Fully automated point-of-care differential diagnosis of acute febrile illness

Author summary Infectious diseases in tropical regions may have a variety of viral, bacterial or parasitic origins and a patient may suffer from several diseases simultaneously, each presenting with acute fever as a clinical symptom. This makes it difficult to determine the origin of the pathogen causing the disease(s). In addition to the endemic infectious diseases, outbreaks of epidemics frequently complicate diagnostic demands. Accurate diagnosis for proper patient management requires the utilization of highly sensitive and specific, rapid, easy-to-use diagnostic tools compatible with point-of-care settings. We describe the use of a disk-shaped microfluidic platform, the FeverDisk, for differential diagnosis of acute fever. Our FeverDisk platform demonstrated its capability to detect bacteria, viruses and parasites that are typical of tropical single and co-infections from biobanked samples within only 2 hours and in very good agreement with reference method results. This, in combination with its easy-to-use and point-of-care compatible nature, render our platform a promising candidate for detection of tropical diseases and precise identification of the cause of acute fever, in endemic and epidemic settings. Future work will involve extensive clinical characterization of the platform in prospective studies. Background In this work, a platform was developed and tested to allow to detect a variety of candidate viral, bacterial and parasitic pathogens, for acute fever of unknown origin. The platform is based on a centrifugal microfluidic cartridge, the LabDisk (FeverDisk for the specific application), which integrates all necessary reagents for sample-to-answer analysis and is processed by a compact, point-of-care compatible device. Methodology/Principal findings A sample volume of 200 mu L per FeverDisk was used. In situ extraction with pre-stored reagents was achieved by bind-wash-elute chemistry and magnetic particles. Enzymes for the loop-mediated isothermal amplification (LAMP) were pre-stored in lyopellet form providing stability and independence from the cold chain. The total time to result from sample inlet to read out was 2 h. The proof-of-principle was demonstrated in three small-scale feasibility studies: in Dakar, Senegal and Khartoum, Sudan we tested biobanked samples using 29 and 9 disks, respectively; in Reinfeld, Germany we tested spiked samples and analyzed the limit of detection using three bacteria simultaneously spiked in whole blood using 15 disks. Overall during the three studies, the FeverDisk detected dengue virus (different serotypes), chikungunya virus, Plasmodium falciparum, Salmonella enterica Typhi, Salmonella enterica Paratyphi A and Streptococcus pneumoniae. Conclusions/Significance The FeverDisk proved to be universally applicable as it successfully detected all different types of pathogens as single or co-infections, while it also managed to define the serotype of un-serotyped dengue samples. Thirty-eight FeverDisks at the two African sites provided 59 assay results, out of which 51 (86.4%) were confirmed with reference assay results. The results provide a promising outlook for future implementation of the platform in larger prospective clinical studies for defining its clinical sensitivity and specificity. The technology aims to provide multi-target diagnosis of the origins of fever, which will help fight lethal diseases and the incessant rise of antimicrobial resistance.

Automated summary

The study presents a LabDisk platform for differential diagnosis of acute fever caused by various pathogens, demonstrating promising results for detecting tropical diseases and precise identification of the cause of acute fever.