A Pilot Study on the Feasibility of an Extended Suckling System for Pasture-Based Dairies.

This study investigated cow-calf productivity in a 10-week, pasture-based, extended suckling system featuring part-time cow-calf contact and once-a-day milking. A total of 30 dairy cows and their calves were assigned to two treatments: (1) cow and calf managed in an extended suckling system; or (2) cow and calf separated at birth and managed as usual. Cow-calf pairs grazed together during the day and spent the night separated by fence-line contact. The dams were reunited with the calves after once-a-day milking every morning. The commercial treatment pairs were separated after birth, and cows were milked twice a day and managed within the farm herd. Commercial calves were reared and managed as per commercial Australian practices. Cow-calf dams yielded 9 L/cow/day less saleable milk (p < 0.001), and their milk had lower fat (p = 0.04) but a higher protein percentage (p < 0.001) than commercial cows during pre-weaning. However, milk yield and composition were comparable post-weaning. Dam-suckled calves gained weight faster and were therefore weaned 2 weeks earlier than commercial calves, which were offered 8 L/day milk. This study has demonstrated a novel system of extended cow-calf suckling that could be practical to implement in pasture-based dairies. The long-term effects and scalability of the extended suckling system described here require further validation.

Acid-base and hematological regulation in chicken embryos during internal progressive hypercapnic hypoxia.

Development of the capacity to mitigate potential disturbances to blood physiology in bird embryos is incompletely understood. We investigated regulation of acid-base and hematology in day 15 chicken embryos exposed to graded intrinsic hypercapnic hypoxia created by varying degrees of water submersion. Metabolic acidosis with additional respiratory or metabolic acidosis occurred at 2 h according to magnitude of submersion. Acid-base disturbance was partially compensated by metabolic alkalosis at 6 h, but compensatory metabolic alkalosis was absent at 24 h. Following submersion with only air cell exposed to air, both hypercapnic respiratory acidosis and metabolic acidosis occurred within 10 min. Subsequently, both forms of acidosis created lethal levels of [HCO3-] at ∼120 min. Blood hematology showed small but significant effects associated with induced acid-base disturbance. Increased Hct occurring during partial egg submersion lasting 24 h was attributed to an increase in MCV. By day 15 of development chicken embryos are able to partially compensate for and withstand all but severe induced internal hypoxic hypercapnia.

Hypoxia during incubation does not affect aerobic performance or haematology of Atlantic salmon (Salmo salar) when re-exposed in later life.

Hypoxia in aquatic ecosystems is becoming increasingly prevalent, potentially reducing fish performance and survival by limiting the oxygen available for aerobic activities. Hypoxia is a challenge for conserving and managing fish populations and demands a better understanding of the short- and long-term impacts of hypoxic environments on fish performance. Fish acclimate to hypoxia via a variety of short- and long-term physiological modifications in an attempt to maintain aerobic performance. In particular, hypoxia exposure during early development may result in enduring cardio-respiratory modifications that affect future hypoxia acclimation capacity, yet this possibility remains poorly investigated. We incubated Atlantic salmon (Salmo salar) in normoxia (~100% dissolved oxygen [DO, as percent air saturation]), moderate hypoxia (~63% DO) or cyclical hypoxia (100-25% DO daily) from fertilization until 113 days post-fertilization prior to rearing all groups in normoxia for a further 8 months. At ~11 months of age, subsets of each group were acclimated to hypoxia (50% DO) for up to 44 days prior to haematology, aerobic metabolic rate and hypoxia tolerance measurements. Hypoxia exposure during incubation (fertilization to 113 days post-fertilization) did not affect the haematology, aerobic performance or hypoxia tolerance of juvenile salmon in later life. Juveniles acclimated to hypoxia increased maximum aerobic metabolic rate and aerobic scope by ~23 and ~52%, respectively, when measured at 50% DO but not at 100% DO. Hypoxia-incubated juveniles also increased haematocrit and haemoglobin concentration but did not affect acute hypoxia tolerance (critical oxygen level and DO at LOE). Thus, while Atlantic salmon possess a considerable capacity to physiologically acclimate to hypoxia by improving aerobic performance in low oxygen conditions, we found no evidence that this capacity is influenced by early-life hypoxia exposure.

