Viruses are common in asexual Aspergilli but not in sexual Aspergilli. We found no viruses in 112 isolates of the sexual Aspergillus nidulans. We have investigated factors that could play a role in preventing the spread of mycoviruses through populations of A. nidulans. Experiments were performed with A. nidulans strains infected with viruses originating from A. niger. Horizontal virus transmission was restricted but not prevented by somatic incompatibility. Viruses were transmitted vertically via conidiospores but not via ascospores. Competition experiments revealed no effect of virus infection on host fitness. Outcrossing was found to limit the spread of viruses significantly more than selfing. It is concluded that the exclusion of viruses from sexual Aspergilli could be due to the formation of new somatic incompatibility groups by sexual recombination.

Homozygous or hemizygous expression of an X-linked wing mutant of Drosophila robusta varies from a rudimentary wing that does not reach the tip of the abdomen (called ‘club’) to forms with full-sized but curled or crumpled wings (called ‘curly’). Homozygous club females crossed to flies from natural populations or laboratory stocks derived from wild flies invariably produce significantly less club male progeny than the 100% expected, most of them exhibiting less severe phenotypes: ‘curly’ forms and wild-type. The male progeny from similar crosses using curly females tend to be predominantly normal. By contrast, the male progeny of outcrossed females homozygous for an X-linked eye colour mutant, vermilion, are all vermilion. The data indicate that natural populations of D. robusta contain suppressors of the wing mutant but not of the eye colour mutant studied. Activity of the suppressors differs by gender: in experiments in which genetic theory expects similar results in the two sexes, males consistently show stronger effects of the suppressors than females.

RNA viruses consist of populations of extremely high genetic heterogeneity called quasispecies. Based on theoretical considerations, it has been suggested that the unit of selection in such complex genetic populations is not the single viral particle but a set of genetically related particles which form the quasispecies. In the present study we carried out a set of experiments with the vesicular stomatitis virus (VSV) dealing with the evolution of life-history characters under selection acting at two factors either in the same or in opposite directions. The two factors at which selective pressure is applied are the individual and the group. We show evidence that group selection modulates the virulence of VSV populations, in opposition to an unlimited increase in virulence by competitive optimization promoted by individual selection. The results are of relevance for understanding the evolution of parasite virulence.

Genetic diversity of natural populations of the two species Naegleria gruberi and N. australiensis collected at the same time in the same area was studied using multilocus enzyme electrophoresis (MLEE analysis). Reference strains and related species were also analysed for comparison. Five loci were found to be polymorphic and allowed identification of 19 different multilocus genotypes among 67 isolates – eight for N. gruberi and 11 for N. australiensis – none being common to the two species. Large departures from Hardy–Weinberg equilibrium were found at all the loci, with significant heterozygote deficiencies in most cases. The two species exhibited strong linkage disequilibrium and the predominance of a few multilocus genotypes within each species was observed. These lines of evidence strongly suggest non-random association of alleles, leading to the conclusion that N. gruberi and N. australiensis have predominantly clonal genetic structures in the wild. Populations consist of a few major clones with some closely related ones – a structure found for several parasitic protozoans and bacteria. This contrasts with the panmictic structure found for N. lovaniensis, another species of this genus, and suggests that closely related species within the genus have evolved to have different reproductive strategies.

We have used the method of disequilibrium pattern analysis to examine associations between the threonine-glycine (Thr-Gly) encoding repeat region of the clock gene period (per) of Drosophila melanogaster, and polymorphic sites both upstream and downstream of the repeat, in a number of European fly populations. The results are consistent with the view that selection may be operating on various haplotypes which share the Thr-Gly length alleles encoding 17, 20 and 23 dipeptide pairs, and that the repeat itself may be the focus for selection. These conclusions lend support to a number of other population and behavioural investigations which have provided evidence that selection is acting on the Thr-Gly region. The linkage analysis was also used to infer an approximate mutation rate (μ) for the repeat, of 10−5<μ<4×10−5 per gamete per generation. Direct measurements of the mutation rate using the polymerase chain reaction in a pedigree analysis of tens of thousands of individuals do not contradict this value. Consequently, the Thr-Gly repeat does not have a mutation rate that is as high as some of the non-coding minisatellites, but it is several orders of magnitude higher than the nucleotide substitution rate. The implications of this elevated mutation rate for linkage disequilibria and selection are discussed.

