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A 7-year cytogenetic study on the leaf litter frog Craugastor fitzingeri from Costa Rica and Panama revealed the existence of highly mobile nucleolus organizing regions (NORs) in their genomes. Silver (Ag)-staining of the active NORs demonstrated an exceptional interindividual pattern of NOR distribution at the telomeres of the chromosomes. All individuals examined showed a different and specific NOR location in their karyotypes. Furthermore, intraindividual variation in the NOR sites was found. This observation suggested the existence of mobile NORs in C. fitzingeri. Confirmation of this phenomenon was possible by systematic FISH analysis using an 18S + 28S rDNA probe. The extremely variable number and position of the NORs in C. fitzingeri is best explained by highly mobile NORs that move freely between the telomeres of the chromosomes. These transpositions must occur preferentially in premeiotic, meiotic, or postmeiotic stages, but also at a lower incidence in the somatic tissues of the animals. It is hypothesized that transposable (mobile) elements are closely linked to the NORs or are inserted into the major 18S + 28S rDNA spacers of C. fitzingeri. When such transposable elements spread by transpositions, they can carry with them complete or partial NORs. The present study provides detailed information on various differential chromosome banding techniques, in situ hybridization experiments, chromosomal hypermethylation patterns, determination of the genome size, and analyses of restriction fragment length polymorphisms of the DNA.

Human amniotic fluid stem cells (hAFSCs) harbor high proliferative capacity and high differentiation potential and do not raise the ethical concerns associated with human embryonic stem cells. The formation of three-dimensional aggregates known as embryoid bodies (EBs) is the principal step in the differentiation of pluripotent embryonic stem cells. Using c-Kit-positive hAFSC lines, we show here that these stem cells harbor the potential to form EBs. As part of the two kinase complexes, mTORC1 and mTORC2, mammalian target of rapamycin (mTOR) is the key component of an important signaling pathway, which is involved in the regulation of cell proliferation, growth, tumor development and differentiation. Blocking intracellular mTOR activity through the inhibitor rapamycin or through specific small interfering RNA approaches revealed hAFSC EB formation to depend on mTORC1 and mTORC2. These findings demonstrate hAFSCs to be a new and powerful biological system to recapitulate the three-dimensional and tissue level contexts of in vivo development and identify the mTOR pathway to be essential for this process.

The mitotic chromosomes of the pocket mouse Heteromys anomalus (Rodentia, Heteromyidae) from Venezuela were analyzed by means of conventional staining and various banding techniques. The karyotype of this species is distinguished by a Y chromosome with exceptional banding properties. The very large heterochromatic Y long arm shows a remarkable bright quinacrine fluorescence, but only moderate or dull fluorescence when labeled with other AT base pair-specific fluorochromes. The implications of these findings are discussed.

We conducted a chromosome banding analysis (heterochromatin, nucleolus organizer regions, DAPI fluorescence, distamycin A/mithramycin fluorescence) of six phylogenetically basal dendrobatid frog species (Colostethus chalcopis, C. leopardalis, Mannophryne herminae, M. neblina, M. olmonae, M. trinitatis). With the exception of C. chalcopis (2n = 22), all examined species had a chromosome complement of 2n = 24 chromosomes. The C- and Q-band analyses showed that constitutive heterochromatin is present at the centromeres of all species, with Q−-regions occurring at the positions of the nucleolus organizer regions (NORs) of C. leopardalis, M. olmonae, and M. trinitatis. The C-band polymorphisms were detected in M. herminae and M. neblina on chromosome No. 6 and in M. herminae on chromosome No. 7. Silver-staining and distamycin A/mithramycin fluorescence resolved a single pair of NORs in each species. The DAPI fluorescence revealed pericentromeric bands on chromosome No. 1 and 5 in M. trinitatis and on chromosom...

