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In domestic goats, the polled intersex syndrome (PIS) refers to XX female-to-male sex reversal associated with the absence of horn growth (polled). The causal variant was previously reported as a 11.7 kb deletion at approximately 129 Mb on chromosome 1 that affects the transcription of both FOXL2 and several long non-coding RNAs. In the meantime the presence of different versions of the PIS deletion was postulated and trials to establish genetic testing with the existing molecular genetic information failed. Therefore, we revisited this variant by long-read whole-genome sequencing of two genetically female (XX) goats, a PIS-affected and a horned control. This revealed the presence of a more complex structural variant consisting of a deletion with a total length of 10 159 bp and an inversely inserted approximately 480 kb-sized duplicated segment of a region located approximately 21 Mb further downstream on chromosome 1 containing two genes, KCNJ15 and ERG. Publicly available short-read whole-genome sequencing data, Sanger sequencing of the breakpoints and FISH using BAC clones corresponding to both involved genome regions confirmed this structural variant. A diagnostic PCR was developed for simultaneous genotyping of carriers for this variant and determination of their genetic sex. We showed that the variant allele was present in all 334 genotyped polled goats of diverse breeds and that all analyzed 15 PIS-affected XX goats were homozygous. Our findings enable for the first time a precise genetic diagnosis for polledness and PIS in goats and add a further genomic feature to the complexity of the PIS phenomenon.

Black and tan animals have tan-coloured ventral body surfaces separated by sharp boundaries from black-coloured dorsal body surfaces. In the at mouse mutant, a retroviral 6 kb insertion located in the hair cycle-specific promoter of the murine Asip gene encoding agouti signalling protein causes the black and tan phenotype. In rabbits, three ASIP alleles are thought to exist, including an at allele causing a black and tan coat colour that closely resembles the mouse black and tan phenotype. The goal of our study was to identify the functional genetic variant causing the rabbit at allele. We performed a WGS-based comparative analysis of the ASIP gene in one black and tan and three wt agouti-coloured rabbits. The analysis identified 75 at -associated variants including an 11 kb deletion. The deletion is located in the region of the hair cycle-specific ASIP promoter and thus in a region homologous to the site of the retroviral insertion causing the at allele in mice. We observed perfect association of the genotypes at this deletion with the coat colour phenotype in 49 rabbits. The comparative analysis and the previous knowledge about the regulation of ASIP expression suggest that the 11 kb deletion is the most likely causative variant for the black and tan phenotype in rabbits.

Chromosomal abnormalities are notable causes of infertility in horses. Mares show various degrees of estrous behavior, and ultrasound examination often reveals an underdeveloped genital tract. This article reports investigations on fertility in a Haflinger sibship with a healthy, normally developed, fertile mare with at least three healthy offspring. Chromosomal analysis performed incidentally and blinded for this mare revealed 63,X/64,XX/65,XXX mosaicism. Two closely related mares were also mosaics (63,X/64,XX), and one of them was a carrier of a marker chromosome. Repeated examinations of the mare and seven relatives (four mares and three stallions) did not provide evidence for sub- or in-fertility. They had no developmental abnormalities or conspicuous body conditions. Peripheral blood samples were collected for analysis of the karyotype and molecular analyses. Chromosomes were Giemsa stained and 4',6-diamidino-2-phenylindole banded to identify numerical or structural aberrations of chromosomes and identification of sex chromosomes, respectively. Fluorescence in situ hybridization was performed with an equine Y-chromosome painting probe to identify and count the sex chromosomes, and polymerase chain reaction analysis was used to test for the presence of the SRY gene and investigating chimerism. The present article demonstrates the necessity of further studies analyzing chromosomal X0 mosaics to improve the predictive value of chromosomal aberrations on fertility.

An ectopic ureter is a congenital anomaly which may lead to urinary incontinence and without a surgical intervention even to end-stage kidney disease. A genetic component contributes to the development of this anomaly in Entlebucher mountain dogs (EMD); however, its nature remains unclear. Using the Illumina CanineHD bead chip, a case-control genome-wide association study was performed to identify SNPs associated with the trait. Six loci on canine chromosomes 3, 17, 27 and 30 were identified with 16 significantly associated SNPs. There was no single outstanding SNP associated with the phenotype, and the association signals were not close to known genes involved in human congenital anomalies of the kidney or lower urinary tract. Additional research will be necessary to elucidate the potential role of the associated genes in the development of ectopic ureters in the EMD breed.