The rolA gene of Agrobacterium rhizogenes contains in its untranslated leader region a spliceosomal intron, which is spliced in Arabidopsis and in Nicotiana tabacum. Expression under the control of the 35S promoter from cauliflower mosaic virus of a rolA gene derivative defective in splicing still causes alterations of growth in transgenic tobacco plants. Splicing of rolA mRNA is required for efficient expression of the rolA phenotype in vivo. Moreover, splicing is required for efficient in vitro translation of the rolA mRNA. In contrast, expression of a 35S-rolA gene derivative with the ATG initiation codon replaced by ATA does not cause any phenotypical alteration. Mutations leading to amino acid substitutions at positions 37 and 40 of the rolA coding region were isolated as null mutants in Arabidopsis plants transgenic for the rolA gene. However, when expressed in tobacco under the control of the 35S promoter, they cause a rolA phenotype reduced in the expressivity of its traits. The molecular characterization of rolA mutants might be useful for understanding the biochemical function of the rolA protein.

In the evolutionary process during which Drosophila sechellia became specialized on a toxic fruit (morinda), a spectacular decrease in female reproductive capacity took place when compared with the species' generalist relatives D. mauritiana and D. simulans. Comparisons of species and interspecific crosses showed that two different traits were modified: number of ovarioles and rate of egg production. During the conservation of a D. sechellia strain on usual food, adaptation to laboratory conditions led to an increase in the rate of oogenesis but not in ovariole number. Comparison of F1 and backcross progeny also suggests that the two traits are determined by different genes (ovariole number has already been shown to be polygenic). When morinda is available as a resource, the low rate of egg production in D. sechellia is partly compensated by a stimulating effect, while an inhibition occurs in D. simulans. It is assumed that D. sechellia progressively adapted itself from rotten, non-toxic morinda to a fresher and more toxic resource. During this process the rate of oogenesis evolved from an inhibition to a stimulation by morinda. Simultaneously a spectacular decrease in ovariole number took place, either as a consequence of stochastic events related to the small population size of D. sechellia and a metapopulation dynamics, or as an adaptive process favouring dispersal capacities of the female.

In vitro studies using highly purified calf thymus RNA polymerase II and a fragment spanning the first intron of H3.3 as template DNA have demonstrated the existence of a strong transcription termination site consisting of thymidine stretches. In this study, nuclear run-on experiments have been performed to assess the extent to which transcription elongation is blocked in vivo using DNA probes corresponding to regions 5′ and 3′ of the in vitro termination sites. These studies suggest that H3.3 expression is stimulated following the inhibition of DNA synthesis through the elimination of the transcription elongation block. Interestingly, both the in vivo and in vitro experiments have revealed that the transcriptional block/termination sites are positioned immediately downstream of a 73 bp region that has been over 90% conserved between the chicken and human H3.3 genes. The extreme conservation of this intronic region suggests a possible role in maintaining cis-acting function. Electrophoretic mobility shift experiments show that HeLa cell nuclear extracts contain protein factors that bind specifically to the region of transcription elongation block. Furthermore, we demonstrate a correlation between the protein binding activity and the transcriptional block in cells that have been either arrested at the initiation of S phase or were replication-interrupted by hydroxyurea. DNA footprinting experiments indicate that the region of protein binding is at the 3′ end of the conserved region and overlaps with one of the three in-vitro-mapped termination sites.

Genetic instability is associated with aging in many species. One of the initiating factors for genetic instability is the movement of transposable elements (TEs), which occur in all prokaryotic and eukaryotic organisms. The hypothesis that TEs could be involved in the aging process is discussed and the correlation between aging and activity of TEs is analysed in a variety of biological systems.

The theory developed by Takahata and colleagues for estimating the effective population size of ancestral species using homologous sequences from closely related extant species was extended to take account of variation of evolutionary rates among loci. Nuclear sequence data related to the evolution of modern humans were reanalysed and computer simulations were performed to examine the effect of rate variation on estimation of ancestral population sizes. It is found that the among-locus rate variation does not have a significant effect on estimation of the current population size when sequences from multiple loci are sampled from the same species, but does have a significant effect on estimation of the ancestral population size using sequences from different species. The effects of ancestral population size, species divergence time and among-locus rate variation are found to be highly correlated, and to achieve reliable estimates of the ancestral population size, effects of the other two factors should be estimated independently.

