GENETIC DIFFERENTIATION BETWEEN TWO FORMS
OF SHORT-FINNED PILOT WHALES OFF
THE PACIFIC COAST OF JAPAN*

SHIRO WADA**ABSTRACTThe northern and southern forms of the short-finned pilot whale off
the Pacific coast of Ja pan were examined by horizontal starch gel electrophoresis on 36 enzyme loci using 371 specimens. Two and five loci werevariable in the northern and southern forms, respectively. The proportionof polymorphic loci under the 0.95 criterion and the value of the average
heterozygosity per locus were 0.028 and 0.009 in the northern form and0.056 and 0.008 in the southern form, respectively. No significant deviationin genotype frequencies from the Hardy-Weinberg expectations was seenin either form. Significant between-form difference in genie proportions
revealed that the northern form is genetically isolated from the southern
form. From the value of Nei's genetic distance, 0.0004, the degree ofgenetic differentiation between two forms was considered to be of the local
population level.
INTRODUCTION

The short-finned pilot whale, Globicephala macrorhynchus Gray, 1846, is one of
the important objective species for the coastal whaling in Japan, which has
been harvested in two different fisheries. Two forms segregate geographicallyand thermally off the Pacific coast of Japan (Kasuya, Miyashita and Kasama~su,
1987). The northern form, tappanaga, having been harvested by the small-typewhaling (Kasuya and Tai, 1986), distributes off the Pacific coast of northern
Japan between 35°N and 42°N in the surface water temperature range of
12-24°C, while the southern form, magondo, having been harvested by thesmall-type whaling and the drive fishery at Taiji (Kasuya, l 986b), distributes
in the waters south of the major range of the northern form and waters over
18°C.
Form-identification is easy from the difference of external characteristics.
The northern form has a larger body size, a distinct saddle mark and a roundcontour of the head, while the southern form has a smaller body size, an* An earlier version of this manuscript was submitted to the Scientific Committee of the International Whaling
Commission, Bournemouth, May 1986 (Document No. SC/38/SM18).

** Far Seas Fisheries Research Laboratory, 5-7-1, Orido, Shimizu-ski, Shizuoka-ken, 424 Japan
Sci. Rep. Whales Res. Inst.,
No. 39, 1988, 91-101

92

WADA

indistinct saddle mark and a square contour of adult male head (Kasuya et al.,1987). However, the life history parameters of the two forms differ significantly
only in those concerning to body size and to breeding season (Kasuya, l 986a).
Both forms share the skull characteristics of G. macrorhynchus described byBree (1971 ). From these, Kasuya (l 986a) suggested that they can be dealt
with as separate geographical forms or subspecies.
Electrophoretic survey on a large number of loci for genetic variability or
genetic differentiation in cetacea have been reported on rather limited numberof species; on minke whale by Wada and Numachi (1979), Simonsen, Kapeland Larsen (1982), and Wada (1982; 1983a; 1984); on fin and sei whales by
Danielsd6ttir and Amason (1987); on fin, sei, Bryde's and minke whales by
Wada ( 1987); on striped dolphin by Wada ( l 983b); on Dall's porpoise by
Winans and Jones (in press); on long-finned pilot whale by Andersen (1987);
on 12 species of toothed. whale by Shimura and Numachi (1987).
The aim of the present study is to evaluate the magnitude of genetic
differentiation between two forms of short-finned pilot whales and to consider
their taxonomic relationship from the viewpoint of biochemical genetics.
MATERIALS AND METHODS

Tissue samples, liver and skeletal muscle, of 204 northern forms werecollected by Dr T. Kasuya and his co-workers as a part of their biologicalsamplings from the whales caught by the small-type whaling off the Sanriku
region and landed at Ayukawa in October and November 1983 ( 105 specimens)
and 1984 (99 specimens).
Tissue samples of 167 southern forms were collected by myself from thewhales driven at Taiji and flensed on 7-8 October 1984 (68 specimens fromfour schools in a single drive of 92 whales) and 11-12 November 1985 (99specimens from a single drive of about 120 whales which might have been
mixed with a few members of a previous drive). Form-identification for the
specimens in 1984 was done by Dr N. Miyazaki and for those in 1985 bymyself. All tissue samples were stored at - 20°C until use.
Methods of sample and gel preparations, and electrophoresis followed
the procedures described in Numachi (1974), and Shimura and Numachi( 1987), and that of gel preservation in N umachi ( 1981 ). Of the 32 enzymes
examined, the following three loci and five enzymes showed an indistinct
electrophoretic pattern, and therefore I didn't use them in the present study
though the last three enzymes were found to be variable; Adh-2, Ak-1,
Pgm-2, Aconitase, Fumarase, Peptidases B, C and D. Table 1 shows buffers
and tissues used for 36 loci on which reliable genotyping was possible. Recipes
for the 100 ml reaction mixture for staining enzymes, prepared according toHarris and Hopkinson (1976) in principle are listed in Table 2.
In case of multi-loci enzymes, a most anodally migrating locus wasnamed as 1, lesser anodal locus as 2, and so on. At a variable locus, alleles
Sci. Rep. Whales Res. Inst.,
No. 39, 1988

