Speciation : the case of fungi - Wiley Online Library · 2017. 10. 31. · MvSl (S. latifolia) MvSd (S. dioica) • Divergence date: ca. 420 000 years ago • Significant gene flow

Speciation : the case of fungi

Tatiana Giraud

University of South Paris

France

Importance of host shift speciation by fungal pathogens for agriculture and ecosystems

Emerging diseases

Phytophthora

Cryphonectria Chestnut blight

Mycosphaerella Septora blotch of wheat

Rhynchosporium secalis Scald on rye

Darwin’s doodles of anther smut

Fungi - and anther smut fungi in particular - are great models for studying eukaryote speciation

-experimentally tractable -access to haploid phase -short generation times -small genomes -applied consequences -some important model species

Fungi - and anther smut fungi in particular - are great models for studying eukaryote speciation

Microbotryum

Saccharomyces

Neurospora

Coyne & Orr

Coyne & Orr

How does reproductive isolation evolve?


(Gourbière & Mallet 2009)

Linear model: constant substitution rate and single gene incompatibilities

Snowball model: constant substitution rate and Dobzansky-Muller incompatibilities (multiple loci)

Slowdown model: decreasing substitution rate and single gene incompatibilities: reinforcement

How does reproductive isolation evolve in fungi?

0

0.25

0.5

0.75

1

0 0.05 0.1 0.15 0.2

Rep

roduct

ive

isola

tion

Genetic distance (ITS)

Pre

zygotic isola

tion

Sympatry

Allopatry


In basidiomycetes:

- increase of prezygotic isolation with genetic distance

- stronger in sympatry than in allopatry

- Best fit to a slowdown model: reinforcement in sympatry

(Le Gac & Giraud JEB 2008, Giraud & Gourbière Heredity 2012)

How does reproductive isolation evolve in the anther smut fungi?

S. dioica

S. latifolia

•Increase with genetic distance

•Best fit by the slow down model (Le Gac et al. Evolution 2007b, Giraud & Gourbière Heredity 2012)


Ability of F1 and F2 hybrids to cause disease


-No evidence of snowball effects in other fungi either

-No evidence for nuclear DBM genetic incompatibilities in genome scans

and genetic engineering

(e.g., Saccharomyces, Kao et al. PLoS Genet. 2010; Greig et al. 2007 PloS Genet,

Chou et al. 2010 PLoS Biol)

(Gourbière & Mallet 2009)

Identification of DBM genetic incompatibilities in fungi

-Some examples of mtDNA-nuclear DBM genetic incompatibilities and nuclear DBM pairs

-Evidence for the role of the mismatch repair system and karyotypic differences in post-zygotic infertility

-We need more studies in more fungi to draw generalities about the existence of DBM or other causes for post-zygotic isolation in fungi

Saccharomyces

Anderson et al. Curr Biol 2010

Lee et al. Cell 2008

Neurospora

Dettman et al. Evolution 2010

In ascomycetes


Apple scab

Grey mould on grapes

Ascochyta blight

0

0.25

0.5

0.75

1

0 0.01 0.02 0.03 0.04 0.05

Genetic distance (ITS)

Rep

rod

uct

ive

iso

lati

on

Sympatry

Allopatry

In ascomycetes:

• many close species have high interfertility in vitro,

even when in sympatry

(Le Gac & Giraud JEB 2008, Giraud et al. FGB 2008, Giraud & Gourbière Heredity 2012)


Apple scab

Grey mould on grapes

Ascochyta blight

Host adaptation Host

adaptation

a b a

b

A B

a b

A b

A B

a b

a B

Host 1 Host 2

Mating within hosts: pleiotropy between host adaptation and reproductive isolation

(« automatic magic trait », Servedio et al. 2011)

A: Host adaptation locus B: neutral locus Giraud, Gladieux and Gavrilets 2010 Trends in Ecology and Evolution

A B

A B

Conditions conducive to host shift speciation in ascomycete fungi

-Pleiotropy of host adaptation and mate choice: « automatic magic trait »

- Huge selection on single genes (gene-for-gene system in plant-fungi interactions: all-or-nothing)

Giraud, Gladieux and Gavrilets 2010 Trends in Ecology and Evolution

Allele Avr Allele Vr

Conditions conducive to host shift speciation in ascomycete fungi

-Pleiotropy of host adaptation and mate choice: magic trait

- Huge selection on single genes (gene-for-gene system in plant-fungi interactions)

- Billions of spores: -Opportunities for mutations -Allow for selection load

Giraud, Gladieux and Gavrilets 2010 Trends in Ecology and Evolution

• Resistance breakdown 10 years ago

• Magic trait

• Sympatric speciation within 40 years?

