Snail husbandry for maintaining the Schistosoma mansoni life cycle

Label clean aquarium with appropriate information in permanent marker (i.e. date, owner, age, numbers of individuals, exposure date, expected patent date, experiment number, etc). Example labels are attachedinfected_new tanks.avery uninfected_new tanks.avery

Rinse a generous handful of autoclaved oyster shells with water until it runs clear and add to the base of the tank

Fill aquarium 1/2-3/4 full with 1x Aquarium Water (see recipe in "MATERIALS" section)

For infected snails, keep water level at or just below 1/2 full to reduce chance of splashing when handling

Place a clean airline and air stone in aquarium and put in appropriate heated incubator

Connect airline and assure that the appropriate amount of air is going to the tank and all other tanks connected to the same airline. Note that changing the number of tanks in a connected airline results in decreased or increased air pressure to other tanks so adjusted accordingly

Add one 15ml falcon tube styrofoam rack to breeder tanks (snails also love to lay eggs on artificial lotus leaves)

Add snails to the tank (<100 baby snails/10L water; <10 adult breeder snails/10L; <30 juvenile parasite-exposed snails/10L)

Place one cube (15mm x 15mm) of snail gel per 6-8 adult snails or two per 100 baby snails (use size and number of snails to estimate amount of snail gel to give to each tank)

To prevent overgrowth of bacteria, do not overfeed snails. If any snail gel is leftover >1 day after a feeding, then the snails are being overfed and reduce as necessary

Visually inspect the snail tanks during feedings for heavy bacterial and/or waste contamination (turbid and/or smelly water, large amount of debris collected in oyster shells) and/or heavy rotifer infestations. If any of these is detected, rinse the snails thoroughly with fresh 1x Aquarium Water and prepare a new tank, airline, and bubbler for the snails

Once a week top up water on tanks to their appropriate levels using MilliQ H₂O. To top up the water use the hose connected to the MilliQ water source

Replace 10% of the dirtiest tanks each week

Fill a 100ml beaker or small dish with 1x Aquarium Water

Working with one styrofoam rack at a time, use featherweight forceps to gently remove the egg masses from the rack and place in the beaker (slide forceps under egg mass and lift away gently)

Repeat for all styrofoam racks from breeder tanks

Check the egg masses under the microscope for rotifer infestation. Discard individual egg masses with heavy rotifer infestations. Rinse egg masses if rotifers are present in the water until none are seen under microscope

When finished collecting, empty the beaker into a prepared nursery/baby tank for that particular week (1 new nursery/baby tank per week) rinse container well with 1x Aquarium Water to remove stuck baby snails/egg masses

Split any tanks that have >100 baby snails to promote growth

Remove airline with air stone and place in tank. If applicable, keep styrofoam rack in tank.

Remove any snails that might be kept – leave dead shells. Add 20ml of 5000ppm Klorsept and leave overnight (minimum 6 hours)

After this period, dispose of the decontaminated water using the sieve or colander to collect oyster shells and dead snails

Rinse tank with diH₂O and then spray tank with 70% EtOH. Scrub tank with green scouring pad, rinse thoroughly with diH₂O and allow to air dry

Styrofoam racks, air stones, and airline should be sprayed with 70% EtOH, rinsed liberally with diH₂O, and air dried

Put decontaminated and rinsed oyster shells and dead snails in an autoclave tray and autoclave when full