The additive genetic variance after bottlenecks is affected by the number of loci involved in epistatic interactions.

We investigated the role of the number of loci coding for a neutral trait on the release of additive variance for this trait after population bottlenecks. Different bottleneck sizes and durations were tested for various matrices of genotypic values, with initial conditions covering the allele frequency space. We used three different types of matrices. First, we extended Cheverud and Routman's model by defining matrices of "pure" epistasis for three and four independent loci; second, we used genotypic values drawn randomly from uniform, normal, and exponential distributions; and third we used two models of simple metabolic pathways leading to physiological epistasis. For all these matrices of genotypic values except the dominant metabolic pathway, we find that, as the number of loci increases from two to three and four, an increase in the release of additive variance is occurring. The amount of additive variance released for a given set of genotypic values is a function of the inbreeding coefficient, independently of the size and duration of the bottleneck. The level of inbreeding necessary to achieve maximum release in additive variance increases with the number of loci. We find that additive-by-additive epistasis is the type of epistasis most easily converted into additive variance. For a wide range of models, our results show that epistasis, rather than dominance, plays a significant role in the increase of additive variance following bottlenecks.

A non-circadian role for clock-genes in sleep homeostasis: a strain comparison.

BACKGROUND: We have previously reported that the expression of circadian clock-genes increases in the cerebral cortex after sleep deprivation (SD) and that the sleep rebound following SD is attenuated in mice deficient for one or more clock-genes. We hypothesized that besides generating circadian rhythms, clock-genes also play a role in the homeostatic regulation of sleep. Here we follow the time course of the forebrain changes in the expression of the clock-genes period (per)-1, per2, and of the clock-controlled gene albumin D-binding protein (dbp) during a 6 h SD and subsequent recovery sleep in three inbred strains of mice for which the homeostatic sleep rebound following SD differs. We reasoned that if clock genes are functionally implicated in sleep homeostasis then the SD-induced changes in gene expression should vary according to the genotypic differences in the sleep rebound. RESULTS: In all three strains per expression was increased when animals were kept awake but the rate of increase during the SD as well as the relative increase in per after 6 h SD were highest in the strain for which the sleep rebound was smallest; i.e., DBA/2J (D2). Moreover, whereas in the other two strains per1 and per2 reverted to control levels with recovery sleep, per2 expression specifically, remained elevated in D2 mice. dbp expression increased during the light period both during baseline and during SD although levels were reduced during the latter condition compared to baseline. In contrast to per2, dbp expression reverted to control levels with recovery sleep in D2 only, whereas in the two other strains expression remained decreased. CONCLUSION: These findings support and extend our previous findings that clock genes in the forebrain are implicated in the homeostatic regulation of sleep and suggest that sustained, high levels of per2 expression may negatively impact recovery sleep.

Genotypic features of lentivirus transgenic mice.

Lentivector-mediated transgenesis is increasingly used, whether for basic studies as an alternative to pronuclear injection of naked DNA or to test candidate gene therapy vectors. In an effort to characterize the genetic features of this approach, we first measured the frequency of germ line transmission of individual proviruses established by infection of fertilized mouse oocytes. Seventy integrants from 11 founder (G0) mice were passed to 111 first generation (G1) pups, for a total of 255 events corresponding to an average rate of transmission of 44%. This implies that integration had most often occurred at the one- or two-cell stage and that the degree of genotypic mosaicism in G0 mice obtained through this approach is generally minimal. Transmission analysis of eight individual proviruses in 13 G2 mice obtained by a G0-G1 cross revealed only 8% of proviral homozygosity, significantly below the 25% expected from purely Mendelian transmission, suggesting counter-selection due to interference with the functions of targeted loci. Mapping of 239 proviral integration sites in 49 founder animals revealed that about 60% resided within annotated genes, with a marked tendency for clustering in the middle of the transcribed region, and that integration was not influenced by the transcriptional orientation. Transcript levels of a set of arbitrarily chosen target genes were significantly higher in two-cell embryos than in embryonic stem cells or adult somatic cells, suggesting that, as previously noted in other settings, lentiviral vectors integrate preferentially into regions of the genome that are transcriptionally active or poised for activation.

