Jaw muscle development as evidence for embryonic repatterning in direct-developing frogs.

The Puerto Rican direct-developing frog Eleutherodactylus coqui (Leptodactylidae) displays a novel mode of jaw muscle development for anuran amphibians. Unlike metamorphosing species, several larval-specific features never form in E. coqui; embryonic muscle primordia initially assume an abbreviated, mid-metamorphic configuration that is soon remodelled to form the adult morphology before hatching. Also lacking are both the distinct population of larval myofibres and the conspicuous, larval-to-adult myofibre turnover that are characteristic of muscle development in metamorphosing species. These modifications are part of a comprehensive alteration in embryonic cranial patterning that has accompanied life history evolution in this highly speciose lineage. Embryonic ''repatterning'' in Eleutherodactylus may reflect underlying developmental mechanisms that mediate the integrated evolution of complex structures. Such mechanisms may also facilitate, in organisms with a primitively complex life cycle, the evolutionary dissociation of embryonic, larval, and adult features.     

THE GROWTH AND DIFFERENTIATION OF CULTURED BARLEY EMBRYOS

Norstog , Knut . (Wittenberg U., Springfield, Ohio.) The growth and differentiation of cultured barley embryos. Amer. Jour. Bot. 48(10): 876–884. Illus. 1961.—Cultures of excised embryos of barley, Hordeum vulgare L., were made on a number of different media. Growth on White's medium was promoted by adding coconut milk. In the absence of coconut milk, amino acids did not promote growth and differentiation. Embryos as small as 60 μ were successfully grown in vitro. Smaller embryos had the capacity to initiate root and shoot primordia but did not possess the ability to form such embryonic organs as the scutellum and epiblast. Proembryos developed shoots and roots only after a period of irregular growth in which unorganized masses of cells were formed. Multiple centers of shoot initiation were observed in such embryos. The results of the study suggest that, in barley at least, embryonic form may result from an interaction between the embryo and nutritional, spatial and other factors within the ovule.     

An evolutionary correlate of genome size change in plethodontid salamanders

Variation in the amount of nuclear DNA, the C-value, does not correlate with differences in morphological complexity. There are two classes of explanations for this observation, which is known as the ''C-value paradox''. The quantity of DNA may serve a ''nucleotypic'' function that is positively selected. Alternatively, large genomes may consist of junk DNA, which increases until it negatively affects fitness. Attempts to resolve the C-value paradox focus on the link between genome size and fitness. This link is usually sought in life history traits, particularly developmental rates. I examined the relationship among two life history traits, egg size and embryonic developmental time and genome size, in 15 species of plethodontid salamanders. Surprisingly, there is no correlation between egg size and developmental time, a relationship included in models of life history evolution. However, genome size is positively correlated with embryonic developmental time, a result that is robust with respect to many sources of variation in the data. Without information on the targets of natural selection it is not possible with these data to distinguish between nucleotypic and junk DNA explanations for the C-value paradox.     

Temperature sensitivity of muscle and neuron differentiation in embryonic cell cultures from the Drosophila mutant, shibire.

The sex-linked temperature-sensitive mutation, shibire^ts1, which causes, at the restrictive temperature, adult paralysis and pleiotropic morphological defects in embryonic, larval, and pupal development, has been shown to exhibit temperature-sensitive inhibition of differentiation in embryonic cultures in vitro. When shi cultures were incubated at 30°C for 24 hr, both muscle and neuron differentiation were inhibited more than 90% compared to control shi cultures incubated at 20°C. Heat shift experiments showed that the temperature-sensitive periods for neuron and muscle differentiation occurred at 11 to 18 and 14 to 16 hr, respectively, where zero time was the initiation of gastrulation in donor embryos. Short heat pulses (4 and 8 hr) which extended into the temperature-sensitive period resulted in moderate inhibition of differentiation; greater inhibition occurred as the duration of the pulses increased. In contrast, heating wild-type Oregon-R cultures at 30°C for 24 hr did not inhibit muscle cell differentiation and inhibited neuron differentiation relatively little. The temperature-sensitive period in shibire for muscle differentiation occurred well after myoblast division, during the period of myocyte elongation, aggregation, and fusion, whereas that for neuron differentiation took place during a period of enzyme synthesis (acetylcholinesterase and choline acetyltransferase) and axon elongation. Thus, the shi temperature-sensitive gene product affects at least two different cell types, in vitro, at different times during differentiation.     

