A nonsense variant in the KRT14 gene in a domestic shorthair cat with epidermolysis bullosa simplex.

Epidermolysis bullosa simplex (EBS) is a hereditary blistering disease affecting the skin and mucous membranes. It has been reported in humans, cattle, buffaloes and dogs, but so far not in cats. In humans, EBS is most frequently caused by variants in the KRT5 or KRT14 genes. Here, we report a case of feline epidermolysis bullosa simplex and describe the causative genetic variant. An 11-month-old male domestic shorthair cat presented with a history of sloughed paw pads and ulcerations in the oral cavity and inner aspect of the pinnae, starting a few weeks after birth. Clinical and histopathological findings suggested a congenital blistering disease with a split formation within the basal cell layer of the epidermis and oral mucous epithelium. The genetic investigation revealed a homozygous nonsense variant in the KRT14 gene (c.979C>T, p.Gln327*). Immunohistochemistry showed a complete absence of keratin 14 staining in all epithelia present in the biopsy. To the best of our knowledge, this is the first report of feline EBS, and the first report of a spontaneous pathogenic KRT14 variant in a non-human species. The homozygous genotype in the affected cat suggests an autosomal recessive mode of inheritance.

A missense variant in the NSDHL gene in a Chihuahua with a congenital cornification disorder resembling inflammatory linear verrucous epidermal nevi.

Congenital hemidysplasia with ichthyosiform nevus and limb defects syndrome in humans is a genodermatosis characterized by inflammatory linear verrucous epidermal nevi (ILVEN), often showing a striking lateralization pattern. It is caused by variants in the NSDHL gene encoding a 3ß-hydroxysteroid dehydrogenase involved in the cholesterol biosynthesis pathway. In the present study, we investigated a female Chihuahua, which showed clinical and histological signs of ILVEN. We performed a candidate gene analysis in the affected animal. This analysis revealed a single missense variant in the NSDHL gene in the affected dog (XM_014111859.2:c.700G>A). The variant is predicted to cause a non-conservative amino acid change from glycine to arginine, XP_013967334.1:p.(Gly234Arg). The mutant allele was absent from WGS data of 594 genetically diverse dogs and eight wolves. Sanger sequencing confirmed that the variant was heterozygous in the affected dog and absent from 22 control Chihuahuas. Based on the knowledge about the functional impact of NSDHL variants in dogs and other species, c.700G>A is probably pathogenic and a convincing candidate causative variant for the observed skin lesions in the affected Chihuahua.

A second KRT71 allele in curly coated dogs.

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 151 Trp 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.

A frameshift variant in the EDA gene in Dachshunds with X-linked hypohidrotic ectodermal dysplasia.

X-linked hypohidrotic ectodermal dysplasia (XLHED) is a genetic disease characterized by hypoplasia or absence of hair, teeth and sweat glands. The EDA gene, located on the X chromosome, encodes the type II transmembrane protein ectodysplasin A. Variants in the EDA gene can lead to XLHED in humans, mice, cattle and dogs. In the present study, we investigated a litter of Dachshund puppies, of which four male puppies showed clinical signs of XLHED. We performed a candidate gene analysis in one affected puppy and several non-affected relatives. This analysis revealed a single base-pair deletion in the coding sequence of the EDA gene in the affected puppy (NM_001014770.2:c.842delT). The deletion is predicted to cause a frameshift, NP_001014770.1:p.(Leu281HisfsTer22), leading to a premature stop codon which truncates more than one quarter of the EDA protein. Sanger sequencing results confirmed that this variant was inherited from the dam. Based on knowledge about the functional impact of EDA variants in dogs and other species, c.842delT is a convincing candidate causative variant for the observed XLHED in the male puppies.

A splice site variant in the SUV39H2 gene in Greyhounds with nasal parakeratosis.

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.

Wattles in goats are associated with the FMN1/GREM1 region on chromosome 10.

