Abstract Neuroaxonal dystrophy (NAD) is a group of inherited neurodegenerative disorders characterized primarily by the presence of spheroids (swollen axons) throughout the central nervous system. In humans, NAD is heterogeneous, both clinically and genetically. NAD has also been described to naturally occur in large animal models, such as dogs. A newly recognized disorder in Miniature American Shepherd dogs (MAS), consisting of a slowly progressive neurodegenerative syndrome, was diagnosed as NAD via histopathology. Affected dogs were typically young adults and displayed an abnormal gait characterized by pelvic limb weakness and ataxia. A combined GWAS and autozygosity mapping approach, together with whole-genome sequencing, identified the underlying genetic cause as a 1-bp deletion in RNF170 (ring finger protein 170), which perfectly segregates in an autosomal recessive pattern. This deletion is predicted to create a frameshift (XM_038559916.1:c.367delG) and early truncation of the RNF170 protein (XP_038415844.1:(p.Ala123Glnfs*11). A significant LOD score of 9.70 in an extended pedigree confirms the linkage of the deletion variant with the canine phenotype. Several RNF170 variants have been identified in human patients with analogous clinical syndromes, indicating that this novel MAS NAD serves as an excellent large animal model for equivalent human diseases, particularly since affected dogs demonstrate a relatively long lifespan, which represents an opportunity for therapeutic trials. The age of this canine RNF170 variant is estimated at approximately 30 years, before the reproductive isolation of the MAS breed. This carries implications for the standard Australian Shepherd, the breed from which MAS were developed.

BACKGROUND:Autosomal recessive ichthyosis leads to structural or biochemical changes that impair skin barrier function. HYPOTHESIS/OBJECTIVES:To assess (1) the phenotype and genotype in a litter of Jack Russell Terriers with autosomal recessive congenital ichthyosis (ARCI), and (2) the defective skin barrier and determine if a topical ceramide can modulate the barrier. ANIMALS:A healthy dam and litter of Jack Russell Terrier puppies (healthy male, affected male and female), one affected adult Jack Russell Terrier and one unrelated healthy Jack Russell Terrier. MATERIALS AND METHODS:A severe cornification defect was identified via examination of affected puppies. As the phenotype worsened, the affected puppies received a topical application of ω-0-acylceramide for 10 days. Before humane euthanasia, the skin barrier was evaluated via transepidermal water loss (TEWL), corneometry and pH in affected dogs. Genomic testing was performed, and skin samples were analysed by light and electron microscopy. RESULTS:Affected puppies were homozygous for the 1980 bp LINE-1 insertion in the TGM1 (transglutaminase 1) gene; the unaffected littermate and the dam were heterozygous carriers. ARCI puppies were underweight and had a severe hyperkeratotic phenotype that impaired mobility. TEWL was markedly higher in affected dogs. The cutaneous pH of affected puppies was higher than the normal littermate. Treatment of the skin with ω-0-acylceramide normalised the pH to match the littermate and decreased TEWL. Electron microscopy revealed marked attenuation of the cornified envelope. CONCLUSIONS AND CLINICAL RELEVANCE:Dogs with TGM1-deficient ARCI have an impaired skin barrier. Topical therapy can partially repair the barrier defect.

Autosomal recessive ichthyosis leads to structural or biochemical changes that impair skin barrier function. To assess (1) the phenotype and genotype in a litter of Jack Russell Terriers with autosomal recessive congenital ichthyosis (ARCI), and (2) the defective skin barrier and determine if a topical ceramide can modulate the barrier. A healthy dam and litter of Jack Russell Terrier puppies (healthy male, affected male and female), one affected adult Jack Russell Terrier and one unrelated healthy Jack Russell Terrier. A severe cornification defect was identified via examination of affected puppies. As the phenotype worsened, the affected puppies received a topical application of ω-0-acylceramide for 10 days. Before humane euthanasia, the skin barrier was evaluated via transepidermal water loss (TEWL), corneometry and pH in affected dogs. Genomic testing was performed, and skin samples were analysed by light and electron microscopy. Affected puppies were homozygous for the 1980 bp LINE-1 insertion in the TGM1 (transglutaminase 1) gene; the unaffected littermate and the dam were heterozygous carriers. ARCI puppies were underweight and had a severe hyperkeratotic phenotype that impaired mobility. TEWL was markedly higher in affected dogs. The cutaneous pH of affected puppies was higher than the normal littermate. Treatment of the skin with ω-0-acylceramide normalised the pH to match the littermate and decreased TEWL. Electron microscopy revealed marked attenuation of the cornified envelope. Dogs with TGM1-deficient ARCI have an impaired skin barrier. Topical therapy can partially repair the barrier defect.

