%0 Journal Article
%T Improvement of the observational method for Plasmodium berghei oocysts in the midgut of mosquitoes
%A Miho Usui
%A Shinya Fukumoto
%A Noboru Inoue
%A Shin-ichiro Kawazu
%J Parasites & Vectors
%D 2011
%I BioMed Central
%R 10.1186/1756-3305-4-118
%X In the improved technique, the parasite-infected midgut was first stained with mercurochrome, and then fixed with formalin. The specimens were finally observed using light microscopy. To evaluate the accuracy in the oocyst counting with the improved technique, mosquitoes were infected with the green fluorescent protein (GFP)-expressing parasite. Then, the midgut oocysts were counted using both the GFP marker and the improved technique. Results were then compared and showed that the improved technique retrieved 78%-123% (arithmetic mean = 97%) of the oocysts counted using the GFP marker. Furthermore, it was also possible to evaluate the oocyst development with a green filter using the light microscopy.The improved technique for oocyst counting will be a useful tool for evaluating midgut oocyst numbers and determining the developmental stage of oocysts in parasite-infected mosquitoes.Malaria is a disease caused by the infection with protozoan parasites of the genus Plasmodium and transmitted by Anopheles mosquitoes. To study the life cycle of Plasmodium berghei in mosquitoes, qualitative and quantitative evaluation of midgut oocysts in experimental infections is needed [1]. There are two methods currently used for counting oocysts. One technique used is the formalin fixation method [2]. In this method, the midgut is fixed with 10% formalin and opened along the median line to prepare a single-layered specimen. The advantage of using this technique is that it can preserve the sample for storage and later observation. However, this technique leaves the oocysts unstained, making them indistinguishable from insect tissues (Figure 1A). Another technique is the mercurochrome staining method [3-5]. In this method, the midgut is stained with 0.5% mercurochrome in water which makes the oocysts be easily distinguished from insect tissues (Figure 1B). However, the resulting specimen from this technique should be observed at the same day since there was no included preservative. T
%U http://www.parasitesandvectors.com/content/4/1/118