Focus on Hereditary Diseases & Genetic Predispositions: Clinical & Laboratory Diagnostic Approach to Hereditary Diseases. Part I (104) and II (105)
ACVIM 2008
Urs Giger, DACVIM, ECVIM (Internal Medicine)
Philadelphia, PA, USA

Because of the increased awareness of breeders, pet owners, and veterinarians of genetic defects and the improved diagnostic abilities in clinical practice, the number of reported hereditary diseases in small animals is rapidly growing. At present, >900 and >200 hereditary diseases in dogs and cats, respectively, have been adequately documented, and every year over a dozen new defects are being reported. For the small animal practitioner, it can be a daunting, nearly impossible task to remember all these diseases and be aware of the many novel tests and their appropriate management and control. The recent advances in canine clinical genetics will be covered and illustrated with clinical case examples.

Many of the characteristic breed traits and common and rare diseases seen in veterinary practice have a heritable basis. Recent exciting advances in our current knowledge of the completed dog and feline genome sequences offer the opportunity to clinicians to use these emerging tools in clinical practice and have a positive impact on the health of dogs as well as cats and in particular the diagnosis, management, and control of hereditary diseases. Indeed, hereditary diseases and genetic predisposition to diseases have taken on an increasing important role in veterinary medicine as many infectious diseases, nutritional deficiencies, and intoxications have been controlled. Genetic screening tests permit the identification of animals at risk for many of the single-gene hereditary diseases prior to the development of clinical signs, mitigating the suffering of dogs and cats carrying 2 mutant alleles for autosomal recessive traits. This advancement has far-reaching benefits for promoting canine and feline health permitting the elimination of deleterious gene defects, while preserving desirable traits in a breed. Identification of complex trait markers, such as those responsible for temperament and trainability, will likely prove extremely valuable to guide- and service-dog organizations, but also the common pet dog and cats. For the first time, the investigation and identification of polygenic diseases is a realistic proposition. Lastly, collaborative comparative veterinary-human studies will serve to accelerate the rate of discovery and the extent to which both human and veterinary medicine accrues benefits from gene-based research.

Genetic diseases are caused by chromosomal alterations or gene mutations. Disease-causing mutations are heritable changes in the sequence of genomic DNA that alter the expression, structure, and function of the coded protein. The genotype refers to the animal's genetic makeup, reflected by its DNA sequence, whereas the phenotype relates to the clinical manifestation of specific gene(s) and environment, or both. The molecular genetic defect is now known for > 50 hereditary disorders in dogs and >15 in cats, some of them are listed below. These molecular genetic changes include point mutations, deletions, and insertions in the DNA sequence that result in a missense or nonsense sequence with an altered codon sequence. For approximately half of the disorders suspected to be of a genetic nature the mode of inheritance remains however unknown. The canine genome (7.5x) has been sequenced during the past years, which has and will continue to greatly facilitate the characterization of molecular bases of simple and complex hereditary diseases in dogs and cats. The high quality canine DNA genome sequence of a Boxer makes up a total of ~20,000 genes. Similarly the recently published low density (1.9 x) feline genome sequence has discovered 19,000 genes. Based upon the vast variety of polymorphic markers (microsatellites and Single Nucleotide Polymorphisms [SNPs]) spread throughout the genome new genes can be discovered and associated with disease traits. Moreover, certain SNP panels can be used to group breeds of dogs and cats. Recently this technology has been applied to develop a mixed breed test (Veterinary-based Mars Veterinary Wisdom Panel Mx mixed breed analysis based upon 130 AKC breeds or MetaMorphix Canine Heritage Breed Test based upon 38 breeds).

The dog has 76 autosomes (38 pairs) and 2 sex chromosomes (78XX or 78XY), and the cat's karyotype is 38XX or 38XY. The pattern of inheritance depends mainly on two factors: 1) whether the mutation is located on an autosome (autosomal) or on the X-chromosome (X-linked), and 2) whether the phenotype, the observable expression of a genotype as a disease trait, is dominant, i.e., expressed when only one chromosome of a pair carries the mutation, or recessive, i.e., expressed when both chromosomes of a pair carry the mutation. Thus, it is the phenotype rather than the mutant gene or protein that is dominant or recessive. Whereas in humans most diseases are dominantly inherited, recessive traits are favored by the common inbreeding practices in small animals. In addition, complex genetic traits where more than one gene alteration (polygenic) and environmental factors play are role in the expression and severity of a disease. Many susceptibilities to disease, such as inflammatory, immune-mediated, and neoplastic diseases as well as drug reactions, are considered to be transmitted by a complex trait.

