Laboratory Diagnosis of Malaria: Comparison of Manual and Automated Diagnostic Tests

Abstract

Malaria is the second most prevalent disease in Pakistan resulting in ~30,000 annual deaths. In endemic countries like Pakistan precise and timely diagnosis of malaria is imperative to overcome the associated risks of fatal outcomes. Malarial parasite was screened in 128 malaria suspected patients and 150 healthy controls, by species-specific PCR, microscopy of blood smears, hemoanalyzer Sysmex XE-2100, and rapid test devices (First Response Malaria® and ICT Malaria Combo®). The microscopy detected MP in 126 samples (parasite load/µl 386-53712/µl); 71.094% were infected with Plasmodium vivax and 14.844% with P. falciparum while 14.062% had mixed P. vivax and P. falciparum infection. The mean parasite load for P. vivax and P. falciparum was 14496/µl and 24410/µl, respectively. The abnormal scattergrams of DIFF, WBC/ Baso, IMI channel, and RET-EXT on Sysmex XE-2100 supported 99.2% parasite detection, whereas only 93% of confirmed malaria cases were detected by both rapid tests. About 127 samples were positive by PCR. Since Sysmex XE-2100 automatically detected the presence of malarial parasite with high sensitivity, it can be a good option for presumptive diagnosis in endemic areas. Microscopy remains the gold standard to confirm MP in suspected patients. Rapid diagnostic tests have acceptable sensitivity and specificity.

Figure 1

Parasitemia (P. falciparum and P. vivax) associated abnormalities in Sysmex XE2100 channels compared with normal sample at the top. (a) DIFF channel: the blue arrow indicates abnormal events depicting pseudoeosinophilia and graying of neutrophil cluster and double neutrophil and eosinophil populations; (b) WBC/BASO channel: yellow box shows parasite containing ghost RBCs; (c) IMI channel: comparatively more immature cells in area below yellow line in malaria positive cases; and (d) RET- EXT channel: presence of cluster of abnormal cells in any of the sectors extending vertically down and moving horizontally towards the y-axis.

Figure 2

Abnormal scattergram in Sysmex XE2100 of malaria positive cases.

Figure 3

Agarose gel electrophoresis of amplified product obtained by 16S rRNA PCR using Plasmodium species-specific primers (Padley et al.). Lane 1: P. vivax; Lane 2: Mixed P. vivax + P. falciparum; Lane 3: P. falciparum; Lane 4: negative; Lane 5: P. falciparum; Lane 6: P. vivax; Lane 7: 100 bp ladder.