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 O’Corry-Crowe, and Joseph I. Hoffman
ACKNOWLEDGEMENTSACKNOWLEDGEMENTS
Project began in 1992Project began in 1992
I thank the following for providing I thank the following for providing samples: samples:
T. Loughlin, T. Gelatt, V. Burkanov T. Loughlin, T. Gelatt, V. Burkanov and D. Calkinsand D. Calkins
Southeastern Alaska and PWS-Southeastern Alaska and PWS-ADF&GADF&G
Gulf of Alaska and Aleutian Is.-NOAA Gulf of Alaska and Aleutian Is.-NOAA sponsored cruisessponsored cruises
ACKNOWLEDGEMENTSACKNOWLEDGEMENTS
Russia: Vladimir Burkanov, Nikolay Pavlov, Russia: Vladimir Burkanov, Nikolay Pavlov, Vladimir Vertynkin, Evgeny Mamaev. Vladimir Vertynkin, Evgeny Mamaev.
The Alaska SeaLife Center, Don CalkinsThe Alaska SeaLife Center, Don Calkins
For help in the laboratory and for data For help in the laboratory and for data analysis I thank Alyson Baker, Robert analysis I thank Alyson Baker, Robert Trujillo, John Patton, Cole Matson and Trujillo, John Patton, Cole Matson and Dmitri Politov.Dmitri Politov.
Population Estimates for Steller Sea Lions
(1960-1994)
Pop
ula
tion
Est
imat
es in
Th
ousa
nd
s
0
20
40
60
80
100
120
140
160
180
200
1960s 1970s 1980s 1990s 1994
Eastern StockEast of 144°W
Western StockWest of 144°W
Population TrendsPopulation Trends
The total SSL population numbered between 240,000 and 300,000 during the late 1950’s and early 1960’s.
Only 116,000 were estimated range-wide in 1989.
-1969--1969- -1979--1979- -1989--1989-
30°
45°
60°
KAMOKH
BER
PWS
NCA
ORE
SEACOM
WAL CALEAL
WGA
CGA
BRC
KUR
RUSSIA ALASKA
135°W180°135°E
Sampled Rookeries
JAPAN
Previous Genetic StudiesPrevious Genetic Studies
Bickham et al. (1996):Bickham et al. (1996): Discovered high genetic differentiation Discovered high genetic differentiation
between populations on either side of 144˚W; between populations on either side of 144˚W; subsequently recognized an eastern stock subsequently recognized an eastern stock (threatened) and western stock (endangered)(threatened) and western stock (endangered)
Also found high diversity of “mtDNA Also found high diversity of “mtDNA haplotypes and a clear pattern of haplotypes and a clear pattern of macrogeographic variation”macrogeographic variation”
30°
45°
60°
KAMOKH
BER
PWS
NCA
ORE
SEACOM
WAL CALEAL
WGA
CGA
BRC
KUR
RUSSIA ALASKA
135°W180°135°E
Sampled Rookeries
JAPAN
Cape Suckling
144°W
WESTERN STOCK
EASTERN
STOCK
The Western and Eastern StocksThe Western and Eastern Stocks
Previous Genetic Studies (cont)Previous Genetic Studies (cont)
Bickham et al. (1998)Bickham et al. (1998) Found evidence for three possible groups: Found evidence for three possible groups:
Asian and central groups within the western Asian and central groups within the western stock and an eastern stockstock and an eastern stock
Could have arisen due to two, possibly three, Could have arisen due to two, possibly three, glacial refugia that separated the populations glacial refugia that separated the populations during the Pleistocene eraduring the Pleistocene era
A Third Stock?A Third Stock?
Previous research suggests that animals sampled from Russian rookeries are genetically distinct from other western stock sea lions.
