Laboratory Evaluation of a Dual-Path Platform Assay for Rapid Point-of-Care HIV and Syphilis Testing



TEXT

Rapid multiplex point-of-care tests are increasingly available means to screen for syphilis and HIV infections (1,–4). Dual screening tests for antibodies to HIV and Treponema pallidum have been evaluated in serum and venipuncture whole-blood samples, showing excellent sensitivity and specificity (5,–7). We assessed the laboratory performance of the Chembio dual-path platform (DPP) HIV-syphilis rapid test and the Chembio digital electronic reader with stored serum samples collected from a cohort of men who have sex with men (MSM) and transwomen at high risk for HIV and syphilis infections.

Serum specimens were collected between 2013 and 2014 from MSM and transwomen who had been recruited into an ongoing cohort study in Lima, Peru, and the specimens were stored at −20°C (8). HIV positivity was determined with the Genscreen Ultra HIV Ag-Ab test (Bio-Rad, Hercules, CA), a novel enzyme immunoassay (EIA) which detects HIV p24 antigen (Ag) and HIV antibodies in the same test (9). Positive EIA results were confirmed by Western blotting (New Lav Blot I; Bio-Rad). Specimens also underwent rapid plasma reagin (RPR) testing, using the BD Macro-Vue RPR card test kit (BD, Franklin Lakes, NJ), and Treponema pallidum particle agglutination (TPPA) testing (Serodia; Fujirebio Diagnostics Inc., Tokyo, Japan). All tests were used according to the manufacturers' instructions.

The Chembio DPP HIV-syphilis test is a single-use, visual and qualitative immunochromatographic, dual rapid test for the detection of antibodies to HIV types 1 and 2 and Treponema pallidum in human serum, plasma, or venous or fingerprick whole-blood samples (10). A red control line confirms test validity. Visual observation of a red line in the HIV and/or syphilis detection zone is interpreted as a reactive result (10).

Immediately after visual interpretation, tests were analyzed using the small, battery-powered, Chembio electronic reader, which was designed specifically to complement the Chembio DPP technology. The electronic reader scans the DPP test cartridge and displays a numerical value based on the test line intensity. If the electronic reader value is higher than the set cutoff value, then the result for the sample is reported as positive; the test result is reported as negative if the measured value is lower than the cutoff value.

We estimated the sensitivity, specificity, and 95% confidence intervals (CIs) using the exact binomial method, and we calculated the concordance between the visual results of the Chembio DPP HIV-syphilis rapid test and the results of the reference tests using Cohen's kappa coefficient. Specimens were defined as HIV positive on the basis of Western blotting results. Specimens were defined as Treponema pallidum antibody positive on the basis of TPPA test results. For electronic reader data, we estimated the sensitivity, specificity, and 95% CIs using the exact binomial method and we evaluated the performance of different cutoff values.

Of the 450 specimens, 100 were confirmed by Western blotting to be HIV-1 positive only, 99 were positive for Treponema pallidum antibodies by TPPA testing only (of which 79 [80%] had RPR titers between 1:1 and 1:64), and 51 were positive for both HIV and Treponema pallidum antibodies by Western blotting and TPPA testing. Of the dual-antibody-reactive specimens, 72% (37/51 specimens) had RPR titers between 1:1 and 1:64. The remaining 200 specimens tested negative for HIV and syphilis antibodies. Additionally, positive and negative controls were used with the tested specimens.

With visual interpretation of HIV antibody reactivity, the test showed 155 positive and 295 negative results (Table 1). There were 4 false-positive results (DPP test positive and Western blot negative) and no false-negative results. The sensitivity of the HIV antibody component was 100% (95% CI, 97.6% to 100.0%), and the specificity was 98.7% (95% CI, 96.6% to 99.6%). The kappa coefficient for correlation between the reference HIV-1 Western blot test results and the Chembio DPP HIV-syphilis rapid test results was 0.98 (95% CI, 0.96 to 1.0).

TABLE 1

Laboratory performance for detection of HIV antibodies using a dual HIV-syphilis rapid immunodiagnostic test in Lima, Peru, in 2015 (n = 450)

^aDPP, dual-path platform; CI, confidence interval.