Extensive cytogenetic analyses on a population of the leptodactylid frog Eleutherodactylus riveroi in northern Venezuela revealed the existence of multiple XXAA♂/XYAA♀/XAAY♀ sex chromosomes. The XAAY karyotype originated by a centric (Robertsonian) fusion between the original, free Y chromosome and an autosome. 46.2% of the male individuals in this population are carriers of this Y-autosome fusion. In male meiosis the XAAY sex chromosomes pair in the expected trivalent configuration. In the same population 53.8% of the male animals still possess the original, free XY sex chromosomes. E. riveroi is only the second vertebrate species discovered in which a derived Y-autosome fusion coexists with the ancestral free XY sex chromosomes. The free XY sex chromosomes, as well as the multiple XAY sex chromosomes are still in a very primitive (homomorphic) stage of differentiation. With no banding technique applied it is possible to distinguish the Y from the X. Various banding techniques and in situ hybridizations have been carried out to characterize the karyotypes. DNA flow cytometric measurements show that the genome size of E. riveroi resembles that of other Eleutherodactylus species. The cytogenetic data obtained in E. riveroi are compared with those of the sole other vertebrate known to possess the extremely rare, multiple XXAA♂/XYAA♀/XAAY♀ sex chromosomes. Surprisingly enough, this vertebrate again is a frog belonging to the genus Eleutherodactylus [E. (maussi) biporcatus] which lives exactly in the same habitat in northern Venezuela as does E. riveroi.

The physical ends of mammalian and other vertebrate chromosomes consist of tandemly repeated (TTAGGG)(n) hexamers, nucleating a specialized telomeric structure. However, (TTAGGG)(n) sequences can also occur at non-telomeric sites, providing important insights into karyotypic evolution. By fluorescence in situ hybridization (FISH) we studied the chromosomal distribution of (TTAGGG)(n) sequences in 16 bird species, representing seven different orders. Many species, in particular the ratites, display (TTAGGG)(n) hybridization signals in interstitial and centromeric regions of their macrochromosomes in addition to the typical telomeric signals. In some but not all species these non-telomeric sites coincide with C-band-positive heterochromatin. The retention and/or amplification of telomeric (TTAGGG)(n) repeats at interstitial and centromeric sites may indicate the fusion of ancestral chromosomes. Compared with the macrochromosomes, the microchromosomes of most species are enriched with (TTAGGG)(n) sequences, displaying heterogeneous hybridization patterns. We propose that this high density of (TTAGGG)(n) repeats contributes to the exceptionally high meiotic recombination rate of avian microchromosomes.

A 15-year cytogenetic survey on one population of the leaf litter frog Eleutherodactylus maussi in northern Venezuela confirmed the existence of multiple XXAA♂/XAAY♀ sex chromosomes which originated by a centric (Robertsonian) fusion between the original Y chromosome and an autosome. 95% of the male individuals in this population are carriers of this Y-autosome fusion. In male meiosis the XAAY sex chromosomes pair in the expected trivalent configuration. In the same population, 5% of the male animals still possess the original, free XY sex chromosomes. In a second population of E. maussi analyzed, all male specimens are characterized by these ancestral XY chromosomes which form normal bivalents in meiosis. E. maussi apparently represents the first vertebrate species discovered in which a derived Y-autosome fusion still coexists with the ancestral free XY sex chromosomes. The free XY sex chromosomes, as well as the multiple XAY sex chromosomes are still in a very primitive (homomorphic) stage of differentiation. With no banding technique applied it is possible to distinguish the Y from the X. DNA flow cytometric measurements show that the genome of E. maussi is among the largest in the anuran family Leptodactylidae. The present study also supplies further data on differential chromosome banding and fluorescence in situ hybridization experiments in this amphibian species.

Highly differentiated, heteromorphic ZZ♀/ZW♂ sex chromosomes were found in the karyotypes of the neotropical leptodactylid frogs Eleutherodactylus euphronides and E. shrevei. The W chromosomes are the largest heterochromatic, female-specific chromosomes so far discovered in the class Amphibia. The analyses of the banding patterns with AT- and GC base-pair specific fluorochromes show that the constitutive heterochromatin in the giant W chromosomes consists of various categories of repetitive DNA sequences. The W chromosomes of both species are similar in size, morphology and banding patterns, whereas their Z chromosomes exhibit conspicuous differences. In the cell nuclei of female animals, the W chromosomes form very prominent chromatin bodies (W chromatin). DNA flow cytometric measurements demonstrate clear differences in the DNA content of male and female erythrocytes caused by the giant W chromosome, and also shows that these Eleutherodactylus genomes are among the smallest of all amphibian genomes. The importance of the heteromorphic ZW sex chromosomes for the study of Z-linked genes, the similarities and differences of the two karyotypes, and the significance of the exceptionally small genomes are discussed.

