Phylogeny reconstruction of the yellowhammer (Emberiza citrinella) and pine bunting (Emberiza leucocephala) based on song and morphological characters

The yellowhammer and pine bunting represent a quite rare case of mass hybridization in a broad secondary contact zone. Previously, it was shown that the two forms did not differ by mitochondrial DNA (Irwin et al., 2009). This result allowed suggesting either a very close evolutionary relationship between these species or a consequence of borrowing the mitochondrial genome due to long-term hybridization. An attempt to reconstruct the phylogeny of these species was made on the basis of cluster analysis performed relative to the song characters and morphological features (male coloration and morphometric characters). The geographical variability of songs in yellowhammers and pine buntings was also investigated. The cladistic analysis included another 5 closely related bunting species: cirl (E. cirlus), chestnut-breasted (E. stewarti), ortolan (E. hortulana), grey-necked (E. buchanani), and Cretzschmar??s bunting (E. caesia). Based on the results of our analysis, the yellowhammer and pine bunting show no sister relationship. They compose the smallest clusters: one with the cirl bunting and the other one with the chestnut-breasted bunting. Thus, in this paper we attempt to prove the hypothesis of a high divergence level between the species and mtDNA introgression from the pine bunting into the yellowhammer following long term hybridization. The analysis of song dialects agrees with the assumption that there existed an ancient hybrid zone between the yellowhammer and the pine bunting in Europe.     

Mapping species differences for adventitious rooting in a <Emphasis Type="Italic">Corymbia torelliana</Emphasis> × <Emphasis Type="Italic">Corymbia citriodora</Emphasis> subspecies <Emphasis Type="Italic">variegata</Emphasis> hybrid

Quantitative trait loci (QTL) detection was carried out for adventitious rooting and associated propagation traits in a second-generation outbred Corymbia torelliana × Corymbia citriodora subspecies variegata hybrid family (n = 186). The parental species of this cross are divergent in their capacity to develop roots adventitiously on stem cuttings and their propensity to form lignotubers. For the ten traits studied, there was one or two QTL detected, with some QTL explaining large amounts of phenotypic variation (e.g. 66% for one QTL for percentage rooting), suggesting that major effects influence rooting in this cross. Collocation of QTL for many strongly genetically correlated rooting traits to a single region on linkage group 12 suggested pleiotropy. A three locus model was most parsimonious for linkage group 12, however, as differences in QTL position and lower genetic correlations suggested separate loci for each of the traits of shoot production and root initiation. Species differences were thought to be the major source of phenotypic variation for some rooting rate and root quality traits because of the major QTL effects and up to 59-fold larger homospecific deviations (attributed to species differences) relative to heterospecific deviations (attributed to standing variation within species) evident at some QTL for these traits. A large homospecific/heterospecific ratio at major QTL suggested that the gene action evident in one cross may be indicative of gene action more broadly in hybrids between these species for some traits.     

Outbreeding causes developmental instability in <Emphasis Type="Italic">Drosophila subobscura</Emphasis>

A possible effect of interpopulation hybridization is either outbreeding depression, as a consequence of breakdown of coadapted gene complexes which can increase developmental instability (DI) of the traits, or increased heterozygosity, which can reduce DI. One of the principal methods commonly used to estimate DI is the variability of fluctuating asymmetry (FA). We analysed the effect of interpopulation hybridization in Drosophila subobscura through the variability in the wing size and the FA of wing length and width for both sexes in parental, F1 and F2 generations. The results of the wing size per se in intra- and interpopulation hybrids of D. subobscura do not explicitly reveal the significance of either of the two hypotheses. However, the results of the FA of the wing traits give a different insight. The FA of wing length and width generally increases in interpopulation crosses in F1 with respect to the FA in the parental generation, which suggests the possibility that outbreeding depression occurred in the first generation after the hybridization event. We generally observed that the FA values for the wing length and width of interpopulation hybrids were higher in F1 and F2 generations, compared to intrapopulation hybrids in same generations. These results suggest that the association between coadaptive genes with the same evolutionary history are the most probable mechanism that maintains the developmental homeostasis in Drosophila subobscura populations.     

