Population and Evolutionary Genetic Analyses Using Multiple Nuclear and Mitochondrial Loci in Steller Sea Lions.

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Population and Evolutionary Genetic Analyses
Using Multiple Nuclear and Mitochondrial Loci in
Steller Sea Lions.
John W. Bickham, Ryan M. Huebinger, Thomas R.
Gelatt, Greg OCorry-Crowe, and Joseph I. Hoffman

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ACKNOWLEDGEMENTS

• Project began in 1992
• I thank the following for providing samples
• T. Loughlin, T. Gelatt, V. Burkanov and D.
  Calkins
• Southeastern Alaska and PWS-ADFG
• Gulf of Alaska and Aleutian Is.-NOAA sponsored
  cruises

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ACKNOWLEDGEMENTS

• Russia Vladimir Burkanov, Nikolay Pavlov,
  Vladimir Vertynkin, Evgeny Mamaev.
• The Alaska SeaLife Center, Don Calkins
• For help in the laboratory and for data analysis
  I thank Alyson Baker, Robert Trujillo, John
  Patton, Cole Matson and Dmitri Politov.

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Eastern Stock East of 144W
Western Stock West of 144W
Population Estimates in Thousands
1960s
1970s
1980s
1990s
1994
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Population Trends

• The total SSL population numbered between 240,000
  and 300,000 during the late 1950s and early
  1960s.
• Only 116,000 were estimated range-wide in 1989.

-1969-
-1979-
-1989-
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Previous Genetic Studies

• Bickham et al. (1996)
• Discovered high genetic differentiation between
  populations on either side of 144W subsequently
  recognized an eastern stock (threatened) and
  western stock (endangered)
• Also found high diversity of mtDNA haplotypes
  and a clear pattern of macrogeographic variation

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The Western and Eastern Stocks
RUSSIA
ALASKA
PWS
60
BER
OKH
KAM
CGA
SEA
COM
WGA
EAL
WAL
CAL
BRC
45
KUR
ORE
NCA
JAPAN
30
135W
180
135E
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Previous Genetic Studies (cont)

• Bickham et al. (1998)
• Found evidence for three possible groups Asian
  and central groups within the western stock and
  an eastern stock
• Could have arisen due to two, possibly three,
  glacial refugia that separated the populations
  during the Pleistocene era

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A Third Stock?

• Previous research suggests that animals sampled
  from Russian rookeries are
• genetically distinct from other western stock
  sea lions.

RUSSIA
ALASKA
PWS
Cape Suckling
60
BER
OKH
KAM
CGA
SEA
COM
WGA
EAL
WAL
CAL
BRC
45
KUR
ORE
144W
NCA
JAPAN
30
135W
180
135E
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Previous Genetic Studies (cont)

• Baker et al. (2005)
• Subdivided the Western Stock into Asian Stock
  (OKH, KUR, KAM) and Western Stock (COM east
  through PWS)

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In this presentation we will

• Summarize a 14-year range-wide population-genetic
  analysis based upon the mtDNA control region of
  Steller sea lions.
• Present preliminary results from an ongoing
  complementary study of cytochrome b sequences.
• Compare phylogeographic and population genetic
  patterns for control region vs. cyt b sequences.
• Present data from nuclear microsatellite loci
  that corroborate some of the findings of stock
  structure in Steller sea lions.

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Control Region Sample Sizes
Stock Region Sample Size
EASTERN STOCK NCA (1 ROOKERY) 51
ORE (1 ROOKERY) 85
BRC (2 ROOKERIES) 23
SEA (3 ROOKERIES) 76
TOTAL 235

WESTERN STOCK PWS (2 ROOKERIES) 153
BER (1 ROOKERY) 42
CGA (6 ROOKERIES) 197
WGA (5 ROOKERIES) 108
EAL (6 ROOKERIES) 275
CAL (10 ROOKERIES) 247
WAL (3 ROOKERIES) 98
COM (1 ROOKERY) 126
TOTAL 1246

ASIAN STOCK KAM (2 ROOKERIES) 71
KUR (5 ROOKERIES) 207
OKH (2 ROOKERIES) 237
TOTAL 515

GRAND TOTAL 1,710
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Cytochrome b Sample Sizes
Stock Region Sample Size
EASTERN STOCK NCA (1 ROOKERY) 27
ORE (1 ROOKERY) 32
BRC (2 ROOKERIES) 15
SEA (3 ROOKERIES) 341
TOTAL 415

