Cytogenetic diversity of SSR motifs within and between Hordeum species carrying the H genome: H. vulgare L. and H. bulbosum L.

Non-denaturing FISH (ND-FISH) was used to compare the distribution of four simple sequence repeats (SSRs)—(AG) _n , (AAG) _n , (ACT) _n and (ATC) _n —in somatic root tip metaphase spreads of 12 barley (H. vulgare ssp. vulgare) cultivars, seven lines of their wild progenitor H. vulgare ssp. spontaneum, and four lines of their close relative H. bulbosum, to determine whether the range of molecular diversity shown by these highly polymorphic sequences is reflected at the chromosome level. In both, the cultivated and wild barleys, clusters of AG and ATC repeats were invariant. In contrast, clusters of AAG and ACT showed polymorphism. Karyotypes were prepared after the identification of their seven pairs of homologous chromosomes. Variation between these homologues was only observed in one wild accession that showed the segregation of a reciprocal translocation involving chromosomes 5H and 7H. The two subspecies of H. vulgare analysed were no different in terms of their SSRs. Only AAG repeats were found clustered strongly on the chromosomes of all lines of H. bulbosum examined. Wide variation was seen between homologous chromosomes within and across these lines. These results are the first to provide insight into the cytogenetic diversity of SSRs in barley and its closest relatives. Differences in the abundance and distribution of each SSR analysed, between H. vulgare and H. bulbosum, suggest that these species do not share the same H genome, and support the idea that these species are not very closely related. Southern blotting experiments revealed the complex organization of these SSRs, supporting the findings made with ND-FISH.     

Ground-beetles of the subgenus Cryptophonus Brandm. et Z. Brandm., genus Harpalus Latr. (Coleoptera, Carabidae)

A taxonomic review of ten species of the subgenus Cryptophonus Brandmayr et Zetto Brandmayr, 1982, the genus Harpalus Latreille, 1802 is given. In addition to nine Palaearctic species, the Ethiopian H. agnatus Reiche, 1849 is also included in Cryptophonus. Harpalus cyrenaicus Koch, 1939, stat. n. from Libya, which was originally described as a subspecies of H. litigiosus Dejean, 1829, is treated as a distinct species. Harpalus tenebrosus Dejean, 1829 is treated as a polytypical species with two subspecies: H. t. tenebrosus (West Palaearctic) and H. t. paivanus Wollaston, 1867, stat. n. (Cabo Verde). A new substitute name H. melancholicus reicheianus nom. n. is proposed to replace the objective homonyms H. reichei Jacobson, 1907 (non Desbrochers des Loges, 1867) and H. ovalis Reiche, 1861 (non Motschulsky, 1844). Lectotypes are designated for H. tenebrosus Dejean, 1829, H. paivanus Wollaston, 1867, H. litigiosus Dejean, 1829, H. agnatus Reiche, 1849, H. fulvus Dejean, 1829, H. melancholicus Dejean, 1829, and H. ineditus Dejean, 1829. The status and diagnosis of Cryptophonus are discussed and a key to all the species of this subgenus is provided. Data on distribution of each species are provided. The following species are recorded for the first time: H. tenebrosus from Afghanistan, H. grilli Kataev, 2002 from India (Uttarhand), H. agnatus Reiche, 1849 from Tanzania, and H. fulvus Dejean, 1829 from Portugal.     

Copper tolerance in the cuprophyte Haumaniastrum katangense (S. Moore) P.A. Duvign. & Plancke

Cu tolerance and accumulation have been studied in Haumaniastrum katangense, a cuprophyte from Katanga (DR Congo), previously described as a copper hyperaccumulator. Nicotiana plumbaginifolia, a well-known non-tolerant and non-accumulator species, was used as a control. The germination rate of H. katangense was enhanced by copper and fungicide addition, suggesting that fungal pathogens, which restrain germination in normal conditions, are limiting. In hydroponic culture in the Hoagland medium, H. katangense did not grow well, in contrast to N. plumbaginifolia. Better growth was achieved by adding fungicide or higher copper concentrations. The maximal non-effective concentration (NEC) was 12 µM CuSO₄ for H. katangense grown in hydroponics, i.e. 24 times greater than Cu concentration in the Hoagland medium. By comparison, copper concentrations greater than 0.5 µM had a negative effect on the growth of N. plumbaginifolia. EC₅₀ (50% effective concentration) in hydroponics was 40 µM CuSO₄ for H. katangense and 6 µM CuSO₄ for N. plumbaginifolia. EC₁₀₀ (100% effective concentration) was 100 µM CuSO₄ for H. katangense and 15 µM CuSO₄ for N. plumbaginifolia. In soil, growth was also stimulated by Cu addition up to 300 mg kg^-1 CuSO₄. Surplus copper was also required for cultivating H. katangense in sterile conditions, suggesting that Cu excess may be necessary for needs other than pathogen defence. Cu accumulation in the shoot has been measured for N. plumbaginifolia and H. katangense at their respective NEC. Cu allocation in the two species showed a similar response to increasing Cu concentrations, i.e. root/shoot concentration ratio well above 1. In conclusion, H. katangense is highly tolerant to copper and has elevated copper requirement even in the absence of biotic interactions. Its accumulation pattern is typical of an excluder species.     