GENETICS OF SHORT-FINNED PILOT WHALES

93

were named alphabetically as c, s, n andf from cathode to anode. Then allele
denotes the most frequent allele in a variable locus, or denotes the single one
in a monomorphic locus.
Genotype and allele frequencies were analysed using the G-test (Sokal
and Rohlf, 1969) for a deviation in genotype frequencies from the HardyWeinberg expectations within a form (Gn), and for between-form heterogeneity
in gene frequencies (CH) based on the R (number of alleles) x C (number offorms) contingency table format. G-values were compared with the criticalvalues of Chi-square under (number of genotypes)-(number of alleles)
degrees of freedom for Gn, and under (R-1) x (C-1) degrees of freedom for
CH.

RESULTS

Genetic variability
Genetic variation was found at two loci, Ada and Sod-I, in the northern
form and at five loci, Ada, Mdh-1, 6Pgd, Pgm-3 and Sod-I, in the southern
form. These loci were all biallelic. The northern form had the same alleles as
the southern form at all loci except Ada, where a variant allele in the former(Adas gene) was different from that in the latter (Adac gene). Heterozygotes at
Ada and Pgm-3 showed a two-banded pattern and those at Mdh-1, 6Pgd andSod-I showed a three-banded pattern, suggesting that the enzymes controlled
by the former two and the latter three loci are monomer and dimer, respectively (Fig. 1). The remaining 31 loci were fixed for a single allele in bothforms, so that the subunit structure of enzymes controlled by these loci was
unknown. These results on the southern form agreed substantially with thosein Shimura and Numachi (1987) except a considerable gene frequency
difference at Ldh-A (their Ldh-1) and Pgm-2, probably due to difference ofsample size between the two studies.
The observed genotype frequencies at five variable loci agreed well with
the Hardy-Weinberg expectations. Although the expectations for rare
genotypes were less than five, the Gn values for 7 cases, ranged from 0.01 to
0. 97, were all less than the critical values of Chi-square at the 5% level ofsignificance (Table 3). This suggests that each sample is homogeneous.
Number of polymorphic loci under the 0.95 criterion (P.95: loci on which
the most frequent allele is less than or equal to 0.95) was very small, only one(Ada) in the northern form and two (Ada and Sod-I) in the southern form,
though Sod-I of the former was nearly polymorphic. The P.95 values were
therefore 0.028 and 0.056 in the northern and southern forms, respectively.
The level of average heterozygosity per locus (H) was also low due to low
frequencies of variant genes. The estimates of H value calculated in thenorthern and southern forms were 0.009 ± 0.007 (SE) and 0.008 ± 0.005(SE), respectively. Standard error (SE) of H was calculated using a formula of
Nei and Roychoudhury (1974).
Sci. Rep. Whales Res. Inst.,
No. 39, 1988

94

WADA
TABLE I. LOCI EXAMINED AND ELECTROPHRETIC CONDITIONSFOR THE SHORT-FINNED PILOT WHALES
Enzyme (abbreviation)

Adenosine deaminase (ADA)
Alcohol dehydrogenase (ADH)
Adenylate kinase (AK)
Aldolase (ALD)
Creatine kinase (CK)
Glyceraldehyde-3-phosphate
dehydrogenase (GAPDH)
a-Glycerophosphate dehydrogenase (GDH)
Glutamate dehydrogenase (GLUDH)
Glutamate oxaloacetate transaminase (GOT)
Glucosephophate isomerase (GPI)
Hexokinase (HK)Isocitrate dehydrogenase (IDH)
Lactate dehydrogenase (LDH)
Malate dehydrogenase (MDH)
Malic enzyme (ME)
Mannosephosphate isomerase (MPI)
NADP-dependent dehydrogenase (NDH)
Peptidase-A (PEPA)
Peptidase-E (PEPE)
Peptidase-S (PEPS)
6-Phophogluconate dehydrogenase (6PGD)
Phophoglucomutase (PGM)
Pyruvate kinase (PK)Sorbitol dehydrogenase (SDH)Superoxide dismutase (SOD)

