Open Access

In vitro susceptibility to quinine and microsatellite variations of the Plasmodium falciparum Na+/H+ exchanger (Pfnhe-1) gene: the absence of association in clinical isolates from the Republic of Congo

  • Sébastien Briolant1,
  • Stéphane Pelleau2,
  • Hervé Bogreau1,
  • Philippe Hovette3,
  • Agnès Zettor1,
  • Jacky Castello3,
  • Eric Baret1,
  • Rémy Amalvict1,
  • Christophe Rogier1 and
  • Bruno Pradines1Email author
Malaria Journal201110:37

https://doi.org/10.1186/1475-2875-10-37

Received: 22 October 2010

Accepted: 11 February 2011

Published: 11 February 2011

Abstract

Background

Quinine is still recommended as an effective therapy for severe cases of Plasmodium falciparum malaria, but the parasite has developed resistance to the drug in some cases. Investigations into the genetic basis for quinine resistance (QNR) suggest that QNR is complex and involves several genes, with either an additive or a pairwise effect. The results obtained when assessing one of these genes, the plasmodial Na+/H+ exchanger, Pfnhe-1, were found to depend upon the geographic origin of the parasite strain. Most of the associations identified have been made in Asian strains; in contrast, in African strains, the influence of Pfnhe on QNR is not apparent. However, a recent study carried out in Kenya did show a significant association between a Pfnhe polymorphism and QNR. As genetic differences may exist across the African continent, more field data are needed to determine if this association exists in other African regions. In the present study, association between Pfnhe and QNR is investigated in a series of isolates from central Africa.

Methods

The sequence analysis of the polymorphisms at the Pfnhe-1 ms4760 microsatellite and the evaluation of in vitro quinine susceptibility (by isotopic assay) were conducted in 74 P. falciparum isolates from the Republic of Congo.

Results

Polymorphisms in the number of DNNND or NHNDNHNNDDD repeats in the Pfnhe-1 ms4760 microsatellite were not associated with quinine susceptibility.

Conclusions

The polymorphism in the microsatellite ms4760 in Pfnhe-1 that cannot be used to monitor quinine response in the regions of the Republic of Congo, where the isolates came from. This finding suggests that there exists a genetic background associated with geographic area for the association that will prevent the use of Pfnhe as a molecular marker for QNR. The contribution of Pfnhe to the in vitro response to quinine remains to be assessed in other regions, including in countries with different levels of drug pressure.

Background

Quinine (QN) remains a first-line drug for the treatment of severe malaria that is still used as a second-line therapy for uncomplicated malaria in many countries [1]. Despite the efficacy of QN against chloroquine-resistant Plasmodium falciparum isolates, reports of QN resistance (QNR) have been increasing. In the 1980s, the frequency of clinical failures increased in Southeast Asia and Africa [24]. The mechanism of QNR is complex, multigenic, and not well understood. QNR has been associated with mutations in both the P. falciparum multidrug resistance gene mdr1 (Pfmdr1) [5] and the chloroquine resistance transporter gene Pfcrt[6]. Other genetic polymorphisms, such as mutations in the resistance protein gene Pfmrp[7] and variations of microsatellite length in the sodium/hydrogen exchanger gene Pfnhe-1[8], may contribute to QNR. Using quantitative trait loci (QTL) on the genetic cross of HB3 and Dd2 strains, Ferdig et al identified genes on chromosome 5, encoding Pfmdr1, on chromosome 7, encoding Pfcrt and on chromosome 13, encoding Pfnhe-1, which were associated with QN reduced susceptibility [8]. Sequences of Pfnhe-1 showed multiple and complex variations such as point polymorphisms at three separate codons (790, 894 and 950) and as microsatellite variations in three different repeat sequences (msR1, ms3580 and ms4760). However, the three point polymorphisms and microsatellite polymorphisms msR1 and ms3580 showed no significant association with QN susceptibility. The investigations of the microsatellite ms4760 polymorphisms in culture-adapted isolates from around the world showed that an increased number of the amino acid motif DNNND was associated with a decreased QN susceptibility, and that an increased number of NHNDNHNNDDD motifs was associated with an increased QN susceptibility [8, 9]. The association of two repeats with a high QN inhibitory concentration of 50% (IC50) was found in a case of QN clinical failure in a traveller from Senegal [10]. In contrast, a recent multivariate analysis performed on 83 freshly collected clinical isolates from Madagascar and 13 African countries did not find an association between QN susceptibility and Pfnhe-1 microsatellite polymorphisms [11]. A similar absence of association was observed in 91 isolates from various countries on different continents (Pelleau S. et al. submitted). Given that the influence of Pfnhe on QN susceptibility has been shown to be strain-dependent, these apparently conflicting results may be explained, in part, by differences in the geographic origin of the parasites analysed, as their local selection history and genetic background varies.