Beyond physical examination and imaging tools, genetic, metabolic, and other laboratory techniques are used to diagnose hereditary disorders in companion animals. Most genetic defects cause clinical signs early in life. The term congenital only implies that the disease is present at birth, however, and does not necessarily mean it is inherited. A common presentation is failure to thrive. They are poor doers, often fade (hence the term fading puppy or kitten syndrome), and finally die. Failure-to-thrive should not be confused with growth retardation. In addition to these relatively unspecific clinical signs, some defects may cause specific clinical manifestations. Easy to recognize are malformations that involve any part of the skeleton and lead to disproportionate dwarfism, gait abnormalities, and/or facial dysmorphia. A large number of hereditary eye diseases have been described in dogs, some of which are not recognized until adulthood. Neuromuscular signs may vary from exercise intolerance to ataxia and seizures. Defects of many other internal organs are associated with unspecific clinical signs.

Diagnostic tests are generally required to further support a genetic disorder in a diseased animal. Radiology and other imaging techniques may reveal skeletal malformations or cardiac anomalies, and an ophthalmologic examination may further define an inherited eye disease, although some are not recognized until several years of age. Routine tests such as complete blood cell count, chemistry screen, and urinalysis may suggest some specific hematological or metabolic disorders or rule out many acquired disorders. Furthermore, clinical function studies may more clearly define a gastrointestinal, liver, kidney, or endocrine problem. Histopathology and/or electron microscopy of a tissue biopsy from an affected animal or from the necropsy of a littermate or relative may give the first clue to a genetic defect.

A few laboratories provide special diagnostic tests that allow a specific diagnosis of an inborn error of metabolism. Inborn errors of metabolism include all biochemical disorders due to a genetically determined, specific defect in the structure and/or function of a protein molecule. Disorders of intermediary metabolism typically produce a metabolic block in a biochemical pathway leading to product deficiency, accumulation of substrates, and production of substances via alternative pathways. The most useful specimen to detect biochemical derangements is urine because abnormal metabolites in the blood will be filtered through the glomeruli, but fail to be reabsorbed, as no specific renal transport system exist for most abnormal metabolites. Once the failing system has been identified, the defect can be determined at the protein level. Homozygously affected animals have very low protein activity and/or quantities (0-10%). These tests may also be used to detect carriers (heterozygotes), who typically have intermediate quantities at the protein level (30-70%), but no clinical signs. Unfortunately, protein assays require submission of appropriate tissue or fluid under special conditions to specialized laboratories along with a control sample, and are labor intensive.

The molecular defect has been identified for >60 canine and >15 feline hereditary diseases, and thus DNA screening tests have been developed. These tests are mutation or DNA marker specific and can therefore only be used in animals suspected to have the exact same gene defect. Small animals within the same or a closely related breed will likely have the same disease-causing mutation for a particular disease. However, dogs and cats as well as unrelated breeds of a species with the same disorder will likely have different mutations. On the other hand a few mutations have been found in a couple of breeds or may be widespread within the canine population. For many inherited disorders, the defective gene remains unknown; however, for a few, a polymorphic DNA marker that is linked to the mutant allele has been discovered. At present, mutation-specific and linkage tests are available only for single gene defects in small animals; however, complex genetic traits may also soon be approached by these methods as they are for humans. Many predispositions such as inflammatory, immune-mediated, malignant disorders have a genetic basis. While many more single gene defects are being studied from clinical signs to the molecular defect, current investigations are shifting toward complex genetic traits. The many breed predispositions for various complex genetic traits are particularly attractive to further define their molecular bases.

DNA tests have several advantages over other biochemical tests. The test results are independent of the age of the animals, thus, the tests can be performed at birth or at least long before an animal is placed in a new home as well as before clinical signs become apparent. DNA is very stable and only the smallest quantities are needed; hence, there are no special shipping requirements as long as one follows the specific mailing instructions for biological products. DNA can be extracted from any nucleated cells, e.g., blood, buccal mucosa (using cheek swabs), hair follicle, semen, and even formalinized tissue. For instance, blood can be sent in an EDTA tube or a drop of blood can be applied to a special filter paper; buccal swabs can be obtained with special cytobrushes--the cheek cells and not the saliva is needed and swabs need to be completely dried. The DNA segment of interest, which is surrounding the mutation, is amplified with appropriate DNA primers utilizing the polymerase chain reaction (PCR). The mutant and/or normal alleles are identified by DNA fragment size or base pair differences. These tests are generally simple, robust, and accurate as long as appropriate techniques and controls are used. Furthermore, they can be used not only for the detection of affected animals, but also for carriers from birth on, and thus are extremely valuable to select breeding animals that will not cause disease or further spread the disease-causing allele. If an animal with all the desirable qualities is found to be a carrier, it could be safely bred to a clear animal (homozygous normal), as this would not result in any affected offspring. However, all offspring should be tested and only clear animals should be used in the next generation.

