Diagnostic potential for IgM antibody detection by the DPP Syphilis TnT assay in neonates at risk for congenital syphilis

ABSTRACT

Neonatal IgM antibodies reflect an in utero immune response to Treponema pallidum and may offer added diagnostic value. This study evaluated the test performance of treponemal IgM levels measured by the research-use-only (RUO) Dual Path Platform (DPP) Syphilis TnT point-of-care (POC) assay for congenital syphilis (CS) risk stratification. Conducted from May 2023 to May 2025, this study tested neonatal serum samples from infants born to mothers with syphilis using the DPP Syphilis TnT RUO POC assay, which reports treponemal and nontreponemal IgM levels as relative light units (RLU). Neonates were classified as confirmed proven or highly probable CS, possible CS, or CS less likely per guidelines; 23 neonates without maternal syphilis served as controls. Treponemal IgM levels were compared across categories using nonparametric tests and ordinal logistic regression. Diagnostic performance used prespecified cutoffs, with agreement assessed against neonatal rapid plasma reagin (RPR). Twenty-two maternal-neonatal dyads were included. Mean treponemal IgM levels rose with CS severity, peaking in the high-risk group (possible or confirmed proven/highly probable CS: 29.9 ± 20.6 RLU) versus CS less likely (17.5 ± 20.8 RLU) and controls (3.5 ± 0.8 RLU; P < 0.05). Higher IgM levels independently linked to elevated CS risk (OR 1.10 per 1 RLU; P = 0.0025). At ≥10 RLU cutoff, treponemal IgM detected 88.9% of high-risk neonates, with 76% agreement to neonatal RPR. The DPP Syphilis TnT RUO POC assay’s treponemal IgM levels discriminated CS risk categories effectively and may supplement current algorithms to improve neonatal CS stratification.

IMPORTANCE

Congenital syphilis (CS) continues to rise in the United States and globally, yet diagnosis at birth remains difficult because no single laboratory test definitively confirms infection in newborns. Clinical decisions often rely on maternal history and indirect serologic comparisons, which can result in both missed cases and unnecessary treatment of low-risk infants. IgM antibodies are produced by the fetus in response to infection in utero and therefore represent a biologically meaningful marker of congenital infection. This study evaluates the diagnostic potential of the Dual Path Platform Syphilis TnT research-use-only point-of-care assay to detect treponemal IgM in at-risk neonates. We demonstrate that IgM levels increase across CS less likely, possible CS, and confirmed proven or highly probable CS categories and are independently associated with disease risk. These findings provide early evidence that neonatal IgM testing may improve risk stratification and support more precise clinical decision-making in CS management.

INTRODUCTION

According to 2022 World Health Organization (WHO) global surveillance data, the estimated number of congenital syphilis (CS) cases is increasing, with over 700,000 cases estimated worldwide, exceeding the WHO global mother-to-child transmission (MTCT) threshold by over 10-fold (1). The adverse birth outcomes related to maternal and CS are profound, with over 390,000 adverse birth outcomes reported in 2022, including 150,000 fetal deaths resulting from untreated maternal infections worldwide (1–4). For infected infants born alive, the estimated cost of neonatal intensive care for one newborn with CS in the United States exceeds $55,000, excluding quality of life or activities-of-daily-living expenses for affected families (5). Although guidance for adult infection is well established and evidence based (6, 7), the current diagnostic strategies for neonatal infection rely on a risk-based algorithm dependent on maternal diagnosis and treatment history, exam findings, placental evaluation, and comparative levels of maternal and neonatal nontreponemal (NT) antibodies detected by Venereal Disease Research Laboratory (VDRL) or rapid plasma reagin (RPR) tests at delivery (6, 7). All these individual criteria lack strong test performance and, when taken together, are used to estimate scenario-based CS risk categories to inform treatment strategies (e.g., confirmed proven or highly probable CS, possible CS, CS less likely, and CS unlikely), according to the current Centers for Disease Control and Prevention (CDC) sexually transmitted infection (STI) treatment guidelines (6, 7). Results of RPR testing are often challenging to interpret due to maternal IgG antibody interference, therefore necessitating between 2 and 15 months of lengthy follow-up with RPR monitoring to ensure clearance of maternal IgG antibody and absence of congenital infection (6–8). The NT serologic assays require venipuncture, specialized equipment, and off-site moderately complex laboratory settings, which often lead to incomplete care cycles with poor follow-up for results management or treatment (6, 7). In contrast, point-of-care (POC) tests like the Food and Drug Administration (FDA)-cleared Syphilis Health Check or Dual Path Platform (DPP) HIV-Syphilis Assay enable decentralized testing in clinics, emergency departments, shelters, and other nonclinical environments by trained staff (9–11). These are considered first-line options for adult syphilis testing in resource-limited global settings, given the advantage of a 15–20-minute turnaround time from testing to results, which can facilitate same-day treatment for early infection, prevent MTCT, and potentially cure fetal disease when used in antenatal screening (9–11).

