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Naked foal syndrome (NFS) is a genodermatosis in the Akhal-Teke horse breed. We provide the first scientific description of this phenotype. Affected horses had almost no hair and showed a mild ichthyosis. So far, all known NFS affected horses died between a few weeks and 3 years of age. It is not clear whether a specific pathology caused the premature deaths. NFS is inherited as a monogenic autosomal recessive trait. We mapped the disease causing genetic variant to two segments on chromosomes 7 and 27 in the equine genome. Whole genome sequencing of two affected horses, two obligate carriers, and 75 control horses from other breeds revealed a single non-synonymous genetic variant on the chromosome 7 segment that was perfectly associated with NFS. The affected horses were homozygous for ST14:c.388G>T, a nonsense variant that truncates more than 80% of the open reading frame of the ST14 gene (p.Glu130*). The variant leads to partial nonsense mediated decay of the mutant transcript. Genetic variants in the ST14 gene are responsible for autosomal recessive congenital ichthyosis 11 in humans. Thus, the identified equine ST14:c.388G>T variant is an excellent candidate causative variant for NFS and the affected horses represent a large animal model for a known human genodermatosis. Our findings will enable genetic testing to avoid the non-intentional breeding of NFS affected foals.

The Ehlers-Danlos syndromes (EDSs) are a heterogeneous group of inherited connective tissue disorders characterized by skin hyperextensibility, joint hypermobility and tissue fragility. Inherited disorders similar to human EDS have been reported in different mammalian species. In the present study, we investigated a female mixed-breed dog with clinical signs of EDS. Whole-genome sequencing of the affected dog revealed two missense variants in the TNXB gene, encoding the extracellular matrix protein tenascin XB. In humans, TNXB genetic variants cause classical-like EDS or the milder hypermobile EDS. The affected dog was heterozygous at both identified variants. Each variant allele was transmitted from one of the case's parents, consistent with compound heterozygosity. Although one of the variant alleles, XM_003431680.3:c.2012G>A, p.(Ser671Asn), was private to the family of the affected dog and absent from whole-genome sequencing data of 599 control dogs, the second variant allele, XM_003431680.3:c.2900G>A, p.(Gly967Asp), is present at a low frequency in the Chihuahua and Poodle population. Given that TNXB is a functional candidate gene for EDS, we suggest that compound heterozygosity for the identified TNXB variants may have caused the EDS-like phenotype in the affected dog. Chihuahuas and Poodles should be monitored for EDS cases, which might confirm the hypothesized pathogenic effect of the segregating TNXB variant.

Two clinical forms of ichthyosis in cattle have been reported, ichthyosis fetalis and congenital ichthyosis. Ichthyosis poses animal welfare and economic issues and the more severe form, ichthyosis fetalis, is lethal. A Shorthorn calf with ichthyosis fetalis was investigated and a likely causal missense variant on chromosome 2 in the ABCA12 gene (NM_001191294.2:c.6776T>C) was identified by whole genome sequencing. Mutations in the ABCA12 gene are known to cause ichthyosis fetalis in cattle and Harlequin ichthyosis in humans. Sanger sequencing of the affected calf and the dam confirmed the variant was homozygous in the affected calf and heterozygous in the dam. Further genotyping of 130 Shorthorn animals from the same property revealed an estimated allele frequency of 3.8%. The presented findings enable genetic testing for breeding and diagnostics.

