Charna Albert
August 2026—In a summer of more ticks, more diseases, and more tick species expanding to new areas and bringing diseases with them, what Bobbi Pritt, MD, MSc, would like to see are more—and better—diagnostics in early Lyme disease.
Laboratory scientists are inching toward improved solutions, though nothing is commercially available now. “I’m cautiously optimistic. I wouldn’t say I’m excited yet,” says Dr. Pritt, co-director of vector-borne diseases laboratory services and chair of the Division of Clinical Microbiology at Mayo Clinic in Rochester.
Across the U.S., tick bites were reported earlier in the year than usual, Dr. Pritt told CAP TODAY in mid-June. “And we continue to see more visits to the emergency department and walk-in clinics for concerns about tick-borne diseases,” she says. Though incidence is rising for all tick-borne illnesses, Lyme disease remains the most common vector-transmitted disease in the country. “There are tens of thousands of cases reported to the CDC each year, but the actual number looks like it’s about 500,000. That’s a huge burden.”
To add to that burden, laboratory testing is insensitive in early Lyme disease, when treatment is most effective. Blood-based PCR testing isn’t recommended; Borrelia burgdorferi, the main causative agent, is present in the bloodstream for a vanishingly brief period and only in minute quantities. “You’re lucky if you’ll detect half the cases, even in patients with a clear-cut bull’s-eye rash,” Dr. Pritt says. Serology is the mainstay of diagnosis, but an antibody response is generally undetectable for several days to weeks after infection. “We’ve struggled with this as a branch of medicine, trying to find something that can detect that early phase.”
The Food and Drug Administration in 2019 approved the modified two-tiered testing algorithm for Lyme disease, in which two or more ELISAs are used for confirmatory testing—an update to the standard two-tiered testing algorithm’s ELISA followed by confirmatory IgM and IgG immunoblotting. In a recent study Dr. Pritt coauthored, the MTTT confirmed more early cases than the STTT, though both testing algorithms were shown to be insensitive in early-stage Lyme disease (Horn EJ, et al. J Clin Microbiol. 2026;64[5]:e0118725). “They’re detecting antibodies, and that relies on the individual producing those antibodies to a level that can be detectable. So there’s always going to be a lag,” Dr. Pritt says.
In their study, Dr. Pritt and her coauthors compared the performance of four FDA-cleared STTT and MTTT algorithms using serum samples from 251 participants (107 cases, 69 with a convalescent draw; 144 endemic controls). Only 39 percent (42/107) of the first-draw samples from participants with early Lyme disease were STTT or MTTT positive. There was also discordance between algorithms: Only 22 of the 45 samples classified as laboratory confirmed by Lyme Disease Biobank testing were positive using all algorithms evaluated.
A clinical diagnosis sans laboratory testing can be made in an endemic area when a patient presents with erythema migrans. “The problem is not all physicians know that and sometimes they won’t give a full treatment,” Dr. Pritt says. “The other problem is that rash may not always be seen by the patient. It may be on their back or scalp. And there’s still the 30 percent or more of people who don’t get the rash. That’s where this study gets into the discussion of needing something for that early diagnostic period.”
Other tick-borne pathogens—the bacteria that cause anaplasmosis or the parasites that cause babesiosis, for instance—can be detected by PCR, says Rafal Tokarz, PhD, associate professor of epidemiology in the Center for Infection and Immunity, Columbia University Mailman School of Public Health. “Those are present in enormous numbers in the bloodstream, so they’re very easy to detect. To detect Borrelia burgdorferi in blood by a molecular test like qPCR is extremely difficult,” he says. “It’s the biology of the pathogen we are limited by—not necessarily the lack of technology.”
New diagnostics are on the way, Dr. Pritt says. “There are some things in the literature already. They’re just not FDA cleared.” She has greater interest in the rapid antigen and rapid antibody tests in development and less in the more esoteric methods. “I’m not so excited about a complex molecular shotgun metagenomics approach that’s going to take several days to send to the reference lab and get the result,” she says. “I’m excited about anything that’s going to be rapid, affordable, and easily available.”
