Large Offspring Research Articles

Investigation of Imprinted Gene Expression for In Vitro Fertilized Mouse Embryos

Abstract: It has been shown that epigenetic abnormalities are involved in both abnormal fetal development, as represented by large offspring syndrome, and the onset of diseases affecting vital prognosis, such as metabolic syndrome and malignancy. And if assisted reproductive technology (ART) causes epigenetic abnormalities, then this must be avoided. We compared the expression of imprinted genes (Igf2, Peg3, Snrpn and Kcnq1ot1), which are expressed in early embryos and are involved in fetal and placental development, using in vivo and in vitro mouse blastocysts. In order to compare the changes in gene expression under different culture conditions, in vitro embryos were incubated using two culture media (KSOM/AA and M16) and two gas phases (O2 5% and 20%). Between in vivo embryos and KSOM/AA embryos, no marked differences were seen in gene expression. However, between in vivo embryos and M16 embryos, changes in gene expression patterns were confirmed. By changing the incubator's O2 concentration, even when...

Recombinational landscape of porcine X chromosome and individual variation in female meiotic recombination associated with haplotypes of Chinese pigs

BackgroundVariations in recombination fraction (θ) among chromosomal regions, individuals and families have been observed and have an important impact on quantitative trait loci (QTL) mapping studies. Such variations on porcine chromosome X (SSC-X) and on other mammalian chromosome X are rarely explored. The emerging assembly of pig sequence provides exact physical location of many markers, facilitating the study of a fine-scale recombination landscape of the pig genome by comparing a clone-based physical map to a genetic map. Using large offspring of F1 females from two large-scale resource populations (Large White ♂ × Chinese Meishan ♀, and White Duroc ♂ × Chinese Erhualian ♀), we were able to evaluate the heterogeneity in θ for a specific interval among individual F1 females.ResultsAlignments between the cytogenetic map, radiation hybrid (RH) map, genetic maps and clone map of SSC-X with the physical map of human chromosome X (HSA-X) are presented. The most likely order of 60 markers on SSC-X is inferred. The average recombination rate across SSC-X is of ~1.27 cM/Mb. However, almost no recombination occurred in a large region of ~31 Mb extending from the centromere to Xq21, whereas in the surrounding regions and in the Xq telomeric region a recombination rate of 2.8-3.3 cM/Mb was observed, more than twice the chromosome-wide average rate. Significant differences in θ among F1 females within each population were observed for several chromosomal intervals. The largest variation was observed in both populations in the interval UMNP71-SW1943, or more precisely in the subinterval UMNP891-UMNP93. The individual variation in θ over this subinterval was found associated with F1 females' maternal haplotypes (Chinese pig haplotypes) and independent of paternal haplotype (European pig haplotypes). The θ between UMNP891 and UMNP93 for haplotype 1122 and 4311 differed by more than fourteen-fold (10.3% vs. 0.7%).ConclusionsThis study reveals marked regional, individual and haplotype-specific differences in recombination rate on SSC-X. Lack of recombination in such a large region makes it impossible to narrow QTL interval using traditional fine-mapping approaches. The relationship between recombination variation and haplotype polymorphism is shown for the first time in pigs.

Congenital malformations after assisted reproduction: risks and implications for prenatal diagnosis and fetal medicine

Congenital malformations after assisted reproduction: risks and implications for prenatal diagnosis and fetal medicine

Genetic evaluation of Devon Cattle using a reaction norms model

The objective of this study was to evaluate differences in sire genetic values obtained by a conventional animal model (AM) and by a reaction norms hierarchical model (RNHM) that considers the genotype by environment interaction. A total of 25,500 records was used collected by PROMEBO - beef cattle improvement program of the National Breeders Association "Herd Book Collares" on Devon cattle born from 1980 though 2005. Post weaning gain 345 adjusted (PWG345) was the evaluated trait and the INTERGEN program was employed for data analysis. Reaction norms of the 25 sires with larger offspring showed ranking exchanges and scale effect in the environmental gradient. Spearman rank correlations between genetic values obtained by AM and RNHM at low, medium and high environmental levels were used to assess selection decision based on the different models. These correlations ranged from 0.12 and 0.99, therefore showing substantial changes in ranking, particularly between those genetic values obtained by the RNHM at low level compared to the same RNHM at medium and high levels and to AM, when considering the top 5% bulls. The results indicated that the selection process should consider the genotype by environmental interaction to maximize genetic gain and production of this population in each specific environment.

