Age, reproduction and fecundity of the spined loach Cobitis taenia L. (Pisces, Cobitidae) from Lake Klawój (Poland)

This is the first study concerning the features of the reproduction process of the karyologically identified spined loach C. taenia (2n=48). The histology of 71 ovaries, and gonadosomatic index (GSI) of karyologically identified spined loach Cobitis taenia L. from Lake Klawój (Northern Poland) were examined. The absolute and relative fecundity of 25 females was estimated by gravimetric method. The age of fish was determined according to the annual increments of otholits. The spawning of C. taenia from Lake Klawój took place from May to July, at a water temperature exceeding 18.5 degrees C. The GSI values at the beginning of the reproduction period ranged from 7 to 19%. The average absolute fecundity of females was 2078 eggs, with the number ranging from 869 to 3371 eggs. High individual variability in the gonad histology and the GSI values during the reproductive period was observed. Such variability could be the result of beginning the reproduction process in the fish at various times and, probably, due to the various numbers of batches laid and various numbers of eggs per batch.     

Cobitis pontica sp. nova—a new spined loach species (Cobitidae) from the Bulgarian waters

Spined loaches from the Veleka River (Black Sea coast of Bulgaria), earlier confirmed to represent a separate evolution lineage within a 50-chromosome morph occurring in the Black Sea basin, are described as a new species. This species has the karyotype (2n = 50, NF = 90) similar to one from earlier described C. taurica occurring in the Crimean Peninsula, but differs from it by larger and less numerous spots in the fourth Gambetta’s zone of pigmentation and more anterior position of suborbital spine.     

Distribution of 5S and 18S–28S rDNA loci in a tetraploid cotton (Gossypium hirsutum L.) and its putative diploid ancestors

The most widely cultivated species of cotton,Gossypium hirsutum, is a disomic tetraploid (2n=4x=52). It has been proposed previously that extant A- and D-genome species are most closely related to the diploid progenitors of the tetraploid. We used fluorescent in situ hybridization (FISH) to determine the distribution of 5S and 18S-28S rDNA loci in the A-genome speciesG. herbaceum andG. arboreum, the D-genome speciesG. raimondii andG. thurberi, and the AD tetraploidG. hirsutum. High signal-to-noise, single-label FISH was used to enumerate rDNA loci, and simultaneous, dual-label FISH was used to determine the syntenic relationships of 5S rDNA loci relative to 18S–28S rDNA loci. These techniques provided greater sensitivity than our previous methods and permitted detection of six newG. hirsutum 18S–28S rDNA loci, bringing the total number of observed loci to 11. Differences in the intensity of the hybrizization signal at these loci allowed us to designate them as major, intermediate, or minor 18–28S loci. Using genomic painting with labeled A-genome DNA, five 18S–28S loci were localized to theG. hirsutum A-subgenome and six to the D-subgenome. Four of the 11 18S–28S rDNA loci inG. hirsutum could not be accounted for in its presumed diploid progenitors, as both A-genome species has three loci and both D-genome species had four.G. hirsutum has two 5S rDNA loci, both of which are syntenic to major 18S–28S rDNA loci. All four of the diploid genomes wer examined contained a single 5S locus. InG. herbaceum (A₁) andG. thurberi (D₁), the 5S locus is syntenic to a major 18S–28S locus, but inG. arboreum (A₂) andG. raimondii (D₅), the proposed D-genome progenitor ofG. hirsutum, the 5S loci are syntenic tominor and intermediate 18S–28S loci, respecitively. The multiplicity, variation in size and site number, and lack of additivity between the tetraploid species and its putative diploid ancestors indicate that the behavior of rDNA loci in cotton is nondogmatic, and considerably more complex and dynamic than previously envisioned. The relative variability of 18S–28S rDNA loci versus 5S rDNA loci suggests that the behavior of tandem repearts can differ widely. Edited by: R. Appels     

Gene mapping of 5S rDNA sites in eight fish species from the Paraíba do Sul river basin, Brazil

The use of improved cytogenetic techniques such as fluorescent in situ hybridization (FISH) has offered important methodologies for cytotaxonomic and evolutionary studies. In particular, the mapping of 5S rDNA sites has proved to be an excellent marker in the study of different organisms and, more recently, in fish. In the present work, the FISH technique was used to map the 5S rDNA sites in the chromosomes of eight neotropical fish species from the Paraíba do Sul river basin, four of these belonging to the order Characiformes, family Characidae, genus Astyanax (A. scabripinnis, A. parahybae, A. giton and A. intermedius) and four to the order Siluriformes, family Loricariidae (Neoplecostomus microps, Harttia loricariformis, Hypostomus affinis and Upsilodus sp.). Karyotype evolution aspects of the analyzed groups are discussed.     

