Abstract
Abstract
Objective
The study was conducted to assess the diagnostic performance of the Hightop Syphilis Rapid Diagnostic Test (RDT) in comparison with the ELISA test used as a reference method.
Design
A laboratory-based cross-sectional and comparative study was conducted to assess the diagnostic performance of the Hightop Syphilis RDT.
Setting
Blood samples obtained from adult participants in eight health facilities were analysed at the National Public Health Laboratory (NPHL), Ministry of Public Health, Yaounde, Cameroon.
Participants
From 29 April to 25 August 2023, 583 adult participants of both sexes (aged ≥21 years), including both syphilis positive and syphilis negative, were recruited consecutively in eight health facilities in eight regions of Cameroon.
Outcome measures
Blood samples were screened for the detection of anti-Treponema pallidum antibodies using the One Step Rapid Test (Qingdao Hightop Biotech), a non-treponemal test and ELISA (Biorex Diagnostics, UK), a treponemal test used as a reference method. Diagnostic performance of the Syphilis RDT was analysed using Epi Info V.7 and validated through online statistical tools such as StatPages, GraphPad, QuickCalcs and MedCalc software.
Results
Of the 583 samples tested, the Hightop Syphilis RDT revealed a sensitivity of 84.6% (95% CI: 74.8% to 91.1%) and specificity of 98.5% (95% CI: 97.5% to 99.1%). The positive predictive value (PPV) and negative predictive value (NPV) were 84.6% (95% CI: 74.8% to 91.1%) and 98.5% (95% CI: 97.5% to 99.1%), respectively. Regarding the stratification of diagnostic performance by clinical stage, the test showed a sensitivity of 100.0% (95% CI: 71.51% to 100.0%) and specificity of 99.06% (95% CI: 94.86% to 99.98%). The PPV and NPV were 91.67% (95% CI: 61.00% to 98.72%) and 100.0% (95% CI: 96.55% to 100.0%), respectively, in symptomatic individuals. Among asymptomatic individuals, sensitivity was 97.56% (95% CI: 87.14% to 99.94%) and specificity was 100.0% (95% CI: 99.14% to 100.0%). The PPV and NPV were 100.0% (95% CI: 91.19% to 100.0%) and 99.77% (95% CI: 98.40% to 99.97%), respectively.
Conclusions
The Hightop Syphilis RDT demonstrated adequate diagnostic performance, particularly among symptomatic individuals, supporting its utility as a reliable tool for syphilis detection in clinical settings.
Keywords: Syphilis, Cross-Sectional Studies, INFECTIOUS DISEASES, HIV & AIDS, MICROBIOLOGY, Diagnostic microbiology
STRENGTHS AND LIMITATIONS OF THIS STUDY.
The study involved a large sample of blood drawn from syphilis-positive and syphilis-negative individuals in health facilities in 8 of the 10 regions of Cameroon.
The Hightop Syphilis Rapid Diagnostic Test exhibits excellent diagnostic capabilities in both symptomatic and asymptomatic individuals.
The study does not differentiate between syphilis stages (primary, secondary and tertiary disease) due to the design.
