By Everard L. Napier, M.R.C.S., L.R.C.P. (Lond.). In charge Kala-azar research, Calcutta School of Tropical Medicine. Second edition. 185 pages of text with 15 charts in the text, 18 plates, and an appendix of references to literature, author index and subject index. Oxford University Press. London, Bombay, Calcutta, Madras, 1927
Malaria Research and Training Center, National School of Medicine and Pharmacy, Malaria Genetics Section, Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bamako, Mali
As chloroquine resistance spreads across Africa, the dihydrofolate reductase (DHFR) inhibitors pyrimethamine and proguanil are being used as alternative first-line drugs for the treatment and prevention of Plasmodium falciparum malaria. Resistance to these drugs is conferred by point mutations in parasite DHFR. These point mutations can be detected by polymerase chain reaction (PCR) assays, but better methods for sample collection, DNA extraction, and a diagnostic PCR are needed to make these assays useful in malaria-endemic areas. Here we report methods for collecting fingerstick blood onto filter paper strips that are air-dried, then stored and transported at room temperature. Cell lysis and DNA extraction are accomplished by boiling in Chelex-100. We also report a nested PCR technique that has improved sensitivity and specificity. These procedures readily detect mixed infections of parasites with both sensitive and resistant genotypes (confirmed by direct sequencing) and are reliable at parasite densities less than 250/mm3 in field surveys.