Genetic structure of species complex of the spined loach Cobitis auc. (Cypriniformes: Cobitidae) in Severskiĭ Donets river basin

Biochemical, genetic, cytometric and morphological analyses of spined loaches of the middle stream of Severskiy Donetz river revealed 3 bisexual species: Cobitis taenia s.l. (68% of the sample); C. melanoleuca (11%); Sabanejewia aurata (9%) and 2 hybrid forms: triploid C. taenia(2)-sp. (9%) and diploid C. taenia-melanoleuca (3%). Distinctive features of genetic structure of polyploid hybrids C. taenia(2)-sp. as well as taxonomic identity of diploid C. taenia s.l. of Severskiy Donetz river were discussed in regard to the Dnieper population ones.     

The Genetic Structure of the Diploid–Polyploid Complex of the Spined Loach Cobitis taenia (Cypriniformes: Cobitidae) from the Middle Dnieper Basin

Biochemical genetic typing and cytometry showed that polyploid females account for 87% of the spined loachCobitis taeniapopulation from the middle Dnieper basin. The polyploidy series included triploids, tetraploids, and, possibly, a few pentaploids. A characteristic feature of the genetic structure of polyploids was that their genetic variation was due to the clonal variation in the haploid portion of the genome originating from Cobitis sp. and to polymorphism of the diploid portion originating fromC. taenia. The results are discussed with regard to comparative evolution of alloploid complexes in fish and terrestrial vertebrates.     

Karyotype and chromosomal characteristics of Ag–NOR sites and 5S rDNA in European smelt, Osmerus eperlanus

Karyotype and cytogenetic characteristics of European smelt Osmerus eperlanus were investigated using different staining techniques (sequential Ag-, CMA3 and DAPI banding) and PRINS to detect 5S rDNA and telomeric sites. The diploid chromosome number was invariably 2n = 56 and karyotype composed of 5 pairs of metacentrics, 9 pairs of subtelocentrics and 14 pairs of subtelo- to acrocentrics. The DAPI-positive heterochromatic regions were found in centromeric positions on bi-armed chromosomes and few acrocentrics. Additionally, some interstitial DAPI-positive bands were identified on three pairs of submetacentric chromosomes. The nucleolar organizer regions (NORs) were detected in the short (p) arms of the largest metacentric pair of chromosomes No. 1. Sequential banding (Giemsa-, AgNO₃ and CMA₃ stainings) revealed NOR sites corresponding to achromatic regions but not associated with CMA₃-positive blocks of heterochromatin located on either side of NORs. Individuals from the analyzed population had this conspicuous pair of chromosomes always in heterozygous combination. A complex inversion system was hypothesized to be involved in the origin of the observed variation but analysis with telomeric PRINS and PNA-FISH did not reveal any Interstitial Telomeric Sites (ITS). Hybridization signals were confined exclusively to terminal chromosomal regions. The 5S ribosomal sites as revealed by PRINS were found to be invariably located in the short (p) arms of four pairs of subtelocentric chromosomes. Cytotaxonomic comparisons of the present results with the voluminous available cytogenetic data-set from salmoniform and esociformes fishes appear to support the recent view, based on robust molecular-based phylogeny, that salmoniform and osmeriform fishes are not as closely related as previously assumed.     

<Emphasis Type="Italic">Cobitis faridpaki</Emphasis> sp. nova—a New spined loach species (Cobitidae) from the southern Caspian Sea basin (Iran)

Spined loaches from the Siahrud River of the southern coast of the Caspian Sea in the province of Mazandaran, north of Iran, are described as a new species Cobitis faridpaki. This species belongs to Cobitis species group with a single lamina Canestrini at the base of the second pectoral-fin ray in males and subdorsal scales with reduced eccentric focal zone. These characters differentiate C. faridpaki from both southern Iranian species C. linea and C. melanoleuca distributed in the northern Caspian Sea basin. The stout body, small and numerous (25–30) lateral spots and the presence of 14–16 branched caudal-fin rays separate C. faridpaki from Caucasian species C. satunini.     