Introduction
Syphilis is a sexually transmitted infection (STI) caused by Treponema pallidum, a pathogenic bacterium belonging to the genus Treponema within the phylum Spirochaetes.1 2 Syphilis remains a global public health priority, with an estimated 17.7 million individuals aged 15–49 years infected in 2012 and approximately 5.6 million new cases occurring annually worldwide.3 In 2023, the Centers for Disease Control and Prevention (CDC) recorded over 2.4 million STI cases, including 209 000 of syphilis, 600 000 of gonorrhoea and 1.6 million of chlamydia.4 Alarmingly, 3882 congenital syphilis cases were documented, resulting in 279 stillbirths and neonatal deaths.4 Sub-Saharan Africa bears the highest global burden of syphilis, with more than 60% of newly diagnosed cases worldwide each year.3 5 T. pallidum is transmitted through several routes, including transfusion of contaminated blood, unprotected sexual contact both heterosexual (men who have sex with women) and homosexual (men who have sex with men) as well as vertical transmission from mother to child during pregnancy and breastfeeding.1 6 7 Syphilis progresses through four clinical stages, of which primary, secondary and early latent stages are infectious. Untreated, primary syphilis can progress towards the secondary, latent and tertiary stages of the disease.8
Serological testing remains the keystone of syphilis diagnosis, serving as the primary method for detecting asymptomatic infections and for confirming clinical suspicion in symptomatic patients.2 Accordingly, syphilis diagnosis and staging require the combined use of two types of serological tests: non-treponemal tests (NTTs) and treponemal tests (TTs).2 NTTs, such as the rapid plasma reagin (RPR) and Venereal Disease Research Laboratory (VDRL) method, lack specificity due to cross-reactivity with antibodies produced in various other pathological conditions, which can result in false-positive test outcomes.2 9 When one or both NTTs were positive results, confirmation is performed using TTs such as the fluorescent treponemal antibody absorption (FTA-ABS), T. pallidum particle agglutination (TPPA) or enzyme immunoassay.10
In Cameroon, the conventional syphilis screening algorithm recommends initial testing with an NTT (RPR or VDRL), followed by confirmation using a TT if the initial test is reactive.11 A study conducted by Solaimalai et al12 in India reported that the ELISA showed a sensitivity of 98% and specificity of 97.5%. In contrast, the VDRL test demonstrated a sensitivity of 71.6% and specificity of 89.5%, while the RPR test showed a sensitivity of 73.5% and specificity of 90.5%. Furthermore, ELISA demonstrated excellent concordance with the T. pallidum haemagglutination assay (TPHA), reflected by a kappa coefficient of 0.9, whereas VDRL and RPR showed moderate concordance with TPHA (kappa=0.6).12 Several studies worldwide have evaluated the diagnostic performance of the syphilis rapid diagnostic tests (RDTs) across diverse population groups.13,15 To date, there is a limited number of studies globally that have evaluated the diagnostic performance of the Hightop Syphilis RDT relative to the reference ELISA, particularly in Cameroon.
Rationale
Syphilis remains a major public health concern, particularly in resource-limited settings where diagnostic capacity is constrained. While RDTs offer a promising alternative for early detection, their utility must be validated in real-world contexts. This study aimed to assess the diagnostic performance of the Hightop Syphilis RDT compared with ELISA, with a view to informing syphilis screening strategies in Cameroon and similar settings.
Study objective
This current study was conducted to assess the diagnostic performance of the Hightop Syphilis RDT in comparison with the ELISA used as a reference method.
Materials and methods
Study design and setting
This laboratory-based cross-sectional study was designed to assess the laboratory performance of the Hightop Syphilis RDT compared with ELISA used as a reference method for the diagnosis of syphilis in Cameroon. The sample (serum and plasma) was collected in eight regions of the country and performed at the National Public Health Laboratory (NPHL).
Participant and data collection
From 29 April to 25 August 2023, a total of 583 adult participants (aged ≥21 years) of both sexes were consecutively enrolled in eight healthcare facilities across the country. Prior to participation, all individuals were provided with a written information note outlining the study’s objectives, procedures, potential risks and benefits. Written informed consent was obtained from each participant. Demographic and clinical data were collected through structured, face-to-face interviews using a coded questionnaire (online supplemental file S1). Venous blood samples were collected using sterile needles and vacuum collection systems. Specimens were drawn into dry tubes for serum and EDTA tubes for plasma. Blood samples were centrifuged at 1500 rpm for 5 min; serum and plasma were aliquoted into colour-coded cryovials (red for serum, blue for plasma). Samples were transported under cold chain conditions in ice-packed coolers by ground to the NPHL. On receipt, samples were stored at −20°C and −80°C until laboratory analysis. Each specimen was labelled with a unique identifier and accompanied by the participant’s coded questionnaire data to maintain data integrity and traceability.
Inclusion and exclusion criteria
The study included adult participants aged 21 years and above, of both sexes who voluntarily agreed to participate. Participants with incomplete questionnaire data, insufficient and non-compliant biological samples were excluded (figure 1).
Figure 1. A chart flow diagram showing the process of participant recruitment. NTT, non-treponemal test; RDT, Rapid Diagnostic Test; TT, treponemal test.