30°
45°
60°
KAMOKH
BER
PWS
NCA
ORE
SEACOM
WAL CALEAL
WGA
CGA
BRC
KUR
RUSSIA ALASKA
Cape Suckling
135°W180°135°E
Sampled Rookeries
JAPAN
144°W
EASTERN STOCK
WESTERN STOCK
ASIAN STOCK
Previous Genetic Studies (cont)Previous Genetic Studies (cont)
Baker et al. (2005)Baker et al. (2005) Subdivided the Western Stock into Asian Subdivided the Western Stock into Asian
Stock (OKH, KUR, KAM) and Western Stock Stock (OKH, KUR, KAM) and Western Stock (COM east through PWS)(COM east through PWS)
In this presentation we will:In this presentation we will:
Summarize a 14-year range-wide population-Summarize a 14-year range-wide population-genetic analysis based upon the mtDNA control genetic analysis based upon the mtDNA control region of Steller sea lions.region of Steller sea lions.Present preliminary results from an ongoing Present preliminary results from an ongoing complementary study of cytochrome complementary study of cytochrome bb sequences. sequences. Compare phylogeographic and population Compare phylogeographic and population genetic patterns for control region vs. cyt genetic patterns for control region vs. cyt bb sequences.sequences.Present data from nuclear microsatellite loci that Present data from nuclear microsatellite loci that corroborate some of the findings of stock corroborate some of the findings of stock structure in Steller sea lions.structure in Steller sea lions.
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
Control Region 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
Cytochrome b 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
Microsatellite Sample Sizes
Individual Haplotype TotalsN= 1,433 IndividualsN= 132 Haplotypes
0
10
20
30
40
50
60
Number of Individuals per Haplotype
Nu
mb
er o
f H
aplo
typ
esL-shaped distribution of control-region haplotypes
No evidence of a significant bottleneck or reduction in variation
Haplotype LLL
OKHKUR
KAMCOM
WAL
CALEAL
BERW
GACGA
PWS
SEABRC
ORENCA
Hap
loty
pe F
requ
ency
0.00
0.02
0.04
0.06
0.08
0.10
Haplotype E
OKHKUR
KAMCOM
WAL
CALEAL
BERW
GACGA
PWS
SEABRC
ORENCA
Hap
loty
pe F
requ
ency
0.00
0.02
0.04
0.06
0.08
0.10
0.12
Haplotype H
OKHKUR
KAMCOM
WAL
CALEAL
BERW
GACGA
PWS
SEABRC
ORENCA
Hap
loty
pe F
requ
ency
0.00
0.10
0.20
0.30
0.40
0.50
Haplotype BB
Hap
loty
pe F
requ
ency
0.00
0.10
0.20
0.30
0.40
0.50
OKHKUR
KAMCOM
WAL
CALEAL
BERW
GACGA
PWS
SEABRC
ORENCA
Control Region Haplotype Distributions
Haplotype 1
0.00
0.10
0.20
0.30
0.40
0.50
0.60
0.70
0.80
0.90
1.00
Ha
plo
typ
e F
req
ue
nc
y
Haplotype 4
0.00
0.10
0.20
0.30
0.40
0.50
0.60
0.70
0.80
0.90
1.00
Ha
plo
typ
e F
req
ue
nc
y
Haplotype 2
0.00
0.10
0.20
0.30
0.40
0.50
0.60
