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Lyme Testing

Explaining Lyme Disease Testing Techniques - YouTube

Christian Perronne, MD, PhD - Current Lyme diagnostic testing, NorVect 2014 - YouTube

Lyme Disease Testing: Bob Giguere from IGeneX - YouTube

Unconventional diagnostic tests for Lyme borreliosis: a systematic review.

Validation of cellular tests for Lyme borreliosis (VICTORY) study.

Detection of Borrelia burgdorferi Cell-free DNA in Human Plasma Samples for Improved Diagnosis of Early Lyme Borreliosis.

2019 - Appropriate laboratory testing in Lyme disease.

A limitation of lab testing is that we look for evidence of Lyme in body fluids or tissues where spirochetes do not reside. For example, dark-field microscopy, conventional PCR tests, or the new "Borrelia culture" look for microbes or their DNA in the blood. Once infected with Lyme, Lyme has a brief blood-borne phase and concentrates in the connective tissue, inside neurons or ganglia, inside lymph nodes, in the fascia, and in other tissues or organs. Even more questionable is the attempt to search for Lyme spirochetes in the cerebrospinal fluid after doing a spinal tap.

As far back as 1998, a study already showed that spirochetes immediately give up their spirochete-form and shift into the "round body" shape when they get in contact with spinal fluid (Brorson, & Brorson, 1998). The round body is not detectable with the typical tests the lab applies when searching for living spirochetes. (Source)

Recommendations for Lab Tests

The blood tests currently used to detect the presence of the Borrelia burgdorferi bacterium are the enzyme-linked immunosorbent assay (ELISA) and the Western blot test. There aren’t any good reliable tests. Tests, like Western Blott (which can give a lot of false negatives), are tests for the antibodies to bacteria and you can’t get that relationship unless they’re getting to each other…bacteria hidden in biofilm won’t be reached for antibody production. At the earliest stage, the acute stage before biofilms are created, ELISA, Western blot and other tests that look for antibodies to Borrelia may pick it up. Once it's become chronic something like the CD57 test may be necessary.

Other problems with the serologic lab tests include:

Serologies are problematic because all of them are based on only one or two lab strains.

Low Sensitivity –

Paradox – the more ill the patient, the weaker the serologic response and LESS likely you are to get a positive test.

Serologies cannot detect an active infection, only prior exposure.

The CD57 test isn't looking for antibodies but rather, takes a count of a patient's CD57 cells. CD57 is a natural killer cell, CD stands for cluster designation, which is simply a molecule in the cell that gives it its identity. The “57” is just an implication of the numerical order it was discovered in. Its designed to kill many types of bacteria but is uniquely attracted to borrelia burgdordferidue to its cell walls which consist of lipopolysaccharides- which is a lattice type of structure. Unlike other CD natural killer cells, CD57 specifically responds to this type of foreign invader. Borrelia are believed to be capable of reducing the total amount of CD57 cells. Its unknown how it does this but the results of this action are used as a means of confirming a diagnosis or determining remission of Lyme by some physicians. Testing CD57 cell counts can be done through Labcorp. Its generally believed that the lower the amount of CD57 natural killer cells, the more chronic or progressive Lyme disease is, and the higher the amount, the closer a person is to remission. There have been instances where high amounts of C57 have been found in those with chronic Lyme and low amounts in those who were believed to be in remission. While the C57 test isn’t 100% accurate in terms of diagnosing chronic lyme it is an incredibly useful test in doing so when it is compared with other diagnostic lyme tests, such as a western blot test or the c3a and c3a complement protein test as well as the person’s symptoms.

Dr. Zubcevik said research has shown there are 10 different strains of Lyme disease in the United States, and many of them do not test positive on the traditional Western blot or ELISA tests. In a previous email to The Times, she wrote that with current testing, 69 out of 100 patients who have Lyme disease may go untreated.

“The bull’s-eye rash only happens 20 percent of the time,” she said. “It can often look like a spider bite or a bruise. If you get a bull’s-eye it’s like winning the lottery. Borrelia miyamotoi, which we have a lot in Massachusetts, will not test positive on either test. That’s a huge problem, so the CDC is moving toward a different kind of test.”

Borrelia miyamotoi also has the potential to spread rapidly, since it’s transmitted directly from mother to offspring. Nymphal deer ticks need to feed on a mammal, most likely the white-footed mouse, to contract the virulent Borrelia burgdorferi bacterium.

Per wikipedia:

Several forms of laboratory testing for Lyme disease are available, some of which have not been adequately validated. The most widely used tests are serologies, which measure levels of specific antibodies in a patient's blood. These tests may be negative in early infection as the body may not have produced a significant quantity of antibodies, but they are considered a reliable aid in the diagnosis of later stages of Lyme disease.[91] Serologic tests for Lyme disease are of limited use in people lacking objective signs of Lyme disease because of false positive results and cost.[92]

The serological laboratory tests most widely available and employed are the Western blot and ELISA. A two-tiered protocol is recommended by the Centers for Disease Control and Prevention: the sensitive ELISA test is performed first, and if it is positive or equivocal, then the more specific Western blot is run.[93] The reliability of testing in diagnosis remains controversial.[43] Studies show the Western blot IgM has a specificity of 94–96% for people with clinical symptoms of early Lyme disease.[94][95] The initial ELISA test has a sensitivity of about 70%, and in two-tiered testing, the overall sensitivity is only 64%, although this rises to 100% in the subset of people with disseminated symptoms, such as arthritis.[96]