It has been demonstrated that IgM antibodies detectable in neonates at risk for CS are indicative of a fetal immune response to active Treponema pallidum infection in utero (12). While IgM-specific assays for CS have demonstrated sensitivities of 83%–100% in symptomatic cases and specificities up to 100%, their overall performance varies (12–15). This reported variability of diagnostic accuracy can be explained by antitreponemal testing performed prior to seroconversion in cases of early CS or after the clearance of antibodies following treatment for early-stage fetal infection. Additionally, there is no gold-standard comparator or FDA-cleared antitreponemal IgM test available in the United States (12–15). Given these limitations, the use of a treponemal IgM test for CS diagnosis has not been adopted in the United States or recommended by the CDC or WHO (1, 7). Further validation is needed for POC applications in neonates to account for maternal IgG interference to potentially optimize clinical algorithms using a rapid treponemal IgM test.

The objective of this pilot study was to assess the diagnostic value of IgM responses detected by the Chembio DPP Syphilis TnT research-use-only (RUO) point-of-care assay in neonates categorized at birth into various CS risk scenarios, per the 2021 CDC STI treatment guidelines (7). The neonatal IgM antibody levels and reactivity rates across the risk groups were also analyzed in comparison with the newborn RPR test results used routinely for CS diagnosis.

MATERIALS AND METHODS

The study was conducted at the University of Texas Health Science Center—obstetric tertiary care center in Houston, Texas, from May 2023 to May 2025. Pregnant individuals aged 14–45 with a confirmed live intrauterine pregnancy at greater than 16 weeks gestational age, presenting for routine prenatal care, and with a diagnosis of syphilis during the index pregnancy were eligible for enrollment. The diagnosis of syphilis was made using the laboratory-based reverse syphilis testing algorithm, which involves a treponemal test, such as an enzyme immunoassay (EIA), followed by RPR test for confirmation of active infection. A secondary treponemal test, T. pallidum particle agglutination, was used when the reverse algorithm produced discrepant results (7). All patients underwent a detailed health history and physical exam for the staging of syphilis and were treated with benzathine penicillin G according to stage (primary, secondary, early latent, and late latent) per the CDC STI treatment guidelines (7). Infection status was confirmed with the regional health department. Exclusion criteria included pregnancies with syphilis treated prior to the pregnancy or if pregnancy was complicated by a fetal demise (7). All consented participants underwent repeat testing at 28 weeks and delivery according to the Texas Health and Safety Code 81.090 unless more frequent testing was warranted. All pregnancies were managed according to the standard of care, regardless of participation in the study. All maternal and neonatal infection-related data were recorded.