Xantine oxidase (XOD)
Xylulose reductase (XR)

Locus

Tissueusedbuffer

Ada
Adh-1
Ak-2
Aid
Ck-1
Ck-2
Gapdh

Liver
Liver
MuscleMuscleLiver
MuscleLiver

AC70
AC70, Mg, NAD
AC70
AC70, Mg
TEB87
TC86
TC86, EDTA, NAD

Gdh
Gludh
Got-I
Got-2
Gpi
Hk
Idh-1
Idh-2
Ldh-A
Ldh-B
Mdh-1
Mdh-2
Me
Mpi
Ndh
Pep A
PepE
Peps
6Pgd
Pgm-1
Pgm-3
Pk
Sdh
Sod-1
Sod-2
Sod-3
Sod-4
Xod
Xr

Liver
Liver
Liver
Liver
Liver
Liver
Liver
Liver
Liver
Liver
Liver
Liver
Liver
Liver
Liver
Liver
Muscle
Liver
Liver
Liver
Liver
Muscle
Liver
Liver
Liver
Liver
MuscleLiver
Liver

AC70, Mg, NAD
TEB87
TC80
AC70
TCSO
TC86
AC70, Mg
AC70
AC60
AC60
AC60
AC60
AC60
AC70
AC70, NADP
AC60
AC60
TCSO, Mg
AC70, NADP
AC70
TC80
AC60, Mg
AC70, Mg, NAD
AC60
AC60
AC60
AC60
TEB87
AC70, Mg, NADP

AC: amine-citrate buffers originally used in Clayton annd Tretiak (1972), slightly modified by Numachi,
Nagahora and Iwata (1979).
Gel: 0.002 M citric acid. Electrode: 0.04 M citric acid.
AC60: adjusted with N-(3-Aminopropyl)-morpholine to pH 6.0.
AC70: adjusted with N-(3-Aminopropyl)-diethanolamine to pH 7.0.
TC: tris-citrate buffers.
TC80: originally used in Clayton and Tretiak (1972). Gel: 0.002 M citric acid. Electrode: 0.04 M citricacid. Both solutions were adjusted with Tris (hydroxymethyl) aminomethane to pH 8.0.
TC86: described in Harris and Hopkinson (1976). Electrode: 0.083 M citric acid adjusted with 0.661 M
Tris (hydroxymethyl) amino-methane to pH 8.6. Gel: 27 .5 times dilution of electrode buffer.

Sci. Rep. Whales Res. Inst.,
No. 39, 1988

GENETICS OF SHORT-FINNED PILOT WHALES

®

..
®

Mdh-1

--

sn

6Pgd

®

..

:

- -- SS

sn

nf

nn

8

Ada

en

Pgm-3

-- - r~:
nn

nn

95

nn

EB

Sod-1

-SS

sn

nn

..
Fig. I. Electrophoretic patterns at five variable loci in the short-finned pilot whales
off the Pacific coast of Japan. The genotype of each phenotype is shown in italic.
~: sample origin, (±): anode, 8: cathode

Genetic differentiation
A significant difference in gene frequencies between the two forms wasfound in the following three loci, Ada; CH = 48.03, d.f. = 2, P<0.001,
Mdh-1; CH = 10.55, d.f. = 1, P<0.01, 6Pgd; CH = 4.80, d.f. = 1, P<0.05,
but the differences at Pgm-3 and Sod-1 were not significant. An overall Gttvalue for the five variable loci was 66.96 (d.f. = 6, P<0.001), which indicatesthat the two forms are genetically isolated stocks.
The extent of genetic differntiation between two forms was measured by
Nei's (1972) genetic distance (D), which represents the average number ofelectrophoretically detectable allelic substitutions per locus. It ranges from 0(all populations share the same alleles in the same frequencies at all loci) to
infinite (each population is fixed for an unique allele at all loci). The estimate
of D value calculated using 36 loci was 0.0004.

(Footnote continued)
TEB:

described in Harris and Hopkinson (1976). Stock: 0.9 M Tris (hydroxymethyl) aminomethane0.02 M ethylenediamine tetraacetic acid disodium salt (EDTA-2Na)-0.5 M boric acid, pH 8.7.
Anodal electrode: 7 times dilution of stock; cathodal electrode: 5 times dilution of stock. Gel: 20
times dilution of stock.
EDTA: gel contains 0.242 mM or 9 mg/100 ml EDTA-2Na.
gel contains 10 mM magnesium chloride.
Mg:
NAD: gel and electrode buffers contain 4 mg/200 ml nicotinamide adenine dinucleotide. For GAPDH
only, 15 mg/200 ml was used.
NADP: gel and electrode buffers contain 4 mg/200 ml nicotinamide adenine dinucleotide phosphate.