Mitotic and meiotic chromosomes of 5 species of the reptile genus Gonatodes are described by means of conventional staining, banding analyses and in situ hybridization using a synthetic telomeric DNA probe. The amount, location and fluorochrome affinities of constitutive heterochromatin, the number and positions of nucleolus organizer regions, and the patterns of telomeric DNA sequences were determined for most of the species. The karyotypes of G. falconensis and G. taniae from northern Venezuela are distinguished by their extraordinarily reduced diploid chromosome number of 2n = 16, which is the lowest value found so far in reptiles. In contrast to most other reptiles, both species have exclusively large biarmed (meta- and submetacentric) chromosomes. Comparison of the karyotypes of G. falconensis and G. taniae with those of other Gonatodes species indicates that the exceptional 2n = 16 karyotype originated by a series of 8 centric fusions. The karyotypes of G. falconensis and G. taniae are further characterized by the presence of considerable amounts of (TTAGGG)n telomeric sequences in the centromeric regions of all chromosomes. These are probably not only relics of the centric fusion events, but a component of the highly repetitive DNA in the constitutive heterochromatin of the chromosomes. The genome sizes of 4 Gonatodes species were determined using flow cytometry. For comparative purposes, all previously published cytogenetic data on Gonatodes and other sphaerodactylids are included and discussed.

The cell lines SW480 and SW620, derived from different stages of colon carcinoma in the same patient, have been used for a number of biochemical, immunological, and genetic studies on colon cancer. A comparative analysis of their karyotypes may identify chromosomal aberrations that might represent markers for metastatic spread. In the present study spectral karyotyping (SKY) was applied to these two colon cancer cell lines. Compared to previously reported G-banded karyotypes, 9 (SW480) and 7 (SW620) markers were identical, 3 (SW480) and 3 (SW620) markers could be redefined, 5 (SW480) and 8 (SW620) markers were newly identified, and 4 (SW480) and 5 (SW620) of the previous described markers could not be confirmed. The redefined aberrations include very complex rearrangements, such as a der(16) t(3;16;1;16;8;16; 1;16;10) and a der(18)t(18;15;17)(q12; p11p13;??) in SW620 and a der(19)t(19;8;19;5) in SW480, that have not been identified by conventional banding techniques. The resulting chromosome gains (5q11→5q15, 7pter→q22, 11, 13q14→qter, 20pter→p12, X) and losses (8pter→p2, 18q12→qter, Y) found in both SW480 and SW620 were in good agreement with those frequently described in colorectal tumors as primary changes in the stem cell. Abnormalities found exclusively in SW620 cells only (gains of 5pter→5q11, 12q12→q23, 15p13→p11, and 16q21→q24 and losses of 2pter→2p24, 4q28→qter, and 6q25→qter) can be viewed as changes that occurred in a putative metastatic founder cell.

Homozygotes for the rare folate-sensitive autosomal fragile sites have never been recorded. Two non-folate-sensitive rare fragile sites (FRA10B and FRA17A) have been previously recorded in normal individuals. We document two unrelated normal individuals who are homozygotes for the rare fragile site FRA16B and record the patterns of induction of this fragile site with berenil. The existence of normal homozygotes for FRA16B suggests that this fragile site is not within a gene essential for normal development.

An unusual NOR-bearing chromosome 7 was detected in a phenotypically normal, healthy 29-year-old male proband. Differential banding techniques as well as in situ hybridization employing various DNA-probes were performed in order to characterize the chromosome in detail. The nucleolus organizer region was found to be located between bands 7q21.3 and 7q22.1. No further rearrangements were detected in this chromosome. Analysis of spontaneously occurring micronuclei revealed 9% of them to contain a 7q fragment distal to (or including) the inserted NOR, suggesting that the inserted secondary constriction represents a potential hot spot for chromosomal breakage and rearrangement. Segregation analysis of the variant chromosome 7 in 51 members of the probands’ family showed transmission in a Mendelian fashion. 27 individuals were found to be heterozygous for the inserted chromosome. A three-year-old child in the consanguineous marriage of two heterozygous carriers exhibited the NOR-insertion in both of his chromosome 7 homologues. To our knowledge, this is the first report on a homozygous carrier of a non-acrocentric NOR-bearing chromosome.