The study population consisted of 45 newborns, including 22 neonates at risk for CS (born to mothers diagnosed with syphilis during pregnancy) and 23 no-risk neonates (no exposure to maternal syphilis) who were used a negative control group. Within 48 hours of delivery, 50–75 μL of neonatal serum was collected and stored frozen at −70°C until testing by the DPP Syphilis TnT RUO POC Assay (Chembio Diagnostics, Inc., Medford, NY) and Trep-Sure Syphilis Total Antibody EIA (Trinity Biotech, Jamestown, NY).

The Chembio DPP Syphilis TnT Assay, originally developed for syphilis testing in adults, is designed to have four separate test lines for differential semi-quantitative detection of IgM and IgG antibodies to treponemal and NT antigens. The treponemal test lines use the same proprietary multiepitope recombinant protein as employed in the DPP HIV-Syphilis Assay product (PMA approved, BP180191;10/2020), whereas the NT test lines use the CDC-licensed VDRL antigen with in-house improved liposomal formulation. To run the assay, 10 μL of serum was mixed with five drops (150 μL) of DPP Syphilis TnT Running Buffer in the test kit provided sample vial. A 125 uL portion of this sample mixture was transferred to test device Well #1 using the transfer pipette provided in the test kit. After waiting for 5 minutes, 10 drops (300 μL) of DPP Syphilis TnT Running Buffer were added to test device Well #2. Results were interpreted using the Chembio DPP Micro Reader II at 15 minutes after the running buffer was added. The DPP Micro Reader II was configured to report a qualitative result for neonatal IgM results interpreted as reactive or nonreactive based on pre-established cutoff values of 10 relative light units (RLU) for each test antigen and the level for each antibody class (IgM and IgG) for treponemal and NT antibodies as independent outputs, expressed as RLU. All results were recorded for analysis.

At birth, neonatal providers assigned the clinical CS risk scenario following the CDC guidelines and American Academy of Pediatrics standards (7, 16) prior to experimental testing. All aliquots were run in duplicates, and the investigators were blinded to the neonatal CS risk category assigned at birth. The protocol was approved by the Institutional Review Board of the McGovern Medical School (HSC-MS-25-0170). All participants provided written informed consent to participate in the research study.

Descriptive statistics were used to summarize participant data comparing all at-risk categories (confirmed proven or highly probable CS, possible CS, and CS less likely) and the negative controls. For data analysis, the possible CS and confirmed proven or highly probable CS results were combined in one group, given their relatively low numbers and high-risk status (7). Summary statistics (median and range) were calculated for maternal and neonatal RPR test results. Kruskal-Wallis tests were used to evaluate differences in treponemal and NT mean IgM levels detected by the Chembio DPP Syphilis TnT Assay across CS categories. Ordinal regression was performed to determine the relationship between treponemal IgM values and CS risk category. Pairwise Mann-Whitney U tests with Bonferroni correction were performed for post hoc contrasts. Receiver operating characteristic (ROC) analysis was used to evaluate the diagnostic performance of treponemal and NT IgM individually. A composite IgM predictor was constructed using multivariable logistic regression incorporating both, and ROC analysis of the resulting score was performed to assess incremental discrimination. Diagnostic performance was assessed using 2 × 2 contingency tables, with sensitivity and specificity calculated for treponemal IgM cutoffs. Given the lack of a diagnostic gold-standard test for CS, the clinical CDC diagnostic criteria were used as the comparator for analysis, and the possible or confirmed proven or highly probable CS group was considered test-positive as described above. Exact 95% confidence intervals were estimated using the Wilson method. Agreement between neonatal treponemal IgM and RPR test results at delivery was evaluated using categorical agreement analysis. Overall percent agreement was calculated, and agreement beyond chance was quantified using Cohen’s kappa (κ). All statistical analysis was performed using STATA software, version 17 (College Station, TX).

