A break down of our latest BMC Evolutionary Biology paper

As promised, in this lab note I will break down our latest paper, published open access in BMC Evolutionary Biology, for you. Since it appeared online last month it hasalready been accessed 7863 times! Something that would never have happened ifthe paper were published behind a pay wall. Open access is really paying off. Addto this the many media outlets that have reported about it and a recommendationon F1000 Prime, and quite some people across the globe have at least picked upsomething about the work we have done!

Here youcan find 3 of my favorite write-ups:

 Turns Out the U.S. Has Its Very Own Species of Ant-Zombifying Fungus 

 Zombie Fungus Enslaves Only Its Favorite Ant Brains 

 Zombifying Fungus Recognizes the Brain of its Favorite Host 

The paperstarts with us introducing that the fungi of the complex Ophiocordyceps unilateralis aren’t just found in the tropics. TheUS has its very own species as well! The species that we use in this paper isone that we have found in South Carolina. In fact, citizen scientist KimFleming did. Kim is a mathematics teacher who loves to go for walks in thewoods behind her house with her camera to photograph all kinds of critters. Assuch, she discovered the zombie ants in her very own back yard! Since then shehas been working together with our lab and we get to visit her yearly to do ourfieldwork. And because of her invaluable work, she is also an author on ourpublished paper. Here are two of the many beautiful pictures she has taken:

 

 

[Camponotus americanus and Camponotus castaneus infected with Ophiocordyceps unilateralis, credit: Kim Fleming]

While doingour fieldwork we realized that 2 Carpenter ant species in the area are infectedand manipulated to bite by Ophiocordycepsbut another, very frequently encountered species, is not. Of course thiscan have many reasons: maybe these ants never encounter the spores, or thefungal spores are simply not able to infect these ants. But since we came upwith a way to infect ants in the lab by injecting them we were able to test ifthese environmental factors would be the (only) barrier that prevents this fungusfrom infecting all Carpenter ant species in the area.

What wefound after injecting was quite remarkable: the fungus was able to kill allthree ant species as a result of the infection, but only manipulated the 2 thatit is able to manipulate in nature as well. So, even when we take away all theenvironmental factors and make sure infection takes place, the non-host speciesis still not manipulated! In fact, the fungus was not even able to grow out ofthe ant cadaver after death, as is seen for the natural host species.Behavioural manipulation therefore seems to be rather species-specific.

Here’s afigure to recap this quickly for you, before we move on to the more complicatedmolecular biology stuff:

 

[Experimental set up used for the infection experiments, showing that the non-host ant species tested cannot be manipulated while the natural hosts are. C. castaneus photo credit: Roel Fleuren. C. americanus and C. pennsylvanicus photo's © Alex Wild,used by permission. Naturally infected C. castaneus and C. americanus photo credit: Kim Fleming]

So, it thusappears that the species-specificity of the manipulation is at least partly dueto the interaction of the fungus with the host while growing inside the ants’body. Now, brain manipulation is most likely achieved by the fungus secretingbioactive compounds onto the ants’ brains. Therefore, as a next step, we let Ophiocordyceps interact with the brainsof different ant species. To do this, we kept ant brains alive in “jars” andthrew in the fungus. To measure what the fungus would secrete as a reaction tothese different species brains, we then harvested the medium in which they werekept and measured all the compounds that were in there. These compounds werecompared to control samples in which we only had the fungus, only had the antbrains, and only had the medium to be able to tell which compounds were therebecause the fungus secreted them to affect the different brains it waspresented with.

 

[The "brain in the jar" experiment. The inserted picture shows you what the drawing of the ant brains and the fungus look like in real life]

Then, wecompared the results we got from the different species interactions to see ifthe fungus reacted the same way or differently to them. What we found was thatapparently Ophiocordyceps is able torecognize that it is dealing with a different ant brain because most of thecompounds it would secrete were different for the various interactions. Thiscould explain why we don’t find every old ant species turned intozombie ants. The parasite-host interaction to achieve control of the host isapparently a very specific one.

[Video of a still alive Camponotus castaneus ant infected in the lab with O. unilateralis, recorded during manipulated biting behavior]

Now, lastbut not least, we have tried to identify what the candidate compounds are thatwe have found. This is a very difficult process at the moment, so most of themstay undiscovered for now, but we have been able to identify 2: guanidinobutyricacid (GBA) and sphingosine. When searching the literature for both of these,they appear to be involved in neurological diseases. This makes theminteresting candidates as ants infected with Ophiocordyceps have some sort of neuro-disease as well (as theirbrains tell them to go bite a leaf instead of forage for food!). But, it alsoshows us the potential of discovering that this fungus indeed secretessome very interesting neuromodulators that might be good candidates for noveldrug discovery.

[Mass spectra of the 2 candidate compounds we have identified. To verify if the identification was indeed correct we bought standards of the compounds. The top is what we have found in the fungal secretion samples and the mirror images are the standards of those compounds we have used to show the spectra are indeed identical.]