Major characteristics of coat variation in dogs can be explained by variants in only a few genes. Until now, only one missense variant in the KRT71 gene, p.Arg151Trp, has been reported to cause curly hair in dogs. However, this variant does not explain the curly coat in all breeds as the mutant 151Trp allele, for example, is absent in Curly Coated Retrievers. We sequenced the genome of a Curly Coated Retriever at 22× coverage and searched for variants in the KRT71 gene. Only one protein‐changing variant was present in a homozygous state in the Curly Coated Retriever and absent or present in a heterozygous state in 221 control dogs from different dog breeds. This variant, NM_001197029.1:c.1266_1273delinsACA, was an indel variant in exon 7 that caused a frameshift and an altered and probably extended C‐terminus of the KRT71 protein NP_001183958.1:p.(Ser422ArgfsTer?). Using Sanger sequencing, we found that the variant was fixed in a cohort of 125 Curly Coated Retrievers and segregating in five of 14 additionally tested breeds with a curly or wavy coat. KRT71 variants cause curly hair in humans, mice, rats, cats and dogs. Specific KRT71 variants were further shown to cause alopecia. Based on this knowledge from other species and the predicted molecular consequence of the newly identified canine KRT71 variant, it is a compelling candidate causing a second curly hair allele in dogs. It might cause a slightly different coat phenotype than the previously published p.Arg151Trp variant and could potentially be associated with follicular dysplasia in dogs.

The skin is the largest organ of the body. It provides a barrier that protects the body from harmful environmental factors as well as from a loss of fluids. Impaired skin integrity can lead to a disruption of the skin barrier and to disease. Genodermatoses are a heterogenous group of mostly rare single gene disorders affecting the skin and its appendages. After decades of genetic and due to recent advances in technology, the genetic cause of numerous genodermatoses was uncovered. Based on the function of the involved genes and the affected skin compartments, monogenic skin disorders can be grouped into ichthyoses, epidermolysis and blistering disorders, pigmentation disorders, disorders of ectodermal appendages, vascular disorders, connective tissue defects, dermal mosaics, and genodermatoses with tumor predisposition. Many human genodermatoses have a very similar counterpart in animals. Spontaneous mutants in purebred animals such as dogs that have a unique population structure and physical similarity to humans, are therefore valuable models for human genodermatoses. In this thesis, I took part in the analysis of 12 canine, equine and feline phenotypes with manifestations in skin or its appendages applying different genetic mapping techniques and whole genome sequencing. In cats, a frameshift variant in the COL5A1 gene was identified in a single case with Ehlers-Danlos syndrome. In a female cat with inflammatory linear epidermal lesions, a missense variant in the X-chromosomal NSDHL gene was found, explaining the observed cutaneous mosaicism. Interestingly, I identified a large deletion in the same gene, NSDHL, underlying a related congenital cornification disorder in a Labrador Retriever and her equally affected crossbred daughter. In a single canine case of ichthyosis, whole genome sequencing revealed a de novo variant in the ASPRV1 gene, probably disturbing filaggrin processing during cornification. A deregulation in cornification of nasal keratinocytes was also suspected in a litter of Greyhounds with nasal parakeratosis, in which I identified a splice defect in the SUV39H2 gene. Pigmentation and hair texture are diverse in dogs. During a study in Chow Chows with coat colour dilution I identified a variant in the MLPH gene encoding melanophilin, and found that this variant segregated in other dog breeds as well. In dogs with oculocutaneous albinism, variants in OCA2 and SLC45A2 encoding transporters in membranes of melanosomes were identified. In dogs with a previously unknown cause for their curly coat, a candidate gene approach led to the identification of a second KRT71 allele for curls, which has a potential predisposing role for hair loss that warrants further investigation. Hairless skin patches on the head and back was also a prominent feature in male Dachshund puppies with X-linked hypohidrotic ectodermal dysplasia, where a single base deletion in the EDA gene was found using targeted Sanger sequencing. Lethal acrodermatitis in Bull terriers is a disorder causing early death of affected animals. All cases in our cohort were homozygous for a splice defect in MKLN1, a gene not yet described in human genodermatoses. Finally, in horses from the Akhal-Teke breed, I identified a nonsense variant in the ST14 gene most likely causing the lethal naked foal syndrome in a monogenic autosomal recessive mode. The identification of these candidate causative variants enables genetic testing, controlled breeding and in the long term eradication of the corresponding disorders from the animal population. The majority of identified variants was located in genes already known from human disorders. However, ASPRV1 and MKLN1 variants have never been reported as cause for human genodermatoses. This thesis therefore demonstrates that genetic anlysis of spontaneous animal mutants offers the chance to gain new biological knowledge and candidate genes for rare human genodermatoses.