Thus, further epidemiological investigation is required to determine the context in which Pfnhe can be used as a molecular marker of QNR. In this work, ms4760 polymorphism was analized and its association tested with in vitro susceptibility to QN in African isolates from a single geographical region, Pointe-Noire in the Republic of Congo.

Methods

Reference culture-adapted strains and clinical isolates of Plasmodium falciparum

Between March 2005 and January 2006, 74 P. falciparum clinical isolates were collected from patients with uncomplicated malaria at the Medical Service of Total Exploration et Production, Pointe-Noire (Republic of Congo) [12]. Two cloned strains of P. falciparum were used for quality control (3D7 Africa and W2 Indochina). These clones were obtained from MR4-ATCC (Manassas, VA, USA). They were maintained in culture in RPMI 1640 (Invitrogen, Paisley, UK) that was supplemented with 10% human serum (Abcys S.A. Paris, France) and buffered with 25 mM HEPES and 25 mM NaHCO3. The parasites were grown in type A+ human blood under controlled atmospheric conditions (10% O2, 5% CO2, and 85% N2) at 37°C with a humidity of 95%. They were synchronized twice with sorbitol before use [13]. Clonality was verified using PCR genotyping of the polymorphic genetic markers, msp1 and msp2, and the microsatellite loci [14, 15].

In vitro assay

The QN was purchased from Sigma (Saint Louis, MO, USA). The QN was first dissolved in methanol and then diluted in water to obtain final concentrations ranging from 5 to 3200 nM.

For the in vitro isotopic microtest, 200 μl of the suspension of synchronous parasitized red blood cells (final parasitaemia, 0.5%; final haematocrit, 1.5%) per well were plated in 96-well plates that contained serial QN concentrations. Parasite growth was assessed by adding 1 μCi of tritiated hypoxanthine with a specific activity of 14.1 Ci/mmol (Perkin-Elmer, Courtaboeuf, France) to each well at time zero. The plates were then incubated for 48 h under controlled atmospheric conditions. Immediately after incubation, the plates were frozen and then thawed to lyse the erythrocytes. The contents of each well were collected on standard filter microplates (Unifilter GF/B; Perkin-Elmer) and washed using a cell harvester (Filter-Mate Cell Harvester; Perkin-Elmer). The filter microplates were dried and 25 μl of a scintillation cocktail (Microscint O; Perkin-Elmer) was added to each well. The radioactivity incorporated by the parasites was then measured using a scintillation counter (Top Count; Perkin-Elmer). The drug concentration that could inhibit 50% of the parasite growth (IC50) was calculated assuming that it corresponded to the drug concentration at which the incorporation of tritiated hypoxanthine by the parasite was 50% of that incorporated in the drug-free, control wells. IC50 values were determined using a non-linear regression analysis of log-based dose-response curves (Riasmart, Packard, Meriden, USA).

Genotyping of the Pfnhe ms4760 microsatellite polymorphisms

Parasite DNA from 100 μl of infected blood was extracted using the E.Z.N.A. Blood DNA kit (Omega Bio-Tek, GA, USA). A sequence containing the previously described ms4760 microsatellite [8] was amplified using the pfnhe-3802F 5'-TTATTAAATGAATATAAAGA-3' and pfnhe-4322R 5'-TTTTTTATCATTACTAAAGA-3' primers. Sequencing was performed using ABI Prism Big Dye Terminator v1.1 Cycle Sequencing Ready Reaction Kits (Applied Biosystems, CA, USA), as directed by the manufacturer. Sequences were analysed with BioEdit sequence alignment editor (version 7.0.9.0) software.

