Apr 12, 2023
Version 2
Infection of Biomphalaria glabrata snails with Schistosoma mansoni miracidia V.2
- Sarah K Buddenborg1,
- Geetha Sankaranarayanan1,
- Magda E Lotkowska1,
- Catherine McCarthy1,
- Carmen L Dias Soria1,
- Lisa Seymour1,
- Gabriel Rinaldi1,
- Matt Berriman1
- 1Wellcome Sanger Institute
Protocol Citation: Sarah K Buddenborg, Geetha Sankaranarayanan, Magda E Lotkowska, Catherine McCarthy, Carmen L Dias Soria, Lisa Seymour, Gabriel Rinaldi, Matt Berriman 2023. Infection of Biomphalaria glabrata snails with Schistosoma mansoni miracidia. protocols.io https://dx.doi.org/10.17504/protocols.io.36wgqjkkxvk5/v2Version created by Sarah K Buddenborg
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: April 11, 2023
Last Modified: April 12, 2023
Protocol Integer ID: 80330
Keywords: miracidia, schistosoma, biomphalaria
Funders Acknowledgement:
Wellcome Trust
Grant ID: 098051
Wellcome Trust
Grant ID: 206194
Abstract
To infect Biomphalaria glabrata snails with miracidia hatched from Schistosoma mansoni eggs
Materials
MilliQ waterContributed by users
24-well Clear TC-treated Multiple Well Plates Individually Wrapped SterileCostarCatalog #3524
Blunt featherweight forceps wide tipBioQuipCatalog #4750
1x DPBSGibco - Thermo FischerCatalog #14190144
Glass Pasteur pipettes with rubber bulbsContributed by users
1x Aquarium Water (diluted from 10X; see recipes)
1-2L flask
Aluminum foil
B. glabrata snails 5-8mm
S. mansoni-infected livers in PBS
28°C room or incubator
Incubator at 37°C and 5% CO2
Bucket of ice
Equipment
Zoom stereomicroscope with diascopic illumination stand
NAME
stereomicroscope
TYPE
Nikon
BRAND
SMZ800N
SKU
10X AQUARIUM WATER
5.56g CaCl2
12.28g MgSO4-7H2O
0.43g K2SO4
4.2g NaHCO3
480µl FeCl3-6H2O (0.5g/100ml water)
Fill to 10L and store in 1L bottles
Dilute to 1x: 9L MQ water + 1L 10x
Liver preparation
Liver preparation
Collect livers from patent mice into 50ml Falcon tubes containing pre-warmed 37°C 1x DPBS
Remove livers from PBS and place in large mortar or laboratory blender
Gently homogenise liver tissue
Place homogenised liver slurry into a flask (size of flask depends on how many livers you have) and fill with diH2O or aquarium water
Hatching miracidia
Hatching miracidia
Cover flask in aluminum foil and shine light horizontally across the opening of the flask for 1-2 hrs
Take a small aliquot of water from the top of the flask using a glass Pasteur pipette and place into a petri dish. Using a stereomicroscope, check for swimming miracidia
Exposing snails to miracidia for regular life cycle maintenance
Exposing snails to miracidia for regular life cycle maintenance
Under a microscope, collect 15 miracidia with a glass Pasteur pipette per well
Alternatively (but less preferred) estimate the number of miracidia by counting 12-5µl aliquots in a petri dish. After collecting (shaking the tube in between) and placing the aliquots in the petri dish add 5µl of Lugol to each drop (this kills and stains the miracidia)
Place individual 5-8mm snails into 24-well culture plates and cover snails completely with 1x aquarium water
Expose snails to miracidia for at least 3 hours (up to overnight)
After exposure, remove the snails carefully using featherweight forceps and put them in a new tank with food
Exposing snails to miracidia for monomiracidium infections
Exposing snails to miracidia for monomiracidium infections
Dilute miracidia so that one miracidium can easily be collected in ~3-5µl of water
Collected single miracidium using fresh 10µl pipette each collection and place in 24-well plate
After a plate is filled, check each well under a microscope to verify there is a single miracidium in each well
Place individual 5-8mm snails in the wells containing confirmed single miracidium and cover snails completely with 1x aquarium water
Leave the plate overnight (inside incubator or room at 28°C)
The following day, transfer the snails to a new tank with food