The present report describes a 4-year-old Trakehner mare which was referred to the clinic for a breeding soundness evaluation. Clinical, histological, and postmortem examination revealed an underdeveloped genital tract, the absence of a cervix uteri, and small inactive ovaries without male gonadal tissue. Blood lymphocyte analysis revealed an unusual mosaic karyotype consisting of 2 cell lines. For the majority of cells (70%), monosomy X (63,X) was observed. The remaining cells (30%) contained 64 chromosomes including one X chromosome and a small rudimentary Y chromosome consisting mostly of heterochromatin. The centromere was retained, but its full functionality was questionable. PCR analysis revealed that the entire male-specific region of Y (Yq14), including the SRY gene, was deleted. It remained unclear if the pseudoautosomal region (Yq15) and parts of the heterochromatic region (Yq13) were affected by this deletion. The phenotype of the mare with this disorder of sex development associated with sex chromosome abnormalities is genetically comparable to 63,X monosomy which fully explains the clinical findings.

The present case report describes a female cotton-top tamarin which was taken over by the Zoo Basel to form a new breeding pair. The animal demonstrated increased marking behavior without any obvious medical reasons, dominant behavior, failure to become pregnant and its external genitals were ambiguous. A disorder of the sexual development (DSD) was suspected by the zoo veterinarians and in a first step, the chromosomes of the monkey were analyzed. Six cell lines with different karyotypes were observed. The two most frequent cell lines had a 46,XX or a 46,XY karyotype which are normal chromosome complements of female and male cotton-top tamarins, respectively. The other much less frequent cell lines showed numerical aberrations with and without a marker chromosome. Specific biological features of the Callitrichidae, such as natural twinning and genetic chimerism impeded the clarification of the pathogenesis and prevented a reliable prognosis on the fertility of the cotton-top tamarin.Dieser Fallbericht beschreibt ein als Weibchen registriertes Lisztäffchen (Saguinus oedipus), das vom Zoo Basel als Zuchttier übernommen wurde. Aufgrund häufigen Markierens ohne ersichtlichen medizinischen Grund, dominanten Verhaltens, fehlender Fruchtbarkeit und eines phänotypisch nicht eindeutigen äusseren Genitales wurde von den Zootierärzten die vorläufige Verdachtsdiagnose auf eine Störung der Geschlechtsentwicklung gestellt. In einem ersten Schritt wurden die Chromosomen des Äffchens untersucht. Es konnten 6 Zell-Linien mit unterschiedlichen Karyotypen nachgewiesen werden. Am häufigsten wurden die beiden Karyotypen 46,XX und 46,XY beobachtet, die für weibliche bzw. männliche Lisztäffchen normal sind. Die anderen weit weniger häufigen Zell-Linien wiesen numerische Veränderungen der diploiden Chromosomenzahl mit und ohne Marker-Chromosom auf. Biologische Besonderheiten der Krallenaffen, wie Mehrlingsträchtigkeit und genetischer Chimärismus erschwerten die Aufklärung der Pathogenese und verhinderten eine verlässliche Prognose zur Fruchtbarkeit des Lisztäffchens.