Genetics of cadmium hyperaccumulation variance in Thlaspi caerulescens natural populations from Switzerland

SUMMARY Heavy metal presence in the environment is a serious concern since some of them can be toxic to plants, animals and humans once accumulated along the food chain. Cadmium (Cd) is one of the most toxic heavy metal. It is naturally present in soils at various levels and its concentration can be increased by human activities. Several plants however have naturally developed strategies allowing them to grow on heavy metal enriched soils. One of them consists in the accumulation and sequestration of heavy metals in the above-ground biomass. Some plants present in addition an extreme strategy by which they accumulate a limited number of heavy metals in their shoots in amounts 100 times superior to those expected for a non-accumulating plant in the same conditions. Understanding the genetic basis of the hyperaccumulation trait - particularly for Cd - remains an important challenge which may lead to biotechnological applications in the soil phytoremediation. In this thesis, Thlaspi caerulescens J. & C. Presl (Brassicaceae) was used as a model plant to study the Cd hyperaccumulation trait, owing to its physiological and genetic characteristics. Twenty-four wild populations were sampled in different regions of Switzerland. They were characterized for environmental and soil parameters as well as intrinsic characteristics of plants (i.e. metal concentrations in shoots). They were as well genetically characterized by AFLPs, plastid DNA polymorphism and genes markers (CAPS and microsatellites) mainly developed in this thesis. Some of the investigated genes were putatively linked to the Cd hyperaccumulation trait. Since the study of the Cd hyperaccumulation in the field is important as it allows the identification of patterns of selection, the present work offered a methodology to define the Cd hyperaccumulation capacity of populations from different habitats permitting thus their comparison in the field. We showed that Cd, Zn, Fe and Cu accumulations were linked and that populations with higher Cd hyperaccumulation capacity had higher shoot and reproductive fitness. Using our genetic data, statistical methods (Beaumont & Nichols's procedure, partial Mantel tests) were applied to identify genomic signatures of natural selection related to the Cd hyperaccumulation capacity. A significant genetic difference between populations related to their Cd hyperaccumulation capacity was revealed based on somè specific markers (AFLP and candidate genes). Polymorphism at the gene encoding IRTl (Iron-transporter also participating to the transport of Zn) was suggested as explaining part of the variation in Cd hyperaccumulation capacity of populations supporting previous physiological investigations. RÉSUMÉ La présence de métaux lourds dans l'environnement est un phénomène préoccupant. En effet, certains métaux lourds - comme le cadmium (Cd) -sont toxiques pour les plantes, les animaux et enfin, accumulés le long de la chaîne alimentaire, pour les hommes. Le Cd est naturellement présent dans le sol et sa concentration peut être accrue par différentes activités humaines. Certaines plantes ont cependant développé des stratégies leur permettant de pousser sur des sols contaminés en métaux lourds. Parmi elles, certaines accumulent et séquestrent les métaux lourds dans leurs parties aériennes. D`autres présentent une stratégie encore plus extrême. Elles accumulent un nombre limité de métaux lourds en quantités 100 fois supérieures à celles attendues pour des espèces non-accumulatrices sous de mêmes conditions. La compréhension des bases génétiques de l'hyperaccumulation -particulièrement celle du Cd - représente un défi important avec des applications concrètes en biotechnologies, tout particulièrement dans le but appliqué de la phytoremediation des sols contaminés. Dans cette thèse, Thlaspi caerulescens J. & C. Presl (Brassicaceae) a été utilisé comme modèle pour l'étude de l'hyperaccumulation du Cd de par ses caractéristiques physiologiques et génétiques. Vingt-quatre populations naturelles ont été échantillonnées en Suisse et pour chacune d'elles les paramètres environnementaux, pédologique et les caractéristiques intrinsèques aux plantes (concentrations en métaux lourds) ont été déterminés. Les populations ont été caractérisées génétiquement par des AFLP, des marqueurs chloroplastiques et des marqueurs de gènes spécifiques, particulièrement ceux potentiellement liés à l'hyperaccumulation du Cd (CAPS et microsatellites). La plupart ont été développés au cours de cette thèse. L'étude de l'hyperaccumulation du Cd en conditions naturelles est importante car elle permet d'identifier la marque, éventuelle de sélection naturelle. Ce travail offre ainsi une méthodologie pour définir et comparer la capacité des populations à hyperaccumuler le Cd dans différents habitats. Nous avons montré que les accumulations du Cd, Zn, Fe et Cu sont liées et que les populations ayant une grande capacité d'hyperaccumuler le Cd ont également une meilleure fitness végétative et reproductive. Des méthodes statistiques (l'approche de Beaumont & Nichols, tests de Martel partiels) ont été utilisées sur les données génétiques pour identifier la signature génomique de la sélection naturelle liée à la capacité d'hyperaccumuler le Cd. Une différenciation génétique des populations liée à leur capacité d'hyperaccumuler le Cd a été mise en évidence sur certains marqueurs spécifiques. En accord avec les études physiologiques connues, le polymorphisme au gène codant IRT1 (un transporteur de Fe impliqué dans le transport du Zn) pourrait expliquer une partie de la variance de la capacité des populations à hyperaccumuler le Cd.

