Conservation Genetics:

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Conservation Genetics

• the link between population genetics and the
  conservation of biodiversity.

Dr. Marie L. Hale
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Conservation genetics

• Conservation genetics is the application of
  genetics to preserve species as dynamic entities
  capable of coping with environmental change.
• The development of conservation genetics has been
  driven by what many consider to be a global
  environmental crisis the sixth extinction
• Very young science
• First journal in 2000
• First textbook in 2002

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Molecular tools

• Many different markers
• mtDNA sequences
• chloroplast DNA sequences
• nuclear DNA sequences
• microsatellites
• minisatellites
• AFLP (amplified fragment length polymorphisms)
• RAPD (random amplified polymorphic DNA)
• Marker chosen depends on the question to be
  answered.

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What are microsatellites?

• Short tandem repeats (2 6 bp).
• Highly variable among individuals in the number
  of repeats.
• High mutation rate (replication slippage).
• Scattered throughout the genome.
• Believed to be selectively neutral.

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Microsatellite variation

• Microsatellite region amplified with PCR
  (Polymerase Chain Reaction).

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Microsatellite data
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Why are they so useful?

• High level of polymorphism means we can generally
  tell individuals apart.
• Very short fragments so can amplify really small
  amounts of DNA which means can devise
  non-invasive sampling methods (e.g. hair and
  faeces), or old museum specimens.
• Lots of microsatellites spread throughout the
  genome, so we can get lots of independent
  information for each individual. Makes these a
  very powerful tool for resolving relationships.
• Disadvantages?
• Have to identify microsatellite regions for each
  new species and design primers to amplify (with
  PCR) the specific microsatellite regions in each
  species. (This takes anything from 2-6 months
  work and costs around 10,000 per species).

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What sort of questions can we answer with them?
Source of invasions / colonisation routes
Dispersal / structure
Hybridisation / introgression
Impact of habitat change
Parentage / mating systems
Level of diversity
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Red squirrel conservation genetics.

• Red squirrels (Sciurus vulgaris) native to
  Britain but severe population decline due to loss
  of habitat and introduction of American grey
  squirrel (Sciurus carolinensis).
• Little known about remaining populations level
  and pattern of diversity, dispersal, habitat use,
  impact of habitat fragmentation etc.
• This knowledge is essential to devise a
  management plan to conserve remaining populations.

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Squirrel data collected.

• 102 individuals genotyped at 4 microsatellite
  loci.
• Location of each sample recorded (Ordnance survey
  grid reference).
• Date collected recorded (many were museum
  specimens).

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Structure dispersal.

• Question 1 How many populations are there?
• Question 2 What is the minimum distance between
  habitat patches that acts as a barrier to
  dispersal between patches?

Habitat patches plotted from GIS data and
satellite photos. Habitat highly fragmented
approx 174,000 separate forest fragments
(separated by more than 25m) in study area.
0 20 40km
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Structure dispersal.

• populations are groups of interbreeding
  individuals.
• So natural populations should be in
  Hardy-Weinberg equilibrium at selectively neutral
  loci (assuming little migration, mutation,
  drift).

If we include samples from two or more real
populations as a single population in our
analysis, we find a deficit of heterozygotes from
that expected under Hardy-Weinberg equilibrium
(the Wahlund effect).
10km
10km
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Structure dispersal.
We can determine maximum usual dispersal
distance between habitat patches by grouping
sampling locations based on distance between the
patches.
The maximum distance between patches at which
samples are in Hardy-Weinberg equilibrium is the
maximum usual dispersal distance.
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Dispersal in squirrels

• Squirrels collected in habitat patches separated
  by ? 1.5km of open ground formed populations in
  Hardy-Weinberg equilibrium.
• When samples collected in habitat patches gt 1.5km
  apart were included as a single population
  there was a deficit of heterozygotes i.e.
  non-random mating.

deficit of heterozygotes significantly gt
0. Usual dispersal distance ? 1.5km.