Erroneous test results have been widely reported in both early and late stages of the disease, and can be caused by several factors, including antibody cross-reactions from other infections, including Epstein–Barr virus and cytomegalovirus,[97] as well as herpes simplex virus.[98] The overall rate of false positives is low, only about 1 to 3%, in comparison to a false-negative rate of up to 36% in the early stages of infection using two-tiered testing.[96]

Polymerase chain reaction (PCR) tests for Lyme disease have also been developed to detect the genetic material (DNA) of the Lyme disease spirochete. PCR tests are susceptible to false positive results from poor laboratory technique.[99] Even when properly performed, PCR often shows false negative results with blood and cerebrospinal fluid specimens.[100]Hence, PCR is not widely performed for diagnosis of Lyme disease, but it may have a role in the diagnosis of Lyme arthritis because it is a highly sensitive way of detecting ospADNA in synovial fluid.[101]

Culture or PCR are the current means for detecting the presence of the organism, as serologic studies only test for antibodies of Borrelia. OspA antigens, shedded by live Borrelia bacteria into urine, are a promising technique being studied.[102] The use of nanotrap particles for their detection is being looked at and the OspA has been linked to active symptoms of Lyme.[103][104] High titers of either immunoglobulin G (IgG) or immunoglobulin M (IgM) antibodies to Borrelia antigens indicate disease, but lower titers can be misleading, because the IgM antibodies may remain after the initial infection, and IgG antibodies may remain for years.[105]

Western blot, ELISA, and PCR can be performed by either blood test via venipuncture or cerebrospinal fluid (CSF) via lumbar puncture. Though lumbar puncture is more definitive of diagnosis, antigen capture in the CSF is much more elusive; reportedly, CSF yields positive results in only 10–30% of affected individuals cultured. The diagnosis of neurologic infection by Borrelia should not be excluded solely on the basis of normal routine CSF or negative CSF antibody analyses.[106]

New techniques for clinical testing of Borrelia infection have been developed, such as LTT-MELISA,[107] although the results of studies are contradictory. The first peer reviewed study assessing the diagnostic sensitivity and specificity of the test was presented in 2012 and demonstrated potential for LTT to become a supportive diagnostic tool.[108] In 2014, research of LTT-MELISA concluded that it is "sensible" to include the LTT test in the diagnostic protocol for putative European-acquired Lyme borreliosis infections.[109] Other diagnostic techniques, such as focus floating microscopy, are under investigation.[110] New research indicates chemokine CXCL13 may also be a possible marker for neuroborreliosis.[111]

Some laboratories offer Lyme disease testing using assays whose accuracy and clinical usefulness have not been adequately established. These tests include urine antigen tests, PCR tests on urine, immunofluorescent staining for cell-wall-deficient forms of B. burgdorferi, and lymphocyte transformation tests. The CDC does not recommend these tests, and stated their use is "of great concern and is strongly discouraged".[100]

Imaging

Neuroimaging is controversial in whether it provides specific patterns unique to neuroborreliosis, but may aid in differential diagnosis and in understanding the pathophysiology of the disease.[112] Though controversial, some evidence shows certain neuroimaging tests can provide data that are helpful in the diagnosis of a patient. Magnetic resonance imaging(MRI) and single-photon emission computed tomography (SPECT) are two of the tests that can identify abnormalities in the brain of a patient affected with this disease. Neuroimaging findings in an MRI include lesions in the periventricular white matter, as well as enlarged ventricles and cortical atrophy. The findings are considered somewhat unexceptional because the lesions have been found to be reversible following antibiotic treatment. Images produced using SPECT show numerous areas where an insufficient amount of blood is being delivered the cortex and subcortical white matter. However, SPECT images are known to be nonspecific because they show a heterogeneous pattern in the imaging. The abnormalities seen in the SPECT images are very similar to those seen in people with cerebral vacuities and Creutzfeldt–Jakob disease, which makes them questionable

New Lyme Test

C. S. Cheung and Maryland colleagues have found a way to detect Borrelia burgdorferi membrane proteins in human serum shortly after infection. B. burgdorferi membrane proteins are ≈107 lower in abundance than serum proteins, according to their abstract. The new test permits early detection of Lyme disease. The researchers focused on a membrane protein that was easily differentiated from human serum proteins.

The researchers used a high-speed centrifuge to separate the serum proteins from the B. burgdorferi membrane proteins and identified the spirochete's proteins using multiple reaction monitoring mass spectrometry.

Cheung and colleagues detected the membrane protein in serum samples taken from three patients with unconfirmed Borrelia infection. The researchers were able to detect Borrelia infection in one patient 3 weeks before the standard blood test came back positive. Results from the new test and the standard test simultaneously confirmed infection in the other two patients. Although a characteristic bull's-eye rash is a defining symptom of Borrelia infection, 20% to 30% of people with Lyme never exhibit the rash. The standard blood test for Lyme measures accumulated antibodies to the infection. It takes 4 to 6 weeks before antibodies reach detectable levels. Because the new test measures bacteria membrane proteins, it can confirm diagnosis shortly after infection occurs, which means earlier treatment. The research team believes that its technique can be used for early detection in other bacterial infections as well.

Cheung CS, Anderson KW, Benitex KY, et al. Quantification of Borrelia burgdorferi membrane proteins in human serum: a new concept for detection of bacterial infection [abstract]. Anal Chem. November 17, 2015; 87(22):11383–11388.

New experimental test detects signs of Lyme disease near time of infection [press release]. National Institute of Standards and Technology (NIST). February 12, 2016.

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