Physiological effects of dissolved oxygen are stage-specific in incubating Atlantic salmon (Salmo salar).

Oxygen availability is highly variable during salmonid incubation in natural redds and also in aquaculture incubation systems. Hypoxia generally decreases growth and aerobic metabolism prior to hatching, in parallel with eliciting physiological modifications that enhance oxygen delivery. However, it is less-well known whether developmental hyperoxia can drive the opposite effect. Moreover, there is insufficient understanding of stage-specific developmental windows during which ambient oxygen availability may be of greater or lesser impact to incubating embryos. Here, we tested the effects of hypoxia (50% dissolved oxygen: DO, % air saturation) and hyperoxia (150% DO) on the growth, routine aerobic metabolism ([Formula: see text]) and hypoxia tolerance (O2crit) of Atlantic salmon (Salmo salar) during seven developmental windows throughout incubation. Embryos exposed to hyperoxia (150% DO) did not differ from the normoxic group in growth, [Formula: see text] or O2crit at any developmental window. In contrast, embryos exposed to hypoxia grew slower and had a lower [Formula: see text], but had higher hypoxia tolerance (lower O2crit) than normoxic and hyperoxic counterparts. Interestingly, these differences were only apparent when the embryos were measured prior to hatching. Larvae (alevins) incubated in hypoxia following hatching grew similarly to normoxia-incubated alevins. Our results provide evidence that Atlantic salmon embryos are most sensitive to hypoxia prior to hatching, probably due to increasing (absolute) oxygen requirements concurrent with restricted oxygen diffusion through the egg. Moreover, the similarities between normoxia- and hyperoxia-incubated salmon demonstrate that embryos are not oxygen-limited under normoxic conditions.

Negligible differences in metabolism and thermal tolerance between diploid and triploid Atlantic salmon (Salmo salar).

The mechanisms that underlie thermal tolerance in aquatic ectotherms remain unresolved. Triploid fish have been reported to exhibit lower thermal tolerance than diploids, offering a potential model organism to better understand the physiological drivers of thermal tolerance. Here, we compared triploid and diploid juvenile Atlantic salmon (Salmo salar) in freshwater to investigate the proposed link between aerobic capacity and thermal tolerance. We measured specific growth rates (SGR) and resting (aerobic) metabolic rates (RMR) in freshwater at 3, 7 and 9 weeks of acclimation to 10, 14 and 18°C. Additionally, maximum metabolic rates (MMR) were measured at 3 and 7 weeks of acclimation, and critical thermal maxima (CTmax) were measured at 9 weeks. Mass, SGR and RMR differed between ploidies across all temperatures at the beginning of the acclimation period, but all three metrics were similar across ploidies by week 7. Aerobic scope (MMR-RMR) remained consistent across ploidies, acclimation temperatures and time. At 9 weeks, CTmax was independent of ploidy, but correlated positively with acclimation temperature despite the similar aerobic scope between acclimation groups. Our findings suggest that acute thermal tolerance is not modulated by aerobic scope, and the altered genome of triploid Atlantic salmon does not translate to reduced thermal tolerance of juvenile fish in freshwater.

Developmental Hypoxia Has Negligible Effects on Long-Term Hypoxia Tolerance and Aerobic Metabolism of Atlantic Salmon (Salmo salar).