Importance of genotypic and phenotypic tolerance in the treatment of experimental endocarditis due to Streptococcus gordonii.

Genotypic and phenotypic tolerance was studied in penicillin treatment of experimental endocarditis due to nontolerant and tolerant Streptococcus gordonii and to their backcross transformants. The organisms were matched for in vitro and in vivo growth rates. Rats with aortic endocarditis were treated for 3 or 5 days, starting 12, 24, or 48 h after inoculation. When started at 12 h, during fast intravegetation growth, 3 days of treatment cured 80% of the nontolerant parent compared with <30% of the tolerant derivative (P < .005). When started at 24 or 48 h and if intravegetation growth had reached a plateau, 3 days of treatment failed against both bacteria. However, a significant difference between the 2 organisms was restored when treatment was extended to 5 days. Thus, genotypic tolerance conferred a survival advantage in both fast- and slow-growing bacteria, demonstrating that the in vitro-defined tolerant phenotype also carried the risk of treatment failure in vivo.

Limited clinical benefit of minority K103N and Y181C-variant detection in addition to routine genotypic resistance testing in antiretroviral therapy-naive patients.

OBJECTIVE: The presence of minority nonnucleoside reverse transcriptase inhibitor (NNRTI)-resistant HIV-1 variants prior to antiretroviral therapy (ART) has been linked to virologic failure in treatment-naive patients. DESIGN: We performed a large retrospective study to determine the number of treatment failures that could have been prevented by implementing minority drug-resistant HIV-1 variant analyses in ART-naïve patients in whom no NNRTI resistance mutations were detected by routine resistance testing. METHODS: Of 1608 patients in the Swiss HIV Cohort Study, who have initiated first-line ART with two nucleoside reverse transcriptase inhibitors (NRTIs) and one NNRTI before July 2008, 519 patients were eligible by means of HIV-1 subtype, viral load and sample availability. Key NNRTI drug resistance mutations K103N and Y181C were measured by allele-specific PCR in 208 of 519 randomly chosen patients. RESULTS: Minority K103N and Y181C drug resistance mutations were detected in five out of 190 (2.6%) and 10 out of 201 (5%) patients, respectively. Focusing on 183 patients for whom virologic success or failure could be examined, virologic failure occurred in seven out of 183 (3.8%) patients; minority K103N and/or Y181C variants were present prior to ART initiation in only two of those patients. The NNRTI-containing, first-line ART was effective in 10 patients with preexisting minority NNRTI-resistant HIV-1 variant. CONCLUSION: As revealed in settings of case-control studies, minority NNRTI-resistant HIV-1 variants can have an impact on ART. However, the implementation of minority NNRTI-resistant HIV-1 variant analysis in addition to genotypic resistance testing (GRT) cannot be recommended in routine clinical settings. Additional associated risk factors need to be discovered.

Replicative phenotyping adds value to genotypic resistance testing in heavily pre-treated HIV-infected individuals--the Swiss HIV Cohort Study.

BACKGROUND: Replicative phenotypic HIV resistance testing (rPRT) uses recombinant infectious virus to measure viral replication in the presence of antiretroviral drugs. Due to its high sensitivity of detection of viral minorities and its dissecting power for complex viral resistance patterns and mixed virus populations rPRT might help to improve HIV resistance diagnostics, particularly for patients with multiple drug failures. The aim was to investigate whether the addition of rPRT to genotypic resistance testing (GRT) compared to GRT alone is beneficial for obtaining a virological response in heavily pre-treated HIV-infected patients. METHODS: Patients with resistance tests between 2002 and 2006 were followed within the Swiss HIV Cohort Study (SHCS). We assessed patients' virological success after their antiretroviral therapy was switched following resistance testing. Multilevel logistic regression models with SHCS centre as a random effect were used to investigate the association between the type of resistance test and virological response (HIV-1 RNA <50 copies/mL or ≥1.5 log reduction). RESULTS: Of 1158 individuals with resistance tests 221 with GRT+rPRT and 937 with GRT were eligible for analysis. Overall virological response rates were 85.1% for GRT+rPRT and 81.4% for GRT. In the subgroup of patients with >2 previous failures, the odds ratio (OR) for virological response of GRT+rPRT compared to GRT was 1.45 (95% CI 1.00-2.09). Multivariate analyses indicate a significant improvement with GRT+rPRT compared to GRT alone (OR 1.68, 95% CI 1.31-2.15). CONCLUSIONS: In heavily pre-treated patients rPRT-based resistance information adds benefit, contributing to a higher rate of treatment success.