The presence of congenital appendages (wattles) on the throat of goats is supposed to be under genetic control with a dominant mode of inheritance. Wattles contain a cartilaginous core covered with normal skin resembling early stages of extremities. To map the dominant caprine wattles (W) locus, we collected samples of 174 goats with wattles and 167 goats without wattles from nine different Swiss goat breeds. The samples were genotyped with the 53k goat SNP chip for a subsequent genome-wide association study. We obtained a single strong association signal on chromosome 10 in a region containing functional candidate genes for limb development and outgrowth. We sequenced the whole genomes of an informative family trio containing an offspring without wattles and its heterozygous parents with wattles. In the associated goat chromosome 10 region, a total of 1055 SNPs and short indels perfectly co-segregate with the W allele. None of the variants were perfectly associated with the phenotype after analyzing the genome sequences of eight additional goats. We speculate that the causative mutation is located in one of the numerous gaps in the current version of the goat reference sequence and/or represents a larger structural variant which influences the expression of the FMN1 and/or GREM1 genes. Also, we cannot rule out possible genetic or allelic heterogeneity. Our genetic findings support earlier assumptions that wattles are rudimentary developed extremities.

EcPV2 DNA in equine papillomas and in situ and invasive squamous cell carcinomas supports papillomavirus etiology.

Equine penile papillomas, in situ carcinomas, and invasive carcinomas are hypothesized to belong to a continuum of papillomavirus-induced diseases. The former ones clinically present as small grey papules, while the latter 2 lesions are more hyperplasic or alternatively ulcerated. To test the hypothesis that these lesions are papillomavirus-induced, samples of 24 horses with characteristic clinical and histologic findings of penile papillomas or in situ or invasive squamous cell carcinomas were collected. As controls, 11 horses with various lesions--namely, Balanoposthitis (6 cases), melanoma (3 cases), follicular cyst (1 case), and amyloidosis (1 case)--were included. DNA was extracted and polymerase chain reaction applied to amplify papillomavirus DNA. The respective primers were designed to amplify DNA of the recently discovered equine papillomavirus EcPV2. All tested papilloma and squamous cell carcinoma samples were found to contain DNA of either of 2 previously published EcPV2 variants. Among the other samples 6 of 11 were found to contain EcPV2 DNA. To further support the findings and to determine where the papillomavirus DNA was located within the lesions, an in situ hybridization for the detection of EcPV2 DNA was established. The samples tested by this technique were found to clearly contain papillomavirus nucleic acid concentrated in the nucleus of the koilocytes. The findings of this study support previous data and the hypothesis that papillomaviruses induce the described penile lesions in horses.

[Juvenile sterile granulomatous dermatitis and lymphadenitis in the dog]. / Juvenile sterile granulomatöse Dermatitis und Lymphadenitis beim Hund.

Juvenile sterile granulomatous dermatitis and lymphadenitis is a rare immune-mediated skin disease in young dogs. History, signalment, diagnostics, treatment, and outcome in 10 dogs are described. The age ranged from 8 - 36 weeks. The lymph nodes were enlarged in all dogs, especially the mandibular and prescapular lymph nodes. Systemic signs including fever were present in 8 dogs. Seven dogs suffered from blepharitis and painful edema of the muzzle with hemorrhagic discharge, pustules and papules. Cytology of pustules and lymph node aspirates revealed a pyogranulomatous inflammation. In 7 cases the diagnosis of juvenile sterile granulomatous dermatitis and lymphadenitis was confirmed by histology. Nine dogs were treated with prednisolone (0.5 - 1.25 mg/kg BID), H2-receptor antagonists and analgetics; all dogs were treated with antibiotics. Four dogs were treated with eye ointment containing antibiotics and glucocorticoids. The prednisolone dosage was tapered over 3 - 8 weeks. One dog had a relapse.

Scleromyxedema-like syndrome with systemic involvement in a cat.

Scleromyxedema--the generalized form of lichen myxedematosus, a primary mucinosis--is a rare disease in human patients. It is characterized by dermal mucin deposits, increased numbers of fibroblasts, and variable fibrosis in the absence of thyroid disease. It is accompanied in 80% of cases by a monoclonal gammopathy. To date, scleromyxedema with systemic involvement has not been documented in domestic animals. This is the first report of a scleromyxedema-like syndrome in a cat, which had a substantial deposition of mucin in the dermis of the head and paws with a mild gammaglobulinemia of 2.25 g/dl (reference range, 1.39-2.22 g/dl). At necropsy, multiple nodules of connective tissue intermingled with mucin deposits were conspicuous on the surface of thoracic and abdominal organs. Such severe systemic accumulations of mucin have not been reported in human or veterinary medicine.