Sci. Rep. Whales Res. Inst.,
No. 39, 1988

96

WADA
TABLE 2. STAIN RECIPIES FOR THE IOOml REACTION MIXTURE

Enzyme 6>

Substrate(s) I)

ADA

40mg Adenosine

ADH!ml 1-octanol
9ml 95% ethanol
500mg D-glucose
IOOmg ADP
200mg Fructose-1,6di phosphate
I OOmg Creatine
phosphate
500mg D-glucose
IOOmgADP
400mg Fructose-1,6diphosphate
2g a-glycerophosphate

AK
ALD
CK

GAPDH
GDH
GLUDH

2g L-glutamate

GOT

75mg 2-oxoglutaric acid
225mg L-aspartic acidGP!

IOOmg Fructose-6phosphate
2g D-glucose
200mgATP200mg DL-isocitrate

HK
IDH
LDH
MDH
ME
MPI
NDH
PEPA
PEPE
PEPS
6PGD
PGM
PK

SDH
SOD
XOD
XR

Buffer
0.05M Phosphate,
pH7.4
O.IM Tris-HCl,
pH8.7
O.IM Tris-HCl,
pH8.0
O.IMTris-HCl,
pH8.0
O.IMTris-HCl,
pH8.0
O. IM Tris-HGl,
pH8.0
O.IM Tris-HCl,
pH8.7
O.IM Tris-HCl,
pH8.0
adjusted to pH7.5
with2MKOH

Coenzyme andcoupling dye 2)

Iron 3)

NBT,PMS

Linking
enzyme4)

Other
components 5)

luXOD
Ju NP

2g Agar

IOOu HK
60u G6PD
IOOuGAPDH

2g Agar

NAD*, NBT, PMS
NADP, NBT, PMS+

NAD, NBT, PMS

500mg
Na arsenate

NADP, NBT, PMS

NAD*, NBT, PMS+

lOOu HK
60u G6PD

2g Agar

25u ALD

500mg
Na arsenate

NAD, NBT, PMS

lgEDTA-2Na

NAD, NBT, PMS
500mg Fast blue BB

O.IM Tris-HCl,
NADP, NBT, PMS
60u G6PD
pH7.4
O.IMTris-HCl,
NADP, NBT, PMS+ 60u G6PD
pH8.0
O.IM Tris-HCl,
NADP, NBT, PMS+
pH8.0
O.IM Tris-HCl,
NAD, NBT, PMS
2ml 50% DL-lactate
pH8.7
NAD, NBT, PMS
2g DL-malate
O.IM Tris-HCl,
pH8.7
2g DL-malate
O.IM Tris-HCl,
NADP, NBT, PMS+
pH8.0
IOOmg D-mannose-6O.IM Tris-HCl,
NADP, NBT, PMS
60u G6PD
phosphate
pH7.4
50u GP!
O.IMTris-HCl,
NADP, NBT, PMS+
pH8.0
I OOmg Glycyl-DL0.05M Phosphate,
75mg Dianisidine-HCl
IOuAAO+IOOu POD
pH7.4
glycine
IOOmg L-leucyl-~-naphtyl-amide as substrate in the same buffer and reagent mixture as PEPA
IOOmg L-leucyl-1-leucine as substrate in the same buffer and reagent mixture as PEPA
IOOmg 6-phosphoO.IMTris-HCl,
NADP, NBT, PMS+
gluconate
pH8.0
200mg D-glucose-1O.IM Tris-HCl,
NADP, NBT, PMS+ 60u G6PD
phosphate*
pH8.0
O.IM Tris-HCI,
NADP, NBT, PMS
I OOmg Phosphoenol++ IOOu HK
pH8.0
pyruvate
60u G6PD
500mg D-glucose
IOOmg ADP
O.IM Tris-HCl,
NAD, NBT, PMS
3g D-sorbitol
pH8.0
NBT,PMS
O. IM Tris-HCl,
pH8.7
O.IMTris-HCI,
NBT,PMS
250mg Hypoxanthine
pH8.0
3g D-xylitol
O.IM Tris-HCl,
NADP, NBT, PMS+
pH8.0

