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Syphilis testing

16 min

Skills you'll build3 skills
  • Choose the next test for a reactive treponemal screen with a nonreactive RPR
  • Call a fourfold RPR or VDRL titer change only between results from one method
  • Report a negative CSF VDRL without excluding neurosyphilis

At the bench: Follow the laboratory algorithm for discordant syphilis results

Try first

Try first

In a reverse-sequence algorithm, a patient's automated treponemal immunoassay is reactive and the rapid plasma reagin (RPR) test is nonreactive. What can this pattern mean?

Keep going to see the reasoning.

Right. Keep going to follow the reasoning.

Keep going to see the reasoning.

Keep going to see the reasoning.

Keep going to see the reasoning.

Get the idea

Use the explanation to check your first decision.

Nontreponemal and treponemal tests

Nontreponemal tests, the RPR and the Venereal Disease Research Laboratory (VDRL) test, detect antibody to cardiolipin, lecithin and cholesterol. Their titers rise within 1 to 4 weeks of the chancre, peak in secondary syphilis and fall after treatment. Every reactive result is reported as an endpoint titer, the highest dilution that still reacts.1

Treponemal tests, such as automated enzyme or chemiluminescence immunoassays and the Treponema pallidum particle agglutination (TP-PA) test, detect antibody to treponemal proteins. They usually stay reactive for life, so they show exposure and cannot monitor treatment response.1

Two algorithms

The traditional sequence starts with an RPR or VDRL and confirms a reactive result with a treponemal test. The reverse sequence starts with a treponemal immunoassay and reflexes a reactive screen to a quantitative RPR.1 The two sequences agree in about 99% of specimens. Cost, volume, staffing and the population served determine which algorithm a laboratory uses.1

Treponemal screenRPRSecond treponemal testInterpretation
NonreactiveNot doneNot doneSyphilis unlikely
ReactiveReactiveNot donePreviously treated or untreated syphilis
ReactiveNonreactiveReactivePast or current syphilis
ReactiveNonreactiveNonreactiveLikely a false-positive screen, when clinical and exposure probability is low

1,2

The second treponemal test uses a different format and different antigens, preferably TP-PA. The treatment and exposure history then determine how a reactive pattern is interpreted.1

Comparing titers

A fourfold nontreponemal titer change is two twofold dilution steps, such as 1:8 to 1:32 or 1:8 to 1:2. Serial titers use the same test method and specimen type, because RPR and VDRL titers differ. A VDRL titer cannot be compared with an RPR titer to count dilution steps.1

CSF VDRL

The VDRL is the only FDA-cleared test for cerebrospinal fluid (CSF). A reactive CSF VDRL is highly specific. Its sensitivity ranges from 13% to 87% depending on the reference standard, so a nonreactive result does not exclude neurosyphilis when CSF pleocytosis or a raised protein supports it.1

Follow the bench demonstration

Sources for this step
References
  1. Papp JR, Park IU, Fakile Y, Pereira L, Pillay A, Bolan GA. CDC laboratory recommendations for syphilis testing, United States, 2024. MMWR Recomm Rep. 2024;73(1):1-32. doi:10.15585/mmwr.rr7301a1
  2. Centers for Disease Control and Prevention. Syphilis. STI Treatment Guidelines. Accessed September 27, 2026. https://www.cdc.gov/std/treatment-guidelines/syphilis.htm

Watch one

Read each decision and its explanation.

Lionel Triplett, 45, was treated for secondary syphilis 6 months ago. His serum nontreponemal results:

  • At treatment: RPR 1:64, on serum in this laboratory.
  • Three months later: VDRL 1:8, on serum at an urgent care laboratory.
  • Today: RPR 1:16, on serum in this laboratory.

Has his nontreponemal titer fallen fourfold since treatment?

  1. Check each result's method and specimen. The two RPRs are on serum in the same laboratory.

    Titers are compared only between results from one method and one specimen type.

  2. Set the VDRL 1:8 aside. It cannot be included in the RPR series.

    RPR and VDRL titers differ on the same specimen.

  3. Count the steps between the RPRs: 1:64 to 1:32 to 1:16 is two steps.

    Each twofold dilution step halves the titer, and a fourfold change is two steps.

  4. 64 ÷ 16 = 4, a fourfold decline.

    The ratio of the reciprocal titers gives the size of the change.

Yes. His RPR fell from 1:64 to 1:16, two dilution steps and a fourfold decline. The VDRL result is not part of the comparison.

Practice this skill

Your turn

Problem 1 of 3 · Hints available

A reverse-sequence treponemal screen is reactive and the quantitative RPR is nonreactive. Which test resolves this discordance?

