Subtelomeric chromosome deletions in field isolates of Plasmodium falciparum and their relationship to loss of cytoadherence in vitro.

Abstract

Subtelomeric deletions are responsible for the loss of expression of several Plasmodium falciparum antigens, including the knob-associated histidine-rich protein (KAHRP). Such deletions are detectable by two-dimensional pulsed-field gradient electrophoresis (PFGE) in which the chromosomes separated in dimension 1 are cleaved with Apa I, and the sizes of telomeric fragments are determined in dimension 2. This sensitive technique has enabled us to examine the role of subtelomeric deletions in two aspects of the biology of Plasmodium falciparum. First, we show that similar subtelomeric deletions to those that occur in vitro also occur in field isolates. Second, we demonstrate a correlation between subtelomeric deletions and loss of the phenotype of "cytoadherence" in cultured isolates. Subclones were generated from the cytoadherent cloned isolate ItG2F6, and their phenotypes were examined with respect to cytoadherence, the expression of "knobs," and agglutination of infected erythrocytes with rabbit antiserum. The only chromosomal change detectable by two-dimensional PFGE among subclones that differ from wild type in each of these three characteristics is a deletion of approximately 100 kilobases at one end of chromosome 2. This deletion includes the gene coding for KAHRP and the subtelomeric repeat designated rep20.Subtelomeric deletions in cultured isolates are known to result in the loss of a number of functional genes of Plasmodium falciparum (1-7). To examine whether this phenomenon occurs in nature or is an artifact of growth in vitro, we used two-dimensional pulsed-field gradient electrophoresis (PFGE) to examine chromosomal DNA from fresh field isolates. This technique allows resolution of telomeric fragments after digestion of separated chromosomes with the restriction endonuclease Apa I. It relies on the observation that a conserved complex consisting of telomeric repeats, an Apa I site(s), and a block of rep20 subtelomeric repeats is common to all chromosomes of P. falciparum (Fig.

False-Negative Rapid Diagnostic Tests for Malaria and Deletion of the Histidine-Rich Repeat Region of the hrp2 Gene

“…These results confirm that hrp2 deletions occur under field conditions as well as in the laboratory [14][15][16] and show that P. falciparum parasites without the histidine-rich repeat region of the hrp2 gene or its HRP2 gene product produce bloodstream infection in sub-Saharan Africa. In addition, recent reports indicate that hrp2 deletions are present in both Papua New Guinea and the Amazon region of Peru 16,17 and thus, suggest that false-negative results for RDTs based on HRP2 may also occur in those regions. However, it is not clear whether parasites without hrp2 are at a selective disadvantage or whether they are more frequent at times of intense transmission, such as the rainy season, when these studies were performed in Mali.…”

“…Although the negative PCR results could have resulted from sequence variability in the upstream and downstream regions used for the primers, that finding alone would not produce a negative RDT for HRP2. For that reason, because we are not aware of sequence variation in those regions that interfere with this PCR and because previous reports detail hrp2 deletions from Papua New Guinea 16 and Peru, 17 these results suggest that spontaneous deletions of hrp2 occur in many (perhaps most) areas with P. falciparum transmission.…”

Selection for high-level chloroquine resistance results in deamplification of the pfmdr1 gene and increased sensitivity to mefloquine in Plasmodium falciparum.

“…Plasmodiumfalciparum FAC8 (Biggs et al, 1989) is a chloroquine resistant clone of ITG2F6 (gift of L.Miller) which is itself a cloned line. Isolates 3D7 and HB3 are both chloroquine sensitive.…”

“…In this paper three moderately chloroquine resistant P.falciparum isolates which carried an amplification of the pfindrl gene (Foote et al, 1989;Wilson et al, 1989) and which also over-produced Pghl (Biggs et al, 1989) and it can be grown in 15 ng/ml chloroquine without inhibition of growth. FAC8 cells were subjected to increasing drug pressure by the addition of 30 ng/ml chloroquine to the medium, resulting in a cell line designated FAC815.…”

Infectivity of Plasmodium falciparum in Malaria-Naive Individuals Is Related to Knob Expression and Cytoadherence of the Parasite

“…The major structural protein in knobs is the knob-associated histidinerich protein (KAHRP) (2,6,7). Deletion of the gene encoding KAHRP results in the loss of knobs (8,9).…”