Breeding system and genetic variance in the monogamous, semi-social shrew, Crocidura russula

The population-genetic consequences of monogamy and male philopatry (a rare breeding system in mammals) were investigated using microsatellite markers in the semisocial and anthropophilic shrew Crocidura russula. A hierarchical sampling design over a 16-km geographical transect revealed a large genetic diversity (h = 0.813) with significant differentiation among subpopulations (F-ST = 5-6%), which suggests an exchange of 4.4 migrants per generation. Demic effective-size estimates were very high, due both to this limited gene inflow and to the inner structure of subpopulations. These were made of 13-20 smaller units (breeding groups), comprising an estimate of four breeding pairs each. Members of the same breeding groups displayed significant coancestries (F-LS = 9-10%), which was essentially due to strong male kinship: syntopic males were on average related at the half-sib level. Female dispersal among breeding groups was not complete (similar to 39%), and insufficient to prevent inbreeding. From our results, the breeding strategy of C. russula seems less efficient than classical mammalian systems (polygyny and male dispersal) in disentangling coancestry from inbreeding, but more so in retaining genetic variance.

Sample size and statistical power in the hierarchical analysis of variance: applications in morphometry of the nervous system.

Analysis of variance is commonly used in morphometry in order to ascertain differences in parameters between several populations. Failure to detect significant differences between populations (type II error) may be due to suboptimal sampling and lead to erroneous conclusions; the concept of statistical power allows one to avoid such failures by means of an adequate sampling. Several examples are given in the morphometry of the nervous system, showing the use of the power of a hierarchical analysis of variance test for the choice of appropriate sample and subsample sizes. In the first case chosen, neuronal densities in the human visual cortex, we find the number of observations to be of little effect. For dendritic spine densities in the visual cortex of mice and humans, the effect is somewhat larger. A substantial effect is shown in our last example, dendritic segmental lengths in monkey lateral geniculate nucleus. It is in the nature of the hierarchical model that sample size is always more important than subsample size. The relative weight to be attributed to subsample size thus depends on the relative magnitude of the between observations variance compared to the between individuals variance.

Effects of common origin and common rearing environment on variance in ectoparasite load and phenotype of nestling Alpine swifts

Knowledge of the quantitative genetics of resistance to parasitism is key to appraise host evolutionary responses to parasite selection. Here, we studied effects of common origin (i.e. genetic and pre-hatching parental effects) and common rearing environment (i.e. post-hatching parental effects and other environment effects) on variance in ectoparasite load in nestling Alpine swifts (Apus melba). This colonial bird is intensely parasitized by blood sucking louse-flies that impair nestling development and survival. By cross-fostering half of the hatchlings between pairs of nests, we show strong significant effect of common rearing environment on variance (90.7% in 2002 and 90.9% in 2003) in the number of louse-flies per nestling and no significant effect of common origin on variance in the number of louse-flies per nestling. In contrast, significant effects of common origin were found for all the nestling morphological traits (i.e. body mass, wing length, tail length, fork length and sternum length) under investigation. Hence, our study suggests that genetic and pre-hatching parental effects play little role in the distribution of parasites among nestling Alpine swifts, and thus that nestlings have only limited scope for evolutionary responses against parasites. Our results highlight the need to take into consideration environmental factors, including the evolution of post-hatching parental effects such as nest sanitation, in our understanding of host-parasite relationships.

Temperature-dependent turnovers in sex-determination mechanisms: a quantitative model.

