Chloroquine resistant Plasmodium vivax: in vitro characterisation and association with molecular polymorphisms.
BACKGROUND
Treatment failure of chloroquine for P. vivax infections has reached high levels in the eastern provinces of Indonesia, however, in vitro characterization of chloroquine resistance and its associated molecular profile have yet to be determined.


METHODS
Using a modified schizont maturation assay we investigated the in vitro chloroquine susceptibility profile and molecular polymorphisms of P. vivax isolates collected from Papua, Indonesia, where high levels of clinical chloroquine treatment failure have been reported, and from Thailand, where chloroquine treatment is generally effective.


RESULTS
The geometric mean chloroquine IC(50) for P. vivax isolates from Papua (n = 145) was 312 nM [95%CI: 237-411 nM] compared to 46.8 nM [95%CI: 34.7-63.1 nM] from Thailand (n = 81); p<0.001. Correlating with the known clinical efficacy of the area, a cut off for chloroquine resistance was defined as 220 nM, a level exceeded in 13.6% (11/81) of Thai isolates and 65% (94/145) of Papuan isolates; p<0.001. Several sequence polymorphisms in pvcrt-o and pvmdr1, and difference in pvmdr1 copy number were identified. A Y976F mutation in pvmdr1 was present in 96% (123/128) of Papuan isolates and 25% (17/69) of Thai isolates; p<0.001. Overall, the geometric mean chloroquine IC(50) in isolates with the Y976F mutation was 283 nM [95%CI: 211-379], compared to 44.5 nM [95%CI: 31.3-63.4] in isolates with the wild type; p< 0.001. Pvmdr1 amplification occurred in 23% (15/66) of Thai isolates compared to none (0/104) of Indonesian isolates (p<0.001), but was not associated with increased chloroquine resistance after controlling for geographical location.


CONCLUSIONS
In vitro susceptibility testing of P. vivax discriminates between populations with differing levels of clinical efficacy of chloroquine. The pvmdr1 polymorphism at Y976F may provide a useful tool to highlight areas of emerging chloroquine resistance, although further studies defining its clinical correlates are needed.
Suwanarusk
Rossarin
R
International Health Program, Infectious Diseases Division, Menzies School of Health Research and Charles Darwin University, Darwin, Australia.
Russell
Bruce
B
Chavchich
Marina
M
Chalfein
Ferryanto
F
Kenangalem
Enny
E
Kosaisavee
Varakorn
V
Prasetyorini
Budi
B
Piera
Kim A
KA
Barends
Marion
M
Brockman
Alan
A
Lek-Uthai
Usa
U
Anstey
Nicholas M
NM
Tjitra
Emiliana
E
Nosten
François
F
Cheng
Qin
Q
Price
Ric N
RN
D000818:Animals; D000962:Antimalarials / Q000494:pharmacology*; D002738:Chloroquine / Q000494:pharmacology*; D004351:Drug Resistance; D014644:Genetic Variation; D006801:Humans; D007214:Indonesia; D020128:Inhibitory Concentration 50; D016780:Malaria, Vivax / Q000469:parasitology*; D008954:Models, Biological; D021261:Parasitic Sensitivity Tests; D010966:Plasmodium vivax / Q000378:metabolism*; D011110:Polymorphism, Genetic; D020641:Polymorphism, Single Nucleotide; D015800:Protozoan Proteins / Q000737:chemistry
D000962:Antimalarials; D015800:Protozoan Proteins; D002738:Chloroquine