[Mucormycosis due to Mycocladus corymbiferus in a fattening pig]. / Mucormykose durch Mycocladus corymbiferus bei einem Mastschwein.

A fattening pig with enlarged head and abdominal lymph nodes was examined. An aspirate of the abscesses did not produce a conclusive diagnosis. Only an excision with subsequent histological and bacteriological examination showed the mould Mycocladus corymbiferus (syn. Absidia corymbifera) to be present. Similar abscesses should be examined as actinomycosis and leucosis are the main differential diagnoses.

Porcine ulcerative dermatitis syndrome in sows: a form of vesicular cutaneous lupus erythematosus?

Severe ulcerative lesions were observed in the skin of two sows in a herd of 540 hybrid sows. Annular to polycyclic, severe crusting dermal ulcerations were found on the abdomen and flanks; moderate lesions were also found at the base of the tail and on the perineum. The lesions were histologically characterised as cell-poor interface dermatitis and folliculitis, basal cell vacuolisation, vesicle formation at the dermal-epidermal junction and serocellular crusts. A subepidermal mild to moderate band, characterised as a mixed inflammatory infiltrate, was present. A test for antinuclear antibodies was negative; however, immunofluorescence testing revealed a linear pattern of IgG precipitation in the skin. Staphylococcus hyicus was demonstrated in the serocellular crusts of one sow. Treatment with antibiotics, topical antiseptics and corticosteroids did not improve the sows' condition. Porcine circovirus and porcine respiratory and reproductive syndrome virus were not isolated from samples taken at postmortem examination. The observed gross lesions, the absence of response to treatment and the exclusion of other skin diseases suggested that the sows were affected with porcine ulcerative dermatitis syndrome.

Feline Injection Site Sarcomas: Data from Switzerland 2009-2014.

Feline injection site sarcomas (FISS) were first described in the early 1990s. Despite extensive research, the pathogenesis of these tumours has not been elucidated conclusively. Their appearance and the marked increase in their incidence has been mainly connected to the injection of vaccines, and it is assumed that a chronic inflammatory reaction at the injection site triggers subsequent malignant transformation. The role of alum-based adjuvants has been discussed, but is controversial. The present study of the Swiss Feline Cancer Registry (SFCR) with data from 2009 to 2014 revealed a marked decrease of the incidence of fibrosarcomas compared with the previous observation period. Notably, this drop occurred after a non-adjuvanted feline leukaemia virus vaccine was introduced in Switzerland in 2007. This observation, together with the previous findings of the SFCR, further supports the notion that alum-adjuvanted vaccines are involved in the genesis of FISS and that non-adjuvanted vaccines might be safer for cats.

Evaluation of the CTSL2 gene as a candidate gene for alopecia X in Pomeranians and Keeshonden.

Alopecia X is a noninflammatory, progressive, bilateral symmetric alopecia in dogs. The disease is mainly found in Nordic breeds. The breed predisposition and a strong familial accumulation suggest a hereditary background. We analyzed the cathepsin L2 gene (CTSL2) as a candidate for alopecia X. The comparative sequencing of 14 affected and 18 control animals revealed ten polymorphisms; however, none of these polymorphisms affected the coding sequence. Haplotype analysis did not reveal an association of one particular CTSL2 haplotype with the disease phenotype; therefore, we conclude that the CTSL2 gene is probably not the causative gene for alopecia X.

Comparative quantitative assessment of GnRH- and LH-receptor mRNA expression in the urinary tract of sexually intact and spayed female dogs.