A review of flying fishes of the subgenus Hirundichthys (genus Hirundichthys, exocoetidae). 1. Oceanic species: H. speculiger, H. indicus sp. nova

A systematic revision of flying fishes of the subgenus Hirundichthys s.str was carried out based on a study of meristic and morphometric traits and characteristics of pigmentation of fishes from the local populations of species belonging to the subgenus. It is found out that the subgenus includes four species: oceanic H. speculiger from the Atlantic, Pacific and Indian Oceans, oceanic H. indicus sp.n. from the waters of the Indian Ocean, nerito-oceanic H. oxycephalus from the waters of the Indo-West Pacific and nerito-oceanic H. affinis from the Atlantic Ocean. The first part of the review focuses on two oceanic species with a large “mirror” on the pectoral fins: H. speculiger and H. indicus. A comparison of local populations showed that the species H. indicus is polytypic and consists of two subspecies. One of the subspecies—nominative H. indicus indicus—is distributed in the western and the central parts of the Indian Ocean and the other—H. indicus orientalis ssp.n.—in the Eastern Indian Ocean. Maps showing a geographical distribution of the species and the subspecies in the World Ocean are drawn up.     

二倍体杂交种棱果沙棘双向杂交起源及其母本主要来源于中国沙棘的分子证据简

棱果沙棘为同域分布的中国沙棘和肋果沙棘同倍化自然杂交形成的。本文利用母系遗传的cpDNA trnS-G序列检测青海祁连棱果沙棘及其亲本中国沙棘和肋果沙棘同域分布的两个地区（拱北湾、八宝河滩）共93个个体的遗传关系。结果表明棱果沙棘及其亲本在拱北湾和八宝河滩分别有12个和7个单倍型,两地区的棱果沙棘都与其亲本共享单倍型,其中拱北湾棱果沙棘共36个个体中有28个与中国沙棘共享3个单倍型(H2,H4,H5),有2个个体与肋果沙棘共享单倍型(H11),八宝河滩的棱果沙棘共10个个体中有7个与中国沙棘共享一个单倍型(H4),3个与肋果沙棘共享单倍型(H7)。应用最大简约法（MP）分别对两地区的棱果沙棘及其亲本trnS-G序列构建的系统发育树中棱果沙棘的大部分个体都与中国沙棘聚在一起,另外,棱果沙棘4种特有单倍型(H3、H7、H8、H9)的6个个体在系统树上也与中国沙棘聚为一支。以上结果进一步证明了二倍体自然杂交种棱果沙棘为双向杂交起源,但其主要母本来源应为中国沙棘。     

Susceptibility of larvae ofHeliothis zea,H. virescens,andH. armigera [lep.: Noctuidae] to 3 baculoviruses

The pattern of virulence (based on inclusion bodies) for 3 baculoviruses ofHeliothis, i.e. a unicapsid, nuclear polyhedrosis virus (HzSNPV); a multicapsid, nuclear polyhedrosis virus (HaMNPV); and a granulosis virus (HaGIV) was the same (HzSNPV>HaMNPV>HaGIV) for 3 species ofHeliothis. Based on numbers of nucleocapsids, however, the HaGIV was ca 2X more virulent than the HaMNPV for larvae ofH. virescens, (F.), and the HaMNPV was about 6X more virulent than the HaGIV for larvae ofH. armigera (Hübner). The fastest rate of larval mortality was obtained with HzSNPV. Although the mortality rate for HaGIV was faster than that of HaMNPV forH. virescens andH. armigera, it was slower than that of HaMNPV for larvae ofH. zea (Boddie). The pattern of susceptibility ofHeliothis species to HzSNPV and HaMNPV wasH. zea>H. virescens>H. armigera. Differences in susceptibility of the least susceptible species (H. armigera) and the most susceptible species (H. zea) to HzSNPV was ca. 1.6 X. Larvae ofH. zea, however, were ca. 4 to 6 X more susceptible to HaMNPV than were larvae ofH. virescens orH. armigera. A different pattern of susceptibility was recorded for HaGIV when larvae were challenged with HzSNPV and HaMNPV. Larvae ofH. virescens were ca. 20 and 35 X more susceptible to HaGIV than were larvae ofH. zea andH. armigera, respectively.     