Data sources and variables
The present study used primary data collected from participants attending routine consultation in health facilities across eight regions of the country following a face-to-face interview. Quantitative variables such as age, qualitative variables such as gender and laboratory results were collected and recorded in a coded questionnaire. The prevalence of syphilis (qualitative variable) was measured using Hightop Syphilis RDT and confirmed by ELISA.
Sampling method and sample size determination
The participants were enrolled using a non-probabilistic sampling method. The sample size was calculated using the Kish and Leslie formula for cross-sectional studies: n=Z²×p×q/m²,16 where n is the required sample size, Z is the standard normal deviate corresponding to a 95% CI (Z=1.96), p is the estimated prevalence (50%), q=1−p and m is the error (5%).
Laboratory testing
Blood samples were tested using the One Step Rapid Test (Qingdao Hightop Biotech; Reference: H133; Lot: TP1220904; date of manufacture: 15 September 2022; expiry date: 14 September 2024). The Hightop test is a lateral flow chromatographic immunoassay based on a sandwich format, designed for the qualitative detection of antibodies to TP in serum, plasma or whole blood. Test results were interpreted in accordance with the manufacturer’s guidelines. A reactive (positive) result was defined by the presence of two distinct red lines: one in the control (C) and one in the test (T) zone. A non-reactive (negative) result was indicated by a single red line in the control (C) zone, with no line in the test (T) zone. Results were considered invalid if (1) a red line appeared in the test (T) zone without a corresponding line in the control (C) zone or (2) no red lines appeared in either zone within the recommended reading window of 15–20 min. The diagnostic performance of the Hightop Syphilis RDT was assessed by comparison with a qualitative sandwich-format ELISA, used as the reference method. The ELISA was manufactured by Biorex Diagnostics (Antrim Technology Park, BT41 1QS, UK), certified under ISO 13485 (Reference: BXE0995A; Lot No.: TP-2210-1; date of manufacture: October 2022; expiry date: June 2024). All procedures were conducted in accordance with the respective manufacturers’ instructions. The ELISA method was selected as the reference standard due to its high sensitivity and reliability in detecting anti-Treponema pallidum antibodies associated with syphilis infection.11 17
Statistical analysis
The data were entered into Microsoft Excel V.2016 and subsequently converted to Excel V.97-2003 sheet. The dataset was imported into Epi Info V.7 (CDC, Atlanta, USA) for statistical analysis. Descriptive statistics, including means, SD, frequencies and percentages, were calculated to summarise demographic and clinical characteristics of the study population. Diagnostic performance metrics including sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), accuracy, likelihood ratios (LR+ and LR⁻) and Cohen’s kappa coefficient (κ) were calculated using Epi Info V.7 and validated through online statistical tools such as StatPages, GraphPad, QuickCalcs and MedCalc software. 95% confidence interval (95%CI) were reported for all diagnostic indices. Furthermore, the receiver operating characteristic (ROC) curve was constructed using MedCalc18 to evaluate the overall discriminative ability of the Hightop Syphilis RDT. The area under the curve (AUC) was computed to quantify test accuracy. ROC curves were also stratified by clinical presentation (symptomatic vs asymptomatic) to assess performance variation across subgroups.
Results
Demographic characteristics of participants
The study enrolled 583 participants aged between 21 and 81 years (mean: 38.96, SD ± 12.46 years). The majority was female sex (70.33%), aged 21–30 years (30.87%), had attained secondary education (37.39%), was married or cohabiting (51.80%), resided in urban settings (75.30%), was asymptomatic at presentation (79.93%) and identified as Christian (61.92%).
Distribution of the Hightop Syphilis RDT results based on reference method
Out of the 583 blood samples analysed, the reference method identified 52 reactive and 531 non-reactive cases. The Hightop Syphilis RDT correctly detected 44 as true positives and 523 as true negatives. However, it yielded 8 false-reactive results (false positives) and missed 8 ELISA-reactive cases (false negatives) (table 1).