0.70
0.80
0.90
1.00
Ha
plo
typ
e F
req
ue
nc
y
Haplotype 10
0.00
0.10
0.20
0.30
0.40
0.50
0.60
0.70
0.80
0.90
1.00
Ha
plo
typ
e F
req
ue
nc
y
OKHKUR
KAMCOM
WAL
CALEAL
BERW
GACGA
PWS
SEABRC
ORENCA
OKHKUR
KAMCOM
WAL
CALEAL
BERW
GACGA
PWS
SEABRC
ORENCA
OKHKUR
KAMCOM
WAL
CALEAL
BERW
GACGA
PWS
SEABRC
ORENCA
OKHKUR
KAMCOM
WAL
CALEAL
BERW
GACGA
PWS
SEABRC
ORENCA
Cytochrome b Haplotype Distributions
Adak Island CALAkun Island EALSeal Rocks PWSClubbing Rocks WGAYunaska Island CAL
Ugamak Island EALChirikof Island CGA
Amak Island EALBogoslof Island EALPinnacle Rock WGA
Chowiet Island CGAMarmot Island CGA
Adugak Island EALAttu Island WAL
Amchitka Island CALSeguam Island CAL
Medney Island COMKasatoshi Island CALSugarloaf Island CGAAkutan Island EAL
Agattu Island WALKozlova Cape RUS
Walrus Island BERFish Island PWS
Kiska Island Lief Cove CALKiska Island Cape St. Stephen CAL
Buldir Island WALGramp Rock CAL
Atkins Island WGATag Island CAL
Ogchul Island EALAntsiferova Island KUR
Zheleznaya Bay KAMRaykoke Island KUR
Iony Island OKHTuleniy Island OKH
Srednego Island KURYamsky Island OKH
Lovushki Island KURChernyye Bratya KUR
Forrester Island SEANorth Danger Rocks BRC
St. George Reef NCATriangle Island BRC
Hazy Islands SEARogue Reef ORE
White Sisters Islands SEA
0.005 changes
Legend
Asian Stock
Western Stock
Eastern Stock
NJ Tree for RookeriesBased on Fst Estimates
Control Region
OKH
KUR
KAM
COM
CAL
EAL
CGA
WGA
PWS
BER
WAL
SEA
BRC
ORE
NCA0.005 changes
NJ Tree RegionsBased on Fst Estimates
Control Region
LegendEastern StockWestern StockAsian Stock
ZalophusAXXXXUUUUU
IIIINNN
EEEEETTTTT
EZZZ
FXX
VVAAAAA
FFFFB
RFFFFF
RRRRUU
XXXXXP
SSSSIII
QC
YYYYYD
HGGGG
SSBBBBB
IYY
JIIIII
LLGGGGG
LCCCCCM
ZZBBBBBB
CCCN
VVVVDDDDD
KO
WWWWUUUU
OOWW
ZQQ
JJJFFF
DDDQQQQQ
AAPP
KKKYYY
RRRRRBBGGGYYYYLLLTTTBBBBMMM
JJJJRRR
SSSVVVVVAAAA
HHHHRR
SMMMMM
OOOOOY
KKKKKNNNNNQQQOOOO
LLLLLKKKKPPPPP
LLLL VW
BBBPPP
XHH MMMM
TTTTII
GGTT
JJMM
PPPPKK
OOOVVV
CCHHHHH
DDHHH
NNNNAAAAAA
JJJJJT
NNU EE
UUUZZZZ
AAAEEE
FFCCCC
WWWWWZZZZZ
QQQQG
XXXWWW
EEEEDDDD
SSSSS
0.001 substitutions/site
NJ
NJ tree of control region haplotypes shows two distinct eastern stock haplotype lineages.
Z
A
BB
S
CC
E
FF
H
G K
WHH
AAA
P
5F
4Y
5B5D
5C
CCC
5G
J
LL
D
B
PP
GG
TTMM
4P
RRR
SSS
OO
4X
NNN
5E
?
?
?
M
N
YY
I
?
?
? ?
?
4R
R
Q
4G
SS
4W
O
4V
L
ZZ
DD
4S
UU
KKK
FFF
JJJ
4A
MMM
LLL AA
JJ
WW
C4I
WWW
4E
4Q
YYY
5R
GGG
TTT4B
4H
4J
?
?
?
?
DDD
4F
VVV
RR
BBBV
4N
VV
OOO
4K
4L
PPP
4T4M
XII
5A
KK
T
U
EE
UUU
ZZZ
QQQ
4O
Y NN4C
5Q
4Z
F
XXX
XX
HHH
EEE
III
4U
4D
Cole W. MatsonApril 2003
Steller Sea Lion mtDNA Control RegionHaplotype Minimum Spanning Network
N = 1798
5W
5V
5U
5Y
6B
6A
5T
5S
5Z
5X
6J
6I
6H
?