Exposure to developmental hypoxia can have long-term impacts on the physiological performance of fish because of irreversible plasticity. Wild and captive-reared Atlantic salmon (Salmo salar) can be exposed to hypoxic conditions during development and continue to experience fluctuating oxygen levels as juveniles and adults. Here, we examine whether developmental hypoxia impacts subsequent hypoxia tolerance and aerobic performance of Atlantic salmon. Individuals at 8°C were exposed to 50% (hypoxia) or 100% (normoxia) dissolved oxygen (DO) saturation (as percent of air saturation) from fertilization for ∼100 d (800 degree days) and then raised in normoxic conditions for a further 15 mo. At 18 mo after fertilization, aerobic scope was calculated in normoxia (100% DO) and acute (18 h) hypoxia (50% DO) from the difference between the minimum and maximum oxygen consumption rates ([Formula: see text] and [Formula: see text], respectively) at 10°C. Hypoxia tolerance was determined as the DO at which loss of equilibrium (LOE) occurred in a constantly decreasing DO environment. There was no difference in [Formula: see text], [Formula: see text], or aerobic scope between fish raised in hypoxia or normoxia. There was some evidence that hypoxia tolerance was lower (higher DO at LOE) in hypoxia-raised fish compared with those raised in normoxia, but the magnitude of the effect was small (12.52% DO vs. 11.73% DO at LOE). Acute hypoxia significantly reduced aerobic scope by reducing [Formula: see text], while [Formula: see text] remained unchanged. Interestingly, acute hypoxia uncovered individual-level relationships between DO at LOE and [Formula: see text], [Formula: see text], and aerobic scope. We discuss our findings in the context of developmental trajectories and the role of aerobic performance in hypoxia tolerance.

Calibration of the HemoCue point-of-care analyser for determining haemoglobin concentration in a lizard and a fish.

Haemoglobin concentration ([Hb]) is measured for a wide variety of animal studies. The use of point-of-care devices, such as the HemoCue, is becoming increasingly common because of their portability, relative ease of use and low cost. In this study, we aimed to determine whether the [Hb] of blue-tongued skink (Tiliqua nigrolutea) blood can be determined accurately using the HemoCue and whether the HemoCue overestimates the [Hb] of reptile blood in a similar manner to fish blood. Additionally, we aimed to test whether ploidy affected [Hb] determined by the HemoCue using blood from diploid and triploid Atlantic salmon (Salmo salar). The HemoCue Hb 201(+) systematically overestimated [Hb] in both blue-tongued skinks and Atlantic salmon, and there was no difference between calibration equations determined for diploid or triploid salmon. The overestimation was systematic in both species and, as such, [Hb] determined by the HemoCue can be corrected using appropriate calibration equations.

Acid-base balance in the developing marsupial: from ectotherm to endotherm.

Marsupial joeys are born ectothermic and develop endothermy within their mother's thermally stable pouch. We hypothesized that Tammar wallaby joeys would switch from α-stat to pH-stat regulation during the transition from ectothermy to endothermy. To address this, we compared ventilation (Ve), metabolic rate (Vo2), and variables relevant to blood gas and acid-base regulation and oxygen transport including the ventilatory requirements (Ve/Vo2 and Ve/Vco2), partial pressures of oxygen (PaO2), carbon dioxide (PaCO2), pHa, and oxygen content (CaO2) during progressive hypothermia in ecto- and endothermic Tammar wallabies. We also measured the same variables in the well-studied endotherm, the Sprague-Dawley rat. Hypothermia was induced in unrestrained, unanesthetized joeys and rats by progressively dropping the ambient temperature (Ta). Rats were additionally exposed to helox (80% helium, 20% oxygen) to facilitate heat loss. Respiratory, metabolic, and blood-gas variables were measured over a large body temperature (Tb) range (∼15-16°C in both species). Ectothermic joeys displayed limited thermogenic ability during cooling: after an initial plateau, Vo2 decreased with the progressive drop in Tb. The Tb of endothermic joeys and rats fell despite Vo2 nearly doubling with the initiation of cold stress. In all three groups the changes in Vo2 were met by changes in Ve, resulting in constant Ve/Vo2 and Ve/Vco2, blood gases, and pHa. Thus, although thermogenic capability was nearly absent in ectothermic joeys, blood acid-base regulation was similar to endothermic joeys and rats. This suggests that unlike some reptiles, unanesthetized mammals protect arterial blood pH with changing Tb, irrespective of their thermogenic ability and/or stage of development.

Acute regulation of hematocrit and acid-base balance in chicken embryos in response to severe intrinsic hypercapnic hypoxia.

