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Four incubation trials were conducted to evaluate the effects of incubation RH on hatchability, allantoic fluid volume, and embryo weight. In Trial 1, White Leghorn eggs were incubated at a RH of 55 or 69% and a dry bulb temperature of 37.8 C and hatchability was determined. In Trial 2, hatchability and allantoic fluid volume were determined (Day 14 of incubation) for eggs incubated at 43, 55, or 69% RH. In Trial 3, allantoic fluid volumes and embryo weights were determined on Days 10, 12, 14, and 16 of incubation for eggs incubated at 40, 55, or 70% RH. In Trial 4, hatchability, allantoic fluid volumes, and embryo weights were determined on Days 13, 14, and 15 of incubation for eggs incubated at 40, 55, or 70% RH from hens 34 and 49 wk of age. There were no differences (P > .05) in hatchability between eggs incubated at 55 or 69% RH in Trial 1, or among eggs incubated at 43, 55, or 69% RH in Trial 2. In Trial 4, hatchability was highest for eggs from 34-wk-old hens incubated at 55% RH. In Trial 2, percentage egg weight loss per day of incubation (.68, .55, and .40%) and projected 19-d weight loss (12.9, 10.45, and 7.6%) decreased significantly with increased incubation RH of 43, 55, and 69%, respectively. Allantoic fluid volume (as a percentage of initial egg weight) on Day 14 of incubation significantly increased with the increase in RH; 13.86, 15.45, and 17.93% for RH of 43, 55, and 69%, respectively. In Trial 3, allantoic fluid volume peaked on Day 14 of incubation and volume increased or decreased with corresponding incubation RH; 14.05, 16.25, and 19.44% for RH of 40, 55, and 70%. In Trial 4, allantoic fluid volume was greatest on Day 13 of incubation and volume increased or decreased with corresponding incubation RH 13.38, 16.91, and 18.68% for eggs from 34-wk-old hens and 11.12, 13.77, and 16.59% for eggs from 49-wk-old hens. Eggs from 49-wk-old hens had lower allantoic fluid volumes, but higher initial egg weights due to greater percentage egg weight loss during incubation. Incubation egg weight loss in different RH appears to result from the corresponding depletion of the allantoic fluid reservoir.
We incubated eggs of Calotes versicolor at four constant temperatures ranging from 24 degrees C to 33 degrees C to assess the effects of incubation temperature on hatching success, embryonic use of energy, and hatchling phenotypes that are likely to affect fitness. All viable eggs increased in mass throughout incubation due to absorption of water, and mass gain during incubation was dependent on initial egg mass and incubation temperature. The average duration of incubation at 24 degrees C, 27 degrees C, 30 degrees C, and 33 degrees C was 82.1 days, 60.5 days, 51.4 days, and 50.3 days, respectively. Incubation temperature affected hatching success, energy expenditure for embryonic development, and several hatchling traits examined, but it did not affect the sex ratio of hatchlings. Hatching success was lowest (3.4%) at 33 degrees C, but a higher incidence of deformed embryos was recorded from eggs incubated at this temperature compared to eggs incubated at lower temperatures. Most of the deformed embryos died at the last stage of incubation. Energy expenditure for embryonic development was, however, higher in eggs incubated at 33 degrees C than those similarly incubated at lower temperatures. A prolonged exposure of eggs of C. versicolor at 33 degrees C appears to have an adverse and presumably lethal effect on embryonic development. Hatching success at 24 degrees C was also low (43.3%), but hatchlings incubated at 24 degrees C did not differ in any of the examined traits from those incubated at two intermediate temperatures (27 degrees C and 30 degrees C). Hatchlings incubated at 33 degrees C were smaller (snout-vent length, SVL) than those incubated at lower incubation temperatures and had larger mass residuals (from the regression on SVL) as well as shorter head length, hindlimb length, tympanum diameter, and eye diameter relative to SVL. Hatchlings from 33 degrees C had significantly lower scores on the first axis of a principal component analysis representing mainly SVL-free head size (length and width) and fore- and hindlimb lengths, but they had significantly higher scores on the second axis mainly representing SVL-free wet body mass. Variation in the level of fluctuating asymmetry in eye diameter associated with incubation temperatures was quite high, and it was clearly consistent with the prediction that environmental stress associated with the highest incubation temperatures might produce the highest level of asymmetry. Newly emerged hatchlings exhibited sexual dimorphism in head width, with male hatchlings having larger head width than females.
