82 
Fishery Bulletin 117(1-2) 
Figure 4 
Image with a ventral view of the rotational behavior of the clasp- 
er of the male whale shark (Rhincodon typus) in captivity at the 
Okinawa Churaumi Aquarium, Japan, taken in June 2012. 
and inflected dorsally while they were crossed. The 
clasper that was placed on the top of the other varied 
among events, and both claspers had the top position 
interchangeably. In addition, starting in August 2012, 
a white milky fluid was released from the claspers 
when this behavior was displayed (Suppl. Video) (online 
only). However, it was not possible to confirm whether 
this fluid was semen because it dispersed and became 
diffused in the tank immediately after release. 
Changes in hormonal levels 
Changes in plasma concentrations of T and P4 were re¬ 
corded among the 3 observable developmental periods 
of clasper development (Fig. 5). There were minimal 
changes in the concentrations of T and P4 during a 
short period (from May 2008 to July 2011), with mean 
T concentrations of 6.23 ng/mL (SE 0.52) and mean P4 
concentrations of 0.50 ng/mL (SE 0.08) (Suppl. Table) 
(online only). However, T concentrations had large fluctu¬ 
ations during the elongating period (from August 2011 
to July 2012), ranging between 31.95 and 8.42 ng/mL 
with a mean of 18.66 ng/mL (SE 3.06). In the elongat¬ 
ing period, no change was notable in the levels of P4, 
with a mean of 0.79 ng/mL (SE 0.16) (Suppl. Table) 
(online only). In the completed period (from August 2012 
to December 2013), T and P4 levels were higher, with 
means of 36.74 ng/mL (SE 9.46) and 1.21 ng/mL (SE 
0.25), respectively, than those in the other 2 periods. 
Among the 3 periods of clasper development, there 
was significant variation in levels of T (Kruskal-Wallis 
test: 29.40, P<0.01) and P4 (Kruskal-Wallis test: 7.30, 
P-0.03) (Fig. 6). Notably, T concentrations were signifi¬ 
cantly lower in the short period than in the elongat¬ 
ing and completed periods (Steel-Dwass test: 
P<0.01 for both periods). However, there were 
no significant differences in T levels between 
the elongating and completed periods (Steel- 
Dwass test: P-0.95). Concentrations of P4 
were also lower in the short period than in the 
completed period (Steel-Dwass test: P=0.03), 
although there were no significant differences 
in P4 levels between the short and elongating 
periods (P=0.23) or between the elongating and 
completed periods (P-0.82). 
There were significant relationships be¬ 
tween water temperature and concentrations 
of T (coefficient of determination [r 2 ]=0.42, 
P<0.01) and P4 (r 2 =0.26, P<0.01) during the 
short period; whereas, these relationships 
were not significant during the elongating (T: 
r 2 =0.06, P=0.59; P4: r 2 =0.14, P=0.40) and com¬ 
pleted (T: r 2 =0.14, P-0.17; P4: r 2 =0.01, P-0.78) 
periods (Fig. 7). 
Discussion 
Abrupt clasper elongation and increased levels 
of sex hormones observed in the male whale 
shark strongly indicate that the individual is sexually 
mature. Increases in hormone levels (T or P4) associ¬ 
ated with male sexual maturation have been reported in 
various elasmobranchs (Rasmussen and Gruber, 1993), 
although the mode of action of these hormones remains 
unclear. Previous studies have hypothesized several 
functions of sex hormones (such as T), including the 
initiation of mating behavior (Rasmussen and Murru, 
1992; Rasmussen and Gruber, 1993), spermiogenesis 
and spermiation (Tricas et al., 2000; Henningsen et ah, 
2008), and clasper elongation (Heupel et al., 1999; Su- 
likowski et al., 2005; Sulikowski et al., 2006; Awruch et 
al., 2008). Hormone levels were slightly correlated with 
changes in water temperature when the shark was im¬ 
mature; however, these correlations were not observed 
during and after the clasper elongation period, and 
hormone levels were also significantly greater after the 
shark started to mature sexually. Such patterns have 
not been previously reported in other elasmobranch spe¬ 
cies. Further studies are needed to clarify the relation¬ 
ships between hormone levels and water temperature 
during each growth stage. 
Our data allow comparisons between wild and cap¬ 
tive whale sharks. The individual studied here was 
considered mature at 8.5 m TL. This finding is consis¬ 
tent with the maturation size estimated for wild male 
whale sharks off the northwestern coast of Australia 
on the Ningaloo Reef, where 50% of 275 wild male 
whale sharks had attained maturity at 8.1 m TL and 
95% of those sharks had done so by 9.1 m TL (Norman 
and Stevens, 2007). We were not able to determine the 
precise age of maturation in our shark because the 
shark’s age was unknown when he was brought into 
the aquarium. 