The regulation of blood acid-base balance and hematology in day 15 chicken embryos in response to partial water submersion (with egg's air cell in air) and complete submersion producing severe intrinsic hypercapnic hypoxia and recovery in air was studied. The acid-base disturbance during submersion was characterized by initial rapid respiratory changes and then superseded by metabolic processes, resulting in a large progressive hysteresis. Throughout submersion and recovery, blood lactate concentration changed swiftly along with the changes in bicarbonate concentration ([HCO3(-)]), indicating that anaerobic glycolysis determined overall acid-base disturbances. Both partial and complete submersion produced large, rapid increases in hematocrit through proportional increases in mean corpuscular volume and red blood cell concentration. Death ensued once the internal pool of O2 was exhausted and/or the acid-base disturbance became too severe for survival (i.e., [HCO3(-)]a<∼10mmolL(-1)). However, embryos recovered from acid-base and hematological disturbances within 120min recovery in air after short bouts of complete (20min) or partial (60min) submersion, suggesting that shorter severe intrinsic hypercapnic hypoxia does not compromise viability of embryos.

Transgenerational variation in metabolism and life-history traits induced by maternal hypoxia in Daphnia magna.

Hypoxic stress can alter conspecific phenotype and additionally alter phenotypes of the filial generation, for example, via maternal or epigenetic processes. Lasting effects can also be seen across development and generations even after stressors have been removed. This study utilized the model of rapidly developing, parthenogenetic Daphnia to examine the intraspecific variability of response of exposure of a parental generation to hypoxia (4 kPa) within a single clone line across development, across broods, and across generations. Body mass across development and reproductive output were monitored in the parental generation and the first three broods of the first filial generation (which were not directly exposed to hypoxia). O(2) consumption across a wide Po(2) range (normoxia to anoxia) was assessed to determine whether exposure of the parental generation to hypoxia conferred hypoxia tolerance on the offspring and whether this transgenerational, epigenetic phenomenon varied intraspecifically. Differences in mass occurred in both the parental generation (hypoxia-exposed smaller during brood 1 and brood 2 neonate production) and the filial generation (e.g., brood 1 and 2 neonates from hypoxic mothers were initially smaller than control neonates). However, differences in mass were not accompanied by changes in reproductive output (assessed by brood number and neonate size). At day 0, first filial generation brood 1 neonates from hypoxia-exposed mothers had a higher metabolic rate than control neonates. However, this effect, like that of body mass, dissipated with development within a brood but also with subsequent broods. An isometric scaling exponent for [Formula: see text] was repeatedly observed across a wide Po(2) range (21-2 kPa) throughout neonatal development.

Acute regulation of hematocrit and blood acid-base balance during severe hypoxic challenges in late chicken embryos (Gallus gallus).

Acid-base and hematocrit (Hct) responses of vertebrate embryos to severe hypoxia are as yet unknown, but may reveal the maturation process of physiological regulatory mechanisms. The present study elucidated how acute, severe hypoxia (10% O2, with and without 5% CO2) affects Hct and blood acid-base balance in late prenatal (days 11-19) chicken embryos. The time-course of the resulting Hct changes and blood acid-base disturbances was examined in detail in day 15 (d15) embryos to further understand the magnitude and time-components of these physiological changes. We hypothesized that Hct of developing embryos increases during severe hypoxia (10% O2) and hypercapnic hypoxia (5% CO2, 10% O2), due to increased mean corpuscular volume (MCV) and red blood cell concentration ([RBC]). We additionally hypothesized that 10% O2 would induce anaerobic glycolysis and the attendant increase in lactate concentration ([La-]) would create a severe metabolic acidosis. Hct increased in all embryos (d11-d19) during severe hypoxia (2h) but, with the exception of d19 embryos, the increase was due to increased MCV and was therefore unlikely related to O2 transport. The time-course of the d15 embryonic Hct response to hypoxic or hypercapnic hypoxic exposure was very rapid with MCV increasing within 30min. Severe metabolic acidosis occurred in all developing embryos (d11-d19) during 2h hypoxic exposure. Additionally, respiratory acidosis was induced in d15 embryos during hypercapnic hypoxia, with acid-base status recovering within 120 min in air. Throughout hypoxic exposure and recovery, changes in [HCO3-] were matched by those in [La-], indicating that anaerobic glycolysis is a key factor determining the metabolic alterations and overall acid-base status. Further, the blood gas and Hct values recovered in air and unchanged embryo mass suggest that the hypoxia-induced disturbances were only transient and may not affect long-term survival.