WESTERN STOCK PWS (2 ROOKERIES) 27
BER (1 ROOKERY) 21
CGA (7 ROOKERIES) 114
WGA (5 ROOKERIES) 32
EAL (6 ROOKERIES) 84
CAL (9 ROOKERIES) 73
WAL (3 ROOKERIES) 51
COM (1 ROOKERY) 46
TOTAL 448

ASIAN STOCK KAM (2 ROOKERIES) 63
KUR (5 ROOKERIES) 74
OKH (2 ROOKERIES) 133
TOTAL 165

GRAND TOTAL 878
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Microsatellite Sample Sizes
Stock Region Sample Size
EASTERN STOCK NCA (1 ROOKERY) 25
ORE (1 ROOKERY) 25
BRC (2 ROOKERIES) 18
SEA (3 ROOKERIES) 45
TOTAL 113

WESTERN STOCK PWS (2 ROOKERIES) 75
BER (1 ROOKERY) 13
CGA (3 ROOKERIES) 43
WGA (3 ROOKERIES) 30
EAL (4 ROOKERIES) 172
CAL (3 ROOKERIES) 70
WAL (1 ROOKERY) 12
COM (1 ROOKERY) 25
TOTAL 440

ASIAN STOCK KAM (1 ROOKERY) 25
KUR (4 ROOKERIES) 40
OKH (2 ROOKERIES) 50
TOTAL 115

GRAND TOTAL 668
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L-shaped distribution of control-region
haplotypes No evidence of a significant
bottleneck or reduction in variation
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Control Region Haplotype Distributions
Haplotype LLL
Haplotype BB
0.10
0.50
0.08
0.40
0.06
0.30
Haplotype Frequency
Haplotype Frequency
0.04
0.20
0.02
0.10
0.00
0.00
KUR
CAL
EAL
BER
BRC
NCA
KUR
CAL
EAL
BER
BRC
NCA
OKH
COM
CGA
SEA
ORE
OKH
COM
CGA
SEA
ORE
KAM
WAL
PWS
KAM
WAL
PWS
WGA
WGA
Haplotype E
Haplotype H
0.12
0.50
0.10
0.40
0.08
0.30
Haplotype Frequency
Haplotype Frequency
0.06
0.20
0.04
0.10
0.02
0.00
0.00
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Cytochrome b Haplotype Distributions
Haplotype 1
Haplotype 4
1.00
1.00
0.90
0.90
0.80
0.80
0.70
0.70
0.60
0.60
0.50
0.50
Haplotype Frequency
Haplotype Frequency
0.40
0.40
0.30
0.30
0.20
0.20
0.10
0.10
0.00
0.00
Haplotype 2
Haplotype 10
1.00
1.00
0.90
0.90
0.80
0.80
0.70
0.70
0.60
0.60
0.50
0.50
Haplotype Frequency
Haplotype Frequency
0.40
0.40
0.30
0.30
0.20
0.20
0.10
0.10
0.00
0.00
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NJ Tree for Rookeries Based on Fst
Estimates Control Region
Adak Island CAL
Akun Island EAL
Seal Rocks PWS
Clubbing Rocks WGA
Yunaska Island CAL
Ugamak Island EAL
Chirikof Island CGA
Amak Island EAL
Bogoslof Island EAL
Pinnacle Rock WGA
Chowiet Island CGA
Marmot Island CGA
Adugak Island EAL
Attu Island WAL
Amchitka Island CAL
Seguam Island CAL
Medney Island COM
Kasatoshi Island CAL
Sugarloaf Island CGA
Akutan Island EAL
Agattu Island WAL
Kozlova Cape RUS
Walrus Island BER
Fish Island PWS
Kiska Island Lief Cove CAL
Kiska Island Cape St. Stephen CAL
Buldir Island WAL
Gramp Rock CAL
Atkins Island WGA
Legend Asian Stock Western Stock Eastern Stock
Tag Island CAL
Ogchul Island EAL
Antsiferova Island KUR
Zheleznaya Bay KAM
Raykoke Island KUR
Iony Island OKH
Tuleniy Island OKH
Srednego Island KUR
Yamsky Island OKH
Lovushki Island KUR
Chernyye Bratya KUR
Forrester Island SEA
North Danger Rocks BRC
St. George Reef NCA
Triangle Island BRC
Hazy Islands SEA
Rogue Reef ORE
White Sisters Islands SEA
0.005 changes
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NJ Tree Regions Based on Fst Estimates Control
