The Traditional or Reverse Algorithm for Diagnosis of Syphilis: Pros and Cons

Abstract We reviewed relevant syphilis diagnostic literature to address the question “What diagnostic considerations should be taken into account when screening for syphilis using the traditional or reverse algorithm?” Improved laboratory diagnosis of syphilis is an important element of the effort to reduce syphilis rates. Screening for syphilis is performed using either a nontreponemal or treponemal test (part of the traditional or reverse algorithm, respectively). Both syphilis algorithms are used by laboratories. However, there are limited data on the performance and cost-effectiveness of the algorithms. An expert panel generated “key questions” in the laboratory diagnosis of syphilis. This paper pertains to the key factors that should be considered when deciding whether to screen for syphilis using either the traditional or the reverse algorithm. A systematic literature review was performed, and tables of evidence were created to address this question.

Screening for syphilis is performed using serological assays that detect treponemal and nontreponemal antibodies. The sequence in which these tests are performed differentiates the traditional from the reverse algorithm. In the traditional algorithm, a nontreponemal test (eg, rapid plasma reagin [RPR] or Venereal Disease Research Laboratory test [VDRL]) is used as the initial screen, and reactive samples are confirmed with a treponemal test. Conversely, the reverse algorithm uses a treponemal test for screening with reactive samples followed by a nontreponemal test. Discordant results between the treponemal screen and the nontreponemal test are resolved with a second confirmatory treponemal test (eg, Treponema pallidum particle agglutination) that preferably detects different antigens than the treponemal screen. Unfortunately, there is no gold standard for serologic syphilis testing, and therefore, all screening results must be correlated with clinical presentation for a diagnosis of syphilis.
According to a 2015 College of American Pathologists (CAP) survey, approximately 80% of laboratories perform the traditional algorithm and 20% perform the reverse algorithm when a single algorithm is offered at their facility [1]. A more recent review of CAP proficiency testing summary data showed a continuing trend toward increased use of the reverse algorithm. In 2019, 35.7% of CAP survey G (syphilis serology) participants reported use of a treponemal assay (authors' unpublished data). The algorithm utilized is primarily driven by the volume of syphilis testing [2,3]. Most nontreponemal tests are manual assays, so high-volume laboratories have chosen to adopt the reverse algorithm, due to availability of United States Food and Drug Administration (FDA)-cleared, automated treponemal platforms that perform high-throughput testing. Little guidance is provided to laboratories in choosing an algorithm, which is in part due to the limited data on the performance and cost-effectiveness of syphilis testing algorithms.
Improved laboratory diagnosis is crucial to curb the rise in syphilis infections. In 2016, the rate of reported syphilis in the United States reached an all-time high of 27.4 cases per 100 000 population, a > 2-fold increase since 2000 [4]. Increased incidence was seen across all stages of infection, and primarily attributed to men who have sex with men. However, all patient populations have been affected with increased rates reported in heterosexual men, women, and congenital infections. Despite improved laboratory methods, a syphilis diagnosis remains challenging to clinicians, especially with 2 algorithms currently recognized for diagnosis. To provide guidance on which algorithm to use, an expert panel reviewed the literature to determine the most effective approach based on laboratory workflow, cost-effectiveness, diagnostic accuracy, and public health impact.