A 13-year-old male domestic short-hair cat presented for evaluation of labored breathing, hyporexia, and lethargy. Pertinent initial diagnostics yielded leukocytosis, characterized by neutrophilia and monocytosis. Numerous small, round, magenta granules were observed within all neutrophils in Wright-Giemsa-stained blood films on the day of presentation and the day thereafter. No other neutrophil morphologic abnormalities were present, making cytoplasmic toxicity highly unlikely. Hyperadrenocorticism was diagnosed based on the lack of suppression in a low-dose dexamethasone suppression test, and without other diagnostics, the cat was discharged on trilostane therapy. Neutrophil granules did not stain with Alcian blue pH 1.0, periodic acid-Schiff (PAS), PAS and Alcian blue pH 2.5, and toluidine blue. Electron microscopy identified no differences in the morphology of the secretory granules or other neutrophil features. Metabolic screening tests of the cat's urine did not identify a genetic metabolic disorder. However, serum α- and β -hexosaminidase (HexA and HexB) activities were 4.3% and 0% of normal controls, respectively, which is supportive of GM2-gangliosidosis, that is, Sandhoff disorder. However, the historical, clinical, and electron microscopy findings did not provide evidence to confirm this genetic defect. To the author's knowledge, this is the first case of magenta-staining granules within neutrophils in a breed other than a Birman, Siamese, or Himalayan.

Mucopolysaccharidosis (MPS) I is a lysosomal storage disorder characterized by deficient alpha-l-iduronidase activity, leading to abnormal accumulation of glycosaminoglycans (GAGs) in cells and tissues. Synovial joint disease is prevalent and significantly reduces patient quality of life. There is a strong clinical need for improved treatment approaches that specifically target joint tissues; however, their development is hampered by poor understanding of underlying disease pathophysiology, including how pathological changes to component tissues contribute to overall joint dysfunction. Ligaments and tendons, in particular, have received very little attention, despite the critical roles of these tissues in joint stability and biomechanical function. The goal of this study was to leverage the naturally canine model to undertake functional and structural assessments of the anterior (cranial) cruciate ligament (CCL) and Achilles tendon in MPS I. Tissues were obtained postmortem from 12-month-old MPS I and control dogs and tested to failure in uniaxial tension. Both CCLs and Achilles tendons from MPS I animals exhibited significantly lower stiffness and failure properties compared to those from healthy controls. Histological examination revealed multiple pathological abnormalities, including collagen fiber disorganization, increased cellularity and vascularity, and elevated GAG content in both tissues. Clinically, animals exhibited mobility deficits, including abnormal gait, which was associated with hyperextensibility of the stifle and hock joints. These findings demonstrate that pathological changes to both ligaments and tendons contribute to abnormal joint function in MPS I, and suggest that effective clinical management of joint disease in patients should incorporate treatments targeting these tissues.

Background: Neuroaxonal dystrophy (NAD) is a group of inherited neurodegenerative disorders characterized primarily by the presence of spheroids (swollen axons) throughout the central nervous system. In humans, NAD is heterogeneous, both clinically and genetically. NAD has also been described to naturally occur in large animal models, such as dogs. A newly recognized disorder in Miniature American Shepherd dogs (MAS), consisting of a slowly progressive neurodegenerative syndrome, was diagnosed as NAD via histopathology. Objectives: To describe the clinical and pathological phenotype together with the identification of the underlying genetic cause. Methods: Clinical and postmortem evaluations, together with a genome-wide association study and autozygosity mapping approach, followed by whole-genome sequencing. Results: Affected dogs were typically young adults and displayed an abnormal gait characterized by pelvic limb weakness and ataxia. The underlying genetic cause was identified as a 1-bp (base pair) deletion in RNF170 encoding ring finger protein 170, which perfectly segregates in an autosomal recessive pattern. This deletion is predicted to create a frameshift (XM_038559916.1:c.367delG) and early truncation of the RNF170 protein (XP_038415844.1:(p.Ala123Glnfs*11)). The age of this canine RNF170 variant was estimated at similar to 30 years, before the reproductive isolation of the MAS breed. Conclusions: RNF170 variants were previously identified in human patients with autosomal recessive spastic paraplegia-85 (SPG85); this clinical phenotype shows similarities to the dogs described herein. We therefore propose that this novel MAS NAD could serve as an excellent large animal model for equivalent human diseases, particularly since affected dogs demonstrate a relatively long lifespan, which represents an opportunity for therapeutic trials. (c) 2024 The Author(s). Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.

