College of Public Health, Medical and Veterinary Sciences, James Cook University, PO Box 6811, Cairns, QLD, 4870, Australia.
Australian Institute of Tropical Health and Medicine, James Cook University, PO Box 6811, Cairns, QLD, 4870, Australia.
Parasit Vectors. 2019 Jul 18;12(1):355. doi: 10.1186/s13071-019-3610-9.
Malaria is the most important vector-borne disease in the world. Epidemiological and ecological studies of malaria traditionally utilize detection of Plasmodium sporozoites in whole mosquitoes or salivary glands by microscopy or serological or molecular assays. However, these methods are labor-intensive, and can over- or underestimate mosquito transmission potential. To overcome these limitations, alternative sample types have been evaluated for the study of malaria. It was recently shown that Plasmodium could be detected in saliva expectorated on honey-soaked cards by Anopheles stephensi, providing a better estimate of transmission risk. We evaluated whether excretion of Plasmodium falciparum nucleic acid by An. stephensi correlates with expectoration of parasites in saliva, thus providing an additional sample type for estimating transmission potential. Mosquitoes were exposed to infectious blood meals containing cultured gametocytes, and excreta collected at different time points post-exposure. Saliva was collected on honey-soaked filter paper cards, and salivary glands were dissected and examined microscopically for sporozoites. Excreta and saliva samples were tested by real time polymerase chain reaction (RT-rtPCR).
Plasmodium falciparum RNA was detected in mosquito excreta as early as four days after ingesting a bloodmeal containing gametocytes. Once sporogony (the development of sporozoites) occurred, P. falciparum RNA was detected concurrently in both excreta and saliva samples. In the majority of cases, no difference was observed between the C values obtained from matched excreta and saliva samples, suggesting that both samples provide equally sensitive results. A positive association was observed between the molecular detection of the parasites in both samples and the proportion of mosquitoes with sporozoites in their salivary glands from each container. No distinguishable parasites were observed when excreta samples were stained and microscopically analyzed.
Mosquito saliva and excreta are easily collected and are promising for surveillance of malaria-causing parasites, especially in low transmission settings or in places where arboviruses co-circulate.
疟疾是世界上最重要的虫媒传染病。传统上，疟疾的流行病学和生态学研究通过显微镜检查或血清学或分子检测来检测整只蚊子或唾液腺中的疟原虫孢子。然而，这些方法劳动强度大，并且可能高估或低估蚊子传播的潜力。为了克服这些限制，已经评估了替代样本类型用于疟疾研究。最近的研究表明，按蚊可以在吸食涂有蜂蜜的卡片上的唾液中检测到疟原虫，从而更好地估计传播风险。我们评估了按蚊排泄的恶性疟原虫核酸是否与唾液中寄生虫的咳出有关，从而为估计传播潜力提供了另一种样本类型。蚊子暴露于含有培养配子的感染性血餐中，并在暴露后不同时间收集排泄物。在涂有蜂蜜的滤纸上收集唾液，并通过实时聚合酶链反应（RT-rtPCR）检测唾液腺。
在摄入含有配子的血餐四天后，最早可以在蚊子排泄物中检测到恶性疟原虫 RNA。一旦发生孢子生殖（孢子形成），即可在排泄物和唾液样本中同时检测到恶性疟原虫 RNA。在大多数情况下，从配对的排泄物和唾液样本中获得的 C 值之间没有差异，这表明两种样本均提供了同样敏感的结果。在这两种样本中检测到寄生虫的分子检测与每个容器中带有唾液腺孢子的蚊子比例之间存在正相关关系。当对排泄物样本进行染色和显微镜分析时，没有观察到可区分的寄生虫。
蚊子唾液和排泄物易于采集，非常适合监测引起疟疾的寄生虫，尤其是在低传播环境或虫媒病毒共同传播的地方。
