EA7510, ESCAPE, Laboratoire de Parasitologie Mycologie, Université de Reims Champagne Ardenne, SFR Cap Santé FED 4231, Reims, France.
ACTALIA Food Safety Department, Saint-Lô, France.
Appl Environ Microbiol. 2019 Oct 1;85(20). doi: 10.1128/AEM.01189-19. Print 2019 Oct 15.
is a ubiquitous foodborne protozoan that can infect humans at low dose and displays different prevalences among countries in the world. Ingestion of food or water contaminated with small amounts of oocysts may result in human infection. However, there are no regulations for monitoring oocysts in food, mainly because of a lack of standardized methods to detect them. The objectives of this study were (i) to develop a reliable method, applicable in biomonitoring, for the rapid detection of infectious oocysts by cell culture of their sporocysts combined with quantitative PCR (sporocyst-CC-qPCR) and (ii) to adapt this method to blue and zebra mussels experimentally contaminated by oocysts with the objective to use these organisms as sentinels of aquatic environments. Combining mechanical treatment and bead beating leads to the release of 84% ± 14% of free sporocysts. The sporocyst-CC-qPCR detected fewer than ten infectious oocysts in water within 4 days (1 day of contact and 3 days of cell culture) compared to detection after 4 weeks by mouse bioassay. For both mussel matrices, oocysts were prepurified using a 30% Percoll gradient and treated with sodium hypochlorite before cell culture of their sporocysts. This assay was able to detect as few as ten infective oocysts. This sporocyst-based CC-qPCR appears to be a good alternative to mouse bioassay for monitoring infectious oocysts directly in water and also using biological sentinel mussel species. This method offers a new perspective to assess the environmental risk for human health associated with this parasite. The ubiquitous protozoan is the subject of renewed interest due to the spread of oocysts in water and food causing endemic and epidemic outbreaks of toxoplasmosis in humans and animals worldwide. Displaying a sensitivity close to animal models, cell culture represents a real alternative to assess the infectivity of oocysts in water and in biological sentinel mussels. This method opens interesting perspectives for evaluating human exposure to infectious oocysts in the environment, where oocyst amounts are considered to be very small.
是一种普遍存在的食源性原生动物，在低剂量下即可感染人类，并且在世界各国的流行率不同。摄入被少量 卵囊污染的食物或水可能导致人类感染。然而，目前并没有监测食品中卵囊的规定，主要是因为缺乏标准化的检测方法。本研究的目的是（i）开发一种可靠的方法，适用于通过其孢子囊的细胞培养结合定量 PCR（孢子囊-CC-qPCR）快速检测传染性卵囊，（ii）将该方法应用于蓝贻贝和斑马贻贝，这些贻贝通过卵囊实验污染，目的是使用这些生物作为水生环境的哨兵。结合机械处理和珠磨可使游离孢子囊的释放率达到 84%±14%。与 4 周后通过小鼠生物测定法相比，孢子囊-CC-qPCR 在 4 天（接触 1 天和细胞培养 3 天）内即可检测到水中少于 10 个传染性卵囊。对于这两种贻贝基质，卵囊先用 30%的聚蔗糖梯度预纯化，然后用次氯酸钠处理，再培养其孢子囊。该检测方法能够检测到低至 10 个感染性卵囊。基于孢子囊的 CC-qPCR 似乎是替代小鼠生物测定法的良好方法，可直接在水中监测传染性 卵囊，也可使用生物指示贻贝物种。该方法为评估与这种寄生虫相关的人类健康的环境风险提供了新的视角。由于卵囊在水和食物中的传播导致全世界人类和动物的弓形虫病地方性和流行性病暴发，普遍存在的原生动物 再次引起人们的兴趣。细胞培养与动物模型的敏感性接近，代表了评估水中和生物指示贻贝中卵囊感染性的真正替代方法。该方法为评估环境中人类对传染性 卵囊的暴露提供了有趣的前景，在这种情况下，卵囊数量被认为非常小。
