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Acridine Orange Fluorescent Microscopy and the Detection of Malaria in Populations with Low-Density Parasitemia

Detection of low-density malaria parasites with Giemsa-stained thick smears (G-TS) requires time and experience and becomes impractical with high sample loads. Acridine orange fluorescent microscopy (AO/FM) of capillary centrifuged blood may offer an alternative technique. We compared AO/FM readings with G-TS in 290 specimens from asymptomatic people in Thai villages endemic for malaria. AO/FM specimens were prepared in modified capillary tubes coated with acridine orange (Quantitative Buffy Coat or "QBC tubes") and examined under a fluorescent microscope. Twenty-three (85.2%) of the 27 specimens found positive by G-TS had under 100 parasites/µl blood (< 35 parasites/200 microscopic fields). The overall AO/FM sensitivity was 78.9% [range: 66.7% (10/15) – 86.7% (13/15)]. For Plasmodium falciparum, regardless of stages, the sensitivities varied from 66.7% (8/12) to 91.7% (11/12). AO/FM performed better for P. falciparum than for Plasmodium vivax and for asexual than for sexual stages of the parasite. However, the species- and stage-specific results must be interpreted with caution because of the small sample sizes and very low parasite densities involved. The test specificity was 96.6% [range: 95.6% (263/275)–97.1% (263/271)]. These levels of accuracy plus the known advantages of AO/FM suggest that the test, supplemented with G-TS to improve species and stage differentiation, is also useful for screening low-density parasitemias.