RESULTS

From May 2023 to May 2025, a total of 22 maternal-neonatal dyads met inclusion criteria and had complete clinical data along with available serum for both experimental assays. These at-risk neonates were classified as confirmed proven or highly probable CS (n = 6), possible CS (n = 3), and CS less likely (n = 13). Table 1 demonstrates baseline infection-related characteristics of the enrolled maternal-neonatal dyads. Most of the infected women were diagnosed and treated for syphilis between 16 and 26 weeks gestational age (n = 14, 64%) and delivered at term (n = 17, 77%). Twenty-three neonates without exposure to maternal syphilis were used as negative controls.

TABLE 1.

Maternal and neonatal syphilis characteristics and RPR titers at delivery^a

Maternal information
Maternal syphilis stage	N	Median RPR titer	RPR titer range
Early syphilis	3	16	4–128
Late/late latent	19	2	NR–32

^a NR = nonreactive.

Neonatal treponemal IgM antibodies were detected by the Chembio DPP Syphilis TnT Assay in all at-risk categories, with the highest mean levels observed in the high-risk CS group (possible CS and confirmed proven or highly probable CS combined) (29.9 ± 20.6) compared to the CS less likely cohort (17.5 ± 20.8) or the negative controls (3.5 ± 0.8) (P < 0.05) (Table 2). NT IgM also differed significantly across CS categories but with less pronounced differences. Figure 1 displays treponemal and NT IgM values according to CS category.

TABLE 2.

Treponemal and nontreponemal IgM levels by congenital syphilis category (possible and confirmed proven or highly probable CS combined)^a

CS category	n	Treponemal IgM mean (SD)	Treponemal IgM median	Treponemal IgM min-max	NT IgM mean (SD)	NT IgM median	NT IgM min-max
Negative control	23	3.5 (0.8)	3	3–6	2.1 (0.8)	2	1–4
CS less likely	13	17.5 (20.8)	12	3–79	5.2 (7.8)	2	1–27
Possible/confirmed/highly probable CS	9	29.9 (20.6)	25	5–72	6.0 (3.2)	5	2–11

^a Possible and confirmed proven or highly probable CS categories were combined for analysis. Treponemal IgM and NT IgM levels are reported as relative light units.

Fig 1.

Box plots display neonatal treponemal and nontreponemal IgM levels, expressed as relative light units, stratified by congenital syphilis category (control, CS less likely, and possible/confirmed/highly probable CS). The possible and highly probable/confirmed CS categories were combined for analysis. Boxes represent the interquartile range (IQR), center lines indicate medians, and whiskers extend to 1.5× the IQR. Mean values with SD are shown above each category. Differences across categories were assessed using the Kruskal-Wallis test.

In ordinal logistic regression including controls, higher mean treponemal IgM levels were significantly associated with increasing CS risk category (OR 1.10 per 1 RLU, 95% CI 1.04–1.18; P = 0.0025), corresponding to an OR of 1.64 per 5 RLU (95% CI 1.19–2.26).

Treponemal IgM was the primary driver of diagnostic sensitivity for CS detection. Using a cutoff of ≥10 RLU, the treponemal IgM-positive results obtained with the Chembio DPP Syphilis TnT Assay identified 8/9 (88.9%) neonates in the high-risk groups and 8/13 (61.5%) newborns in the CS less likely scenario. Risk stratification among high-risk neonates (possible CS and confirmed proven or highly probable CS) could be enhanced by applying a higher treponemal IgM cutoff of ≥19 RLU. At this threshold, test specificity increased to 94.4%, with a corresponding reduction in sensitivity to 77.8%, potentially resulting in two missed high-risk CS cases. Neonates classified as CS less likely were included in the negative group for diagnostic performance analyses; however, these infants represent an exposed population, and misclassification cannot be fully excluded.