Regulators of Trypanosoma brucei Cell Cycle Progression and Differentiation Identified Using a Kinome-Wide RNAi Screen

The African trypanosome, Trypanosoma brucei, maintains an integral link between cell cycle regulation and differentiation during its intricate life cycle. Whilst extensive changes in phosphorylation have been documented between the mammalian bloodstream form and the insect procyclic form, relatively little is known about the parasite''s protein kinases (PKs) involved in the control of cellular proliferation and differentiation. To address this, a T. brucei kinome-wide RNAi cell line library was generated, allowing independent inducible knockdown of each of the parasite''s 190 predicted protein kinases. Screening of this library using a cell viability assay identified ≥42 PKs that are required for normal bloodstream form proliferation in culture. A secondary screen identified 24 PKs whose RNAi-mediated depletion resulted in a variety of cell cycle defects including in G1/S, kinetoplast replication/segregation, mitosis and cytokinesis, 15 of which are novel cell cycle regulators. A further screen identified for the first time two PKs, named repressor of differentiation kinase (RDK1 and RDK2), depletion of which promoted bloodstream to procyclic form differentiation. RDK1 is a membrane-associated STE11-like PK, whilst RDK2 is a NEK PK that is essential for parasite proliferation. RDK1 acts in conjunction with the PTP1/PIP39 phosphatase cascade to block uncontrolled bloodstream to procyclic form differentiation, whilst RDK2 is a PK whose depletion efficiently induces differentiation in the absence of known triggers. Thus, the RNAi kinome library provides a valuable asset for functional analysis of cell signalling pathways in African trypanosomes as well as drug target identification and validation.     

Embryological development of Salvator merianae (Squamata: Teiidae)

Here we describe the embryonic development of Salvator merianae external morphologic features, as based on observation of 28 embryos across different days of incubation at 31 ± 0.5°C. Observed developmental stages were grouped and classified into the early, middle, and late periods. The early period (Stages 3–11) is distinguished by the origin of the encephalic vesicles, sensory placodes, pharyngeal arches, and degree of body flexion and rotation. The medium period (Stages 8–15) is distinguished by limb differentiation and by cranium‐facial characteristics. The late period (Stages 15–18) is determined by scale patterns, pigmentation, and embryo growth.     

Influence of Environment and Mitochondrial Heritage on the Ecological Characteristics of Fish in a Hybrid Zone

Background

Ecological characteristics (growth, morphology, reproduction) arise from the interaction between environmental factors and genetics. Genetic analysis of individuals'' life history traits might be used to improve our understanding of mechanisms that form and maintain a hybrid zone.

Methodology/Principal Findings

A fish hybrid zone was used to characterize the process of natural selection. Data were collected during two reproductive periods (2001 and 2002) and 1117 individuals (nase, Chondrostama nasus nasus, sofie C. toxostoma toxostoma and hybrids) were sampled. Reproductive dates of the two parental species overlapped at sympatric sites. The nase had an earlier reproductive period than the sofie; males had longer reproductive periods for both species. Hybridisation between female nase and male sofie was the most likely. Hybrids had a reproductive period similar to the inherited parental mitochondrial type. Growth and reproductive information from different environments has been synthesised following a bayesian approach of the von Bertalanffy model. Hybrid life history traits appear to link with maternal heritage. Hybrid size from the age of two and size at first maturity appeared to be closer to the size of the maternal origin species (nase or sofie).Median growth rates for hybrids were similar and intermediate between those of the parental species. We observed variable life history traits for hybrids and pure forms in the different parts of the hybrid zone. Geometrical analysis of the hybrid fish shape gave evidence of two main morphologies with a link to maternal heritage.