Among those working on new solutions is Andrew Levin, PhD, chief executive and chief scientific officer of Kephera Diagnostics, which has developed the Hybrid Lyme ELISA, a novel single-tier methodology. Dr. Levin and his colleagues reported last year in the Journal of Clinical Microbiology that the test showed greater sensitivity for early Lyme disease than the STTT and MTTT algorithms and equivalent specificity, and it demonstrated more than 90 percent sensitivity in patients with erythema migrans (Levin AE, et al. J Clin Microbiol. 2025;63[9]:e0048325).
Dr. Levin sees the Hybrid Lyme ELISA, a simple ELISA in principle, as an eventual Lyme testing workhorse. “It’s something most laboratories that do any volume of testing are set up to do. They have all the equipment to run ELISAs as a conventional method. People know it well. They know the steps.” The assay is undergoing a multicenter clinical trial now for FDA review. At the same time, Kephera is developing and validating an in-house version of the test in its CLIA-certified laboratory; Dr. Levin and his colleagues intend to offer it as a laboratory-developed test while they await regulatory approval.
Dr. Levin hopes to dispense with the two-tier protocol if an equivalent level of specificity can be reached with a single test. The data generated so far, published last year, “showed we did hit that goal.” Further studies are needed to demonstrate how high the specificity is in different control populations. If statistically equivalent to the two-tier protocol, “we could make the whole process more streamlined, more manageable, and more cost-effective,” he says. The company’s other objective: develop a test sensitive enough to detect infection in the erythema migrans stage.
With the Hybrid Lyme ELISA, Kephera’s approach differs from that of the commercially available serologic assays, which target combinations of the antigens expressed early in infection, such as VlsE and OspC, using a conventional indirect ELISA format. The Kephera assay relies on the simultaneous binding of individual antibody molecules to the B. burgdorferi surface protein VlsE and to the C6 peptide derived from it. This dual binding requirement makes the assay’s high degree of specificity possible—a discovery that came from years of experimentation, Dr. Levin says.
“I had run a lot of ELISA assays using the C6 peptide as the antigen. I also ran a lot of ELISAs using the VlsE protein from which the peptide is derived,” he says. He was studying sera from negative control subjects when he realized that using both antigens in a single assay could potentially reduce the number of false-positives. “We found a few false-positives from the C6 peptide ELISA, and we also found a few false-positives from the VlsE ELISA. The interesting finding was that they were not the same sera that were false-positive.
“That’s what made me think,” he continues, “whatever it is that’s reacting and causing the false-positivity, maybe it’s not the true Borrelia-specific epitope in this antigen.” That epitope, he hypothesized, is shared by the C6 peptide and the VlsE protein. Whether the assay design—with the protein on one side of an antibody and the peptide on the other—would work was far from assured; the peptide, he explains, is largely concealed within the structure of the protein and thus not easily accessed by the antibody. But when he and his colleagues tested serum from Lyme patients, “we found it did work. We started detecting positive signals in Lyme serum, and the controls were all very clean.”
In their study, Dr. Levin and his coauthors tested a subset of 32 serum samples from patients presenting with erythema migrans for whom the dates of onset of symptoms and sample draw were known. For samples collected within seven days of onset, the assay detected all 15 serum samples, compared with five of 15 detected by the STTT. The MTTT detected 12 of 15.
The test has low background reactivity, Dr. Levin says, enabling a very low cutoff that allows the detection of weak signals. “That’s our theory that we believe explains how it’s able to pick up serum samples from patients with very low levels of antibodies that would otherwise be lost in the background.”
One potential limitation is that the test does not distinguish between IgM and IgG reactivity, a shortcoming in situations that might call for a diagnosis of early- versus late-stage Lyme disease. Cross-reactivity is another potential weakness: B. miyamotoi infection, Dr. Levin and his coauthors note, could lead to a false-positive, though this may not be clinically important as the treatment is the same for Lyme and B. miyamotoi infections.
Dr. Levin’s take on the Lyme disease testing landscape: Don’t discount serology.
“It has gotten better and better for Lyme disease. It has further to go, but it offers a lot of potential in delivering high-quality, accurate results for Lyme.”
“People might have the impression we need a change,” he says. “But technologies develop over time and get better. The computers we use today are not the same computers we used 20 years ago, but they’re still computers. And serology is like that.”
The rarer and more serious tickborne diseases have been on display at Mayo Clinic this summer.
“I’ve been quite surprised to see a good number of Powassan virus-positive cases,” Dr. Pritt says. “That’s a tick-borne virus that can cause severe neurologic disease and can be fatal. We’re also seeing what I would consider to be an unusually high number of Heartland virus PCR-positives,” indicating acute, not past, infection.
