C. Zijlstra

Exposure of Oocytes to the Fusarium Toxins Zearalenone and Deoxynivalenol Causes Aneuploidy and Abnormal Embryo Development in Pigs

Abstract Fungi of the Fusarium species can infect food and feed commodities and produce the mycotoxins zearalenone (ZEA) and deoxynivalenol (DON). Since both toxins have been reported to reduce fertility, the mechanisms of ZEA and DON on inhibition of oocyte maturation were examined. Pig oocytes were matured in the presence of ZEA (a mycotoxin with estrogenlike activity), 17beta-estradiol, and DON (all 3.12 μmol/L). Zearalenone, 17beta-estradiol, and DON inhibited oocyte maturation and caused approximately 34% of the oocytes to form an aberrant spindle. Different ratios of ZEA:DON did not lead to a more severe inhibition of oocyte maturation. Both mycotoxins caused abnormal formation of the meiotic spindle. The developmental competence of oocytes matured in the presence of mycotoxins was further investigated after in vitro fertilization. Presence of ZEA (3.12 μmol/L) during maturation reduced the percentages of oocytes that cleaved and formed a blastocyst to about 12%, compared with 25% of control oocytes. Maturation in the presence of equimolar concentrations of DON was not compatible with development. The ploidy of blastomeres from blastocysts derived from mycotoxin-exposed oocytes was analyzed with fluorescent in situ hybridization. All blastocysts, even those from the control group, contained at least one blastomere with abnormal ploidy, but the variation in the percentages of aneuploid blastomeres was significantly larger in embryos from oocytes exposed to mycotoxins. It is concluded that ZEA and DON can lead to abnormal spindle formation, leading to less fertile oocytes and embryos with abnormal ploidy, and that the effects of ZEA and DON are not synergistic.

The PiGMaP consortium cytogenetic map of the domestic pig (Sus scrofa domestica)

llNRA, Laboratoire de Grnrtique Cellulaire, BP27, 31326 Castanet-Tolosan, France 2Department of Functional Morphology, Faculty of Veterinary Medicine, Utrecht University, Utrecht, The Netherlands 3Department of Animal Breeding and Genetics, Swedish University of Agricultural Sciences, Uppsala, Sweden 4Division of Anatomy, Department of Anatomy and Physiology, The Royal Veterinary and Agricultural University, Copenhagen, Denmark 5Division of Animal Genetics, Department of Animal Science and Animal Health, The Royal Veterinary and Agricultural University, Copenhagen, Denmark 6Department of Clinical Genetics, University of Ulm, Ulm, Germany 7Laboratoire de Cytomrtrie, CEA, Fontenay-aux Roses, France

Establishment of an R-banded rabbit karyotype nomenclature by FISH localization of 23 chromosome-specific genes on both G- and R-banded chromosomes

Direct detection of fluorescent in situ hybridization signals on R-banded chromosomes stained with propidium iodide is a rapid and efficient method for constructing cytogenetic maps for species with R-banded standard karyotypes. In this paper, our aim is to establish an R-banded rabbit karyotype nomenclature that is in total agreement with the 1981 G-banded standard nomenclature. For this purpose, we have produced new GTG- and RBG-banded mid-metaphase karyotypes and an updated version of ideograms of R-banded rabbit chromosomes. In addition, to confirm correlations between G- and R-banded chromosomes, we have defined a set of 23 rabbit BAC clones, each containing a specific gene, one marker gene per rabbit chromosome, and we have localized precisely each BAC clone by FISH on both G- and R-banded chromosomes.

Localization of 18S + 28S and 5S ribosomal RNA genes in the dog by fluorescence in situ hybridization.

The gene clusters encoding 18S + 28S and 5S rRNA in the dog (Canis familiaris) have been localized by using GTG-banding and fluorescence in situ hybridization. The 18S + 28S rDNA maps to chromosome regions 7q2.5-->q2.7, 17q1.7, qter of a medium-sized, not yet numbered autosome, and Yq1.2-->q1.3. Our data show that there is one cluster of 5S rDNA in the dog, which maps to chromosome region 4q1.4.

The canine sarcoglycan delta gene: BAC clone contig assembly, chromosome assignment and interrogation as a candidate gene for dilated cardiomyopathy in Dobermann dogs.

Dilated cardiomyopathy (DCM) is a common disease of the myocardium recognized in human, dog and experimental animals. Genetic factors are responsible for a large proportion of cases in humans, and 17 genes with DCM causing mutations have been identified. The genetic origin of DCM in the Dobermann dogs has been suggested, but no disease genes have been identified to date. In this paper, we describe the characterization and evaluation of the canine sarcoglycan delta (SGCD), a gene implicated in DCM in human and hamster. Bacterial artificial chromosomes (BACs) containing the canine SGCD gene were isolated with probes for exon 3 and exons 4–8 and were characterized by Southern blot analysis. BAC end sequences were obtained for four BACs. Three of the BACs overlapped and could be ordered relative to each other and the end sequences of all four BACs could be anchored on the preliminary assembly of the dog genome sequence (www. ensembl.org). One of the BACs of the partial contig was localized by fluorescent in situ hybridization to canine chromosome 4q22, in agreement with the dog genome sequence. Two highly informative polymorphic microsatellite markers in intron 7 of the SGCD gene were identified. In 25 DCM-affected and 13 non DCM-affected dogs seven different haplotypes could be distinguished. However, no association between any of the SGCD variants and the disease locus was apparent.

