Genome‐specific histories of divergence and introgression between an allopolyploid unisexual salamander lineage and two ancestral sexual species

Quantifying introgression between sexual species and polyploid lineages traditionally thought to be asexual is an important step in understanding what drives the longevity of putatively asexual groups. Here, we capitalize on three recent innovations—ultraconserved element (UCE) sequencing, bioinformatic techniques for identifying genome‐specific variation in polyploids, and model‐based methods for evaluating historical gene flow—to measure the extent and tempo of introgression over the evolutionary history of an allopolyploid lineage of all‐female salamanders and two ancestral sexual species. Our analyses support a scenario in which the genomes sampled in unisexual salamanders last shared a common ancestor with genomes in their parental species ～3.4 million years ago, followed by a period of divergence between homologous genomes. Recently, secondary introgression has occurred at different times with each sexual species during the last 500,000 years. Sustained introgression of sexual genomes into the unisexual lineage is the defining characteristic of their reproductive mode, but this study provides the first evidence that unisexual genomes have undergone long periods of divergence without introgression. Unlike other sperm‐dependent taxa in which introgression is rare, the alternating periods of divergence and introgression between unisexual salamanders and their sexual relatives could explain why these salamanders are among the oldest described unisexual animals.     

Evolution of clonality and polyploidy in a weevil system

The increased interest in asexual organisms calls for in-depth studies of asexual complexes that actively give rise to new clones. We present an extensive molecular study of the Otiorhynchus scaber (Coleoptera, Curculionidae) weevil system. Three forms have traditionally been recognized: diploid sexuals, triploid, and tetraploid parthenogens. All forms coexist in a small central area, but only the polyploid parthenogens have colonized marginal areas. Analyzing the phylogenetic relationship, based on three partial mitochondrial genes, of 95 individuals from 19 populations, we find that parthenogenesis and polyploidy have originated at least three times from different diploid lineages. We observe two major mitochondrial lineages, with over 2.5% sequence divergence between the most basal groups within them, and find that current distribution and phylogenetic relationships are weakly correlated. Quite unexpectedly, we also discover diploid clones that coexist with, and are morphologically indistinguishable from, the diploid sexual females. Our results support that these diploid clones are derived directly from the diploid sexuals. We also find that it is mainly an increase in ploidy level and not the benefits of asexual reproduction that confers to polyploid parthenogens the advantage over their diploid sexual relatives.     

Adrenergic receptor homologies in vertebrate and invertebrate species examined by DNA hybridization

The deduced protein sequences of the mammalian adrenergic receptors (ARs) suggest that these proteins have evolved by several ancient gene duplication events. To investigate in what species these events may have occurred DNA fragments encoding the family of adrenergic receptors from human (beta 1AR and alpha 2AR) and hamster (beta 2AR and alpha 1AR) were used to detect homologous sequences in other vertebrates, invertebrates and unicellular organisms by Southern blot hybridization analysis. Sequences homologous to hamster beta 2AR were detected in lower vertebrates, invertebrates and Dictyostelium, but not in yeast or bacteria. Within vertebrates, sequences strongly homologous to human beta 1AR and human platelet alpha 2AR were confined to the higher vertebrates only. In the invertebrates, only Drosophila contained sequences homologous to hamster alpha 1AR. Our results suggest that non-mammalian species may contain receptors homologous to the mammalian adrenergic receptors and that the sequences homologous to human beta 2AR have been the most strongly conserved.     

