Timely, differential tick-borne disease diagnosis: The benefits of modified two-tier testing (MTTT) for Lyme disease

Caused by infection with the Borellia burgdorferi bacteria transmitted through the bite of infected ticks, Lyme disease or Lyme borreliosis is by far the most common tick-borne disease in the US. In 2023, healthcare providers reported approximately 90,000 cases of Lyme disease to the CDC.¹ Based on insurance claims data, however, the annual number of patients diagnosed and treated is estimated to be as high as 476,000 per year.²

An analysis of nationwide inpatient data found about 261,630 hospitalizations with tick-borne diseases between 2002 and 2021, with Lyme disease making up about 65% of these admissions.​ Over that 20-year period, total admissions for tick-borne diseases increased about 2.5-fold, showing a meaningful upward trend.​³

Understanding the growing menu of diagnostic tools available—and when to use them—can support earlier diagnoses and help slow the rising incidence of serious, chronic illness related to tick-borne diseases.

In this article:

Clinical challenge | Why it matters | Ordering recommendations | Interpreting test results | Next steps | Supporting resources

Clinical challenge: Tick-borne diseases, coinfections, and the challenge of diagnosis

Tick-borne infections now account for more than 75% of all reported vector‑borne disease cases in the US, making them the dominant vector‑borne threat nationally.⁴

The annual number of tick-borne disease cases has more than doubled in the US in the past 2 decades.⁵ While Lyme disease is the most common of those diseases—affecting approximately 476,000 Americans each year^6,7—there are 17 other known pathogens associated with tick-borne illnesses, with 6 new illnesses discovered in the past 20 years alone.⁸

Due to its vague symptoms, Lyme disease is difficult to diagnose—and the frequency of coinfections further complicates diagnosis. Ticks that transmit Lyme disease frequently carry multiple pathogens such as Babesia microti and Anaplasma phagocytophilum, making coinfections biologically common in endemic areas.⁹

A 2015–2022 analysis of babesiosis cases found a 42% prevalence of one or more coinfections, and 41% of babesiosis patients were coinfected specifically with Borellia burgdorferi, the pathogen that causes Lyme disease.¹⁰

An understanding of coinfection and geographic factors combined with appropriate testing approaches can help primary care providers (PCPs) achieve accurate diagnosis more promptly, increasing the chances of preventing serious, chronic illness.

Choosing the right test at the right time: Molecular vs serologic testing

Testing for tick-borne diseases includes CDC-recommended and FDA-approved serologic laboratory tests as well as molecular testing, such as PCR. For most tick-borne infections, serology can be useful for detecting antibodies in response to a tick-borne infection. Molecular testing can be useful for detecting genetic presence of a tick-borne pathogen within the first few days of infection before antibodies have developed. ^{11, 12}

For Lyme disease, the CDC recommends FDA-approved 2-step serologic testing, using a validated standard two-tier test (STTT) or modified two-tier test (MTTT) algorithm.¹² Quest offers both.

It is important to note that serologic testing during the acute phase of Lyme disease is less sensitive than at later stages of the disease.¹³ Testing done too soon (≤14 days) following infection may produce negative or equivocal results due to the time needed for the immune system to develop a serologic response.¹³ Positive or repeat testing on another sample collected in 7 to 14 days is recommended.¹³

Know which test to choose and when

Detecting more cases of Lyme with MTTT

During early-stage Lyme disease (first 30 days of infection), the MTTT has been shown to have improved sensitivity and detect more cases of Lyme compared to STTT.

Other diagnostic advantages of MTTT include

• Does not include the Western blot, which can be time-consuming and require subjective interpretation of results
• Has a higher sensitivity than the STTT in the early stages of infection and maintains specificity similar to the STTT
• Its algorithm has a higher positive predictive value than the STTT

Why it matters: Timing is critical in improving treatment of tick-borne diseases

Achieving an accurate diagnosis sooner after a tick bite is imperative to more effective treatment and management of tick-borne diseases.

For Lyme disease, babesiosis, and other tick-borne infections, earlier diagnosis and treatment are consistently associated with better short‑term recovery, lower risk of severe complications, and a lower risk of long‑term, persistent symptoms.¹⁴

Most patients treated for Lyme disease in the early localized stage (for example, erythema migrans rash with prompt antibiotics) recover fully and return to their baseline health. Prognosis worsens, however, when diagnosis and treatment are delayed into later stages.¹⁵

To help prevent serious chronic illness and lower the risk of long-term symptoms, providers should prioritize timely, appropriate testing in alignment with current CDC recommendations.

Ordering recommendations: Tick-borne disease testing

The CDC recommends 2-step serologic testing for Lyme disease, using a validated standard two-tier test (STTT) or modified two-tier test (MTTT) algorithm.¹² During early-stage Lyme disease (first 30 days of infection), the MTTT has been shown to have improved sensitivity and detect more cases of Lyme compared to STTT.

Recommended test: Tick-Borne Disease Antibodies Panel with Reflexes

Tick-Borne Disease Antibodies Panel with Reflexes

• Test code: 16220
• CPT^® code(s): 86618, 86666 (x4), 86753 (x2)

Related test

Quest’s comprehensive tick-borne disease test portfolio provides healthcare professionals with diagnostic tools to identify Lyme, tick-borne diseases, and coinfections as recommended based on geographic and symptom indicators.

Lyme Disease Antibody with Reflex to Immunoassay (IgG, IgM) (MTTT)

• Test code: 39733
• CPT^® code: 86618

Reflex tests are performed at an additional charge.

Note: The CPT codes provided are based on American Medical Association guidelines and are for informational purposes only. CPT coding is the sole responsibility of the billing party. Please direct any questions regarding coding to the payer being billed.

Next steps: Navigating treatment pathways of Lyme and other tick-borne diseases

The science of diagnosing and treating Lyme and other tick-borne diseases continues to evolve. These links provide current recommendations for laboratory support and other information.

Diagnosis and management of Lyme Disease

Diagnosis and management of Tick-Borne Disease

Lyme Disease FAQ

Supporting resources