A high-efficiency HeLa cell nuclear transcription extract.

A HeLa cell nuclear transcription extract that is approximately 20 times more efficient than standard HeLa cell transcription extracts was developed. Transcription of the strong adenovirus II major late promoter by this extract results in the synthesis of 1.5-4 molecules of product RNA per molecule of template, indicating that the extract is capable of multiple rounds of initiation. Standard HeLa cell nuclear extracts transcribe closed circular and linear adenovirus major late promoter templates with equal efficiency. In contrast, the new extract exhibits an increase of approximately twofold on transcription of a closed circular, as opposed to a linear, major late promoter template.

Twofold cost of reproduction: an increase in parental effort leads to higher malarial parasitaemia and to a decrease in resistance to oxidative stress.

Parental effort is usually associated with high metabolism that could lead to an increase in the production of reactive oxidative species giving rise to oxidative stress. Since many antioxidants involved in the resistance to oxidative stress can also enhance immune function, an increase in parental effort may diminish the level of antioxidants otherwise involved in parasite resistance. In the present study, we performed brood size manipulation in a population of great tits (Parus major) to create different levels of parental effort. We measured resistance to oxidative stress and used a newly developed quantitative PCR assay to quantify malarial parasitaemia. We found that males with an enlarged brood had significantly higher level of malarial parasites and lower red blood cell resistance to free radicals than males rearing control and reduced broods. Brood size manipulation did not affect female parasitaemia, although females with an enlarged brood had lower red blood cell resistance than females with control and reduced broods. However, for both sexes, there was no relationship between the level of parasitaemia and resistance to oxidative stress, suggesting a twofold cost of reproduction. Our results thus suggest the presence of two proximate and independent mechanisms for the well-documented trade-off between current reproductive effort and parental survival.

When several queens reign: how polygyny affects social life in ants

Astract: The aim of this thesis was to investigate how the presence of multiple queens (polygyny) affects social organization in colonies of the ant Formica exsecta. This is important because polygyny results in reduced relatedness among colony members and therefore reflects a potential paradox for altruistic cooperation being explained by inclusive fitness theory. The reason for this is that workers in polygynous colonies rear no longer only their siblings (high inclusive fitness gain) but also more distantly ox even unrelated brood (low or no inclusive fitness gain). All research projects conducted in this thesis are novel and significant contributions to the understanding of the social evolution of insect societies. We used a mixture of experimental and observational methodologies in laboratory and field colonies of F. exsecta to examine four important aspects of social life that are impacted by polygyny. First, we investigated the influence of queen number on colony sex allocation and found that the number of queens present in a colony significantly affects colony sex ratio investment. The data were consistent with the queen-replenishment hypothesis, which is based on the observation that newly mated queens are often recruited back to their parental nest. According to this theory, colonies containing many queens should only produce males due to local resource competition (i.e. related queens compete for common resources), whereas colonies hosting few queens benefit most from producing new queens to ensure colony survival. Second, we examined how reproduction is partitioned among nestmate queens. We detected a novel pattern of reproductive partitioning whereby a high proportion of queens were completely specialized in the production of only a subset of offspring classes produced within a colony, which might translate into great differences in reproductive success between queens. Third, we could demonstrate that F. exsecta workers indiscriminately reared highly related and unrelated brood although such nepotistic behaviour (preferential rearing of relatives) would be predicted by inclusive fitness theory. The absence of nepotism is probably best explained by its negative effects on overall colony efficiency. Finally, we conducted a detailed population genetic analysis, which revealed that the genetic population structure is different for queens and workers. Our data were best explained with queens forming family-based groups (multicolonial population structure), whereas workers from several nests seemed to be grouped into larger unites (unicolonial population structure) with workers moving freely between neighbouring nests. Altogether, the presented work significantly increased our understanding of the complex organization of polygynous social insect colonies and shows how an important life history trait such as queen number affects social organization at various levels. Résumé: Le but de cette thèse était d'étudier comment la présence de plusieurs reines par colonie (polygynie) influence la vie sociale chez la fourmi Formica exsecta. Ce sujet est important parce que la polygynie chez les insectes sociaux présente un passible paradoxe au niveau de la théorie du "fitness inclusive". Ce paradoxe est basé sur le fait que les ouvrières n'élèvent plus uniquement leurs frères et soeurs (gain de "fitness inclusive" maximale), mais également des individus moins ou pas du tout apparentés (gain de "fitness inclusive" réduit ou absent). Tous les projets de recherche présentés au cours de cette thèse apportent une meilleure compréhension et connaissance au niveau de l'organisation des colonies chez les insectes sociaux. Nous avons employé des méthodes d'observation et de laboratoire afin de mettre en évidence des aspects importants de la vie sociale chez les fourmis influencés par la polygynie. Quatre aspects ont été caractérisés : (1) l'influence du nombre de reines sur le sexe ratio produit par la colonie. Nous avons démontré que les colonies contenant beaucoup de reines produisaient rarement des reines tandis que les colonies contenant peu de reines souvent investissaient beaucoup de ressources dans la production des reines. Ces résultats sont en accord avec la "queen-replenishment hypothesis" qui est basé sur l'observation que les nouvelles reines sont recrutées dans la colonie où elles étaient nées. Cette hypothèse postule que la production des reines est défavorable dans les colonies contenant beaucoup de reines, parce que ces reines apparentées, rentrent en compétition pour des ressources communes. Au contraire, la production des reines est favorable dans des colonies contenant peu de reines afin d'assurer la survie de la colonie ; (2) comment les reines dans une colonie répartissent leur reproduction. Nous avons mis en évidence un nouveau pattern de cette répartition où une grande proportion de reines est complètement spécialisée dans la production d'un seul type de couvain ce qui probablement aboutit à des différences significatives entre reines dans le succès reproducteur ; (3) la capacité des ouvrières à discriminer un couvain de soeur d'un couvain non apparenté. Les résultats ont montré que les ouvrières ne font pas de discrimination entre le couvain de soeur et le couvain non apparenté ce qui n'est pas en accord avec la théorie de la "fitness inclusive". Cette absence de discrimination est probablement due à des effets négatifs comme par exemple la diminution de la production du couvain; (4) la structure génétique d'une population de F. exsecta. Nous avons mis en évidence que la structure génétique entre des groupes de reines est significativement différente de la structure génétique entre des groupes d'ouvrières. Les données suggèrent que les reines forment des groupes basés sur une structure familiale tandis que les ouvrières sont groupées dans des unités plus grandes.