The mitotic chromosomes of 11 species from the anuran families Centrolenidae and Allophrynidae were analyzed by means of conventional staining, banding techniques, and in situ hybridization. The amount, location, and fluorochrome affinities of constitutive heterochromatin, the number and positions of nucleolus organizer regions, and the patterns of telomeric DNA sequences were determined for most of the species. The karyotypes were found to be highly conserved with a low diploid chromosome number of 2n = 20 and morphologically similar chromosomes. The sister group relationship between the Centrolenidae and Allophrynidae (unranked taxon Allocentroleniae) is clearly corroborated by the cytogenetic data. The existence of heteromorphic XY♂/XX♀ sex chromosomes in an initial stage of morphological differentiation was confirmed in Vitreorana antisthenesi. The genome sizes of 4 centrolenid species were determined using flow cytometry. For completeness and for comparative purposes, all previously published cytogenetic data on centrolenids are included.

A cytogenetic investigation of Hyla ebraccata Cope, H. microcephala Cope, and H. phlebodes Stejneger revealed that the karyotypes of these 30-chromosome Hyla are very conservative. With the exception of some structural rearrangements, only few differences in chromosomal morphology could be discerned. Based on our results, we hypothesize that the telomeric position of nucleolus organizer regions (NOR) on chromosome no. 10 may represent a derived condition in 30-chromosome Hyla. This cytotype was found only in the Caribbean population of H. ebraccata, Such within-species disparity has not been observed previously among amphibians. This phenomenon can most readily be explained by a translocation or insertion that rapidly drifted to high frequency in a small population.

Cytogenetic analyses were performed on a triploid hybrid individual produced in the laboratory by mating Poecilia formosa, an all-female gynogenetic diploid species, with a normal male of melanistic ornamental Poecilia sphenops (black molly). Direct chromosome preparations revealed 69 (3n) chromosomes in the somatic complement. This cytogenetic observation in conjunction with the pigmentation phenotype leads us to conclude that occasional failure of sperm exclusion may result in the presence of triploid hybrids even in the natural habitats. ? 1995 The Fisheries Society of the British Isles

Cytogenetic analyses were performed on several populations of the Central American tree frog Agalychnis callidryas, using conventional methods and banding techniques. The karyotype of this species is distinguished by an inversion polymorphism in chromosome 9, which is either submetacentric or telocentric. The populations examined are in Hardy-Weinberg equilibrium with respect to the two alternative morphs of chromosome 9. This is the first report of the occurrence of an intrapopulational chromosomal inversion polymorphism in the order Anura. In male meiosis, the two chromosomes 9 form a bivalent exhibiting a ring-like pairing configuration with terminal chiasmata in both arms, regardless of whether the paired homologs are heteromorphic or homomorphic. Furthermore, individual specimens of A. callidryas exhibit one or two unexpected 18S + 28S ribosomal RNA gene clusters, in addition to the standard nucleolus organizers. The chromosomal localization of these extra nucleolus organizers is identical in all metaphases from the same specimen and shows a specific intraindividual pattern. The karyotype evolution in the phyllomedusine hylids, the structure of the various classes of heterochromatin, and the occurrence and possible origin of the rare inversion polymorphisms and multiple nucleolus organizers in A. callidryas and a few other amphibian species are discussed.

The laser has gained increasing importance during recent years as a useful addition to the physician's therapeutic armoury. Reports on successful prostatectomy by ultrasound-guided laser coagulation in an animal model encouraged us to evaluate the possibility of using this technique for gynecological indications (for example, ectopic pregnancy, endometriosis, ovarian cysts, and other areas of medicine). We used two different laser systems. The devices consisted of a real-time abdominal scanner (4–5 MHz) or a transvaginal sector scanner (5–7.5 MHz) coupled to a neodymium : YAG laser with a 1064 nm wavelength or a holmium : YAG laser with 2120 nm wavelength. An experimental set-up using a water bath revealed clear visibility of the fiber tip and the laser irradiation in real-time ultrasound at 7.5 MHz. Subsequently, a needle (routinely used in transvaginal puncture of ectopic pregnancy) was placed in animal and human tissue specimens and the laser fiber was inserted. The tissue was exposed to both laser systems at different energy densities, pulse rates and pulse widths. The sonographic appearance was recorded and correlated with the macroscopic findings. Results indicate that the holmium : YAG laser at a pulse rate of 5 Hz and a power of 2 watts is effective in removing substantial amounts of tissue. Moreover, it offers immediate visualization and control of the damaged area by producing a strongly echogenic focus. The neodymium : YAG laser showed comparatively poorer sonographic visibility and a precarious thermal mode of action, which are important considerations influencing its applicability. The holmium : YAG laser system with its non-thermal mode of action seems promising for the further clinical application of ultrasound-guided laser surgery. Copyright © 1993 International Society of Ultrasound in Obstetrics and Gynecology