Ovariectomy interrupts the regulatory loop in the hypothalamus-pituitary-gonad axis, leading to a several-fold increase in gonadotropin levels. This rise in hormonal secretion may play a causal role in ovariectomy-related urinary incontinence. The purpose of this study was to examine the effect of ovariectomy in bitches on the expression of GnRH- and LH-receptors in the lower urinary tract, and assess the relationship between receptor expression and plasma gonadotropin concentrations. Plasma gonadotropins were measured in 37 client-owned bitches. Biopsies were harvested from the mid-ventral bladder wall in all dogs, and from nine further locations within the lower urinary tract in 17 of the 37 animals. Messenger RNA of the LH and GnRH receptors was quantified using RT-PCR with the TaqMan Universal PCR Master Mix. Gonadotropins were measured with a canine-specific FSH-immunoradiometric assay and LH-radioimmunoassay. The hierarchical mixed ANOVA model using MINITAB, Mann-Whitney U-test, unpaired means comparison and linear regressions using StatView were applied for statistical analyses. Messenger RNA for both receptors was detected in all biopsy samples. Age was negatively correlated to mRNA expression of the LH and the GnRH receptors. A relationship between the mRNA values and the plasma gonadotropin concentrations was not established. Evaluation of results within each of the biopsy locations revealed greater LH-receptor expression in the proximal second quarter of the urethra in spayed bitches than in intact bitches (P=0.0481). Increased mRNA expression of LH receptors in this location could possibly play a role in the decrease in closing pressure of the urethra following ovariectomy.

[Inherited alopecia X in Pomeranians]. / Vererbte Alopezie X beim Zwergspitz.

Four male Pomeranians that showed alopecia with an age of onset between five months and eight years were investigated. The aim of the investigation was to clarify whether the affected dogs had alopecia X and whether their symptoms might be due to a hereditary defect. The four affected dogs showed hairless patches at the root of the tail, at the back, at the limbs from the thigh to the tarsus and at the abdomen. Within the hairless patches some islets with sparse hair were present. In hairless patches the skin was dark pigmented. Besides the alopecia and hyperpigmentation no other symptoms were found according to anamnestic and clinical examination. History, clinical examinations, laboratory diagnostics, and histopathology of skin biopsies allowed the diagnosis of alopecia X in three affected male dogs. The last one of the affected dogs additionally had slightly reduced thyroid hormone levels. Based on identical symptoms and the close relatedness of all four animals, it was assumed that the fourth affected dog also had alopecia X. The available data possibly indicate a monogenic autosomal dominant inheritance, however a recessive inheritance can not be excluded at this time.

Congenital Cutaneous Panadnexal Papillomatous Hamartomas in a Calf.

Several cauliflower-like alopecic masses arose on the head of an otherwise healthy, full-term newborn Jersey × Belgian blue heifer, which was humanely destroyed shortly after birth due to the severity of the skin lesions. Microscopically, the masses were composed of multiple papillary projections displaying well-differentiated sebaceous glands surrounded by a moderate number of well-developed sweat glands, as well as embryonic and fully developed, but dysplastic hair follicles. Thick branching connective tissue stalks supported these adnexal components. The papillated surface, the predominance of sebaceous glands, the presence of embryonic hair follicles and the well-differentiated sweat glands were compatible with nevus sebaceous, a rare form of cutaneous hamartoma described in man, dogs, cats and cattle. However, the cauliflower-like growth pattern, the presence of supportive thick branching connective tissue stalks and the relative abundance of dysplastic hair follicles in association with nevus sebaceous has not been described in the human or veterinary literature. A diagnosis of panadnexal papillomatous hamartoma was made in this case.

Immune dysregulation in flea allergy dermatitis--a model for the immunopathogenesis of allergic dermatitis.

