Kurita et al.: The effect of temperature on the duration of spawning markers in Paralichthys olivaceus 
85 
Figure 4 
(A) Duration of oocytes at each developmental stage and new postovulatory follicles and 
(B) combined developmental stages in relation to temperature for Japanese flounder 
( Paralichthys olivaceus). MN(E) = oocyte at early migratory nucleus stage; MN(L) = at late 
migratory nucleus stage; HD = at hydrated stage; POF(new)=new postovulatory follicles; 
MN(E)-HD = combined stages of MN(E), MN(L), and HD; MN(L)-HD = combined stages 
of MN(L) and HD. Statistics for regression lines are shown in Table 3. 
G hd = 8.66 e 0 07997 , 
(9.2<T<19.6°C, n= 20, r 2 = 0.937, PcO.OOl). (2) 
Thus, growth rate increased by a factor of 2.2 as tem- 
perature increased by 10°C. Body size and individual 
effects on the relationship between the growth rate of 
hydrated oocytes and temperature were not observed 
(body size effect; multiple regression analysis, P=0.12: 
individual effect; ANCOVA, slope, P= 0.29; intercept, 
P= 0.75). 
Ovulated egg diameters (ED, pm), as a substitution 
for the maximum hydrated oocyte diameter, decreased 
from 980 to 830 pm as temperature increased (Fig. 6) 
as follows: 
ED = -11.8 T + 1099, 
(9.2<T<22.6°C, n=7, r 2 = 0.975, PcO.OOl). (3) 
The largest diameter of oocytes at the MN(L) stage, 
as a substitution for the minimum hydrated oocyte 
diameter, did not decrease significantly with tem- 
perature (9.2<T , <19.6°C, n = 6) and averaged 645 pm. 
Thus, the ranges of hydrated oocyte diameter also 
decreased linearly from 345 to 220 pm as tempera- 
ture increased from 9.2° to 19.7°C (Fig. 6). As a 
result, the duration of the HD stage, i.e., the range 
of hydrated oocyte diameter divided by the growth 
rate of hydrated oocytes, also decreased exponen- 
tially from 19.1 to 5.3 h as temperature increased 
from 9.2° to 19.7°C (Fig. 4, Table 3). Duration of this 
stage decreased by a factor of 0.30 as temperature 
increased by 10°C. 
The duration of combined spawning markers, i.e., 
MN(L) + HD stages and MN(E) + MN(L) + HD stag- 
es, also decreased exponentially from 35.1 to 10.3 h 
(9.2-19.7°C ) and 53.0 to 13.3 h (11.3-19.7°C ), respec- 
tively (Fig. 4, Table 3). 
40 
P 
SZ 
E 
3 
o/ 
CD 
cc 30 
JZ 
% 
o 
Q/cP 
0 
20 
yO 
o, 
10 15 20 25 
Temperature (°C) 
Figure 5 
Relationship between growth rate of 
oocyte diameter during hydration and 
temperature for Japanese flounder 
( Paralichthys olivaceus) . Each data 
point represents one hydration event. 
See Equation 2 in the main text. 
Discussion 
In this study, we clearly show that duration of spawn- 
ing markers, i.e., oocytes at the MN(E), MN(L), and HD 
stages, and the POF(new) stage, largely varied because 
of water temperature. The duration of a marker affects 
estimates of spawning frequency according to Equation 
1 and can have large effects if fixed spawning mark- 
ers are used for a range of ambient temperatures. The 
duration of spawning markers decreased exponentially 
as temperature increased. Thus, estimates of spawning 
frequency per day in colder and warmer waters tend 
to be overestimated and underestimated, respectively, 
if the effect of temperature on marker duration is not 