Natives on susceptible

apple trees

Emerging disease

on resistant apple

trees

Example: emerging disease on a resistant variety of apple

Apple scab

(Gladieux et al. 2011 Mol Ecol)

Coalescence and ABC methods: Current degree of gene flow and history of speciation

Coalescence and ABC methods: Current degree of gene flow and history of speciation

MvSl (S. latifolia) MvSd (S. dioica)

• Divergence date: ca. 420 000 years ago

• Significant gene flow from MvSd to MvSl (2 Nm=0.099)

• But only recently: secondary contact (Gladieux et al. MBE 2011)

Using NGS for population genomics

Stress response

Transmembrane proteins

Nutrient uptake

Secreted proteins

Hyphae growth

Expression regulation

(Aguileta et al. Mol Ecol 2010)

•EST libraries in 4 Microbotryum species specialized on different host plants

•42 genes under positive selection (dN/dS)

Genes of speciation /adaptation

Identification of genomic islands of introgression in fungi

(Neafsey et al. Genome Res 2010)

Cryptococcus Human pathogen

Adaptive introgression

Identification of genomic islands of divergence in fungi

(Ellison et al. PNAS 2011)

Neurospora Burnt vegetation

Climatic adaptation

Mickaël Le Gac

Odile Jonot

Lionel Saunois

Guislaine Refrégier

Jacqui Shykoff Damien de Vienne

Michael Hood

Léo and Romain

Michael Fontaine Elodie Vercken

Sergey Gavrilets

Pierre Gladieux

Credits

Bruno Le Cam

Amanda Gibson

Gabriela Aguileta Sébastien Gourbière

Selfing as a barrier to gene flow

• Selfing reduces gene flow

• But cannot be regarded as an “isolating barrier between species”?

Coyne and Orr, Speciation

Experimental test for selfing, combined to sibling selection arena, as barriers to gene flow

•Does the possibility of selfing reduce the rate of hybridization beyond what is expected based on intrinsic hybrid inviability and selfing rates?

•Would be evidence that competition between hybrid and selfed progeny further reduce gene flow: a true “isolating barrier”

(Gibson et al. Evolution 2012)

Experimental test for selfing, combined to sibling selection arena, as barriers to gene flow

•The possibility of selfing did reduce the rate of hybridization beyond what was expected based on intrinsic hybrid inviability and selfing rates

•Evidence that competition between hybrid and selfed progeny further reduce gene flow: a true “isolating barrier”


0

0,2

0,4

0,6

0,8

1

S-pair S-mixRat

e o

f H

ybri

d In

fect

ion

Expectedrate

-75%

-51%

Sporidia Teliospores

Microbotryum anther-smut fungi

Meiosis

Conjugation

(A1 and A2)

Dikaryotic hyphae invade meristems

Dispersal by pollinators

Spore production in anthers

Winter

No « magic trait »

0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1

0 0.01 0.02 0.03 0.04 0.05 0.06 0.07

Genetic distance between parents

Ab

no

rma

l p

he

rom

on

e r

ece

pto

r

co

nte

nt

0,00

0,20

0,40

0,60

0,80

1,00

1,20

1,40

1,60

1,80

0 0,01 0,02 0,03 0,04 0,05 0,06 0,07

Re

lati

ve

ge

no

me

siz

e

Genetic distance between parents

rho = 0.83; P = 0.0002

Karyotypic differences between Microbotryum species

rho = 0.71; P < 0.001

F1 hybrid sterility is associated with abnormal genome content

Genetic distance Genetic distance

(de Vienne et al. JEB 2009, Giraud & Gourbière Heredity 2012)