Table 1. Distribution of the Hightop Syphilis RDT results based on reference method.
| Reference method using ELISA | |||
|---|---|---|---|
| Hightop syphilis RDT | Reactive | Non-reactive | Total |
| Reactive | 44 (TP) | 8 (FP) | 52 |
| Non-reactive | 8 (FN) | 523 (TN) | 531 |
| Total | 52 | 531 | 583 |
FN, false negative; FP, false positive; RDT, Rapid Diagnostic Test; TN, true negative; TP, true positive.
Diagnostic performance of the Hightop Syphilis RDT based on the reference method
The diagnostic performance of the Hightop Syphilis RDT, as compared with the reference ELISA method, demonstrated a sensitivity of 84.6% (95% CI: 74.8% to 91.1%) and a specificity of 98.5% (95% CI: 97.5% to 99.1%). The PPV and NPV were also 84.6% (95% CI: 74.8% to 91.1%) and 98.5% (95% CI: 97.5% to 99.1%), respectively. The positive likelihood ratio (LR+) was estimated between 30.26 and 105.16, while the negative likelihood ratio (LR⁻) was 0.1562 (95% CI: 0.089 to 0.259). The overall diagnostic accuracy and Cohen’s kappa coefficient were similar with 83.1% (95% CI: 72.3% to 90.3%) (table 2).
Table 2. Diagnostic performance of the Hightop Syphilis RDT based on the reference method.
| Value | 95% CI | |
|---|---|---|
| Sensitivity | 84.6% | 74.8% to 91.1% |
| Specificity | 98.5% | 97.5% to 99.1% |
| PPV | 84.6% | 74.8% to 91.1% |
| NPV | 98.5% | 97.5% to 99.1% |
| LR+ | 56.41 | 30.26 to 105.16 |
| LR- | 0.1562 | 0.089 to 0.259 |
| Accuracy | 83.1% | 72.3% to 90.3% |
| Kappa | 83.1% | 72.3% to 90.3% |
LR+, positive likelihood ratio; LR−, negative likelihood ratio; NPV, negative predictive value; PPV, positive predictive value; RDT, Rapid Diagnostic Test.
An ROC curve of the Hightop Syphilis RDT
The ROC curve illustrates the balance between sensitivity (true positive rate) and 1–specificity (false positive rate). In this study, the Hightop Syphilis RDT exhibited a sensitivity of 84.6% and a specificity of 98.5%, positioning its performance point well above the line of no discrimination. The AUC was 0.92, indicating high overall diagnostic accuracy (figure 2).
Figure 2. An ROC curve connects coordinate points with 1−specificity (= false positive rate) as the x-axis and sensitivity as the y-axis at all cut-off values measured from the test results. AUC, area under the curve; RDT, Rapid Diagnostic Test; ROC, receiver operating characteristic.
Distribution of the results of both tests according to the clinical stage
Among the 583 participants enrolled, 20.07% (117/583) were symptomatic, while 79.93% (466/583) were asymptomatic. In the symptomatic group, the Hightop Syphilis RDT identified 100% (11/11) of true positive cases, with no false negatives and a single false positive result (0.94%, 1/106), yielding 99.06% (105/106) true negatives. Among asymptomatic individuals, the test demonstrated a true positive rate of 97.56% (40/41), with one false negative (2.44%) and no false positives, resulting in 100% (425/425) specificity (table 3).
Table 3. Distribution of the results of both tests according to the clinical stage.
| Hightop Syphilis RDT | Reference method using ELISA | ||
|---|---|---|---|
| Reactive | Non-reactive | Total | |
| Symptomatic | |||
| Reactive | 11 (TP) | 1 (FP) | 12 |
| Non-reactive | 0 (FN) | 105 (TN) | 105 |
| Total | 11 | 106 | 117 |
| Asymptomatic | |||
| Reactive | 40 (TP) | 0 (FP) | 40 |
| Non-reactive | 1 (FN) | 425 (TN) | 426 |
| Total | 41 | 425 | 466 |
FN, false negative; FP, false positive; RDT, Rapid Diagnostic Test; TN, true negative; TP, true positive.