57 RELATIONSHIP BETWEEN FETAL ABNORMALITIES AND PERIPHERAL STEROID CONCENTRATIONS DURING GESTATION IN COWS TRANSFERRED WITH EMBRYOS PRODUCED BY SOMATIC CELL NUCLEAR TRANSFER

Cloning of mammals by nuclear transfer frequently results in gestational or neonatal failure with a variety of abnormalities that are likely caused by inappropriate epigenetic reprogramming. Early diagnosis of fetal abnormality is important for efficient production of cloned animals. Sex steroids are produced in the bovine placenta and their levels in the blood might be useful as a measure of fetal well-being, as in humans. The objective of this study was to investigate whether changes in peripheral levels of progesterone and estrogens reflect fetal abnormalities. Donor cells for nuclear transfer were obtained from subculture of cumulus cells retrieved from ovarian follicles of a Japanese Black cow. Recipient oocytes were derived from ovaries obtained at an abattoir and matured in vitro. Metaphase II oocytes were enucleated and each fused with a donor cell by DC pulses. Nuclear transferred oocytes were activated and cultured for 7 days. Embryos developed to the blastocyst stage were transferred into the uterine horn ipsilateral to the ovary bearing the CL of 32 multiparous Japanese Black and Holstein crossbred cows at 7 to 8 days after the day of standing estrus (Day 0). Blood was collected from Day 40 until parturition. Progesterone and estrogens in the blood plasma of 6 recipient cows with full-term delivery were measured by RIA. These profiles were compared with each other and with the changes in a cow made pregnant by MOET. Statistical differences were analyzed with repeated measures ANOVA. Parturition was induced on Day 290. Stillborn, dead, or euthanized calves were subjected to necropsy and histopathological analysis. Pregnant cows were 14, 13, and 9 on Days 30, 60, and 90, respectively. Thereafter, 3 aborted around Days 110, 120, and 190. Six cows delivered calves weighing 45.8 ± 2.1 kg (mean ± SEM) on Days 291 to 293. Their birth weights were greater than those of female calves (31.2 ± 0.4 kg, n = 6) produced by MOET in the same breed. Three calves grew normally until weaning. One was stillborn due to dystocia, but no abnormalities were observed except for large offspring syndrome. One was euthanized 2 days after natural delivery due to ananastasia. Thymic and thyroid hypoplasia, left ventricular dysplasia, and pulmonary fibrosis were found by necropsy. Another calf delivered by Caesarean section died of infirmity 2 days later and had thymic and thyroid hypoplasia. Changes in plasma steroid concentrations were consistent with each other and with those in a MOET cow except the last one, in which progesterone levels tended to be higher during the first trimester and estrogens were lower during the last half of gestation. Progesterone levels tended to be lower in cows bearing a healthy calf than in cows bearing a weak calf before parturition. These results imply that peripheral steroid levels may reflect fetal normality, although large offspring syndrome does not affect their concentrations.