Molecular characterization and mating type analysis of oyster mushroom (<Emphasis Type="Italic">Pleurotus</Emphasis> spp<Emphasis Type="Italic">.</Emphasis>) using single basidiospores for strain improvement

Existence of variability in morphological traits and growth rate of mycelium of homokaryotic single basidiospores can be exploited for the development of inter-strainal hybrids. We isolated 182 single basidiospores from mushroom bodies of P. sajor-caju, P. florida, P. eous and one wild relative of Pleurotus called Hypsizygus ulmaris. The single spores were isolated using a new technique that is less prone to contamination and more efficient than the common techniques used by earlier workers. All the isolates showed a varied range of cultural morphology. Mating types of all the isolates within the species were identified on the basis of hyphal fusion via anastomosis with the tester strains. Four compatible pairs of isolates with well prominent tuft in the contact zone were selected for dikaryon isolation. Dikaryons were used for spawn preparation and mushroom cultivation. The dikaryotic isolates with their replicates were evaluated for spawn run period, yield and biological efficiency. 42 isolates (10 di- and 32 mono-karyotic isolates) were analyzed with RAPD genetic markers. Phenotypic characters and mating types of all the 42 isolates analyzed genetically were correlated with their genetic polymorphism data. The isolates showed very large diversity both at the phenotypic and the genotypic level. Available phenotypic and genotypic data can further help in the selection of monosporous isolates for developing inter-strainal hybrids which can lead to better prospects for genetic improvement in different species of Pleurotus.     

Vocal Diversity and Taxonomy of <Emphasis Type="Italic">Nomascus</Emphasis> in Central Vietnam and Southern Laos

Previous researchers suggested that gibbon song repertoire is genetically determined and song characteristics are useful for assessing systematic relationships. The southern white-cheeked crested gibbon is regarded as either a subspecies of Nomascus leucogenys or its own species (Nomascus siki). I studied vocal diversity among different wild populations of Nomascus in central Vietnam and southern Laos to assess their taxonomic relationships and to examine whether their vocal patterns correspond to forms previously described for Nomascus siki. I examined the songs of 7 Nomascus populations in Vietnam and Laos. I analyzed 192 song bouts from different gibbon groups including 173 phrases of 42 females and 192 phrases of 42 males. Linear discriminant analysis, classification trees, and multidimensional scaling revealed marked separation of groups in the northern and southern populations. Within the 2 geographic populations, there is little variability and the vocal characteristics exhibited no apparent cline. I conclude that the northern and southern geographic populations may represent 2 distinct taxa. I postulate that a taxonal boundary such as large rivers existing between southern Quang Binh province and northern Thua-Thien Hue province in Vietnam and northern Phou Xang He NBCA and southern Dong Phou Vieng NBCA in Laos has limited gene flow between the populations. Differing topographic features could also serve as a selective force for improved sound transmission in a highly territorial species, driving the divergence between the 2 populations.     

Genetic analysis of sable (<Emphasis Type="Italic">Martes zibellina</Emphasis>) and pine marten (<Emphasis Type="Italic">M. martes</Emphasis>) populations in sympatric part of distribution area in the northern Urals

Analysis of nucleotide sequences of the mitochondrial DNA (mtDNA) control region (495 bp) of sables (Martes zibellina) and pine martens (M. martes) from allopatric parts of the species ranges has shown a considerable interspecific genetic distance (>3%). In sympatric populations of these species in the northern Urals, differences between two species-specific mtDNA lineages are still large; however, classification of each individual nucleotide sequence with one of the two lineages is not correlated with whether the given animal is phenotypically a sable, a pine marten, or a potential hybrid (the so-called “kidas”). This indicates a high degree of reciprocal introgression of the sable and pine marten mtDNA in the northern Urals and suggests that their interspecific hybridization is common in the sympatric zone.     

Revision of “<Emphasis Type="Italic">Septonema ochraceum</Emphasis>” revealed three new species of <Emphasis Type="Italic">Venturiaceae</Emphasis> and <Emphasis Type="Italic">Herpotrichiellaceae</Emphasis>

Survey of seven strains determined as Septonema ochraceum (Dothideomycetes, inc. sed.) isolated from pine litter or obtained from public collections revealed three new species, Fusicladium cordae, F. sicilianum (Venturiaceae), Cladophialophora matsushimae (Herpotrichiellaceae) and a cryptic species morphologically identical to Devriesia americana (Teratosphaeriaceae), but phylogenetically distinct. Morphological survey and phylogenetic analysis using nucleotide sequence data from the nuclear ribosomal subunit genes indicate a close relationship within three species colonising pine litter needles, F. cordae, F. pini and F. ramoconidii. F. sicilianum is most related to F. rhodense. C. matsushimae represents a species belonging to one of the lineages of the polyphyletic genus Cladophialophora. None of the strains observed can be classified morphologically as S. ochraceum, of which the type material does not exist.     