Mzabimyces algeriensis gen. nov., sp. nov., a halophilic filamentous actinobacterium isolated from a Saharan soil,and proposal of Mzabimycetaceae fam. nov.

Three halophilic mycelium-forming actinobacteria, strains H195^T, H150 and H151, were isolated from a Saharan soil sample collected from Béni-isguen in the Mzab region (Ghardaïa, South of Algeria) and subjected to a polyphasic taxonomic characterisation. These strains were observed to show an aerial mycelium differentiated into coccoid spore chains and fragmented substrate mycelium. Comparative analysis of the 16S rRNA gene sequences revealed that the highest sequence similarities were to Saccharopolyspora qijiaojingensis YIM 91168^T (92.02 % to H195^T). Phylogenetic analyses showed that the strains H195^T, H150 and H151 represent a distinct phylogenetic lineage. The cell-wall hydrolysate was found to contain meso-diaminopimelic acid, and the diagnostic whole-cell sugars were identified as arabinose and galactose. The major cellular fatty acids were identified as iso-C_15:0, iso-C_16:0, iso-C_17:0 and anteiso-C_17:0. The diagnostic phospholipid detected was phosphatidylcholine and MK-9 (H₄) was found to be the predominant menaquinone. The genomic DNA G+C content of strain H195^T was 68.2 mol%. On the basis of its phenotypic features and phylogenetic position, we propose that strain H195^T represents a novel genus and species, Mzabimyces algeriensis gen. nov., sp. nov., within a new family, Mzabimycetaceae fam. nov. The type strain of M. algeriensis is strain H195^T (=DSM 46680^T = MTCC 12101^T).     

Structure and synthesis of a lipid-containing bacteriophage. XIX. Reconstitution of bacteriophage PM2 in vitro.

In order to construct a physical map of the bacteriophage fd genome, the doubly closed replicative form (RFI) DNA of phage fd was cleaved into unique fragments by four different restriction endonucleases (Hap, Hga, HinH and Hind) prepared from Haemophilus strains H. aphirophilus, H. gallinarum, H. influenzae H-I and H. influenzae Rd, respectively. As Hind cleaved RFI DNA at a single site, this site was used as a reference point for mapping. HinH cleaved RFI DNA at three sites, Hga at six sites and Hap at 13 sites, respectively. The 5′-termini of the fragments produced by either HinH or Hga were labelled with ³²P in the polynucleotide kinase reaction. The labelled fragments were separated and further cleaved by other enzymes. The re-digestion products of partially digested fragments were also analysed. On the basis of these data and estimates of the size of each fragment, a cleavage map of the phage fd genome was constructed.     

Determination of the ploidy level in the genusHieracium subgenusPilosella (Hill) S.F. Gray by flow cytometric DNA analysis

Ploidy levels of 8 taxa of the genusHieracium subgenusPilosella collected from the same locality were determined by means of flow cytometric estimation of relative nuclear DNA content. The DNA anaysis of isolated nuclei stained with propidium iodide was performed with a flow cytometer equipped with an argon-ion laser. The diploid speciesH. lactucella (2n=18) was used as internal standard. As a rule 20 individuals per taxon were analysed. The flow cytometric analysis proved to be a rapid, accurate and sensitive method for screening of ploidy levels. Considering the known cytological data, the following ploidy levels were determined:H. aurantiacum 4x,H. pilosella 4x,H. piloselloides 4x,H. bauhini 5x,H. caespitosum 5x,H. stoloniflorum 4x,H. brachiatum 4x and 6x,H. leptophyton 7x.H. piloselloides, H. bauhini andH. caespitosum yield relative DNA contents which are the exact multiples of the haploid DNA content ofH. lactucella. In the case ofH. pilosella the haploid DNA content is lower than that predicted from the diploid, namely 0.43 instead of the expected 0.5. The haploid 1x DNA contents of bothH. brachiatum andH. leptophyton range between the value found forH. pilosella and those for the other investigated species, confirming their hybrid origin.     