Made available in DSpace on 2020-12-12T01:07:41Z (GMT). No. of bitstreams: 0 Previous issue date: 2020-02-01 Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) In the past two decades, average litter size (ALS) in Entlebucher Mountain dogs decreased by approximately 0.8 puppies. We conducted a GWAS for ALS using the single-step methodology to take advantage of 1632 pedigree records, 892 phenotypes and 372 genotypes (173 662 markers) for which only 12% of the dogs had both phenotypes and genotypes available. Our analysis revealed associations towards the growth differentiation factor 9 gene (GDF9), which is known to regulate oocyte maturation. The trait heritability was estimated at 43.1%, from which approximately 15% was accountable by the GDF9 locus alone. Therefore, markers flanking GDF9 explained approximately 6.5% of the variance in ALS. Analysis of WGSs revealed two missense substitutions in GDF9, one of which (g.11:21147009G>A) affected a highly conserved nucleotide in vertebrates. The derived allele A was validated in 111 dogs and shown to be associated with decreased ALS (−0.75 ± 0.22 puppies per litter). The variant was further predicted to cause a proline to serine substitution. The affected residue was immediately followed by a six-residue deletion that is fixed in the canine species but absent in non-canids. We further confirmed that the deletion is prevalent in the Canidae family by sequencing three species of wild canids. Since canids uniquely ovulate oocytes at the prophase stage of the first meiotic division, requiring maturation in the oviduct, we conjecture that the amino acid substitution and the six-residue deletion of GDF9 may serve as a model for insights into the dynamics of oocyte maturation in canids. Department of Preventive Veterinary Medicine and Animal Reproduction School of Agriculture and Veterinarian Sciences São Paulo State University (Unesp), Via de acesso Prof. Paulo Donato Castellane s/n International Atomic Energy Agency Collaborating Centre on Animal Genomics and Bioinformatics, Clóvis Pestana, 793 Department of Support Production and Animal Health School of Veterinary Medicine São Paulo State University (Unesp), Clóvis Pestana, 793 Clinic of Reproductive Medicine Vetsuisse-Faculty University of Zurich, Eschikon 27 Clinic of Reproductive Medicine Vetsuisse-Faculty University of Zurich, Winterthurerstr. 260 Institute of Genetics Vetsuisse-Faculty University of Bern, Bremgartenstrasse 109A Department of Clinics Surgery and Animal Reproduction School of Veterinary Medicine São Paulo State University (Unesp), Clóvis Pestana, 793 Division of Livestook Sciences Department of Sustainable Agriculture System BOKU - University of Natural Resource and Live Sciences, Gregor-Mendel-Straße 33 Department of Preventive Veterinary Medicine and Animal Reproduction School of Agriculture and Veterinarian Sciences São Paulo State University (Unesp), Via de acesso Prof. Paulo Donato Castellane s/n Department of Support Production and Animal Health School of Veterinary Medicine São Paulo State University (Unesp), Clóvis Pestana, 793 Department of Clinics Surgery and Animal Reproduction School of Veterinary Medicine São Paulo State University (Unesp), Clóvis Pestana, 793 FAPESP: 2016/05787-7 FAPESP: 2017/08373-1

Abnormalities of chromosomes are an important and well documented cause of disorders of sexual development, fertility problems and congenital anomalies in mammals. Detection of low-level 63,X/64,XX mosaicism during routine cytogenetic evaluation is a challenge because its clinical significance is not yet fully clear. This study describes the prevalence and levels of 63,X mosaicism for a cohort of fertile mares and compares the results with eight problem mares for which no clinical cause of sub-fertility was found. The study design allowed for the analysis of micronuclei which are biomarkers of genomic instability and can disturb cell divisions, drive cancer development or cause congenital diseases. Although 27% of the fertile mares were identified to be 63,X mosaics, the results showed that the rates of abnormal cells were very low (1–3%). Levels of abnormal cells in problem mares with 63,X mosaicism were similar or higher. The average rate of micronuclei in the blood of the fertile mares was ∼1%, well below the baseline (5%) which was proposed for peripheral blood of normal healthy humans. We found weak to modest, but not significant, correlations between the age of fertile mares and 63,X cells (Kendall’s tau b = 0.2905; p > 0.05) as well as the rate of micronuclei (Kendall’s tau b = 0.1896; p > 0.05). Likewise, the correlation between presence of a 63,X cell line and micronuclei rate was not significant (Kendall’s tau b = 0.3201; p > 0.05). The presence of 63,X cells in rates greater than 3% may indeed indicate a higher risk for sub-fertility and eventually for associated health problems in such mares. Detection and elimination of mares with high level of X aneuploidies from breeding may have a positive effect on the fertility within the general horse population. This data may support the evaluation of problem mares with mosaic karyotypes involving the X chromosome.

