Zhou Zhiyong, Mitchell Rebecca Mans, Gutman Julie, Wiegand Ryan E, Mwandama Dyson A, Mathanga Don P, Skarbinski Jacek, Shi Ya Ping
Malaria Branch and Division of Parasitic Diseases and Malaria, Center for Global Health, Centers for Disease Control and Prevention, Atlanta, GA, USA.
Malar J. 2014 Dec 18;13:509. doi: 10.1186/1475-2875-13-509.
Low malaria parasite densities in pregnancy are a diagnostic challenge. PCR provides high sensitivity and specificity in detecting low density of parasites, but cost and technical requirements limit its application in resources-limited settings. Pooling samples for PCR detection was explored to estimate prevalence of submicroscopic malaria infection in pregnant women at delivery. Previous work uses gold-standard based methods to calculate sensitivity and specificity of tests, creating a challenge when newer methodologies are substantially more sensitive than the gold standard. Thus prevalence was estimated using Bayesian latent class models (LCMs) in this study.
Nested PCR (nPCR) for the 18S rRNA gene subunit of Plasmodium falciparum was conducted to detect malaria infection in microscopy-negative Malawian women on IPTp. Two-step sample pooling used dried blood spot samples (DBSs) collected from placenta or periphery at delivery. Results from nPCR and histology as well as previously published data were used to construct LCMs to estimate assay sensitivity and specificity. Theoretical confidence intervals for prevalence of infection were calculated for two-step and one-step pooling strategies.
Of 617 microscopy-negative Malawian women, 39 (6.3%) were identified as actively infected by histology while 52 (8.4%) were positive by nPCR. One hundred forty (22.7%) individuals had past infection assessed by histology. With histology as a reference, 72% of women in the active infection group, 7.1% in the past infection group and 3.2% in histology-negative group were nPCR positive. Using latent class models without a gold standard, histology had a median sensitivity of 49.7% and specificity of 97.6% for active infection while PCR had a median sensitivity of 96.0% and specificity of 99.1%. The true prevalence of active infection was estimated at 8.0% (CI: 5.8-10.5%) from PCR. PCR also had similar sensitivity for detecting either peripheral or placental malaria for submicroscopic infections. One-step pooling would give similar confidence intervals for pool sizes less than 20 while reducing the number of tests performed.
Pooled nPCR testing was a sensitive and resource-efficient strategy and LCMs provided precise prevalence estimates of submicroscopic infections. Compared to two-step pooling, one-step pooling could provide similar prevalence estimates at population levels with many fewer tests required.
孕期疟原虫密度低是一个诊断难题。聚合酶链反应（PCR）在检测低水平寄生虫密度方面具有高灵敏度和特异性，但成本和技术要求限制了其在资源有限环境中的应用。本研究探讨了采用样本合并进行PCR检测，以估计分娩时孕妇亚显微疟原虫感染的患病率。以往的研究采用基于金标准的方法来计算检测方法的灵敏度和特异性，当新方法比金标准灵敏得多时，就会产生问题。因此，本研究使用贝叶斯潜在类别模型（LCM）来估计患病率。
对接受孕期间歇性预防治疗（IPTp）的马拉维显微镜检查阴性的妇女，采用针对恶性疟原虫18S rRNA基因亚单位的巢式PCR（nPCR）检测疟原虫感染情况。两步样本合并使用分娩时从胎盘或外周采集的干血斑样本（DBS）。nPCR和组织学检查结果以及先前发表的数据用于构建LCM，以估计检测方法的灵敏度和特异性。计算了两步法和一步法合并策略下感染患病率的理论置信区间。
在617名显微镜检查阴性的马拉维妇女中，39名（6.3%）经组织学检查被确定为有活动性感染，52名（8.4%）nPCR检测呈阳性。140名（约22.7%）个体经组织学检查有既往感染。以组织学检查为参照，活动性感染组中72%的女性、既往感染组中7.1%的女性以及组织学检查阴性组中3.2%的女性nPCR检测呈阳性。在没有金标准的情况下，使用潜在类别模型，组织学检查对活动性感染的中位灵敏度为49.7%，特异性为97.6%，而PCR的中位灵敏度为96.0%，特异性为99.1%。根据PCR检测结果，活动性感染的实际患病率估计为8.0%（置信区间：5.8 - 10.5%）。对于亚显微感染，PCR检测外周或胎盘疟疾的灵敏度相似。对于样本量小于20的情况，一步法合并能给出相似的置信区间，同时减少检测次数。
合并nPCR检测是一种灵敏且资源高效的策略，LCM能提供亚显微感染患病率的精确估计。与两步法合并相比，一步法合并在人群水平上能提供相似的患病率估计，且所需检测次数少得多。
