Studies on bucephalid digeneans parasitising molluscs and fishes in Finland I. Ecological data and experimental studies

Two types of bucephalid cercariae are reported from the bivalve Anodonta anatina in two Finnish lakes. One, Type A, resembles in gross morphology the cercaria of Bucephalus polymorphus, and the other, Type B, resembles the cercaria of Rhipidocotyle campanula. Type A daughter-sporocysts develop more slowly, have a greater cercarial productivity and exhibit a differential diurnal rhythm to that of Type B. Cercariae of Type A have a shorter longevity than Type B and tend to encyst in the fins rather than the gill-arches of fish intermediate hosts. The main definitive host of Type A is pike Esox lucius and, in the case of Type B, perch Perca fluviatilis. Adults of Types A and B are morphologically very similar and both species belong to the genus Rhipidocotyle.     

Studies on bucephalid digeneans parasitising molluscs and fishes in Finland. II. The description of Rhipidocotyle fennica n. sp. and its discrimination by principal components analysis

Rhipidocotyle fennica n. sp. (= Rhipidocotyle Type A of Taskinen et al., 1991) from the intestine of Esox lucius in central Finland is described and compared by means of a principal components analysis (PCA) with R. campanula (= Rhipidocotyle Type B of Taskinen et al., 1991). Its cercaria develops in the bivalve Anodonta anatina and the metacercaria occurs in the skin and fins of Rutilus rutilus. The metacercaria is discriminated from that of R. campanula by PCA and is described along with aspects of the chaetotaxy of the cercaria. The new species is distinguished from R. campanula, R. kovalae, R. papillosa and R. septpapillata.     

Clarification of the systematic position of Cercariaeum crassum Wesenberg-Lund, 1934 (Digenea), based on karyological analysis and DNA sequences

Chromosome set and rDNA sequences of the larval digenean Cercariaeum crassum were analysed in order to clarify its systematic position and possible adult form. Parasites were obtained from the sphaeriid bivalve Pisidium amnicum, collected in Lithuanian and Finnish rivers. The karyotype is shown to consist of five pairs (2n = 10) of large, up to 14 μm, chromosomes. Complement, composed of a low diploid number of exclusively bi-armed elements, presumably arose through Robertsonian fusions of acrocentric chromosomes. Consistent with a Robertsonian-derived karyotype, one or two small, metacentric, mitotically stable B chromosomes were detected in the cells of parthenitae isolated from some host individuals. A phylogenetic analysis using rDNA internal transcribed spacer 2 (ITS2) and 28S sequences corroborates the allocation of C. crassum to the family Allocreadiidae. In neighbour-joining and maximum parsimony phylogenetic trees C. crassum clusters into one clade with Allocreadium spp., and is the closest sister group in relation to A. isoporum; the level of rDNA sequence divergence between them (2.67% for ITS2 and 1.16% for 28S) is consistent with the level expected for intrageneric variation. The present study adds significant information to a database for establishing species-specific characters for confident characterization of different developmental stages of allocreadiid species, clarification of their life cycles and evaluation of intra- and interspecific variability.     

Four closely related but forgotten species of Rhipidocotyle Diesing, 1858 (Digenea: Bucephalidae) in fishes from European seas

The following species of Rhipidocotyle are described: R. minima (Wagener, 1852) from Chelidonichthys gurnardus, C. lastoviza and Aspitrigla cuculus at various localities off the British Isles; R. nicolli n. sp. from A. cuculus off Plymouth, SW England; R. triglae (van Beneden, 1870) from C. lucernus in the Gulf of Marseilles, western Mediterranean; and R. viperae (van Beneden, 1870) from Echiichthys vipera at various localities off the British Isles. The distinguishing features of the species are discussed in detail. A list of the bucephalid species reported from the Mediterranean Sea is appended.     

Samia cynthia versus Bombyx mori: comparative gene mapping between a species with a low-number karyotype and the model species of Lepidoptera

We performed gene-based comparative FISH mapping between a wild silkmoth, Samia cynthia ssp. with a low number of chromosomes (2n = 25-28) and the model species, Bombyx mori (2n = 56), in order to identify the genomic components that make up the chromosomes in a low-number karyotype. Mapping of 64 fosmid probes containing orthologs of B. mori genes revealed that the homologues of either two or four B. mori chromosomes constitute the S. c. ricini (Vietnam population, 2n = 27??/28??, Z0/ZZ) autosomes. Where tested, even the gene order was conserved between S. c. ricini and B. mori. This was also true for the originally autosomal parts of the neo-sex chromosomes in S. c. walkeri (Sapporo population, 2n = 26??/26??, neo-Wneo-Z/neo-Zneo-Z) and S. cynthia subsp. indet. (Nagano population, 2n = 25??/26??, neo-WZ₁Z₂/Z₁Z₁Z₂Z₂). The results are evidence for an internal stability of lepidopteran chromosomes even when all autosomes had undergone fusion processes to form a low-number karyotype.     

