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Elucidating the molecular mechanisms behind the behavioural manipulation of ‘zombie’-flies by the entomopathogenic fungus, Entomophthora muscae

Nunn, Adam (2018) BINP32 20171
Degree Projects in Bioinformatics
Abstract
The basal fungus Entomophthora muscae is an obligate insect pathogen specific to adult dipterans such as Musca domestica. It is ostensibly regarded as a parasite that induces aberrant behaviour in the host species which is adaptive to the fungus at the expense of the host. Specifically, during the later stages of infection flies exhibit a behaviour sequence just prior to death which is considered to aid in the transmission of fungal spores. The timing of this phenomenon has been shown previously to be governed in a “gated-pattern” under a circadian rhythm, whereby an unknown condition has to be met by a specific time each day before the behaviour sequence is triggered. It is suggested herein that this unknown condition may be related to... (More)
The basal fungus Entomophthora muscae is an obligate insect pathogen specific to adult dipterans such as Musca domestica. It is ostensibly regarded as a parasite that induces aberrant behaviour in the host species which is adaptive to the fungus at the expense of the host. Specifically, during the later stages of infection flies exhibit a behaviour sequence just prior to death which is considered to aid in the transmission of fungal spores. The timing of this phenomenon has been shown previously to be governed in a “gated-pattern” under a circadian rhythm, whereby an unknown condition has to be met by a specific time each day before the behaviour sequence is triggered. It is suggested herein that this unknown condition may be related to resource depletion, as the fungus exhibits dimorphism and a switch to an invasive growth pattern immediately prior to host death.
A dual RNA sequencing experiment was conducted on infected houseflies collected at various stages of infection, including immediately before and after behaviour manipulation, to characterise the test system and elucidate molecular mechanisms in the fungus that are differentially expressed between these stages. Gene set enrichment analysis revealed a suite of M16 metalloproteases upregulated just prior to host manipulation, and trypsin-like serine proteases that were upregulated just after. The MAPK signal transduction pathway was also differentially expressed during this time, which is a known precursor to filamentous/invasive growth in fungi such as Saccharomyces cerevisiae following resource depletion. (Less)
Popular Abstract
Manipulation of ‘zombie’-flies by an insect pathogenic fungus

The fungus Entomophthora muscae is a highly specialised parasite of the housefly Musca domestica. Infections typically take hold over the course of a single week, during which time the host exhibits aberrant behaviour that appears to benefit the fungi at the expense of the host species. To determine how the fungus might be influencing the fly in this manner, the variation in fungal gene expression was studied over the course of the infection by sequencing mRNA at different stages of the disease progression.

There is something inherently terrifying about seeing others lose control of their own faculties, especially if it means that the deadly fungus feeding on their insides... (More)
Manipulation of ‘zombie’-flies by an insect pathogenic fungus

The fungus Entomophthora muscae is a highly specialised parasite of the housefly Musca domestica. Infections typically take hold over the course of a single week, during which time the host exhibits aberrant behaviour that appears to benefit the fungi at the expense of the host species. To determine how the fungus might be influencing the fly in this manner, the variation in fungal gene expression was studied over the course of the infection by sequencing mRNA at different stages of the disease progression.

There is something inherently terrifying about seeing others lose control of their own faculties, especially if it means that the deadly fungus feeding on their insides might take over and find a way to get you! Fortunately, to our knowledge evolution has so far spared humans from such peril; however the same cannot be said for the common housefly, M. domestica. In the final hours of infection an unknown trigger will cause the fly to climb steadily to an elevated position where it then transfixes to a surface, thereby maximising the chance of spreading fungal spores to new hosts. There is no discernible reason for the fly to do this but indeed this behaviour, regarded ostensibly as summit disease, has been observed in several similar host-parasite interactions involving insects.

