GENETIC DIVERSITY OF WILD vs. CULTIVATED/NATURALIZED ...biodiv.iptpo.hr/wp-content/uploads/2016/11/Satovic... · P30 S709 161 165 185 195 135 150. GENETIC DIVERSITY Wild Cultivated

Zlatko Zlatko ŠŠatoviatoviććUniversity of Zagreb, Faculty of AgricultureUniversity of Zagreb, Faculty of Agriculture,, Zagreb, CroatiaZagreb, Croatia

Centre of Excellence for Biodiversity and Molecular Plant BreediCentre of Excellence for Biodiversity and Molecular Plant Breeding, ng, Zagreb, CroatiaZagreb, Croatia

EE--mail: [email protected]: [email protected]

GENETIC DIVERSITY OFGENETIC DIVERSITY OFWILD WILD vvs. s. CCULTIVATEDULTIVATED//NATURALIZED NATURALIZED POPULATIONS OF DALMATIAN SAGEPOPULATIONS OF DALMATIAN SAGE

(1) Dalmatian sage (Salvia officinalis L.)

(2) Genetic diversity and relationships

(3) Genetic structure

(4) Ecological niche modelling

(5) Demographic history

PLANPLAN






PLANPLAN

DALMATIAN SAGEDALMATIAN SAGE

- Salvia officinalis L.

- outcrossing, insect-pollinated

- perennial subshrub- widely used since ancient times

for medicinal, culinary and ornamental purposes

- natural distribution:coastal region of the western Balkan and central and southern ApenninePeninsulas

- cultivation:throughout the Mediterranean region, in Australia,Germany, USA etc.

- naturalized populations:plants that have escaped from earlier cultivation

NATURAL DISTRIBUTION OF DALMATIAN SAGENATURAL DISTRIBUTION OF DALMATIAN SAGE

SVNHRV

BIH

MNE

MKDALB

GRC

ITA

KOS

SRB

ROM

MDA

BLG

HUN






PLANPLAN

GGENETIC DIVERSITYENETIC DIVERSITY

(1) Collecting

- leaf tissue for DNA extraction

- 30 populations / 709 samples (20 to 25 per populations) - origin:

11 Balkan countries

- status:23 wild populations

7 cultivated/naturalized populations

(2) DNA extraction

(3) Molecular marker analysis

- 8 microsatellite markers developed for Dalmatian sage

(simple sequence repeats; SSRs)

- 165 alleles

COLLECTINGCOLLECTING

Populations:

wild

cultivated

(naturalized)

KomiKomižža, a, iisland of sland of VVisis

CCaavtatvtat

(A) Isolation and characterization of microsatellites- construction of microsatellite libraries from genomic

DNA for GA, GT, AGA and ACA repeats- primer pairs flanking microsatellite repeats were

designed for 29 clones- 29 microsatellites were tested using 25 plants(B) Microsatellite marker analysis- eight most polymorphic markers: 165 alleles

MOLECULAR MAMOLECULAR MARRKER ANALYSISKER ANALYSIS

sequencing scoring

microsatellite region

ind2

ind3

ind1130100

172

142130

130

MOLECULAR DATAMOLECULAR DATA

Population Individual SoUZ001 SoUZ001 SoUZ002 SoUZ002 ... SoUZ019 SoUZ019

P01 S001 173 189 183 185 135 150

P01 S002 165 199 185 185 150 153

P01 S003 183 185 195 197 150 159

P01 S004 163 187 189 195 150 150

P01 S005 165 167 187 195 135 150

P01 S006 165 183 185 187 135 156

P01 S007 173 183 185 195 150 159

P01 S008 165 167 183 195 135 135

P01 S009 165 171 181 183 147 150

P01 S010 165 173 195 201 150 159

...