METHODS
A literature review was conducted based on the key question of factors to consider when deciding to screen for syphilis using either the traditional or reverse algorithm. Medline, Embase, Cumulative Index to Nursing and Allied Health Literature (CINAHL), Cochrane, and Scopus databases from January 2000 to June 2017 were queried with the following search terms: "Treponema pallidum" or "neurosyphilis" or "syphilis" and "serodiagnose" or "serodiagnose" or "serology" and "test" or "exam" or "assay" or "screen" or "lab" or "diagnose" or "nontreponemal" or "treponemal" or "algorithm" or "antibodytiter" or "serofast" and yielded 4702 abstracts. Excluded from the search results were duplicate, animal model, non-English-language, non-FDA-cleared, and nonsyphilis abstracts resulting in 1851 total abstracts. The retrieved articles were then manually curated for potentially relevant papers using the terms "diagnosis, " "sero diagnosis, " "diagnostics, " "serology, " "serological test, " "treponemal, " "enzyme immunoassay, " "CLIA/CIA, " "IgG, " "IgM, " "TPPA, " "TPHA, " "nontreponemal, " "RPR, " "VDRL, " "laboratory screening, " or "algorithm" and yielded 251 abstracts. Meta-analysis, opinion, guidelines, letter to the editor, editorial, concepts, current trend/new trend, observations, guidance, reviews, non-FDA-approved tests, and international studies with non-FDA tests abstracts were excluded, finally giving 69 abstracts of which all full articles were pulled out and reviewed. Findings from the relevant article/data were summarized in form of a "table of evidence" for the key question. The data collected were presented to a group of syphilis experts convened in Atlanta, Georgia, 28-29 November 2017. Answers to the key questions were developed based on the tables of evidence, as well as expert opinion. Among the 69 articles in the "table of evidence, " a focus was placed on articles pertaining to the following: use of the traditional or reverse algorithm, evaluation of multiple FDA-approved assays, clearly stated gold standards (laboratory and/or clinical), signal strength-to-cutoff ratio, and cost-effectiveness.

RESULTS AND DISCUSSION
Both the traditional and reverse syphilis algorithms are used by laboratories today. Low-volume laboratories typically utilize the traditional algorithm due to the low cost of the manual nontreponemal tests. The lack of automated nontreponemal platforms-until very recently-makes it difficult for highvolume laboratories to provide adequate turnaround times. High-volume laboratories have instead opted to perform the reverse algorithm as automated treponemal assays that increase throughput are widely available and FDA-cleared. With a shortage of medical laboratory technicians, automated systems also reduce labor costs and provide an ergonomic benefit by eliminating repetitive pipetting steps of manual assays. Recently, 3 automated nontreponemal platforms have been FDA-cleared (AIX1000, Gold Standard Diagnostics, Davis, California; BioPlex 2200 syphilis total and RPR, Bio-Rad Laboratories, Hercules, California; and ASI Evolution, Arlington Scientific, Springville, Utah), but there are limited peer-reviewed data evaluating their performance. As data become available, more high-volume laboratories may continue to use the traditional algorithm depending on the diagnostic accuracy of these automated assays and test cost.
Today, laboratories are constantly faced with pressure to increase output at a reduced cost, so test cost has become an important consideration when choosing an algorithm. However, data supporting the cost-effectiveness of either algorithm are sparse and conflicting. In 2 cost-analysis studies, Owusu-Edusei et al found that the traditional algorithm was more cost-effective in a low-prevalence setting (0.5%) and generated more cost savings in a high-prevalence setting (10%), which was largely attributed to less confirmatory testing compared to the reverse algorithm (Table 1) [5,6]. Both analyses concluded that the amount of syphilis cases detected and treated was essentially the same when performing either algorithm in low-and highprevalence settings. In contrast, Chuck et al found that a treponemal screen and confirmation was more cost-effective when screening prenatal and nonprenatal patient populations, with prevalence rates of 0.076% and 1.94%, respectively (Table 1) [7]. The Chuck et al analysis took into account more healthcareassociated costs (nurses, clerical services, infectious disease consultation) with a substantially higher cost for congenital syphilis ($16 017 vs $760.36) and neurosyphilis ($77 149 vs not assessed). The cost savings generated from preventing congenital and neurosyphilis favor the reverse algorithm due to more correct diagnoses (51 517 vs 51 510 in the prenatal population; 38 035 vs 37 876 in the nonprenatal population). However, the Chuck et al report assumed a sensitivity rate of 70.6% for RPR across all stages of infection. These conflicting reports suggest that the reverse algorithm may be more costly, but the socioeconomic impact of missing a syphilis diagnosis needs to be determined/standardized for cost-analysis studies to be useful.