Diagnostic performance of the Hightop Syphilis RDT based on the reference method according to the clinical stage
In symptomatic individuals, the Hightop Syphilis RDT revealed a sensitivity of 100.0% (95% CI: 71.51% to 100.0%) and specificity of 99.06% (95% CI: 94.86% to 99.98%). The PPV and NPV were 91.67% (95% CI: 61.00% to 98.72%) and 100.0% (95% CI: 96.55% to 100.0%), respectively. The test showed an LR+ of 106.00 (95% CI: 15.07 to 745.59), an LR⁻ of 0.00, an overall accuracy of 99.15% (95% CI: 95.33% to 99.98%) and a Cohen’s kappa coefficient of 95.20% (95% CI: 68.0% to 96.0%) (table 4).
Table 4. Diagnostic performance of the Hightop Syphilis RDT based on the reference method according to the clinical stage.
| Clinical stage | ||||
|---|---|---|---|---|
| Symptomatic | Asymptomatic | |||
| Value | 95% CI | Value | 95% CI | |
| Sensitivity | 100.0% | 71.51% to 100.0% | 97.56% | 87.14% to 99.94% |
| Specificity | 99.06% | 94.86% to 99.98% | 100.0% | 99.14% to 100.0% |
| PPV | 91.67% | 61.00% to 98.72% | 100.0% | 91.19% to 100.0% |
| NPV | 100.0% | 96.55% to 100.0% | 99.77% | 98.40% to 99.97% |
| LR+ | 106.00 | 15.07 to 745.59 | Na | Na |
| LR- | 0.00 | 0.0 | 0.02 | 0.00 to 0.17 |
| Accuracy | 99.15% | 95.33% to 99.98% | 99.79% | 98.81% to 99.99% |
| Kappa | 95.20% | 68.0% to 96.0% | 98.6% | 90.50% to 99.10% |
LR+, positive likelihood ratio; LR-, negative likelihood ratio; NPV, negative predictive value; PPV, positive predictive value; RDT, Rapid Diagnostic Test.
Among asymptomatic individuals, sensitivity was 97.56% (95% CI: 87.14% to 99.94%) and specificity was 100.0% (95% CI: 99.14% to 100.0%). The PPV and NPV were 100.0% (95% CI: 91.19% to 100.0%) and 99.77% (95% CI: 98.40% to 99.97%), respectively. LR+ was not applicable due to the absence of false positives, while LR⁻ was 0.02 (95% CI: 0.00 to 0.17). The test showed an accuracy of 99.79% (95% CI: 98.81% to 99.99%) and a kappa value of 98.6% (95% CI: 90.50% to 99.10%) (table 4).
ROC curve of the Hightop Syphilis RDT according to clinical stage
The ROC curve illustrates the diagnostic performance of the Hightop Syphilis RDT in both symptomatic and asymptomatic individuals. The blue line represents the symptomatic group, with an AUC of approximately 1.00, indicating perfect diagnostic accuracy. The green line represents the asymptomatic group, also with an AUC near 1.00, demonstrating excellent performance even in the absence of symptoms. The diagonal grey dashed line represents a non-informative test (AUC=0.5) (figure 3).
Figure 3. The ROC curve showing the diagnostic performance of the Hightop Syphilis RDT in both symptomatic and asymptomatic individuals. AUC, area under the curve; RDT, Rapid Diagnostic Test; ROC, receiver operating characteristic.