With visual interpretation of Treponema pallidum antibody reactivity, the test showed 142 positive and 308 negative results (Table 2). There were 8 false-negative results (DPP test negative and TPPA test positive) and no false-positive results. The sensitivity of the Treponema pallidum antibody component was 94.7% (95% CI, 89.8% to 97.7%), and the specificity was 100.0% (95% CI, 98.8% to 100.0%). The kappa coefficient for correlation between the reference TPPA syphilis test results and the Chembio DPP HIV-syphilis rapid test results was 0.96 (95% CI, 0.93 to 0.99).

TABLE 2

Laboratory performance for detection of Treponema pallidum antibodies using a dual HIV-syphilis rapid immunodiagnostic test in Lima, Peru, in 2015 (n = 450)

^aDPP, dual-path platform; TPPA, Treponema pallidum particle agglutination; CI, confidence interval.

Using the Chembio electronic reader with the default cutoff value of 10 for HIV antibody detection, the sensitivity was 100.0% (95% CI, 97.6% to 100.0%), the specificity was 98.7% (95% CI, 96.6% to 99.6%), and the kappa coefficient was 0.98 (95% CI, 0.96 to 1.00) (Table 1). When the cutoff value was changed to 21, the specificity was 100.0% (95% CI, 98.8% to 100.0%), the sensitivity was 100.0% (95% CI, 97.6% to 100.0%), and the kappa coefficient for correlation was 1.0. No statistical difference was found when use of a cutoff value of 10 was compared with visual interpretation (P > 0.1).

Using the Chembio electronic reader with the default cutoff value of 10 for Treponema pallidum antibody detection, the sensitivity was 94.7% (95% CI, 89.8% to 97.7%), the specificity was 99.7% (95% CI, 98.2% to 100.0%), and the kappa coefficient was 0.95 (95% CI, 0.93 to 0.98) (Table 2). Various cutoff values for the Treponema pallidum antibody component showed that the cutoff value of 10 demonstrated the best performance. No statistical difference was found when use of a cutoff value of 10 was compared with visual interpretation (P > 0.1).

Repeat laboratory testing was performed for the specimens with discordant test results. All 4 specimens with initial false-positive results for HIV antibodies were found to be negative in repeat testing with the Chembio DPP HIV-syphilis rapid test, with both visual interpretation and the electronic reader.

Of the 8 specimens with false-negative results for treponemal antibodies, repeat Chembio DPP HIV-syphilis rapid tests were weakly positive for 4 samples using visual observation; 4 samples remained negative. The electronic reader classified 2 of the 4 samples that were weakly reactive using visual interpretation as positive.

We conducted a laboratory-based performance study of the Chembio DPP HIV-syphilis rapid test and the electronic reader with characterized stored serum samples in Lima, Peru. With visual interpretation, we found outstanding test performance. Different electronic reader cutoff values were tested, and the cutoff value of 21 had 100% specificity and sensitivity for HIV antibody detection. For Treponema pallidum antibody detection, the electronic reader with the default cutoff value of 10 had the best sensitivity and specificity.

Our results are comparable to those of laboratory evaluations that were conducted previously with different point-of-care tests for HIV and treponemal antibody detection in sera and whole blood, which also yielded high performance values for sensitivity and specificity (1, 5,–7, 11,–13). In our evaluation, the sensitivity of the Chembio DPP HIV-syphilis rapid test to detect HIV antibodies was somewhat greater than its sensitivity to detect syphilis antibodies, in concurrence with several other studies (5, 6, 11, 12, 14).

Our study had some limitations. We used specimens from a single population in Lima, Peru, which did not include any pregnant women or low-risk populations. The population had high risks for HIV and syphilis acquisition; therefore, our values for the sensitivity and specificity of the test might not be representative. The tested specimens were stored serum samples instead of the ideal samples, i.e., fingerprick whole-blood specimens. Since the evaluation of stored serum specimens seemed to be highly accurate, further studies using fingerprick whole-blood specimens are warranted, to evaluate the field performance of the Chembio DPP HIV-syphilis rapid test and the electronic reader.

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ACKNOWLEDGMENTS

We thank all of the participants in the study, as well as the staff at Barton Health Center, Epicentro, and Cayetano Heredia University. All test materials were donated by Chembio Diagnostic Systems, Inc.

The funders and donors had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

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