Flavonoids from Haplophyllum pedicellatum, H. robustum AND H. glabrinum

Haplophyllum pedicellatum, H. robustum and H. glabrinum all yielded the known compound gossypetin 8,3′-dimethyl ether 3-rutinoside. In addition the first two species afforded isorhamnetin and its 3-rutinoside. A new glycoside, gossypetin 8,3′-dimethyl ether 3-glucoside was obtained from H. pedicellatum together with the 3-malonylrutinoside, 3-malonylglucoside and 3-galactoside of isorhamnetin plus kaempferol 3-malonylglucoside. H. robustum yielded isorhamnetin 7-glucoside and 3-glucoside and quercetin 3-galactoside, while H. glabrinum was found to contain gossypetin 8-methyl ether 3-malonylrutinoside in addition to kaempferol and isorhamnetin 3-glucoside.     

Essential oil composition of Hypericum L. species from Southeastern Serbia and their chemotaxonomy

The essential oils of the aerial parts of nine species of Hypericum (Hypericum barbatum, Hypericum hirsutum, Hypericum linarioides, Hypericum maculatum, Hypericum olympicum, Hypericum perforatum, Hypericum richeri, Hypericum rumeliacum and Hypericum tetrapterum), collected from different locations in Southeast Serbia, were obtained by steam distillation and analyzed by GC and GC–MS. The essential oils investigated were characterized by a high content of non-terpene compounds and a low content of monoterpenes. The contents of non-terpenes, monoterpenes and sesquiterpenes in oils of the species H. barbatum, H. richeri and H. rumeliacum (section Drosocaprium) were similar and these oils were characterized by high contents of fatty acids. The oils of H. hirsutum and H. linarioides (section Taeniocarpium) contained a high percentage of n-nonane. There were similarities in contents of non-terpenes and sesquiterpenes in oils of species that belong to the section Hypericum (H. maculatum, H. perforatum and H. tetrapterum). The oil of H. olympicum differed from others by higher terpene content. A comparison was also carried out of the chemical composition of the essential oils from flower, leaf and stem of H. perforatum and it revealed that the highest concentration of non-terpene compounds was found in the flower and stem oil, while a high concentration of sesquiterpenes was characteristic for leaf oil. There were significant differences in the concentrations of the same compounds in the essential oils of H. maculatum, H. olympicum and H. perforatum, collected in different years from the same location which could be explained by seasonal differences. All data were statistically processed with principal component analysis and cluster analysis. The main conclusion from the above data is that genetic and environmental factors both play a role in determining the composition of essential oils of the Hypericum species studied.     

Development of T. aestivum L.-H. californicum Alien Chromosome Lines and Assignment of Homoeologous Groups of Hordeum californicum Chromosomes

Hordeum californicum （2n = 2x = 14, HH） is resistant to several wheat diseases and tolerant to lower nitrogen. In this study, a molecular karyotype of H. californicum chromosomes in the Triticum aestivum L. cv. Chinese Spring （CS）-H. californicum amphidiploid （2n = 6x = 56, AABBDDHH） was established. By genomic in situ hybridization （GISH） and multicolor fluorescent in situ hybridization （FISH） using repetitive DNA clones （pTa71, pTa794 and pSc119.2） as probes, the H. californicum chromosomes could be differentiated from each other and from the wheat chromosomes unequivocally. Based on molecular karyotype and marker analyses, 12 wheat--alien chromosome lines, including four disomic addition lines （DAH1, DAH3, DAH5 and DAH6）, five telosomic addition lines （MtH7L, MtHIS, MtH1L, DtH6S and DtH6L）, one multiple addition line involving H. californicum chromosome H2, one disomic substitution line （DSH4） and one translocation line （TH7S/1BL）, were identified from the progenies derived from the crosses of CS-H. californicum amphidiploid with common wheat varieties. A total of 482 EST （expressed sequence tag） or SSR （simple sequence repeat） markers specific for individual H. californicum chromosomes were identified, and 47, 50, 45, 49, 21, 51 and 40 markers were assigned to chromosomes H1, H2, H3, H4, H5, H6 and H7, respectively. According to the chromosome allocation of these markers, chromosomes H2, H3, H4, H5, and H7 of H. californicum have relationship with wheat homoeologous groups 5, 2, 6, 3, and 1, and hence could be designated as 5Hc, 2He, 6Hc, 3Hc and 1Hc, respectively. The chromosomes H1 and H6 were designated as 7Hc and 4Hc, respectively, by referring to SSR markers located on rye chromosomes.     