P30 S709 161 165 185 195 135 150

GGENETIC DIVERSITYENETIC DIVERSITY

Wild Cultivated

No. 23 7

Nav 8.696 3.821

Nar 7.920 3.672

Range 5.13-10.30 2.71-4.19

P(Nar) P < 0.001

Npr(total per population)

20 0

Npr(wild vs.

cultivated)

115 0

(1) Average no. of alleles per population (Nav)

(2) Allelic richness (Nar)- average no. of alleles per population independent of sample size

(3) No. of private alleles (Npr)- number of allelesdetected in a single population

(or in a group of populations: wild vs. cultivated)

AALLELIC RICHNESSLLELIC RICHNESS

Allelic richness:

< 5

5 - 7

7 - 9

> 9

GRC 1

GRC 2

ALB 2

MNE 2

MKD 1

MKD 2HRV 6

MNE 1

SRB 2

BIH 3BIH 2

BIH 1

ALB 1

BIH 4

HRV 8HRV 7

HRV 5

HRV 3

HRV 4

HRV 2

HRV 1

SVN 2

SVN 1

SRB 1 ROU 2

MDA 1

MDA 2

ROU 1KOS 1

KOS 2

0.01

81

99

99

100

99

72

CultivatedNaturalized

- distance matrix:Cavalli-Sforza and Edwards' chord distance

- tree algorithm:Neighbour-joining

- unrooted tree

- seven cultivated/naturalized populations groupedseparately from the restand formed a well-supported clade (bootstrap support 99%) suggesting the common origin of the cultivated plant material

GENETIC RELATIONSHIPSGENETIC RELATIONSHIPS

Bootstrap support:






PLANPLAN

- Bayesian model-based clustering method for inferring population structure

Assumption:

- there are K populations of origin each of which is characterized by a set of allele frequencies at each locus

Goal:

- assign individuals to populations of origin in such a way that within each population the departures from:

(1) Hardy-Weinberg equilibrium (HWE) and

(2) linkage equilibrium (LE)

are minimized

GENETIC STRUCTUREGENETIC STRUCTURE

PROCEDUREPROCEDURE

(1) Estimation of the number of Ks

populations of origin

(1.1) Choose different values of K

(1.2) Compute posterior probabilities for each K

(1.3) Choose the most likely number of populations (K)

(2) Assignment

(2.1) Assign individuals to populations (K)

(2.2) For each individal, estimate the proportion of genome derived from the different population of origin

GGENETIC STRUCTURE AT K = 2ENETIC STRUCTURE AT K = 2

100

75

50

25

0

K = 2

Q

Cluster: A B

P30

P29

P28

P27

P26

P25

P24

P23

P22

P21

P20

P19

P18

P17

P16

P15

P14

P13

P12

P11

P10

P09

P08

P07

P06

P05

P04

P03

P02

P01

BCultivatedpopulations

AWild

populations

- proportions of membership (Q) of each individual plant in each of the two clusters (population of origin):- each individual plant is represented by a single

vertical line divided into colors representing different clusters

- the length of the colored segment shows the individual’s estimated proportion of membership in that cluster

P26Sićevo Gorge

Serbia

SVN HRV BIH MNE ALB MKD GRE KOS SRB ROM MDA

100

75

50

25

0

100

75

50

25

0

K = 3

K = 2

Pro

port

ion o

f m

embers

hip (Q

)

Cluster: A B

Cluster: A1 A2 B

P30

P29

P28

P27

P26

P25

P24

P23

P22

P21

P20

P19

P18

P17

P16

P15

P14

P13

P12

P11

P10

P09

P08

P07

P06

P05

P04

P03

P02

P01


A1North-Westpopulations

A2South-Eastpopulations

- the cluster A (Wild populations) is at K = 3 split intotwo clusters according to geographical locationsA1 North-West: Adriatic populationsA2 South-East: Montenegro-Albania-Macedonia-Greece

SVN HRV BIH MNE ALB MKD GRE KOS SRB ROM MDA

GGENETIC STRUCTURE AT K = 3ENETIC STRUCTURE AT K = 3

100

75

50

25

0

100

75

50

25

0

100

75

50

25

0

K = 4

K = 3

K = 2

Pro

port

ion o

f m

embers

hip (Q

)