IOOmg EDTA-2Na
3g Na phosphatemonobasic
2g Agar
2g Agar

2g Agar

2g Agar

500mg EDTA-2NaI) ADP: Adenosine-5'-diphosphate; ATP: Adenosine-5'-triphosphate; *:containing D-glucose-1,6-diphosphate. Ca. 1%.
2) NAD: 30mg Nicotinamide adenine dinucleotide; NAO*: 90mg; NADP: Nicotinamide adenine dinucleotide phosphate;
NBT: 20mg Nitroblue tetrazolium; PMS: 5mg Phenazine methosulfate.
3) +: IOmM MgCl2; ++: !OmM MgCh, 150mg KC!.
4) NP: Nucleoside phospholylase; G6PD: Glucose-6-phosphate dehydrogenase; AAO: Amino acid oxidase; POD: Peroxidase.
5) EDTA: Ethylenediaminetetraacetic acid.
6) see Table I

Sci. Rep. Whales Res. Inst.,
No. 39, 1988

97

GENETICS OF SHORT-FINNED PILOT WHALESTABLE 3. GENOTYPE AND ALLELE FREQUENCIES FOR FIVE VARIABLE LOCIIN THE NORTHERN AND SOUTHERN FORMS OF THE SHORT-FINNEDPILOT WHALE OFF THE PACIFIC COAST OF JAPAN
Locus

Form

Ada

N

s

en
0
13

sn
17
0

nn
51
86

Total
69
99

0
0

sn
0
8

nn
154
159

nn
204
164

nf
0
3

SS

SS

0
SS

Mdh-1

N

s
6Pgd

N

s
Pgm-3

N

s

0
0
SS

Sod-I

Allelefrequencies

Genotypes

N

s

0

Gn
value

.138
.OOO

n
.862
.934

0.64
0.91

Total
154
167

.OOO
.024

n
I.OOO
.976

0.20

ff
0
0

Total
204
167

n
I.OOO
.991

.OOO
.009

0.03

sn
0
2

nn
204
165

Total
204
167

.OOO
.006

n
I.OOO
.994

0.01

sn
18
16

nn
186
150

Total
204
167

.044
.054

n
.956
.946

0.83
0.97

.OOO
.066

f

S: southern form. N: northern form. Gn: G-value for deviation in genotype frequencies from the HardyWeinberg expectations within a form.

DISCUSSION

Reliable estimates of average heterozygosity for marine mammals have beenreported from nine species of pinniped, four species of baleen whale and 12
species of toothed whale. Table 4 summarises the level of their genetic
variabilities. Mean H value for pinnipeds is 0.019, ranging from 0 in northernelephant seal, Mirounga angustirostris (Bonnell and Selander, 1974) to 0.047 in
ribbon seal, Phoca fasciata (Fujio and Saito, 1986). Wada (1987) calculated H
values for three stocks of fin whale, Balaenoptera physalus, two stocks of sei
whale, B. borealis, five stocks of Bryde's whale, B. edeni, and three stocks ofminke whale, B. acutorostrata, which allowed to calculate the mean values of
H for fin, sei, Bryde's and minke whales as 0.017, 0.021, 0.022 and 0.051,
respectively. Simonsen et al. (1982) reported H = 0.046 for the Atlantic minke
whales. Using these values, the mean H values for four stocks of minke whales
and four baleen whale species have been calculated as 0.050 and 0.028,
respectively.
On toothed whales, Wada (l 983b) reported the H value in striped dolphin,
Stenella coeruleoalba, off the Pacific coast of Japan as 0.021. Shimura andNumachi (1987) calculated H values on 12 species in 11 genera from 4 families,
which ranged from 0 in finless porpoise, Neophocaena phocaenoides, to 0.154 in
Dall's porpoise, Phocoenoides dalli (dalli-type). Winans and Jones (in press) also
Sci. Rep. Whales Res. Inst.,
No. 39, 1988

98

WADA
TABLE 4. SUMMARY OF THE GENETIC VARIABILITY IN MARINE MAMMALS
No. loci
examined

P.95

Seals
Mirounga angustirostrisM.
leonina
Odobenus rosmarus rosmarus
Pagophilus groenlandicus
Cystophora cristata
Pusa hispida
Phoea f asciata
P.
viturina stejnegeriP.
largha

24
18
32
21
21
21
34
34
34

0.000
0.278
0.094*
0.048*
0.048*
0.095*
0.147
0.059
0.059

0.000 Bonnell and Selander (1974)
0.030 McDermid, Ananthakrishnan and Agar (1972)
0.026 Simonsen, Born and Kristensen (1982)
0.007 Simonsen, Allendorf, Eanes and Kapel (1982)
0.009 Simonsen, et al. (1982)
0.009 Simonsen, et al. (1982)
0.047 Fujio and Saito (1986)
0.023 Fujio and Saito (1986)
0.017 Fujio and Saito ( 1986)

Baleen whales
Balaenoptera physalus 1l
B.
borealis 2l
edeni 3lB.
acutorostrata 1l
B.
acutorostrata
B.