Hereditary nasal parakeratosis (HNPK), described in the Labrador Retriever breed, is a monogenic autosomal recessive disorder that causes crusts and fissures on the nasal planum of otherwise healthy dogs. Our group previously showed that this genodermatosis may be caused by a missense variant located in the SUV39H2 gene encoding a histone 3 lysine 9 methyltransferase, a chromatin modifying enzyme with a potential role in keratinocyte differentiation. In the present study, we investigated a litter of Greyhounds in which six out of eight puppies were affected with parakeratotic lesions restricted to the nasal planum. Clinically and histologically, the lesions were comparable to HNPK in Labrador Retrievers. Whole genome sequencing of one affected Greyhound revealed a 4-bp deletion at the 5'-end of intron 4 of the SUV39H2 gene that was absent in 188 control dog and three wolf genomes. The variant was predicted to disrupt the 5'-splice site with subsequent loss of SUV39H2 function. The six affected puppies were homozygous for the variant, whereas the two non-affected littermates were heterozygous. Genotyping of a larger cohort of Greyhounds revealed that the variant is segregating in the breed and that this breed might benefit from genetic testing to avoid carrier × carrier matings.

Ehlers-Danlos syndrome (EDS) is a group of heritable connective tissue disorders caused by defective collagen synthesis or incorrect assembly of the collagen triple helical structure. EDS is characterised by joint hypermobility, skin hyperextensibility, abnormal scarring, poor wound healing and tissue friability. Human EDS may be caused by variants in several different genes including COL5A1, which encodes the collagen type V alpha 1 chain. For the present study we investigated a 1.5-year-old, spayed female, domestic shorthair cat with EDS. The affected cat showed multiple recurrent skin tears, hyperextensibility of the skin and joint abnormalities. We obtained whole genome sequencing data from the affected cat and searched for variants in candidate genes known to cause EDS. We detected a heterozygous single base-pair deletion in exon 43 of the COL5A1 gene, namely c.3420delG. The deletion was predicted to result in a frameshift and premature stop codon: p.(Leu1141SerfsTer134). Sanger sequencing confirmed that the variant was present in the affected cat and absent from 103 unaffected cats from different breeds. The variant was also absent from a Burmese cat with EDS. Based on knowledge about the functional impact of COL5A1 variants in other species, COL5A1:c.3420delG represents a compelling candidate causative variant for the observed EDS in the affected cat.

Coat colour dilution may be the result of altered melanosome transport in melanocytes. Loss-of-function variants in the melanophilin gene (MLPH) cause a recessively inherited form of coat colour dilution in many mammalian and avian species including the dog. MLPH corresponds to the D locus in many domestic animals, and recessive alleles at this locus are frequently denoted with d. In this study, we investigated dilute coloured Chow Chows whose coat colour could not be explained by their genotype at the previously known MLPH:c.-22G>A variant. Whole genome sequencing of such a dilute Chow Chow revealed another variant in the MLPH gene: MLPH:c.705G>C. We propose to designate the corresponding mutant alleles at these two variants d1 and d2 . We performed an association study in a cohort of 15 dilute and 28 non-dilute Chow Chows. The dilute dogs were all either compound heterozygous d1 /d2 or homozygous d2 /d2 , whereas the non-dilute dogs carried at least one wildtype allele D. The d2 allele did not occur in 417 dogs from diverse other breeds. However, when we genotyped a Sloughi family, in which a dilute coloured puppy had been born out of non-dilute parents, we again observed perfect co-segregation of the newly discovered d2 allele with coat colour dilution. Finally, we identified a blue Thai Ridgeback with the d1 /d2 genotype. Thus, our data identify the MLPH:c.705G>C as a variant explaining a second canine dilution allele. Although relatively rare overall, this d2 allele is segregating in at least three dog breeds, Chow Chows, Sloughis and Thai Ridgebacks.

