Evaluation of Plasmodium falciparum gametocyte detection in different patient material

Background: For future eradication strategies of malaria it is
important to control the transmission of gametocytes from humans to
the anopheline vector which causes the spread of the disease.
Sensitive, non-invasive methods to detect gametocytes under field
conditions can play a role in monitoring transmission potential.
Methods: Microscopically Plasmodium falciparum-positive patients
from Jimma, Ethiopia donated finger-prick blood, venous blood,
saliva, oral mucosa and urine samples that were spotted on filter
paper or swabs. All samples were taken and stored under equal,
standardized conditions. RNA was extracted from the filter paper
and detected by real-time QT-NASBA. Pfs16-mRNA and Pfs25-mRNA were
measured with a time to positivity to detect gametocyte specific
mRNA in different gametocyte stages. They were compared to
18S-rRNA, which is expressed in all parasite stages. Results were
quantified via a known dilution series of artificial RNA copies.
Results: Ninety-six samples of 16 uncomplicated malaria patients
were investigated. 10 (66.7%) of the slides showed gametocyte
densities between 0.3-2.9 gametocytes/mu l. For all RNA-targets,
molecular detection in blood samples was most sensitive;
finger-prick sampling required significantly smaller amounts of
blood than venous blood collection. Detection of asexual 18S-rRNA
in saliva and urine showed sensitivities of 80 and 67%,
respectively. Non-invasive methods to count gametocytes proved
insensitive. Pfs16-mRNA was detectable in 20% of urine samples,
sensitivities for other materials were lower. Pfs25-mRNA was not
detectable in any sample. Conclusions: The sensitivity of
non-invasively collected material such as urine, saliva or mucosa
seems unsuitable for the detection of gametocyte-specific mRNA.
Sensitivity in asymptomatic carriers might be generally even lower.
Finger-prick testing revealed the highest absolute count of RNA
copies per mu L, especially for Pfs25-mRNA copies. The method
proved to be the most effective and should preferably be applied in
future transmission control and eradication plans. A rapid test for
gametocyte targets would simplify efforts.