Statistical analysis

Data were analysed using R software (version 2.10.1) and GraphPad Prism (version 5.01). Differences between the median QN IC50 values of isolates harbouring one, two or three DNNND repeats were tested using the Kruskal-Wallis test. The median QN IC50 values of isolates with one or two NHNDNHNNDDD repeats were compared using the Mann Whitney test.

Results

The mean QN IC50 of the 74 isolates was 381.9 nM, CI95% [330.7-433.1] (min-max: 36-1097 nM). Four isolates (5.4%) had a QN IC50 > 800 nM (Table 1) that corresponds to the in vitro QNR threshold. Twenty-seven different Pfnhe-1 genotypes were observed among the 74 isolates, and eighteen were identified as new genotypes (GenBank accession numbers FJ392810 to FJ392827). The DNNND repeats varied from 1 to 3. There was no statistically significant association (p = 0.84) between the median QN IC50 value and the number of DNNND repeats (medians at 335, 316 and 330 nM, for isolates harbouring one, two or three repeats, respectively) (Figure 1). No statistically significant association was observed (p = 0.92) between the median QN IC50 value of isolates and the number of NHNDNHNNDDD repeats (medians at 328 and 329 nM, for isolates with one and two repeats, respectively).
Table 1

In vitro susceptibility and Pfnhe-1 polymorphisms of the 74 Plasmodium falciparum isolates from the Republic of Congo

