To estimate the accuracy of RDTs for diagnosing malaria infection in pregnant women, the results of 49 studies were analysed. Few studies exist that fully evaluate microscopy, RDTs and PCR against the gold standard, placental histology, and each other. This makes it difficult to directly compare the accuracies of the different tests, and therefore currently no reliable data about the accuracy of RDTs and PCR for the detection of placental malaria is available. The present study shows that microscopy of both placental and peripheral blood do not detect many placental infections that are identified by histology (sensitivities of 54% and 44%) and cannot reliably replace histology as a reference standard for placental P. falciparum infection. Despite these limitations RDTs, especially HRP2-based tests, appear to have reasonable accuracy compared to microscopy. The World Health Organization (WHO) together with the Foundation for Innovative New Diagnostics (FIND) have performed extensive testing of many RDT devices and there is a great need to compare the best tests from that evaluation with histology, microscopy and PCR in pregnant women .
When using placental microscopy as a reference test, PCR has the best sensitivity, followed by RDT and both have higher sensitivity than peripheral microscopy. Peripheral microscopy, however, has the highest specificity, followed closely by RDT. Based on these results, RDTs seem a good alternative for diagnosis of placental malaria. For the determination of the accuracy of RDT and PCR compared to placental blood microscopy, however, tests performed on both peripheral and placental blood were pooled. This might have introduced bias, since the starting material is not the same, and therefore, the accuracy might be different. There were, however, too few studies performed on peripheral blood alone to perform meta-analysis. The pooled summary sensitivities and specificities are not that different from the summary values of these tests using placental blood, but this may be due to the fact that these tests are a large proportion of the pooled analysis.
Compared to the imperfect reference standard peripheral microscopy, the proportion of microscopy positives detected by any RDT ("sensitivity") was 81% [51-95 CI]. The proportion microscopy negatives, with a negative RDT ("specificity"), was 94% [76-99 CI]. As the RDTs seem to miss patients that are positive in microscopy, this is not very reassuring for the value of RDTs for the diagnosis of malaria in pregnant women. Nevertheless, HRP2-based RDTs might have adequate sensitivity (94% [91-96 CI]).
The results in this study suggest that the proportion of microscopy positives detected by HRP2-based RDTs compared to peripheral microscopy is higher than for pLDH-based RDTs. The proportion microscopy negatives with a negative RDT, however, is lower for HRP2 RDTs compared to pLDH RDTs. This pattern is similar to the results of a meta-analysis of RDTs for uncomplicated malaria, and the sensitivity of HRP2-based RDTs for pregnant women (94% [91-96 CI]) is very similar to that for uncomplicated malaria (95% [93-96 CI]) . The specificity compared to peripheral microscopy for HRP2-based RDTs for pregnant women (81% [71-88 CI]), however, is much lower than for uncomplicated malaria (95% [93-99 CI]). A possible explanation for this observation is that peripheral microscopy is not a suitable reference test for placental malaria and does not detect all placental infections, whereas the HRP2 RDTs might be able to detect these infections, resulting in a lower specificity when compared to peripheral microscopy. This is strengthened by the specificity of HRP2 RDTs (90% [84-95 CI]) compared to placental blood microscopy, which detects more placental infections than peripheral microscopy. Direct comparisons of HRP2 RDTs with histology are needed to confirm this.
For PCR, the proportion of microscopy positives detected ("sensitivity") was 98% [91-99 CI] and the proportion microscopy negatives, with a negative PCR ("specificity"), was 65% [59-72 CI]. PCR may seem to miss fewer patients with peripheral P. falciparum parasites than an RDT, but does indicate a number of women without parasites detected by microscopy as having malaria. Whether these are cases that were missed by microscopy or whether these were false positive PCR results, resulting in low specificity, needs to be further investigated.
The sensitivity and specificity of tests vary with parasite density, and many included studies report lower sensitivities at lower parasite densities (two studies for PCR and RDT; seven for RDTs; four for microscopy). This factor is a particular challenge for malaria infections during pregnancy in both high and low transmission settings, especially for multigravid women who are often reported to have lower parasite densities. Of the included studies, 13 report higher parasite densities in peripheral and/or placental blood for primi- and/or secundigravidae. In high transmission areas, women have acquired immunity during their life, and although they might have substantial placental sequestration (especially in primi- and secundigravidae), they can have a lower amount of parasites in the circulation by clearance of infected red blood cells by the immune system. In low transmission areas, women have low or no immunity and women can get sick at initially low parasite densities. Therefore it is very important for the diagnosis of malaria in pregnant women that the test has sufficient accuracy at lower parasite densities, and low parasite density might partially explain the poor performance of pLDH RDTs.
In a previous report, prevalence of placental malaria estimated by different tests was compared within each study . However, even if the prevalence detected by one test is higher than the prevalence detected with the other test, it does not necessarily mean that the same cases are detected. Additionally, in this way it is not clear whether the positives found by each test are true positives or false positives. Nevertheless, the conclusions of that report and this review are broadly similar. Histology found a higher prevalence than placental blood microscopy and placental blood microscopy in turn found a higher prevalence than peripheral microscopy in four out of six studies. In both peripheral and placental blood, in nearly all studies, higher prevalence was found using PCR than using RDT and higher prevalence was found using RDT than using microscopy. No studies compared prevalence estimated by RDT and PCR to prevalence found by histology.
The results presented in this review are mainly applicable to sub-Saharan Africa, as 85% of the included studies were conducted in that area. It is surprising that only a few studies were available evaluating diagnostics in the Asian-Pacific region, considering that most pregnancies at risk in the world are located in this region . In line with this observation, most studies described P. falciparum infections, and only six studies reported P. vivax infections. Meta-analysis was based on studies of which the majority described P. falciparum infections and studies with P. vivax infections had little influence on the outcome. Therefore, conclusions in this review are mainly applicable to P. falciparum infections. Due to the potential pathological differences between P. falciparum and P. vivax infection in pregnancy a difference in diagnostic accuracy of the various tests is expected between the species and more studies should aim at evaluating diagnostics in P. vivax-infected pregnant women in the future. Too few studies with P. vivax infections were included in the meta-analysis to be able to determine if there is a difference in accuracy.
The use of PCR for diagnosis of (placental) malaria remains a matter of discussion, especially in the submicroscopic cases, because it is difficult to determine what the PCR is detecting. It is very sensitive in detecting parasite nucleic acids, but it is unclear if this is a residual from a non-viable sequestered parasite, or a viable parasite or gametocyte. Additionally, there is some discussion whether these low parasite levels detected by PCR are clinically relevant. Several studies have tried to address this issue by researching the association of a positive (submicroscopic) PCR result and outcome measures such as anaemia, low birth weight and premature delivery. A systematic review has been conducted to summarize this data and concluded that the frequency of anaemia is significantly lower in uninfected women compared to women with a submicroscopic infection, although the risk is lower than with microscopic infections, and a similar pattern is found for low birth weight . The review on the effects of submicroscopic infections did not include three studies that have been described in the current study [[60, 90, 93]]. One of these three studies described a significantly increased risk of anaemia with submicroscopic infections compared to PCR- and microscopy negative women, as well . One of the other studies describes that submicroscopic infection was predictive of low birth weight in HIV positive, but not HIV negative women . More comparisons with histology might shed more light on this issue and show that a significant proportion of the peripheral submicroscopic infections, are in fact placental infections. As mentioned before, pregnant women often have low parasite densities, and tests should have good sensitivity at these low densities; PCR techniques often have better sensitivities at low parasite densities than RDTs and microscopy.