Background In the past 10 years, the frequency of unplanned cesarean sections in the Labrador Retriever breeding colony at Guiding Eyes for the Blind stayed around 10% (range 5% to 28%). To reduce the number of cesarean sections, factors influencing the occurrence of a cesarean section need to be known. The goal of this study was to identify factors that contribute to the decision to perform a cesarean section. Results Of the 688 Labrador Retriever litters whelped between 2003 and 2016, 667 litters had sufficient data and remained in the analysis. The target trait was ordinal with the three levels “normal whelping”, “assisted whelping” and “cesarean section”. A general ordinal logistic regression approach was used to analyze the data. Model selection with possible predictors resulted in a final model including weight of the dam, the weight of the heaviest puppy of a litter, the number of fetuses malpositioned and the quality of uterine contractions. Weight and size of a litter, parity, maternal inbreeding coefficient, whelping season, dam and sire were dropped from the model because they were not significant. The risk of a cesarean section was influenced by the combination of the weight of the dam and the weight of the heaviest puppy in the litter, as well as by the number of malpositioned fetuses and the quality of the contractions. Larger puppies increased the risk of cesarean section especially when the dam had a lighter weight. For dams weighing 23.6 kg and 32.8 kg the predicted probability of a cesarean section was low, with 0.06 and 0.02, respectively, when the heaviest puppy in a litter was light (0.42 kg), contractions were normal and no fetus was malpositioned. However, the probability of a cesarean section was much higher, ranging from 0.24 to 0.08, when the heaviest puppy in a litter was heavy (0.66 kg). Conclusions Means to reduce the cesarean section frequency in this Labrador Retriever breeding colony should include genetic selection for ideal puppy weight. In addition, dams with an adult body weight substantially below average should not be selected as breeders in this colony. Electronic supplementary material The online version of this article (10.1186/s12917-018-1381-8) contains supplementary material, which is available to authorized users.

The present case report describes a 6-year old subfertile pony mare, which became pregnant after the eleventh artificial insemination. The examination of the ovaries and the uterus did not reveal any abnormal clinical findings and the mare showed a regular oestrous cycle. Based on cytogenetic and molecular genetic analyses it became possible to elucidate the observed subfertility. The mosaic karyotype of the mare consisted of 63,X (20%) and 64,XX (80%) cells. A PCR analysis failed to amplify sequences from the equine SRY gene. The observed classic 63,X/64,XX mosaicism is a plausible explanation for the subfertility of the mare.Der vorliegende Fallbericht beschreibt eine 6 Jahre alte subfertile Stute der Rasse Ponystute, die erst nach der elften künstlichen Besamung tragend wurde. Die klinische Untersuchung der Ovarien und des Uterus waren ohne besonderen Befund und die Stute zeigte einen regelmässigen Zyklus. Aufgrund von zytogenetischen und molekulargenetischen Untersuchungen wurde es möglich, eine plausible Ursache für die Subfertilität zu ermitteln. Der Karyotyp der Ponystute setzte sich aus zwei Zelllinien zusammen: In 20% der Zellen wurde eine Monosomie X nachgewiesen (63,X) und in 80% der Zellen war der Karyotyp normal (64,XX). Eine PCR-Analyse für den Nachweis von Sequenzen des equinen SRY Gen war negativ. Es handelte sich also um einen klassischen Fall einer mosaiken Form von Monosomie X, die eine offensichtliche Erklärung für die Subfertilität der Stute liefert.

A good reproductive performance is a central element of animal breeding. The breeders of Entlebucher Mountain dogs observed a decrease of the mean litter size and an increase of the number of unsuccessful matings in the past years. The aim of the present study was to identify factors with an influence on fertility in this breed. In total, 915 litters from 202 sires and 348 dams from 1986 to 2013 entered the analyses. The total puppy losses (7.4%) reduced the mean litter size at birth of 5.49 ± 2.13 to a mean litter size at registration of 5.08 ± 2.05. There was no deviation from the expected equal sex distribution for puppies at birth and at registration, as well as for puppy losses consisting of stillborn puppies and puppies which died or had to be euthanized before registration. The mean annual litter inbreeding coefficient increased from 0.37 in 1986 to 0.40 in 2013 and was correlated with the year of birth of the litter (Kendall's tau b = 0.46). The age of the dam and parental inbreeding were identified as significant predictors with a negative effect on litter size at birth. For the litter size at registration the age and inbreeding of the dam had a significant negative effect and a 1% increase of dam inbreeding is expected to decrease the litter size at birth and registration by 0.1 and 0.09 puppies, respectively. The occurrence of total puppy losses decreased during the years and was more frequent in larger litters. In addition, in litters of older parents the occurrence of puppy losses was more frequent than in litters from younger parents. The final generalized linear mixed-effects models for litter size at birth, litter size at registration and for total puppy losses explained 36%, 33% and 22% of the total variance, respectively. The impact of inbreeding and parental age on fertility of the Entlebucher Mountain dog was small and the influence of the dam was much bigger than the one of the sire. Other factors must be responsible for the variability of litter sizes not explained by the models. Without changes of breeding circumstances, a further increase of inbreeding must be expected. Therefore, a close monitoring and minimizing of inbreeding must be followed up by the breeding community.

