Logo of aacPermissionsJournals.ASM.orgJournalAAC ArticleJournal InfoAuthorsReviewers
Antimicrob Agents Chemother. 2009 Oct; 53(10): 4570–4571.
Published online 2009 Aug 17. doi:  10.1128/AAC.00684-09
PMCID: PMC2764200

New PfATP6 Mutations Found in Plasmodium falciparum Isolates from Vietnam

Lionel Bertaux*
UMR MD3
Institut de Recherche Biomédicale des Armées
Antenne de Marseille
Institut de Médecine Tropicale du Service de Santé des Armées
Allée du Médecin Colonel Eugène Jamot
Parc du Pharo, BP60109
13262 Marseille cedex 07, France
Le Hong Quang
Military Center of Preventive Medicine
168 Bis Duong Phan Van Tri Phuong 5—Quan Go Vap TP HCM
Ho Chi Minh City, Vietnam
Véronique Sinou
UMR MD3
Institut de Recherche Biomédicale des Armées
Antenne de Marseille
Institut de Médecine Tropicale du Service de Santé des Armées
Allée du Médecin Colonel Eugène Jamot
Parc du Pharo, BP60109
13262 Marseille cedex 07, France
Nguyen Xuan Thanh
Military Institute of Hygiene and Epidemiology of Hanoi
21 Trung Liet Street
Dong Da, Hanoi, Vietnam
Daniel Parzy
UMR MD3
Institut de Recherche Biomédicale des Armées
Antenne de Marseille
Institut de Médecine Tropicale du Service de Santé des Armées
Allée du Médecin Colonel Eugène Jamot
Parc du Pharo, BP60109
13262 Marseille cedex 07, France

Artemisinin and its derivatives have been used against malaria in Vietnam since 1991 (4). An increase in clinical artemisinin resistance would be disastrous for malaria treatment. All possible indicators of this potential resistance must be monitored. The sarco/endoplasmic reticulum Ca2+-ATPase ortholog of Plasmodium falciparum (PfATP6) has been suggested to be the target of artemisinins (3). Consequently, the polymorphism of PfATP6 is being monitored by several scientific research teams (2, 6, 7, 9, 10, 15) We report here the genotyping results of PfATP6 from 98 P. falciparum field isolates collected in 2006 to 2007 in South Vietnam.

Parasite samples were taken from patients (28.82 ± 12.31 years old) with uncomplicated P. falciparum infections before drug treatment. They were collected in Binh Phuoc and Dak Nong provinces in South Vietnam. Patients did not follow a chemoprophylaxis before sampling. Diagnosis was carried out by microscopic examination and confirmed by real-time PCR as previously described (14). The whole PfATP6 gene was sequenced once in both directions with five primer pairs (adapted from Jambou et al. [7]) and compared to the reference sequence of the 3D7 strain (“PFA0310c” in the genome annotation).

We found a total of eight mutations (Table (Table1):1): four nonsynonymous (I89T, N463S, N465S, and N683K), three synonymous (N460N, I898I, and C1031C), and one double deletion leading to the loss of two asparagines (Δ463-464). Five of these have not been described previously (N460N, N463S, Δ463-464, N465S, and C1031C). All of the mutations were detected on different isolates, except for I898I, which was found alone or associated with others. Like Mugittu et al. in Tanzania (10) and Zhang et al. in China (15), we did not find either the S769N mutation or the A623E E431K double mutation, associated with reduced susceptibility to artemether (7). Previously, the N683K mutation was only found in Cambodia (2), suggesting that it may be specific to P. falciparum from South-East Asia. However, we did not detect this mutation in the South-East Asiatic strains W2 and Dd2 (both from Indochina, Malaria Research and Reference Reagent Resource Center), IMT-A4 (Vietnam), and IMT-K2 (Cambodia; data not shown). Interestingly, the N460N, N463S, N465S, and N683K mutations and the Δ463-464 double deletion are in a stretch of nine asparagines located in the interspecies variable region of PfATP6, a domain specific for Plasmodium species (8). Consequently, these modifications could be adaptive changes that might alter susceptibility to artemisinins.

TABLE 1.
Diversity of PfATP6 in P. falciparum samples from Vietnam

Cojean et al. found the S769N mutation in an isolate from Africa that was susceptible to dihydroartemisinin (1), while Noedl et al. did not find this mutation in Cambodian samples that were less susceptible to artesunate (11). Consequently, we speculated on whether the correlation between the S769N mutation and the increased artemether 50% inhibitory concentration found in six isolates from French Guyana (7) should be regarded as a local case. Like other investigators, we did not detect any polymorphism in codon 263, described as the key amino acid for the interaction between PfATP6 and artemisinins (13). Mutations observed in our sequences and in those of previous studies (2, 6, 9) could be implicated indirectly in this interaction, in the case of association with artemisinin susceptibility. Considering the development of artemisinin combined therapies and the possible implication of PfATP6 in artemisinin resistance, the molecular variability of this gene should be carefully monitored.

Acknowledgments

We thank Yannick Le Priol for discussions; Nicolas Benoit, Julien Cren, and Marc Desbordes for technical help; and the Military Institute of Hygiene and Epidemiology of Hanoi, the Military Center of Analysis and Research of Hanoi, and the Military Center of Preventive Medicine of Ho Chi Minh for helpful collection of samples.