Weitere Untersuchungen über Chromosomenverhältnisse und DNS-Gehalt bei Anuren (Amphibia)

The karyotypes of the toad Bufo marinus L. (2n=22) and the frogs Limnodynastes tasmaniensis Gthr. (2n=24), Rana temporaria L., R. esculenta L. (both 2n=26) and R. arvalis Nills. (2n=24) were analysed in colchicine treated leukocyte and spermatogonial metaphases and/or embryonic and larval mitoses. The DNA content of Feulgen stained erythrocyte nuclei was measured microspectrophotometrically. Heteromorphic sex chromosomes are absent in all species. L. tasmaniensis has the lowest DNA content among these species. The south American toad B. marinus shows a karyotype similar to the other known toad species and contains the same amount of DNA as the European species B. calamita with the lowest DNA amount among the European toads. In southern German populations of R. temporaria besides animals with the “standard”-karyotype (2n=26) individuals with 1 or 2, in rare cases with 3 or 4 supernumerary chromosomes have been found. The supernumeraries are heterochromatic and smaller than the smallest chromosome of the “standard”-karyotype. If only 1 or 2 supernumerary chromosomes are present, they seem to show normal mendelian inheritance as a rule. The observation of a few tadpoles with intraindividual different numbers of supernumeraries points to the occurrence of unequal distribution of these chromosomes in individuals containing a higher number of supernumerary chromosomes. The karyotype of R. esculenta is very similar to the “standard”-karyotype of R. temporaria, but the chromosomes of R. esculenta are somewhat longer than those of R. temporaria. R. esculenta contains about 54% more DNA than R. temporaria in the erythrocyte nuclei, so that it must be assumed that all chromosomes of R. esculenta contain more DNA than their homologues in R. temporaria. R. arvalis possesses about 28% more DNA than R. temporaria. It is supposed that these interspecific differences in DNA content of the Rana species — as observed earlier in Bufo species — are not a consequence of differential polyteny but are caused during evolutionary processes by local increase in DNA in the chromosomes of R. esculenta and R. arvalis.     

Repetitive DNA Sequences and Evolution of ZZ/ZW Sex Chromosomes in Characidium (Teleostei: Characiformes)

Characidium constitutes an interesting model for cytogenetic studies, since a large degree of karyotype variation has been detected in this group, like the presence/absence of sex and supernumerary chromosomes and variable distribution of repetitive sequences in different species/populations. In this study, we performed a comparative cytogenetic analysis in 13 Characidium species collected at different South American river basins in order to investigate the karyotype diversification in this group. Chromosome analyses involved the karyotype characterization, cytogenetic mapping of repetitive DNA sequences and cross-species chromosome painting using a W-specific probe obtained in a previous study from Characidium gomesi. Our results evidenced a conserved diploid chromosome number of 2n = 50, and almost all the species exhibited homeologous ZZ/ZW sex chromosomes in different stages of differentiation, except C. cf. zebra, C. tenue, C. xavante and C. stigmosum. Notably, some ZZ/ZW sex chromosomes showed 5S and/or 18S rDNA clusters, while no U2 snDNA sites could be detected in the sex chromosomes, being restricted to a single chromosome pair in almost all the analyzed species. In addition, the species Characidium sp. aff. C. vidali showed B chromosomes with an inter-individual variation of 1 to 4 supernumerary chromosomes per cell. Notably, these B chromosomes share sequences with the W-specific probe, providing insights about their origin. Results presented here further confirm the extensive karyotype diversity within Characidium in contrast with a conserved diploid chromosome number. Such chromosome differences seem to constitute a significant reproductive barrier, since several sympatric Characidium species had been described during the last few years and no interespecific hybrids were found.     