DNA flow cytophotometric measurements demonstrate that the quantity of nuclear DNA of the South American leptodactylid frog Pleurodema brachyops is 3.4 times greater than that of P. thaul. Nevertheless, the conventionally stained karyotypes of both species are nearly identical. In the metaphase chromosomes of P. brachyops, the chromatin has a distinctly higher degree of packaging than in those of P. thaul. C-banding reveals that almost 6 times the constitutive heterochromatin is present in the karyotype of P. brachyops than in the karyotype of P. thaul. Analysis of fluorescence banding patterns shows that the chromosomes of P. brachyops contain AT- and GC-rich heterochromatin, whereas the karyotype of P. thaul is devoid of brightly fluorescing heterochromatin. The substantial differences in the genome sizes of Pleurodema is explained by homogeneous, symmetrical changes in the amounts of all DNA sequence classes along all chromosomes, which preserved the ancestral morphology of the chromosomes.

"The Chromosomes of Terraranan Frogs" is the most comprehensive, original and comparative cytogenetic study of vertebrates ever performed, presenting results obtained over a 36-year period of more than 70 expeditions to the Neotropics (Central and South America and the Caribbean) and from several laboratories (Canada, Costa Rica, Cuba, Germany, USA, and Venezuela). The karyotypes and genomes of 227 recognized species and 2,548 individual specimens of Terrarana were examined using a variety of staining techniques, molecular probe in situ hybridization, and genome size measurements. Furthermore, all previously published data on the cytogenetics of Terrarana have been included. The karyotype characters were mapped onto the molecular phylogeny obtained by analyses of several mitochondrial and nuclear genes. This allowed reconstruction of the various karyophylogenies as well as calculation of the chromosomal mutation rates. The results were always interpreted in context with the cytogenetic data published for other amphibian taxa or vertebrates. The general properties of eukaryote chromosomes, the rules governing structural and numerical chromosome repatterning as well as the principles of chromosome evolution are explained. An extensive 'Material and Methods' section was compiled with the intention of providing the necessary guidance for everyone interested in studying amphibian chromosomes. This monograph provides detailed current and archival information on terraranan karyotypes and genomes to colleagues who are already recognized specialists in amphibian or vertebrate cytogenetics. It is also valuable reading for herpetologists involved in the evolution of neotropical amphibians and provides comprehensive insights into vertebrate cytogenetics for students.

Animal sex chromosome evolution has started on different occasions with a homologous pair of autosomes leading to morphologically differentiated gonosomes. In contrast to other vertebrate classes, among fishes cytologically dernonstrahle sex chromosomes are rare. In reptiles, certain motifs of simple tandemly repeated DNA sequences like (gata)\(_n\)/(gaca)\(_m\) are associated with the constitutive heterochromatin of sex chromosomes. In this study a panel of simple repetitive sequence probes was hybridized to restriction enzyme digested genomic DNA of poeciliid fishes. Apparent male heterogamety previously established by genetic experiments in Poecilia reticulata (guppy) was correlated with male-specific hybridization using the (GACA)\(_4\) probe. The (GATA)\(_4\) oligonucleotide identifies certain male guppies by a Y chromosomal polymorphism in the outbred population. In cantrast none of the genetically defined heterogametic situations in Xiphophorus could be verified consistently using the collection of simple repetitive sequence probes. Only individuals from particular populations produced sex-specific patterns of hybridization with (GATA)\(_4\). Additional poeciliid species (P. sphenops, P. velifera) harbour different sex-specifically organized simple repeat motifs. The observed sex-specific hybridization patterns were substantiated by banding analyses of the karyotypes and by in situ hybridization using the (GACA)\(_4\) probe.

