way to minimize cases. In effect, using the newer testing criteria excluded as many as 69% of the cases of pediatric LD. Basically by eliminating these bands from the WB, kids became discriminated against and undercounted in the statistics.
It’s unconscionable that these bands were never put back nor to my knowledge has there been significant discussion to reinstate them. This basically leaves doctors in the position of missing more youngsters (as well as non-vaccinated adults) who really have LD.
There is more to consider in how the official change in testing criteria short-changed children, adolescents and even adults. Common sense dictates that testing for bands 31 and 34 in anyone born after 2002 (when the vaccine was taken off the market) would result in higher sensitivity and specificity. The more bands available for identifying the illness, the more likely you’ll identify positive individuals. Why would any medical professional argue against this?
The omission of bands 31 and 34 presents yet another problem for kids. Evidence in the medical literature indicates that positivity for these bands, is associated with the presence of neuropsychiatric issues such as autism. Wouldn’t one want to include in the testing, markers that might alert one to these issues, especially in children?
Continuing to omit these bands for more than two decades, has only served to deny the real number of people who have in the past, and perhaps still continue, to suffer from LD. By removing these two bands, we may be removing years of optimum health, as well as impairing social and cognitive development for some children for their lifetime.
3. Another criticism is that present two-tiered testing is based on indirect evidence of the spirochete.
Since the bacteria is difficult to isolate, much of the current testing is designed to look for reactions indicating that the bacteria is present, further complicating the diagnostic process. Let me explain. Present approved testing looks for the presence of and intensity level of an individual’s immune response to the LD bacterial proteins. Testing would be much less controversial if the organism could be directly cultured or identification of the bacterial DNA occurred. Most scientists would agree that the indirect approach of looking at antibody response with an illness that can cause immunosuppression is inherently fraught with problems and limitations.
Interestingly, at present some research laboratories are focused on developing better and more direct methods of testing for BB. This is sorely needed. Consider that in 2020, the National Institute of Health allotted only 13% ($5.3 million) of their total LD total budget to advance diagnostic Lyme testing.5 Given that testing is the foundation of much research on the diagnosis and treatment, and present recommendations are problematic, 13% is a small amount. It is disturbing to realize that 30% of this allotted money went toward more antibody testing research (i.e. more indirect testing). This only serves to perpetuate the present problem of using an indirect method to identify an immunosuppressive bacterium.
4. Another limitation of presently available laboratory testing is that it is designed to identify only a few species of BB that can cause Lyme disease.
There are multiple strains or types of Borrelia bacteria that can cause Lyme as well as other diseases (e.g. Borrelia miyamotoi causes tick-borne relapsing fever.) Most labs use the strain BB 31 for LD testing. Igenex laboratory uses two strains: BB 31 and BB 297. The more strains used as possibilities, the increased likelihood of getting a positive test result. This is part of why Igenex Laboratory gets more positive testing for LD than bigger labs like Quest and LabCorp. One question is really how many types of Borrelia that cause illness are we missing just because we are using these limited tests?
As you can see, there are multiple problems inherent in the recommended testing. It is crucial to acknowledge that the present faulty testing, which serves as the foundation for many studies, creates bias in all the research that is dependent upon it. This is a major flaw that affected and colored too many aspects of our knowledge about LD. The present system of testing results in a significant underestimation of the number of individuals who suffer from LD. Our people, and especially our children, deserve much better from American medicine.
References
1 Stricker RB and Johnson L. Lyme disease diagnosis and treatment: lessons from the AIDS epidemic. Minerva Med 2010 Dec;101(6):419-25. PMID: 21196901.
2 Hasley CJ, Eisner RA, Barthold SW and Baumgarth N. Delays and Diversions Mark the Development of B Cell Responses to Borrelia burgdorferi Infection. J Immunol June 1, 2012, 188 (11) 5612-5622;
3 https://doi.org/10.4049/jimmunol.1103735.
Cook MJ and Puri BK. Application of Bayesian decision-making to laboratory testing for Lyme disease and comparison with testing for HIV.
Int J Gen Med. 2017; 10: 113–123. Published online 2017 Apr 10. doi: 10.2147/IJGM.S131909 PMCID: PMC5391870.
4 Paul Fawcett et al. Rheumatology Symposia Abstract # 1254. 1995 Rheumatology Conference in Texas.
5 https://www.documentcloud.org/projects/nih-lyme-research-funding-206251/