6G
6F
6E
6D
6C
5I
5H
5O
5P5K
5L
5N
5M
5J
6O
6N
6M
6L
6K
WEST EAST
NON-ROOKERYSAMPLE
WIDESPREAD
ASIA
20-29
30-49
10-19
2-9
1
200+
100-199
50-99SCAT
WEST EAST
NON-ROOKERYSAMPLE
WIDESPREAD
ASIA
WEST EAST
NON-ROOKERYSAMPLE
WIDESPREAD
ASIA
WEST EAST
NON-ROOKERYSAMPLE
WIDESPREAD
ASIA
20-29
30-49
10-19
2-9
1
200+
100-199
50-99SCAT
20-29
30-49
10-19
2-9
1
200+
100-199
50-99SCAT
2
4
5
610
7
8
9
11
12
1314
15
17
16
23
18
19
20
21
22
24 25
26
27
?
33
30
29
28
34
31
32
?
1
3
Minimum Spanning Network of Steller Sea Lion Cytochrome b haplotypesn = 878, 1040 bp29 Variable PositionsTi/Tv = 26/313 Silent, 13 Nonsynonymous Transitions1 Silent, 2 Nonsynonymous Transversions1st Position: 10 (9 AAS)2nd Position: 6 (6 AAS)3rd Position: 13 (0 AAS)14 Total Variable Amino Acids
200+
100-200
50-100
25-50
10-25
2-10
1ALL
EASTWEST
ASIANWhite Line: Silent MutationRed Line: Amino Acid Shift
Regional Cytochrome b Slatkin’s Linearized FST Midpoint Rooted NJ Phylogram
OKH
KUR
KAM
COM
CAL
CGA
WAL
WGA
BER
EAL
PWS
BRC
ORESEA
NCA
0.05 changes
LegendEastern StockWestern StockAsian Stock
AsianStock
Ogchul Island EAL
Rogue Reef ORESt. George Island NCA
North Danger Rocks BRC
Forrester Island SEATriangle Island BRC
White Sisters Island SEAHazy Island SEA
Chirikof Island CGAAkutan Island EAL
Ugamak Island EALPinnacle Rocks WGAMedny Island COMAdugak Island EALAntsiferova Island KURClubbing Rocks WGAWalrus Island BER
Seal Rocks PWSBuldir Island WAL
Chowiet Island CGAFish Island PWS
Atkins Island WGA
0.01
EasternStock
Yunaska Island CALSugarloaf Island CGA
Seguam Island CALKiska Island CAL
Lovushki Island KURIony Island OKH
Kozlova Cape KAMRaykoke Island KUR
Srednego Island KURYamsky Island OKH
AsianStock
Western Stock
NJ Phylogram of RookeriesBased on Microsatellite Data
LegendEastern StockWestern StockAsian Stock
OKH
KAM
KUR
WAL
CAL
EasternStock
WesternStock
AsianStock
COM
EAL
BER
CGA
WGA
PWS
NCA
ORE
SEA
BRC
0.01
NJ Phylogram of RegionsBased on Microsatellite Data.
LegendEastern StockWestern StockAsian Stock
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.
ConclusionsConclusionsControl Region PhylogeographyControl Region Phylogeography
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.
The Asian StockThe Asian Stock
The recognition of three management units may be appropriate for Steller sea lion conservation.
30°
45°
60°
KAMOKH
BER
PWS
NCA
ORE
SEACOM
WALCAL
EALWGA
CGA
BRC
KUR
RUSSIA ALASKA
Cape Suckling
135°W180°135°E
Sampled Rookeries
JAPAN
144°W
EASTERN STOCK
WESTERN STOCK
ASIAN STOCK