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Squirrel populations

• Squirrel populations include all samples
  collected within groups of forest patches
  separated by ? 1.5km of open ground.
• 10 populations in sample area.

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Genetic differentiation

• Question 3 Are the populations genetically
  differentiated?
• Question 4 What is the geographic pattern of any
  differentiation?

AMOVA (Analysis of Molecular Variance)
Fixation index FST 0.161 p lt 0.0001
Squirrel populations are genetically
differentiated
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Pairwise genetic distance

• Calculate pairwise genetic distance between all
  possible pairs of populations.
• A distance of 0.0 no differentiation a
  distance of 1.0 complete differentiation.

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Genetic differentiation
Unrooted UPGMA Neis genetic distance

Hale et al. (2001) Science 293 2246-2248.
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Regional differentiation
Do the 3 regions represent real distinct
groupings of squirrels, or are they just an
artificial construct of the clustering
analysis? At what level is the genetic
differentiation important? Is there more genetic
differentiation among regions or among
populations within regions? (Should we just
conserve regions or are individual populations
important?)
Hale et al. (2001) Science 293 2246-2248.
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Are the regions real?

• 3 level AMOVA for hierarchical structure

Fixation indices FST 0.215 p lt 0.0001
(total differentiation) FSC 0.059 p lt
0.0001 (differentiation among
populations within regions) FCT 0.165 p lt
0.0001 (differentiation among regions)
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Genotype assignment

• Genotype assignment methods use the entire
  multilocus genotype to determine the likelihood
  of each individual being a member of each
  predefined population.

Individual multilocus genotypes
Allele frequencies in populations for Scv10
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Genotype assignment

• Example of likelihood values for squirrels caught
  in the Northern region

For each region, significantly more squirrels
were assigned to the region they were collected
in than the other two regions (P lt 0.001).
This will only occur if there are real genetic
differences between the regions.
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c)
Regions are the basic conservation unit for
squirrels.
Hale et al. (2001) Science 293 2246-2248.
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Are squirrels using corridors?

• Question 5 How have changes to the landscape
  affected red squirrel populations?

Kielder Forest planted 1945 1960. Trees
matured during 1980s. Has this new conifer
forest created a biological corridor connecting
squirrel populations?
Kielder Forest
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Temporal variation
Collection dates (decades) for squirrel specimens
Range 1861 to 2000, 110 individuals.
Hale et al. (2001) Science 293 2246-2248.
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Temporal variation
Eastern no change in gene frequencies over time.
FST -0.015, P 0.728, df
3,56. Western gene frequencies have changed
over time. FST 0.128, P lt 0.001,
df 4,55.
Significant pairwise FST values are indicated in
red (P lt 0.05). All decades prior to 1980 are
significantly different from all decades after
the 1980s.
Hale et al. (2001) Science 293 2246-2248.
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Temporal variation

FST 0.128, P lt 0.001, df 4,55.

Hale et al. (2001) Science 293 2246-2248.
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Change in Cumbria
Pre 1980.
Hale et al. (2001) Science 293 2246-2248.
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Red squirrel conclusions

• Habitat fragments separated by ? 1.5km of open
  ground can be considered a single habitat
  containing a single population.
• The three regions should be treated as distinct
  entities for management.
• The integrity of the 3 regions has been lost with
  the recent connection of the Northern and Western
  regions through Kielder Forest. Genetic diversity
  in the Western region is in danger of being lost
  due to swamping by migrants from the Northern
  region.

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Resources

• Frankham R., Ballou J.D. Briscoe D.A. (2002)
  Introduction to Conservation Genetics. Cambridge
  Uni Press.
• Issues of the journal Conservation Genetics.
  Publisher Springer Netherlands.
• Hedrick P.W. (2001) Conservation genetics where
  are we now? Trends in Ecology Evolution 16
  629-636.
• DeSalle R. (2005) Genetics at the brink of
  extinction. Heredity 94 386-387.