Ancient hybridization and mitochondrial capture between two species of chipmunks

Models that posit speciation in the face of gene flow are replacing classical views that hybridization is rare between animal species. We use a multilocus approach to examine the history of hybridization and gene flow between two species of chipmunks ( Tamias ruficaudus and T. amoenus ). Previous studies have shown that these species occupy different ecological niches and have distinct genital bone morphologies, yet appear to be incompletely isolated reproductively in multiple areas of sympatry. We compared data from four sequenced nuclear loci and from seven microsatellite loci to published cytochrome b sequences. Interspecific gene flow was primarily restricted to introgression of the T. ruficaudus mitochondrial genome into a sympatric subspecies of T. amoenus , T. a. canicaudus , with the four sequenced nuclear loci showing little to no interspecific allele sharing. Microsatellite data were consistent with high levels of differentiation between the species and also showed no current gene flow between broadly sympatric populations of T. a. canicaudus and T. ruficaudus . Coalescent analyses date the mtDNA introgression event from the mid-Pleistocene to late Pliocene. Overall, these data indicate that introgression has had a minimal impact on the nuclear genomes of T. amoenus and T. ruficaudus despite multiple independent hybridization events. Our findings challenge long-standing assumptions on patterns of reproductive isolation in chipmunks and suggest that there may be other examples of hybridization among the 23 species of Tamias that occur in western North America.     

Distribution, mechanisms and evolutionary significance of clonality and polyploidy in weevils

Abstract 1 Genetical mtDNA relationships of 41 taxa of weevils were examined using cladistics. Ingroup taxa belong to Otiorhynchus scaber and O. nodosus and outgroup comparison was made with O. singularis. All three species are minor forest pests. 2 Otiorhynchus scaber specimens are either diploid sexuals or diploid, triploid and tetraploid clones, from two different populations (Slovenia and Austria) that belong to two different evolutionary lineages. Otiorhynchus nodosus specimens are tetraploid clones. Both species show geographical parthenogenesis, as do many other Otiorhynchus species. 3 Mitochondrial data indicate that O. nodosus clones are more closely related to Slovenian sexuals of O. scaber than these are to sexuals from Austria. It also shows that almost all clones of O. scaber collected in one of the two regions where sexuals are found are more closely related to sexuals from the other region. 4 Three different hypotheses that may explain the distribution of O. scaber, mechanisms important for the evolution of the clones and implications of the presence of Wolbachia are discussed. 5 We conclude that parthenogenesis is likely to be linked to hybridization in O. scaber and that hybridization events between ancestors of O. nodosus and O. scaber are the probable cause of the presence of O. nodosus in the ingroup. We also find that polyploid clones are superior colonizers compared to sexuals and diploid clones, in O. scaber. 6 The results suggest that systems where both sexuals and clones exist are more complex than previously suggested. The mapping of genetic variation in clonal complexes and the tracing of clonal origins may be useful in pest management.     

Existence of two sexual races in the planarian species switching between asexual and sexual reproduction

In certain planarian species that are able to switch between asexual and sexual reproduction, determining whether a sexual has the ability to switch to the asexual state is problematic, which renders the definition of sexuals controversial. We experimentally show the existence of two sexual races, acquired and innate, in the planarian Dugesia ryukyuensis. Acquired sexuals used in this study were experimentally switched from asexuals. Inbreeding of acquired sexuals produced both innate sexuals and asexuals, but inbreeding of innate sexuals produced innate sexuals only and no asexuals. Acquired sexuals, but not innate sexuals, were forced to become asexuals by ablation and regeneration (asexual induction). This suggests that acquired sexuals somehow retain asexual potential, while innate sexuals do not. We also found that acquired sexuals have the potential to develop hyperplastic and supernumerary ovaries, while innate sexuals do not. In this regard, acquired sexuals were more prolific than innate sexuals. The differences between acquired and innate sexuals will provide a structure for examining the mechanism underlying asexual and sexual reproduction in planarians.     

Isolation of microsatellites in a parthenogenetic lizard Menetia greyii (Scincidae) and their utility in sexual species of the Menetia greyii complex

Tetra and tri‐nucleotide microsatellite loci were isolated from a parthenogenetic form of the Australian lizard Menetia greyii, using an enrichment method with polymerase chain reaction (PCR)‐based colony screening. Primers for 23 loci were designed and 11 of these loci amplified well in a panel of 10 parthenogenetic individuals from a single population. Seven loci were further characterized and six successfully amplified and were polymorphic in all five sexual species of the Menetia greyii complex they were tested in. These loci will be used to investigate the parental ancestry and clonal diversity of various parthenogenetic forms within the Menetia greyii species complex.     