Conclusions/Significance

Selective mating seemed to be the underlying process which, with mitochondrial heritage, could explain the evolution of the studied hybrid zone. More generally, we showed the importance of studies on hybrid zones and specifically the study of individuals'' ecological characteristics, to improve our understanding of speciation.     

An analysis of temporal homogenisation and differentiation in Central European village floras

Agriculture and urbanisation shape biodiversity through extirpation of species and facilitation of species introductions. These processes include changes in the functional composition of species assemblages and can result in taxonomic and functional homogenisation. Especially the spread of non-native species has been discussed as a driver of homogenisation. However, no consensus has been reached so far; instead, both homogenisation and differentiation by non-native species have been shown. This inconsistency can partly be attributed to the lack of temporal data: Most homogenisation studies rely on purely spatial analyses, while homogenisation develops over time. We studied vascular plant species occurrences in 59 villages in the West of Germany in the 1980s and twenty years later. Within this period, the villages experienced changes in agriculture and trends towards urbanisation. We asked whether the villages’ floras became more similar to each other within the study period, and whether this process differed between selected plant groups. We based plant groups on leaf traits, life form, species native/non-native status, and mode of introduction. This enabled us to discuss changes in the flora in the context of land-use changes. We used Simpson's index of dissimilarity as a measure of β-diversity among villages and calculated species turnover and homogenisation in time. Overall, village floras became more similar to each other within the study period. However, neophytes became less similar to each other across villages. Turnover between sampling periods was largest for species promoted by horticulture and for species with helomorphic leaves (suggesting an effect of habitat loss on turnover). Neophytes will likely continue to differentiate floras on regional scales due to on-going and various introductions.     

Culture of blastomeres from in vitro-matured,fertilized, and cultured bovine embryos

A culture system was devised to study the differentiation of bovine blastomeres. Blastomeres (2–13 per well) from embryos produced by in vitro maturation, fertilization, and culture of oocytes obtained from slaughterhouse ovaries were cultured for 10 days in 24-well culture plates on feeder layers in blastomere culture medium (BCM: equal parts tissue culture medium 199 and low-glucose Dulbecco's modified Eagle's medium with 10% fetal bovine serum). Ovine embryonic fibroblasts and STO cells were superior to bovine and mouse embryonic fibroblasts as mitotically inactivated feeder cells. Over five studies in which four blastomeres from an embryo were added to each culture well, an average of one colony per well formed from the blastomeres. The colonies continued to grow throughout the culture period, and most colonies resembled trophectoderm in their cellular characteristics, although some cultures contained a mixture of trophectoderm and endoderm. When the number of blastomeres cultured in each well was varied from 2–8, the number of colonies formed was proportional to the number of blastomeres added. Blastomeres from day 5 and day 6 embryos produced fewer colonies than did those from day 4 embryos, perhaps as a result of differentiation and tighter blastomere adhesion resulting in damage during their separation. The absence of serum did not alter the number of colonies formed. A number of growth factors, including LIF, OM, PDGFα, and FGF4, had no effect on the number of colonies, the size of colonies, or their alkaline phosphatase staining score beyond that provided by the feeder layer or serum when present. Blastomeres did not form colonies in the absence of feeder layers. Mol. Reprod. Dev. 48:238–245, 1997. © 1997 Wiley-Liss, Inc.     

Fast myosin heavy chain expression during the early and late embryonic stages of chicken skeletal muscle development