Matrotrophy in the cave molly: an unexpected provisioning strategy in an extreme environment

Maternal provisioning of animal embryos may be entirely through yolk deposited in the unfertilized egg (lecithotrophy) or may include post-fertilization nutrient transfer (matrotrophy) in varying degrees. Current theory suggests that the extent of post-fertilization provisioning is resource-dependent, with higher levels of matrotrophy being advantageous in more productive environments. In this study, we investigated post-fertilization embryo provisioning in a livebearing fish, Poecilia mexicana, from two different habitats (a toxic cave and a non-toxic surface habitat) that impose different energetic demands and therefore differ in resources available for reproduction. We predicted that fish in the benign habitat would be more matrotrophic than those from the toxic cave. We used two different techniques for this assay: (1) the matrotrophy-index analysis (MI) for field-collected fish and (2) both MI and radio-tracer assay for laboratory-reared females. According to the interpretation of the matrotrophy index, both populations are purely lecithotrophic, while the radio-tracer assay found females from both populations to actively transfer nutrients to developing embryos at approximately the same rate. Our results suggest that P. mexicana, which was traditionally classified as lecithotrophic, is capable of incipient matrotrophy, and that matrotrophy can contribute to embryo provisioning even in populations from resource-limited environments. Furthermore, the analysis of laboratory-reared animals provides evidence for a genetic component to the large offspring size in cave mollies, which had so far only been described from the field. Specifically, our results suggest matrotrophy occurs in species interpreted as lecithotrophic using the MI approach. Hence, to avoid misclassification, both techniques should ideally be employed in concert, rather than individually. Finally, our results provide further insights into the possible evolutionary pathway from lecithotrophic oviparity to matrotrophic viviparity.

Convergent life-history shifts: toxic environments result in big babies in two clades of poeciliids

The majority of studies on ecological speciation in animals have investigated the divergence caused by biotic factors like divergent food sources or predatory regimes. Here, we examined a system where ecological speciation can clearly be ascribed to abiotic environmental gradients of naturally occurring toxic hydrogen sulfide (H(2)S). In southern Mexico, two genera of livebearing fishes (Poeciliidae: Poecilia and Gambusia) thrive in various watercourses with different concentrations of H(2)S. Previous studies have revealed pronounced genetic differentiation between different locally adapted populations in one species (Poecilia mexicana), pointing towards incipient speciation. In the present study, we examined female reproductive life-history traits in two species pairs: Gambusia sexradiata (from a nonsulfidic and a sulfidic habitat) and Gambusia eurystoma (sulfide-endemic), as well as P. mexicana (nonsulfidic and sulfidic) and Poecilia sulphuraria (sulfide endemic). We found convergent divergence of life-history traits in response to sulfide; most prominently, extremophile poeciliids exhibit drastically increased offspring size coupled with reduced fecundity. Furthermore, within each genus, this trend increased with increasing sulfide concentrations and was most pronounced in the two endemic sulfur-adapted species. We discuss the adaptive significance of large offspring size in toxic environments and propose that divergent life-history evolution may promote further ecological divergence through isolation by adaptation.

Epigenetics: Definition, Mechanisms and Clinical Perspective

A vast array of successive epigenetic modifications ensures the creation of a healthy individual. Crucial epigenetic reprogramming events occur during germ cell development and early embryogenesis in mammals. As highlighted by the large offspring syndrome with in vitro conceived ovine and bovine animals, any disturbance during germ cell development or early embryogenesis has the potential to alter epigenetic reprogramming. Therefore the complete array of human assisted reproductive technology (ART), starting from ovarian hormonal stimulation to embryo uterine transfer, could have a profound impact on the epigenetic state of human in vitro produced individuals. Although some investigators have suggested an increased incidence of epigenetic abnormalities in in vitro conceived children, other researchers have refuted these allegations. To date, multiple reasons can be hypothesized why irrefutable epigenetic alterations as a result of ART have not been demonstrated yet.

Genotypic selection shapes patterns of within‐species diversity in experimental plant populations

Summary 1. An increasing body of evidence suggests that within‐species diversity plays an important role for community and ecosystem functioning, alters complex trophic interactions and affects patterns of species diversity and coexistence. Nonetheless, we lack a good understanding of how genotypic trait variation translates into shifts in the relative abundance of genotypes within populations. 2. In this study, we show that genotypic selection strongly alters dominance relationships among genotypes over a period of 5 years. This resulted in remarkably consistent changes in the proportional representation of genotypes, and in a concomitant decline of diversity and evenness in our experimental populations. 3. High growth rates and the production of large offspring were positively associated with genotypic performance. Vegetative abundances of genotypes translated monotonically into flowering frequencies. 4. Synthesis. We conclude that genotypic selection markedly affects patterns of diversity and consistently alters genotypic abundance and mean trait distributions in plant populations over a relatively short period of time.