Species-genomic relationships among the tribasic diploid and polyploid Carthamus taxa based on physical mapping of active and inactive 18S–5.8S–26S and 5S ribosomal RNA gene families,and the two tandemly repeated DNA sequences

In the genus Carthamus (2n = 20, 22, 24, 44, 64; x = 10, 11, 12), most of the homologues within and between the chromosome complements are difficult to be identified. In the present work, we used fluorescent in situ hybridisation (FISH) to determine the chromosome distribution of the two rRNA gene families, and the two isolated repeated DNA sequences in the 14 Carthamus taxa. The distinctive variability in the distribution, number and signal intensity of hybridisation sites for 18S–26S and 5S rDNA loci could generally distinguish the 14 Carthamus taxa. Active 18S–26S rDNA sites were generally associated with NOR loci on the nucleolar chromosomes. The two A genome taxa, C. glaucus ssp. anatolicus and C. boissieri with 2n = 20, and the two botanical varieties of B genome C. tinctorius (2n = 24) had diagnostic FISH patterns. The present results support the origin of C. tinctorius from C. palaestinus. FISH patterns of C. arborescens vis-à-vis the other taxa indicate a clear division of Carthamus taxa into two distinct lineages. Comparative distribution and intensity pattern of 18S–26S rDNA sites could distinguish each of the tetraploid and hexaploid taxa. The present results indicate that C. boissieri (2n = 20) is one of the genome donors for C. lanatus and C. lanatus ssp. lanatus (2n = 44), and C. lanatus is one of the progenitors for the hexaploid (2n = 64) taxa. The association of pCtKpnI-2 repeated sequence with rRNA gene cluster (orphon) in 2–10 nucleolar and non-nucleolar chromosomes and the consistent occurrence of pCtKpnI-1 repeated sequence at the subtelomeric region in all the taxa analysed indicate some functional role of these sequences.     

Linkage disequilibrium mapping in the Newfoundland population: a re-evaluation of the refinement of the Bardet–Biedl syndrome 1 critical interval

Genetically isolated populations, such as Newfoundland, have contributed greatly to the identification of disease-causing genes. A linkage disequilibrium (LD) study involving six Newfoundland families predicted a critical interval for Bardet-Biedl syndrome 1 (BBS1) (Young et al. in Am J Hum Genet 65:1680–1687, 1999), but the subsequent identification of BBS1 revealed that it lies outside this region. This suggested that either there is another gene responsible for BBS in these families or the Newfoundland population may not be ideal for LD studies. We screened these six Newfoundland families for mutations in BBS1 and found that affected individuals in five of them were homozygous for the same M390R mutation. There was no evidence for any BBS1 mutation in the affected individual in the sixth family. Therefore, one of the criteria for LD mapping was not met; namely, there should be a single disease-causing allele in the population. Haplotype analysis of unaffected individuals from south-west Newfoundland and English BBS1 patients homozygous for M390R, revealed that a second criterion for LD mapping was violated. The M390R mutation occurred in a common haplotype and both of these chromosomes, the ancestral wild-type and disease-causing haplotypes, were introduced to Newfoundland and spread by a founder effect. Moreover, it was found that disease-associated alleles occurred at relatively high frequencies in normal haplotypes and this probably accounted for the incorrect prediction in the previous LD study. Knowing the amount of genetic variation and its distribution in the Newfoundland population would be useful to maximize its potential for mapping hereditary disorders.     

Identification, characterization and mapping of EST-derived SSRs from the cacao–Ceratocystis cacaofunesta interaction

Ceratocystis cacaofunesta is an ascomycete responsible for the lethal wilt disease of cacao (Theobroma cacao L.). Marker-assisted selection combined with conventional breeding is one powerful approach to improve cacao resistance to Ceratocystis wilt. In this study we screened a set of ESTs obtained from cacao elicited with C. cacaofunesta to identify EST-SSRs and test their efficacy for mapping. Among the 3,432 ESTs analysed, 384 contained SSRs and 428 EST-SSRs were identified, mainly dinucleotides (78.5 %), with four repeats (75.23 %), and preferentially AG/CT motif (25.47 %). Gene ontology function was assigned to the ESTs containing SSRs: 4.04 % belonged to “defense response” category, with 20.69 % of them to the sub-category “defense response to fungi”. In relation to the ORF, the same quantity of EST-SSRs was observed in the 5′ UTR, ORF and the 3′ UTR (about 30 %). From the 428 EST-SSRs identified, 12 were polymorphic, revealing a total of 41 alleles. The number of alleles per locus ranged from 2 to 6, with an average of 3.41. Four EST-SSRs were mapped on the F₂ Sca 6 × ICS 1 population segregating for Ceratocystis wilt, and were distributed on the 2, 3, 4 and 8 linkage groups. These markers will have potential applications in linkage mapping and will be valuable for the research community to improve the cacao breeding program.     