Sex determination is often seen as a dichotomous process: individual sex is assumed to be determined either by genetic (genotypic sex determination, GSD) or by environmental factors (environmental sex determination, ESD), most often temperature (temperature sex determination, TSD). We endorse an alternative view, which sees GSD and TSD as the ends of a continuum. Both effects interact a priori, because temperature can affect gene expression at any step along the sex-determination cascade. We propose to define sex-determination systems at the population- (rather than individual) level, via the proportion of variance in phenotypic sex stemming from genetic versus environmental factors, and we formalize this concept in a quantitative-genetics framework. Sex is seen as a threshold trait underlain by a liability factor, and reaction norms allow modeling interactions between genotypic and temperature effects (seen as the necessary consequences of thermodynamic constraints on the underlying physiological processes). As this formalization shows, temperature changes (due to e.g., climatic changes or range expansions) are expected to provoke turnovers in sex-determination mechanisms, by inducing large-scale sex reversal and thereby sex-ratio selection for alternative sex-determining genes. The frequency of turnovers and prevalence of homomorphic sex chromosomes in cold-blooded vertebrates might thus directly relate to the temperature dependence in sex-determination mechanisms.

The evolution and consequences of sex-specific reproductive variance.

Natural selection favors alleles that increase the number of offspring produced by their carriers. But in a world that is inherently uncertain within generations, selection also favors alleles that reduce the variance in the number of offspring produced. If previous studies have established this principle, they have largely ignored fundamental aspects of sexual reproduction and therefore how selection on sex-specific reproductive variance operates. To study the evolution and consequences of sex-specific reproductive variance, we present a population-genetic model of phenotypic evolution in a dioecious population that incorporates previously neglected components of reproductive variance. First, we derive the probability of fixation for mutations that affect male and/or female reproductive phenotypes under sex-specific selection. We find that even in the simplest scenarios, the direction of selection is altered when reproductive variance is taken into account. In particular, previously unaccounted for covariances between the reproductive outputs of different individuals are expected to play a significant role in determining the direction of selection. Then, the probability of fixation is used to develop a stochastic model of joint male and female phenotypic evolution. We find that sex-specific reproductive variance can be responsible for changes in the course of long-term evolution. Finally, the model is applied to an example of parental-care evolution. Overall, our model allows for the evolutionary analysis of social traits in finite and dioecious populations, where interactions can occur within and between sexes under a realistic scenario of reproduction.

Natural selection on fecundity variance in subdivided populations: kin selection meets bet hedging.

In a series of seminal articles in 1974, 1975, and 1977, J. H. Gillespie challenged the notion that the "fittest" individuals are those that produce on average the highest number of offspring. He showed that in small populations, the variance in fecundity can determine fitness as much as mean fecundity. One likely reason why Gillespie's concept of within-generation bet hedging has been largely ignored is the general consensus that natural populations are of large size. As a consequence, essentially no work has investigated the role of the fecundity variance on the evolutionary stable state of life-history strategies. While typically large, natural populations also tend to be subdivided in local demes connected by migration. Here, we integrate Gillespie's measure of selection for within-generation bet hedging into the inclusive fitness and game theoretic measure of selection for structured populations. The resulting framework demonstrates that selection against high variance in offspring number is a potent force in large, but structured populations. More generally, the results highlight that variance in offspring number will directly affect various life-history strategies, especially those involving kin interaction. The selective pressures on three key traits are directly investigated here, namely within-generation bet hedging, helping behaviors, and the evolutionary stable dispersal rate. The evolutionary dynamics of all three traits are markedly affected by variance in offspring number, although to a different extent and under different demographic conditions.

Rapid genotypic change and plasticity in arbuscular mycorrhizal fungi is caused by a host shift and enhanced by segregation.

Arbuscular mycorrhizal fungi (AMF) are among the most abundant symbionts of plants, improving plant productivity and diversity. They are thought to mostly grow vegetatively, a trait assumed to limit adaptability. However, AMF can also harbor genetically different nuclei (nucleotypes). It has been shown that one AMF can produce genotypically novel offspring with proportions of different nucleotypes. We hypothesized that (1) AMF respond rapidly to a change of environment (plant host) through changes in the frequency of nucleotypes; (2) genotypically novel offspring exhibit different genetic responses to environmental change than the parent; and (3) genotypically novel offspring exhibit a wide range of phenotypic plasticity to a change of environment. We subjected AMF parents and offspring to a host shift. We observed rapid and large genotypic changes in all AMF lines that were not random. Genotypic and phenotypic responses were different among offspring and their parents. Even though growing vegetatively, AMF offspring display a broad range of genotypic and phenotypic changes in response to host shift. We conclude that AMF have the ability to rapidly produce variable progeny, increasing their probability to produce offspring with different fitness than their parents and, consequently, their potential adaptability to new environmental conditions. Such genotypic and phenotypic flexibility could be a fast alternative to sexual reproduction and is likely to be a key to the ecological success of AMF.