Interspecific hybridization in wild roses (<Emphasis Type="Italic">Rosa</Emphasis> L. sect. Caninae DC.)

Variability of morphological characters and Inter Simple Sequence Repeat (ISSR) markers is studied in a mixed multispecies sympatric growth of wild roses in Lugansk province of Ukraine. Several methods of data analysis are used, such as Principal Coordinates Analysis, cluster analysis (UPGMA), and Bayesian analyses of population structure with the Structure 2.2 and NewHybrids programs. The results indicate that there are four species growing at the site, viz. R. canina, R. villosa, R. dimorpha, and R. podolica, as well as various hybrids between them. The hybrid nature of the specimens is confirmed by molecular data, however they are not intermediate between the parental species as to their morphology. The hybrids either coincide morphologically with one of the parents, or demonstrate novel combinations of diagnostic characters.     

Chromosomes are eliminated in the symmetric fusion between <Emphasis Type="Italic">Arabidopsis </Emphasis><Emphasis Type="Italic">thaliana</Emphasis> L. and <Emphasis Type="Italic">Bupleurum scorzonerifolium</Emphasis> Willd.

In order to investigate chromosome elimination in symmetric somatic hybridization between Bupleurum scorzonerifolium and Arabidopsis thaliana, protoplasts were isolated from suspension cultures of both A. thaliana and B. scorzonerifolium parents. Biparental protoplasts were mixed at a rate of 1.5:1 and fused with PEG-method. After protoplast fusion, the products were cultured in the P5 liquid medium for microcallus formation. Single cell lines formed from microcalli after subculturing on the MB1 (Xia and Chen, Plant Sci 120:197–203, 1996) solid medium. The putative somatic hybrid cell lines were identified by cytological and molecular analysis. Of the 132 somatic cell lines generated, 16 were identified as somatic hybrids, with the phenotypes resembled B. scorzonerifolium parent. These hybrids showed a complete set of B. scorzonerifolium chromosome and 0–2 small chromosome(s) of A. thaliana. A few of them showed nuclear and cytoplasmic SSR fragments of A. thaliana. These hybrid cell lines could differentiate to green spots, buds/leaves through complementation of regeneration ability. The chromosomes elimination of A. thaliana was discussed. Wang Minqin and Zhao Junsheng contributed equally to the work.     

The viability of the first generation roach (<Emphasis Type="Italic">Rutilus rutilus</Emphasis> L.), bream (<Emphasis Type="Italic">Abramis brama</Emphasis> L.), and blue bream (<Emphasis Type="Italic">Abramis ballerus</Emphasis> L.) hybrids at early stages of development

The viability of bream, roach, and blue bream F ₁hybrids at early stages of their development is analyzed. Viability is controlled according to parameters of the survival rate at stages from fertilized eggs to yearlings. During embryogenesis, significant stages (blastula—gastrula and hatching) are revealed by the amount of losses. The viability of hybrids of the first generation (compared to pure species) from the hatching stage and in the subsequent development constantly increases. At the stage of fingerlings, the viability of F ₁hybrids significantly exceeds that of pure species, which points to the heterozygous effect according to the parameters of hybrid survival for the first generation, which is absent in embryogenesis.     

Observations on a hybrid between the whooper swan (<Emphasis Type="Italic">Cygnus cygnus</Emphasis>) and the mute swan (<Emphasis Type="Italic">Cygnus olor</Emphasis>)

The swan obtained by hybridization between Cygnus cygnus and C.olor was delivered to the Moscow Zoo from the Leningrad Zoo in 1989. Between 2002 and 2004, it permanently remained with a female whooper swan in a large pond. The two swans behaved as a breeding pair, although the sex of the hybrid bird remained unknown. We observed no courtship displays addressed by the hybrid swan either to the female whooper swan or to other swans living in the pond. The hybrid swan was exteriorly closer to C. cygnus rather than to C. olor and showed increased aggressiveness. Its threatening behavior was intermediate between parental species in terms of threat displays and motor activity, although behavioral patterns characteristic of C.olor prevailed. The most frequent sounds in the vocal repertoire of the hybrid swan were close to the vibrant throbbing call of C.olor in terms of duration and frequency range. In contrast to these sounds, the call of the hybrid swan was noiselike, with distinct harmonic structure (as in the whooper swan except for extremely narrow interharmonic intervals). These vocal signals of the hybrid swan were shown to be intermediate between those of the parental species. In addition, the acoustic repertoire of the hybrid swan included rarely emitted sounds closely resembling elements of vocalizations of both C. cygnus and C. olor.     