Asexual and sexual hybrids between Fundulus diaphanus and F. heteroclitus in the Canadian Atlantic region

Levels and origins of clonal diversity in asexual hybrid animals are critical to understanding how they can coexist with their sexual progenitor species. In this study, asexual gynogenetic hybrids between Fundulus diaphanus and Fundulus heteroclitus known from two sites in Nova Scotia (Canada) were characterized using discriminant morphological traits, eight microsatellite loci, and mitochondrial DNA. Fifteen clonal genotypes were uncovered, all bearing the same F. diaphanus maternal haplotye. Each site harboured a different dominant clone along with several rarer clones that all appear to be of recent origin. Unexpectedly, highly introgressed sexually reproducing hybrids (0.25 > q > 0.75) were also detected. Sexual hybrids with maternal ascendance in either species were also found at three other sites in the Atlantic region. Based on a single meristic trait (scale counts), it is shown that asexual clones can be significantly more variable than populations of sexual parental species. Also, species are morphologically more alike when living in sympatry, suggesting that introgression may occur via sexual hybrids. Altogether, these results confirm and refine the available knowledge on this hybrid system, and indicate that hybridization is probably a more widespread phenomenon than suspected, with implications for the phenotypic variability of a widely used model species, F. heteroclitus.     

Difference in parasite load and nonspecific immune reaction between sexual and gynogenetic forms of Carassius auratus

For coexistence, the sexual form in sexual/asexual complexes needs short-term advantages that can compensate for the two-fold disadvantage of sex. Higher mortality in the asexual form due to a higher parasite load will provide an advantage to the sexual form. In Lake Suwa, Japan, the parasite load (Metagonnimus sp.; Trematoda) of triploid gynogenetic females of Carassius auratus was significantly higher than that of diploid sexual females. In an immunoassay using healthy wild fish that were conditioned for 1 month in laboratory tanks, the nitroblue tetrazolium (NBT) immune reaction of sexual females was significantly higher than that of gynogenetic females. The NBT activity indicates the abundance of oxygen radicals from phagocytes, and hence the level of immune activity of the phagocytes. We suggest that the higher parasite load of the gynogenetic form is in part due to the lower immune activity of the phagocytes (nonspecific immune reaction) in the gynogenetic form compared to the sexual form.     

Patterns of reproductive isolation within and between two Lygaeus species characterized by sexual conflicts over mating

Theory suggests that, under some circumstances, sexual conflict over mating can lead to divergent sexually antagonistic coevolution among populations for traits associated with mating, and that this can promote reproductive isolation and hence speciation. However, sexual conflict over mating may also select for traits (e.g. male willingness to mate) that enhance gene flow between populations, limiting population divergence. In the present study, we compare pre‐ and post‐mating isolation within and between two species characterized by male–female conflict over mating rate. We quantify sexual isolation among five populations of the seed bug Lygaeus equestris collected from Italy and Sweden, and two replicates of a population of the sister‐species Lygaeus simulans, also collected from Italy. We find no evidence of reproductive isolation amongst populations of L. equestris, suggesting that sexual conflict over mating has not led to population divergence in relevant mating traits in L. equestris. However, there was strong asymmetric pre‐mating isolation between L. equestris and L. simulans: male L. simulans were able to mate successfully with female L. equestris, whereas male L. equestris were largely unable to mate with female L. simulans. We found little evidence for strong post‐mating isolation between the two species, however, with hybrid F₂ offspring being produced. Our results suggest that sexual conflict over mating has not led to population divergence, and indeed perhaps supports the contrary theoretical prediction that male willingness to mate may retard speciation by promoting gene flow.     