The development of embryonic skeletal muscles in the chick can be divided into two periods of fiber specialization--an early one during which the different muscles of the limb are formed and an initial round of fiber specialization occurs and a late or fetal period during which there is extensive growth of this previously established fiber pattern. This latter period of growth is dependent on the establishment and maintenance of functional neuromuscular contacts. As has been described for other developmental stages, we show here that there are different embryonic fast skeletal muscle myosin heavy chain (MHC) isoforms expressed during the different embryonic periods of muscle growth. The identification of these isoforms was based on differences in their reactivity with various fast MHC monoclonal antibodies and on their different peptide banding patterns. The in ovo accumulation of the late embryonic MHC isoform pattern was similar to the time course of the previously described changes in alpha-actin and troponin T isotype switching during embryogenesis. The appearances of the late embryonic isoforms were blocked by chronic treatment with the neuromuscular blocking agent, d-tubocurarine, and cell cultures of embryonic chicken skeletal muscle which differentiated in the absence of motorneurons expressed little of the late embryonic isoform, indicating that the expression of the late embryonic isoform was dependent on functional nerve-muscle interactions. These different embryonic fast MHC isoforms provide important markers for monitoring the progression of muscle through its embryonic stages and its interaction with motorneurons.     

Animal survival strategies in Neoproterozoic ice worlds

The timing of the first appearance of animals is of crucial importance for understanding the evolution of life on Earth. Although the fossil record places the earliest metazoans at 572–602 Ma, molecular clock studies suggest a far earlier origination, as far back as ~850 Ma. The difference in these dates would place the rise of animal life into a time period punctuated by multiple colossal, potentially global, glacial events. Although the two schools of thought debate the limitations of each other's methods, little time has been dedicated to how animal life might have survived if it did arise before or during these global glacial periods. The history of recent polar biota shows that organisms have found ways of persisting on and around the ice of the Antarctic continent throughout the Last Glacial Maximum (33–14 Ka), with some endemic species present before the breakup of Gondwana (180–23 Ma). Here we discuss the survival strategies and habitats of modern polar marine organisms in environments analogous to those that could have existed during Neoproterozoic glaciations. We discuss how, despite the apparent harshness of many ice covered, sub-zero, Antarctic marine habitats, animal life thrives on, in and under the ice. Ice dominated systems and processes make some local environments more habitable through water circulation, oxygenation, terrigenous nutrient input and novel habitats. We consider how the physical conditions of Neoproterozoic glaciations would likely have dramatically impacted conditions for potential life in the shallows and erased any possible fossil evidence from the continental shelves. The recent glacial cycle has driven the evolution of Antarctica's unique fauna by acting as a “diversity pump,” and the same could be true for the late Proterozoic and the evolution of animal life on Earth, and the existence of life elsewhere in the universe on icy worlds or moons.     

基于生长测量仪监测的亚热带地区马尾松多时间尺度径向变化及其与环境因子的关系

树干径向变化的多尺度研究提供了树木生长及其和环境因子关系的详细信息,有助于准确评估全球气候变化背景下森林生态系统碳汇变异。以往树干径向变化研究主要集中在温带和热带地区,且大多数研究方法基于时间分辨率较粗的树木年轮法,然而缺少亚热带地区高时间分辨率树干径向变化的研究。利用树干径向变化记录仪连续监测亚热带地区马尾松13个月的树干径向变化动态,探索不同时间尺度树干径向变化规律及与环境因子的关系。结果表明：（1）在日尺度,马尾松径向变化模式为白天收缩夜晚膨胀,秋冬季节夜晚膨胀没有春夏季明显。（2）在季节尺度,马尾松树干径向变化可分为4个时期,其中3-8月是主要生长月份,4月是累计生长量最大的月份。（3）在日尺度上,相对湿度和饱和水汽压亏缺是调节马尾松径向变化主要环境因素;在季节尺度上,土壤温度对树干径向变化的影响大于空气温度,降水量与相对湿度等水分因素对树干径向生长的促进作用在生长季中后期更为明显。研究结果有助于深入理解亚热带季风气候区树干径向变化及其对环境变化的响应,为气候变化背景下亚热带地区的植树造林设计和森林可持续管理提供依据。     

Effects of temperature during early life history on embryonic and larval development and growth in haddock

