Characterization of anti-malarial resistance genes in pregnant and non-pregnant subjects of the northwest of Colombia
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KeywordsMalaria Primaquine Intermittent Preventive Treatment Resistance Associate Gene Falciparum Sample
Surveillance of resistance to antimalarials is a priority in the light of limited therapeutic options against P. falciparum and the emergence of artemisinin resistant strains. In some countries, antifolates and 4-aminoquinolines might remain a second line choice therapy, mainly in particular clinical situations or when availability of artemisinin combined therapy (ACT) is limited. During the past decade, we have monitored the status of antimalarial resistance associated genes in the northwest region of Colombia, where both vivax and falciparum malaria are highly endemic. Treatment for P. falciparum infection in Colombia is based on ACT, and for P. vivax, on choloroquine plus primaquine. Some authors have reported pregnant subjects as an important reservoir of P. falciparum resistant clones, mainly as a consequence of Intermittent preventive treatment in pregnancy (IPTp). In the light of the recent reports made in Colombia of a higher than expected frequency of pregnancy associated malaria, efforts were directed to study the genetic characteristics of the P. falciparum and P. vivax alleles associated with antifolate resistance in pregnant and non-pregnant populations of the country.
Materials and methods
A sample size sample of 20 isolates of P. vivax and 20 of P. falciparum was proposed and grouped into 1) parturient with a positive peripheral blood test, 2) positive placental blood, 3) gestational malaria (non-parturient) and 4) positive non-pregnant subject. The genes dhfr, dhps and mdr1 were assessed for both species. For this, PCR followed by direct sequencing was performed in each gene. Analysis of the sequences was carried out using Genious 6.1.3.
Successful DNA amplification was observed in 20 P. vivax and in 15 P. falciparum samples. In 19/20 P. vivax, the triple mutant Pvdhfr (58R, 117N) and Pvdhps (383G) was detected. Similarly, for P. falciparum, 14/15 isolates exhibited the triple mutant genotype Pfdhfr (51I, 108N) and Pfdhps (437G). Assessment of the Pvmdr1 gene confirmed presence of the wild type alleles Y976 and F1076 in 19/20 samples. A double mutant (976F, 1076L) was observed in a pregnant woman. As for Pfmdr1, all samples presented the haplotype N86, 184F. Distribution of the different genotypes was similar in all groups.
Over 90% of P. falciparum samples, regardless of the group of study, exhibited mutant alleles of Pfdhps, Pfdhfr y Pfmdr1. Pvmdr1 remains wild-type in all groups studied. A triple-mutant pattern of resistance in antifolat-related genes was observed in P. vivax and P. falciparum. This renders antifolates a non-viable option for a IPTp policy in the country.
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