Introgressive hybridization and morphological transgression in the contact zone between two Mediterranean Solea species

Hybrid zones provide natural experiments where new combinations of genotypes and phenotypes are produced. Studying the reshuffling of genotypes and remodeling of phenotypes in these zones is of particular interest to document the building of reproductive isolation and the possible emergence of transgressive phenotypes that can be a source of evolutionary novelties. Here, we specifically investigate the morphological variation patterns associated with introgressive hybridization between two species of sole, Solea senegalensis and Solea aegyptiaca. The relationship between genetic composition at nuclear loci and individual body shape variation was studied in four populations sampled across the hybrid zone located in northern Tunisia. A strong correlation between genetic and phenotypic variation was observed among all individuals but not within populations, including the two most admixed ones. Morphological convergence between parental species was observed close to the contact zone. Nevertheless, the samples taken closest to the hybrid zone also displayed deviant segregation of genotypes and phenotypes, as well as transgressive phenotypes. In these samples, deviant body shape variation could be partly attributed to a reduced condition index, and the distorted genetic composition was most likely due to missing allelic combinations. These results were interpreted as an indication of hybrid breakdown, which likely contributes to postmating reproductive isolation between the two species.     

Asymmetric hybridization between two species of sunfishes (Lepomis: Centrarchidae)

Many of the sunfishes (Centrarchidae) hybridize in natural systems, yet little is known about the interactions among hybridizing individuals in nature. We used allozyme electrophoresis to identify interspecific hybrids between bluegill (Lepomis macrochirus) and pumpkinseed (L. gibbosus) in Lake Opinicon, Ontario, and restriction fragment length polymorphism analysis of mitochondrial DNA (mtDNA) to identify the maternal parentage of those individuals. All 44 individuals collected and identified electrophoreti-cally as F₁ hybrids had a pumpkinseed mtDNA haplotype, indicating that hybridization between these species in Lake Opinicon is asymmetrical; F₁ hybrids result only from matings between female pumpkinseed and male bluegill.     

Cytogenetic relationships,polyploid origin and taxonomic issues in Paspalum species: inter‐ and intraspecific hybrids between a sexual synthetic autotetraploid and five wild apomictic tetraploid species

• Paspalum is a noteworthy grass genus due to the forage quality of most species, with approximately 330 species, and the high proportion of those that reproduce via apomixis. Harnessing apomictic reproduction and widening knowledge about the cytogenetic relationships among species are essential tools for plant breeding.
• We conducted cytogenetic analyses of inter‐ and intraspecific hybridisations involving a sexual, colchicine‐induced autotetraploid plant of P. plicatulum Michx. and five indigenous apomictic tetraploid (2n = 40) species: P. compressifolium Swallen, P. lenticulare Kunth, two accessions of P. nicorae Parodi, P. rojasii Hack. and two accessions of P. plicatulum. Fertility of the hybrids was investigated and their reproductive system was analysed considering the relative embryo:endosperm DNA content from flow cytometry. Morphological, nomenclatural and taxonomic issues were also analysed.
• Cytogenetic analysis suggested that all indigenous tetraploid accessions of five apomictic species are autotetraploid or segmental allotetraploid. If segmental allotetraploids, they probably originated through autoploidy followed by diploidisation processes. Autosyndetic male chromosome pairing observed in all hybrid families supported this assertion. Allosyndetic chromosome associations were also observed in all hybrid families. In the hybrids, the proportion of male parent chromosomes involved in allosyndesis per pollen mother cell varied from 5.5% to 35.0% and the maximum was between 25% and 60%. The apomictic condition of the indigenous male parents segregated in the hybrids.
• These results confirm a strong association between autoploidy and apomixis in Paspalum, and the existence of cytogenetic relationships between different species of the Plicatula group. Allosyndetic chromosome pairing and seed fertility of the hybrids suggest the feasibility of gene transfer among species.

