The experimental system consisted of 10 glass aquaria (17 l) in
an open, filtered (10 μm) and UV-treated seawater system where
temperature and salinity were computer controlled. Each aquarium
was stocked with 1800 taurine or non-taurine electroporated control
eggs (ca 100 eggs l−1
). This allowed the testing of the taurine treatment
and control in replicates of 5 aquaria treatment−1
. The NCM (National
Center for Mariculture) gilthead sea bream larval rearing protocol was
implemented (Tandler et al., 1989) which briefly included a 16 h/8 h
day/night photoperiod regime where 2 dph larvae were offered rotifers
(Brachionus rotundiformis) twice daily maintaining a prey concentration
of 10 rotifers ml−1
. The “greening” of the tanks was carried out by
adding the microalgae Nannochloropsis sp. twice daily to maintain a
tank concentration of 0.5 × 106 cells ml−1 until 10 dph. In parallel,
samples of treated and untreated eggs were incubated (as described
previously) to determine the rates of hatching and survival after 24 h.
In order to determine taurine content after electroporation, 8 l mesh
covered (275 μm) cages floating in 1500 l tanks were stocked with
electroporated or unelectroporated eggs (100 eggs l−1
) and sampled
for taurine determination. Larvae were also sampled at 2, 4, 6, 8 and
10 dph, 1.5 h after rotifer feeding. During sampling 10 larvae from
each experimental aquarium (5 larvae at 8 dph, due to small biomass
remaining in the aquaria) were measured using light microscopy
(OPTIKA B-350) for total body length (±0.1 mm), mastax number in
gut, (mastax is an indigestible part of the rotifer mouth apparatus that
can be used as a measure of rotifer feeding rate) and for the presence
of calculi in the urinary bladder which were recorded as small
(≤50 μm) or large (N50 μm). At the end of the experiment at 10 dph,
the remaining larvae in the experimental aquaria were counted to
determine survival rates and collected for taurine content analysis.