Analysis of chromosomal polymorphism in barley (Hordeum vulgare L. ssp. vulgare) and between H. vulgare and H. chilense using three-color fluorescence in situ hybridization (FISH)

The aim of the present work was to study chromosomal polymorphism within cultivated barley (Hordeum vulgare ssp. vulgare) using three-color fluorescence in situ hybridization (FISH). The physical distribution of the most frequently used, highly repetitive DNA sequences (GAA)₇ specific for pericentromeric heterochromatic regions, the ribosomal DNA clone pTa71, specific for the 45S rDNA, and the barley-specific telomere-associated sequence HvT01, was investigated to reveal genetic diversity in metaphase spreads of ten barley genotypes with diverse geographical origin, growth habit and row number. A wild relative of barley, Hordeum chilense was also studied in order to compare the polymorphism between and within Hordeum species. Significant differences in the hybridization patterns of all three DNA probes could be detected between the two related species, but only probes pTa71 and HvT01 showed variation in the intensity and/or position of hybridization sites among genotypes of H. vulgare ssp. vulgare. The extent of polymorphism was less than that earlier reported for molecular markers and was restricted to the long chromosome arms, with differences between the chromosomes. 1H and 3H proved to be the most variable chromosomes and 4H and 6H the most conserved.     

Helicotylenchus oleae n. sp. and H. neopaxilli n. sp. (Hoplolaimidae), Two New Spiral Nematodes Parasitic on Olive Trees in Italy

Helicotylenchus oleae n. sp. and H. neopaxilli n. sp., from olive roots and soil in Italy, are described and illustrated. Helicotylenchus oleae can be distinguished from the related species H. canadensis and H. tunisiensis especially by the smaller styler, its distinctive tail shape, and a tail longer than one anal body width. Helicotylenchus neopaxilli differs from the close species H. paxilli by having a conical, anteriorly truncated labial region, shorter stylet, and phasmids always anterior to level of anus. Also illustrated and discussed are histopathological changes within feeder roots of olive caused by the feeding activity of the semi-endoparasitic H. oleae.     

Interactions between embryogenic callus of Abies alba and Heterobasidion spp. in dual cultures

Interactions between three genotypes of a silver fir (Abies alba Mill.) embryogenic callus and Heterobasidion abietinum, H. parviporum, and H. annosum were examined in dual cultures. The aim of this study was to determine whether dual cultures can be used to evaluate the degree of fungal virulence at an embryogenic level, and whether different genotypes of a callus show different susceptibility. The dual cultures were performed on Schenk and Hildebrandt medium. Mycelial growth of H. parviporum and H. annosum was significantly stimulated in the presence of the callus but was not directional in nature. The embryogenic callus died between six and nine days after being colonized by H. parviporum suggesting that this was the most virulent species. By contrast, the callus remained healthy for up to 50 d after colonization with H. abietinum suggesting that this was the least virulent species. The callus of the A. alba genotype which originated in the mountain region of Poland remained healthy and alive for a significantly longer period than the other two genotypes in the dual cultures with all three Heterobasidion species even though overgrown by mycelium suggesting that the mountain genotype had the strongest defence response to Heterobasidion infection.     

Interactions of Concomitant Species of Nematodes and Fusarium oxysporum f. sp. vasinfectum on Cotton

Meloidogyne incognita, Hoplolaintus galeatus, and North Carolina and Georgia populations of Belonolaimus longicaudatus were introduced singly and in various combinations with Fusarium oxysporum f. sp. vasinfectum on wilt-susceptible ''Rowden'' cotton. Of all the nematodes, the combination of the N. C. population of B. longicaudatus with Fusarium promoted greatest wilt development. H. galeatus had no effect on wilt. With Fusarium plus M. incognito or B. longicaudatus, high nematode levels promoted greater wilt than low levels. The combination of either population of B. longicaudatus with M. incognita and Fusarium induced greater wilt development than comparable inoculum densities of either nematode alone or where H. galeatus was substituted for either of these nematodes. Nematode reproduction was inversely related to wilt development. Without Fusarium, however, the high inoculum level resulted in greater reproduction of all nematode species on cotton. Combining M. incognita with B. longicaudatus or H. galeatus gave mutually depressive effects on final nematode populations. The interactions of H. gateatus with B. longicaudatus varied with two populations of the latter.     