This study was supported by the French Ministry of Foreign Affairs and by the French and Vietnamese Military Health Services.

Footnotes

Published ahead of print on 17 August 2009.

REFERENCES

1. Cojean, S., V. Hubert, J. Le Bras, and R. Durand. 2006. Resistance to dihydroartemisinin. Emerg. Infect. Dis. 12:1798-1799. [PMC free article] [PubMed]
2. Dahlström, S., M. I. Veiga, P. Ferreira, A. Martenson, A. Kaneko, B. Andersson, A. Björkman, and J. P. Gil. 2008. Diversity of the sarco/endoplasmic reticulum Ca2+-ATPase orthologue of Plasmodium falciparum (PfATP6). Infect. Genet. Evol. 8:340-345. [PubMed]
3. Eckstein-Ludwig, U., R. J. Webb, I. D. A. van Goethem, J. M. East, A. G. Lee, M. Kimura, P. M. O'Neill, P. G. Bray, S. A. Ward, and S. Krishna. 2003. Artemisinins target the SERCA of Plasmodium falciparum. Nature 424:957-961. [PubMed]
4. Ettling, M. B. 2002. The control of malaria in Viet Nam from 1980 to 2000: what went right? Report of Consultancy for W.H.O. Regional Office for Western Pacific 1-20. World Health Organization, Geneva, Switzerland.
5. Ferreira, I. D., D. Lopes, A. Martinelli, C. Ferreira, V. E. do Rosario, and P. Cravo. 2007. In vitro assessment of artesunate, artemether, and amodiaquine susceptibility and molecular analysis of putative resistance-associated mutations of Plamsodium falciparum from São Tomé and Príncipe. Trop. Med. Int. Health 12:353-362. [PubMed]
6. Ibrahim, M. L., N. Khim, H. H. Adam, F. Ariey, and J. B. Duchemin. 2009. Polymorphism of PfATPase in Niger: detection of three new point mutations. Malar. J. 8:28. [PMC free article] [PubMed]
7. Jambou, R., E. Legrand, M. Niang, N. Khim, P. Lim, B. Volney, M. T. Ekala, C. Bouchier, P. Esterre, T. Fandeur, and O. Mercereau-Puijalon. 2005. Resistance of Plasmodium falciparum field isolates to in-vitro artemether and point mutations of the SERCA-type PfATPase6. Lancet 366:1960-1963. [PubMed]
8. Kimura, M., Y. Yamaguchi, S. Takada, and K. Tanabe. 1993. Cloning of a Ca2+-ATPase gene of Plasmodium falciparum and comparison with vertebrate Ca2+-ATPase. J. Cell Sci. 104:1129-1136. [PubMed]
9. Menegon, M., A. R. Sannella, G. Majori, and C. Severini. 2008. Detection of novel point mutations in the Plasmodium falciparum ATPase6 candidate gene for resistance to artemisinins. Parasitol. Int. 57:233-235. [PubMed]
10. Mugittu, K., B. Genton, H. Mshinda, and H. P. Beck. 2006. Molecular monitoring of Plasmodium falciparum resistance to artemisinin in Tanzania. Malar. J. 5:126-128. [PMC free article] [PubMed]
11. Noedl, H., Y. Se, K. Schaecher, B. L. Smith, D. Socheat, M. M. Fukuda, et al. 2008. Evidence of artemisinin-resistant malaria in western Cambodia. N. Engl. J. Med. 359:2619-2620. [PubMed]
12. Price, R. N., A. C. Uhlemann, A. Brockman, R. McGready, E. Ashley, L. Phaipun, R. Patel, K. Laing, S. Looareesuwan, N. White, F. Nosten, and S. Krishna. 2004. Mefloquine resistance in Plasmodium falciparum and increased pfmdr1 gene copy number. Lancet 364:438-447. [PMC free article] [PubMed]
13. Uhlemann, A. C., A. Cameron, U. Eckstein-Ludwig, J. Fischbarg, P. Iserovich, F. A. Zuniga, M. East, A. Lee, L. Brady, R. K. Hayes, and S. Krishna. 2005. A single amino acid residue can determine the sensitivity of SERCAs to artemisinins. Nat. Struct. Mol. Biol. 12:628-629. [PubMed]
14. Vo, T. K., P. Bigot, P. Gazin, V. Sinou, J. J. De Pina, D. C. Huynh, F. Fumoux, and D. Parzy. 2007. Evaluation of a real-time PCR assay for malaria diagnosis in patients from Vietnam and in returned travellers. Trans. R. Soc. Trop. Med. Hyg. 101:422-428. [PubMed]
15. Zhang, G., Y. Guan, B. Zheng, S. Wu, and L. Tang. 2008. No PfATPase6 S769N mutation found in Plasmodium falciparum isolates from China. Malar. J. 7:122-125. [PMC free article] [PubMed]

Articles from Antimicrobial Agents and Chemotherapy are provided here courtesy of American Society for Microbiology (ASM)
PubReader format: click here to try

Formats:

Save items

Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...

Links

  • PubMed
    PubMed
    PubMed citations for these articles
  • Substance
    Substance
    PubChem chemical substance records that cite the current articles. These references are taken from those provided on submitted PubChem chemical substance records.

Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...