Asexual reproduction in introduced and native populations of the ant Cerapachys biroi.

Asexual reproduction is particularly common among introduced species, probably because it helps to overcome the negative effects associated with low population densities during colonization. The ant Cerapachys biroi has been introduced to tropical and subtropical islands around the world since the beginning of the last century. In this species, workers can reproduce via thelytokous parthenogenesis. Here, we use genetic markers to reconstruct the history of anthropogenic introductions of C. biroi, and to address the prevalence of female parthenogenesis in introduced and native populations. We show that at least four genetically distinct lineages have been introduced from continental Asia and have led to the species' circumtropical establishment. Our analyses demonstrate that asexual reproduction dominates in the introduced range and is also common in the native range. Given that C. biroi is the only dorylomorph ant that has successfully become established outside of its native range, this unusual mode of reproduction probably facilitated the species' worldwide spread. On the other hand, the rare occurrence of haploid males and at least one clear case of sexual recombination in the introduced range show that C. biroi has not lost the potential for sex. Finally, we show that thelytoky in C. biroi probably has a genetic rather than an infectious origin, and that automixis with central fusion is the most likely underlying cytological mechanism. This is in accordance with what is known for other thelytokous eusocial Hymenoptera.

Effect of an ectoparasite on reproduction in great tits.

1. The effect of a haematophageous ectoparasite, the hen flea, on quality an number of offspring was experimentally investigated in the great tit. The experiment consisted of a controlled infestation of a random sample of nests with the parasitic flea and of a regular treatment of control nests with Microwaves in order to eliminate the naturally occurring fleas. 2. To assess the effects of fleas on variables related to offspring number, we considered the number of hatchlings and fledglings, the mortality between hatching and fledging, and the hatching and fledging success. For assessment of offspring quality, we measured body mass, tarsus and wing length, and calculated the nutritional condition of, nestlings as the ratio of body mass to tarsus length. A physiological variable, the haematocrit level, was also measured. 3. Hatching success and hatchling numbers did not differ between the two experimental groups. Offspring mortality between hatching and fledging was significantly higher in the infested broods (xBAR = 0.22 chicks dead per day) than in the parasite-free broods (xBAR = 0.07 dead per day). Fledging success was 83% in the parasite-free broods, but only 53% in the infested ones. The number of fledglings in infested broods (xBAR = 3.7 fledglings +/-2.1 SD) was significantly lower than in the parasite-free (xBAR = 4.9 +/- 1.1 SD) broods. 4. Body mass of chicks in the infested broods was significantly smaller than in the parasite-free broods both 14 days and 17 days after hatching. The chicks in the infested broods reached a significantly smaller tarsus length than the ones in the parasite-free broods. Close to fledging, the nutritional condition of chicks was significantly lower in infested broods. Haematocrit levels were significantly lower in the infested broods. 5. Brood size correlated differently with body mass and condition of chicks in infested and parasite-free nests. In parasite-free broods both body mass and condition of chicks at age 17 days, i.e. close to fledging, were significantly higher in small broods than in large ones. However, in the infested broods chicks were of the same body mass and condition in large as in small broods. Therefore, in parasite-free broods fitness can potentially be gained through offspring quality or number or both, whereas in infested broods it can be gained through offspring quantity only. In other words, a trade-off between quality and number of offspring is feasible only in the absence of the parasitic hen flea. 6. These results emphasize the need to study the effects of ectoparasites on ecological, behavioural and evolutionary traits of their bird hosts. A knowledge of these effects is essential for the understanding of population dynamics, behaviour and life-history traits of the hosts.