45
45
45
45
21

0.138*
0.158*
0.114*
0.207*
0.095*

0.017 Wada(l987)
0.021 Wada (1987)
0.022 Wada (1987)
0.051 Wada (1987)
0. 046 Simonsen, et al. (1982)

Toothed whales
Berardius bairdii
Globicephala macrorhynchus (N. F.)
G.
macrorhynchus ( S.F.)
macrorhynchus ( S.F.)
G.
Peponocephala electra
Pseudorca crassidens
Stenella coeruleoalba
S.
coeruleoalbas.
attenuataTursiops truncatus
Lagenorhynchus obliquidens
Steno bredanensis
Phocoena phocoena
Phocoenoides dalli (dalli-type)dalli (dalli-type)P.
P.
dalli (truei-type)
Neophocaena phocaenoides

18
36
36
19
19
19
15
19
19
19
19
19
18
26
19
19
18

0.056
0.028
0.056
0.263
0.105
0.211
0.130
0.263
0.263
0.105
0.316
0.053
0.167
0.231
0.421
0.474
0.000

0.016 Shimura and Numachi (1987)
0.009 Present study
0.008 Present study
0.054 Shimura and Numachi (1987)
0.035 Shimura and Numachi (1987)
0.051 Shimura and Numachi (1987)
0.021 Wada (1983b)
0.089 Shimura and Numachi (1987)
0.089 Shimura and Numachi (1987)
0.039 Shimura and Numachi (1987)
0.093 Shimura and Numachi (1987)
0.007 Shimura and Numachi (1987)
0.047 Shimura and Numachi (1987)
0.058 Winans and Jones (in press)
0.154 Shimura and Numachi (1987)
0.147 Shimura and Numachi (1987)
0.000 Shimura and Numachi (1987)

SpeciesH

Reference* under the 0.99 criterion
1) mean value for three stocks. 2) mean value for two stocks. 3) mean value for five stocks.
N.F. = the northern form. S.F. = the southern form.

examined other specimens of the latter species from three areas in the North
Pacific and Bering Sea, and calculated H value as 0.058. However, there arelarge differences in the reported values of P.95 and H for S. coeruleoalba, P.
dalli (dalli-type) and G. macrorhynchus between authors (Table 4). However,comparing the common loci only, these studies show very similar results apartfrom a few loci where sample size is small. The mean of the above 17 H values
for 13 toothed whale species (Table 4) is 0.054, which is higher than that for
Sci. Rep. Whales Res. Inst.,
No. 39, 1988