In the present thesis the total gene expression in E. muscae was characterised for the first time over the course of the disease progression, using RNA sequencing and de novo assembly of the fungal transcriptome. Samples were taken at several time points including both immediately before and after behavioural manipulation had taken place in the host. Previous studies which reported attributes of E. muscae during fungal pathogenesis were presently corroborated with regards to gene expression. Specifically, focus was given to signal transduction pathways involved in fungal dimorphism. This refers to the way in which the fungus interprets the environment and responds to this information by changing its pattern of growth. As the timing of fungal dimorphism coincides with summit disease in the housefly, the mechanisms regulating it may also be candidates for regulating behavioural manipulation.

Differential expression analysis
By considering the genes that were expressed differently before and after behaviour manipulation had taken place, a suite of protein-digesting enzymes were identified as potentially having involvement in this phenomenon. Additionally, genes involved in the signal transduction pathway “MAPK” were identified; a known precursor to fungal dimorphism in fungi such as Saccharomyces cerevisiae which can trigger in response to resource depletion. Perhaps the fungus triggers summit disease in the host after reaching a plateau in resource utilisation? Perhaps pathways such as “MAPK” are responsible for stimulating invasive growth, which in turn utilise protein-digesting enzymes for penetrating host critical systems and tissues? Such morphological observations have been made in previous studies regarding E. muscae infections of M. domestica, but the specific molecular mechanisms driving these processes are seldom reported.

Messenger RNA sequencing and differential expression analysis allows for a glimpse at the different molecular mechanisms that are active in the pathogen during the manipulation of ‘zombie’-flies. Deriving biologically meaningful conclusions, however, requires further testing of these mechanisms in the lab. Complex interaction networks often exist between genes, and genes frequently have multiple functions within different cells and tissues. Mechanisms for behavioural manipulation therefore cannot be elucidated by gene expression alone, but it does provide a molecular roadmap for those that may be worth closer consideration.

Future insights into intricate host-pathogen interactions such as these may lead to a greater understanding of underlying disease archetypes in modern medicine. In the meantime, if a microbial ecologist tells you that something is “driving them up the wall” then you’ll just have to hope for the best.

Master’s Degree Project in Biology & Bioinformatics 60 credits 2018
Department of Biology, Lund University

Advisor: Dag Ahrén
Molecular Cell Biology, Department of Biology (Less)
Please use this url to cite or link to this publication:
author
Nunn, Adam
supervisor
organization
course
BINP32 20171
year
type
H2 - Master's Degree (Two Years)
subject
language
English
id
8935584
date added to LUP
2018-02-12 11:27:19
date last changed
2018-02-12 11:27:19
@misc{8935584,
  abstract     = {{The basal fungus Entomophthora muscae is an obligate insect pathogen specific to adult dipterans such as Musca domestica. It is ostensibly regarded as a parasite that induces aberrant behaviour in the host species which is adaptive to the fungus at the expense of the host. Specifically, during the later stages of infection flies exhibit a behaviour sequence just prior to death which is considered to aid in the transmission of fungal spores. The timing of this phenomenon has been shown previously to be governed in a “gated-pattern” under a circadian rhythm, whereby an unknown condition has to be met by a specific time each day before the behaviour sequence is triggered. It is suggested herein that this unknown condition may be related to resource depletion, as the fungus exhibits dimorphism and a switch to an invasive growth pattern immediately prior to host death. 
A dual RNA sequencing experiment was conducted on infected houseflies collected at various stages of infection, including immediately before and after behaviour manipulation, to characterise the test system and elucidate molecular mechanisms in the fungus that are differentially expressed between these stages. Gene set enrichment analysis revealed a suite of M16 metalloproteases upregulated just prior to host manipulation, and trypsin-like serine proteases that were upregulated just after. The MAPK signal transduction pathway was also differentially expressed during this time, which is a known precursor to filamentous/invasive growth in fungi such as Saccharomyces cerevisiae following resource depletion.}},
  author       = {{Nunn, Adam}},
  language     = {{eng}},
  note         = {{Student Paper}},
  title        = {{Elucidating the molecular mechanisms behind the behavioural manipulation of ‘zombie’-flies by the entomopathogenic fungus, Entomophthora muscae}},
  year         = {{2018}},
}