Juan Pedro M. Camacho

Possible origin of a B chromosome deduced from its DNA composition using double FISH technique

Double fluorescentin situ hybridization (FISH) with two DNA probes (a 180 bp tandemly repeated DNA and ribosomal DNA) was performed in embryo cells of the grasshopperEyprepocnemis plorans. Repetitive DNA was present in most standard chromosomes (excepting 7, 8 and 10) and in the proximal two-thirds of the B chromosome, which was its major location in the complement. Ribosomal DNA was present distally on the B, and in the active nucleolar organizer regions (NORs) of the X, 9, 10 and 11 chromosomes. A small number of rRNA gene clusters was also observed in the pericentromeric regions of chromosomes 1–8. The double FISH technique showed that the B chromosome (B2 type) is mainly composed of a 180 bp tandem repeat and ribosomal DNA, the minute short arm being the only region that does not hybridize with them. The location and order of the centromere and both the DNA sequences on the B chromosome coincide only with those in the X chromosome, indicating that the B most probably derives from the X.

A geographic selection mosaic in a generalized plant–pollinator–herbivore system

The concept of Selection Mosaic is central to the Geographic Mosaic Theory of Coevolution. Most information on coevolving interactions, however, comes from specialized organisms. In contrast, an accurate understanding of the effect of geographically varying evolutionary dynamics on the evolution of generalist organisms is lacking, although these kinds of organisms are the most frequent in nature. In flowering plants, pollinators and herbivores are important selective agents. In this study we investigate whether a geographic selection mosaic for floral traits in a generalist plant, Erysimum mediohispanicum (Brassicaceae), can be mediated by the interplay of mutualistic and antagonistic interacting organisms. In eight populations we quantified the selection exerted by these organisms on several plant traits. We found significant spatial variation in pollinator assemblage. In different populations, the main pollinators belonged to different functional groups (beeflies, large bees, small bees, and beetles). Damage by ungulates also varied among populations. Consequently, we found that different populations were under different selective regimes, and the traits affected by selection depended on the local interaction intensity with pollinators and mammal herbivores. Some traits, such as flower number and stalk height, were selected similarly in most populations. Other traits, such as corolla diameter and tube length, were selected only in some populations. Finally, we found divergent selection for some traits, such as corolla tube width and corolla shape, which were selected in contrasting directions in different localities. This spatial variation in selective scenarios results in populations with strong selective regimes (hot spots) intermingled with populations with weak selective regimes (cold spots). Four important outcomes emerge from the E. mediohispanicum selection mosaic. (1) Interactions with generalist organisms may produce strong selection. (2) Spatial changes in main pollinators result in divergent selection across populations. (3) Geographic mosaics depend on a balance between mutualistic and antagonistic selection. (4) Selection mosaics operate at fairly small spatial scales. These findings will surely contribute to expanding the conceptual framework of the Geographic Mosaic Theory of Coevolution.

Location and expression of ribosomal RNA genes in grasshoppers: Abundance of silent and cryptic loci

We investigate regularities and restrictions in chromosome location of ribosomal RNA genes, analysed by fluorescent in situ hybridization (FISH), and their phenotypic expression assessed by nucleolus formation at first meiotic prophase cells, analysed by silver impregnation, in 49 grasshopper species. High variation was found for rDNA location between species within most genera analysed. The mean haploid number of rDNA loci detected by FISH was 2.47, but some species had up to 10 loci. Chromosome distribution of rDNA loci differed between the Gomphocerinae and Oedipodinae subfamilies, most loci being proximal to the centromere in the former and distal to it in the latter. Chromosomes 2, 3 and X frequently carried rDNA in Gomphocerinae species with 2n♂=17 chromosomes, whereas chromosomes 6 and 9 were the most frequent rDNA locations in the Oedipodinae. About 13% of the 126 rDNA loci detected by FISH were silent, although this figure might be even higher. The comparison of FISH and silver-impregnation results also suggested the existence of cryptic NORs, i.e. those forming small nucleoli with no apparent presence of rDNA revealed by FISH. This was especially clear after the same cells in two species were sequentially treated with both silver impregnation and FISH. The abundance of silent and cryptic loci might thus suggest that rDNA spreads through grasshopper genomes by the Dubcovsky and Dvorak mechanism—that is, the transposition of a few rRNA genes to new chromosome locations, their amplification giving rise to new NORs, and the elimination of the old NORs. The cryptic NORs might correspond to nascent NORs, i.e. a few rRNA gene copies moved to new locations, whereas the inactive rDNA loci might correspond to those being in the process of elimination.

