432 
Fishery Bulletin 109(4) 
Counts were made on the dorsal-fin rays, anal-fin 
rays, pectoral-fin rays, pored lateral line scales, gill 
rakers, and dorsal-fin spines without basal scales. Sig- 
nificant differences in these characters were tested 
with the Mann-Whitney U- test. The presence or ab- 
sence of small scales on the lower jaw was noted and 
the difference in frequencies between species assessed 
by Fisher’s exact test. All statistical analyses were 
conducted using R language, vers. 2.11.1 (R Develop- 
ment Core Team, 2010). Differences were considered 
significant at PcO.Ol. 
Results 
Genetic analysis 
The 12 primer sets yielded 364 AFLP fragments, of 
which 127 (34.9%) were polymorphic. All specimens 
tested displayed unique AFLP fragment patterns, indi- 
cating a high level of genetic variability. A PCoA based 
on the pairwise distance matrix clearly separated the 
specimens into two groups along the first principal 
coordinates (PCo) axis (accounting for 41.83% of the 
total variance) with no overlap (Fig. 3), which corre- 
sponded well with the initial identifications of S. vulpes 
and S. zonatus based on body coloration, except for 
two specimens (FAKU 82515 and FAKU 130236). The 
latter were initially identified as S. vulpes, but geneti- 
cally assigned to S. zonatus. Because S. vulpes and S. 
zonatus were clearly distinguished by the PCol scores 
without any intermediate specimens, we regarded the 
above specimens as S. zonatus. Comparisons below were 
made between the two genetically assigned species (S. 
vulpes : 37 specimens, S. zonatus: 28 specimens). In 
contrast, the PCo2 and PCo3 scores (accounting for 
13.52% and 12.56% of the total variance, respectively) 
did not result in separation of the two species (not 
shown). Despite the high polymorphism evident in the 
AFLP fragments, no diagnostic differences in fragment 
patterns were observed between genetically assigned 
S. vulpes and S. zonatus. Mean (± standard deviation) 
pairwise genetic distances estimated by the algorithm 
of Huff et al. (1993) were 22.63 ±3.95 within S. vulpes, 
28.69 ±5.16 within S. zonatus, and 32.21 ±5.52 between 
them . 
Within the amplified region of mtDNA, continuous se- 
quences of part of the tRNA Thr gene (24 bp), the proline 
transfer RNA (tRNA Pr0 ) gene (70 bp), and part of the 
mtCR (358 bp) were aligned. The sequences contained 
45 variable sites with three indels among 
65 specimens, 29 of which were parsimony 
informative, defining a total of 41 haplo- 
types. Twenty-one haplotypes were found 
in S. vulpes and 22 in S. zonatus, two be- 
ing shared by the two species. No transver- 
sions were observed. The nucleotide compo- 
sition was AT-biased (A=38.4%, C = 19.4%, 
G = 13.6%, T=28.6%), as is common for fish 
mtDNA (McMillan and Palumbi, 1997). 
Pairwise sequence divergences between S. 
vulpes and S. zonatus varied from 0% to 
3.1% (mean 1.5%). The haplotype diversities 
for S. vulpes and S. zonatus were 0.94 ±0.03 
and 0.98 ±0.02, respectively, and nucleotide 
diversities (in %), 1.41 ±0.76 and 1.47 ±0.80, 
respectively. In the MSN inferred from mtD- 
NA sequence variations, 41 haplotypes were 
connected to each other by one to six muta- 
tional steps, revealing a rather expanded 
topology in the network (Fig. 4). Sebastes 
vulpes and S. zonatus were not clearly sepa- 
rated in the network, but restricted gene 
flow between them was indicated by the low 
but significant pairwise <f> sr value at a=0.05 
level (<J> ST = 0.053, P=0.011 ±0.001). 
Morphological analysis 
Measurements of S. vulpes and S. zonatus 
and results of the ANCOVA are shown in 
Table 1. Among measurements meeting the 
statistical assumptions for ANCOVA, four 
characters out of 23 differed significantly 
between the two species. 
Figure 3 
Distribution of first principal coordinate (PCo) scores based on 364 
amplified fragment length polymorphisms fragments for Sebastes 
zonatus and S. vulpes. The amount of variance explained by PCol 
is given in parentheses. The body coloration of each specimen is 
designated by the fill pattern of the bars (grayish specimens, lateral 
stripes; brownish specimen, diagonal stripes). The present specimens 
were separated completely into two genetically distinct groups. 
