Isolation and characterization of microsatellite markers in the nuclear genome of the brown alga Laminaria digitata (Phaeophyceae).
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Biomedical subjects
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Ten microsatellite loci and a partial sequence of the COII mitochondrial gene were used to investigate genetic differentiation in B. terrestris, a bumble bee of interest for its high-value crop pollination. The analysis included eight populations from the European continent, five from Mediterranean islands (six subspecies altogether) and one from Tenerife (initially described as a colour form of B. terrestris but recently considered as a separate species, B. canariensis). Eight of the 10 microsatellite loci displayed high levels of polymorphism in most populations. In B. terrestris populations, the total number of alleles detected per polymorphic locus ranged from 3 to 16, with observed allelic diversity from 3.8 +/- 0.5 to 6.5 +/- 1.4 and average calculated heterozygosities from 0.41 +/- 0.09 to 0.65 +/- 0.07. B. canariensis showed a significantly lower average calculated heterozygosity (0.12 +/- 0.08) and observed allelic diversity (1.5 +/- 0.04) as compared to both continental and island populations of B. terrestris. No significant differentiation was found among populations of B. terrestris from the European continent. In contrast, island populations were all significantly and most of them strongly differentiated from continental populations. B. terrestris mitochondrial DNA is characterized by a low nucleotide diversity: 0.18% +/- 0.07%, 0.20% +/- 0.04% and 0.27% +/- 0.04% for the continental populations, the island populations and all populations together, respectively. The only haplotype found in the Tenerife population differs by a single nucleotide substitution from the most common continental haplotype of B. terrestris. This situation, identical to that of Tyrrhenian islands populations and quite different from that of B. lucorum (15 substitutions between terrestris and lucorum mtDNA) casts doubts on the species status of B. canariensis. The large genetic distance between the Tenerife and B. terrestris populations estimated from microsatellite data result, most probably, from a severe bottleneck in the Canary island population. Microsatellite and mitochondrial DNA data call for the protection of the island populations of B. terrestris against importation of bumble bees of foreign origin which are used as crop pollinators.
Complex eusocial insect societies are generally matrifilial, suggesting kin selection has been of importance in their development. For simpler social systems, factors favouring their existence, in particular kin selection, have rarely been studied. Communal nesting is one of these simple social organizations, and is found in a diversity of insect species. To examine whether kin selection may play a role in the evolution and maintenance of communality, we estimated genetic relatedness of nestmate females of the facultatively communal bee, Andrena jacobi. Microsatellite loci were developed for this species and used to analyse individuals from two populations. Loci were variable, they were in heterozygote deficit and showed positive inbreeding coefficients. This may arise from nonrandom mating; previous observations (Paxton & Tengö 1996) indicate that a large proportion of females mate intranidally with nestmate males in their natal nests before first emerging. Nestmate relatedness was low, no different from zero for all loci in one population and for three of four loci in the other population. The large number of nestmates sharing a common nest (up to 594) may explain the low relatedness estimates, although relatedness was also independent of the number of females sharing a nest. Lack of inclusive fitness payoffs could constrain social evolution in this communal species.
Samples from nine populations belonging to three African (intermissa, scutellata and capensis) and four European (mellifera, ligustica, carnica and cecropia) Apis mellifera subspecies were scored for seven microsatellite loci. A large amount of genetic variation (between seven and 30 alleles per locus) was detected. Average heterozygosity and average number of alleles were significantly higher in African than in European subspecies, in agreement with larger effective population sizes in Africa. Microsatellite analyses confirmed that A. mellifera evolved in three distinct and deeply differentiated lineages previously detected by morphological and mitochondrial DNA studies. Dendrogram analysis of workers from a given population indicated that super-sisters cluster together when using a sufficient number of microsatellite data whereas half-sisters do not. An index of classification was derived to summarize the clustering of different taxonomic levels in large phylogenetic trees based on individual genotypes. Finally, individual population x loci data were used to test the adequacy of the two alternative mutation models, the infinite allele model (IAM) and the stepwise mutation models. The better fit overall of the IAM probably results from the majority of the microsatellites used including repeats of two or three different length motifs (compound microsatellites).
Similar microsatellite electromorphs (PCR products of the same size) can arise from independent mutational events. Such alleles are not identical by descent. This phenomenon, termed size homoplasy, was studied by sequencing electromorphs of two microsatellite loci in which the stretch of basic repeats is interrupted by different short (1-2 bp) DNA motifs. The number and position of these interruptions were established for electromorphs from closely and distantly related populations of honeybees and bumblebees. No sequence difference was found when electromorphs came from the same subspecies or from closely related subspecies, suggesting that they were probably identical by descent. In contrast, sequence differences were often detected in distantly related subspecies, showing that size homoplasy frequently occurs at this level of population differentiation. Size homoplasy is increased by limits to free length variation of alleles, a phenomenon that seems to act on interrupted microsatellites when comparing distantly related taxa, that is, honeybee subspecies from different evolutionary lineages. Electromorph sequences suggest that, within the scope of these limits, large mutation events have occurred frequently at both interrupted loci studied. In good agreement with the molecular data, computations based on the observed heterozygosity and number of electromorphs and simulation studies showed that neither locus fits the one-step stepwise mutant model (SMM). We speculate that interrupted microsatellites in general could be characterized by a higher variance in repeat number and consequently a lower homoplasy rate than pure ones. Hence, interrupted microsatellites should be most appropriate for investigating population differentiation and evolutionary relationship between relatively distant populations.