Incorrect. Repeating the original immunoassay reproduces its reactivity, including a false-positive screen. Resolving the discordance needs a treponemal test with a different format and antigens.

Incorrect. The 2- to 4-week repeat RPR follows a reactive second treponemal test in a previously treated person with possible reexposure. Resolve this discordance first with a different treponemal test.

Correct. A treponemal test with a different format and antigens separates a false-positive screen from true antibody in treated past infection or early primary disease. The combined results are then read with treatment and exposure history.

Hint
  1. Repeating the same test reproduces the cause of the reaction.
  2. Ask which kind of test could confirm the treponemal antibody.

Review Testing algorithms

Problem 2 of 3 · Hints available

Comparable rapid plasma reagin (RPR) endpoint titers change from 1:8 to 1:32 using the same method and specimen type. What is the magnitude of the increase?

Correct. 32 ÷ 8 = 4, reached in two twofold steps (1:8 to 1:16 to 1:32). A fourfold change between results from the same test and specimen type is clinically significant.

Incorrect. One twofold step from 1:8 reaches 1:16, and a second step reaches 1:32.

Incorrect. The three titers 1:8, 1:16, and 1:32 span two twofold steps, and 32 ÷ 8 = 4.

Hint
  1. Write out each twofold dilution between 1:8 and 1:32.
  2. Divide the larger reciprocal titer by the smaller one.

Review Nontreponemal (lipoidal antigen) tests

Problem 3 of 3 · On your own

A man with reactive serum syphilis tests has new confusion. His CSF VDRL is nonreactive. The CSF white cell count is 48 cells/µL (reference interval 0 to 5 cells/µL) and the CSF protein is 92 mg/dL (reference interval 15 to 45 mg/dL). How is the CSF VDRL reported?

The CSF VDRL misses many cases. His raised cell count and protein can still support neurosyphilis.

Excluded neurosyphilis with a negative CSF VDRL

The CSF VDRL is highly specific but insensitive, with reported sensitivity from 13% to 87%, so a nonreactive result can accompany neurosyphilis. Reporting neurosyphilis as excluded on that result ignores the CSF cell count and protein, which can support the diagnosis.

The result is released as tested. The comment tells the provider that the raised cell count and protein can still support neurosyphilis.

The VDRL is the FDA-cleared nontreponemal test for CSF. An RPR on CSF would not resolve the question.

Serum and CSF tests are interpreted separately. The CSF result is valid and is released.

Review Neurosyphilis and congenital syphilis

Use it

Read the case evidence, then make each decision. The step is complete without it.

  • Rosa Dillon, 36, was treated for secondary syphilis 3 years ago. Her record documents the treatment.
  • Your laboratory uses the reverse sequence. Today's serum gives a reactive treponemal chemiluminescence immunoassay and a reactive RPR at 1:16.
  • Her last nontreponemal result, 1 year ago at another laboratory, was a serum VDRL of 1:4.
Decision 1 of 2

What does the reactive treponemal immunoassay add?

Treponemal antibody usually stays for life after treatment. Her documented treatment explains it.

Read a reactive treponemal screen as untreated syphilis

Treponemal antibody usually persists after treatment, and a reactive screen with a nonreactive RPR can mean a treated past infection, early primary disease, or a false-positive screen. Reporting untreated syphilis skips the second treponemal test, such as TP-PA, that distinguishes among them.

In a person with treated syphilis, treponemal results stay reactive. The quantitative RPR and the history show whether infection is active.

A second treponemal test resolves a reactive screen with a nonreactive RPR. After documented treatment, more treponemal testing cannot detect reinfection.

Review Testing algorithms

Decision 2 of 2

The provider asks whether 1:16 is a fourfold rise from last year's 1:4. What do you tell the provider?

The earlier titer is a VDRL and today's is an RPR. Their titers differ on the same specimen, so the steps cannot be counted across them.

Compared an RPR titer directly with a VDRL titer

RPR and VDRL titers differ for the same specimen, so a VDRL of 1:8 followed by an RPR of 1:32 cannot be read as a fourfold rise. Comparing across methods can report a significant change, or miss one, that the same test would show correctly.

From 1:4 to 1:16 would be two steps. The two titers come from different methods, so no change can be calculated from them.

A fourfold change is assessed only between results from one method and specimen type. Later RPRs on serum can be compared with today's 1:16.

Review Nontreponemal (lipoidal antigen) tests

The finding that matters is the method used for last year's titer. It was a VDRL, so today's RPR of 1:16 cannot be compared with it.

For reference

Sources checked