Interactions of acid-base balance and hematocrit regulation during environmental respiratory gas challenges in developing chicken embryos (Gallus gallus).

How the determinants of hematocrit (Hct) - alterations in mean corpuscular volume (MCV) and/or red blood cell concentration ([RBC]) - are influenced by acid-base balance adjustments across development in the chicken embryo is poorly understood. We hypothesized, based on oxygen transport needs of the embryos, that Hct will increase during 1 day of hypercapnic hypoxia (5%CO(2), 15%O(2)) or hypoxia alone (0%CO(2), 15%O(2)), but decrease in response to hyperoxia (0%CO(2), 40%O(2)). Further, age-related differences in acid-base disturbances and Hct regulation may arise, because the O(2) transport and hematological regulatory systems are still developing in embryonic chickens. Our studies showed that during 1 day of hypoxia (with or without hypercapnia) Hct increased through both increased MCV and [RBC] in day 15 (d15) embryo, but only through increased MCV in d17 embryo and therefore enhancement of O(2) transport was age-dependent. Hypercapnia alone caused a ≈ 14% decrease in Hct through decreased [RBC] and therefore did not compensate for decreased blood oxygen affinity resulting from the Bohr shift. The 11% (d15) and 14% (d17) decrease in Hct during hyperoxia in advanced embryos was because of an 8% and 9% decrease, respectively, in [RBC], coupled with an associated 3% and 5% decrease in MCV. Younger, d13 embryos were able to metabolically compensate for respiratory acidosis induced by hypercapnic hypoxia, and so were more tolerant of disturbances in acid-base status induced via alterations in environmental respiratory gas composition than their more advanced counterparts. This counter-intuitive increased tolerance likely results from the relatively low [Formula: see text] and immature physiological functions of younger embryos.

The accessory role of the diaphragmaticus muscle in lung ventilation in the estuarine crocodile Crocodylus porosus.

Crocodilians use a combination of three muscular mechanisms to effect lung ventilation: the intercostal muscles producing thoracic movement, the abdominal muscles producing pelvic rotation and gastralial translation, and the diaphragmaticus muscle producing visceral displacement. Earlier studies suggested that the diaphragmaticus is a primary muscle of inspiration in crocodilians, but direct measurements of the diaphragmatic contribution to lung ventilation and gas exchange have not been made to date. In this study, ventilation, metabolic rate and arterial blood gases were measured from juvenile estuarine crocodiles under three conditions: (i) while resting at 30°C and 20°C; (ii) while breathing hypercapnic gases; and (iii) during immediate recovery from treadmill exercise. The relative contribution of the diaphragmaticus was then determined by obtaining measurements before and after transection of the muscle. The diaphragmaticus was found to make only a limited contribution to lung ventilation while crocodiles were resting at 30°C and 20°C, and during increased respiratory drive induced by hypercapnic gas. However, the diaphragmaticus muscle was found to play a significant role in facilitating a higher rate of inspiratory airflow in response to exercise. Transection of the diaphragmaticus decreased the exercise-induced increase in the rate of inspiration (with no compensatory increases in the duration of inspiration), thus compromising the exercise-induced increases in tidal volume and minute ventilation. These results suggest that, in C. porosus, costal ventilation alone is able to support metabolic demands at rest, and the diaphragmaticus is largely an accessory muscle used at times of elevated metabolic demand.

Hematocrit and blood osmolality in developing chicken embryos (Gallus gallus): in vivo and in vitro regulation.