Climate change is the main culprit, says Dr. Tokarz of Columbia. With the increases in temperature, tick species are expanding beyond their usual range. “Ticks that we normally found in the South historically we now find up in Maine,” he says. “And because the range is expanding, you’re going to have more ticks in new places and the pathogens they carry will expand as well.”
Take Heartland virus, initially identified in 2009 in Missouri. “Most of the cases historically over the past decade were in the South, but the tick that transmits this virus is now present throughout the Northeast,” Dr. Tokarz says. “There have been reports of Heartland virus in New York State. That’s not something you would hear about 10 years ago.”
Like Lyme disease, many of the tick-borne infections can be treated with antibiotics such as doxycycline. But for others, like the viruses and Babesia, treatment is different. “It’s imperative that patients, and particularly elderly patients who might be more susceptible to babesiosis, get tested for both,” he says.
With tick-borne diseases on the rise, a diagnostic strategy that considers coinfection is another new and needed frontier.
“That’s ultimately where we’re going,” Dr. Tokarz says, noting that more physicians are becoming aware of the potential for coinfection, especially in areas of high endemicity.
How often coinfection occurs is difficult to estimate, though tick surveillance studies can help researchers extrapolate prevalence rates. It’s easier to test the ticks themselves than it is to test patients, he explains. “You can do it by PCR, often multiplex PCR,” and get an answer quickly. To test patients, “you would have to perform several costly tests to exclude other pathogens, but also to establish some kind of a prevalence rate.”
“We found that on Long Island, approximately 25 percent of ticks infected with Borrelia burgdorferi . . . were also infected with Babesia.” A patient bitten by one of these ticks could be thought to have a 25 percent chance of coinfection. “If you look at the actual numbers of Lyme disease cases and Babesia cases, that’s nowhere near true,” Dr. Tokarz says, because Babesia testing trails testing for Lyme disease.
In his analyses of whole blood samples of patients with Lyme disease, 10 to 15 percent of patients were estimated to also have Babesia. To establish a true prevalence rate, though, “you would have to have a very large patient cohort and test for everything. For a regular laboratory, that’s a very costly thing to do, and for a public health laboratory, it’s just not feasible.”
More laboratories are developing multiplex serological tests for Lyme disease and other tick-borne pathogens, he says, including his own. “Before it was always Lyme serology, PCR everything else, pretty much. Now it’s becoming more uniform.” His laboratory’s test is a 30-minute lateral flow antibody test for Lyme disease, babesiosis, and anaplasmosis. It has slightly higher sensitivity for Lyme disease than the STTT, he says. For now, it’s a research-grade test, though they’re seeking partners to take it through the regulatory approval process. “It works quite well,” he says. “We want to take it further, but we just haven’t gotten there yet.”
Could a multiplex serological test include the tick-borne viruses, such as Powassan and Heartland virus? “Possibly, but it’s very difficult to do,” Dr. Tokarz says. “Serology is highly cross-reactive, so for every agent, you would need to have a very thorough evaluation to ensure your test is specific.” He expects additional targets to be added over time as multiplex serology tests become available. “It’s baby steps,” he says.
Another area of study for Dr. Tokarz is how strain diversity affects disease. Work in animal models has implicated several strains of B. burgdorferi with increased likelihood of systemic dissemination, he and his coauthors note in a study (Jain K, et al. Sci Rep. 2021;11[1]:12384). And different strains within B. burgdorferi can be distinguished that affect pathogenicity and virulence in humans. B. burgdorferi RST1 strains, for example, which account for about 40 percent of Lyme borreliosis cases in the northeastern U.S., are more likely to disseminate hematogenously and are associated with a higher risk of antibiotic-refractory Lyme arthritis. “Diagnostic tests tell us only whether a person is positive or negative,” he says. “We generally don’t know anything about the strains that are circulating.”
High-throughput sequencing studies could help answer questions about strain diversity and how it might be linked to Lyme disease-associated syndromes such as neuroborreliosis. The main limiting factor—aside from cost, Dr. Tokarz notes—is inadequate sensitivity. To address that gap, Dr. Tokarz and his colleagues have developed a capture sequencing assay for enhanced detection and genotyping of tick-borne pathogens. The assay, TBDCapSeq, uses hybridization capture probes that cover the complete genomes of the 11 most common tick-borne agents found in the U.S.