Best fit by the linear model

F2 hybrid genotypes

(de Vienne et al. JEB 2009)

Genotyping of F2 hybrids Heterozygous loci

Homozygous loci with the allele corresponding to the inoculated plant

Homozygous loci with the allele NOT corresponding to the inoculated plant

Inter-specific cross

MvSlxMvSd

Inter-specific cross

MvSlxMvLfc

Intra-specific cross

MvSlxMvSl

Increasing genetic distance between parents

(de Vienne et al. JEB 2009,

Giraud & Gourbière Heredity 2012)

• Best fit by the slowdown model


Ability of F2 hybrids to cause disease


Fungi, and anther smut fungi in particular, are fantastic models for stuying speciation

Mickaël Le Gac

Odile Jonot

Lionel Saunois

Guislaine Refrégier

Jacqui Shykoff Damien de Vienne

Michael E Hood

Léo and Romain

Michael Fontaine Elodie Vercken

Sergey Gavrilets

Pierre Gladieux

Credits

Bruno Le Cam

Identification of DBM genetic incompatibilities in fungi

-Some examples of mtDNA-nuclear DBM genetic incompatibilities and one nuclear DBM pair

-ATP-driven proton pump responsible for maintening pH gradient across the cell membrane

-A cluster of ATP-driven sodium efflux pumps

Saccharomyces

Anderson et al. Curr Biol 2010

Lee et al. Cell 2008

Inab

ilit

y to

pro

duc

e h

yphae


Ability of hybrids to grow hyphae:

-Increase of prezygotic isolation with genetic distance

-Best fit to a linear model

(Le Gac & Giraud JEB 2008, Giraud & Gourbière Heredity 2012)

rho = 0. 77, P = 0. 003

(de Vienne et al. JEB 2009, Giraud & Gourbière 2012 Heredity)

0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1

0 0.01 0.02 0.03 0.04 0.05 0.06 0.07

F1 g

amet

e s

teri

lity

• Best fit by the linear model Genetic distance


Ability of hybrids to mate

Stress response

Transmembrane proteins

Nutrient uptake

Secreted proteins

Hyphae growth

Expression regulation

(Gladieux et al. Heredity 2012)

•EST libraries in 4 Microbotryum species specialized on different host plants

•42 genes under positive selection (dN/dS)

•But under purifying selection at the within species level

Genes of speciation /host adaptation?

Cospeciations or host shifts?

•Congruence between host and parasite phylogenies?

Caryophyllaceae Microbotryum

Stellaria

Soffinialis

Grepens

Dcarthusianorum

Dmonspessulanus

Dsylvestris

Dgratianopolitanus

Lfloscuculi

Lflosjovis

Snutans

Sotites

Sacaulis

Sdioica

Slatifolia

Svulgaris

Slemmonii

Scaroliniana

Svirginica

MvStellaria

MvSaoff

MvDc

MvDsp1

MvDsp2

MvSot

MvLfc

MvSn

MvSv2

MvSd

MvSl

MvSlem

MvLfj

MvSv1

MvSa

MvSvir&caro

(Refrégier et al. BMC Evol Biol 2008)



Stellaria

Soffinialis

Grepens

Dcarthusianorum

Dmonspessulanus

Dsylvestris

Dgratianopolitanus

Lfloscuculi

Lflosjovis

Snutans

Sotites

Sacaulis

Sdioica

Slatifolia

Svulgaris

Slemmonii

Scaroliniana

Svirginica

MvStellaria

MvSaoff

MvDc

MvDsp1

MvDsp2

MvSot

MvLfc

MvSn

MvSv2

MvSd

MvSl

MvSlem

MvLfj

MvSv1

MvSa

MvSvir&caro

Frequent host shifts

But between close hosts




Stellaria

Soffinialis

Grepens

Dcarthusianorum

Dmonspessulanus

Dsylvestris

Dgratianopolitanus

Lfloscuculi

Lflosjovis

Snutans

Sotites

Sacaulis

Sdioica

Slatifolia

Svulgaris

Slemmonii

Scaroliniana

Svirginica

MvStellaria

MvSaoff

MvDc

MvDsp1

MvDsp2

MvSot

MvLfc

MvSn

MvSv2

MvSd

MvSl

MvSlem

MvLfj

MvSv1

MvSa

MvSvir&caro


Mostly host shift speciations

Pollinators and habitats are somehow different, but ecological isolation is incomplete (Van Putten et al. 2007, Goulson 1997, Karrenberg et al. 2008, Minder et al. 2007)

Intersterility? Pre-zygotic? Post-zygotic? Function of genetic distance between species?