Objective To correlate the appearances of intrafollicular cumulus-like structures using three-dimensional (3D) ultrasound technology with the recovery of mature oocytes. Materials and methods A prospective, observational study using 3D ultrasound examination was performed in 50 women undergoing in vitrofertilization (IVF) after ovarian stimulation. Examination was performed on the day of human chorionic gonadotropin (hCG)-ovulation induction. Estradiol-17-β (E2) levels were measured on the same day. The ovaries were examined for the presence of cumuli in all follicles greater than 15 mm in diameter. Only cumuli demonstrable in all three planes, or in a computer-animated 3D reconstruction, were recorded as ‘visible’. The number, maturity, and fertilization rate of retrieved oocytes were compared with the pre-ovulatory assessment of cumuli by 3D ultrasound. Statistics Correlation coefficients (Pearson) between the variables were calculated and stepwise regression analyses performed. Results A total of 262 cumuli were visualized within 318 follicles from which 296 mature oocytes were retrieved and 218 fertilized. There was a significant correlation between the number of detected cumuli and the number of retrieved oocytes (r2 = 0.83; P < 0.0001), mature oocytes (r2 = 0.78; P < 0.0001), and the number of the fertilized oocytes (r2 = 0.65; P < 0.0001). There was a marked correlation between E2 and the number of follicles and the number of fertilized oocytes. Conclusions Cumulus visualization by 3D ultrasound appears to be an indicator for mature oocytes and successful fertilization. Follicles in which the cumulus cannot be visualized in all three planes are unlikely to contain mature oocytes or oocytes in which fertilization is achieved. Copyright © 2000 International Society of Ultrasound in Obstetrics and Gynecology

Thrombophilia was implicated in the development of pregnancy complications [1, 2, 3, 4, 5, 6], among these recurrent idiopathic pregnancy loss and repeated implantation failure in patients undergoing in-vitro-fertilization (IVF). The endogenous thrombin potential (ETP) has been reported to be increased during hypercoagulable states [6, 7, 8], such as in hereditary thrombophilia and during the intake of oral contraceptives (OC). Aim of this pilot-study was to evaluate the ETP as a global coagulation screening test for a presumptive thrombophilic state in women with recurrent miscarriage and/or repeated implantation failure in patients undergoing IVF.

Hybridization of restriction enzymedigested genomic guppy (Poecilia reticulata, Poeciliidae) DNA with the oligonucleotide probe (GACA)4 revealed a male-specific simple tandem repeat locus, which defines the Y chromosome in outbred populations. The related (GATA)4 probe identifies certain males with the red color phenotype. In contrast only in two out of eight laboratory guppy strains was the typical (GACA)4 band observed. By specific staining of the constitutive heterochromatin one pair of chromosomes could also be identified as the sex chromosomes, confirming the XX/XY mechanism of sex determination. All males exhibit Y chromosomes with a large region of telomeric heterochromatin. Hybridization in situ with nonradioactively labeled oligonucleotide probes localized the (GACA)n repeats to this heterochromatic portion. Together these results may be regarded as a recent paradigm for the differentiation of heteromorphic sex chromosomes from a pair of autosomes during the course of evolution. According to the fish model system, this may have happened in several independent consecutive steps.

Antibodies against annexin-V, a potent anticoagulant abundant in placental tissues, were recently controversially reported to be associated with recurrent miscarriages or failures of in-vitro-fertilization (IVF) attempts.We screened 56 women (34.7 +/- 4.3 years of age: mean +/- 1 SD) with recurrent IVF failures and/or early pregnancy losses for resistance against activated protein C, lupus anticoagulant and antibodies against annexin V, cardiolipin or beta(2)-glycoprotein-1. Among them the prevalence of APC-R (8/56, 14%) and elevated levels of IgG- or IgM-anti-cardiolipin antibodies (7/56, 12%) were more common than elevated levels of (IgG or IgM) antibodies against beta(2)-glycoprotein-1 (3/56, 5%) or annexin-V (1/56, 2%). 42 (75%) of the women had another IVF-attempt after this haemostaseological evaluation and received low molecular weight heparin and/ or acetylsalicylic acid in the case of positivity for APC-resistance, lupus anticoagulant or antibodies against annexin V, cardiolipin or beta(2)-glycoprotein-1.The outcome of these IVF-attempts were 19 pregnancies (34%): 4 early miscarriages (7%) and 15 so far uncomplicated pregnancies (27%). The only woman with an elevated anti-annexin V (IgG) level had had 7 IVF before and received 40 mg Enoxaparin (Lovenox) subcutaneously once daily during the 8(th) IVF, which resulted in a healthy pregnancy.Our findings suggest that among women with recurrent IVF failures anti-annexin V antibody positivity is less prevalent than APC-resistance, lupus anticoagulant (LA) or elevated levels of antibodies against cardiolipin, beta(2)-glycoprotein-1 and that the IVF-result of women with APC-R, LA or with elevated levels of antibodies against annexin V, cardiolipin or beta(2)-glycoprotein might be positively influenced by low molecular weight heparin.

