28 Minutes Later: Investigating the role of aflatrem-like compounds in Ophiocordyceps parasite manipulation of zombie ants

Abstract

Parasites and their hosts often share intimate coevolutionary relationship that can lead to bizarre phenotypes over time. Ophiocordyceps is one such genus that has evolved to manipulate the behavior of its various insect and arachnid hosts, the clearest examples of which are found in ants of the Camponotus tribe, colloquially known as zombie ants. While the unusual behaviors induced during infection are well described, the molecular driving force behind these changes are still unknown. Recent genomic and transcriptomics analyses have identified several candidate secreted proteins and secondary metabolites highly upregulated during infection. Of particular interest amongst these compounds is an aflatrem-like ergot alkaloid related to the mycotoxin aflatrem known to cause neurological issues in other animals, issues collectively called staggers syndrome. To test if aflatrem-like compounds play a role in Camponotus floridanus manipulation, healthy ants were injected with purified aflatrem and their behavior was monitored for 30 minutes. RNA-Seq was also performed on the heads of aflatrem-injected ants to compare any changes at the transcriptomic level to data available for ants infected by Ophiocordyceps camponoti-floridani. Behavioral assay footage was analyzed using the machine learning tool MARGO, which showed a negative correlation between activity and speed, and dose of aflatrem. Behaviors were also scored manually, identifying a positive coloration between aflatrem dose and staggering behavioral phenotypes. RNA-seq discovered 262 genes significantly up or down regulated compared to sham injected ants. Of these genes, 108 were also differentially regulated in the heads of late-stage Ophiocordyceps-infected ants. Together, these results indicate that aflatrem-like compounds may play an important role during late-stages of infection by reducing activity in ants, keeping them in elevated microclimates optimal for the production and dispersal of spores.

Neural mechanisms of parasite-induced summiting behavior in “zombie”Drosophila

“…Seven other compounds were significantly more abundant in summiting versus non-summiting flies across our replicate experiments: three of these could not be identified (MW 276.08, 179.08 and 429.15 g/mol) and the other four were putatively identified as guanosine, uridine, cytosine and 5-methylcytosine. Future collection of large quantities of summiting flies and fractionation approaches could be used to home in on compounds of interest and determine their chemical structure such that these compounds can be produced synthetically and assayed for behavioral effects (Beckerson et al, 2022). Cytosine is a pyrimidine nucleobase used in both DNA and RNA, a core molecular building block.…”

“…On the host side, evidence in NPV-infected L. dispar and Helicoverpa armigera point to changes in the host phototactic pathway underlying summiting behavior (Bhattarai et al, 2018; Liu et al, 2022). Outside of NPVs, work in Ophiocordyceps suggests that the parasitic fungus may use enterotoxins and small secreted proteins to mediate end-of-life “zombie” behaviors (Beckerson et al, 2022; de Bekker et al, 2015; Will et al, 2020), potentially targeting host phototaxis (Andriolli et al, 2019), circadian rhythm, chemosensation, and locomotion (de Bekker et al, 2015; Trinh et al, 2021; Will et al, 2020).…”

Using machine learning to predict protein-protein interactions between a zombie ant fungus and its carpenter ant host

“…Behavioral changes preceding this final summit could include host-adaptive responses, parasite-adaptive manipulations, or general symptoms of disease that span hyperactivity, uncoordinated foraging, decreased nestmate communication, and convulsions [9,[14][15][16][17]. While these modified ant behaviors have been observed in nature for some time [18], explorations of the fungal effectors involved are relatively more recent endeavors [14,[19][20][21][22][23][24][25]. These multi "-omics" studies have provided hypotheses about the parasite and host molecules that play a role in establishing the behavioral phenotypes observed, but leave their potential interactions open to interpretation.…”