In contrast, NT IgM demonstrated significantly lower seroreactivity rates and weaker differentiation between the risk categories. Using a cutoff ≥10 RLU, this biomarker detected 2/9 (22%) high-risk neonates, 2/13 (15.4%) low-risk neonates, and none among 23 no-risk neonates. The four newborns with NT IgM antibodies found were also RPR reactive (titers ranging from 1:1 to 1:4) and positive for treponemal IgM antibodies detected by the Chembio DPP Syphilis TnT Assay at relatively high levels (50.5 ± 11). Although a combined treponemal and NT IgM metric improved overall discrimination as assessed by ROC analysis, it did not enhance the test sensitivity beyond treponemal IgM alone. Additionally, treponemal IgG levels did not significantly differ across CS risk categories, whereas NT IgG levels showed statistical differences with substantial overlap between groups, likely reflecting higher passively transferred maternal antibody levels among higher-risk dyads. Because neonatal IgG may represent maternal antibody rather than fetal immune activation, IgG measures were not pursued for risk stratification analyses.

Among the 22 at-risk neonates, 15 tested RPR reactive at birth, of which 13 (86.7%) had treponemal IgM antibodies. In contrast, out of the remaining seven RPR nonreactive newborns, only three (42.9%) produced this serologic biomarker. When the treponemal IgM results were stratified by RPR titers, the IgM component of Chembio DPP Syphilis TnT Assay detected all at-risk neonates (5/5) with RPR titers ≥1:4, as compared to lower treponemal IgM reactivity rate of 80% (8/10) found in the low-titer subgroup (≤1:2). At the 10 RLU cutoff, categorical agreement between treponemal IgM positivity and neonatal NT positivity at delivery was 76%, with moderate agreement beyond chance (Cohen’s κ = 0.44), indicating concordance with the RPR titers used for clinical management. All syphilis-exposed neonatal samples were positive by the Trep-Sure Syphilis Total EIA test, confirming the presence of maternal IgG or both isotypes of treponemal antibody in neonatal serum.

Seven of 12 (58%) CS less likely neonates received a single dose of benzathine penicillin G, and 5 (42%) were managed with follow-up alone. All remained disease-free on subsequent serologic and clinical evaluation.

DISCUSSION

In this pilot study of neonates at risk for syphilis infection, treponemal IgM reactivity rates and levels measured using the Chembio DPP Syphilis TnT Assay increased across clinical CS categories. Mean RLU values were significantly higher among neonates classified as possible CS and confirmed proven or highly probable CS compared with the negative controls (~8.5-fold) and with those classified as CS less likely (~1.7). This was also demonstrated in ordinal regression analysis, where each one-unit increase in RLU conveyed an increased level of CS risk. Median values and overall distributional shifts also demonstrated progressively higher treponemal IgM levels with increasing CS severity. Minor nonmonotonicity in mean values was observed, attributable to small sample sizes and greater variability within the high-risk CS group.

NT IgM levels also differed across CS categories, although less significantly if compared with treponemal IgM. Overall, treponemal IgM demonstrated superior discrimination across CS risk categories and aligned with laboratory-based RPR test performed at delivery, supporting its potential utility for neonatal CS risk stratification.

Two neonates had notable clinical contexts. One neonate classified as possible CS was born to a mother with systemic lupus erythematosus and demonstrated a treponemal IgM response within the range observed for this category. Another neonate classified as CS less likely was exposed to antenatal corticosteroids for fetal lung maturation approximately 2 weeks prior to delivery and exhibited a relatively high treponemal IgM level (72 RLU). These observations suggest that maternal autoimmune disease or exposure to antenatal corticosteroids did not preclude development of treponemal IgM responses in this neonatal cohort.

For the one neonate classified as high risk for CS with a nonreactive treponemal IgM, the RPR titer was 1:4, and the infant exhibited clinical findings consistent with CS, and treatment was initiated based on clinical assessment. This case highlights the important point that although a reactive treponemal IgM strongly supports congenital infection, a nonreactive IgM does not exclude it. A negative result may reflect early infection in which IgM antibody levels have not yet reached detectable thresholds, a recognized limitation of IgM-based assays. Accordingly, IgM testing should be interpreted as an adjunct to clinical and serologic assessment rather than a standalone diagnostic tool. In clinical practice, the DPP TnT assay may be particularly informative in asymptomatic neonates with diagnostic uncertainty, where additional objective evidence could meaningfully support management decisions. In contrast, neonates with clear clinical findings consistent with CS often present less diagnostic ambiguity.