Hematocrit (Hct) regulation is a complex process involving potentially many factors. How such regulation develops in vertebrate embryos is still poorly understood. Thus, we investigated the role of blood pH in the regulation of Hct across developmental time in chicken embryos. We hypothesized that blood pH alterations in vitro (i.e., in a test tube) would affect Hct far more than in vivo because of in vivo compensatory regulatory processes for Hct. Large changes in Hct (through mean corpuscular volume (MCV)) and blood osmolality (Osm) occur when the blood was exposed to varying ambient temperatures (T(a)'s) and P(CO2) in vitro alongside an experimentally induced blood pH change from ~7.3 to 8.2. However, homeostatic regulatory mechanisms apparently limited these alterations in vivo. Changes in blood pH in vitro were accompanied by hydration or dehydration of red blood cells depending on embryonic age, resulting in changes in Hct that also were specific to developmental stage, due likely to initial blood gas and [HCO(3)(-)](v) values. Significant linear relationships between Hct and pH (Hct/ΔpH=-21.4%/(pH unit)), Hct and [HCO(3)(-)] (ΔHct/Δ[HCO(3)(-)]=1.6%/(mEq L(-1))) and the mean buffer value (Δ[HCO(3)(-)]/ΔpH=-13.4 (mEq L(-1))/(pH unit)) demonstrate that both pH and [HCO(3)(-)] likely play a role in the regulation of Hct through MCV at least in vitro. Low T(a) (24°C) resulted in relatively large changes in pH with small changes in Hct and Osm in vitro with increased T(a) (42°C) conversely resulting in larger changes in both Hct and Osm. In vivo exposure to altered T(a) caused age-dependent changes in Hct, demonstrating a trend towards increased Hct at higher T(a). Further, exposing embryos to a gas mixture where P(CO2) = 5.1 kPa for >4 h period at T(a) of 37 or 42°C also did not elicit a change in Hct or Osm. Presumably, homeostatic mechanisms ensured that in vivo Hct was stable during a 4-6 h temperature and/or hypercapnic stress. Thus, although blood pH decreases (induced by acute T(a) increase and exposure to CO(2)) increase MCV and, consequently, Hct in vitro, homeostatic mechanisms operating in vivo are adequate to ensure that such environmental perturbations have little effect in vivo.

Embryonic control of heart rate: examining developmental patterns and temperature and oxygenation influences using embryonic avian models.

Long-term measurements (days and weeks) of heart rate (HR) have elucidated infradian rhythms in chicken embryos and circadian rhythms in chicken hatchlings. However, such rhythms are lacking in emu embryos and only rarely observed in emu hatchlings. Parasympathetic control of HR (instantaneous heart rate (IHR) decelerations) occurs at ∼60% of incubation in both precocial and altricial avian embryos, with sympathetic control (IHR accelerations) becoming more prevalent close to hatching. A large increase in avian embryonic HR occurs during hatching (presumably an energetically expensive process, i.e. increased oxygen consumption M(O) 2), beginning during pipping when a physical barrier to O(2) conductance is removed. Alterations in ambient O(2) have little effect on early embryonic HR, likely due to the low rate of M(O)2 of early embryos and the fact that adequate O(2) delivery can occur via diffusion. As M(O)2 increases in advanced embryos and circulatory convection becomes important for O(2) delivery, alterations in ambient O(2) have more profound effects on embryonic HR. Early embryos demonstrate a wide ambient temperature (T(a)) tolerance range compared with older embryos. In response to a rapid decrease in T(a), embryonic HR decreases (stroke volume and blood flow are preserved) in an exponential fashion to a steady state (from which it can potentially recover if re-warmed). A more severe decrease in T(a) results in complete cessation of HR; however, depending on developmental age, embryos are able to survive severe cold exposure and cessation of HR for up to 24h in some instances. The development of endothermy can be tracked by measuring baseline HR during T(a) changes. HR patterns change from thermo-conformity to thermoregulation (reverse to T(a) changes). Further, IHR low frequency oscillations mediated by the autonomic nervous system are augmented at low T(a)s in hatchlings. Transitions of baseline HR during endothermic development are unique to individual avian species (e.g. chickens, ducks and emu), reflecting differences in life history.

Development of hematological respiratory variables in late chicken embryos: the relative importance of incubation time and embryo mass.