We investigated a German Spitz family where the mating of a black male to a white female had yielded three puppies with an unexpected light brown coat color, lightly pigmented lips and noses, and blue eyes. Combined linkage and homozygosity analysis based on a fully penetrant monogenic autosomal recessive mode of inheritance identified a critical interval of 15 Mb on chromosome 3. We obtained whole genome sequence data from one affected dog, three wolves, and 188 control dogs. Filtering for private variants revealed a single variant with predicted high impact in the critical interval in LOC100855460 (XM_005618224.1:c.377+2T>G LT844587.1:c.-45+2T>G). The variant perfectly co-segregated with the phenotype in the family. We genotyped 181 control dogs with normal pigmentation from diverse breeds including 22 unrelated German Spitz dogs, which were all homozygous wildtype. Comparative sequence analyses revealed that LOC100855460 actually represents the 5'-end of the canine OCA2 gene. The CanFam 3.1 reference genome assembly is incorrect and separates the first two exons from the remaining exons of the OCA2 gene. We amplified a canine OCA2 cDNA fragment by RT-PCR and determined the correct full-length mRNA sequence (LT844587.1). Variants in the OCA2 gene cause oculocutaneous albinism type 2 (OCA2) in humans, pink-eyed dilution in mice, and similar phenotypes in corn snakes, medaka and Mexican cave tetra fish. We therefore conclude that the observed oculocutaneous albinism in German Spitz is most likely caused by the identified variant in the 5'-splice site of the first intron of the canine OCA2 gene.

Oculocutaneous albinism type 4 (OCA4) in humans and similar phenotypes in many animal species are caused by variants in the SLC45A2 gene, encoding a putative sugar transporter. In dog, two independent SLC45A2 variants are known that cause oculocutaneous albinism in Doberman Pinschers and several small dog breeds respectively. For the present study, we investigated a Bullmastiff with oculocutaneous albinism. The affected dog was highly inbred and resulted from the mating of a sire to its own grandmother. We obtained whole genome sequence data from the affected dog and searched specifically for variants in candidate genes known to cause albinism. We detected a single base deletion in exon 6 of the SLC45A2 gene (NM_001037947.1:c.1287delC) that has not been reported thus far. This deletion is predicted to result in an early premature stop codon. It was confirmed by Sanger sequencing and perfectly co-segregated with the phenotype in the available family members. We genotyped 174 unrelated dogs from diverse breeds, all of which were homozygous wildtype. We therefore suggest that SLC45A2:c.1287delC causes the observed oculocutaneous albinism in the affected Bullmastiff.

