Periodic Reporting for period 1 - GenoSpec (Insights from Population Genomics to the Evolution of Host Specificity in Insect Fungi)
Periodo di rendicontazione: 2015-11-16 al 2017-11-15
The project address the problematics of host-specific adaptation in Ophiocordyceps unilateral sensu lato, known as “zombie-ant” fungi. This fungal species complex is pathogenic specifically to ants and well known for its outstanding feature to manipulate the behaviour of the ant hosts in order to favour its own dispersion. Infected ants become erratic and climb into vegetation before fixing themselves to vegetal materials until death. This fungal complex encompasses multiple sibling species specialised to different ant species. The objectives of the Genospec project were to identify genes underlying host-specific adaptation in O. unilateral sensu lato by using the approach of population genomics. We have identified species-specific genes and genes under diversifying selection between closely related, host-specific species of O. unilateralis, and we found that these were enriched in pathogenesis functions, and mostly in heat-labile enterotoxins.
Some entomopathogenic fungi are widely used as biocontrol agents in developing countries. However, their use is limited by the lack of knowledge on the evolution of virulence toward different host ranges. The knowledge gained from the Genospec project contributes to a better understanding of the host-specific adaptation in entomopathogenic fungi and could thus enhance the use of these organisms in organic farming.
Some entomopathogenic fungi are widely used as biocontrol agents in developing countries. However, their use is limited by the lack of knowledge on the evolution of virulence toward different host ranges. The knowledge gained from the Genospec project contributes to a better understanding of the host-specific adaptation in entomopathogenic fungi and could thus enhance the use of these organisms in organic farming.
Overview of the results, their exploitation and dissemination
Comparative genomic analyses have identified species-specific genes and genes under diversifying selection between closely related, host-specific species of the O. unilateralis complex, and we found that these were enriched in pathogenesis functions, and mostly in heat-labile enterotoxins. These results were reported in a manuscript, which is under revision for the journal Molecular Ecology. Population genomic analyses revealed population subdivision showing that the sibling species of the O. unilateralis complex were highly divergent with very low shared polymorphism. We found population subdivisions within the recognized sibling species, which could be explained by the presence of further cryptic species or phylogeographic patterns. The data of this part is still being analysed with a planned manuscript in 6 months.
Work performed from the beginning of the project
A total of 59 strains of three species of the O. unilateralis complex, specific to three different ant species, were collected across Thailand. They were sequenced with Illumina HiSeq3000 at high coverage. Initially, the raw reads of these samples were mapped back to the unique genome reference of O. polyrhachis-furcata (Wichadakul et al. 2015) for detection of single nucleotide polymorphisms (SNPs). This first set of analyses showed that the different species shared so little polymorphism that the scan of differentiation between species along the genome could not reveal any outlier regions that would allow us to detect genes under divergent selection. Therefore, we undertook first a study of comparative genomics in order to detect species-specific genes and genes with high rates of non-synonymous substitutions among species, likely revealing divergent selection. By comparing the genomes of the three closely related species from Thailand and a more distant taxon from the USA associated to distinct ant host species, we found that species-specific genes were enriched in pathogenesis functions, and mostly in heat-labile enterotoxins. Furthermore these genes were over-represented among those with significant footprints of positive selection; other categories of genes suspected to be important for virulence and pathogenicity in entomopathogenic fungi (e.g. chitinases, lipases, proteases, core genes of secondary metabolites) were much less represented, although a few candidates were found to evolve under positive selection. An analysis including orthologs from a broader phylogenetic context in entomopathogenic fungi showed that positive selection on enterotoxins was specific to the O. unilateralis complex. Together with previous studies showing overexpression of an enterotoxin during behavior manipulation of diseased ants, our findings support the view that heat-labile enterotoxins are important effectors involved in host adaptation and coevolution in Ophiocordyceps entomopathogenic fungi.
The de novo and improved assemblies of different species were then used as references to call SNPs in their respective species. SNPs for each species were used to infer intra-specific population structures and will be used in the next six months to detect selective sweeps.
Comparative genomic analyses have identified species-specific genes and genes under diversifying selection between closely related, host-specific species of the O. unilateralis complex, and we found that these were enriched in pathogenesis functions, and mostly in heat-labile enterotoxins. These results were reported in a manuscript, which is under revision for the journal Molecular Ecology. Population genomic analyses revealed population subdivision showing that the sibling species of the O. unilateralis complex were highly divergent with very low shared polymorphism. We found population subdivisions within the recognized sibling species, which could be explained by the presence of further cryptic species or phylogeographic patterns. The data of this part is still being analysed with a planned manuscript in 6 months.
Work performed from the beginning of the project
A total of 59 strains of three species of the O. unilateralis complex, specific to three different ant species, were collected across Thailand. They were sequenced with Illumina HiSeq3000 at high coverage. Initially, the raw reads of these samples were mapped back to the unique genome reference of O. polyrhachis-furcata (Wichadakul et al. 2015) for detection of single nucleotide polymorphisms (SNPs). This first set of analyses showed that the different species shared so little polymorphism that the scan of differentiation between species along the genome could not reveal any outlier regions that would allow us to detect genes under divergent selection. Therefore, we undertook first a study of comparative genomics in order to detect species-specific genes and genes with high rates of non-synonymous substitutions among species, likely revealing divergent selection. By comparing the genomes of the three closely related species from Thailand and a more distant taxon from the USA associated to distinct ant host species, we found that species-specific genes were enriched in pathogenesis functions, and mostly in heat-labile enterotoxins. Furthermore these genes were over-represented among those with significant footprints of positive selection; other categories of genes suspected to be important for virulence and pathogenicity in entomopathogenic fungi (e.g. chitinases, lipases, proteases, core genes of secondary metabolites) were much less represented, although a few candidates were found to evolve under positive selection. An analysis including orthologs from a broader phylogenetic context in entomopathogenic fungi showed that positive selection on enterotoxins was specific to the O. unilateralis complex. Together with previous studies showing overexpression of an enterotoxin during behavior manipulation of diseased ants, our findings support the view that heat-labile enterotoxins are important effectors involved in host adaptation and coevolution in Ophiocordyceps entomopathogenic fungi.
The de novo and improved assemblies of different species were then used as references to call SNPs in their respective species. SNPs for each species were used to infer intra-specific population structures and will be used in the next six months to detect selective sweeps.
The molecular mechanisms underlying host-specificity between hosts and parasites is an active research field. For entomopathogenic fungi, knowledge on the genetic basis of host-specificity is still limited. This question has rarely been considered under the aspect of evolutionary comparative and population genomics. Our project used such approaches for identifying genes underlying host-specific adaptation in a group of pathogenic fungi manipulating insect behaviors. Our findings contribute not only to a better understanding of adaptation in entomopathogenic fungi but also in eukaryotes in general. Indeed, the small genomes of fungi allowed us to perform genome-wide evolutionary genomic analyses that are still restricted to a handful of plant and animal model species so far.
We have now obtained all required data, we have performed comparative genomic analyses and we will perform population genomic analyses in the next six months, which will reveal further outstanding insights. the manuscript currently under revision, we expect to be able to submit two additional manuscripts.
We have now obtained all required data, we have performed comparative genomic analyses and we will perform population genomic analyses in the next six months, which will reveal further outstanding insights. the manuscript currently under revision, we expect to be able to submit two additional manuscripts.