BACKGROUND: Flea allergy dermatitis (FAD) is a common skin disease in dogs and can be induced experimentally. It often coexists with other allergic conditions. So far no studies have investigated the quantitative production of cytokine mRNA in skin biopsies and peripheral blood mononuclear cells (PBMC) in flea allergic dogs. OBJECTIVE: The aim of our study was to improve the understanding of the immunopathogenesis of allergic dermatitis as a response to fleabites. MATERIAL AND METHODS: Allergic and non-allergic dogs were exposed to fleas. Before and after 4 days of flea exposure mRNA was isolated from biopsies and PBMC. Production of chymase, tryptase, IL-4, IL-5, IL-13, TNF-alpha and IFN-gamma mRNA was measured by real-time RT-PCR. The inflammatory infiltrate in the skin was scored semi-quantitatively. The number of eosinophils, mast cells (MC) and IgE+ cells/mm2 was evaluated to complete the picture. RESULTS: FAD was associated with a higher number of MC before flea exposure and with a significant increase of eosinophils after flea exposure as compared to non-allergic dogs. The number of IgE+ cells was higher in allergic dogs before and after flea exposure. In allergic dogs mRNA for most cytokines and proteases tested was higher before flea exposure than after flea exposure. After exposure to fleas an increased mRNA production was only observed in non-allergic dogs. In vitro stimulation with flea antigen resulted in a decreased expression of most cytokines in allergic dogs before flea exposure. In contrast, in PBMC, only increased levels of IL-4 and IL-5 mRNA were observed in allergic dogs before flea exposure. However, after flea exposure and additional stimulation with flea antigen the production of mRNA for all cytokines tested was significantly increased in allergic dogs. CONCLUSION: We demonstrated that the response in biopsies and PBMC is different and that FAD is associated with a TH2 response.

Treatment with gonadotropin-releasing hormone after first timed artificial insemination improves fertility in noncycling lactating dairy cows.

Lactating Holstein cows were assigned randomly to treatments to improve fertility after first postpartum timed artificial insemination (TAI). In Experiment 1, cows received no treatment (control; n = 9), a controlled internal drug releasing (CIDR) insert from 5 to 12 d after TAI (CIDR; n = 9), or 100 microg of GnRH 5 d after TAI (G5; n = 7). Although treatments did not affect circulating progesterone (P4) concentrations from 5 to 19 d after TAI, there was a tendency for CIDR cows to have greater P4 compared with control or G5 cows within 24 h after treatment. In 2 field trials, cows received either control (n = 223), CIDR (n = 218), or G5 (n = 227) treatments (Experiment 2), or control (n = 160), G5 (n = 159), or treatment with 100 microg of GnRH 7 d after TAI (G7; n = 163; Experiment 3). Treatment did not affect pregnancies per AI (P/AI) in Experiments 2 or 3; however, when data were combined to compare control (n = 383) and G5 (n = 386) treatments, P/AI tended to be greater for G5 (49.1%) than for control (45.8%) cows. This effect resulted from a GnRH treatment x cyclicity status interaction in which P/AI for noncycling cows receiving G5 was greater than for noncycling control cows (45.5 vs. 31.1%). In conclusion, treatment with CIDR inserts after TAI had no effect on P/AI, whereas treatment with GnRH 5 d after TAI improved P/AI for noncycling, but not for cycling cows.

Effect of interval from timed artificial insemination to initiation of resynchronization of ovulation on fertility of lactating dairy cows.

To compare 2 strategies for systematically resynchronizing ovulation, lactating Holstein cows (n = 763) at various days in milk and prior artificial insemination services were assigned randomly at timed AI (TAI) to receive the first GnRH injection of Ovsynch 26 (D26) or 33 (D33) d after TAI to resynchronize ovulation (Resynch) in cows failing to conceive. Cows in the D26 treatment received GnRH 26 d after TAI and continued Resynch only when diagnosed not pregnant by using ultrasonography 33 d after TAI, whereas D33 cows initiated Resynch only when diagnosed not pregnant 33 d after TAI. Cows were classified based on the presence or absence of a corpus luteum (CL) at the not-pregnant diagnosis, and cows without a CL received an intravaginal progesterone-releasing insert during Resynch. When analyzed as a systematic strategy, pregnancy rate per AI (PR/AI) was greater for cows assigned to the D33 than the D26 Resynch treatment (39.4 vs. 28.6%). A treatment x parity interaction was detected for PR/AI after Resynch for nonpregnant cows having a CL in which primiparous cows had a greater PR/AI than multiparous cows when Resynch was initiated 33 d after the initial TAI, and primiparous and multiparous cows when Resynch was initiated 26 d after the initial TAI. Pregnancy loss for Resynch was 6.4% between 33 and 40 d, and 2.6% between 40 and 61 d after Resynch TAI. We concluded that delaying initiation of Resynch until 33 d after TAI increased PR/AI for primiparous cows.