In humans, the prevalence of congenital microphthalmia is estimated to be 0.2-3.0 for every 10,000 individuals, with nonocular involvement reported in ∼80% of cases. Inherited eye diseases have been widely and descriptively characterized in dogs, and canine models of ocular diseases have played an essential role in unraveling the pathophysiology and development of new therapies. A naturally occurring canine model of a syndromic disorder characterized by microphthalmia was discovered in the Portuguese water dog. As nonocular findings included tooth enamel malformations, stunted growth, anemia, and thrombocytopenia, we hence termed this disorder Canine Congenital Microphthalmos with Hematopoietic Defects. Genome-wide association study and homozygosity mapping detected a 2 Mb candidate region on canine chromosome 4. Whole-genome sequencing and mapping against the Canfam4 reference revealed a Short interspersed element insertion in exon 2 of the DNAJC1 gene (g.74,274,883ins[T70]TGCTGCTTGGATT). Subsequent real-time PCR-based mass genotyping of a larger Portuguese water dog population found that the homozygous mutant genotype was perfectly associated with the Canine Congenital Microphthalmos with Hematopoietic Defects phenotype. Biallelic variants in DNAJC21 are mostly found to be associated with bone marrow failure syndrome type 3, with a phenotype that has a certain degree of overlap with Fanconi anemia, dyskeratosis congenita, Shwachman-Diamond syndrome, Diamond-Blackfan anemia, and reports of individuals showing thrombocytopenia, microdontia, and microphthalmia. We, therefore, propose Canine Congenital Microphthalmos with Hematopoietic Defects as a naturally occurring model for DNAJC21-associated syndromes.

Objectives This study aimed to determine thyroid hormone concentrations in a cohort of healthy kittens due to the paucity of information in the literature, and the potential for congenital hypothyroidism (CH) to contribute to fading kitten syndrome (FKS). Methods The serum concentrations of total thyroxine (TT4), free thyroxine (fT4), total triiodothyronine (TT3), free triiodothyronine (fT3) and thyroid-stimulating hormone (TSH) were measured in 19 healthy kittens aged 2–16 weeks. Results Mean TT4, fT4, TT3 and fT3 concentrations significantly differed across age groups. Mean TT4 and fT4 concentrations peaked at 6 and 5 weeks of age, respectively. The TT4 concentration exceeded the adult cat reference interval (ACRI) in 54% (32/59) of samples at week 6, with the highest TT4 concentration being 7.1 µg/dl (91 nmol/l). Mean TT3 and fT3 concentrations also peaked at 6 weeks of age. Mean TT3 concentration started below the ACRI until 4 weeks of age, after which it remained within the ACRI. The mean fT3 concentration was within the ACRI at all ages. The mean TSH concentration did not differ across age groups and remained within the ACRI in nearly 100% of samples. Conclusions and relevance Peak TT4, fT4, TT3 and fT3 concentrations in healthy kittens at 5 and 6 weeks of age are likely due to changes in the maternal transfer of thyroid hormones after weaning and organ system development. Knowing healthy neonatal and pediatric thyroid hormone concentrations in a cohort of kittens might help a veterinarian interpret thyroid hormone levels when trying to rule out CH in a kitten with FKS.

Mucopolysaccharidosis (MPS) VII is an inherited lysosomal storage disorder characterized by deficient activity of the enzyme b-glucuronidase. Skeletal abnormalities are common in patients and result in diminished quality of life. Enzyme replacement therapy (ERT) for MPS VII using recombinant human b-glucuronidase (vestronidase alfa) was recently approved for use in patients; however, to date there have been no studies evaluating therapeutic efficacy in a large animal model of MPS VII. The objective of this study was to establish the effects of intravenous ERT, administered at either the stan-dard clinical dose (4 mg/kg) or a high dose (20 mg/kg), on skel-etal disease progression in MPS VII using the naturally occur-ring canine model. Untreated MPS VII animals exhibited progressive synovial joint and vertebral bone disease and were no longer ambulatory by age 6 months. Standard-dose ERT-treated animals exhibited modest attenuation of joint dis-ease, but by age 6 months were no longer ambulatory. High -dose ERT-treated animals exhibited marked attenuation of joint disease, and all were still ambulatory by age 6 months. Vertebral bone disease was recalcitrant to ERT irrespective of dose. Overall, our findings indicate that ERT administered at higher doses results in significantly improved skeletal disease outcomes in MPS VII dogs.

