Using Principles of Conservation Genetics to Inform Propagation

David J. BergDepartment of BiologyMiami University, Ohio

Use of Propagation for Conservation

• Save species from extinction• Reintroduce extirpated populations• Augment extant populations• Provide individuals for research / education

Modified from IUCN (1987)

Goals of Propagation

Create populations that:• survive in captivity• have a high probability of survival in the wild• retain evolutionary potential

Problem

How do wemaximize “good” variation

increased survival in the wildmaintenance of evolutionary potential

whileminimizing “bad” variation?

loss of variationadaptation to captivity

Population Genetics Refresher

Gene: nucleotide sequence coding for a protein

Allele: one version of a geneuniparental: haploidbiparental: diploidhomozygous vs. heterozygous

Locus: physical location on a chromosome

Population Genetics RefresherMutation: change in nucleotide sequence

SubstitutionAATCCTAA AATCCAAA

DeletionAATCCTAA AATCCAA

InsertionAATCCTAA AATCCTCAA

Neutral: no change in protein

Population Genetics Refresher

Evolution: change in allele frequency over time

Hardy-Weinberg equilibrium = no change IF• No mutation• Infinite population size• Random mating• Closed population• No selection

Modified from Frankham et al. (2010), page 433

Six stages of captive breeding1. Observe decline, characterize wild population(s)2. Found captive population(s)3. Expand captive population(s)4. Manage captive population(s) over generations5. Choose individuals for reintroduction6. Manage reintroduced population(s)

1. Characterize variation

Total variation within a population isimportant

Variation among populations must bemaintained

Common Markers

DNA sequences

– Mitochondrial DNAhaploid, maternal, neutral, 1 locus

– Nuclear DNA~ haploid, neutral or under selection, ≥ 1 locus

http://www.mtdnatest.com/reagents-kits/mtdnatest-human/

Common Markers

Microsatellitesdiploidneutral10s of loci

http://genomics.cafs.ac.cn/ssrdb/index.php?do=about

Future Markers

• SNPs (genomics)diploid, neutral or under selection1000s of loci

Measures ofwithin-population variation

• MicrosatellitesNA = allelic richnessH = heterozygosity (HO and HE)Ne = genetically effective population size

• SequencesNumber of haplotypesk = average number of nucleotide differences

Genetically EffectivePopulation Size

Target population with census size (Nc) = xhas genetic variation equal to an “ideal”population of size y.

Constant Nc

50:50 sex ratioSmall, random variation in family size

Measures ofamong-population variation

• MicrosatellitesFST = (pooled variation – avg. variation)/pooled variation

variation among populations within riversvariation among rivers within regionsvariation among regions

Assignment tests

• SequencesΦST = FST analogue, accounting for sequence similarity

Black River(n = 154 / 6 sites)

Devils River(n = 3 / 3 sites)

Rio Grande(n = 58 / 5 sites)

Collection sites

Parsimony Network of COINhaplotype = 34

mtDNA COI

2

12

1

12

1

1

1

1

13

4 3

3

1

12

1

2

1

1

13

Black River (n = 146)Rio Grande (n = 10)

1

2

12

1

11

1

1

Black River (n = 146)Devils River (n = 3)Rio Grande (n = 58)

Inoue et al. (2015)

Microsatellites

BlackRiverN:154NA:5.9HE:0.506Ne:5870

DevilsRiverN:3NA:3.45HE:0.733

RioGrandeN:58NA:15.8HE:0.899Ne:22,600

Within-population genetic diversityNA = allelic richness

HE = expected heterozygosity

3%Among

populations

73%Within

populations

24%Among

drainages

Inoue et al. (2015)

CC Fall

DV RF1

BS RF3

DR RG1

RG2

RG3

RG4

RG5

DevilsRiver

Black River Rio Grande

0

0.5

1.0

Microsatellites Structure2 distinct clusters (k = 2)

Post

erio

r Pro

babi

lity

Group 1Group 2

Inoue et al. (2015)

St.Croix(SC)

Clinch(CR)

Gasconade(GR)

Meramec(MR)

Ouachita(OR)

COISequencesN=#ofindividualswith

Lineage1/Lineage2haplotypes

N=34/14

N=33/7

N=28/6

N=0/20

N=45/5SC

GR MR

CR

OR

3

251

1

1

5

1

111

2

11

23

2

2

1

52

11

1

2

11

11

28

1

17

1

2

12

Lineage1

Lineage2

COISequences

Inoue et al. (2014)

