In vitro assessment of artesunate, artemether and amodiaquine susceptibility and molecular analysis of putative resistance‐associated mutations of Plasmodium falciparum from São Tomé and Príncipe

Abstract

Summaryobjective To evaluate the basal in vitro responses of Plasmodium falciparum isolates collected in The Democratic Republic of Sã o Tomé and Príncipe to artemether (ATH), artesunate (ATN) and amodiaquine (AMQ).methods The prevalence of given single nucleotide polymorphisms in the pfmdr1, pfcrt, pftctp and pfATPase6 genes 1 was assessed by PCR-RFLP or DNA sequencing, and gene copy numbers were estimated by real-time PCR.results Mean IC50s to ATH and ATN were relatively low (1.12 nm and 0.58 nm, respectively). However, 10% of parasites displayed AMQ IC50 values above the accepted resistance threshold of 60 nm and there was a positive association between susceptibility to all three drugs (ATH vs. ATN: R ¼ 0.84; ATH vs. AMQ: R ¼ 0.68; ATN vs. AMQ: R ¼ 0.72). Mutations in the pfcrt and pfmdr1 genes were highly prevalent, while only one synonymous polymorphism was detected in the pfATPase6 gene and no mutations were found in pftctp. All isolates harboured a single copy of the genes studied.conclusions Artemisinin combination treatment in the Sã o Tomé and Príncipe should be efficacious, although a significant number of AMQ-resistant parasites were detected and the susceptibility to each drug was positively associated with that of the other two. Mutations in the pfcrt and pfmdr1 genes are near fixation, most likely because of high levels of chloroquine resistance, whereas only one protein type of the artemisinin resistance candidate, PfATPase6, was identified.

In vitro susceptibility of Plasmodium falciparum Welch field isolates to infusions prepared from Artemisia annua L. cultivated in the Brazilian Amazon

“…The IC 50 values obtained for artemisinin against Amazonian field isolates exhibited low variation (average IC 50 = 2.1 ± 1.0 nM). Given the number of samples studied, the levels of sensitivity of Amazonian P. falciparum field isolates to artemisinin are in agreement with results for other field isolates reported in the literature (Jambou et al 2005, Ferreira et al 2007, Gama et al 2011). In Fig.…”

“…However, two samples revealed high IC 50 values (224 and 259 nM) that were lower than the in vitro threshold value for quinine resistance (500 nM) established by the WHO (2007). The levels of field isolate parasite inhibition (IC 50 = 2.1 ± 1.0 nM) by artemisinin are generally in agreement with data from other parts of South America and Brazil (Jambou et al 2005, Ferreira et al 2007. The drug susceptibility of field isolates was evaluated as a means to characterise the resistance profiles of the samples of Amazonian parasites utilised to evaluate their susceptibility to infusions.…”

“…Information on the epidemiology of resistance can be applied to the monitoring and control of the appearance and propagation of chemoresistance in populations of natural parasites. This monitoring can be performed with drugs in use in a particular location or with new drug alternatives, especially when the parasite exhibits cross-resistance to drugs (Ferreira et al 2007, Chiyaka et al 2009). The stabilised field isolates developed and examined in the present study exhibited similar sensitivity profiles to chloroquine, quinine and artemisinin.…”

Polymorphism of Pfatpase6 in Cte dIvoire: Detection of a four new point mutations

“…These point mutations are new compared to the data available on the polymorphism in PfATPase6 gene. Similar studies were conducted in Sao Tome and Principe and showed a silent mutation of T2694A (Ferreira et al, 2007). Ferreira et al (2008) also published a similar paper with Brazilian samples (Ferreira et al, 2008).…”

Molecular and Biological Aspects of Antimalarial Resistance in Plasmodium falciparum and Plasmodium vivax

“…IC 50 values observed in many studies from South America appear to be similar to those observed in Africa and different from those in SE-Asia, where chemotolerance appears to be higher. As an example, a recent study from Laos has reported an IC 50 to Artesunate of 5.0 nm among local P. falciparum populations, a value that is more than fivefold higher than the one observed in Brazil (0.85 nm) by Ferreira et al [109,110]. In contrast, African parasites appear to be more susceptible to ART derivatives with IC 50 s to Artesunate of 0.58, 0.73 and 2.2 nm from Sao Tome, Gabon and Senegal, respectively [109,111].…”