280
E. A. G. Vidal and S. von Boletzky
normal or late; see below) or on their subsequent survival and performance to attest
that infestation by the worms is indeed beneficial during embryonic development.
In any case, infestation can be prevented or greatly reduced by removing the sand
from the apical tip of the egg strands. Thus, the relationship between the squid eggs
and these worms remains not fully explained, as well as other ecological relationships during the egg phase. It has been shown that the egg capsules of D. opalescens
and D. pealeii hold a complex bacterial community via the accessory nidamental
glands, recognized as the host organ and inocula for bacterial species (Barbieri et al.
2001). The precise function of the egg capsule-associated bacteria is unknown, but
it was suggested by Biggs and Epel (1991) that the defence of the eggs against microscopic and macroscopic predators is a possible explanation, as demonstrated by
the antifungal effects of D. pealeii-associated Pseudoalteromonas strains (Barbieri
et al. 1997).
16.5.2 Incubation Conditions and Influence of Environmental
Factors during Embryogenesis
Both natural and artificial seawater have been used to incubate L. vulgaris and D.
opalescens eggs, although developmental problems have been observed mainly in
artificial seawater. The perivitelline fluid inside the egg chorion is in osmotic equilibrium with seawater and will reflect changes in the concentration of major ions of
seawater; therefore, these ions must be within the required concentration for normal
egg development (i.e. calcium and potassium 9–15 mM, magnesium 48–68 mM
and sulphate 15–37 mM) (D’Aniello et al. 1987). Even though late-stage eggs developed normally when incubated in artificial seawater containing these major ions
in proper concentrations, early stage eggs did not, suggesting that other elements
are required. This was later addressed by Hanlon et al. (1989), who demonstrated
that the embryo statoliths do not develop normally when strontium levels are below
8 mg L
−1
. Paralarvae hatching without or with abnormal statoliths were termed
‘spinners’, as they displayed erratic swimming patterns making continuous loops
at the surface and being unable to descend in the water column. The importance of
strontium in the biomineralization process during embryogenesis remains unknown
and other factors must play a role in the formation of statoliths, since the spinners
are occasionally found from natural seawater cultures. Natural seawater is therefore
recommended for egg incubation.
Eggs of both species have been incubated in closed and open, flow-through culture systems (Tables 16.1 and 16.2) and exposed to both constant and fluctuating
water temperatures and salinity conditions. Temperature strongly influences the rate
of embryonic development, incubation time and hatching rate (Boletzky 1987) and
there is some evidence that it might also have an effect on the hatching rhythm
(Paulij et al. 1990). Sen (2005a) evaluated thermal tolerance limits of L. vulgaris intermediate stage eggs (11–13) from Izmir Bay, Turkey. When eggs were exposed to
temperatures from 6 to 28 °C (at 37 psu), hatching rates (defined as number of viable
E. A. G. Vidal and S. von Boletzky
normal or late; see below) or on their subsequent survival and performance to attest
that infestation by the worms is indeed beneficial during embryonic development.
In any case, infestation can be prevented or greatly reduced by removing the sand
from the apical tip of the egg strands. Thus, the relationship between the squid eggs
and these worms remains not fully explained, as well as other ecological relationships during the egg phase. It has been shown that the egg capsules of D. opalescens
and D. pealeii hold a complex bacterial community via the accessory nidamental
glands, recognized as the host organ and inocula for bacterial species (Barbieri et al.
2001). The precise function of the egg capsule-associated bacteria is unknown, but
it was suggested by Biggs and Epel (1991) that the defence of the eggs against microscopic and macroscopic predators is a possible explanation, as demonstrated by
the antifungal effects of D. pealeii-associated Pseudoalteromonas strains (Barbieri
et al. 1997).
16.5.2 Incubation Conditions and Influence of Environmental
Factors during Embryogenesis
Both natural and artificial seawater have been used to incubate L. vulgaris and D.
opalescens eggs, although developmental problems have been observed mainly in
artificial seawater. The perivitelline fluid inside the egg chorion is in osmotic equilibrium with seawater and will reflect changes in the concentration of major ions of
seawater; therefore, these ions must be within the required concentration for normal
egg development (i.e. calcium and potassium 9–15 mM, magnesium 48–68 mM
and sulphate 15–37 mM) (D’Aniello et al. 1987). Even though late-stage eggs developed normally when incubated in artificial seawater containing these major ions
in proper concentrations, early stage eggs did not, suggesting that other elements
are required. This was later addressed by Hanlon et al. (1989), who demonstrated
that the embryo statoliths do not develop normally when strontium levels are below
8 mg L
−1
. Paralarvae hatching without or with abnormal statoliths were termed
‘spinners’, as they displayed erratic swimming patterns making continuous loops
at the surface and being unable to descend in the water column. The importance of
strontium in the biomineralization process during embryogenesis remains unknown
and other factors must play a role in the formation of statoliths, since the spinners
are occasionally found from natural seawater cultures. Natural seawater is therefore
recommended for egg incubation.
Eggs of both species have been incubated in closed and open, flow-through culture systems (Tables 16.1 and 16.2) and exposed to both constant and fluctuating
water temperatures and salinity conditions. Temperature strongly influences the rate
of embryonic development, incubation time and hatching rate (Boletzky 1987) and
there is some evidence that it might also have an effect on the hatching rhythm
(Paulij et al. 1990). Sen (2005a) evaluated thermal tolerance limits of L. vulgaris intermediate stage eggs (11–13) from Izmir Bay, Turkey. When eggs were exposed to
temperatures from 6 to 28 °C (at 37 psu), hatching rates (defined as number of viable