Ultraweak photon emission in assessing bone growth factor efficiency using fibroblastic differentiation.

Photons participate in many atomic and molecular interactions and changes. Recent biophysical research has shown the induction of ultraweak photons in biological tissue. It is now established that plants, animal and human cells emit a very weak radiation which can be readily detected with an appropriate photomultiplier system. Although the emission is extremely low in mammalian cells, it can be efficiently induced by ultraviolet light. In our studies, we used the differentiation system of human skin fibroblasts from a patient with Xeroderma Pigmentosum of complementation group A in order to test the growth stimulation efficiency of various bone growth factors at concentrations as low as 5 ng/ml of cell culture medium. In additional experiments, the cells were irradiated with a moderate fluence of ultraviolet A. The different batches of growth factors showed various proliferation of skin fibroblasts in culture which could be correlated with the ultraweak photon emission. The growth factors reduced the acceleration of the fibroblast differentiation induced by mitomycin C by a factor of 10-30%. In view that fibroblasts play an essential role in skin aging and wound healing, the fibroblast differentiation system is a very useful tool in order to elucidate the efficacy of growth factors.

Fish oil supplementation does not alter energy efficiency in healthy males

BACKGROUND AND AIMS: Fish oil (FO) supplementation prevents the development of obesity and insulin resistance, and upregulate the expression of UCP3 in skeletal muscle in rodents. This may represent indirect evidence that FO promotes fat oxidation and/or alter energy efficiency. The aim of this study was to evaluate whether such effects can be observed in humans. The metabolic effects of FO were assessed during exercise in order to obtain a direct measurement of energy efficiency. METHODS: Eight healthy male volunteers were studied with and without supplementation with 7.2 g/day FO (including 1.1 g/day eicosopentaenoic acid and 0.7 g/day decosahexaenoic acid) during 14 days. Their VO(2 max) was measured on cycle ergometer. Thereafter, energy metabolism (substrate oxidation, energy expenditure and energy efficiency) was assessed during a 30 min cycling exercise at 50% VO(2 max) performed 2 h 30 after a standardized, high carbohydrate breakfast. RESULTS: VO(2 max) was 38.6+/-2.2 after FO and 38.4+/-2.0 (mL x kg(-1) x min(-1)) in control conditions (NS). Basal plasma glucose, insulin and NEFA concentrations, and energy metabolism were similar with FO and in controls. During exercise, the increases in plasma NEFA concentrations, energy expenditure, glucose and lipid oxidation, and the decreases in glycaemia and insulinemia were not altered by FO intake. Energy efficiency was 22.4+/-0.6% after FO vs 21.8+/-0.7% in controls. In order to ascertain that the absence of effects of FO was not due to consumption of a carbohydrate meal immediately before exercise, 4 of the 8 subjects were re-studied in fasting conditions, FO also failed to alter energy efficiency in this subset of studies. CONCLUSION: FO supplementation did not significantly alter energy metabolism and energy efficiency during exercise in healthy humans.

Improved SNR efficiency in gradient echo coronary MRA with high temporal resolution using parallel imaging.

Contemporary coronary magnetic resonance angiography techniques suffer from signal-to-noise ratio (SNR) constraints. We propose a method to enhance SNR in gradient echo coronary magnetic resonance angiography by using sensitivity encoding (SENSE). While the use of sensitivity encoding to improve SNR seems counterintuitive, it can be exploited by reducing the number of radiofrequency excitations during the acquisition window while lowering the signal readout bandwidth, therefore improving the radiofrequency receive to radiofrequency transmit duty cycle. Under certain conditions, this leads to improved SNR. The use of sensitivity encoding for improved SNR in three-dimensional coronary magnetic resonance angiography is investigated using numerical simulations and an in vitro and an in vivo study. A maximum 55% SNR enhancement for coronary magnetic resonance angiography was found both in vitro and in vivo, which is well consistent with the numerical simulations. This method is most suitable for spoiled gradient echo coronary magnetic resonance angiography in which a high temporal and spatial resolution is required.