At present, the therapeutic options in the treatment of hereditary diseases are limited and ethical principles need to be carefully considered. Many hereditary diseases are progressive with currently only palliative therapy available, and thus lead to the early demise of a diseased animal or euthanasia. Surgical interventions may correct some malformations including some orthopedic and eye problems as well as hepatic shunts, but such animals should be altered to prevent them from being used for breeding. In a few cases a deficient protein, cofactor, substrate, or metabolite can be supplemented to correct the defect. For instance, vitamin B12 deficiency in cachectic and lethargic giant schnauzers and Border collies with an ileal receptor defect can be helped by monthly cobalamin injections. Pancreatic enzyme supplementation and daily insulin injections are used to manage animals with exocrine or endocrine pancreatic insufficiency, respectively. Fresh frozen plasma is administered in the treatment of hereditary coagulopathies and von Willebrand disease whenever animals excessively bleed. Other enzyme and protein replacements are also experimentally attempted. For instance, recombinant coagulation factors such as human recombinant factor VIIa has been successfully used for factor VII deficiency in Beagles and has also been tried as a bypassing agent in other coagulopathies and von Willebrand disease.

Although kidney transplants have been established in clinical practice for chronic renal failure in cats, they have not been applied in animals with hereditary (juvenile) renal disorders. Several hereditary disorders of hematopoietic cells have been experimentally corrected by bone marrow transplantation, e.g., pyruvate and phosphofructokinase deficiency, cyclic hematopoiesis, and interleukin-2 (IL-2) receptor defects. Furthermore, bone marrow transplantation is being attempted to deliver functional cells or active proteins to other tissues including liver, bone, and brain, e.g., in lysosomal storage disease. Finally, gene therapy, the integration of a functional gene into the patient's own defective cells, will likely be clinically feasible in the twenty-first century. Experiments in rodent models have provided encouraging results. However, effective gene therapy has proven more difficult in larger mammals, and the technology needs to be further improved to achieve persistent and regulated gene expression in larger mammals including humans, dogs and cats. One of the first and most promising canine gene therapy experiments has been the correction of mucopolysaccharidosis type VII in neonatal puppies with a retroviral vector carrying the beta-glucuronidase gene; these treated animals remain ambulatory, whereas affected become tetraparetic by a few months. Other examples include feline mannosidosis, hemophilia, and severe x-linked combined immunodeficiency. Such treatments are being developed for humans and once the technique is established it may also be applied in companion animals in the near future. Much more important than the treatment of hereditary disorders is the control of these traits in breeding programs.

In conclusion, it is most exciting to learn about many recent advances for many hereditary disorders and genetic predispositions in small animal practice, be it for the diagnostic approach to a hereditary disease, the understanding of its pathophysiology, or its control. In addition to the clinician's responsibility to suspect a genetic disease and to appropriately diagnose it with modern specific techniques, clinicians must become involved in the control of these disorders in the breeders' kennels or catteries. Practitioners thus can make an important contribution toward controlling the further spread of mutant genes and reducing future suffering of animals.

Examples of hereditary diseases in dogs with known mutations (excluding eye diseases).

Disease

Breeds affected

Alport Syndrome

Samoyed, mixed breed dog

Centronuclear Myopathy (CNM)

Labrador retriever

Ceroid lipofuscinosis

Border Collie, English Setter, Dachshund

Cobalamin Malabsorption

Giant Schnauzer, Australian Shepherd

Complement 3 Deficiency

Brittany Spaniel

Congenital Hypothyroidism with Goiter

Toy Fox Terrier, Rat terrier

Copper Toxicosis

Bedlington Terrier

Cyclic Neutropenia/Gray Collie Syndrome

Collie, Rough and Smooth

Cystinuria Type I

Labrador, Newfoundland

X-linked Ectodermal Dysplasia

German Shepherd

Epidermolysis Bullosa

Golden Retriever, German Shorthaired Pointer

Epilepsy (Lafora Type)