Reevaluation of the Evolutionary Position of Opalinids Based on 18S rDNA,and α- and β-Tubulin Gene Phylogenies

Opalinids are enigmatic endosymbiotic protists principally found in the large intestine of anuran amphibians. They are multinucleates and uniformly covered with numerous flagella (or cilia). Their appearance is somewhat similar to that of ciliates, leading to opalinid’s initial classification as ciliates, or later as protociliates. However, on the basis of their monomorphic nuclei, absence of a ciliate-like life cycle characterized by conjugation, and an interkinetal fission mode, opalinids were subsequently transferred in the zooflagellates. As several common ultrastructural characteristics shared with proteromonads were elucidated, in particular of the flagellar base, such as their double-stranded flagellar helix, an alliance with proteromonads was widely accepted. Thus, opalinids are currently favored to be placed in the class Opalinea, within the heterokont kingdom Chromista. However, the question of their classification has not been fully resolved, because of a lack of molecular information. Here, we report their phylogenetic position inferred from 18S rDNA, and α- and β-tubulin gene sequences. The 18S rDNA tree gives the opalinids an ancestral position in heterokonts, together with proteromonads, as suggested by the morphological studies. In great contrast, α- and β-tubulin gene analyses suggest an affiliation of opalinids to alveolates, not to heterokonts. However, the AU test implies that opalinids are not closely related with any of other three phyla in the alveolates, suggesting an occupation of an ancestral position within the alveolates. Based on the present molecular information, in particular rDNA phylogeny, and the ultrastructural character of the double helix common to heterokonts, we conclude that opalinids would have a common origin with heterokonts, although analyses based on two tubulin genes do not as yet completely deny a possible placement outside heterokonts. The ambiguity of the evolutionary position shown by the discrepancy between rDNA and tubulin genes phylogenies might reflect an early emergence of opalinids in ancestral chromalveolates, and an extreme specialization during a lengthy history of parasitism, as suggested by a long branch in the rDNA tree.Reviewing Editor: Dr. Patrick Keeling     

O father where art thou? Paternity analyses in a natural population of the haploid–diploid seaweed Chondrus crispus

The link between life history traits and mating systems in diploid organisms has been extensively addressed in the literature, whereas the degree of selfing and/or inbreeding in natural populations of haploid–diploid organisms, in which haploid gametophytes alternate with diploid sporophytes, has been rarely measured. Dioecy has often been used as a proxy for the mating system in these organisms. Yet, dioecy does not prevent the fusion of gametes from male and female gametophytes originating from the same sporophyte. This is likely a common occurrence when spores from the same parent are dispersed in clumps and recruit together. This pattern of clumped spore dispersal has been hypothesized to explain significant heterozygote deficiency in the dioecious haploid–diploid seaweed Chondrus crispus. Fronds and cystocarps (structures in which zygotes are mitotically amplified) were sampled in two 25 m² plots located within a high and a low intertidal zone and genotyped at 5 polymorphic microsatellite loci in order to explore the mating system directly using paternity analyses. Multiple males sired cystocarps on each female, but only one of the 423 paternal genotypes corresponded to a field-sampled gametophyte. Nevertheless, larger kinship coefficients were detected between males siring cystocarps on the same female in comparison with males in the entire population, confirming restricted spermatial and clumped spore dispersal. Such dispersal mechanisms may be a mode of reproductive assurance due to nonmotile gametes associated with putatively reduced effects of inbreeding depression because of the free-living haploid stage in C. crispus.     

Karyotype and Chromosomal Location of 18S–28S and 5S Ribosomal DNA in the Scallops Pecten maximus and Mimachlamys varia (Bivalvia: Pectinidae)

This work describes the karyotype and chromosomal location of the ribosomal DNA (rDNA) of Pecten maximus and Mimachlamys varia, two commercial scallop species from Europe. According to the chromosome centromeric index values found, the karyotype of P. maximus is composed of 1 metacentric, 2 metacentric–submetacentric, 1 telocentric–subtelocentric and 15 telocentric pairs, and that of M. varia of 4 metacentric, 2 subtelocentric–submetacentric, 9 subtelocentric, 3 subtelocentric–telocentric and 1 telocentric–subtelocentric pairs. In P. maximus, 18S-28S rDNA was located by FISH on a metacentric–submetacentric pair, and in M. varia on a subtelocentric–submetacentric pair using both silver staining and FISH. PCR amplification of the 5S rDNA unit yielded a single product of about 460 bp (P. maximus) and 450 bp (M. varia), that used as probe revealed a 5S rDNA site on a telocentric pair in P. maximus and a subtelocentric pair in M. varia. Two-color FISH or sequential silver staining of 5S rDNA-FISH-metaphases corroborated that the two gene families are located on different chromosomes in both species. A comparative analysis of the data allowed the inference of karyotypic relationships within scallops.     