Our findings suggest that among women with recurrent miscarriages and recurrent IVF failures anti-annexin V antibody positivity is less prevalent than APC-resistance, lupus anticoagulant or elevated levels of antibodies against cardiolipin or β2-glycoprotein-1 and that the IVF-result in women with APC-resistance, lupus anticoagulants or elevated levels of antibodies against annexin V, cardiolipin or β2-glycoprotein-1 might be positively influenced by low molecular weight heparin.

In a Zoo-FISH study chicken autosomal chromosome paints 1 to 9 (GGA1–GGA9) were hybridized to metaphase spreads of nine diverse birds belonging to primitive and modern orders. This comparative approach allows tracing of chromosomal rearrangements that occurred during bird evolution. Striking homologies in the chromosomes of the different species were noted, indicating a high degree of evolutionary conservation in avian karyotypes. In two species, the quail and the goose, all chicken paints specifically labeled their corresponding chromosomes. In three pheasant species as well as in the American rhea and blackbird, GGA4 hybridized to chromosome 4 and additionally to a single pair of microchromosomes. Furthermore, in the pheasants fission of the ancestral galliform chromosome 2 could be documented. Hybridization of various chicken probes to two different chromosomes or to only the short or long chromosome arm of one chromosome pair in the species representing the orders Passeriformes, Strigiformes, and Columbiformes revealed translocations and chromosome fissions during species radiation. Thus comparative analysis with chicken chromosome-specific painting probes proves to be a rapid and comprehensive approach to elucidate the chromosomal relationships of the extant birds.

Extensive cytogenetic analyses on a population of the leptodactylid frog Eleutherodactylus riveroi in northern Venezuela revealed the existence of multiple XXAA male/XYAA female/XAA(Y) female sex chromosomes. The XAA(Y) karyotype originated by a centric (Robertsonian) fusion between the original, free Y chromosome and an autosome. 46.2% of the male individuals in this population are carriers of this Y-autosome fusion. In male meiosis the XAA(Y) sex chromosomes pair in the expected trivalent configuration. In the same population 53.8% of the male animals still possess the original, free XY sex chromosomes. E. riveroi is only the second vertebrate species discovered in which a derived Y-autosome fusion coexists with the ancestral free XY sex chromosomes. The free XY sex chromosomes, as well as the multiple XA(Y) sex chromosomes are still in a very primitive (homomorphic) stage of differentiation. With no banding technique applied it is possible to distinguish the Y from the X. Various banding techniques and in situ hybridizations have been carried out to characterize the karyotypes. DNA flow cytometric measurements show that the genome size of E. riveroi resembles that of other Eleutherodactylus species. The cytogenetic data obtained in E. riveroi are compared with those of the sole other vertebrate known to possess the extremely rare, multiple XXAA male/XYAA female/XAA(Y) female sex chromosomes. Surprisingly enough, this vertebrate again is a frog belonging to the genus Eleutherodactylus [E. ((maussi) biporcatus] which lives exactly in the same habitat in northern Venezuela as does E. riveroi.

The karyotypes of four South American didelphid marsupials, representing diploid numbers of 2n = 14 and 18, have been analyzed by a variety of banding techniques. The 2n = 14 karyotypes display a high degree of homoeology, but there also exist distinct similarities between the 2n = 14 and 2n = 18 karyotypes. The interspecific differences found are due to centric fissions, pericentric inversions, and variations in the amount and composition of the constitutive heterochromatin. Contrary to the evolutionary conservation of the banding patterns in all autosomal arms, there are multiple differences in the number and chromosomal location of the nucleolus organ- izer regions. In species with X-linked nucleolus organizers, the 18S + 28S ribosomal RNA genes escape inactivation in female cells. Measurements on the X chromosomes of Marmosa fuscata and Micoureus demerarae unexpectedly reveal the lowest quantities of euchromatin so far determined in the X chromosomes of mammals: 1.5% and 1.8%, respectively, of their haploid female genomes. This is significantly less than the amount of euchromatin in the basic X chromosomes of other marsupials (3%) or eutherians (5%).