WHO recommends that all infants born to women diagnosed with syphilis during pregnancy receive effective treatment, even if asymptomatic at birth. While a universal “treat all” approach appropriately prioritizes prevention of missed CS, it also reflects persistent diagnostic uncertainty at delivery and underscores the need for improved tools to refine neonatal risk assessment. Similar uncertainty exists in high-resource settings, where low-risk infants may undergo unnecessary hospitalization, parenteral penicillin therapy, and extensive evaluation, which carry both tremendous clinical and economic implications. In addition, inconsistent availability of benzathine penicillin further highlights the importance of judicious antimicrobial use. A rapid assay enabling more precise risk assessment may therefore provide meaningful clinical and economic value.

Importantly, the DPP TnT device provides both a qualitative result and a quantified RLU value simultaneously via the handheld reader, without additional user steps. Thus, while this study examined alternative thresholds for risk stratification, the assay itself remains simple to perform and interpret in clinical practice. Future validation studies will help determine whether a single optimized cutoff or tiered interpretation strategy is most appropriate for routine use.

The DPP Syphilis TnT POC assay is approved for use with finger-prick whole blood, as well as serum and plasma. Although testing was performed on batched frozen serum in the current study, future studies will include evaluation of performance across different specimen types, including capillary whole blood, to assess concordance and operational feasibility in real-world clinical settings.

A treponemal IgM cutoff of ≥10 RLU provided optimal sensitivity (89%), prioritizing detection of all at-risk neonates while minimizing missed cases. These findings are particularly relevant in clinical and public health settings, both globally and within the United States, when maternal infection history, treatment status, or related information may be incomplete, unavailable, or unreliable at the time of delivery. In such contexts, neonatal biomarkers that enable sensitive, objective risk assessment independent of maternal history are critical for timely identification and management of CS. In contrast to lower IgM thresholds optimized for screening, a treponemal IgM value ≥19 RLU may have particular utility as a risk stratification marker. At this threshold, specificity was high (94.4%), suggesting that neonates with IgM levels above this cutoff represent a population at especially high risk for CS. Although sensitivity is reduced, such values may support clinical decisions regarding escalation of evaluation or prioritization for close follow-up, particularly when clinical RPR titers are equivocal.

The serologic response to T. pallidum begins with the development of treponemal IgM antibodies, which emerge earlier than the NT response in fetal, neonatal, and adult infection (12–15). Because IgM is produced in primary or recent infection and does not always persist in late-stage disease, it provides valuable temporal information that IgG cannot. In contrast, the RPR test used for clinical diagnosis and monitoring detects reagin antibodies of both IgG and IgM classes against lipoid (cardiolipin-lecithin-cholesterol) antigen (7, 8). The RPR test reports a total antibody response and, therefore, cannot distinguish between immunoglobulin isotypes or differentiate maternal IgG transferred to fetus from neonatal IgM production. As a result, comparative RPR titers may substantially underestimate early neonatal infection (12). The clinical neonatal RPR results produced in the present study were uniformly of low titers across all CS categories and were essentially nondiagnostic. To meet the CDC-recommended diagnostic criteria for CS, an RPR titer in neonates must be at least fourfold higher than the maternal RPR titer at delivery (7). However, this approach has poor clinical sensitivity as most infected neonates have equivocal RPR titers (12). Most at-risk infants in the present study, including some classified as confirmed proven or highly probable CS by the clinical care team, demonstrated no or very low RPR titers. In our cohort, only one neonate considered confirmed proven or highly probable CS had an RPR titer fourfold higher than the maternal titer at delivery. In contrast, treponemal IgM values showed clear stratification across the CS risk groups and remained elevated even when neonatal RPR titers were low.

