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Comparative Study
. 2009 May 11:8:98.
doi: 10.1186/1475-2875-8-98.

A comparison of the sensitivities of detection of Plasmodium falciparum gametocytes by magnetic fractionation, thick blood film microscopy, and RT-PCR

Stephan Karl  1 , Timothy M E Davis, Tim G St-Pierre
Affiliations
Comparative Study

A comparison of the sensitivities of detection of Plasmodium falciparum gametocytes by magnetic fractionation, thick blood film microscopy, and RT-PCR

Stephan Karl et al. Malar J. .

Abstract

Background: The magnetic properties of Plasmodium-infected erythrocytes have been exploited for different clinical and research purposes. A recent study in a rural clinical setting in Papua New Guinea has demonstrated that Plasmodium falciparum gametocyte detection is facilitated by magnetic deposition microscopy but no study has yet determined the relative sensitivity and limit of detection of a magnetic fractionation technique. The present study compares the detection limit and sensitivity of a technique based on the use of commercially available magnetic fractionation columns with those for thick blood film microscopy and reverse transcriptase polymerase chain reaction (RT-PCR) methods.

Methods: Gametocyte detection in six series of dilutions of cultured P. falciparum parasites with known gametocytaemia was conducted using magnetic fractionation, thick blood film, and RT-PCR techniques.

Results: The preparations obtained by the magnetic fractionation method were of thin film quality allowing easy gametocyte identification by light microscopy. Magnetic fractionation had a higher sensitivity and approximately two orders of magnitude better limit of detection than thick blood film microscopy. Gametocytes were also more readily detectable on the magnetically fractionated preparations. Magnetic fractionation had a similar limit of detection to that of RT-PCR.

Conclusion: Magnetic fractionation is a highly sensitive and convenient method for gametocyte detection in comparison with the standard thick blood film and RT-PCR methods, and could readily be adapted to field application.

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Figures

Figure 1
Figure 1
Gametocytes on thick blood film (TBF) and magnetically fractionated (MF) preparations. Gametocytes as observed on TBF in the ranges of 101–102 (A) and 100–101 (B) as compared with gametocytes as observed on MF preparations in the ranges of 101–102 (C), 100–101 (D), 10-1–100 (E) and 10-2-10-1 (F). Images were obtained on a Nikon Eclipse TE2000 -N Microscope with a 1000 × optical magnification with a Nikon LH-M100CB-1 Camera.
Figure 2
Figure 2
Observed gametocyte increase on MF preparations as compared to TBF preparations. Panel A: Mean gametocyte number per high power microscope field for each of the gametocyte density ranges for MF and TBF preparations. Panel B: Mean gametocyte density observed with MF and TBF for each of the gametocyte density ranges.
Figure 3
Figure 3
RT-PCR for Pfs25 and Pfg377. Example gel showing bands for Pfs25 and Pfg377 from one of the dilution series used in this study. B = Blank, N = uninfected blood. The numbers above the other lanes are the gametocyte densities (μL-1) in the dilutions used in this experiment.
Figure 4
Figure 4
Detection limits for gametocytes using magnetic fractionation (MF), thick blood film (TBF), and RT-PCR techniques. Wilcoxon's matched pairs test was used as the significance test.
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