Gene–Gene and Gene-Sex Epistatic Interactions of MiR146a,IRF5, IKZF1, ETS1 and IL21 in Systemic Lupus Erythematosus

Several confirmed genetic susceptibility loci involved in the interferon signaling and Th17/B cell response for SLE in Chinese Han populations have been described. Available data also indicate that sex-specific genetic differences contribute to SLE susceptibility. The aim of this study was to test for gene–gene/gene-sex epistasis (interactions) in these known lupus susceptibility loci. Six single-nucleotide polymorphisms (SNPs) in MiR146a, IRF5, IKZF1, ETS1 and IL21 were genotyped by Sequenom MassArray system. A total of 1,825 subjects (858 SLE patients and 967 controls) were included in the final analysis. Epistasis was tested by additive model, multiplicative model and multifactor dimensionality reduction (MDR) method. Additive interaction analysis revealed interactions between IRF5 and IKZF1 (OR 2.26, 95% CI 1.48–3.44 [P = 1.21×10⁴]). A similar tendency was also observed between IL21 and ETS1 by parametric methods. In addition, multiple high dimensional gene-gene or gene-sex interactions (three-and four-way) were identified by MDR analysis. Our study identified novel gene–gene/gene-sex interactions in lupus. Furthermore, these findings highlight sex, interferon pathway, and Th17/B cells as important contributors to the pathogenesis of SLE.     

Chromosomal localization of 5S and 18S–5.8S–25S ribosomal DNA sites in five Asian pines using fluorescence in situ hybridization

Fluorescence in situ hybridization (FISH) was employed on mitotic metaphase chromosome preparations of five Asian Pinus species: Pinus tabuliformis, Pinus yunnanensis, Pinus densata, Pinus massoniana and Pinus merkusii, using simultaneously DNA probes of the 18S rRNA gene and the 5S rRNA gene including the non-transcribed spacer sequences. The number and location of 18S rDNA sites varied markedly (5-10 pairs of strong signals) among the five pines. A maximum of 20 major 18S rDNA sites was observed in the diploid genome (2n = 24) of P. massoniana. The 5S rDNA FISH pattern was less variable, with one major site and one minor site commonly observed in each species. The differentiation of rDNA sites on chromosomes among the five pines correlates well with their phylogenic positions in Pinus as reconstructed from other molecular data. P. densata, a species of hybrid origin, resembles its parents ( P. tabuliformis and P. yunnanensis), including some components characteristic of each parent in its pattern. However, the species is unique, showing new features resulting possibly from recombination and genome reorganization.     

Temporal trends in net and crude probability of death from cancer and other causes in the Australian population, 1984–2013

BackgroundWhile net probabilities of death in the relative survival framework ignore competing causes of death, crude probabilities allow estimation of the real risk of cancer deaths. This study quantifies temporal trends in net and crude probabilities of death.MethodsAustralian population-based cohort of 2,015,903 people aged 15-89 years, diagnosed with a single primary invasive cancer from 1984 to 2013 with mortality follow-up to 31 December 2014. Survival was analyzed with the cohort method. Flexible parametric relative survival models were used to estimate both probability measures by diagnosis year for all cancers and selected leading sites.ResultsFor each site, excess mortality rates reduced over time, especially for prostate cancer. While both the 10-year net and crude probability of cancer deaths decreased over time, specific patterns varied. For example, the crude probability of lung cancer deaths for males aged 50 years decreased from 0.90 (1984) to 0.79 (2013); whereas the corresponding probabilities for kidney cancer were 0.64 and 0.18 respectively. Patterns for crude probabilities of competing deaths were relatively constant. Although for younger patients, both net and crude measures were similar, crude probability of competing deaths increased with age, hence for older ages net and crude measures were different except for lung and pancreas cancers.ConclusionsThe observed reductions in probabilities of death over three decades for Australian cancer patients are encouraging. However, this study also highlights the ongoing mortality burden following a cancer diagnosis, and the need for continuing efforts to improve cancer prevention, diagnosis and treatment.