Isolation mechanisms in closely related grasshopper species <Emphasis Type="Italic">Chorthippus albomarginatus</Emphasis> and <Emphasis Type="Italic">Ch. oschei</Emphasis> (Orthoptera: Acrididae)

Two closely related grasshopper species Chorthippus albomarginatus and Ch. oschei are known to hybridize in the narrow contact zone at the territory of Ukraine and Moldova. Different isolaton mechanisms providing reproductive isolation between the two species were studied. In choice mating experiments, females of the both species demonstrated a strong assortative mating (80–90% preference for the conspecific males). Comparison of the parental and hybrid viability revealed a reduced hatching and increased larval mortality in F₁ and F₂ hybrids. In choice mating experiments, the hybrid females mated less assortatively than the parental females. An assymmetry was found in mating preferences and in viability of hybrids. The results demonstrate the existence of pre-and post-mating isolation between Ch. albomarginatus and Ch. oschei. A possible fate of the hybrid zone is discussed.     

Analysis of secondary sex ratio in the fox (<Emphasis Type="Italic">Vulpes vulpes</Emphasis> L.)

Secondary sex ratio and its variability in relation to some paratypic and genetic factors were studied in the silver fox by analysis of data obtained at the Pushkinskii fur farm (Moscow oblast) in 1980–1989. A total of 17285 whelps were examined. The mean proportion of males over the ten years of observation was 0.536 ± 0.004. No effect of parent age or litter size on this proportion was found. Individual analysis of the progeny of a single parent revealed 44 males and 49 females showing significant deviations from the expected sex ratio (1 ♂: 1 ♀). These results can be used for reconstruction of pedigrees whose progeny yields regular deviations from the expected sex ratio.     

Cytogenetic analysis of reciprocal hybrids and their parents between <Emphasis Type="Italic">Larix leptolepis</Emphasis> and <Emphasis Type="Italic">Larix gmelinii</Emphasis>: implications for identifying hybrids

We studied the parental taxa and the interspecific reciprocal hybrids between Larix leptolepis with Larix gmelinii, using classical cytogenetic methods, as well as fluorescence in situ hybridization (FISH) and genomic in situ hybridization. A high frequency (>90%) of complete bivalent formation was observed in reciprocal hybrids. Less than 10% of pollen mother cells exhibited abnormalities. The most frequent abnormalities were bridges. Multivalent chromosome associations were also observed in both reciprocal hybrids, suggesting that some chromosome interchange events did occur, and introgressions from one to the other species were possible. Intergenomic recombination indicates that genes might be readily introgressed into one species from the other in the genus Larix. Interspecific hybridization may be a potential method for genetic improvement in larch. FISH markers documented that the recombinant genomes of reciprocal hybrids were strictly additive and stable, indicating that FISH also might be a useful tool in Larix breeding.     

Intraspecific variability of triterpene content in the leaves of <Emphasis Type="Italic">Betula pendula</Emphasis> Roth

Changes in the content of dammarane-type triterpene alcohols in the leaves of silver birch (Betula pendula Roth) are studied for the first time along a zonal-climatic transect from the northern to southern borders of the species areal in the Trans-Ural region. It is shown that populations of northern and southern regions differ concerning the triterpene content in leaves. A correlation between the content of triterpene alcohols in silver birch leaves and some climatic parameters is revealed. At the same time, the range of the intrapopulation variability of triterpene content is significantly less than that of the interpopulation variability, which indicates a high potential of population selection on the basis of this biochemical parameter.     

Multiple host-ant use by the predatory social parasite <Emphasis Type="Italic">Phengaris</Emphasis> (=<Emphasis Type="Italic">Maculinea</Emphasis>) <Emphasis Type="Italic">arion</Emphasis> (Lepidoptera,Lycaenidae)

Phengaris (=Maculinea) arion is an endangered social parasite of Myrmica ants, and for a very long time was considered as specific to Myrmica sabuleti. Previous studies carried out in Poland suggested some discrepancies within this assumption, and therefore a much more intensive survey was undertaken. The host ant use of P. arion was studied at five sites in different types of biotopes in Poland, i.e. xerothermal grasslands where Thymus pulegioides was used as a larval food plant by the butterfly, and more or less sandy biotopes with Thymus serpyllum. Altogether nine Myrmica species were recorded, and considerable variation in species composition and density of nests was recorded. At four localities M. sabuleti proved to be the most common ant. A total of 529 Myrmica nests were examined, and only 20 of them contained larvae and pupae of P. arion. Host ants belonged to five different species, i.e. M. sabuleti, Myrmica scabrinodis, Myrmica schencki, Myrmica lobicornis and Myrmica hellenica. Only at one site (NE Poland) was a significant heterogeneity in parasitation rates among Myrmica species detected. M. lobicornis was the most often infested ant there, which may suggest local specialisation of the butterfly. Overall low parasitism rates may explain the vulnerability of P. arion in Central Europe but further studies are also necessary.     