Normal development following chromatin transfer correlates with donor cell initial epigenetic state

If the full potential of chromatin transfer (CT) technology is to be realized for both animal production and biomedical applications it is imperative that the efficiency of the reprogramming process be improved, and the potential for deleterious development be eliminated. Generation of the first cloned animals from adult somatic cells demonstrated that development is substantially an epigenetic process (Wilmut I, Schnieke AE, McWhir J, Kind AJ, Campbell KH, 1997. Viable offspring derived from fetal and adult mammalian cells. Nature. 385(6619): 810–813.). In this study, we provide preliminary evidence that the epigenetic state of the donor cell, may be valuable in assessing potential cloning success. We have measured key indicators of cellular epigenetic state in both serially derived cell populations of the same genetic origin, but differing in epigenomic status, and in a distinct cohort of donor cell populations with diverse genetic origins and epigenomic status. Specifically, the relative abundance of particular histone modifications in donor populations prior to manipulation has been correlated with the measurable variance in reprogramming efficiencies observed following CT, as defined by the number of resulting live births and healthy progeny, and the concomitant incidence of deleterious growth measures (notably the appearance of large offspring syndrome (LOS)). Thus, we suggest that the likely outcome and relative success of cloning may be predictable based on the expression of discriminating histone marks present in the donor cell population before CT. This approach may provide the basis of a prognostic signature for the future evaluation and risk assessment of putative donor cells prior to CT, and thus increase future cloning success and alleviate the incidence of abnormal development.

Effects of parity on pelvic size and shape dimorphism in Mus

The pelvis is a sexually dimorphic structure and although the causes of that dimorphism have long been studied, relatively little is known regarding the effects of partuitive events on the magnitude of that dimorphism. Here, we use a sample of Mus musculus domesticus to contrast dimorphism in body length and os coxae size and shape between males and parous and nulliparous females. We also test for correlations between relative litter size (L/M) and relative offspring size (O/M) with body length and os coxae size and shape in parous females. Males had greater body length than nulliparous females but were not different from parous females. Females as a whole had the largest os coxae, with parous females having the largest and males the smallest. Os coxae shape was also significantly different between groups and was most divergent between parous females and males than between nulliparous females and males. Os coxae shape differences between females are associated with differences in body length between females and O/M is correlated with os coxae shape in parous females such that females with the largest offspring have the most divergent shapes along the relative warp one axis. Pelvic shape differences between males and females were consistent with previous findings in other taxa which identify the pubo-ischial complex as the primary region of dimorphism.

Mammal Reproductive Strategies Driven by Offspring Mortality‐Size Relationships

Trade-offs have long been a major theme in life-history theory, but they have been hard to document. We introduce a new method that reveals patterns of divergent trade-offs after adjusting for the pervasive variation in rate of resource allocation to offspring as a function of body size and lifestyle. Results suggest that preweaning vulnerability to predation has been the major factor determining how female placental mammals allocate production between a few large and many small offspring within a litter and between a few large litters and many small ones within a reproductive season. Artiodactyls, perissodactyls, cetaceans, and pinnipeds, which give birth in the open on land or in the sea, produce a few large offspring, at infrequent intervals, because this increases their chances of escaping predation. Insectivores, fissiped carnivores, lagomorphs, and rodents, whose offspring are protected in burrows or nests, produce large litters of small newborns. Primates, bats, sloths, and anteaters, which carry their young from birth until weaning, produce litters of one or a few offspring because of the need to transport and care for them.