Region
OKH
KUR
KAM
COM
CAL
EAL
CGA
WGA
PWS
BER
WAL
SEA
Legend Eastern Stock Western Stock Asian Stock
BRC
ORE
NCA
0.005 changes
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NJ
Zalophus
A
XXXX
UUUUU
IIII
NNN
EEEEE
TTTTT
E
ZZZ
F
NJ tree of control region haplotypes shows two
distinct eastern stock haplotype lineages.
XX
VV
AAAAA
FFFF
B
R
FFFFF
RRRR
UU
XXXXX
P
SSSS
III
Q
C
YYYYY
D
H
GGGG
SS
BBBBB
I
YY
J
IIIII
LL
GGGGG
L
CCCCC
M
ZZ
BBBBBB
CCC
N
VVVV
DDDDD
K
O
WWWW
UUUU
OO
WW
Z
QQ
JJJ
FFF
DDD
QQQQQ
AA
PP
KKK
YYY
RRRRR
BB
GGG
YYYY
LLL
TTT
BBBB
MMM
JJJJ
RRR
SSS
VVVVV
AAAA
HHHH
RR
S
MMMMM
OOOOO
Y
KKKKK
NNNNN
QQQ
OOOO
LLLLL
KKKK
PPPPP
LLLL
V
W
BBB
PPP
X
HH
MMMM
TTTT
II
GG
TT
JJ
MM
PPPP
KK
OOO
VVV
CC
HHHHH
DD
HHH
NNNN
AAAAAA
JJJJJ
T
NN
U
EE
UUU
ZZZZ
AAA
EEE
FF
CCCC
WWWWW
ZZZZZ
QQQQ
G
XXX
WWW
EEEE
DDDD
SSSSS
0.001 substitutions/site
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Minimum Spanning Network of Steller Sea Lion
Cytochrome b haplotypes n 878, 1040 bp 29
Variable Positions Ti/Tv 26/3 13 Silent, 13
Nonsynonymous Transitions 1 Silent, 2
Nonsynonymous Transversions 1st Position 10 (9
AAS) 2nd Position 6 (6 AAS) 3rd Position 13 (0
AAS) 14 Total Variable Amino Acids
29
5
16
34
33
28
18
1
20
22
?
25
24
7
19
9
17
23
21
8
3
27
11
12
200
4
14
13
32
15
25-50
100-200
26
2
10-25
31
50-100
2-10
10
6
30
?
White Line Silent Mutation Red Line Amino Acid
Shift
1
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Regional Cytochrome b Slatkins Linearized FST
Midpoint Rooted NJ Phylogram
OKH
KUR
KAM
COM
CAL
CGA
WAL
WGA
BER
EAL
PWS
Legend Eastern Stock Western Stock Asian Stock
BRC
ORE
SEA
NCA
0.05 changes
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NJ Phylogram of Rookeries Based on Microsatellite
Data
Asian Stock
Western Stock
Legend Eastern Stock Western Stock Asian Stock
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NJ Phylogram of Regions Based on Microsatellite
Data.
Legend Eastern Stock Western Stock Asian Stock
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Conclusions Control Region Phylogeography

• Despite the generally high sample sizes in this
  study, resolution of population substructure
  increased when rookeries were grouped into their
  respective regions.
• Control region sequences clearly support the
  recognition of three stocks.

• Cytochrome b Phylogeography
• Clearly differentiates the eastern and western
  stocks but does not resolve the Asian stock.

• Microsatellite Analysis
• Nuclear microsatellites show a pattern nearly
  the same as for the cytochrome b gene.
• Clearly differentiates the eastern and western
  stocks but does not resolve the Asian stock.

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The Asian Stock

• The recognition of three management units may
  be appropriate for Steller sea
• lion conservation.

RUSSIA
ALASKA
PWS
Cape Suckling
60
BER
OKH
KAM
CGA
SEA
COM
WGA
EAL
WAL
CAL
BRC
45
KUR
ORE
144W
NCA
JAPAN
30
135W
180
135E