Hello!
we have been infecting Anopheles gambiae G3 mosquitoes with malaria parasites, but on the external surface of the midgut of a few mosquitoes at 8-9 days post-infection we observed also yellowish protrusions with a bubbly interior (I have included some pics, midguts were stained with 0.25% mercurochrome). We suspect they may be microsporidia. Has anyone experience with this? We don't know how prevalent are in the colony but I have been reading about ways of getting rid of them, alternative to colony destruction, like rinsing the eggs with water or 1% HCl.
Many thanks for whoever can help with this!
Hi there, I think it may well be.
"Methods in Anopheles Research" is a super useful guide and has some info on how best to detect (handy PCR), and control microsporidia/alternate infections in cultures. Check out chapter 2.2:
https://www.beiresources.org/Portals/2/PDFS/2014MethodsinAnophelesResearchManualFullVersionv2tso.pdf
"Infected colonies may need to be replaced or isolated to ensure the infection does not spread to other insect colonies. Some microsporidian infections are self-limiting and will resolve themselves within 1-2 generations. However, others are persistent due to transmission through the embryos. Routine disinfection of pans and surfaces should limit the number of resistant spores."
Hi,
Yes, it does look like microsporidia. I was going to mention the same thing as Joshua - "Methods in Anopheles Research" is super useful.
Another easy way to check for microsporidia, apart from dissecting adults, is to put a L3/L4 (i.e. big) larva on a slide in a small amount of water (alive) and cover with coverslip. Have a look under the microscope - if you see motile creatures (zoospores), possibly of different sizes, that's a sure sign of infection, and you might also spot spores attached to the intestine.
If you possibly can, get rid of the colony. In any case, decontaminate everything (larval bowls, cages, netting, food for larvae, aspirators, etc) with hot water, ethanol and bleach. Bleach the eggs. Regularly change the water in larval bowls. Rinse pupae. Always prepare new sugar solution (and use autoclaved bottles). And you might want to autoclave larval food in small batches (you can then freeze them and take out of freezer as required).
Hope this helps! Good luck.
I would suggest this is a melanized oocyst. Which means it is an oocyst that not developed fully. Furthermore, the sporozoites already dead inside.
Hope your colony is better by now. What I can see from your midguts pictures are fat bodies and not microsporidia. They are very common in guts and sometimes they have the same shape/size of an oocyst. Melanized oocysts are black.
Must correct myself. These are epithelial cells. See Griffin et al. 2016. S2 supporting material.
Thanks D. Calvo , this is reassuring. I wasn't entirely sure myself about the microsporidia at the time.
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