Establishment of a new hypotrichous genus,Heterotachysoma n. gen. and notes on the morphogenesis of Hemigastrostyla enigmatica (Ciliophora,Hypotrichia)

A marine hypotrich ciliate, Heterotachysoma multinucleatum (Gong and Choi, 2007) n. comb., found in coastal waters near Qingdao, China, was investigated. Heterotachysoma multinucleatum is characterized by its dorsal ciliature arranged in Gonostomum-pattern. Additionally, a new genus, Heterotachysoma n. gen., is established which is mainly characterized by: 18-cirri pattern; flexible body; three dorsal kineties with no dorsomarginal kineties nor kinety fragmentation; one right and one left row of marginal cirri; caudal cirri absent. The genus Tachysoma is redefined, and three new combinations, T. multinucleatum, T. ovatum and T. dragescoi, are proposed. The morphogenesis of Hemigastrostyla enigmatica (Dragesco and Dragesco-Kernéis, 1986) Song and Wilbert, 1997, is also described. Compared with that of its congeners, the differences are mainly in the dorsal ciliature: (1) the dorsal kinety anlagen are formed de novo in H. enigmatica (vs. intrakinetally in H. paraenigmatica and H. elongata); (2) the dorsal kineties anlagen develop in secondary mode in H. enigmatica (vs. primary mode in H. paraenigmatica); (3) the kinetal fragment anterior to the right marginal row in both filial product is absent in both H. enigmatica and H. elongata (vs. present in H. paraenigmatica). These findings suggest that morphogenesis is not uniform among members of the genus Hemigastrostyla.     

Chromosome numbers and DNA ploidy levels of selected species ofHieracium s.str. (Asteraceae)

Chromosome numbers and /or ploidy levels are reported for 44 species and subspecies ofHieracium s.str. from the following European countries: Andorra, Austria, Bulgaria, Czech Republic, France, Italy, Montenegro, Serbia, Slovakia, Slovenia, Spain, Sweden, Switzerland and Ukraine. The chromosome numbers/DNA ploidy levels ofH. bocconei (2n ～ 4x),H. bupleuroides subsp.leviceps (2n = 27),H. caesioides subsp.caesioides (2n = 27),H. basifolium (H. caesium agg., 2n = 36),H. plumbeum (H. caesium agg., 2n = 36),H. glaucum subsp.nipholepium (2n= 27),H.gouanii (2n = 18),H. gymnocerinthe (2n = 27),H. ramondii (2n = 27),H. recoderi (2n = 18),H. stelligerum (2n = 18), andH. tomentosum (2n = 18, 2n ～ 2x, 2n ～ 3x) were determined for the first time. New ploidy levels are reported forH. cerinthoides s.str. (2n = 27),H. humile (2n = 36), andH. tommasinianum (2n = 27).     

Interspecific hybridization in the genus Hieracium s. str.: evidence for bidirectional gene flow and spontaneous allopolyploidization

Although reticulation has indisputably played an important role in the evolutionary history of the genus Hieracium s. str. (Asteraceae), convincingly documented cases of recent interspecific hybridization are very rare. Here we report combined evidence on recent hybridization between two diploid species, Hieracium alpinum and H. transsilvanicum. The hybrid origin of the plants from the Romanian Eastern Carpathians was supported by additive patterns of nuclear ribosomal DNA polymorphism (ITS), an intermediate position of hybrid plants in principal coordinate analysis based on amplified fragment length polymorphism phenotypes (AFLP), and additivity at one allozyme locus. Flow cytometric analyses and chromosome counting showed that two hybrids were diploid (2n ~ 2x ~ 18) while one was surprisingly tetraploid (2n = 4x = 36). To our knowledge, this is the first record of spontaneous polyploidization following interspecific crossing in the genus. Allozyme data, especially the presence of unbalanced heterozygosity at one locus, suggest the origin of this tetraploid via a triploid bridge with subsequent backcrossing to H. alpinum. According to PCR-RFLP analyses of the trnT-trnL intergenic spacer, all H. ×krasani hybrids examined had the H. alpinum haplotype while H. transsilvanicum served as a pollen donor. The hybrids occurred at the locality with abundant H. alpinum plants where paternal H. transsilvanicum was missing. Previously reported instances of interspecific hybridization between the same parental taxa showed an opposite direction of crossing and relative abundance of parental taxa. This suggests that the direction of hybridization might be influenced by the frequency of parental taxa at the locality.