Cluster: A1 A2 A3 B

Cluster: A B

Cluster: A1 A2 B

P30

P29

P28

P27

P26

P25

P24

P23

P22

P21

P20

P19

P18

P17

P16

P15

P14

P13

P12

P11

P10

P09

P08

P07

P06

P05

P04

P03

P02

P01


A1NothernAdriatic

A2SouthernAdriatic

A3MacedoniaGreece

- wild populations split further into three clusters

GENETIC STRUCTURE AT K = 4GENETIC STRUCTURE AT K = 4

A1

A2 A3

B

0.00

0.05

0.10

0.15

0.20

0.25

0.30

0.00 1.00 2.00 3.00 4.00 5.00 6.00 7.00

0.00

0.05

0.10

0.15

0.20

0.25

0.30

0.00 1.00 2.00 3.00 4.00 5.00 6.00 7.00

Geographic distance

Genetic d

ista

nce r = 0.47

PMantel = 0.0001

R2 = 0.23

GENETIC STRUCTURE AT K = 4GENETIC STRUCTURE AT K = 4






PLANPLAN

ECOLOGICAL NICHE MODELLINGECOLOGICAL NICHE MODELLING

- species distribution modelling (SDMs)- estimate the relationship between species records at

sites and the environmental characteristics of those sites

- predict the suitability of sites for occupation or persistence of the species

- produce a modelled distribution of the species(= identify species' suitable environmental space)

- input data:(1) geographic distribution

68 data on occurrence (evenly distributed)(2) environmental characteristics of the sites

19 bioclimatic variables (WorldClim database)- 11 temperature- and 8 precipitation-related- representing the annual trends, seasonal variations and extremes in temperature and precipitation

0.26 - 0.50

0.51 - 0.75

0.76 - 0.94

Suitability

EENM: PRESENT DAY CONDITIONSNM: PRESENT DAY CONDITIONS

0.26 - 0.500.51 - 0.750.76 - 0.94

Suitability

Apennine Peninsula

Adriatic Sea

Balkan Peninsula

0.26 - 0.50

0.51 - 0.75

0.76 - 0.94

Suitability

EENM: PRESENT DAY CONDITIONSNM: PRESENT DAY CONDITIONS

0.26 - 0.500.51 - 0.750.76 - 0.94

Suitability

Apennine Peninsula

Adriatic Sea

Balkan Peninsula

A1

A2A3

PAST DISTPAST DISTRRIBUTIONIBUTION

- to model potential species distribution during the

Last Glacial Maximum (LGM; ~21,000 years BP)

- to identify putative glacial refugia of the species

(= region which made possible the survival of the species and allowed a post-glacial re-colonization)

- input:

(1) the present model

(2) past environmental characteristics of the sites

19 bioclimatic variables

bioclimatic data for the LGM developed by

Paleoclimate Modelling Intercomparison Project Phase II

- two models:

CCSM (Community Climate System Model; USA)

MIROC (Model for Interdisciplinary Research on Climate; Japan)

EENNMM: LAST GLACIAL MAXIMUM: LAST GLACIAL MAXIMUM CCCSM CONDITIONSCSM CONDITIONS

Balkan PeninsulaApennine Peninsula

0.26 - 0.500.51 - 0.750.76 - 0.94

Suitability

A3

Adriatic Sea

- CCSM: severe temperature decline

EENNMM: LAST GLACIAL MAXIMUM : LAST GLACIAL MAXIMUM MMIROCIROC CONDITIONSCONDITIONS

Suitability0.26 - 0.50

0.51 - 0.75

0.76 - 0.84

0.26 - 0.500.51 - 0.750.76 - 0.94

Suitability

Apennine Peninsula

Balkan Peninsula

A3

A1

A2

Adriatic Sea

- MIROC: moderate temperature decline






PLANPLAN

DEMOGRAPHIC HISTORYDEMOGRAPHIC HISTORY

- inference on demographic history of a species based on approximate Bayesian computation (ABC)