Sve više studenata i akademika poseže za mrežnim izvorima kod pisanja znanstvenih i stručnih radova. Iako se njihova vjerodostojnost može lako provjeriti, novi problem koji se javlja jest nestanak tih izvora. Zbog sve bržeg zastarijevanja tematike ili ne održavanja stranica na kojima se mogu naći dotični izvori (bilo zbog financijskog ili nekog drugog razloga), životni vijek istih se skraćuje radi čega ih se ne može uvijek naći na istom mjestu, ili ih se uopće ne može naći. Glavni cilj je istaknuti trend 'nestajanja' mrežnih izvora koji se koriste za pisanje znanstvenih radova, te kakve to posljedice može imati. Istraživanje je provedeno nad podacima prikupljenim za izradu rada Mrežni izvori u diplomskim radovima informacijskih znanosti na Filozofskom fakultetu u Zagrebu (Bauer, Kirin, Turković) kada je primijećeno da neki mrežni izvori ili ne postoje ili su premješteni. Želi se pokazati koliko je mrežnih izvora nestalo ili premješteno u periodu od pristupa mrežnim izvorima do kada je napravljeno prvo istraživanje, te koliko ih je još nestalo ili premješteno u periodu od završetka prvog istraživanja do provedbe sljedećeg istraživanja, tj. ovog rada. S obzirom da je između dva istraživanja prošlo više od godine dana, očekuje se da je određeni broj tada važećih mrežnih izvora nestao ili premješten. Proučavanjem ponašanja životnog vijeka mrežnih izvora moći će se pokazati kolikom brzinom se izmjenjuju i/ili zastarijevaju informacije, što također može pokazati koliko dugo se neki mrežni izvor može smatrati iskoristivim, te mijenja li se konkretna informacija ili samo njena lokacija. Istraživanje je uključivalo analizu mrežnih izvora kroz duži period godina, te je pretpostavljeno da je najviše nestalo onih mrežnih izvora koje su koristili studenti ranijih godina. Isto tako, prikazan je odnos životnog vijeka različitih mrežnih izvora, gdje se očekuje da je vijek mrežnih stranica daleko kraći od vijeka članaka i zapisa unutar e-baza. Uz to se napravila analiza s obzirom na mrežna mjesta, autore i izdavače nevažećih izvora. Vezano za mrežna mjesta, očekuju se većinski nestanak onih mrežnih izvora koji se mogu naći u sklopu komercijalnih, neznanstvenih, rijetko posjećivanih mrežnih mjesta. Što se tiče prisutnosti autora i izdavača, za očekivati je da većina nevažećih izvora nema zastupljene ni autore ni izdavače, što bi bio jedan od razloga njihovog kratkog vijeka. Što se tiče produljenja životnog vijeka mrežnih izvora, u radu su predstavljene neke od ideja drugih autora, poput standardizacije termina, potpunijeg arhiviranja, te kvalitetnog dizajna. S druge strane, ovo istraživanje bi uz tendenciju nestajanja mrežnih izvora, trebalo dati odgovore na pitanja gdje se to najčešće javlja, te s obzirom na kriterije koje zadovoljavaju važeći mrežni izvori, što bi se trebalo učiniti da se osigura njihova dugotrajnost, jer ona rezultira većom razinom vjerodostojnosti korištenih izvora, što izravno djeluje na vjerodostojnost rada u kojem su korišteni i jača kredibilitet znanstvene zajednice, što pak vodi pozitivnoj percepciji javnosti i vrednovanju od strane Vlade, te potencijalnih partnera kulturnog, znanstvenog i obrazovnog sektora.

An increasing amount of students and scholars use web references as a prime source in their papers. The main concern is that those references have a short lifespan. In retrospect to that, the aim of this article is to show how many of the web references, gathered in a corpus of 1947 web references within 362 graduate papers at the Department of Information Sciences at the Faculty of Humanities and Social Sciences in Zagreb in the period from 2003 to 2010, are no longer active, i.e. accessible. It was also substantial to know how many of the web references have disappeared in the period between two researches. The main hypothesis is that there has been an increase in the number of inactive web references within the aforementioned corpus. It is expected that the highest number of inactive web references is present in the graduate papers that have been written during the first half of the analysed period, i.e. more time has passed since those references have been accessed. As well as the availability of the web references, certain bibliographic data were also analysed, where it is predicted that according to the type of web references web pages are more likely to be inactive than any other types such as scientific articles, etc. In response to that, it is expected that the web sites of inactive web references are non-scientific in snature. The analysis of the presence of authors and publishers among inactive web references is also included

Canine progressive retinal atrophies (PRA) are genetically heterogeneous diseases characterized by retinal degeneration and subsequent blindness. PRAs are untreatable and affect multiple dog breeds, significantly impacting welfare. Three out of seven Giant Schnauzer (GS) littermates presented with PRA around four years of age. We sought to identify the causal variant to improve our understanding of the aetiology of this form of PRA and to enable development of a DNA test. Whole genome sequencing of two PRA-affected full-siblings and both unaffected parents was performed. Variants were filtered based on those segregating appropriately for an autosomal recessive disorder and predicted to be deleterious. Successive filtering against 568 canine genomes identified a single nucleotide variant in the gene encoding NECAP endocytosis associated 1 (): c.544G>A (p.Gly182Arg). Five thousand one hundred and thirty canids of 175 breeds, 10 cross-breeds and 3 wolves were genotyped for c.544G>A. Only the three PRA-affected GS were homozygous (allele frequency in GS, excluding proband family = 0.015). In addition, we identified heterozygotes belonging to Spitz and Dachshund varieties, demonstrating c.544G>A segregates in other breeds of German origin. This study, in parallel with the known retinal expression and role of in clathrin mediated endocytosis (CME) in synapses, presents as a novel candidate gene for retinal degeneration in dogs and other species.