Natural Selection on Erysimum mediohispanicum Flower Shape: Insights into the Evolution of Zygomorphy

Paleontological and phylogenetic studies have shown that floral zygomorphy (bilateral symmetry) has evolved independently in several plant groups from actinomorphic (radially symmetric) ancestors as a consequence of strong selection exerted by specialized pollinators. Most studies focused on unraveling the developmental genetics of flower symmetry, but little is known about the adaptive significance of intraspecific flower shape variation under natural conditions. We provide the first evidence for natural selection favoring zygomorphy in a wild population of Erysimum mediohispanicum (Brassicaceae), a plant showing extensive continuous variation in flower shape, ranging from actinomorphic to zygomorphic flowers. By using geometric morphometric tools to describe flower shape, we demonstrate that plants bearing zygomorphic flowers received more pollinator visits and had the highest fitness, measured not only by the number of seeds produced per plant but also by the number of seeds surviving to the juvenile stage. This study provides strong evidence for the existence of significant fitness differences associated with floral shape variation in E. mediohispanicum, thus illuminating a pathway for the evolution of zygomorphy in natural populations.

Structural and functional evidence that a B chromosome in the characid fish Astyanax scabripinnis is an isochromosome

Astyanax scabripinnis possesses a widespread polymorphism for metacentric B chromosomes as large as the largest chromosome pair in the A complement. On the basis of C-banding pattern, it was hypothesized that these B chromosomes are isochromosomes that have arisen by means of centromere misdivision and chromatid nondisjunction. In the present paper we test this hypothesis by analysing (i) the localization of a repetitive DNA sequence on both B chromosome arms, and (ii) synaptonemal complex formation, in order to test the functional homology of both arms. Genomic DNA digested with KpnI and analysed by gel electrophoresis showed fragments in a ladder-like pattern typical of tandemly repetitive DNA. These fragments were cloned and their tandem organization in the genome was confirmed. A 51-bp long consensus sequence, which was AT-rich (59%) and contained a variable region and two imperfect reverse sequences, was obtained. Fluorescence in situ hybridization (FISH) localized this repetitive DNA into noncentromeric constitutive heterochromatin which encompasses the terminal region of some acrocentric chromosomes, the NOR region, and interstitial polymorphic heterochromatin in chromosome 24. Most remarkably, tandem repeats were almost symmetrically placed in the two arms of the B chromosome, with the exception of two additional small clusters proximally located on the slightly longer arm. Synaptonemal complex (SC) analysis showed 26 completely paired SCs in males with 1B. The ring configuration of the B univalent persisting until metaphase I suggests that the two arms formed chiasmata. All these data provided strong support for the hypothesis that the B chromosome is an isochromosome.

Spatial variation in selection on corolla shape in a generalist plant is promoted by the preference patterns of its local pollinators

Summary An adaptive role of corolla shape has been often asserted without an empirical demonstration of how natural selection acts on this trait. In generalist plants, in which flowers are visited by diverse pollinator fauna that commonly vary spatially, detecting pollinator-mediated selection on corolla shape is even more difficult. In this study, we explore the mechanisms promoting selection on corolla shape in the generalist crucifer Erysimum mediohispanicum Polatschek (Brassicaceae). We found that the main pollinators of E. mediohispanicum (large bees, small bees and bee flies) discriminate between different corolla shapes when offered artificial flowers without reward. Importantly, different pollinators prefer different shapes: bees prefer flowers with narrow petals, whereas bee flies prefer flowers with rounded overlapping petals. We also found that flowers with narrow petals (those preferred by bees) produce both more pollen and nectar than those with rounded petals. Finally, different plant populations were visited by different faunas. As a result, we found spatial variation in the selection acting on corolla shape. Selection favoured flowers with narrow petals in the populations where large or small bees are the most abundant pollinator groups. Our study suggests that pollinators, by preferring flowers with high reward, exert strong selection on the E. mediohispanicum corolla shape. The geographical variation in the pollinator-mediated selection on E. mediohispanicum corolla shape suggests that phenotypic evolution and diversification can occur in this complex floral trait even without specialization.