Belted cattle have a circular belt of unpigmented hair and skin around their midsection. The belt is inherited as a monogenic autosomal dominant trait. We mapped the causative variant to a 54 kb segment on bovine chromosome 3. Whole genome sequence data of 2 belted and 130 control cattle yielded only one private genetic variant in the critical interval in the two belted animals. The belt-associated variant was a copy number variant (CNV) involving the quadruplication of a 6 kb non-coding sequence located approximately 16 kb upstream of theTWIST2gene. Increased copy numbers at this CNV were strongly associated with the belt phenotype in a cohort of 239 cases and 1303 controls (p = 1.3 x 10-278). We hypothesized that the CNV causes aberrant expression ofTWIST2during neural crest development, which might negatively affect melanoblasts. Functional studies showed that ectopic expression of bovineTWIST2in neural crest in transgenic zebrafish led to a decrease in melanocyte numbers. Our results thus implicate an unsuspected involvement of TWIST2 in regulating pigmentation and reveal a non-coding CNV underlying a captivating Mendelian character.Author SummaryBelted cattle, a spontaneous coat color mutant, have been recognized at least 600 years ago. The striking pigmentation pattern probably has arisen in medieval cattle of the Alpine region. The belt still segregates in Brown Swiss cattle and it has become a breed-defining character in the Lakenvelder or Dutch Belted cattle. The belted allele has also been introgressed into Galloways to form the Belted Galloways. We report here the causative genetic variant, a non-coding copy number variant (CNV) upstream of theTWIST2gene. We hypothesize that the CNV leads to ectopic expression of TWIST2 in the neural crest, which negatively affects melanocyte development. Overexpression of bovine TWIST2 in transgenic zebrafish embryos led to a decrease in melanocyte numbers, which provides functional support for our hypothesis.

The present case report describes a stray cat with a female appearance. The new owners requested to neuter the animal. During surgery the veterinarian could not find any gonadal tissue. After puberty the cat showed more and more male behaviour. The owners of the cat were interested to know the cause of the abnormal behaviour, but forbid any further clinical tests or surgery. Based upon cytogenetic and molecular genetic experiments a diagnosis became possible. The uniform karyotype (38,XY) was in accordance with the karyotype of a male cat and it was possible to amplify the SR Y gene by PCR. The cat represents a case of SRYpositive sex reversal.

A 23-month-old tomcat was referred to our clinic because of male behavioral problems, cryptorchidism, and an undefined intra-abdominal organ resembling a uterus. Ultrasonography and computed tomography showed 2 fluid-filled tubular structures dorsolaterally to the bladder and connected to the pelvic urethra. The cat was castrated, and the tubular structures were surgically removed. Histology identified them as Müllerian duct remnants. The testes were hypoplastic, the epididymes and deferent ducts were normal. Cytogenetic analyses revealed the presence of a mosaic 37,X/38,XY karyotype which explains the clinical findings.

Belted cattle have a circular belt of unpigmented hair and skin around their midsection. The belt is inherited as a monogenic autosomal dominant trait. We mapped the causative variant to a 37 kb segment on bovine chromosome 3. Whole genome sequence data of 2 belted and 130 control cattle yielded only one private genetic variant in the critical interval in the two belted animals. The belt-associated variant was a copy number variant (CNV) involving the quadruplication of a 6 kb non-coding sequence located approximately 16 kb upstream of the TWIST2 gene. Increased copy numbers at this CNV were strongly associated with the belt phenotype in a cohort of 333 cases and 1322 controls. We hypothesized that the CNV causes aberrant expression of TWIST2 during neural crest development, which might negatively affect melanoblasts. Functional studies showed that ectopic expression of bovine TWIST2 in neural crest in transgenic zebrafish led to a decrease in melanocyte numbers. Our results thus implicate an unsuspected involvement of TWIST2 in regulating pigmentation and reveal a non-coding CNV underlying a captivating Mendelian character.