Detection of Acute Tropical Fever Pathogens via Multiplex Polymerase Chain Reaction: A Cross-Sectional Study to Guide...

Pakistan faces a substantial burden of tropical diseases, including malaria, dengue, and typhoid fever, particularly in Sindh Province, where recent outbreaks and floods have further increased disease incidence, highlighting the urgent need for improved diagnostics and disease control. Currently, diagnosis relies on multiple laboratory tests, contributing to frequent empirical antimicrobial use due to many undiagnosed or misdiagnosed cases. This study aimed to validate a rapid molecular test for the early diagnosis of malaria, dengue, and typhoid infections, thereby facilitating targeted therapy and reducing empirical antimicrobial use.

Patients aged three months or older who presented to Aga Khan Hospital clinical laboratories with fever lasting two to seven days were recruited. From a total of 1,123 collected blood samples, 400 were randomly selected for multiplex real-time polymerase chain reaction (PCR) testing. Multiplex PCR was performed alongside standard blood tests for malaria, dengue, and typhoid. Sensitivity, specificity, positive predictive value, and negative predictive value of PCR were calculated and compared with routine tests. Data analysis was conducted using STATA 17.0 (StataCorp, College Station, Texas).

Among the 1,123 samples collected over one year, 400 blood samples were randomly selected for molecular testing. Of these, 211 were PCR positive. The sensitivity for detecting malaria was 92.4%, specificity was 98.4%, and the positive and negative predictive values were 90.7% and 98.6%, respectively. The specificity of PCR for dengue and typhoid was 94.6% and 93%, respectively; however, the sensitivity for both dengue and typhoid was low (35.3% and 25%, respectively).

While multiplex PCR demonstrates high accuracy in detecting malaria, its limited sensitivity for dengue and typhoid highlights the need for more advanced molecular assays. Developing rapid and reliable diagnostic tests is essential to enable targeted treatment and promote antimicrobial stewardship within the community.