Isolates

Cpm

without drug

Cpm

With highest QN

concentration

QN IC50

No DNNND repeat

No NHNDNHNNDDD

repeat

No genotype profile

16517

23404

1440

1097

3

2

ms4760-27

16612

49650

8273

888

1

2

ms4760-3

16520

6512

1077

882

1

2

ms4760-3

16094

2007

620

862

2

1

ms4760-24

16980

2151

980

782

2

2

ms4760-33

17267

2558

455

776

2

2

ms4760-1

16631

1807

282

750

1

2

ms4760-3

16698

1263

353

740

2

1

ms4760-6

16960

2011

714

689

2

2

ms4760-1

16958

19732

861

645

3

2

ms4760-30

16983

3710

943

627

3

2

ms4760-1

16957

19773

1300

615

1

2

ms4760-29

16966

2264

545

604

2

2

ms4760-31

16941

18540

1175

576

1

2

ms4760-3

16991

2804

815

567

1

1

ms4760-36

16953

5675

1249

564

1

2

ms4760-12

16539

44532

4112

537

1

2

ms4760-3

17250

6558

606

506

1

2

ms4760-3

16942

10927

1163

502

1

2

ms4760-3

16959

9886

914

488

2

2

ms4760-1

17008

2151

961

482

1

2

ms4760-35

16986

2302

624

481

2

2

ms4760-1

16968

8477

962

457

2

2

ms4760-1

17249

1276

412

408

1

2

ms4760-22

17216

3179

539

404

1

2

ms4760-38

17253

9717

1091

396

2

1

ms4760-6

16987

7232

938

388

3

1

ms4760-15

16349

28606

2048

377

3

1

ms4760-25

17231

3545

389

354

1

2

ms4760-39

16518

8912

1361

351

3

1

ms4760-7

16955

9379

1188

346

3

2

ms4760-27

17211

6451

655

346

1

2

ms4760-3

16695

27421

2316

341

1

2

ms4760-3

16996

1347

597

340

2

2

ms4760-34

17208

18356

1208

335

1

2

ms4760-3

16536

6617

847

333

2

2

ms4760-1

16365

4631

724

332

1

2

ms4760-22

16979

7449

990

326

2

2

ms4760-18

17202

11053

929

319

1

2

ms4760-3

16364

12210

1990

316

2

2

ms4760-26

16331

8912

1113

314

3

2

ms4760-9

15868

1530

498

309

1

2

ms4760-22

16610

4063

961

305

3

1

ms4760-7

16535

16007

1274

291

3

1

ms4760-15

16323

43074

5859

290

1

2

ms4760-12

16992

9464

1111

276

1

2

ms4760-12

17237

12385

873

269

1

2

ms4760-3

17305

21667

1398

269

2

2

ms4760-3

16305

11269

917

265

1

2

ms4760-3

17303

18553

1646

264

2

2

ms4760-1

17212

2755

520

259

1

2

ms4760-3

16304

16194

738

254

2

2

ms4760-1

17241

5758

447

254

1

2

ms4760-3

16970

2359

705

247

1

1

ms4760-32

16995

7642

1004

247

2

2

ms4760-1

17210

11018

1370

244

1

2

ms4760-22

17233

12100

956

231

2

1

ms4760-6

17265

1593

238

226

2

2

ms4760-1

17214

2421

316

224

2

2

ms4760-37

17209

3499

680

216

3

2

ms4760-9

17007

3633

1159

212

3

2

ms4760-9

16116

11417

999

204

2

2

ms4760-18

17201

25594

2530

204

1

1

ms4760-36

17304

24453

1982

199

2

2

ms4760-1

16086

10918

699

174

3

1

ms4760-15

17283

4391

315

146

3

2

ms4760-1

17010

2897

1103

141

3

2

ms4760-9

16117

17474

1291

121

2

2

ms4760-1

16078

1654

396

106

2

2

ms4760-1

16085

5229

707

86

2

1

ms4760-6

17301

39337

2016

84

1

2

ms4760-12

15867

6893

739

73

1

2

ms4760-3

16091

17320

1695

67

2

2

ms4760-1

16077

4539

353

36

1

2

ms4760-23

Cpm = count per minute

IC50 = Inhibitory concentration 50% in nM

QN = quinine

Pfnhe- 1 ms4760 microsatellite genotype profiles according to the GenBank

Figure 1

Distribution of quinine (QN) IC 50 versus the repeats number of DNNND or NHNDNHNNDDD. The horizontal bars indicate median values.

Discussion

QN has been used to treat malaria for more than 350 years in Africa, with little emergence and spread of resistance. QN remains the first line anti-malarial drug for the treatment of complicated malaria in Europe and Africa. However, despite the efficacy of QN against chloroquine-resistant strains, the emergence of QN resistance has been observed. The first cases of QN clinical failure were observed in Brazil and Asia in the 1960s; then in the 1980s, clinical failures became more frequent in Southeast Asia, South America and Africa. However, QN resistance is not yet a significant problem in Africa, and QN remains the first-line drug to treat severe malaria and a second-line therapy for uncomplicated malaria in some areas of Africa.

Although some reports of QN treatment failure exist, it is difficult to confirm QN resistance because of the short elimination half-life of the drug, the requirement to administer it three times a day for at least five days, drug intolerance that often leads to poor compliance and a lack of reliable data on the correlation between QN IC50 and clinical failure. Maximizing the efficacy and longevity of QN as a tool for the control of malaria will depend critically on the pursuit of intensive research toward the identification of in vitro markers of QNR and the implementation of in vitro and in vivo surveillance programs, such as those championed by the Worldwide Antimalarial Resistance Network [16, 17]. Specifically, there is a need to identify molecular markers that effectively predict QN resistance and enable the active surveillance of temporal trends in parasite susceptibility [18]. The present study aimed to test the association between the Pfnhe polymorphism and QN susceptibility in clinical isolates from Pointe-Noire in Congo to assess the validity of Pfnhe as a molecular marker of QN susceptibility in this region.

The level of susceptibility to QN remained high, with a mean QN IC50 of 382 nM and with 4/74 isolates (5.4%) that exceed the QNR threshold of 800 nM [19]. The threshold at which QN in vitro susceptibility is considered to be compromised is arbitrary, and different studies have set the following thresholds: 300 nM [20], 500 nM [21] or 800 nM. If a QNR threshold of 500 nM is applied here, 19/74 isolates exceed this value. These results suggest that QN treatment is still effective against the chloroquine-resistant parasites that are found in this area, where the prevalence of chloroquine-resistant parasites has reached 75% [12].

A high level of Pfnhe-1 microsatellite sequence polymorphisms was found (27 genotypes for 74 isolates). Previous studies reported from 8 genotypes for 71 isolates [8] to 35 for 60 isolates [22]. DNNND and NHNDNHNNDDD repeat numbers ranged from 1 to 3 and 1 to 2, respectively. However, neither DNNND nor NHNDNHNNDDD repeat polymorphisms were linked to QN susceptibility.