Within-populationgeneticdiversityNA =AllelicrichnessNE =Effectivepopulationsize

NA:12.8Ne:4,406

NA:13.8Ne:5,547

NA:13.9Ne:4,375

NA:8.0Ne:1,410

NA:13.2Ne:4,559

Microsatellites

Inoue et al. (2014)

Clin

ch

Gasc

onad

e

Mer

amec

Ouac

hita

St. C

roixClinch Gasconade Meramec Ouachita St. Croix

0

0.5

1.0

Post

erio

r pro

babi

lity

Structurek =2Microsatellites

Group1

Group2

Inoue et al. (2014)

Popenaias popeiiRio Grande has much greater within-population

variation than the Black RiverSignificant among-river variation

Manage as separate units

Cumberlandia monodontaOuachita population much lower variationLow among-population variation except for

Ouachita

Manage as two units, one covering large area

2. Found captive population(s)

Number of founders determineswithin-population variation

Demographic features are important

Size of founding population

Frankham et al. (2010), page 436

Ne = Nc, when50:50 sex ratio with random mating

Foos (1986)

3. Expand captive population(s)

Genetic drift = loss of genetic variation

Variance in reproduction must be random

Genetic diversity is lost over timeRate of loss depends on Ne

Foos (1986)

Ne = Nc when50:50 sex ratio with random mating

AND

variance in reproduction is random

Foos (1986)

4. Manage captive population(s)

Minimize inbreeding

Maintain Ne

Minimize adaptation to captivity

Frankham et al. (2010), page 442

Maximum avoidance of inbreeding

Ne = Nc, when50:50 sex ratio with random mating

ANDvariance in reproduction is random

ANDpopulation size is constant

Ne is harmonic mean of N over timelong-term Ne ~ t / Σ(1/N)

Ne and population sizeTime Population Size1 50002 1003 10004 30005 6000

Ne = 427

Selection

Selection is on heritable traits

Selection is on the phenotype:genotype + environment

Selection varies with environmentnatural vs. artificial selection

Frankham et al. (2010), page 460

SLOSS: Single large or several small populations?

Traits under positive selection in captivity can be deleterious in the wild

Frankham et al. (2010), page 457

Genetic deterioration in Drosophila

5. Choose individuals toreintroduce

Distribute evenly among families / lineages

If augmenting, do not swamp out natives

Maintain geographic boundaries

Destroy / repurpose excess individuals

Ne / Nc

Population Ne Nc Ne / Nc

Black River 5,870 48,006 0.12

Rio Grande 22,600 280,000 0.08

Burlakova & Karatayev (2013)Inoue et al. (2015)

6. Manage reintroducedpopulation(s)

Protect from abnormal mortality, allowmortality from “normal” sources

Maintain natural habitat

Maintain geographic integrity

Let evolution happen

Frankham et al. (2010), page 340

Ne is key!No evolution in culture is the goal:

1. No mutation2. Infinite population size3. Random mating4. No gene flow5. No selection

• Maximizing Ne will retain evolutionary potential

• Reintroduce and let nature take its course

Thanks to the following

Curt Elderkin, Kentaro Inoue, Emy Monroe,Ashley Walters

Brian Lang (in memorium)

CitationsBurlakova, L. E., A. Y. Karatayev (2013) Survey of Texas hornshell

populations in Texas. Interim Performance Report for Project #419446.Texas Parks & Wildlife Department, Austin.

Foos, T. J. (1986) Genetics and demography of small populations. ThePrzewalski Horse and Restoration to Its Natural Habitat in Mongolia.Food and Agriculture Organization of the United Nations, Rome.http://www.fao.org/docrep/004/ac148e/AC148E00.htm#TOC

Frankham, R., J. D. Ballou, D. A. Briscoe (2010) Introduction toConservation Genetics, 2nd Ed. Cambridge University Press, England.

Inoue, K., B. K. Lang, D. J. Berg (2015) Past climate change drivescurrent genetic structure of an endangered freshwater mussel.Molecular Ecology 24: 1910-1926.

Inoue, K., E. M. Monroe, C. L. Elderkin, D. J. Berg (2014) Phylogeographicand population genetic analyses reveal Pleistocene isolation followed byhigh gene flow in a wide-ranging, but endangered, freshwater mussel.Heredity 112: 282-290.

IUCN (1987) The IUCN Policy Statement on Captive Breeding, IUCN,Gland, Switzerland.