Structured coalescent processes from a modified Moran model with large offspring numbers

Structured coalescent processes are derived for the finite island model under a migration mechanism that conserves the subpopulation sizes. The underlying population model is a modified Moran model in which the reproducing individual can have very many offspring with some probability. Convergence to a structured coalescent process results when assuming that migration follows a coalescent timescale which can be much shorter than the usual Wright–Fisher timescale. Three different limit processes are possible depending on the coalescent timescale, two of which allow multiple mergers of ancestral lines. The expected time to most recent common ancestor, and the expected total size of the genealogy, of balanced and unbalanced samples can be very similar, even when migration is low, if the coalescent process allows multiple mergers. The expected total size increases almost linearly with sample size in some cases. The results have implications for inference about genetic population structure.

Using Transgenic Animal Models in Neuroendocrine Research

Transgenic animals are commonly employed to explore the function of individual proteins. Transgenic animal models include the mouse, the zebrafish, and the South African clawed toad Xenopus laevis. In contrast to mice and zebrafish, with Xenopus transgenesis DNA integration is mostly achieved in the one-cell stage. Moreover, Xenopus (as well as zebrafish) eggs are relatively large, the embryos are transparent, a large offspring is generated, and maintenance of the offspring is easy. In our transgenic studies in Xenopus, we focus on the well-characterized neuroendocrine melanotrope cells of the pituitary pars intermedia that are regulated during the process of adaptation of Xenopus to a changing environment. When the animal is placed on a black background, the melanotrope cells produce and process large amounts of the prohormone proopiomelanocortin (POMC). We apply stable melanotrope-specific transgenesis that is achieved by mixing a Xenopus POMC-promoter/transgene construct with sperm nuclei and injecting this mixture into unfertilized eggs. Since in the melanotrope cells the POMC promoter is much more active in black-adapted animals, the level of transgene expression can be manipulated by placing the animal on either a black or a white background. In this paper we review the possibilities of the Xenopus melanotrope-specific transgenic approach. Following a brief overview of the functioning of Xenopus melanotrope cells, stable melanotrope-specific transgenesis is discussed and our transgenic studies on brain-derived neurotrophic factor and secretory pathway components are described as examples of the transgenic approach in a physiological context and close to the in vivo situation.

Investigating Contemporary Evolution via Size-Selective Harvest

[ILLUSTRATION OMITTED] Evolution is commonly taught as a slow process that changes gene frequencies over long time periods. These genotypes are changed through natural selection on phenotypes with the fittest individuals spreading more genes into the environment than less-fit phenotypes. What is now well known to science, but still under- emphasized in textbooks and classroom lessons, is that humans can be one of the strongest selection agents on an organism's phenotype through several different pathways. Humans have perhaps the longest history of affecting organism phenotypes through size-selective harvest, or collection of the largest organism. This activity was designed to supplement traditional evolution units and allow students to discover that evolution can occur in relatively short time-periods and be directed by human action. After completing this activity, students will be able to: * predict the consequences of selective harvesting * understand how evolution and natural selection affect genotypes and phenotypes * design and critique experimental design * collect and analyze data * make conclusions based on research results. Students are likely most familiar with size-selective harvest in trophy hunting of large mammals, such as deer. However, marine fisheries are probably the most common arena for size-selective harvest based upon common industry practices. For centuries, fishermen have used large nets pulled through schools of fish to maximize their catch-per-unit effort. In modern times, large commercial fishing fleets use sophisticated technology, including global positioning satellites and sonar, in conjunction with traditional knowledge about behavior and biology to locate the largest concentrations of fish. Nets are still used to haul in the fish and require fish to be of a certain size to be captured. Even with smaller mesh sizes, some fish will escape, but this is less so in the modern era of commercial fisheries. Fish often mature at a genetically-determined size and/or age, which in turn is structured by environmental conditions. These types of traits are called Life History Traits. Growth is highly correlated with age, and older individuals are usually larger. Generally speaking, the larger the mother, the more offspring she will have with each reproductive event. If she is sufficiently large, her offspring will also be larger. These larger offspring often have higher survival rates than small offspring. Darwinian fitness is related to the proportion of the parents' genes in the next generation; fitness is generically the number of offspring you have that live to reproduce. Larger female fish have higher fitness than smaller female fish. Darwin (2004) used changes seen with selective breeding of animals as evidence of natural selection. Selective breeding is common in today's commercial agriculture and food animal production, but is seldom discussed with relevancy to evolutionary processes. Humans can also inadvertently affect the evolution of species. Using the marine fisheries example, fish that escape fishing nets to reproduce are often smaller and thus pass on genes that allow maturity at smaller sizes. If the harvest is strong enough, or repeated over many generations, this selection can result in fish being smaller at maturity (Figure 1). This has fitness consequences and reduces biomass in commercial harvests (Conover & Munch, 2002; Hutchings, 2005). Contemporary evolution is defined as heritable trait evolution observed in contemporary time (i.e., less then a few hundred years) (Stockwell et al., 2003). [FIGURE 1 OMITTED] A simple laboratory exercise using inexpensive and nonperishable materials can demonstrate contemporary evolution. The following activity will allow students to discover how humans can interfere with the natural selection of organisms and do so in relatively little time. …