Population Dynamics of A Selfish B Chromosome Neutralized by the Standard Genome in the Grasshopper Eyprepocnemis Plorans

Effects of the B chromosome polymorphism of the grasshopper Eyprepocnemis plorans were analyzed in two natural populations. Postmating sexual selection, female fertility, and survival were studied. The B chromosome lacks drive and has no detectable effects on fitness. A neutral B cannot invade a population and establish a polymorphism, but the confidence limits on our estimates cannot exclude the possibility that the polymorphism is maintained by a balance between weak drive and weak selection against individuals with two and three Bs. However, other lines of evidence favor the following model of the dynamics of the B in E. plorans. In a newly invaded population, the B has substantial drive, but the evolution of drive suppressor genes in the A chromosomes neutralizes the B drive so that it becomes near‐neutral and begins a random walk toward extinction by stochastic loss. Because the B is common by the time drive disappears, the random walk is likely to continue for a long time. If in the course of the random walk a variant B with greater drive appears, then it will displace the original variant, and a new cycle of drive suppression and drift to extinction occurs. A simulation model of this process suggested that the mean time to extinction is proportional to the two‐thirds power of the population size; it is much less affected by subpopulation size or the number of populations in a subdivided population.

Sex-ratio distortion associated with the presence of a B chromosome in Astyanax scabripinnis (Teleostei, Characidae)

Cytogenetic studies were conducted on 154 specimens of Astyanax scabripinnis collected at three localities in the Campos do Jordão region (State of São Paulo, Brazil). The C-banding pattern suggested that the metacentric B chromosome found in most of the specimens is an isochromosome derived from chromosome 24, the only chromosome in the standard complement that carries interstitial C-bands similar to those present in each arm of the B chromosome. The sex ratio was biased toward females in the Córrego das Pedras and Ribeirão do Casquilho streams and toward males in the Ribeirão das Perdizes stream. In all three populations analyzed, the B chromosome was more frequent in females than in males. In the most exhaustively sampled population (those from Córrego das Pedras), there was a highly significant association between B-chromosome frequency and sex-ratio distortion, with a disproportionately high number of males without B chromosomes and females with one B chromosome.

POLYMORPHISM REGENERATION FOR A NEUTRALIZED SELFISH B CHROMOSOME

Long‐run evolution of B chromosomes is mainly made up by an evolutionary arms race between these selfish genetic elements and the standard genome. The suppression of B drive is one of the clearest expressions of genome defense against B chromosomes. After drive neutralization, the B is condemned to extinction unless a new variant showing drive can emerge and replace it. This paper reports the first empirical evidence for the substitution of a neutralized B variant by a new selfish B variant. Such a polymorphism regeneration has recently taken place in a natural population of the grasshopper Eyprepocnemis plorans.

Chromosome mapping of H3 and H4 histone gene clusters in 35 species of acridid grasshoppers

We analyse chromosome location of H3 and H4 histone gene clusters by fluorescence in-situ hybridization (FISH) in 35 species of Acrididae grasshoppers belonging to seven subfamilies. As in other organisms, H3 and H4 co-localized in the same chromosome region in the 11 species where double FISH was performed with the H3 and H4 DNA probes. Chromosome location of H3-H4 histone gene clusters showed high regularity in the species analysed, with all of them carrying a single H3-H4 cluster in an autosome which, in most cases, was located interstitially in the proximal chromosome third. In 17 out of the 21 species with 2n♂ = 23 acrocentric chromosomes, the H3-H4-carrying autosome was about eighth in order of decreasing size. Two of the four exceptions changed H3-H4 localization to proximal (Pezotettix giornae) or distal (Tropidopola graeca) in the eighth-sized autosome, but the remainder (the two Eyprepocnemis species) showed the H3-H4 cluster distally located in the second-sized autosome. All 14 species with 2n♂ = 17 chromosomes (including three long metacentric autosome pairs, five acrocentric autosome pairs and an acrocentric X chromosome) carried an interstitial H3-H4 cluster in the short arm of the smallest of the three long metacentric pairs. These results suggest that chromosome location of H3-H4 histone gene clusters seem to be highly conservative in Acrididae grasshoppers. The change in H3-H4 location from the acrocentric medium-sized autosome in the 2n♂ = 23 karyotype to the long metacentric autosome in the 2n♂ = 17 karyotype is most parsimoniously explained by common ancestry, i.e. by the involvement of the H3-H4-carrying acrocentric in the centric fusion that gave rise to the smallest of the three long metacentric autosomes of 2n♂ = 17 species.