A high number of (GT)n and (CT)n microsatellites were isolated from a partial genomic library of primitively eusocial wasp, Vespula rufa. On average, (GT)n and (CT)n repeats occur in the genome approximately every 8 kb and 2.5 kb respectively. This is the highest density of dinucleotide microsatellites reported in the literature. Predominant categories of repeats are imperfect and compound repeats for (GT)n microsatellites and perfect repeats for (CT)n microsatellites. Comparisons with other Hymenoptera and with mammalian species showed significant differences in the frequency distribution of these categories of repeats. Many (CT)n and (GT)n repeats are surrounded by various types of microsatellites, suggesting an associative distribution of short tandem repeats. A high level of intracolonial polymorphism is demonstrated with one microsatellite locus, showing that multiple mating of the queen occurs in V. rufa.
Highly variable microsatellites enabled a precise assessment of the number of queen matings in the colonies of five bumble bee species. Fifteen of the sixteen microsatellites initially cloned from B. terrestris had flanking regions similar enough to allow PCR amplification on the other Bombus species analysed. The microsatellites selected for intracolony study (four per species) were characterized by a high heterozygosity (0.58-0.93) and a large number of alleles (3-18) in the local populations from which the colonies originated. A single male appeared to have inseminated the queens in the colonies of four species, B. terrestris, B. lucorum, B. lapidarius and B. pratorum, which belong to three subgenera, whereas two of the three analysed colonies of B. hypnorum were polyandrous (minimum number of two and four patrilines, respectively).
A set of 52 (CT)n and 23 (GT)n microsatellites in honeybee, 24 (CT)n and 2 (GT)n microsatellites in bumble-bee (n > 6) have been isolated from partial genomic libraries and sequenced. On average, (CT)n and (GT)n microsatellites occur every 15 kb and 34 kb in honeybee and every 40 kb and 500 kb in bumble-bee, respectively. The prevailing categories are imperfect repeats for (CT)n microsatellites in bumble-bee, and perfect repeats for both (CT)n and (GT)n microsatellites in honey-bee. Comparisons with data available in vertebrates indicate a lower proportion of perfect repeats in bees but length distributions are very similar regardless the phylum. This result extends to insects the concept of an evolutionary conservation for quantitative and qualitative characteristics of (CT)n and (GT)n microsatellites. Many (CT)n and (GT)n repeats are surrounded with various types of microsatellites, revealing an associative distribution of short repeat sequences. As expected, a high level of intrapopulational polymorphism has been found with one tested honeybee microsatellite. Also, flanking regions of this microsatellite are similar enough to allow PCR amplification in several other species of Apis and Bombus.
Thirteen (GT)n and four (CT)n microsatellite loci (n = 10 or more and n = 20 or more, respectively) have been isolated from a partial genomic library of brown trout and sequenced. On average, a (GT)n repeat sequence occurs approximately every 23 kb and a (CT)n repeat sequence every 76 kb in brown trout genome. Primers for DNA amplifications using the polymerase chain reaction (PCR) were synthesized for three single locus microsatellites. Mendelian inheritance of the observed polymorphisms was confirmed in full-sib families. Four brown trout populations (10 unrelated individuals per population) were screened for polymorphism with these three microsatellite loci. The total number of alleles detected in the four populations is five at one locus, six at the other two microsatellite loci and is three, on average, per population. Heterozygosities range from 0.18 to 0.74. The largest differences in allelic frequencies occurred between the Mediterranean and the Atlantic populations: this result is congruent with previous allozymic data. The gene-centromere distances of the three microsatellite markers were determined on gynogenetic lines: post-reduction rates range from 0.17 to 0.60. For all the three microsatellite loci, the primers designed from brown trout sequences can be used in another closely related species of salmonid, the rainbow trout (Oncorhynchus mykiss). This last aspect supports the view that microsatellite markers may have wide application in genetic studies in salmonid species and fishes in general.
GENECLASS2 is a software that computes various genetic assignment criteria to assign or exclude reference populations as the origin of diploid or haploid individuals, as well as of groups of individuals, on the basis of multilocus genotype data. In addition to traditional assignment aims, the program allows the specific task of first-generation migrant detection. It includes several Monte Carlo resampling algorithms that compute for each individual its probability of belonging to each reference population or to be a resident (i.e., not a first-generation migrant) in the population where it was sampled. A user-friendly interface facilitates the treatment of large datasets.