GENETICS OF SHORT-FINNED PILOT WHALES

99

pinnipeds (H0.019), baleen whales (H = 0.028) and 184 mammalian
species (H = 0.041; Nevo, Beiles and Ben-Shlomo, 1984).
Selander and Kaufman (1973) suggested that large, mobile animals
generally have lower levels of genetic variability than smaller, less mobile
ones. However, both forms of the short-finned pilot whale showed almost thelowest levels of genetic variability among cetacean species, which were lower
than Baird's beaked (Berardius bairdii), Bryde's, sei and fin whales that have alarger body size thereby a lower level of genetic variability than the pilot
whales is predicted.
Nei (1975) showed the D values to correlate with the systematic distance,i.e., 0-0.058 of interpopulation 'level, 0.004-0.351 (mainly 0.1-0.2) of intersubspecies level and 0.05-2.73 (mainly 0.5-2.0) of interspecies level. However,Avise and Aquadro ( 1982) showed that the degree of genetic differentiationdiffer considerably from species to species, and that the mean D value between
species was 1.75 in amphibia, 0.67 in reptilia, 0.41 in mammalia and 0.08
in Aves.
In cetacea, however, much smaller D values have been reported between
populations. Among five stocks_of Bryde's whale in the Indian and Pacific
Oceans, Wada (1987) obtained D = 0.0029, ranging from 0.0003 to 0.0050.
The D value between two color types of the Dall's porpoise was 0.004 (Shimura
and Numachi, 1987). Andersen (1987) calculated the genetic distance amongnine schools of the long-finned pilot whale, Globicephala melaena, around theFaroe Islands. The largest D value estimated from her gene frequency data
is 0.0032. The D value of 0.0004 calculated in the present study was in thelower range of above values.
Nei and Roychoudhury (1974) and Nei (1978) studied on the sampling
variance of D, and decomposed a total variance into the intralocus andinterlocus variances. The former is due to a restricted number of genes
sampled for each locus, and is almost negligible when the sample size is morethan 50. The latter is due to a restricted number of loci sampled from thegenome. Therefore, the reliability of the estimate of D mainly depends on thenumber of loci or chance of sampling particular loci.
In cetacea examined to date, the most frequent allele at each locus wascommon in all populations within a species except in minke whales. This
yielded a small value of a single-locus distance; the largest ones between
populations were 0.0034 at Me in fin whale, 0.0150 at Ada in short-finned
pilot whale, 0.0211 at Pgm-3 in Dall's porpoise, 0.1164 at Sdh in sei whale and0.1316 at Pgm-1 in Bryde's whale. If a hypothetical Dis calculated between
the two forms of the short-finned pilot whales off Japan assuming an additional
37th locus with a single-locus distance value of 0.15, tenfold of the actuallyobserved value, the estimate of D over all loci increases from 0.0004 to
0.0040. Even under this extreme assumption, the D value is still smaller than
the smallest ones obtained from species comparison; D = 0.026 between
striped dolphin, Stenella coeruleoalba and spotted dolphin, S. attenuata, and D
Sci. Rep. Whales Res. Inst.,
No. 39, 1988

100

WADA

= 0.04 7 between sei and Bryde's whales. From this, I conclude that the extent
of genetic differentiation between the two forms of the short-finned pilot
whale is nothing more than the level of interpopulation. It is quite reasonable
to say that their current taxonomic relationship is the two local forms withina species. This conclusion coincides with the results of the comparative studies
on life history parameters (Kasuya, l 986a) and on morphology and segregation(Kasuya et al., 1987) between the two forms.
ACKNOWLEDGEMENTS

I wish to thank the following three persons: Dr T. Kasuya, who identified thewhales at Ayukawa as northern form, collected tissue samples from them, andread the manuscript critically. Dr N. Miyazaki, who identified the whales at
Taiji in 1984 as southern form and helped me in collecting tissue samples
from them. Miss T. Kohno, who helped me in eletrophoresis.
REFERENCES
ANDERSEN, L. W. 1987. An investigation of the population structure of the long-finned pilot whales,
Globicephala melaena, around the Faroe Islands on basis of isozymes. IWC/SC/39/SM13. 14pp.
Av1sE,]. C. and C. F. AQUADRO, 1982. A comparative summary of genetic distances in the vertebrates. Evol.
Biol., 15: 151-185.
BONNELL, M.L. and R.K. SELANDER, 1974. Elephant seals: genetic variation and near extinction. Science,
184: 908-909.
BREE, P.J.H. VAN, 1971. On Globicephala sieboldii Gray, 1946, and other species of pilot whales. Beaufortia,
19(247): 79-87.
CLAYTON,]. W. and D. N. TRETIAK, 1972. Amine-citrate buffers for pH controls in starch gel electrophoresis.
]. Fish. Res. Bd. Can., 29: 1169-1172.
DANIELSDOTTIR, A. K. and A. ARNASON, 1987. Progress report on the study of genetic variation at enzymeloci in fin whales (Balaenoptera physalus) and sei whales (Balaenoptera borealis). IWC/SC/39/Ba8. 15 pp.
FUJIO, Y. and Y. SAITO, 1986. Population genetics in harbor seal, Phoca vitulina, spotted seal, Phoca largha andribbon seal, Phocafasciata. pp. 195-213. Jn: K. WADA, T. lTo, A. NnzvMA, S. HAYAMA, and M. SUZUKI,(eds.) Ecology and Conservation for Harbor Seals. Tokai Univ. Press, Tokyo. 418 pp. (in Japanese)
HARRIS, H. and D.A. HOPKINSON, 1976. Handbook of Enzyme Electrophoresis in Human Genetics. North-Holland,
Amsterdam.
KASUYA, T. 1986a. Comparison of life history parameters between two stock of short-finned pilot whales
off the Pacific coast of Japan. IWC/SC/38/SMIO. 32 pp.
KAsuYA, T. 1986b. Japanese fisheries exploiting southern form short-finned pilot whales. IWC/SC/38/SM19.
12 PP·KASUYA, T., T. MIYASHITA and F. KASAMATSU, 1988. Segregation of two forms of short-finned pilot whales
off the Pacific coast of Japan. Sci. Rep. Whales Res. Inst., 39: 77-90.
KASUYA, T. and S. TAI, 1986. Operation of Japanese small-type whaling on the short-finned pilot whale off
the northeastern Japan. IWC/SC/38/SM16. 16 pp.
McDERMID, E.M., R. ANANTHAKRISHNAN and N.S. AGAR, 1972. Electrophoretic investigation of plasma andred cell proteins and enzymes of Macquarie Island elephant seals. Anim. Blood Grps. Biochem. Genet.,
3: 85-94.
NEI, M. 1972. Genetic distance between populations. Amer. Natur., 106: 282-292.
NEI, M. 1975. Molecular Population Genetics and Evolution. North-Holland, Amsterdam, 288 pp.