Freshwater turtle eggs are normally subjected to fluctuations in incubation temperature during natural incubation. Because of this, developing embryos may make physiological adjustments to growth and metabolism in response to incubation at different temperatures. I tested this hypothesis by incubating eggs of the Brisbane river turtle Emydura signata under four different temperature regimes, constant temperatures of 24 degrees C and 31 degrees C throughout incubation, and two swapped-temperature treatments where incubation temperature was changed approximately halfway through incubation. Incubation at 31 degrees C took 42 d, and incubation at 24 degrees C took 78 d, with intermediate incubation periods for the swapped-temperature treatments. Hatchling mass, hatchling size, and total oxygen consumed during development were similar for all incubation regimes. The pattern of oxygen consumption during the last phase of incubation as reflected by rate of increase of oxygen consumption, peak oxygen consumption, and fall in oxygen consumption before hatching was determined solely by the incubation temperature during the last phase of incubation; that is, incubation temperature during the first phase of incubation had no influence on these factors. Thus there is no evidence of temperature compensation in growth or development during embryonic development of E. signata eggs.
Incubation of tissue slices in physiological buffers gives rise to significant changes in the intracellular ion concentrations, which may disturb subsequent X-ray microanalysis. In the present study it was attempted to design incubation conditions that retain the in vivo conditions better. The following variables were investigated: (1) exchange of Na+ in the incubation medium for K+, and exchange of Cl- for the less permeable gluconate anion; (2) incubation at 4 degrees C rather than at 37 degrees C; and (3) addition of dextran to the incubation medium. Brief exposure (a few seconds) of liver slices to a buffer causes changes in the intracellular Na, Cl and K concentrations, depending on the ionic composition of the buffer. Incubation in a normal physiological (high NaCl) buffer at 37 degrees C results in a further increase of Na and Cl and a further decrease in K in liver cells. The changes reach a maximum at 30 min and the concentrations then remain stable throughout a 2-h incubation. Incubation in sodium gluconate medium or addition of dextran to the physiological buffer somewhat reduces the changes in the intracellular ion composition (compared to the standard physiological incubation medium). Incubation in potassium gluconate medium results in a decrease in cellular Na and an increase in K. Quantitative morphological studies show that tissue oedema is observed to the same extent in hepatocytes incubated in sodium gluconate, potassium gluconate and physiological buffer containing 10% dextran. However, these buffers cause significantly less cell oedema than the physiological (high NaCl) buffer. Incubation of liver, cerebral cortex or submandibular gland slices in physiological (high NaCl) solutions at 4 degrees C for 4 h caused a more extensive increase in Na+ and decrease in K+ than incubation at 37 degrees C for 2 h. This suggests inhibition of the Na+, K(+)-ATPase under these conditions. As compared to incubation at 37 degrees C for 2 h, tissues incubated in potassium gluconate buffer at 4 degrees C for 4 h have a cellular K concentration closer to the in situ value. Cholinergic stimulation of tissue slices from cerebral cortex and submandibular gland at room temperature for 1 min shows the best physiological response in tissue slices preincubated at 4 degrees for 4 h in high KCl, potassium gluconate and high NaCl, in this order. The response can, however, only be seen, when cholinergic stimulation is carried out in a standard physiological buffer with a high NaCl concentration. It is concluded that in vitro storage of tissue for X-ray microanalysis is best carried out at 4 degrees C in a solution with a high K+ concentration.
Eggs of two small Australian lizards, Lampropholis guichenoti and Bassiana duperreyi, were incubated to hatching at 25 degrees C and 30 degrees C. Incubation periods were significantly longer at 25 degrees C in both species, and temperature had a greater effect on the incubation period of B. duperreyi (41.0 days at 25 degrees C; 23.1 days at 30 degrees C) than L. guichenoti (40.1 days at 25 degrees C; 27.7 days at 30 degrees C). Patterns of oxygen consumption were similar in both species at both temperatures, being sigmoidal in shape with a fall in the rate of oxygen consumption just prior to hatching. The higher incubation temperature resulted in higher peak and higher prehatch rates of oxygen consumption in both species. Total amount of oxygen consumed during incubation was independent of temperature in B. duperreyi, in which approximately 50 ml oxygen was consumed at both temperatures, but eggs of L. guichenoti incubated at 30 degrees C consumed significantly more (32.6 ml) than eggs incubated at 25 degrees C (28.5 ml). Hatchling mass was unaffected by either incubation temperature or the amount of water absorbed by eggs during incubation in both species. The energetic production cost of hatchling B. duperreyi (3.52 kJ x g(-1)) was independent of incubation temperature, whereas in L. guichenoti the production cost was greater at 30 degrees C (4.00 kJ x g(-1)) than at 25 degrees C (3.47 kJ g(-1)). Snout-vent lengths and mass of hatchlings were unaffected by incubation temperature in both species, but hatchling B. duperreyi incubated at 30 degrees C had longer tails (29.3 mm) than those from eggs incubated at 25 degrees C (26.2 mm). These results indicate that incubation temperature can affect the quality of hatchling lizards in terms of embryonic energy consumption and hatchling morphology.