Until now, with regard to the hobo system of hybrid dysgenesis, natural populations of Drosophila melanogaster have been investigated using only two criteria: at the molecular level, the presence or absence of XhoI fragments 2·6 kb long or smaller; and/or at the genetic level, the ability to induce gonadal dysgenesis sterility in crosses A (females of an E reference strain crossed with males under test) and A* (females under test crossed with males of an H reference strain). Recently, analyses of laboratory strains using these criteria as well as the mobilization of two reporter genes, the male recombination and the number of ‘TPE’ repeats in the S region, revealed a lack of correlation between the different dysgenic parameters themselves, and also between these parameters and the molecular characteristics of the strains. Thirteen current strains derived from world populations were therefore investigated with regard to all these dysgenic traits, to determine discriminating criteria providing a robust method of classifying natural populations and deducing the dynamics of hobo elements in these populations. We show, as in laboratory strains, a lack of correlation between the parameters studied. Therefore, the significance of each of them as well as the nature of hobo hybrid dysgenesis are discussed, to propose an analysis method of the hobo system applicable to natural populations. According to the geographical distribution of hobo activities in world populations and to the variable polymorphism of the number of ‘TPE’ repeats, we propose a new scenario for the invasion of D. melanogaster by hobo elements.

Koala (Phascolarctos cinereus) populations in southern Australia have a history of bottlenecks – earlier this century the species became extinct in South Australia, and almost so in Victoria. Subsequently large numbers of animals from island populations (founded from very few animals) have been translocated back to mainland sites and to other islands in the region. As part of a larger study of the genetic structure of koala populations in southern Australia, we have undertaken a survey of mitochondrial DNA restriction fragment length polymorphism (mtDNA-RFLP) variability. Genomic DNA from 91 koalas from five populations was examined using 23 restriction enzymes, and mtDNA fragments were detected using a domestic cat full-length mtDNA clone. Only one of the enzymes, TaqI, revealed polymorphism – a relatively low amount of variation compared with other mammals, although low mtDNA-RFLP variation has also been reported in Queensland koalas. French Island and populations established predominantly from French Island immigrant koalas, either directly or via other island populations, were indistinguishable by haplotype frequencies. The mtDNA data are thus consistent with the interpretation that the koala translocation programme has homogenized gene frequencies amongst those populations involved. South Gippsland is not recorded as having received translocated koalas directly, and has significantly different mtDNA-RFLP haplotype frequencies from all other populations examined. The fact that this distinction was not previously observed in nuclear gene frequencies may reflect predominantly male-mediated dispersal in koalas.

A modification of the neighbour joining method of Saitou & Nei (1987) is shown to be applicable to the ordering of genetic markers. This neighbour mapping method is compared with some other procedures for ordering genetic markers using both real and test data sets. The limitations and likely errors associated with the use of neighbour mapping are discussed. The speed and simplicity of this method commend its application, as does its concurrence with other mapping methods.

Although cryopreservation is routinely used for the storage of a range of biological organisms, few studies have been conducted to determine whether cryopreservation increases the frequency of mutation. A procedure for the cryopreservation of Drosophila melanogaster embryos has recently been developed. Cryopreservation of D. melanogaster is of special interest to geneticists and evolutionary biologists because it would make it possible to assay control and experimental populations simultaneously during long-term studies. Before cryopreserved embryos can be used for such studies, it is first necessary to show that cryopreservation is not mutagenic. We tested for mutagenic effects of cryopreservation in D. melanogaster embryos with an X-linked, recessive lethal assay. The mutation rates of cryopreserved and control flies were not significantly different. We can be 95% certain that cryopreservation does not increase mutation by a factor greater than 2·39. This is the first quantitative estimate of the mutagenic effect of cryopreservation on the germ line of a metazoan. The results are reassuring when considering the genetic impact of cryopreservation on mammalian gametes and embryos.

