Contributions to Zoology, 78 (2) - 2009Milena Cvijanovic; Ana Ivanovic; Natasa Tomasevic Kolarov; Georg Dzukic; Milos L. Kalezic: Early ontogeny shows the same interspecific variation as natural history parameters in the crested newt (Triturus cristatus superspecies) (Caudata, Salamandridae)
Materials and methods

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Analysed traits

The eggs deposited by one female over a 24 h period were used as cohorts, and the mean values of analysed traits were calculated separately for each cohort. Eggs were photographed immediately after removal from the plastic strips for measurements. The morphologically defined developmental events (stages of embryonic development) were established according to a description of embryonic stages for M. alpestris (Knight, 1938; see also Epperlein and Junginger, 1982). To estimate the variation in timing of developmental events within and between species, the developing embryos were examined under a binocular microscope by M. C. and N. T. K. at three different time points: at 7, 11 and 15 days following egg laying. The embryonic developmental stage was recorded at each particular checkpoint (at day 7, 11, and 15 of embryonic development), hereafter designated as S7, S11 and S15, respectively. Such experimental design provided a basis for analysing the variation in developmental events and developmental rates within and between species. Undeveloped eggs and dead embryos were removed regularly. Due to the possible effect of signalling between embryos on embryonic induction (Hall, 1999; 2003), median values for recorded stages of embryos per Petri dish (up to 10 embryos) were calculated; these data were used in further analyses. Hatched larvae were photographed with a digital camera (Nikon Coolpix 4500) and a 10-mm scale bar to measure the total length of larvae.

The following eight life-history traits were recorded: time of egg deposition relative to the oviposition period (DO), number of eggs laid per cohort (NE), mean vitellus diameter per cohort (RV), volume of galerta calculated as a difference among egg’s volume and vitellus volume (VG), number of hatched larvae per cohort (NH), total length of hatched larvae per cohort (TL), duration of embryonic period of embryos per cohort (EP) and total hatchling survival rate of embryos per cohort (SR). Measurements of egg and vitellus diameter and TL of hatched larvae were taken by M. C. using UTHSCSA IMAGETOOL version 3.0 (http://ddsdx.uthscsa.edu/dig/itdesc.html).