Allelic switching of the imprinted IGF2R gene in cloned bovine fetuses and calves

Cloned animals often suffer from loss of development to term and abnormalities, typically classified under the umbrella term of Large Offspring Syndrome (LOS). Cattle are an interesting species to study because of the relatively greater success rate of nuclear transfer in this species compared with all species cloned to date. The imprinted insulin-like growth factor receptor ( IGF2R; mannose-6-phosphate) gene was chosen to investigate aspects of fetal growth and development in cloned cattle in the present study. IGF2R gene expression patterns in identical genetic clones of several age groups were assessed in day 25, day 45, and day 75 fetuses as well as spontaneously aborted fetuses, calves that died shortly after birth and healthy cloned calves using single stranded conformational polymorphism gel electrophoresis. A variable pattern of IGF2R allelic expression in major organs such as the brain, cotyledon, heart, liver, lung, spleen, kidney and intercotyledon was observed using a G/A transition in the 3’UTR of IGF2R. IGF2R gene expression was also assessed by real time RT-PCR and found to be highly variable among the clone groups. Proper IGF2R gene expression is necessary for survival to term, but is most likely not a cause of early fetal lethality or an indicator of postnatal fitness. Contrary to previous reports of the transmission of imprinting patterns from somatic donor cells to cloned animals within organs in the same cloned animal the paternal allele of IGF2R can be imprinted in one tissue while the maternal allele is imprinted in another tissue. This observation has never been reported in any species in which imprinting has been studied.