Mucopolysaccharidosis I is a lysosomal storage disorder characterized by deficient alpha-L-iduronidase activity, leading to abnormal accumulation of glycosaminoglycans in cells and tissues. Synovial joint disease is prevalent and significantly reduces patient quality of life. There is a critical need for improved understanding of joint disease pathophysiology in MPS I, including specific biomarkers to predict and monitor joint disease progression, and response to treatment. The objective of this study was to leverage the naturally-occurring MPS I canine model and undertake an unbiased proteomic screen to identify systemic biomarkers predictive of local joint disease in MPS I. Synovial fluid and serum samples were collected from MPS I and healthy dogs at 12 months-of-age, and protein abundance characterized using liquid chromatography tandem mass spectrometry. Stifle joints were evaluated postmortem using magnetic resonance imaging (MRI) and histology. Proteomics identified 40 proteins for which abundance was significantly correlated between serum and synovial fluid, including markers of inflammatory joint disease and lysosomal dysfunction. Elevated expression of three biomarker candidates, matrix metalloproteinase 19, inter-alpha-trypsin inhibitor heavy-chain 3 and alpha-1-microglobulin, was confirmed in MPS I cartilage, and serum abundance of these molecules was found to correlate with MRI and histological degenerative grades. The candidate biomarkers identified have the potential to improve patient care by facilitating minimally-invasive, specific assessment of joint disease progression and response to therapeutic intervention.

German Shorthaired Pointer (GSHP) dogs with a UNC93B1 gene mutation develop exfoliative cutaneous lupus erythematosus (ECLE) and kidney disease resembling lupus nephritis in humans. The objective of this study was to characterize the kidney disease by light microscopy, immunofluorescence, and electron microscopy in a population of GSHP dogs with ECLE. Medical records were reviewed, and light microscopy of kidneys from 7 GSHP dogs with a previous histologic diagnosis of ECLE was performed. Immunofluorescence of fresh-frozen kidney from 1 dog and transmission electron microscopy of kidney from that dog and 2 additional dogs were performed. Five of 7 dogs had proteinuria diagnosed by urinalysis or urine protein-to-creatinine ratio. Two of 7 dogs were intermittently hypoalbuminemic, and none were azotemic. Histologic findings included early (2 dogs) to late (5 dogs) membranous glomerulonephropathy characterized by mild-to-severe glomerular capillary loop thickening and tubular proteinosis. In all 7 cases, trichrome staining revealed red granular immune deposits on the subepithelial surface of the glomerular basement membrane. Immunofluorescence revealed strong granular labeling for immunoglobulins and complement protein C3. Electron microscopy demonstrated subepithelial electron-dense immune deposits encircled by the remodeled glomerular basement membrane. These findings are diagnostic of immune-complex membranous glomerulonephropathy and are similar to class V lupus in humans. This cohort of GSHP dogs with ECLE developed immune-complex membranous glomerulonephropathy, which we hypothesize is a manifestation of systemic lupus erythematosus. GSHP dogs with ECLE should undergo clinical evaluation of renal function for early identification and treatment.

Ichthyoses are hereditary skin disorders characterized by the formation of scales and defects in the outermost layer of the epidermis. In dogs, at least six different breed-specific ichthyoses including a relatively common PNPLA1-related autosomal recessive ichthyosis in Golden Retrievers are known. In this study, we investigated 14 Golden Retrievers with scales that were not homozygous for the mutant PNPLA1 allele suggesting a genetically distinct new form of ichthyosis. Histopathological examinations showed lamellar, orthokeratotic hyperkeratosis, and mildly hyperplastic epidermis that led to the diagnosis of a nonepidermolytic ichthyosis. Combined linkage and homozygosity mapping in 14 cases and 30 nonaffected family members delimited a critical interval of similar to 12.7 Mb on chromosome 23. Whole-genome sequencing of an affected dog revealed a single protein-changing variant within this region that was not present in 795 control genomes. The identified variant is a 14 bp deletion in the ABHD5 gene (c.1006_1019del), leading to a frameshift and altering the last 14 codons p.(Asp336Serfs*6). The genotypes at this variant showed perfect cosegregation with the ichthyosis phenotype in a large family comprising 14 cases and 72 controls. ABHD5 encodes an acyltransferase required for lipid metabolism. In humans, variants in ABHD5 cause Chanarin-Dorfman syndrome, a neutral lipid storage disease with ichthyosis. Our data in dogs together with the knowledge on the effects of ABHD5 variants in humans strongly suggest ABHD5:c.1006_1019del as candidate causative genetic variant for a new canine form of ichthyosis, which we propose to designate as Golden Retriever ichthyosis type 2 (ICH2).