     

Chromosomal inheritance of parental rDNAs distribution pattern detected by FISH in diploid F1 hybrid progeny of Cobitis (Teleostei,Cobitidae) species has non-Mendelian character

This study was conducted to describe the major and the minor rDNA chromosome distribution in the spined loach Cobitis taenia (2n = 48) and the Danubian loach Cobitis elongatoides (2n = 50), and their laboratory-produced diploid reciprocal F₁ hybrid progeny. It was tested by fluorescence in situ hybridisation (FISH) whether the number of 28s and 5s rDNA sites in the karyotypes of diploid hybrids corresponds to the expectations resulting from Mendelian ratio and if nucleolar organiser regions (NOR)were inherited from both parents or nucleolar dominance can be observed in the induced F₁ hybrid progeny. Ten (females) or twelve (males) 28s rDNA loci were located in nine uniarm chromosomes of C. taenia. Two of such loci terminally bounded on one acrocentric chromosome were unique and indicated as specific for this species. Large 5s rDNA clusters were located on two acrocentric chromosomes. In C. elongatoides of both sexes, six NOR sites in terminal regions on six meta-submetacentric chromosomes and two 5s rDNA sites on large submetacentrics were detected. The F₁ hybrid progeny (2n = 49) was characterised by the intermediate karyotype with the sites of ribosome synthesis on chromosomes inherited from both parents without showing nucleolar dominance. 5s rDNA sites were detected on large submetacentric and two acrocentric chromosomes. The observed number of both 28s and 5s rDNAs signals in F₁ diploid Cobitis hybrids was disproportionally inherited from the two parental species, showing inconsistency with the Mendelian ratios. The presented rDNA patterns indicate some marker chromosomes that allow the species of the parental male and female to be recognised in hybrid progeny. The 5s rDNA was found to be a particularly effective diagnostic marker of C. elongatoides to partially discern genomic composition of diploid Cobitis hybrids and presumably allopolyploids resulting from their backcrossing with one of the parental species. Thus, the current study provides insight into the extent of rDNA heredity in Cobitis chromosomes and their cytotaxonomic character.     

Reproductive biology and natural hybridization between two endemic species of Pitcairnia (Bromeliaceae)

We investigated pollination biology and breeding systems in hybridizing populations of Pitcairnia albiflos and P. staminea; both species are endemic to rocky outcrops at Rio de Janeiro, Brazil. These species are morphologically distinct and easily recognized by floral color: white in P. albiflos and red in P. staminea. Putative hybrids show a large range of intermediate pink floral colors. The showy hermaphroditic flowers offer pollen and nectar that attract many visitors including bees, butterflies, hawk moths, and bats. Although the flowers of both parental species and hybrids open at night, only P. albiflos had other adaptations for nocturnal pollination. Flowering times overlapped during three consecutive years of observation. Bees visited both species and putative hybrids. Cross-pollinations were performed within and among parental species and hybrids in a greenhouse using plants transplanted from the field. Pitcairnia staminea and hybrids are self-compatible and could be spontaneously self-pollinated, whereas P. albiflos, though self-compatible, needs pollinators' services for self-pollination. Facultative agamospermy was found in the parental species. Prezygotic and postzygotic reproductive barriers between these taxa were weak. Reciprocal hand-pollinations between parental species and with hybrids yielded high fruit sets with viable seeds. Evaluations of fruit set, seed set, seed germination, and pollen viability were undertaken to compare the fitness of the hybrids relative to their parents. The hybrids showed equivalent fitness, except for lower pollen viability. Some conservation implications are noted.     

