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Annexe III
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corresponding alternatively to the beginning and the middle of each third of the total
incubation time minus the first 5 days. The three periods of incubation were determined as the
totally duration of incubation (estimated approximately at 60 days at this temperature),
divided in three equal periods of time. We defined three water potentials: (i) a “wet substrate”
(-10 KPa; i.e. 2 g of sterilized water/ g of vermiculite) that represents a soil after rainfalls, (ii)
an “intermediate substrate” (-398 KPa; i.e. 0.44 g of sterilized water / g of vermiculite) and
(iii) a “dry substrate” (-1000 KPa; i.e. 0.09 g of sterilized water/ g of vermiculite) that
represents a soil in drought period. At each period of incubation, two groups of three different
and new boxes were transferred from intermediate to the wet and to the dry substrates (see
Table 1). In the middle of the period, the two groups of boxes were transferred again with, for
each group, one box staying in the same substrate and the two others changed for the two
other substrates. One box stayed on intermediate substrate during all the course of incubation.
Finally, each box represented a different treatment of moisture substrate and has undergone a
maximum of two transfers during one period of incubation.
Data collection during incubation
Every other day, from day 5 until hatching, eggs were individually weighed with a Sartorius
electronic balance (to the nearest 0.001g). Boxes without eggs were also weighed to adjust
water potential in replacing water absorbed by eggs or escaped around edges of the box sides.
Eggs covered by Fungus or other aborted eggs were removed and opened to check the
content. Eggs with no viable embryos were excluded from statistical analyses of change in
egg mass during incubation.
After piping and before hatching, each turtle was placed on wet cotton in an individual plastic
cup. Incubation time of each egg was defined as the pip date minus the date where a white
spot appeared on the eggshell.
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