While study microscopists changed over the study period, the procedures were consistent throughout.
For quality control, 10% of the negative samples and 20% of the positive samples were re-examined by at least two microscopist during the study.
Despite regular supervision and the use of the same clinician and microscopist throughout the study (which insured internal consistency of our results), the discrepancies we found between the Amsel's criteria and the accepted gold standard diagnosis (Nugent's method) in this study appear to preclude the use of this method under routine clinical conditions in our setting.
Blood films were stained with 10% Giemsa stain for 10 15 minutes and screened microscopically under a ×100 oil immersion lens using a light microscope by a microscopist trained by the study teams and based at the referral center.
A senior member of the study team reviewed the slides if there was any disagreement between the two microscopists.
In case of a difference in results (positive/negative; species diagnosis) between the two microscopists, the blood smear was re-examined by a third microscopist independent of the earlier findings and the third reading accepted as the final result.
The assessment of agreement between the 2 microscopists revealed discrepant readings of slides made from 3 strongly positive specimens.
Blood smears with discordant results (differences between the two microscopists in species diagnosis, in parasite density of >50% or in the presence of parasites) will be re-examined by a third microscopist, and parasite density will be calculated by averaging the two closest counts.
The final reference diagnosis for malaria was blood slides with concordant results between the two microscopists, and if the slide results differed, the diagnosis was based on polymerase chain reaction of filter paper blood samples.
κ-analysis overall agreement between the six microscopists was moderate (κ = 0.45) for murine typhus IgM IFA titers and fair (κ = 0.32) for scrub typhus IgM IFA titers (Table 1).