Laboratories also should consider their patient population and syphilis risk when performing the traditional and reverse algorithm. Missing a syphilis diagnosis can have devastating effects particularly in a prenatal population where congenital syphilis is a concern. In high-risk patient populations (eg, sexually transmitted disease [STD] clinic patients, people with human immunodeficiency virus, men who have sex with men), routine screening at 3-month intervals, regardless of the algorithm used, is the most effective approach to identifying cases of early syphilis [42][43][44][45]. Screening with a nontreponemal test in the traditional algorithm detects cases of active syphilis, but reports have shown decreased sensitivity of the RPR and VDRL at detecting cases of primary and possibly latent syphilis, although many studies fail to differentiate latent from past treated syphilis based on treponemal confirmatory testing [2,12,15,19,31]. Screening with a treponemal test will identify more cases of syphilis, presumably past treated, but additional confirmatory testing drives costs up [8,16,18,19,30,31,40]. However, the signal strength (a semiquantitative value) of automated treponemal screening assays can predict when a confirmatory    treponemal test will be reactive. Numerous studies have used a high signal strength (signal strength to cutoff ratio) in lieu of confirmatory treponemal testing to reduce unnecessary procedures or laboratory costs [11,26,27,32,36,38,39]. While each algorithm has a role depending on the patient population, these data suggest that a treponemal assay should be used to confirm a negative nontreponemal result if primary or latent syphilis is clinically suspected. Several studies have attempted to compare the traditional and reverse algorithm, but they lack direct comparison because a nontreponemal and treponemal screen were not run in parallel. Only a single high-quality study directly compared the traditional and reverse algorithm by prospectively testing 1000 patient samples in a low-prevalence patient population with both algorithms [12]. The reverse algorithm in this study produced 6 false-reactive results, while the traditional algorithm had none. The false-reactive rate of the reverse algorithm (0.6%) was also consistent with a previous Centers for Disease Control and Prevention (CDC) report of 0.6% [15]. However, the traditional algorithm may have missed 2 patients with latent syphilis, although the definitive diagnosis of these patients was not determined. Recently, the CDC Division of STD Prevention and the Association of Public Health Laboratories have collaborated to developed a syphilis serum repository comprised of laboratory characterized, residual syphilis specimens with reported stages derived from laboratory submission forms, as submitted to public health laboratories [46]. These syphilis specimens serve as a resource for validating existing or new syphilis diagnostic tests and hence support public health, commercial or clinical institutions in the United States. Future studies should compare both algorithms prospectively or use clinically characterized samples stratified by stage of infection to effectively evaluate the performance of syphilis testing algorithms.
Use of the traditional or reverse algorithm is ultimately institution dependent based on patient population, test cost, volume, and workflow. The traditional algorithm is well suited for smaller laboratories with a low-test volume since manual nontreponemal screening assays are typically less expensive and have minimal effect on workflow. On the other hand, the reverse algorithm may be more appropriate for smaller laboratories serving a high-risk population, such as an STD clinic, where patients are more likely to be at risk for primary and latent syphilis that may be missed by the traditional algorithm. In larger laboratories, automated platforms improve workflow efficiency and provide a better turnaround time. Current data support the use of automated treponemal assays for screening, but this may change as more studies with automated nontreponemal assays become available. Regardless of the syphilis testing algorithm used, laboratory results should correlate clinically with patients' symptoms and risk to make an accurate diagnosis. The laboratory can aid in the clinical decision process by collectively reporting all the laboratory results in a composite report that includes the algorithm, test methods, and interpretation.
Notes Conflicts that the editors consider relevant to the content of the manuscript have been disclosed. Owusu-Edusei 2011 [5] Cohort decision analysis model to estimate cost and health outcomes of traditional and reverse algorithms N = 10 000 0.5% and 10% prevalence NA Low-prevalence setting: Reverse algorithm led to ≈2 times the number of confirmatory tests and was only cost-effective when the test was < $5.80. Traditional algorithm more cost-effective per adverse outcome ($1400 vs $1500). High-prevalence setting: Reverse algorithm led to ≈3 times the number of confirmatory tests and was only cost-effective when the test was < $1.80. Both algorithms detected the same number of syphilis cases in low-and high-prevalence settings. Reverse algorithm leads to overtreatment of uninfected patients.