Accuracy and reliability of malaria diagnostic techniques for guiding febrile outpatient treatment in malaria-endemic countries.
ABSTRACT The main purpose of this study was to assess the accuracy of various techniques available for diagnosis of malaria. Blood samples were collected from 313 patients with clinical suspicion of uncomplicated malaria in 2 primary health centers in Madagascar. The presence of Plasmodium parasites was assessed by conventional microscopy, 2 rapid diagnostic tests (one HRP2-based test, PALUTOP(+4), and one pLDH-based test, OptiMAL-IT), and real-time polymerase chain reaction (PCR), which is used as the "gold standard" method. The degree of agreement observed was very high for microscopy (0.99) and the HRP2-based test (0.93) and high for the pLDH-based test (0.82). Public-health implications are also discussed in this paper.
Article: Global sequence variation in the histidine-rich proteins 2 and 3 of Plasmodium falciparum: implications for the performance of malaria rapid diagnostic tests.[show abstract] [hide abstract]
ABSTRACT: Accurate diagnosis is essential for prompt and appropriate treatment of malaria. While rapid diagnostic tests (RDTs) offer great potential to improve malaria diagnosis, the sensitivity of RDTs has been reported to be highly variable. One possible factor contributing to variable test performance is the diversity of parasite antigens. This is of particular concern for Plasmodium falciparum histidine-rich protein 2 (PfHRP2)-detecting RDTs since PfHRP2 has been reported to be highly variable in isolates of the Asia-Pacific region. The pfhrp2 exon 2 fragment from 458 isolates of P. falciparum collected from 38 countries was amplified and sequenced. For a subset of 80 isolates, the exon 2 fragment of histidine-rich protein 3 (pfhrp3) was also amplified and sequenced. DNA sequence and statistical analysis of the variation observed in these genes was conducted. The potential impact of the pfhrp2 variation on RDT detection rates was examined by analysing the relationship between sequence characteristics of this gene and the results of the WHO product testing of malaria RDTs: Round 1 (2008), for 34 PfHRP2-detecting RDTs. Sequence analysis revealed extensive variations in the number and arrangement of various repeats encoded by the genes in parasite populations world-wide. However, no statistically robust correlation between gene structure and RDT detection rate for P. falciparum parasites at 200 parasites per microlitre was identified. The results suggest that despite extreme sequence variation, diversity of PfHRP2 does not appear to be a major cause of RDT sensitivity variation.Malaria Journal 01/2010; 9:129. · 3.19 Impact Factor
Article: Malaria rapid diagnostic tests: Plasmodium falciparum infections with high parasite densities may generate false positive Plasmodium vivax pLDH lines.[show abstract] [hide abstract]
ABSTRACT: Most malaria rapid diagnostic tests (RDTs) detect Plasmodium falciparum and an antigen common to the four species. Plasmodium vivax-specific RDTs target P. vivax-specific parasite lactate dehydrogenase (Pv-pLDH). Previous observations of false positive Pv-pLDH test lines in P. falciparum samples incited to the present study, which assessed P. vivax-specific RDTs for the occurrence of false positive Pv-pLDH lines in P. falciparum samples. Nine P. vivax-specific RDTs were tested with 85 P. falciparum samples of high (>or=2%) parasite density. Mixed P. falciparum/P. vivax infections were ruled out by real-time PCR. The RDTs included two-band (detecting Pv-pLDH), three-band (detecting P. falciparum-antigen and Pv-pLDH) and four-band RDTs (detecting P. falciparum, Pv-pLDH and pan-pLDH). False positive Pv-pLDH lines were observed in 6/9 RDTs (including two- three- and four-band RDTs). They occurred in the individual RDT brands at frequencies ranging from 8.2% to 29.1%. For 19/85 samples, at least two RDT brands generated a false positive Pv-pLDH line. Sixteen of 85 (18.8%) false positive lines were of medium or strong line intensity. There was no significant relation between false positive results and parasite density or geographic origin of the samples. False positive Pv-pLDH lines in P. falciparum samples with high parasite density occurred in 6/9 P. vivax-specific RDTs. This is of concern as P. falciparum and P. vivax are co-circulating in many regions. The diagnosis of life-threatening P. falciparum malaria may be missed (two-band Pv-pLDH RDT), or the patient may be treated incorrectly with primaquine (three- or four-band RDTs).Malaria Journal 01/2010; 9:198. · 3.19 Impact Factor
Article: In vitro susceptibility to pyrimethamine of DHFR I164L single mutant Plasmodium falciparum.[show abstract] [hide abstract]
ABSTRACT: Recently, Plasmodium falciparum parasites bearing Pfdhfr I164L single mutation were found in Madagascar. These new mutants may challenge the use of antifolates for the intermittent preventive treatment of malaria during pregnancy (IPTp). Assays with transgenic bacteria suggested that I164L parasites have a wild-type phenotype for pyrimethamine but it had to be confirmed by testing the parasites themselves. Thirty Plasmodium falciparum clinical isolates were collected in 2008 in the south-east of Madagascar. A part of Pfdhfr gene encompassing codons 6 to 206 was amplified by PCR and the determination of the presence of single nucleotide polymorphisms was performed by DNA sequencing. The multiplicity of infection was estimated by using an allelic family-specific nested PCR. Isolates that appeared monoclonal were submitted to culture adaptation. Determination of IC(50s) to pyrimethamine was performed on adapted isolates. Four different Pfdhfr alleles were found: the 164L single mutant-type (N = 13), the wild-type (N = 7), the triple mutant-type 51I/59R/108N (N = 9) and the double mutant-type 108N/164L (N = 1). Eleven out 30 (36.7%) of P. falciparum isolates were considered as monoclonal infection. Among them, five isolates were successfully adapted in culture and tested for pyrimethamine in vitro susceptibility. The wild-type allele was the most susceptible with a 50% inhibitory concentration (IC(50)) < 10 nM. The geometric mean of IC(50) of the three I164L mutant isolates was 6-fold higher than the wild-type with 61.3 nM (SD = 3.2 nM, CI95%: 53.9-69.7 nM). These values remained largely below the IC(50) of the triple mutant parasite (13,804 nM). The IC(50)s of the I164L mutant isolates were significantly higher than those of the wild-type (6-fold higher) and close from those usually reported for simple mutants S108N (roughly10-fold higher than wild type). Given the observed values, the determination of IC(50)s directly on parasites did not confirm what has been found on transgenic bacteria. The prevalence increase of the Pfdhfr I164L single mutant parasite since 2006 could be explained by the selective advantage of this allele under sulphadoxine-pyrimethamine pressure. The emergence of highly resistant alleles should be considered in the future, in particular because an unexpected double mutant-type allele S108N/I164L has been already detected.Malaria Journal 09/2011; 10:283. · 3.19 Impact Factor