Discussion
The study was conducted to assess the laboratory performance of the Hightop RDT for syphilis diagnosis in Cameroon. A comparison was made between the Hightop Syphilis RDT and ELISA, used as a reference test. A total of 583 venous blood samples were obtained from adult participants attending eight healthcare facilities. The Hightop Syphilis RDT demonstrated a sensitivity of 84.6% (95% CI: 74.8% to 91.1%), indicating a strong capacity to detect true positive cases (table 2). Sensitivity is a key metric in assessing a diagnostic tool’s ability to correctly identify infected individuals, particularly in screening settings where early detection is critical for timely treatment and interruption of transmission. The suboptimal sensitivity observed may be partly explained by the predominance of asymptomatic individuals in the study population, likely associated with lower antibody titres, thus reducing test detectability. Nonetheless, the sensitivity corroborates with findings from previous studies, such as those by Kamolrattana et al,15 Lodiongo et al,19 and Van Den Heuvel et al,20 which reported sensitivities ranging from 85% to 87% for similar RDTs15 19 20 who reported a sensitivity of 85.7% (95%CI 63.7% to 97.0%), 86.4% (95% CI: 65.1% to 97.1%) and 87%, respectively, using Bioline HIV/Syphilis Dual test and Determine Syphilis RDT. In contrast, our findings diverge from those reported by Kamolrattana et al and Van Den Heuvel et al, who documented lower sensitivities of 73.5% and 66.7% (95% CI: 43.0% to 85.4%), respectively, when evaluating the Multiplo TP/HIV Duo Test and the Bioline Syphilis 3.0 assay.15 20 The specificity of the Hightop Syphilis RDT was 98.5% (95% CI: 97.5% to 99.1%) (table 2). Specificity measures the ability of the test to correctly identify individuals without the disease, minimising false positives. The high specificity of the Hightop Syphilis RDT reinforces its capability to accurately identify those without syphilis, which is crucial for reducing unnecessary follow-ups and anxiety for patients. The specificity obtained in our study corroborates the study conducted by Zhang et al who reported a specificity of 98% (95% CI: 96% to 99%) across 17 studies in a meta-analyses using treponemal component.21 Meanwhile, our specificity is lower compared with the study performed by Van Den Heuvel et al20 who reported specificities ranging from 99.0% to 100% using Dual test Multiplo and the Bioline. The difference can be explained by the sample size, the study population and the type of test used (One-step RDT vs Dual test).
In this current study, the sensitivity and PPV of the Hightop Syphilis RDT were similar, with 84.6% (95% CI: 74.8% to 91.1%) (table 2). PPV reflects the proportion of true positives among all positive test results, indicating the likelihood that a positive result correctly identifies the disease. In the same vein, the specificity and the NPV of the Hightop syphilis test were similar with 98.5% (95% CI: 97.5% to 99.1%) (table 2). NPV measures the proportion of true negatives among all negative test results, highlighting the ability of the test to accurately rule out the disease. A similar performance was reported by the study conducted by Shimelis and Tadesse22 reported for the diagnosis of syphilis, a PPV and NPV of 95.4% (95% CI: 89.3% to 98.5%) and 98% (95% CI: 93.4% to 99.7%), respectively, using HIV/Syphilis Dual test. Moreover, our findings reported that the accuracy and kappa (k) were similar with 83.1% (95% CI: 72.3% to 90.3%) (table 2). Accuracy provides a broad measure of overall test performance, integrating both sensitivity and specificity, while kappa measures the agreement between the test results and the actual disease status, accounting for chance agreement. Our findings demonstrate the excellent concordance between Hightop Syphilis RDT and ELISA. The Cohen’s Kappa (κ) obtained in this study was lower compared with the study done by Lodiongo et al who reported a k of 0.92 (95% CI: 0.980 to 0.999) using Dual test.19
Stratified analysis showed that the Hightop Syphilis RDT performed exceptionally in symptomatic individuals, with a sensitivity of 100%, specificity of 99.06% and overall accuracy of 99.15% (table 3). These results are consistent with findings from a Brazilian study evaluating a treponemal RDT, which reported sensitivities of 75–100% depending on VDRL titres, with 91.9% sensitivity in VDRL-reactive samples and specificities up to 100%, including among HIV-positive individuals.23 Notably, pooled sensitivity exceeded 98% in high-titre cases, a trend similarly observed in our symptomatic cohort, indicating that antibody detection is more effective during active syphilis infection.21 Furthermore, meta-analyses of treponemal RDTs report pooled sensitivity and specificity of approximately 93% and 98%, respectively, while non-treponemal components demonstrate sensitivity around 90% and specificity near 97%.21