Women’s health—what’s new world wide

Women’s health—what’s new world wide

32 CHARACTERISTICS OF BOVINE CLONE OFFSPRING (F1): COMPARISON WITH CLONES

Information on clone offspring (F1) remains limited, especially in species with a long intergeneration interval such as cattle. In the present study, we report data on calves born from cloned females at the INRA experimental unit. Characteristics of these offspring are compared to that of clones or AI control animals born and raised in the same farm. Currently, 39 F1 have been obtained after AI of 64 cloned heifers from 3 different Holstein genotypes, with semen from 6 different Holstein sires. All calves were naturally delivered at term with a mean easy calving score of 1.58 and a normal pregnancy length of 282 days. There were 20 males and 19 females, resulting in a normal male/female sex ratio of 51/49%. Male calves were euthanized during the first 3 months after birth, and a group of 18 Holstein single-born female F1 were further considered and compared to female AI controls (n = 68) or clones (n = 50) all Holsteins. The mean birth weight of the female F1 was 40.28 ± 3.32 kg (range 35–44), not significantly different from that of AI controls: 40.57 ± 5.55 kg and significantly lower (P < 0.05) and less variable than that of the clones: 49.27 ± 10.93 kg. No case of large offspring syndrome was observed in the F1 group. Postnatal survival in the clone offspring group was normal as 17/18 of the females (94.4%) remained alive and healthy by 6 months of age. This was comparable to survival in the AI control group: 64/68 (94.1%) and higher than for clones at the same age: 35/50 (70.0%). Animals were weighed every month. There were no statistical differences between the clone offspring group and the AI controls raised in the same conditions, with a total body weight of 188.25 ± 15.17 kg v. 185.30 ± 18.94 kg by 6 months of age and a daily weight gain of 0.820 and 0.803 kg day–1, respectively. The oldest F1 heifers are now mature and their reproductive function is being evaluated. Some animals are already used as recipients in an embryo transfer program. As enlarged internal organs had been reported for clones, we analyzed the data from euthanasia of 20 male F1 by weighing the different organs after postmortem examination. The mean ± SD relative organ weight (% of body weight) was compared to that of somatic clones (n = 18) and a limited number of AI control calves (n = 5) previously autopsied in our institute at similar ages. Figures were very similar between F1 and controls, but significant differences (t-test, P < 0.05) were observed between F1 and clones. F1 had a smaller (and less variable) liver ratio than clones (0.022 ± 0.004 v. 0.028 ± 0.009, respectively), a smaller kidney ratio (0.005 ± 0.001 v. 0.007 ± 0.002), and a lower heart ratio (0.008 ± 0.002 v. 0.010 ± 0.003) than clones. No significant difference was observed between groups for thymus/body ratio. None of the F1 calves presented any of the pathologies observed in clones. Altogether, these results indicate that clone offspring F1 develop similarly to AI controls and differ from the clones themselves.

HEMATOLOGIC AND SERUM BIOCHEMICAL PROFILE OF THE NORTHERN ELEPHANT SEAL (MIROUNGA ANGUSTIROSTRIS): VARIATION WITH AGE, SEX, AND SEASON

The foraging success, and thus the survival and reproductive success, of deep-diving pinnipeds such as the northern elephant seal, Mirounga angustirostris, depends on the ability to withstand repetitive breath-hold dives. Health parameters can be incorporated as potential explanatory variables for differences observed in diving and migratory performance of individual seals. Furthermore, biomedical samples from apparently healthy individuals can provide valuable baseline data for evaluating effects of natural or anthropogenic threats to individuals and populations. We evaluated 42 blood parameters in 134 northern elephant seals during the breeding and molting seasons (1992-1999) to test for age, sex, and seasonal differences and to develop reference ranges. Adult males sampled during the breeding season differed from all other adult groups for a suite of parameters often associated with inflammation, infection, or other stressors: lower hematocrit, higher white blood cell count, higher band neutrophils, higher neutrophil count, lower albumin, and lower serum iron. Adult females during the breeding season differed from all other adult categories for two parameters (lower platelet counts, lower alanine aminotransferase activity). Molting males had higher blood urea nitrogen than all other classes; creatinine did not differ between breeding and molting adult males, but was higher in males than in females in both seasons. We found significant differences among age classes for 24 of 42 parameters measured, including higher levels of triglycerides, total protein, calcium, and iron in pups than we found in juveniles or adults. Unlike other mammals which undergo substantial decreases in energy expenditure during prolonged fasting (e.g., hibernation), northern elephant seals defend territories, give birth and suckle large offspring, mate, and molt during their bi-annual fasts. Nonetheless, many studies have described physiologic homeostasis during fasting in elephant seals. The genus Mirounga is superbly adapted to going without feeding for extended periods, and this is reflected in our hematologic and serum biochemical profiles.

Consequences of In Vitro Culture Conditions on Embryo Development and Quality

Despite major efforts directed at improving the yield of blastocysts from immature oocytes in vitro, the quality of such blastocysts continually lags behind that of blastocysts produced in vivo. These differences are manifested at the level of morphology, metabolism, gene expression and cryotolerance, and may have a knock-on effect further along the developmental axis. Evidence suggesting that in vitro culture conditions, while capable of producing blastocysts in relatively high numbers, are far from optimal with deficiencies being manifested in terms of abnormally large offspring. It is clear nowadays that modification of the post-fertilization culture environment in vitro can improve blastocyst quality to some extent.