Reliability of Rapid Diagnostic Tests in Diagnosing Pregnancy-Associated Malaria in North-Eastern Tanzania.

Accurate diagnosis and prompt treatment of pregnancy-associated malaria (PAM) are key aspects in averting adverse pregnancy outcomes. Microscopy is the gold standard in malaria diagnosis, but it has limited detection and availability. When used appropriately, rapid diagnostic tests (RDTs) could be an ideal diagnostic complement to microscopy, due to their ease of use and adequate sensitivity in detecting even sub-microscopic infections. Polymerase chain reaction (PCR) is even more sensitive, but it is mainly used for research purposes. The accuracy and reliability of RDTs in diagnosing PAM was evaluated using microscopy and PCR. A cohort of pregnant women in north-eastern Tanzania was followed throughout pregnancy for detection of plasmodial infection using venous and placental blood samples evaluated by histidine rich protein 2 (HRP-2) and parasite lactate dehydrogenase (pLDH) based RDTs (Parascreen™) or HRP-2 only (Paracheck Pf® and ParaHIT®f), microscopy and nested Plasmodium species diagnostic PCR. From a cohort of 924 pregnant women who completed the follow up, complete RDT and microscopy data was available for 5,555 blood samples and of these 442 samples were analysed by PCR. Of the 5,555 blood samples, 49 ((proportion and 95% confidence interval) 0.9% [0.7 -1.1]) samples were positive by microscopy and 91 (1.6% [1.3-2.0]) by RDT. Forty-six (50.5% [40.5 - 60.6]) and 45 (49.5% [39.4 - 59.5]) of the RDT positive samples were positive and negative by microscopy, respectively, whereas nineteen (42.2% [29.0 - 56.7]) of the microscopy negative, but RDT positive, samples were positive by PCR. Three (0.05% [0.02 - 0.2]) samples were positive by microscopy but negative by RDT. 351 of the 5,461 samples negative by both RDT and microscopy were tested by PCR and found negative. There was no statistically significant difference between the performances of the different RDTs. Microscopy underestimated the real burden of malaria during pregnancy and RDTs performed better than microscopy in diagnosing PAM. In areas where intermittent preventive treatment during pregnancy may be abandoned due to low and decreasing malaria risk and instead replaced with active case management, screening with RDT is likely to identify most infections in pregnant women and out-performs microscopy as a diagnostic tool

Submicroscopic Gametocytes and the Transmission of Antifolate-Resistant Plasmodium falciparum in Western Kenya

BACKGROUND: Single nucleotide polymorphisms (SNPs) in the dhfr and dhps genes are associated with sulphadoxine-pyrimethamine (SP) treatment failure and gametocyte carriage. This may result in enhanced transmission of mutant malaria parasites, as previously shown for chloroquine resistant parasites. In the present study, we determine the association between parasite mutations, submicroscopic P. falciparum gametocytemia and malaria transmission to mosquitoes. METHODOLOGY/PRINCIPAL FINDINGS: Samples from children treated with SP alone or in combination with artesunate (AS) or amodiaquine were genotyped for SNPs in the dhfr and dhps genes. Gametocytemia was determined by microscopy and Pfs25 RNA-based quantitative nucleic acid sequence-based amplification (Pfs25 QT-NASBA). Transmission was determined by membrane-feeding assays. We observed no wild type infections, 66.5% (127/191) of the infections expressed mutations at all three dhfr codons prior to treatment. The presence of all three mutations was not related to higher Pfs25 QT-NASBA gametocyte prevalence or density during follow-up, compared to double mutant infections. The proportion of infected mosquitoes or oocyst burden was also not related to the number of mutations. Addition of AS to SP reduced gametocytemia and malaria transmission during follow-up. CONCLUSIONS/SIGNIFICANCE: In our study population where all infections had at least a double mutation in the dhfr gene, additional mutations were not related to increased submicroscopic gametocytemia or enhanced malaria transmission. The absence of wild-type infections is likely to have reduced our power to detect differences. Our data further support the use of ACT to reduce the transmission of drug-resistant malaria parasites

Enrolment characteristics and gametocyte carriage in relation to mutations in the <i>dhfr</i> gene.

<p>AUC = area under the curve; GM = geometric mean; IQR = interquartile range; Non-ACT = treatment with sulphadoxine-pyrimethamine (SP) alone or in combination with amodiaquine; ACT = treatment with SP and artesunate (3 days).</p>¥<p>Gametocyte densities were measured by <i>Pfs25</i> QT-NASBA.</p>*<p>Mutations were determined at enrolment: double mutation = mutations in c108+c59 or c108+c51; three mutations = mutations in c51+c59+c108;</p>¶<p>the period from enrolment until time of feeding (day 14) was considered for determining the AUC.</p

Prevalence of mutant genotypes for <i>dhfr</i> and <i>dhps</i> genes prior to treatment and on day 14 after non-ACT or ACT treatment in samples included in the membrane-feeding assays.

<p>Non-ACT = sulphadoxine-pyrimethamine (SP) alone or in combination with amodiaquine; ACT = SP in combination with artesunate; ≥ three mutations = mutations in c51+c59+c108 and possible additional mutations The number of observations on day 0: all mutations (n = 57–58) (see text); on day 14 after non-ACT treatment: c51, c59, c108, c164 (n = 33); c436, c437 (n = 23); c540, c581, c613 (n = 25); ≥ three mutations (n = 33); on day 14 after ACT treatment: c51, c59 (n = 8); c108, c164 (n = 7); c436, c437 (n = 4); c540, c581, c613 (n = 5); ≥ three mutations (n = 7).</p