Review: Genetic diversity of Plasmodium falciparum: asexual stages

keywords malaria, Plasmodium falciparum, polymorphism, drug resistancecorrespondence Christian G. Meyer, Bernhard-Nocht-Institut fu¨r Tropenmedizin, Bernhard-Nocht-Str. 74, 20359 Hamburg, Germany. Fax: +49 40 4281 8512; E-mail: c.g.meyer@bni.uni-hamburg.deIntroductionThe genetic complexity of Plasmodium falciparum, theinfectious agent of malignant malaria, and in particular itsability to generate mutant variants, make it a successfulpathogen. Genetic variants are involved in pathogenicityand in immune responses and have led to the emergence ofresistance against virtually any drug available for causativetreatment. Such variants areunder strongselective pressure.Analysesofsequencevariationingenesegmentsthatarenotsubjectedtoselectionandofintergenicsequencespermitthestudy of additional mutation patterns, interpopulationdiversity of the parasite and its evolutionary origin.The genomeThe genome of P. falciparum consists of 14 linearchromosomes with a total of 25–30 megabases of nuclearDNA with approximately 5000 genes, a mitochondrialfragment of a 6 kb repeat element, and a circular elementof 35 kb within the apicoplast. As a consequence ofcontinuous deletions and crossing-over and rearrangementevents occurring preferably at their telomeric regions, thechromosomes differ considerably in size (Corcoran et al.1986). The genome is extremely A/T-rich (80%), whichhas led to difficulties in conventional sequencing strategiesbecause of the instability of genomic fragments in bacterialEscherichia coli clones. Meanwhile, several yeast artificialclone (YAC) constructs have been established, allowing fora stable maintenance of P. falciparum clone fragments.Essentially based on YAC contiguous sequences (con-tigs), the P. falciparum genome is now subject of a largeDNA-sequencing project, the Malaria Genome Project,which was established in 1996. Contig arrays andrestriction maps have been produced for mapping ofcomplete chromosomes (Rubio et al. 1995). At present, themajority of the sequences of the 14 chromosomes of theP. falciparum clone 3D7 are covered by YAC contigs.Sequencing is performed by a chromosomal shotguntechnique with a 10–15· coverage. The project is fundedby the Wellcome Trust, the Burroughs Wellcome Fund, theUS Department of Defence, and the National Institute ofAllergy and Infectious Diseases (US) and performed byStandford University (US), the Sanger Centre (UK), theInstitute for Genomic Research (TIGR, US), and contri-buting groups. Sequencing is close to completion andsequences are published for chromosome 2 and 3 (Gardneret al. 1998; Bowman et al. 1999; sequence of chromosome5 not yet published). Gaps still remain to be filled in otherchromosomes. The closure of gaps is complicated by theconsiderable number of A/T-repeats. Sequencing of thecytoplasmatic mitochondrial and apicoplast segments hasbeen completed. A draft sequence of the P. falciparum 3D7genome is expected to be published this year.Sequence information is being deposited in severalclickable databases (Table 1). The information available atpresent has, by in silico analysis of genetic similarity withother organisms, allowed to trace and to assign a functionto approximately 40% of the genes. Many of them encodeproteins of metabolic pathways. The sequence informationfinally provided will, after the determination of transcrip-tion patterns and the identification of coding sequences,support detailed analyses of relevant proteins, parasite-specific metabolic pathways and, furthermore, support theidentification of targets for new drugs and for vaccines.Targets for drugs are now continuously being identified.One example described on the basis of P. falciparum DNAsequence information are nuclear genes that are involved inthe pathway of lipid synthesis and the subsequentidentification of the enoyl acyl-carrier protein (Waller et al.