The Hightop Syphilis RDT showed excellent diagnostic performance among asymptomatic individuals, with a sensitivity of 97.6% (95% CI: 87.1% to 99.9%) and a specificity of 100.0% (95% CI: 99.1% to 100.0%). Predictive values were also high, with a PPV of 100.0% (95% CI: 91.2% to 100.0%) and an NPV of 99.8% (95% CI: 98.4% to 99.9%). The overall accuracy rate was 99.8%, and the Cohen’s kappa value (κ=0.986; 95% CI: 0.905 to 0.991) reflected almost perfect agreement with the ELISA reference standard (table 3). These findings are consistent with pooled meta-analyses showing that treponemal RDTs maintain high specificity (approximately 98–99%) and moderate-to-high sensitivity (85–95%) among asymptomatic individuals under laboratory conditions. A recent WHO-supported systematic review similarly reported that dual-component syphilis RDTs achieved pooled sensitivity and specificity of 86% and 97% for treponemal targets, and 80% and 96% for non-treponemal components, respectively.21
In this study, the Hightop Syphilis RDT showed a sensitivity of 84.6% and specificity of 98.5%, with an area under the ROC curve (AUC) of 0.92, indicating high overall diagnostic accuracy and excellent discriminatory ability. The ROC curve analysis, stratified by clinical stage, revealed AUCs nearing 1.0 in both symptomatic and asymptomatic individuals, indicating that the test reliably distinguishes between infected and non-infected cases across different clinical stages. These findings are consistent with data from a systematic review and meta-analysis conducted by Zhang et al, which reported pooled AUCs between 0.90 and 0.95 for treponemal RDTs, confirming their robust diagnostic performance in diverse populations.21 The near-perfect AUC values observed in symptomatic individuals are likely associated with higher antibody titres in active syphilis, a trend similarly reported in other RDT evaluations such as those by Arai et al, where sensitivity improved significantly with increasing VDRL titres.23
Study limitations
The study has several limitations. First, the cross-sectional design limited the ability to track the natural course of syphilis infection or antibody dynamics over time. As a result, diagnostic performance could not be evaluated across different stages of infection (primary, secondary, latent or tertiary). Second, clinical staging of syphilis was based solely on self-reported symptoms and physical examination without laboratory confirmation tests such as dark-field microscopy or PCR, introducing a risk of classification bias. Third, the sample included a high proportion of asymptomatic individuals, which may have led to an underestimation of sensitivity, as low antibody titres are more common in early or latent stages of infection. Furthermore, using ELISA alone as the reference standard without confirmatory tests of the other TT such as FTA-ABS or TPPA may have led to an overestimation of prevalence and diagnostic accuracy. Finally, the lack of verification for recent antibiotic exposure may have resulted in false-negative results, and recruitment from selected health facilities introduces potential selection bias, thereby limiting the generalisability.
Conclusion
The Hightop Syphilis RDT shows strong, satisfactory diagnostic performance, with high specificity and strong concordance with ELISA results. Its ease of use, rapid turnaround time and robustness in both symptomatic and asymptomatic populations support its application as a valuable screening tool in resource-limited settings. These findings contribute evidence for the integration of this test into national syphilis screening strategies. Further research is warranted to evaluate its longitudinal performance and applicability across different stages of infection.
Supplementary material
Acknowledgements
The authors would like to express their thanks to the individuals for their participation in this study. We also like to express our sincere gratitude to the laboratory technicians of the eight regions for the blood samples collection. Moreover, we also like to acknowledge the technical staff of NPHL for the technical support in blood analysis, particularly to Mme Cressence Fouda.
Footnotes
Funding: The authors have not declared a specific grant for this research from any funding agency in the public, commercial or not-for-profit sectors.
Prepublication history and additional supplemental material for this paper are available online. To view these files, please visit the journal online (https://doi.org/10.1136/bmjopen-2024-093330).
Provenance and peer review: Not commissioned; externally peer reviewed.
Patient consent for publication: Consent obtained directly from patient(s).
Ethics approval: Ethical approval for this study was granted by the National Committee for Ethics in Human Health Research (Reference: 2023/02/1526/CE/CNERSH/SP). All participants were provided with detailed information regarding the study’s objectives, potential risks and benefits through an information note. Written informed consent form was obtained from each participant prior to enrolment (online supplemental file 2).
Patient and public involvement: Patients and/or the public were not involved in the design, or conduct, or reporting, or dissemination plans of this research.
Data availability statement
Data are available upon reasonable request.
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