Since the last glacial period forest trees have expanded to their present range very rapidly, with rates up to 500 m yr−1 for oaks in Europe, which can be explained only by the dispersion of acorns over long distances. We used a stratified dispersal model, including both diffusive and long-distance dispersal of seeds, to simulate the colonization of a 100 km×300 km grid by populations of oak trees. An appropriate rate of spread is obtained with rare dispersal at distances of the order of tens of kilometres. We simulated the effect of stratified versus diffusive dispersal of seeds on the spatial genetic structure at a maternally inherited locus. Founding events associated with stratified dispersal generate a high amount of genetic differentiation among populations, which is likely to persist for a long time after colonization. Using autocorrelation methods, we show that diffusive and stratified dispersals create quite different spatial patterns of variation for the maternally inherited locus. Stratified dispersal creates patchy patterns that are concordant with a previous experimental investigation of chloroplast DNA variation at a regional scale in the oaks Quercus petraea and Quercus robur. For plant populations that have passed through recent episodes of range expansion, long-distance dispersal events are probably the most important factors of spatial genetic structuring of maternally inherited genes at small or medium geographic scales.

A new estimator is proposed for the parameter C=4Nc, where N is the population size and c is the recombination rate in a finite population model without selection. The estimator is an improved version of Hudson's (1987) estimator, which takes advantage of some recent theoretical developments. The improvement is slight, but the smaller bias and standard error of the new estimator support its use. The variance of the average number of pairwise differences is also derived, and is important in the formulation of the new estimator.

Based on a deterministic mutation–selection model the concept of error thresholds is critically examined. It has often been argued that genetic information – for instance, an advantageous allele – can be selectively maintained in a population only if the mutation rate is below a certain limit, the error threshold, which is inversely related to the genome size. Here, I will show that such an inverse relationship strongly depends on the fitness model. To produce the error threshold, as given by Eigen (1971), requires that the fitness model is an extreme form of diminishing epistasis. The error threshold, in a strict sense, vanishes as epistasis changes from diminishing to synergistic. In the latter case even the usual definition of error thresholds becomes ambiguous. Initially, a finite sites model has been used to describe error thresholds. However, they can also be defined within the framework of the infinite sites model. I study both models in parallel and compare their properties as far as error thresholds are concerned. It is concluded that error thresholds possibly play a much less important role in molecular evolution than has often been assumed in the past.

To assess the role of genetic changes in sensitivity to leptin hormone in contributing to responses to long-term selection for fatness, leptin was administered to a long-term fat selected (F) and a control line (C) of mice. These lines differ almost three fold in their percentage of fat (fat%) at about 15 weeks of age. Treated (T) animals received twice-daily intraperitoneal injections of 5 mg/kg leptin from 91 to 105 days of age; untreated (U) animals received equivolume injections of phosphate-buffered saline. Treated compared with untreated animals in both lines had significantly (P<0·05) lower mean body weight, food intake and fatness at the end of test (fat%: CT 3%, CU 7·4%, FT 14·9%, FU21·1%). The differences in response between the lines [(CT−CU)−(FT−FU)] were all non-significant (P>0·05), however. There was a very wide range of fatness (estimated from dry matter content) among FT animals (3–29%), much higher than in FU (15–31%), CT (0·7–6·4%) and CU (2–15%) animals. While sensitivity to leptin remains in the fat line, response appears to vary among animals at the dose level used.

A simple model of migration between two populations, each in a balance between mutation and stabilizing selection on a polygenic trait, is explored. Below a critical migration rate, genetic differences between the two populations can be maintained, even if the populations are selected towards the same phenotypic optimum. Gene flow then maintains genetic variance within each population. For this process to account for heritable variation, there must be some mechanism that causes divergence. The possibility that fluctuating selection could lead to the initial differentiation of the populations is explored.

In crosses between inbred lines linear regression can be used to estimate marker effects; these marker effects then allow marker-assisted selection (MAS) for quantitative traits. Weighting of marker and phenotypic information in MAS requires estimation of genetic variance associated with the markers: the usual estimators are biased, resulting in too much weight being placed on marker information relative to phenotypic information. In this paper we develop a cross-validation method to remove this bias, and show by simulation that response to selection using this method is almost as high as that achieved using optimal weighting of marker and phenotypic information.