MvSl

MvSd

MvSsp

MvDsp

MvSv

MvSspA

• Crossing experiments

• Viability and fertility of hybrids as a function of genetic distance between species

Evolution of reproductive isolation in Microbotryum

Gene flow concentrated on recent branches

•Secondary contact?

IM: isolation with migration (coalescence)

(Gladieux et al. MBE in press)

Structure of MvSl populations (Microsats+sequences)

3 clusters, recolonization from glacial refugia?

Selfing rate MvSl: 99%

Selfing rate MvSd : 95%

Structure of MvSd populations (Microsats+sequences)

MvSl

Sf Sx1

Sx2 Sx

MvSd MvSd MvSd

MvSd

MvSl

Shared polymorphisms

•Gene flow? •Ancestral polymorphism?

(Gladieux et al. MBE in press)

Host 1 Host 2

A B

A B

A B

a b a

b

A B

a b

A b

A B

a b

a B

A b

a B

Not the case when mating occurs outside the host

A: Host adaptation locus B: neutral locus

Host adaptation

Host adaptation

Host adaptation

Microbotryum violaceum anther-smut fungus

1. Ancient, easy to find, globally distributed pathogen

2. Fungus and its hosts are easily grown and crossed

3. Agriculturally and socially unimportant, but rich

scientific history 5. >500 host species

6. The existence of “host races” debated since

the early 20th century

7. Few morphological differences

Benefits of the Study System

Criteria for species recognition in fungi

– Morphology

– Intersterility

– Congruence between multiple gene genealogies

Individual 5

Individual 3

Individual 1

Individual 7

Gene B

Individual 6

Gene A

Individual 2

Individual 4

Individual 8

Gene C

Criteria for species recognition in fungi

– Morphology

– Intersterility

– Congruence between multiple gene genealogies

– 4 genes in Microbotryum

Individual 5

Individual 3

Individual 1

Individual 7

Gene B

Individual 6

Gene A

Individual 2

Individual 4

Individual 8

Gene C

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/99

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D. sylvestris

D. carthusianorum

D. monspessulanus

S. vulgaris

S. nutans

L. floscuculi

S. dioica

S. latifolia

S. acaulis

(Le Gac et al. Evolution 2007a)

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99

1.0/95/94

Microbotryum violaceum anther-smut fungus

female

male

aborted ovary

Effect on female and male hosts Diseased flower of Silene latifolia

Basidiomycete, pathogen of Caryophyllaceae


–Reproductive isolation has been found to increase with time/genetic divergence

–Pre-zygotic reproductive isolation is expected to be stronger in sympatry

–Reproductive character displacement has indeed been reported in several species

Coyne & Orr


Abnormal symptoms in hybrids

Mixtures of spores and pollen in anthers

(Sloan et al. JEB 2008)

Spores with ovules

•More than 100 host plants •Host-specialized species •Sympatric species •No « magic trait » •What are the barriers to gene flow? •How did reproductive isolation evolve?



Pro

po

rtio

n o

f co

nju

gati

on

s

Genetic distance

0

0.04

0.08

0.12

0.16

0.2

0 0.02 0.04 0.06 0.08

No evidence of assortative mating

(Le Gac et al. Evolution 2007b)


•No intrinsic assortative mating

•Post-zygotic isolation increasing with genetic distance between species, but remaining low between close species

•Surprising for sympatric species…

•More consistent with allopatry patterns

•Recent secondary contact?

•Role of selfing and a selection arena: fewer hybrids than expected based on selfing rates and hybrid fitness alone


Documents

Speciation : the case of fungi - Wiley Online Library · 2017. 10. 31. · MvSl (S. latifolia) MvSd (S. dioica) • Divergence date: ca. 420 000 years ago • Significant gene flow