Structure and Functional Differentiation of PfCRT Mutation in Chloroquine Resistance (CQR) in Plasmodium falciparum Malaria

Approximately one million deaths are attributed to malaria every year. Latest reports of multi-drug treatment failure of falciparum malaria underscore the desideratum to understand the molecular substratum of drug resistance. The mutations in the digestive vacuole transmembrane protein Plasmodium falciparum chloroquine resistance transporter (PfCRT) are mainly responsible for chloroquine resistance (CQR) in Plasmodium falciparum. Multiple mutations in the PfCRT are concerned in chloroquine resistance, but the evolution of intricate haplotypes is not yet well understood. P. falciparum resistance to chloroquine is the standard antimalarial drug and is mediated primarily by mutant forms of the PfCRT. In this chapter, we present the mechanism of action of the chloroquine, the structural changes of the gene after the mutations as well as different haplotypes of the PfCRT

Molecular characterization of Plasmodium falciparum in Arunachal Pradesh from Northeast India based on merozoite surface protein 1 & glutamate-rich protein

Background & objectives: Northeast (NE) India is one of the high endemic regions for malaria with a preponderance of Plasmodium falciparum, resulting in high morbidity and mortality. The P. falciparum parasite of this region showed high polymorphism in drug-resistant molecular biomarkers. However, there is a paucity of information related to merozoite surface protein 1 (msp-1) and glutamate-rich protein (glurp) which have been extensively studied in various parts of the world. The present study was, therefore, aimed at investigating the genetic diversity of P. falciparum based on msp-1 and glurp in Arunachal Pradesh, a State in NE India. Methods: Two hundred and forty nine patients with fever were screened for malaria, of whom 75 were positive for P. falciparum. Blood samples were collected from each microscopically confirmed patient. The DNA was extracted; nested polymerase chain reaction and sequencing were performed to study the genetic diversity of msp-1 (block 2) and glurp. Results: The block 2 of msp-1 gene was found to be highly polymorphic, and overall allelic distribution showed that RO33 was the dominant allele (63%), followed by MAD20 (29%) and K1 (8%) alleles. However, an extensive diversity (9 alleles and 4 genotypes) and 6-10 repeat regions exclusively of R2 type were observed in glurp. Interpretation & conclusions: The P. falciparum population of NE India was diverse which might be responsible for higher plasticity leading to the survival of the parasite and in turn to the higher endemicity of falciparum malaria of this region

Age and codon wise association of CQ (above) and SP (below) ETF.

<p>Age and codon wise association of CQ (above) and SP (below) ETF.</p

Demographic and clinical characteristics of patients and their responses to treatment with SP and CQ.

<p>ACPR = Adequate Clinical and Parasitological Response; TF = Treatment failure, ETF = Early Treatment Failure, LCF = Late Clinical Failure; LPF = Late Parasitological Failure; CF = Clinical failure, TF = Treatment failure.</p><p>Demographic and clinical characteristics of patients and their responses to treatment with SP and CQ.</p

The frequency distribution of SNPs combination for <i>dhps</i> alleles.

<p>Allelic combinations are in order of S436F/A, A437G, K540E, A581G, and A613S/T, where bold and underlined alleles denotes mutations. Values within bracket are the percentage of occurrences.</p><p>The frequency distribution of SNPs combination for <i>dhps</i> alleles.</p

<i>dhps/dhfr</i> haplotypes associated with CQ and SP early treatment failure (ETF).

<p>Allelic combinations are in order of S436F/A, A437G, K540E, A581G, and A613S/T (<i>dhps</i>)-A16V, C50R N51I, C59R, S108N and I164L (<i>dhfr</i>) where bold alleles denotes mutations.</p><p><i>dhps/dhfr</i> haplotypes associated with CQ and SP early treatment failure (ETF).</p

Map of the studied areas of north east India.

<p>Map of the studied areas of north east India.</p

Molecular Evidence of Increased Resistance to Anti-Folate Drugs in <i>Plasmodium falciparum</i> in North-East India: A Signal for Potential Failure of Artemisinin Plus Sulphadoxine-Pyrimethamine Combination Therapy

<div><p>North-east India, being a corridor to South-east Asia, is believed to play an important role in transmitting drug resistant <i>Plasmodium falciparum</i> malaria to India and South Asia. North-east India was the first place in India to record the emergence of drug resistance to chloroquine as well as sulphadoxine/pyrimethamine. Presently chloroquine resistance is widespread all over the North-east India and resistance to other anti-malarials is increasing. In this study both <i>in vivo</i> therapeutic efficacy and molecular assays were used to screen the spectrum of drug resistance to chloroquine and sulphadoxine/pyrimethamine in the circulating <i>P. falciparum</i> strains. A total of 220 <i>P. falciparum</i> positives subjects were enrolled in the study for therapeutic assessment of chloroquine and sulphadoxine/pyrimethamine and assessment of point mutations conferring resistances to these drugs were carried out by genotyping the isolates following standard methods. Overall clinical failures in sulphadoxine/pyrimethamine and chloroquine were found 12.6 and 69.5% respectively, while overall treatment failures recorded were 13.7 and 81.5% in the two arms. Nearly all (99.0%) the isolates had mutant <i>pfcrt</i> genotype (76T), while 68% had mutant <i>pfmdr</i>-1 genotype (86Y). Mutation in <i>dhps</i> 437 codon was the most prevalent one while <i>dhfr</i> codon 108 showed 100% mutation. A total of 23 unique haplotypes at the <i>dhps</i> locus and 7 at <i>dhfr</i> locus were found while <i>dhps</i>-<i>dhfr</i> combined loci revealed 49 unique haplotypes. Prevalence of double, triple and quadruple mutations were common while 1 haplotype was found with all five mutated codons (<b><u>F/AGEGS/T</u></b>) at <i>dhps</i> locus. Detection of quadruple mutants (51I/59R/108N/164L) in the present study, earlier recorded from Car Nicobar Island, India only, indicates the presence of high levels of resistance to sulphadoxine/pyrimethamine in north-east India. Associations between resistant haplotypes and the clinical outcomes and emerging resistance in sulphadoxine/pyrimethamine in relation to the efficacy of the currently used artemisinin combination therapy are discussed.</p></div