Sci. Rep. Whales Res. Inst.,
No. 39, 1988

GENETICS OF SHORT-FINNED PILOT WHALES

101

NE!, M. 1978. Estimation of average heterozygosity and genetic distance from a small number of individuals.
Genetics, 89: 583-590.
NEI, M. and A. K. ROYCHOUDHURY, 1974. Sampling variances of heterozygosity and genetic distance. Genetics,
76: 379-390.
NEvo, E., A. BEILES and R. BEN-SHLOMO, 1984. The evolutionary significance of genetic diversity:
ecological, demographic and life history correlates. pp. 13-213. In: G.S. MANI (ed.) Lecture Notes in
Biomathematics, 53. Springer-Verlag, Berlin. 312 pp.
NuMACHI, K. 1974. Genetic characteristics of fish population. pp. 5-36. In: M. N1smwAKI (ed.) Shigenseibutsuron. Tokyo University Press, Tokyo, 214 pp. (in Japanese).
NuMACHI, K. 1981. A simple method for preservation and scanning of starch gel. Biochem. Genet., 19: 233-236.
NuMACHI, K., S. NAGAHORA and M. lwATA, 1979. Genetic demonstration of hybrids between chum and pinksalmon in the northwest Pacific. Otsuchi Mar. Res. Cent. Rep., 5: 87-102.
SELANDER, R. K. and D. W. KAUFMAN, 1973. Genetic variability and strategies of adaptation in animals. Proc.
Nat. Acad. Sci. USA, 70: 1875-1877.
SIMONSEN, v., F.W. ALLENDORF, W.F. EANES and F.O. KAPEL, 1982. Electrophoretic variation in large
mammals. III. The ringed seal, Pusa hispida, the harp seal, Pagophilus groenlandicus, and the hooded
seal, Cystophora cristata. Hereditas, 97: 87-90.
SIMONSEN, V., E.W. BORN and T. KRISTENSEN, 1982. Electrophoretic variation in large mammals. IV. The
Atlantic walrus, Odobenus rosmarus rosmarus (L.). Hereditas, 97: 91-94.
SIMONSEN, V., F. KAPEL and F. LARSEN, 1982. Electrophoretic variation in the minke whale, Balaenoptera
acutorostrata Lacepede. Rep. int. Whal. Commn, 32: 275-278.
SHIMURA, E. and K. NuMACHI, 1987. Genetic variability and differentiation in the toothed whales. Sci. Rep.
Whales Res. Inst., 38: 141-163.
SOKAL, R.R. and F.J. ROHLF, 1969. Biometry. W. H. FREEMAN and Co., San Francisco. 859 pp.
WADA, S. l 983a. Genetic structure and taxonomic status of minke whales in the coastal waters of Ja pan.
Rep. int. Whal. Commn, 33: 361-363.
WADA, S. l 983b. Genetic heterozygosity in the striped dolphin off Japan. Rep. int. Whal. Commn, 33:
617-619.
WADA, S. 1987. biochemical analyses of genetic differentiation among the populations and species in
Balaenoptera. Doctoral thesis submitted to Faculty of Agriculture, University of Tokyo, 145 pp. (in Japanese).
WADA, S. and K. NuMACHI, 1979. External and biochemical characters as an approach to stock identification
for the Antarctic minke whale. Rep. int. Whal. Commn, 29: 421-432.
WINANS, G.A. and L. JONES (in press). Electrophoretic variability in the Dall's porpoise (Phocoenoides dalli)in the North Pacific Ocean and Bering Sea.]. Mamm.

Sci. Rep. Whales Res. Inst.,
No. 39, 1988