Genome analysis of intergeneric hybrids of apomictic and sexual Australian Elymus species with wheat,barley and rye: implication for the transfer of apomixis to cereals

Summary Four hybrids were obtained between three Australian Elymus taxa and three cereal grains: wheat, rye, and barley. Mean meiotic metaphase-I configurations were 41.14 I, 0.42 rod II, 0.003 ring II, and 0.01 III for E. scabrus var plurinervis x Triticum aestivum (1 hybrid plant), 22.27 I, 2.63 rod II, 0.06 ring II, and 0.12 III for E. scabrus var scabrus x Secale cereale (4 hybrid plants), and 26.65 I, 0.66 rod II, 0.00 ring II, and 0.01 III for E. scabrus var plurinervis x Hordeum vulgare (13 hybrid plants). The I genome of barley also paired very little in a B_III hybrid of apomictic E. rectisetus x H. vulgare (2 hybrid plants). Megasporogenesis in this B_III hybrid was at least facultatively apomeiotic, with the same sort of nuclear elongation, apomeiotic division, and dyad formation seen previously in E. rectisetus itself. All four hybrid combinations were sterile. While spike morphology in the E. scabrus x T. aestivum and E. scabrus x H. vulgare hybrids were intermediate to their parents, E. scabrus x S. cereale and E. rectisetus x H. vulgare looked like their maternal parents.     

Unidirectional hybridization and reproductive barriers between two heterostylous primrose species in north-west Yunnan,China

Background and Aims

Heteromorphy in flowers has a profound effect on breeding patterns within a species, but little is known about how it affects reproductive barriers between species. The heterostylous genus Primula is very diverse in the Himalaya region, but hybrids there have been little researched. This study examines in detail a natural hybrid zone between P. beesiana and P. bulleyana.

Methods

Chloroplast sequencing, AFLP (amplified fragment length polymorphism) markers and morphological comparisons were employed to characterize putative hybrids in the field, using synthetic F₁s from hand pollination as controls. Pollinator visits to parent species and hybrids were observed in the field. Hand pollinations were conducted to compare pollen tube growth, seed production and seed viability for crosses involving different morphs, species and directions of crossing.

Key Results

Molecular data revealed all hybrid derivatives examined to be backcrosses of first or later generations towards P. bulleyana: all had the chloroplast DNA (cpDNA) of this species. Some individuals had morphological traits suggesting they were hybrids, but they were genetically similar to P. bulleyana; they might have been advanced generation backcrosses. Viable F₁s could not be produced with P. bulleyana pollen on P. beesiana females, irrespective of the flower morphs used. Within-morph crosses for each species had very low (<10 %) seed viability, whereas crosses between pin P. bulleyana (female) and pin P. beesiana had a higher seed viability of 30 %. Thus genetic incompatibility mechanisms back up mechanical barriers to within-morph crosses in each species, but are not the same between the two species. The two species share their main pollinators, and pollinators were observed to fly between P. bulleyana and hybrids, suggesting that pollinator behaviour may not be an important isolating factor.

Conclusions

Hybridization is strongly asymmetric, with P. bulleyana the only possible mother and all detected hybrids being backcrosses in this direction. Partial ecological isolation and inhibition of heterospecific pollen, and possibly complete barriers to F₁ formation on P. beesiana, may be enough to make F₁ formation very rare in these species. Therefore, with no F₁ detected, this hybrid zone may have a finite life span as successive generations become more similar to P. bulleyana.     

Inheritance of allozymes and hybridization in two European Tilia species

Inheritance analysis of seven enzyme systems in hexaploid Tilia cordata Mill. was performed utilizing single trees and their open-pollinated progenies. Genetic analyses of 12 polymorphic gene loci showed that T. cordata had disomic inheritance despite being an allopolyploid. T. platyphyllos Scop. was also used in the investigations although no genetic inheritance analysis was carried out. In comparison with T. cordata, zymograms of 10 spontaneous T. cordata x T. platyphyllos hybrids showed markedly different banding patterns with species-specific alleles at 13 of the 14 described gene loci. Hence, differentiation between both species and their naturally occurring hybrid (T. x europaea) is easily feasible with allozyme studies.