- input:- molecular data

- scenarios describing demographic history

- procedure:- simulate (a large number of) datasets for each scenario

- compare simulated and observed datasets

- the most similar simulated dataset is the most likely

- goal:- compare competing scenarios

posterior probability of each scenario

- estimate parameters for one or more scenarios

effective population size; time of divergence

PPOPULATIONSOPULATIONS

A1

A2A3

Allelic richness:

A1 7.97

A2 8.49

A3 6.25

ABC analysis:

A1: 92 inds from 7 pops



FFIVE SIMPLE IVE SIMPLE HHISTORIC ISTORIC SSCENARIOSCENARIOS

Scenario 1

Population A1 is derived from population A2,

itself derived from population A3

Scenario 2

Population A3 is derived from population A2,

itself derived from population A1

A1

A2

A3

A3 A2 A1

A1 A2 A3

A1

A2

A3

Scenario 3

Both populations A1 and A3 derived independently from

population A2

Scenario 4

Population A2 was generated by admixture of populationsA1 and A3

A1

A2

A3

A3 A2 A1

A1 A2 A3

A1

A2

A3


Scenario 5

All three populations

diverged at the same time

A2 A1 A3

A1

A2

A3


Scenario Posterior probability (PP)

1 0.166

2 0.193

3 0.075

4 0.126

5 0.440

RESULTS:RESULTS:

HISTORICAL RECONSTRUCTION: ANCIENT PASTHISTORICAL RECONSTRUCTION: ANCIENT PAST

?

Vegetation map of Europeduring the last glacial period

Distibution of Dalmatian sage before the last glaciation ?

ABC Effective population sizes:Ancestral 8,550 inds

A1 4,330 indsA2 7,190 indsA3 2,440 inds

Glacial refugia:1. Iberian Peninsula2. Apennine Peninsula3. Balkan Peninsula

Source: http://www.metatech.org/07/ice_age_global_warming.html


0.51 - 0.75

0.76 - 0.84

0.26 - 0.500.51 - 0.750.76 - 0.94

Suitability

Apennine peninsula

Balkan peninsula

A3

A1

A2

Adriatic sea


0.51 - 0.75

0.76 - 0.84

0.26 - 0.500.51 - 0.750.76 - 0.94

Suitability0.26 - 0.500.51 - 0.750.76 - 0.94

Suitability

Apennine peninsula

Balkan peninsula

A3

A1

A2

Adriatic sea

Distribution of Dalmatian sage during Distribution of Dalmatian sage during the Last Glacial Maximumthe Last Glacial Maximum

A2A3

A1

Map of the 52 putative refugia within the Mediterranean region

ABC Time of divergence:572 generations ago(95% CI: 157-1,540)

Generation time (life span):up to 300 years

HISTORICAL RECONSTRUCTION: HISTORICAL RECONSTRUCTION: DDIVERGENCEIVERGENCE

A2 A1 A3

Source: http://onlinelibrary.wiley.com/doi/10.1111/j.1365-2699.2008.02051.x/abstract

A1

A2 A3

B

HISTORICAL RECONSTRUCTION: PRESENT TIMEHISTORICAL RECONSTRUCTION: PRESENT TIME

Rešetnik, I., Baričević, D., Batîr Rusu, D., Carović-Stanko, K., Chatzopoulou, P., Dajić-Stevanović, Z., Gonceariuc, M., Grdiša, M., Greguraš, D., Ibraliu, A., Jug-Dujaković, M., Krasniqi, E., Liber, Z., Murtić, S., Pećanac, D., Radosavljević, I., Stefkov, Gj., Stešević, D., Šoštarić, I., Šatović, Z. 2016. Genetic Diversity and Demographic History of Wild and Cultivated/Naturalised Plant Populations: Evidence from Dalmatian Sage (Salvia officinalis L., Lamiaceae). PLOS One 11(7): e0159545 (doi: 10.1371/journal.pone.0159545)

Documents

GENETIC DIVERSITY OF WILD vs. CULTIVATED/NATURALIZED ...biodiv.iptpo.hr/wp-content/uploads/2016/11/Satovic... · P30 S709 161 165 185 195 135 150. GENETIC DIVERSITY Wild Cultivated