According to this study and previous studies, the association between QN in vitro susceptibility and the Pfnhe-1 microsatellite genotype appears to be geographically dependent.

One explanation for this could be variation in genetic background. A specific genetic background observed in Asia may allow the observed contribution of Pfnhe polymorphism on QN in vitro susceptibility. This explanation is consistent with: i) the first evidence of Pfnhe-QNR association from QTL analysis using Americano-Asian cross strains [8], ii) that most associations identified have been shown among Asian strains [8, 9, 22] and iii) that at least 5 genes spanning the P. falciparum genome influence the QNR in vitro phenotype with an additive effect or with pairwise interactions [8].

Additionally, these genetic dissimilarities between African and Asian plasmodial populations may be accentuated by different local selection histories. The best-documented genomic modifications by a local selection process relate to drug pressure. Several studies have shown that drug pressure may involve extended linkage disequilibrium around a drug resistance associated gene [23]. This is characterized by a strong genetic diversity loss, called a selective sweep. This process stretch may be modified by: i) drug use and ii) malaria transmission level. A reduced drug use and higher malaria transmission level in Africa would be consistent with the lower selective sweep and an absence of linkage disequilibrium between Pfnhe and other cooperative drug response genes or selected compensatory mutations. For example, QTL analysis on chromosome 13 located 60 genes of unknown function as being close to Pfnhe[8]. They would be more or less linked depending on drug pressure and malaria transmission level if QN pressure selects at least one of them. In Kenya there is an association between two DNNND repeats in the ms4760 Pfnhe microsatellite and a reduced susceptibility to quinine in 29 P. falciparum isolates [24]. This is consistent with the historic precedent of the spread of drug resistance around the world. The emergence of chloroquine resistance in Asia was followed by an initial introduction into East Africa and spreading across the African continent. Geographic proximity may explain plasmodial population migration. Moreover, East African plasmodial populations may exhibit genetic dissimilarities to other African populations [14].

As the QN response is controlled by multiple genes with complex interactions, one can expect: i) higher sensitivity to genetic background than if the response was controlled by only one gene, ii) higher sensitivity to parameters that may cause linkage disequilibrium between genes and iii) various combinations of gene polymorphisms may result in similar QNR phenotypes.

Conclusions

In summary, although studies have demonstrated that Pfnhe-1 contributes to QNR [24], the present study and another recent study [11] did not show an association between Pfnhe-1 polymorphism and QNR. Currently, Pfnhe-1 cannot be used as a QNR molecular marker when evaluating field isolates, especially in Africa in the context of a low level of quinine selection pressure. Further studies, using more parasite samples from South East Asia with reduced susceptibility to QN, are required to confirm or reject the use of the Pfnhe-1 gene as a QNR marker in this geographic region.

Conflict of interests

The authors declare that they have no competing interests.

Declarations

Acknowledgements

The authors thank the patients and staff of the Clinique Total of Total Exploration et Production Congo in Pointe-Noire and the Département Médical International of Total in Paris. This work was supported by the Direction Centrale du Service de Santé des Armées. SP was supported by a grant from the Fondation des Treilles.

Authors’ Affiliations

(1)
Unité de Parasitologie, Unité Mixte de Recherche 6236, Institut de Recherche Biomédicale des Armées - Antenne de Marseille
(2)
Unité de Recherche en Pharmacologie et Physiopathologie Parasitaires, UMR MD3 Relation hôte-parasite, Pharmacologie et Thérapeutique, Institut de Recherche Biomédicale des Armées - Antenne de Marseille
(3)
Clinique Total, Division Médicale de Total Exploration et Production Congo Pointe-Noire