Oxygen demand increases during embryonic development, requiring an increase in red blood cells (RBCs) containing hemoglobin (Hb) to transport O(2) between the respiratory organ and systemic tissues. A thorough ontogenetic understanding of the onset and maturation of the complex regulatory processes for RBC concentration ([RBC]), Hb concentration ([Hb]), hematocrit (Hct), mean corpuscular indices (mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH) and mean corpuscular hemoglobin concentration ([MCHb])) is currently lacking. We hypothesize that during the last half of incubation when the respiratory organ (the chorioallantoic membrane) envelops most of the egg contents, mean corpuscular indices will stabilize. Accordingly, Hct, [RBC] and [Hb] must also all change proportionally across development. Further, we hypothesize that the hematological respiratory variables develop and mature as a function of incubation duration, independently of embryonic growth. As predicted, a similar increase in Hct (from 18.7±0.6% on day 10 (d10) to 34.1±0.5% on d19 of incubation), [RBC] (1.13±0.03×10(6)/µL to 2.50±0.03×10(6)/µL) and [Hb] (6.1±0.2 g% to 11.2±0.1 g%) occurred during d10-19. Both [RBC] and [Hb] demonstrated high linear correlation with Hct, resulting in constant [MCHb] (~33 g% from d10 to d19). The decrease in MCV (from ~165 µ(3) on d10 to ~140 µ(3) on d13) and MCH (~55 pg to ~45 pg) during d10-13, may be attributed to a changeover from larger primary to smaller secondary and adult-type erythrocytes with MCV and MCH remaining constant (~140 µ(3) and ~45 pg respectively) for the rest of the incubation period (d13-19). Hematological respiratory values on a given incubation day were identical between embryos of different masses using either natural mass variation or experimental growth acceleration, indicating that the hematological variables develop as a function of incubation time, irrespective of embryo growth.

Does incubation temperature fluctuation influence hatchling phenotypes in reptiles? A test using parthenogenetic geckos.

Many lineages of parthenogenetic organisms have persisted through significant environmental change despite the constraints imposed by their fixed genotype and limited evolutionary potential. The ability of parthenogens to occur sympatrically with sexual relatives may in part be due to phenotypic plasticity in their responses to their environment, especially with respect to incubation temperature--a maternally selected trait. Here we measured the incubation temperatures selected by two lineages of triploid parthenogenic geckos in the Heteronotia binoei complex by allowing them to deposit clutches along a thermal gradient. The average nest temperature selected was 28.4 degrees C, with no significant differences between parthenogenic races or individual clones. To investigate the effect of nest-temperature variability on physiological and morphological traits, we incubated eggs from different races at one of four incubation regimes (32 degrees +/- 0 degrees, +/- 3 degrees , +/- 5 degrees , or +/- 9 degrees C). Embryos incubated at constant 32 degrees C developed faster than embryos reared under increasing extremes of diel temperature fluctuation (+/- 3 degrees , +/- 5 degrees C), and incubation at 32 degrees +/- 9 degrees C was unsuccessful. Incubation regime had no effect on the body size, preferred substrate temperature, or mass-specific .V(O2) of hatchlings. However, parthenogenic race had a significant effect on egg mass, tail length, snout-to-vent length, total length, and .V(O2) . We conclude that developmental traits are strongly influenced by clonal genotypes in this parthenogenic complex but are well buffered against fluctuations in incubation temperature.

Phenotypic differences in terrestrial frog embryos: effect of water potential and phase.

The terrestrial embryos of many amphibians obtain water in two ways; in a liquid phase from the substrate on which eggs are deposited, and in a vapour phase from the surrounding atmosphere. We tested whether the mode of water flux (liquid or vapour) affected the morphology and metabolic traits of the terrestrial Victorian smooth froglet (Geocrinia victoriana) embryos by incubating eggs both with a liquid water source and at a range of vapour water potentials. We found that embryos incubated with a liquid water source (psi(pi)=0 kPa) were better hydrated than embryos incubated with a vapour water source (psi(v)=0 kPa), and grew to a larger size. Eggs incubated in atmospheres with lower psi(v) values showed significant declines in mass and in the thickness of the jelly capsule, while embryos primarily showed reductions in dry mass, total length, tail length and fin height. The most significant deviations from control (psi(v)=0 kPa) values were observed when the psi(v) of the incubation media was less than the osmotic water potential (psi(pi)) of the embryonic interstitial fluid (approximately -425 kPa). Despite the caveat that a psi(v) of 0 kPa is probably difficult to achieve under our experimental conditions, the findings indicate the importance for eggs under natural conditions of contacting liquid water in the nesting substrate to allow swelling of the capsule.