The effect of incubation temperature (2, 4, 6, 8 and 10° C) on haddock Melanogrammus aeglefinus development and growth during the embryonic period and in subsequent ontogeny in a common post‐hatch thermal environment (6° C) was investigated. Hatching times were inversely proportional to incubation temperature and ranged from 20·3 days at 2° C to 9·1 days at 10° C. Growth rates were directly proportional to incubation temperature during both the embryonic and larval periods. There was a significant decline in growth rates following hatch in all temperature groups. Compared to the endogenously feeding embryos, growth rates in the exogenous period declined by 4·4‐fold at 4° C to 3·9‐fold at 8° C, indicative of the demarcation between the endogenous and exogenous feeding periods. Yolk utilization varied from 17 days at 2° C to 6 days at 10° C and followed a three‐stage sigmoidal pattern with the initial lag period inversely proportional to incubation temperature. Time to 50% yolk depletion varied inversely with temperature but occurred 1–1·5 days post‐hatch at all temperatures. Additionally, the period between 10 and 90% yolk depletion also decreased with increased temperature. Overall developmental rate was sequential with and directly proportional (2·3‐fold increase) to incubation temperature while the time spent in each developmental stage was inversely proportional to temperature. Larger embryos tended to be produced at lower temperatures but this pattern reversed following hatch, as larvae from higher temperature groups grew more rapidly than those from other temperature groups. Larvae from all temperatures achieved a similar length (c.total length 4·5 mm) upon complete yolk absorption. The study demonstrated the significant impact that temperature has upon developmental and growth rates in both endogenous and exogenous feeding periods. It also illustrated that temperature changes during embryogenesis had significant and persistent effects on growth in subsequent ontogeny.     

Growth of asthmatic children during treatment with budesonide: a double blind trial.

OBJECTIVE--To determine whether the inhaled glucocorticosteroid budesonide has any adverse effect on short term linear growth in children with mild asthma. SETTING--Outpatient clinic in secondary referral centre. PATIENTS--15 children aged 6-13 years with normal statural growth velocity during the previous year, no signs of puberty, and no use of systemic or topical steroids in the two months before the study. DESIGN OF INTERVENTIONS--Double blind, randomised crossover trial with two active periods in which budesonide was given in divided daily doses of 200 micrograms and 800 micrograms. During run in and two washout periods placebo was given. After the second washout period the children received open treatment with 400 micrograms budesonide daily. All periods were of 18 days'' duration. MAIN OUTCOME MEASURE--Growth of the lower leg as measured twice a week by knemometry. RESULTS--Mean growth velocity of the lower leg was 0.63 mm/week during run in and during washout 0.64 mm/week. Budesonide treatment was associated with a significant dose related reduction of growth velocity: the mean reduction in growth velocity during treatment was 0.11 (95% confidence interval -0.15 0.36 (0.13 to 0.59) mm/week with 800 micrograms budesonide (p less than 0.05; Page''s test). During treatment with 400 micrograms budesonide a reduction of 0.17 (-0.10 to 0.45) mm/week was found. CONCLUSIONS--Treatment with inhaled budesonide is associated with a dose related suppression of short term linear growth in children with mild asthma.     

Generation of Human Induced Pluripotent Stem (iPS) Cells in Serum- and Feeder-Free Defined Culture and TGF-β1 Regulation of Pluripotency

Human Embryonic Stem cells (hESCs) and human induced Pluripotent Stem cells (hiPSCs) are commonly maintained on inactivated mouse embryonic fibroblast as feeder cells in medium supplemented with FBS or proprietary replacements. Use of culture medium containing undefined or unknown components has limited the development of applications for pluripotent cells because of the relative lack of knowledge regarding cell responses to differentiating growth factors. In addition, there is no consensus as to the optimal formulation, or the nature of the cytokine requirements of the cells to promote their self-renewal and inhibit their differentiation. In this study, we successfully generated hiPSCs from human dental pulp cells (DPCs) using Yamanaka''s factors (Oct3/4, Sox2, Klf4, and c-Myc) with retroviral vectors in serum- and feeder-free defined culture conditions. These hiPSCs retained the property of self-renewal as evaluated by the expression of self-renewal marker genes and proteins, morphology, cell growth rates, and pluripotency evaluated by differentiation into derivatives of all three primary germ layers in vitro and in vivo. In this study, we found that TGF-β1 increased the expression levels of pluripotency markers in a dose-dependent manner. However, increasing doses of TGF-β1 suppressed the growth rate of hiPSCs cultured under the defined conditions. Furthermore, over short time periods the hiPSCs cultured in hESF9 or hESF9T exhibited similar morphology, but hiPSCs maintained in hESF9 could not survive beyond 30 passages. This result clearly confirmed that hiPSCs cultured in hESF9 medium absolutely required TGF-β1 to maintain pluripotency. This simple serum-free adherent monoculture system will allow us to elucidate the cell responses to growth factors under defined conditions and can eliminate the risk might be brought by undefined pathogens.     