References

  1. World Health Organization:: WHO guidelines for the treatment of malaria in travellers. chapters 7 (section 9.4) and 8, [http://www.who.int/malaria/docs/TreatmentGuidelines2006.pdf]
  2. Harinasuta T, Bunnag D, Lasserre R: Quinine resistant falciparum malaria treated with mefloquine. Southeast Asian J Trop Med Public Health. 1990, 21: 552-557.PubMedGoogle Scholar
  3. Jelinek T, Schelbert P, Loscher T, Eichenlaub D: Quinine resistant falciparum malaria acquired in east Africa. Trop Med Parasitol. 1995, 46: 38-40.PubMedGoogle Scholar
  4. Palmieri F, Petrosillo N, Paglia MG, Conte A, Goletti D, Pucillo LP, Menegon M, Sannella A, Severini C, Majori G: Genetic confirmation of quinine-resistant Plasmodium falciparum malaria followed by postmalaria neurological syndrome in a traveler from Mozambique. J Clin Microbiol. 2004, 42: 5424-5426. 10.1128/JCM.42.11.5424-5426.2004.PubMed CentralView ArticlePubMedGoogle Scholar
  5. Reed MB, Saliba KJ, Caruana SR, Kirk K, Cowman AF: Pgh1 modulates sensitivity and resistance to multiple antimalarials in Plasmodium falciparum. Nature. 2000, 403: 906-909. 10.1038/35002615.View ArticlePubMedGoogle Scholar
  6. Cooper RA, Lane KD, Deng B, Mu J, Patel JJ, Wellems TE, Su X, Ferdig MT: Mutations in transmembrane domains 1, 4 and 9 of the Plasmodium falciparum chloroquine resistance transporter alter susceptibility to chloroquine, quinine and quinidine. Mol Microbiol. 2007, 63: 270-282. 10.1111/j.1365-2958.2006.05511.x.View ArticlePubMedGoogle Scholar
  7. Mu J, Ferdig MT, Feng X, Joy DA, Duan J, Furuya T, Subramanian G, Aravind L, Cooper RA, Wootton JC, Xiong M, Su XZ: Multiple transporters associated with malaria parasite responses to chloroquine and quinine. Mol Microbiol. 2003, 49: 977-989. 10.1046/j.1365-2958.2003.03627.x.View ArticlePubMedGoogle Scholar
  8. Ferdig MT, Cooper RA, Mu J, Deng B, Joy DA, Su XZ, Wellems TE: Dissecting the loci of low-level quinine resistance in malaria parasites. Mol Microbiol. 2004, 52: 985-997. 10.1111/j.1365-2958.2004.04035.x.View ArticlePubMedGoogle Scholar
  9. Henry M, Briolant S, Zettor A, Pelleau S, Baragatti M, Baret E, Mosnier J, Amalvict R, Fusai T, Rogier C, Pradines B: Plasmodium falciparum Na+/H+ exchanger 1 transporter is involved in reduced susceptibility to quinine. Antimicrob Agents Chemother. 2009, 53: 1926-1930. 10.1128/AAC.01243-08.PubMed CentralView ArticlePubMedGoogle Scholar
  10. Pradines B, Pistone T, Ezzedine K, Briolant S, Bertaux L, Receveur MC, Parzy D, Millet P, Rogier C, Malvy D: Quinine-resistant malaria in traveler returning from Senegal, 2007. Emerg Infect Dis. 2010, 16: 546-548. 10.3201/eid1603.091669.PubMed CentralView ArticlePubMedGoogle Scholar
  11. Andriantsoanirina V, Menard D, Rabearimanana S, Hubert V, Bouchier C, Tichit M, Bras JL, Durand R: Association of microsatellite variations of Plasmodium falciparum Na+/H+ exchanger (Pfnhe-1) gene with reduced in vitro susceptibility to quinine: lack of confirmation in clinical isolates from Africa. Am J Trop Med Hyg. 2010, 82: 782-787. 10.4269/ajtmh.2010.09-0327.PubMed CentralView ArticlePubMedGoogle Scholar
  12. Pradines B, Hovette P, Fusai T, Atanda HL, Baret E, Cheval P, Mosnier J, Callec A, Cren J, Amalvict R, Gardair JP, Rogier C: Prevalence of in vitro resistance to eleven standard or new antimalarial drugs among Plasmodium falciparum isolates from Pointe-Noire, Republic of the Congo. J Clin Microbiol. 2006, 44: 2404-2408. 10.1128/JCM.00623-06.PubMed CentralView ArticlePubMedGoogle Scholar