Canine progressive retinal atrophies (PRA) are genetically heterogeneous diseases characterized by retinal degeneration and subsequent blindness. PRAs are untreatable and affect multiple dog breeds, significantly impacting welfare. Three out of seven Giant Schnauzer (GS) littermates presented with PRA around four years of age. We sought to identify the causal variant to improve our understanding of the aetiology of this form of PRA and to enable development of a DNA test. Whole genome sequencing of two PRA-affected full-siblings and both unaffected parents was performed. Variants were filtered based on those segregating appropriately for an autosomal recessive disorder and predicted to be deleterious. Successive filtering against 568 canine genomes identified a single nucleotide variant in the gene encoding NECAP endocytosis associated 1 (NECAP1): c.544G>A (p.Gly182Arg). Five thousand one hundred and thirty canids of 175 breeds, 10 cross-breeds and 3 wolves were genotyped for c.544G>A. Only the three PRA-affected GS were homozygous (allele frequency in GS, excluding proband family = 0.015). In addition, we identified heterozygotes belonging to Spitz and Dachshund varieties, demonstrating c.544G>A segregates in other breeds of German origin. This study, in parallel with the known retinal expression and role of NECAP1 in clathrin mediated endocytosis (CME) in synapses, presents NECAP1 as a novel candidate gene for retinal degeneration in dogs and other species.

Naked foal syndrome (NFS) is a genodermatosis in the Akhal-Teke horse breed. We provide the first scientific description of this phenotype. Affected horses had almost no hair and showed a mild ichthyosis. So far, all known NFS affected horses died between a few weeks and 3 years of age. It is not clear whether a specific pathology caused the premature deaths. NFS is inherited as a monogenic autosomal recessive trait. We mapped the disease causing genetic variant to two segments on chromosomes 7 and 27 in the equine genome. Whole genome sequencing of two affected horses, two obligate carriers, and 75 control horses from other breeds revealed a single non-synonymous genetic variant on the chromosome 7 segment that was perfectly associated with NFS. The affected horses were homozygous for ST14:c.388G>T, a nonsense variant that truncates more than 80% of the open reading frame of the ST14 gene (p.Glu130*). The variant leads to partial nonsense mediated decay of the mutant transcript. Genetic variants in the ST14 gene are responsible for autosomal recessive congenital ichthyosis 11 in humans. Thus, the identified equine ST14:c.388G>T variant is an excellent candidate causative variant for NFS and the affected horses represent a large animal model for a known human genodermatosis. Our findings will enable genetic testing to avoid the non-intentional breeding of NFS affected foals.

BACKGROUND The feline counterpart of human inflammatory linear verrucous epidermal nevus (ILVEN) has been described; however, the possible underlying developmental defect has not been investigated. OBJECTIVE To report a case of multiple ILVEN-like lesions in a cat with a genetic variant in the NSDHL gene. ANIMALS A 2-year-old, female, domestic short hair cat with a history of multiple alopecic, verrucous, hyperpigmented and erythematous skin lesions, following Blaschko's lines on the head, the limbs, the trunk and paw pads. METHODS AND RESULTS According to the clinical and histopathological findings, a diagnosis of multiple ILVEN-like lesions was made. Genetic investigation revealed a heterozygous missense variant in the X-chromosomal NSDHL gene predicted to lead to a loss-of-function of the NSDHL protein. CONCLUSIONS AND CLINICAL IMPORTANCE To the best of the authors' knowledge, this is the first case of feline ILVEN-like lesions in which a genetic cause has been proposed. Future studies to establish a causal relationship between NSDHL variants and skin lesions might lead to pathogenesis-directed treatments.