Mucopolysaccharidosis (MPS) I is a lysosomal storage disease characterized by deficient activity of the enzyme alpha-L-iduronidase, leading to abnormal accumulation of heparan and dermatan sulfate glycosaminoglycans in cells and tissues. Patients commonly exhibit progressive skeletal abnormalities, in part due to failures of endochondral ossification during postnatal growth. Previously, using the naturally-occurring canine model, we showed that bone and cartilage cells in MPS I exhibit elevated lysosomal storage from an early age and that animals subsequently exhibit significantly diminished vertebral trabecular bone formation. Wnts are critical regulators of endochondral ossification that depend on glycosaminoglycans for signaling. The objective of this study was to examine whether lithium, a glycogen synthase kinase-3 inhibitor and stimulator of Wnt/betacatenin signaling, administered during postnatal growth could attenuate progression of vertebral trabecular bone disease in MPS I. MPS I dogs were treated orally with therapeutic levels of lithium carbonate from 14 days to 6 months-of-age. Untreated heterozygous and MPS I dogs served as controls. Serum was collected at 3 and 6 months for assessment of bone turnover markers. At the study end point, thoracic vertebrae were excised and assessed using microcomputed tomography and histology. Lithium-treated animals exhibited significantly improved trabecular spacing, number and connectivity density, and serum bone-specific alkaline phosphatase levels compared to untreated animals. Growth plates from lithium-treated animals exhibited increased numbers of hypertrophic chondrocytes relative to both untreated MPS I and heterozygous animals. These findings suggest that bone and cartilage cells in MPS I are still capable of responding to exogenous osteogenic signals even in the presence of significant lysosomal storage, and that targeted osteogenic therapies may represent a promising approach for attenuating bone disease progression in MPS I.

Epidermolysis bullosa (EB) is a group of blistering disorders that includes several subtypes, classified according to their level of cleavage. Typical clinical signs are blisters and erosions resulting from minimal trauma. The disease has been described in many mammalian species and pathogenic variants in at least 18 different genes have been identified. In the present study, we investigated a Cardigan Welsh Corgi with congenital clinical signs consistent with epidermolysis bullosa. The puppy had blisters and erosions on the paw pads, and the oral mucosa. Histologic examination demonstrated the typical clefting between the dermis and epidermis and confirmed the clinical suspicion. We obtained whole genome sequencing data from the affected puppy and searched for variants in candidate genes known to cause EB. This revealed a heterozygous missense variant, KRT5:p.(E476K), affecting the highly conserved KLLEGE motif of keratin 5. The mutant allele in the affected puppy arose owing to a de novo mutation event as it was absent from both unaffected parents. Knowledge of the functional impact of KRT5 variants in other species together with the demonstration of the de novo mutation event establishes KRT5:p.(E476K) as causative variant for the observed EBS.

Practical relevance: When compared with the number of individuals that make up a dog breed, the population within a given cat breed is very small. Therefore, to maintain a breed standard, a certain degree of inbreeding is necessary. However, when inbreeding reaches a certain threshold, it can lead to decreased fertility, which manifests as failure to conceive, smaller litter size, increased neonatal illness and neonatal mortality. Breeders should be encouraged to keep comprehensive records on breeding outcomes, including number of kittens born, neonatal vitality, daily kitten weights and kitten health at weaning. Commercially available DNA panels are available to inform and facilitate excellent breeding choices and can estimate the coefficient of inbreeding. Clinicians should include a review of the degree of inbreeding in the work-up for any cattery or cat colony experiencing decreased fertility. Aim: The objective of this article is to provide clinicians, especially those working with cat breeders, with an easy-to-understand guide to genetics and to demonstrate how inbreeding influences fertility and neonatal survival. Equipment and technical skills: Medical records and the pedigree of the cats in question are required to investigate cases of infertility that may be related to inbreeding. A DNA analysis kit that measures genetic diversity and health parameters can also be helpful; those that have been developed by geneticists and veterinarians at universities are preferable, as they include access to highly skilled genetic counselors and researchers who are open to working up newly discovered genetic diseases. Evidence base: The material provided is based on current literature and the author's own studies examining outcomes in a closed cattery.