Localization of the 18S, 5.8S and 28S rRNA genes and the 5S rRNA genes in the babirusa and the white-lipped peccary

The locations of the genes encoding 18S, 5.8S and 28S rRNA and 5S rRNA were studied in two relatives of the domestic pig, the babirusa (Babyrousa babyrussa) and the white-lipped peccary (Tayassu pecari). In the babirusa, the 18S, 5.8S and 28S rDNA is located on chromosomes 6, 8 and 10. The genes on chromosomes 8 and 10 are actively transcribed, in contrast to those on chromosomes 6. In the white-lipped peccary, this rDNA was found to be located on chromosomes 4 and 8. The genes on both of these pairs of chromosomes are actively transcribed. The 5S rDNA was physically mapped to chromosome 16 in the babirusa, and to chromosome 11 in the white-lipped peccary. These data are compared to similar data obtained for the domestic pig, and confirm previously recognized chromosome homologies.

Persistent Mullerian duct syndrome in a Miniature Schnauzer dog with signs of feminization and a Sertoli cell tumour

A 5-year-old male Miniature Schnauzer was presented with unilateral cryptorchidism and signs of feminization. Abdominal ultrasonography revealed an enlarged right testis and a large, fluid-filled cavity that appeared to arise from the prostate. Computed tomography revealed the cavity to be consistent with an enlarged uterine body, arising from the prostate, and showed two structures resembling uterine horns that terminated close to the adjacent testes. The dog had a normal male karyotype, 78 XY. Gonadohysterectomy was performed and both the surgical and the histological findings confirmed the presence of a uterus in this male animal, resulting in a diagnosis of persistent Mullerian duct syndrome (PMDS). The enlarged intra-abdominal testis contained a Sertoli cell tumour. Computed tomography proved to be an excellent diagnostic tool for PMDS.

Comparative chromosome painting between the domestic pig (Sus scrofa) and two species of peccary, the collared peccary (Tayassu tajacu) and the white-lipped peccary (T. pecari): a phylogenetic perspective

The Suidae and the Dicotylidae (or Tayassuidae) are related mammalian families, both belonging to the artiodactyl suborder Suiformes, which diverged more than 37 million years ago. Cross-species chromosome painting was performed between the domestic pig (Sus scrofa; 2n = 38), a representative of the Suidae, and two species of the Dicotylidae: the collared peccary (Tayassu tajacu; 2n = 30) and the white-lipped peccary (T. pecari; 2n = 26). G-banded metaphase chromosomes of the two peccaries were hybridized with whole chromosome painting probes derived from domestic pig chromosomes 1–18 and X. For both peccary species, a total of 31 autosomal segments that are conserved between pig and peccary could be identified. The painting results confirm conclusions inferred from G-band analyses that the karyotypes of the collared peccary and the white-lipped peccary are largely different. The karyotypic heterogeneity of the Dicotylidae contrasts with the relative homogeneity among the karyotypes of the Suidae. For this difference between the Dicotylidae and the Suidae, a number of explanations are being postulated: 1) the extant peccaries are phylogenetically less closely related than is usually assumed; 2) the peccary genome is less stable than the genome of the pigs; and 3) special (e.g. biogeographical or biosocial) circumstances have facilitated the fixation of chromosome rearrangements in ancestral dicotylid populations.

A satellite DNA element specific for roe deer (Capreolus capreolus).

Abstract. An abundant tandem repetitive DNA segment (CCsatIII) with a repeat unit of 2.2 kb has been found in the genome of roe deer (Capreolus capreolus). It accounts for approximately 5%–10% of the genome and is only present in the two species of the genus Capreolus. The sequence has no similarity or common motifes with other deer satellite DNAs and there is no internal repeat structure. A 93 bp fragment is homologous to a bovine repeat. Fluorescent in situ hybridisation showed a predominant centromeric staining of most chromosomes accompanied by a weak interstitial staining of the same chromosomes. On Southern blots, CCsatIII probes do not discriminate between the closely related Capreolus species.

Prostasomes as a source of diagnostic biomarkers for prostate cancer

New biomarkers are needed to improve the diagnosis of prostate cancer. Similarly to healthy cells, prostate epithelial cancer cells produce extracellular vesicles (prostasomes) that can be isolated from seminal fluid, urine, and blood. Prostasomes contain ubiquitously expressed and prostate-specific membrane and cytosolic proteins, as well as RNA. Both quantitative and qualitative changes in protein, mRNA, long noncoding RNA, and microRNA composition of extracellular vesicles isolated from prostate cancer patients have been reported. In general, however, the identified extracellular vesicle-associated single-marker molecules or combinations of marker molecules require confirmation in large cohorts of patients to validate their specificity and sensitivity as prostate cancer markers. Complications include variable factors such as prostate manipulation and urine flux, as well as masking by ubiquitously expressed free molecules and extracellular vesicles from tissues other than the prostate. Herein, we propose that the most promising methods include comprehensive combinational screening for (mutant) RNA in prostasomes that are immunoisolated with antibodies targeting prostate-specific epitopes.