  13. Lambros C, Vanderberg JP: Synchronization of Plasmodium falciparum erythrocytic stages in culture. J Parasitol. 1979, 65: 418-420. 10.2307/3280287.View ArticlePubMedGoogle Scholar
  14. Bogreau H, Renaud F, Bouchiba H, Durand P, Assi SB, Henry MC, Garnotel E, Pradines B, Fusai T, Wade B, Adehossi E, Parola P, Kamil MA, Puijalon O, Rogier C: Genetic diversity and structure of African Plasmodium falciparum populations in urban and rural areas. Am J Trop Med Hyg. 2006, 74: 953-959.PubMedGoogle Scholar
  15. Henry M, Diallo I, Bordes J, Ka S, Pradines B, Diatta B, M'Baye PS, Sane M, Thiam M, Gueye PM, Wade B, Touze JE, Debonne JM, Rogier C, Fusai T: Urban malaria in Dakar, Senegal: chemosusceptibility and genetic diversity of Plasmodium falciparum isolates. Am J Trop Med Hyg. 2006, 75: 146-151.PubMedGoogle Scholar
  16. Sibley CH, Barnes KI, Plowe CV: The rationale and plan for creating a World Antimalarial Resistance Network (WARN). Malar J. 2007, 6: 118-10.1186/1475-2875-6-118.PubMed CentralView ArticlePubMedGoogle Scholar
  17. Sibley CH, Barnes KI, Watkins WM, Plowe CV: A network to monitor antimalarial drug resistance: a plan for moving forward. Trends Parasitol. 2008, 24: 43-48. 10.1016/j.pt.2007.09.008.View ArticlePubMedGoogle Scholar
  18. Plowe CV, Roper C, Barnwell JW, Happi CT, Joshi HH, Mbacham W, Meshnick SR, Mugittu K, Naidoo I, Price RN, Shafer RW, Sibley CH, Sutherland CJ, Zimmerman PA, Rosenthal PJ: World Antimalarial Resistance Network (WARN) III: molecular markers for drug resistant malaria. Malar J. 2007, 6: 121-10.1186/1475-2875-6-121.PubMed CentralView ArticlePubMedGoogle Scholar
  19. Ralaimazava P, Durand R, Godineau N, Keundjian A, Jezic Z, Pradines B, Bouchaud O, Le Bras J: Profile and evolution of the chemosusceptibility of falciparum malaria imported into France in 2000. Euro Surveill. 2002, 7: 113-118.PubMedGoogle Scholar
  20. Brasseur P, Kouamouo J, Moyou-Somo R, Druilhe P: Multi-drug resistant falciparum malaria in Cameroon in 1987-1988. I. Stable figures of prevalence of chloroquine- and quinine-resistant isolates in the original foci. Am J Trop Med Hyg. 1992, 46: 1-7.PubMedGoogle Scholar
  21. Pradines B, Rogier C, Fusai T, Tall A, Trape JF, Doury JC: Sensibilité in vitro de 85 isolats de Plasmodium falciparum au Sénégal. Med Trop. 1996, 56: 141-145.Google Scholar
  22. Meng H, Zhang R, Yang H, Fan Q, Su X, Miao J, Cui L, Yang Z: In vitro sensitivity of Plasmodium falciparum clinical isolates from the China-Myanmar border area to quinine and association with polymorphism in the Na+/H+ exchanger. Antimicrob Agents Chemother. 2010, 54: 4306-4313. 10.1128/AAC.00321-10.PubMed CentralView ArticlePubMedGoogle Scholar
  23. Wootton JC, Feng X, Ferdig MT, Cooper RA, Mu J, Baruch DI, Magill AJ, Su XZ: Genetic diversity and chloroquine selective sweeps in Plasmodium falciparum. Nature. 2002, 418: 320-323. 10.1038/nature00813.View ArticlePubMedGoogle Scholar
  24. Nkrumah LJ, Riegelhaupt PM, Moura P, Johnson DJ, Patel J, Hayton K, Ferdig MT, Wellems TE, Akabas MH, Fidock DA: Probing the multifactorial basis of Plasmodium falciparum quinine resistance: evidence for a strain-specific contribution of the sodium-proton exchanger PfNHE. Mol Biochem Parasitol. 2009, 165: 122-131. 10.1016/j.molbiopara.2009.01.011.PubMed CentralView ArticlePubMedGoogle Scholar

Copyright

© Briolant et al; licensee BioMed Central Ltd. 2011

This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.