Capture sequencing uses agent-specific probes to selectively capture the template of interest prior to sequencing. “Regular sequencing takes everything in the sample and amplifies and sequences it . . . but it’s inherently very insensitive. If you have a pathogen, it’s like looking for a needle in a haystack. With regular next-generation sequencing, you’re not going to find the needle,” Dr. Tokarz says. TBDCapSeq enriches for the pathogenic nucleic acid, capturing the specific desired DNA or RNA and washing away the background. NGS is then performed only on the captured material.
“We find that doing this enriches for the desired template sometimes up to four magnitudes,” he says. “We go from nothing to not only detecting something but recovering a substantial part of the genome, which allows us to study the pathogen more.”
Molecular methods like capture sequencing could potentially complement serology to detect early-stage Lyme disease. “We need to come up with some kind of combination of the two,” Dr. Tokarz says. “Molecular tests for Lyme disease have shortcomings that can be offset by serology tests and vice versa.”
“Using capture sequencing,” he continues, “we can detect more samples as positive than just regular PCR.” It’s still only a portion of samples, he notes. “But in the future, if something like this were to be implemented, it would probably be implemented early in disease, where you’re more likely to find the bacteria in the bloodstream but you won’t be able to detect it serologically.” Later in infection, when the pathogen can’t be found in the bloodstream, “that’s when serology can become useful.”
Also on the docket for Dr. Tokarz and his collaborators is a study of the causes behind Post-Treatment Lyme Disease Syndrome.
No diagnostic test can ascertain whether Lyme disease causes the syndrome’s symptoms, which as many as 10 to 20 percent of people who are successfully treated experience, says Mayo’s Dr. Pritt. Studies have shown that B. burgdorferi is no longer present in these patients, though there’s data to show its proteins could still be in the body, especially in the case of joint disease. “That’s a protected space where antigens could be hard to clear and the body may be forming an immune response to those. So I would say our knowledge continues to expand and we don’t have all the answers yet,” she says.
Dr. Tokarz and his colleagues are among those searching for answers. They reported in 2026 the results of a study in which a high-density peptide array was used to examine antibody responses to more than 60 primary antigens of B. burgdorferi from one cohort of PTLDS patients and another of recovered patients (Marques AR, et al. Sci Rep. 2026;16[1]:13368). “We were hoping to find some kind of serologic marker that’s indicative of Post-Treatment Lyme Disease [Syndrome]. We wanted to see if there’s a particular Borrelia burgdorferi peptide that patients with Post-Treatment Lyme Disease Syndrome have a more robust antibody response to,” indicating a potential autoimmune response. That study didn’t yield a serologic finding, though Dr. Tokarz still believes the cause could be autoimmune. He plans in the future to do further autoimmunity testing with human proteins.
It’s important to rule out other potential causes when a patient with a history of a positive Lyme disease test experiences long-lasting symptoms after treatment, Dr. Pritt says. “There have been cases where people had something else, like leukemia or fibromyalgia or rheumatoid arthritis,” she says, and it flew under the radar because it was assumed the patient had PTLDS.
Many patients get treated effectively in early Lyme disease with a clinical diagnosis.
“The bottom line is that clinicians are still going to give doxycycline right away,” Dr. Pritt says. “And they should if they suspect Lyme disease,” until a highly sensitive and specific point-of-care test is developed.
Some patients, to be sure, do not have Lyme disease and end up taking antibiotics unnecessarily, Dr. Levin says. “Whether a rapid test could be helpful with that would depend on how accurate a test it is. That’s going to be a challenge. It’s not impossible, but it’s not a slam dunk to get to the level of accuracy of a good lab test.”
There are other scenarios, too, where rapid testing is needed—distinguishing potential Lyme arthritis from septic arthritis, for instance, before a patient undergoes surgery. “We’ve learned from ER physicians that that’s something people consider a need,” he says. The rapid test, however, must meet criteria for making a medical decision. “And not all rapid tests fit within those requirements.”
When it comes to Lyme disease, Dr. Pritt says, “the earlier you treat the better.” And right now, “there’s still a gap” in being able to catch patients at those early time points.
“But that’s why we have all this interest in this field now,” she says.
Charna Albert is CAP TODAY senior editor.