Reticulate hybridization of <Emphasis Type="Italic">Alpinia</Emphasis> (Zingiberaceae) in Taiwan

Reticulate hybridization is a complicated and creative mechanism in plant evolution that can cause interference in phylogenetic studies. Based on observations of intermediate morphology, low pollen fertility, and overlapping distributions of putative parent species, Yang and Wang (Proceedings of the cross-strait symposium on floristic diversity and conservation. National Museum of Natural Science, Taichung, Taiwan, pp 183–197, 1998) first proposed reticulate hybridization of Alpinia in Taiwan. In the present study, molecular tools were used to explore relationships between four parental species and their homoploidy hybrids, and the impact of hybridization on phylogeny reconstruction. Based on DNA markers, maternal heritance of the chloroplast genome, and additivity of nuclear ribosomal internal transcribed spacer, the present results provide strong support for the hybridization hypothesis. Co-existence of parental ribotypes within hybrids revealed that these hybridization events were current, while reciprocal and introgressive hybridization were inferred from chloroplast DNA data. Furthermore, iterative hybridizations involving more than two parental species may occur in notorious hybrid zones. Ecological, phenological, and physiological evidence provides insight into why such frequent hybridization occurs in Taiwanese Alpinia. In the phylogenetic tree of the Zerumbet clade reconstructed in this study, the chloroplast sequences from one hybrid species were not grouped into a subclade, implying instability caused by hybridization. Failure to find morphological apomorphies and biogeographical patterns in this clade was likely partially due to reticulate hybridization. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Transgressive segregation of primary and secondary metabolites in F<Subscript>2</Subscript> hybrids between <Emphasis Type="Italic">Jacobaea aquatica</Emphasis> and <Emphasis Type="Italic">J. vulgaris</Emphasis>

Hybridization between plant species can have a number of biological consequences; interspecific hybridization has been tied to speciation events, biological invasions, and diversification at the level of genes, metabolites, and phenotypes. This study aims to provide evidence of transgressive segregation in the expression of primary and secondary metabolites in hybrids between Jacobaea vulgaris and J. aquaticus using an NMR-based metabolomic profiling approach. A number of F₂ hybrid genotypes exhibited metabolomic profiles that were outside the range encompassed by parental species. Expression of a number of primary and secondary metabolites, including jacaronone analogues, chlorogenic acid, sucrose, glucose, malic acid, and two amino acids was extreme in some F₂ hybrid genotypes compared to parental genotypes, and citric acid was expressed in highest concentrations in J. vulgaris. Metabolomic profiling based on NMR is a useful tool for quantifying genetically controlled differences between major primary and secondary metabolites among plant genotypes. Interspecific plant hybrids in general, and specifically hybrids between J. vulgaris and J. aquatica, will be useful for disentangling the ecological role of suites of primary and secondary metabolites in plants, because interspecific hybridization generates extreme metabolomic diversity compared to that normally observed between parental genotypes.     

<Emphasis Type="Italic">Strobilanthes pothigaiensis</Emphasis> (<Emphasis Type="Italic">Acanthaceae</Emphasis>) — a new species from southern Western Ghats,India

Summary   Strobilanthes pothigaiensis, a new species of Strobilanthes (Acanthaceae) allied to S. foliosus T. Anderson from southern Western Ghats, India, is described and illustrated.     

Morphometry of non-native black bullhead <Emphasis Type="Italic">Ameiurus melas</Emphasis> from Slovakia

The study on the external morphology of the non-native black bullhead (Ameiurus melas) was carried out on a population from Slovakia, based on triple regression and geometrical analysis. The breakpoints distribution in distance-based morphometric characters indicated that black bullhead reached its definitive phenotype early in ontogeny. Ontogenetic changes in external morphology occurred continuously throughout the whole size-range of the sample examined, and inter-individual morphological variation was very low. Such uniformity may reflect the possible founder effect and/or little phenotypic plasticity of this non-native population. No sexual dimorphism was observed. This is the first detailed study on external morphology of the species, and as such it contributes to the assessment of phenotypic plasticity and/or overall morphological variability of black bullhead’s invasive populations.