The chromosomes of the rare South American marsupial frogs Gastrotheca walkeri and G. ovifera were extensively reexamined with various banding techniques. The karyotypes of both species are distinguished by a new category of XY female symbol /XX male symbol female sex chromosomes. The unusual Y chromosomes are characterized by containing the least amount of constitutive heterochromatin in the karyotypes. This is in contrast to all previously known amphibian Y chromosomes and does not fit the evolutionary model of early XY differentiation in vertebrates. In male meiosis, the heteromorphic XY chromosomes of both species still exhibit the same pairing configurations as the autosomes. DNA flow cytometric measurements show the nuclear DNA amount of G. walkeri to be 10.90 pg. The significance of the XY/XX sex chromosomes of these marsupial frogs, the various classes of constitutive heterochromatin detected, and the data obtained from meiotic analyses are discussed in detail.

Highly differentiated, heteromorphic ZZ female symbol /ZW male symbol sex chromosomes were found in the karyotypes of the neotropical leptodactylid frogs Eleutherodactylus euphronides and E. shrevei. The W chromosomes are the largest heterochromatic, female-specific chromosomes so far discovered in the class Amphibia. The analyses of the banding patterns with AT- and GC base-pair specific fluorochromes show that the constitutive heterochromatin in the giant W chromosomes consists of various categories of repetitive DNA sequences. The W chromosomes of both species are similar in size, morphology and banding patterns, whereas their Z chromosomes exhibit conspicuous differences. In the cell nuclei of female animals, the W chromosomes form very prominent chromatin bodies (W chromatin). DNA flow cytometric measurements demonstrate clear differences in the DNA content of male and female erythrocytes caused by the giant W chromosome, and also shows that these Eleutherodactylus genomes are among the smallest of all amphibian genomes. The importance of the heteromorphic ZW sex chromosomes for the study of Z-linked genes, the similarities and differences of the two karyotypes, and the significance of the exceptionally small genomes are discussed.

The rare human fragile site 16B (FRA16B) has been found to occur spontaneously. Its expression in lymphocyte cultures can also be induced or greatly enhanced by addition of chemicals which are known to bind to AT-rich DNA regions. Following optimal treatment with 150 µg/ml berenil 24 h prior to fixation, the heterozygote frequency of FRA16B is found to be about 5% in populations of European descent. Thus, FRA16B represents the most common of the rare fragile sites. Consistent with cytogenetic observations, the molecular characterization of FRA16B revealed that it is an amplified 33-base pair AT-rich minisatellite repeat. These interindividually variable, extremely large repeat expansions of 15–70 kb in size do not seem to interfere with the expression of genes essential for human development since heterozygotes and homozygotes for FRA16B are normal.

A 15-year cytogenetic survey on one population of the leaf litter frog Eleutherodactylus maussi in northern Venezuela confirmed the existence of multiple XXAA male symbol /XAA(Y) female symbol sex chromosomes which originated by a centric (Robertsonian) fusion between the original Y chromosome and an autosome. 95% of the male individuals in this population are carriers of this Y-autosome fusion. In male meiosis the XAA(Y) sex chromosomes pair in the expected trivalent configuration. In the same population, 5% of the male animals still possess the original, free XY sex chromosomes. In a second population of E. maussi analyzed, all male specimens are characterized by these ancestral XY chromosomes which form normal bivalents in meiosis. E. maussi apparently represents the first vertebrate species discovered in which a derived Y-autosome fusion still coexists with the ancestral free XY sex chromosomes. The free XY sex chromosomes, as well as the multiple XA(Y) sex chromosomes are still in a very primitive (homomorphic) stage of differentiation. With no banding technique applied it is possible to distinguish the Y from the X. DNA flow cytometric measurements show that the genome of E. maussi is among the largest in the anuran family Leptodactylidae. The present study also supplies further data on differential chromosome banding and fluorescence in situ hybridization experiments in this amphibian species.

The study was aimed to investigate the influence of time factors (day, week, month) on pregnancy rates and in vitro fertilization (IVF) and embryo transfer (ET) parameters.8,184 IVF-ET cycles, taking place in two IVF centers from 1992 to 1999, were analyzed. Multiple logistic and linear regression methods were performed as statistical analyses.Oocyte pickup on Tuesdays achieved a significantly higher mean number of oocytes (median: 7) and the highest pregnancy rate (PR) (33.4%) per ET There was a significant variation over the year, with the lowest PR/ET in July (25.71%) and the highest in December (35.5%). Concerning outcomes, the age factor and the number of embryos transferred had the highest significant (p0.0001) influence.The observed weekly rhythm for oocyte pick-ups is certainly due to preprogrammed ovarian stimulation used in our IVF programs. Age as well as the number of embryos transferred are the main influencing factors on a positive outcome and more predictive than seasonal aspects.