Costs of female reproduction in a conifer tree: a whole-tree level assessment

1. Costs of reproduction lie at the core of basic ecological and evolutionary theories, and their existence is commonly invoked to explain adaptive processes. Despite their sheer importance, empirical evidence for the existence and quantification of costs of reproduction in tree species comes mostly from correlational studies, while more comprehensive approaches remain missing. Manipulative experiments are a preferred approach to study cost of reproduction, as they allow controlling for otherwise inherent confounding factors like size or genetic background. 2. Here, we conducted a manipulative experiment in a Pinus halepensis common garden, removing developing cones from a group of trees and comparing growth and reproduction after treatment with a control group. We also estimated phenotypic and genetic correlations between reproductive and vegetative traits. 3. Manipulated trees grew slightly more than control trees just after treatment, with just a transient, marginally non-significant difference. By contrast, larger differences were observed for the number of female cones initiated 1 year after treatment, with an increase of 70% more cones in the manipulated group. Phenotypic and genetic correlations showed that smaller trees invested a higher proportion of their resources in reproduction, compared with larger trees, which could be interpreted as an indirect evidence for costs of reproduction. 4. Synthesis. This research showed a high impact of current reproduction on reproductive potential, even when not significant on vegetative growth. This has strong implications for how we understand adaptive strategies in forest trees and should encourage further interest on their still poorly known reproductive life-history traits.

A random linear functional approach to efficiency bounds

This paper suggests a method for obtaining efficiency bounds in models containing either only infinite-dimensional parameters or both finite- and infinite-dimensional parameters (semiparametric models). The method is based on a theory of random linear functionals applied to the gradient of the log-likelihood functional and is illustrated by computing the lower bound for Cox's regression model

Efficiency and significance of multiple vocal signals in sibling competition

Animals can compete for resources by displaying various acoustic signals that may differentially affect the outcome of competition. We propose the hypothesis that the most efficient signal to deter opponents should be the one that most honestly reveals motivation to compete. We tested this hypothesis in the barn owl (Tyto alba) in which nestlings produce more calls of longer duration than siblings to compete for priority access to the indivisible prey item their parents will deliver next. Because nestlings increase call rate to a larger extent than call duration when they become hungrier, call rate would signal more accurately hunger level. This leads us to propose three predictions. First, a high number of calls should be more efficient in deterring siblings to compete than long calls. Second, the rate at which an individual calls should be more sensitive to variation in the intensity of the sibling vocal competition than the duration of its calls. Third, call rate should influence competitors' vocalization for a longer period of time than call duration. To test these three predictions we performed playback experiments by broadcasting to singleton nestlings calls of varying durations and at different rates. According to the first prediction, singleton nestlings became less vocal to a larger extent when we broadcasted more calls compared to longer calls. In line with the second prediction, nestlings reduced vocalization rate to a larger extent than call duration when we broadcasted more or longer calls. Finally, call rate had a longer influence on opponent's vocal behavior than call duration. Young animals thus actively and differentially use multiple signaling components to compete with their siblings over parental resources.

The potential effects of social interactions on reproductive efficiency of stallions

The reproductive efficiency of stabled domestic stallions is often lower than what could be expected from observations in feral herds. In the wild, stallions typically live with mares in harem bands, with other stallions in bachelor bands, or occasionally in mixed-sex transitional bands. We, therefore, argue that permanent contact with mares may increase reproductive efficiency of stallions suffering from low libido and/or fertility. We also provide a summary of our present knowledge of natural conditions, management, and husbandry of domestic stallions, and of intra- and intersexual behavioral interactions in horses.

Genomic determinants of the efficiency of internal ribosomal entry sites of viral and cellular origin

Variation in cellular gene expression levels has been shown to be inherited. Expression is controlled at transcriptional and post-transcriptional levels. Internal ribosome entry sites (IRES) are used by viruses to bypass inhibition of cap-dependent translation, and by eukaryotic cells to control translation under conditions when protein synthesis is inhibited. We aimed at identifying genomic determinants of variability in IRES-mediated translation of viral [Encephalomyocarditis virus (EMCV)] and cellular IRES [X-linked inhibitor-of-apoptosis (XIAP) and c-myc]. Bicistronic lentiviral constructs expressing two fluorescent reporters were used to transduce laboratory and B lymphoblastoid cell lines [15 CEPH pedigrees (n = 205) and 50 unrelated individuals]. IRES efficiency varied according to cell type and among individuals. Control of IRES activity has a significant genetic component (h(2) of 0.47 and 0.36 for EMCV and XIAP, respectively). Quantitative linkage analysis identified a suggestive locus (LOD 2.35) on chromosome 18q21.2, and genome-wide association analysis revealed of a cluster of SNPs on chromosome 3, intronic to the FHIT gene, marginally associated (P = 5.9E-7) with XIAP IRES function. This study illustrates the in vitro generation of intermediate phenotypes by using cell lines for the evaluation of genetic determinants of control of elements such as IRES.