Chromosomal Locations of 5S and 45S rDNA in Gossypium Genus and Its Phylogenetic Implications Revealed by FISH

We investigated the locations of 5S and 45S rDNA in Gossypium diploid A, B, D, E, F, G genomes and tetraploid genome (AD) using multi-probe fluorescent in situ hybridization (FISH) for evolution analysis in Gossypium genus. The rDNA numbers and sizes, and synteny relationships between 5S and 45S were revealed using 5S and 45S as double-probe for all species, and the rDNA-bearing chromosomes were identified for A, D and AD genomes with one more probe that is single-chromosome-specific BAC clone from G. hirsutum (A₁D₁). Two to four 45S and one 5S loci were found in diploid-species except two 5S loci in G . incanum (E₄), the same as that in tetraploid species. The 45S on the 7th and 9th chromosomes and the 5S on the 9th chromosomes seemed to be conserved in A, D and AD genomes. In the species of B, E, F and G genomes, the rDNA numbers, sizes, and synteny relationships were first reported in this paper. The rDNA pattern agrees with previously reported phylogenetic history with some disagreements. Combined with the whole-genome sequencing data from G . raimondii (D₅) and the conserved cotton karyotype, it is suggested that the expansion, decrease and transposition of rDNA other than chromosome rearrangements might occur during the Gossypium evolution.     

Dual-color FISH karyotype analyses using rDNAs in three Cucurbitaceae species

Dual-color fluorescence in situ hybridization (FISH) analysis of three Cucurbitaceae species from different genera was conducted using 5S and 45S rDNA probes. In Benincasa hispida (Thunb.) Cogn. (2n=24), the 45S rDNA probe hybridized on two chromosomes, one in the short arm of a medium-sized metacentric chromosome and another at the satellite of a chromosome. The 5S rDNA hybridized at a site proximal to the centromere of the same short arm of the 45S rRNA gene locus that occupied almost the entire short arm. For Citrullus lanatus (Thunb.) Matsum & Nakai (2n=22), the 45S rDNA probe hybridized at sites in the short arms of two chromosomes and the 5S rDNA probe was co-localized with the 45S rRNA locus at the region proximal to the centromere in one chromosome. The 45S rRNA loci occupied almost all of the short arms in both chromosomes. In Cucurbita moschata Duch. (2n=40), the 45S rDNA probe hybridized in five chromosomes in which the 45S rRNA genes occupied almost two-thirds of the chromosomes in two large chromosomes and the entire short arm of a medium-sized chromosome. Two other loci were present in two medium-sized chromosomes, one in the proximal region in the short arm of a chromosome and another at the tip of the long arm of a chromosome. Chromosomes of B. hispida were relatively larger than those of the other two species. The karyotype of B. hispida is composed of two metacentrics and 10 submetacentrics, while that of C. lanatus is composed of seven metacentrics and four submetacentrics and that of C. moschata is composed of 18 metacentrics and two submetacentrics. Comparative chromosome evolution among the three Cucurbitaceae species was attempted using the karyotypes and the chromosomal distribution patterns of the 5S and 45S rDNAs. The results presented herein will be useful in elucidating the phylogenetic relationships among Cucurbitaceae species, and will provide basic data for their breeding programs.     

Comparative cytogenetic analysis of the genomes of the model grass Brachypodium distachyon and its close relatives

Background and Aims

Brachypodium is a small genus of temperate grasses that comprises 12–15 species. Brachypodium distachyon is now well established as a model species for temperate cereals and forage grasses. In contrast to B. distachyon, other members of the genus have been poorly investigated at the chromosome level or not at all.

Methods

Twenty accessions comprising six species and two subspecies of Brachypodium were analysed cytogenetically. Measurements of nuclear genome size were made by flow cytometry. Chromosomal localization of 18–5·8–25S rDNA and 5S rDNA loci was performed by dual-colour fluorescence in situ hybridization (FISH) on enzymatically digested root-tip meristematic cells. For comparative phylogenetic analyses genomic in situ hybridization (GISH) applied to somatic chromosome preparations was used.

Key Results

All Brachypodium species examined have rather small genomes and chromosomes. Their chromosome numbers and genome sizes vary from 2n = 10 and 0·631 pg/2C in B. distachyon to 2n = 38 and 2·57 pg/2C in B. retusum, respectively. Genotypes with 18 and 28 chromosomes were found among B. pinnatum accessions. GISH analysis revealed that B. pinnatum with 28 chromosomes is most likely an interspecific hybrid between B. distachyon (2n = 10) and B. pinnatum (2n = 18). Two other species, B. phoenicoides and B. retusum, are also allopolyploids and B. distachyon or a close relative seems to be one of their putative ancestral species. In chromosomes of all species examined the 45S rDNA loci are distally distributed whereas loci for 5S rDNA are pericentromeric.

Conclusions

The increasing significance of B. distachyon as a model grass emphasizes the need to understand the evolutionary relationships in the genus Brachypodium and to ensure consistency in the biological nomenclature of its species. Modern molecular cytogenetic techniques such as FISH and GISH are suitable for comparative phylogenetic analyses and may provide informative chromosome- and/or genome-specific landmarks.