Comparative analysis of the interaction between habitat and growth form in diatoms

We characterized the evolutionary history of growth form (solitary–colonial) and its interaction with species'' habitat (planktonic–benthic) across a multi-gene phylogeny encompassing a broad sample of the order-level diversity of diatoms. We treated these characters broadly, modeling the evolution of aggregation of cells into a colony irrespective of the way aggregation is achieved, and relating the growth form to a broad concept of niche location: in the plankton or benthos. The results showed that habitat shifts are rare implying conservatism in niche location at the level of large clades. On the other hand, the evolutionary history of growth form is more dynamic with evolutionary rates that vary across the tree. Analyses of a possible interaction revealed that shifts in growth form are independent of habitat and that traversing between habitats does not hinge upon species'' growth form. Our findings help to fill a gap in the understanding of diatom niche and growth form macroevolution and contribute toward a platform for the comparative study of the mechanisms underlying diatom species and functional diversity.     

The origins of larval forms: what the data indicate,and what they don't

What is a larva, if it is not what survives of an ancestor's adult, compressed into a transient pre‐reproductive phase, as suggested by Haeckel's largely disreputed model of evolution by recapitulation? A recently published article hypothesizes that larva and adult of holometabolous insects are developmental expressions of two different genomes coexisting in the same animal as a result of an ancient hybridization event between an onychophoran and a primitive insect with eventless post‐embryonic development. More likely, however, larvae originated from late embryonic or early post‐embryonic stages of ancestors with direct development. Evolutionary novelties would thus be intercalary rather than terminal, with respect to the ancestor's ontogenetic schedule. This scenario, supported by current research on holometabolous insects and marine invertebrates with complex life cycles, offers a serious alternative to the traditional scenario (‘what is early in ontogeny is also early in phylogeny’) underlying the current perception of the evolution of genetic regulatory networks.     

Life history strategies of annual killifish <Emphasis Type="Italic">Millerichthys robustus</Emphasis> (Cyprinodontiformes:Cynolebiidae) in a seasonally ephemeral water body in Veracruz,México

The annual life cycle of Millerichthys robustus is described from an ephemeral pool in Veracruz, Mexico. The state of the pool can be divided into three periods: a flood period lasting from September to March when the pool was filled with water, a drought period from April to June when the pool was dry, and a humid period in July and August when the pool was intermittently filled. Soil substrate was examined in each of these three periods (flood, drought and humid), embryos were found, and the stage of embryonic diapause was determined. During the flood period embryos were in diapause I; during the drought period in diapause II; and during the humid period mainly in diapause III with a small subset in diapause II (an example of a bet-hedging strategy). Two hatching periods (separated by two weeks) were documented during the beginning of the flood period. Fish growth was analyzed in both males and females, with females showing an overall slower growth rate and smaller adult size. In females, ovarian maturity was characterized histologically to understand the reproductive cycle. The onset of sexual maturity began during the third week after hatching (21 days) with the presence of secondary sexual characteristics in females and the beginning of Secondary Growth Stage in some ovarian follicles. All stages of oogenesis, postovulatory follicles and ovulated oocytes were observed from the fourth week post hatching (28 days) until death. M. robustus appears to exhibit similar patterns of embryonic diapause compared to other annual killifish living in seasonal water bodies closer to the equator. This study characterizes (for the first time) the adaptations and life cycle of M. robustus. This information could be useful to evaluate the potential risk of these populations and, if necessary, to develop plans for their conservation.