There are commercial IgM immunoblot and enzyme-linked immunosorbent assay (ELISA) assays used in Europe (e.g., ViraMed and Euroimmun, Germany), all of which are Conformité Européenne (CE) marked and therefore meet regulatory standards for in vitro diagnostics in the European market. Published data show that these CE-marked IgM immunoblot assays (ViraMed, Euroimmun) demonstrate strong diagnostic accuracy, with reported sensitivities ranging from 90.0% to 94.4% and specificities between 97.1% and 99.2% when evaluated against composite or reference standards in infants and neonates with suspected CS (13–15, 17–19). In contrast, the CE-marked Euroimmun IgM ELISA typically shows more modest sensitivity (≈60%) despite specificity approaching 100%. Historical studies, including those using the 19S IgM-Flourescent treponemal antibody-absorption (FTA-ABS) assay, similarly demonstrated that IgM-based methods may offer meaningful diagnostic value in CS (13–15, 18, 19). More recent work, including a 2025 study evaluating a different experimental lateral-flow POC IgM test, suggests that newer technologies may achieve high diagnostic performance with sensitivities in the 88%–92% range and specificities of 96%–98% when compared to a composite reference comparator and clinical CS categories (17, 20).

Despite encouraging performance from CE-marked assays internationally and promising early US data using experimental point-of-care antitreponemal IgM tests, no treponemal-IgM assay is currently FDA cleared for CS diagnostics in the United States. Consistent with this, routine IgM testing is not recommended by the CDC and WHO because of variable accuracy, limited clinical validation, and inconsistent correlation with disease stage (7). These considerations underscore the critical need for improved neonatal-specific diagnostic tools, particularly given that many infected infants are asymptomatic at birth, traditional RPR titers are often nondiagnostic, and that long-term serologic follow-up is difficult to achieve for many families (6, 7). More accurate, rapidly deployable assays, such as the Chembio DPP Syphilis TnT Assay evaluated in the present study, would therefore represent a major advance for timely identification and management of CS.

Strengths

This pilot study has several important strengths. First, it includes a well-characterized cohort of maternal-neonatal dyads with confirmed syphilis infection, with maternal staging performed according to current CDC STI treatment guidelines by clinicians and nurses with established expertise in syphilis management. Second, all neonatal specimens were collected within 48 hours of delivery and processed within 1 hour, ensuring high sample integrity. Third, the Chembio DPP Syphilis TnT Assay was performed by research staff who were blinded to neonatal CS categories, which were independently assigned by pediatric clinicians with expertise in congenital infections.

Weaknesses

This study has limitations. The sample size was modest (n = 21 neonates and 23 controls) and the study period relatively brief, limiting generalizability. However, the consistency of findings, statistically significant differences across CS categories, and rigorous methodology provide confidence in the internal validity of the results. Neonates born to mothers adequately treated prior to pregnancy or those with other maternal infections or inflammatory conditions were not included. Inclusion of these groups would allow for more precise evaluation of assay specificity in real-world clinical contexts. Future studies will incorporate these important populations.

Given the ongoing syphilis epidemic and the urgent clinical need for improved CS diagnostics, dissemination of these preliminary findings remains both relevant and necessary to guide further research and test development (1, 7, 8).

Conclusion

Our findings demonstrate that the detection of treponemal IgM antibodies by the Chembio DPP Syphilis TnT Assay in neonates may have an added diagnostic value for CS risk assessment. The potential integration of a rapid POC treponemal IgM test into the neonatal CS diagnostic algorithm would be paradigm shifting for families and clinicians, particularly given the longstanding absence of a reliable gold-standard diagnostic test for an infection associated with significant neonatal morbidity and mortality.