Authors Yung-san Liang, Mei-Shei Su, Laksiri Karunaratne and Fred Lewis
Materials and Reagents
Petri dishes (60 mm dia)
Aged tap water
Fine forceps or small hypodermic needle (e.g., 25 gauge) placed on a syringe
Hairloop attached to a pasteur pipette
· Primary sporocysts are difficult to see in the headfoot tissues of an exposed Oncomelania hupensis ssp. snail; however at about 3 months after exposure one can see developing secondary sporocysts through the shell (apex area), provided that the shell is not too eroded.
· Cercariae can be obtained by crushing the snail at about 3 ½ months after exposure. It is possible to obtain a few cercariae from the snails by shedding alone, but for large numbers of cercariae it is necessary to crush them and dissect out the tissues.
· Place a few snails on an inverted plastic petri dish top, and gently crush the shells by placing the bottom of the dish on top of the snails and pressing down. Add a small drop of water to the crushed snail, tease out the secondary sporocysts from the tissue, then place them in a 60 mm petri dish with aged tap water.
· Using fine forceps and a small gauge needle, tease the sporocysts apart, releasing the cercariae. The most infective (mature) cercariae will be those that swim to the top of the water and hang there.
· With the aid of a dissecting microscope, pick up the cercariae in the meniscus of the hairloop and place them on the shaved and moistened abdomen of an anesthetized mouse. For most purposes a good exposure number is 20-25 cercariae per mouse.
To obtain S. japonicum cercariae in large numbers, it is necessary to dissect the secondary sporocysts from the tissues in order to release them. This obviously means the snails cannot be used at a later time for cercarial production, as is the case with Bulinus and Biomphalaria snails. Each positive Oncomelania hupensis ssp. snail, though, can typically yield between 300-500 infective cercariae.
Since the pathology of S. japonicum infection in a mouse is considerably greater on a worm pair basis than that of a S. mansoni infection, low cercariae numbers (e.g., 20-30 per mouse) are usually used in experimental situations. It is best to have cercariae from at least 10 snails to expose mice, to ensure mixed sex infections.
The main difficulty of mouse exposures with S. japonicum is related to the extremely sticky nature of the cercariae. Since they readily adhere to any plastic or glass surface, it is best to handle them in the meniscus of a hairloop or similar tool.
Liang, Y-S., Bruce, J.I., and Boyd, D.A. 1987. Laboratory cultivation of schistosome vector snails and maintenance of schistosome life cycles. Proceedings of the First Sino-American Symposium 1: 34-48.
Tucker, M. S., Karunaratne, L. B., Lewis, F. A., Frietas, T. C., and Liang, Y-S. 2013. Schistosomiasis, in Current Protocols in Immunology 19.1.1-19.1.57, John Wiley and Sons, Inc., (R. Coico, Ed). Published online November 2013 in Wiley Online Library (wileyonlinelibrary.com). doi: 10.1002/0471142735.im1901s103.