Diagnostic Accuracy of PCR in gambiense Sleeping Sickness Diagnosis, Staging and Post-Treatment Follow-Up: A 2-year Longitudinal Study

Abstract

Background

The polymerase chain reaction (PCR) has been proposed for diagnosis, staging and post-treatment follow-up of sleeping sickness but no large-scale clinical evaluations of its diagnostic accuracy have taken place yet.

Methodology/Principal Findings

An 18S ribosomal RNA gene targeting PCR was performed on blood and cerebrospinal fluid (CSF) of 360 T. brucei gambiense sleeping sickness patients and on blood of 129 endemic controls from the Democratic Republic of Congo. Sensitivity and specificity (with 95% confidence intervals) of PCR for diagnosis, disease staging and treatment failure over 2 years follow-up post-treatment were determined. Reference standard tests were trypanosome detection for diagnosis and trypanosome detection and/or increased white blood cell concentration in CSF for staging and detection of treatment failure. PCR on blood showed a sensitivity of 88.4% (84.4–92.5%) and a specificity of 99.2% (97.7–100%) for diagnosis, while for disease staging the sensitivity and specificity of PCR on cerebrospinal fluid were 88.4% (84.8–91.9%) and 82.9% (71.2–94.6%), respectively. During follow-up after treatment, PCR on blood had low sensitivity to detect treatment failure. In cerebrospinal fluid, PCR positivity vanished slowly and was observed until the end of the 2 year follow-up in around 20% of successfully treated patients.

Conclusions/Significance

For T.b. gambiense sleeping sickness diagnosis and staging, PCR performed better than, or similar to, the current parasite detection techniques but it cannot be used for post-treatment follow-up. Continued PCR positivity in one out of five cured patients points to persistence of living or dead parasites or their DNA after successful treatment and may necessitate the revision of some paradigms about the pathophysiology of sleeping sickness.

Author Summary

Post-treatment follow-up is crucial for sleeping sickness patient management and still relies on microscopic examination of the cerebrospinal fluid (CSF). Detection of the parasites DNA with the polymerase chain reaction (PCR) is proposed as a promising and possibly non-invasive alternative for monitoring treatment outcome, but has never been evaluated. We performed PCR on blood and CSF of 360 Trypanosoma brucei gambiense sleeping sickness patients, before treatment and during 2 years after treatment, and on blood of 129 controls. We found that performance of PCR to diagnose sleeping sickness and detect brain involvement was better or similar to current diagnostic techniques. However, we observed that PCR was unreliable for monitoring treatment outcome. Continued PCR positivity in cured patients points to persistence of parasites, or their DNA, after successful treatment, challenging the dogma that in sleeping sickness cure equals parasite elimination. In conclusion, we do not recommend PCR for treatment outcome assessment in sleeping sickness.