Miniature Wirehaired Dachshund

Factor VII Deficiency

Beagle, Alaskan Klee Kai, Scottish Deerhounds

Factor XI Deficiency

Kerry Blue Terrier

Fucosidosis

English Springer Spaniel

Glanzmann Thrombasthenia

Great Pyrenees, Otterhound

Globoid Cell Leukodystrophy

West Highland White Terrier, Cairn Terrier

GM1 Gangliosidosis

Portuguese Water Dog, Shiba Inu, Alaskan Husky

Glycogenosis IA

Maltese Terrier

Glycogenosis IIIA

Curly-coated Retriever

Hemophilia A

Mixed Breeds

Hemophilia B

Airedale Terrier, Bull Terrier, Lhasa Apso, Labrador Retriever

L-2 Hydroxyglutaric Aciduria

Staffordshire Bull Terrier, West Highland White Terrier

Leukocyte Adhesion Deficiency (CLAD)

Irish Setter, Irish Red and White Setter

Malignant Hyperthermia

Greyhound

MDR1 Drug Sensitivity (Ivermectin)

Collie related breeds

Mucopolysaccharidosis (MPS) I

Plott Hound

Mucopolysaccharidosis (MPS) IIIA

Dachshund, New Zealand Huntadog

Mucopolysaccharidosis (MPS) IIIB

Schipperke

Mucopolysaccharidosis (MPS) VI

Miniature Pinschers, Miniature Schnauzer

Mucopolysaccharidosis (MPS) VII

German Shepherd, mixed-breed Dog

Muscular Dystrophy (Duchenne)

Golden Retriever, Rottweiler, German Shorthair Pointer, others

Myotonia Congenita

Miniature Schnauzer, Australian Cattle dog

Mitochondrial Myopathy

Clumber and Sussex Spaniel

Narcolepsy

Dachshund, Doberman Pinscher, Labrador Retriever

Osteogenesis Imperfecta, Dominant

Beagle, Golden Retriever

Phosphofructokinase Deficiency

American Cocker Spaniel, English Springer Spaniel, Mixed Breed Dog

Primary Hyperparathyroidism

Kerry Blue Terrier

Pyruvate Kinase Deficiency (PK)

Basenji, Beagle, West Highland White Terrier

Renal Adenocarcinoma and Nodular Hyperplasia

German Shepherd

Renal Dysplasia, Juvenile

Lhasa Apso, Shih Tzu, Soft Coated Wheaton Terrier

X-Linked Severe Combined Immunodeficiency (SCID)

Basset Hound, Cardigan Welsh Corgi

Autosomal Recessive SCID

Jack Russell Terrier

Shaking Puppy

English Springer Spaniel

Von Willebrand's Disease (vWD) Type I

Doberman Pinscher, Drentsche Patrijschond, Kerry Blue Terrier, Manchester Terrier, Papillion, Pembroke Welsh Corgi, Poodles, Shetland Sheepdog, West Highland White

Von Willebrand's Disease (vWD) Type II

German Shorthaired and Wirehaired Pointer

Von Willebrand's Disease (vWD)

Scottish Terrier, Kooiker

Examples of hereditary diseases in cats with known mutations.

Disease

Breeds affected

Acute Intermittent Porphyria

Siamese

Alpha Mannosidase

Persian, Domestic Shorthair

Dominant Cardiomyopathy

Maine Coon

Gangliosidosis GM1

Siamese, Korat

Gangliosidosis GM2

Korat

Glycogenosis Type IV

Norwegian Forest

Hemophilia B

Domestic Shorthair

Hypertriglyceremia/-Chylomicronemia

Domestic Shorthair

Mucolipidosis II

Domestic Shorthair

Mucopolysaccharidosis I

Domestic Shorthair

Mucopolysaccharidosis VI

Siamese

Mucopolysaccharidosis VII

Domestic Shorthair

Muscular Dystrophy, x-chromosomal

Domestic Shorthair

Niemann-Pick Disease C

Domestic Shorthair

Polycystic Kidney Disease

Persian, and others

Pyruvate Kinase Deficiency

Abyssinian, Somali, Domestic Shorthair

Retinitis Pigmentosa

Abyssinian

Spinal Muscular Atrophy

Maine Coon

References

1.  Giger U. Clinical Genetics, in Textbook of Veterinary Internal Medicine, Eds. Ettinger S.J. and Feldman E.C., Philadelphia, PA, Saunders, 264-268, 2005.

2.  Giger U. Erbkrankheiten in Praktikum der Hundeklinik. Ed. Suter PF, Parey Verlag pp 205-215, 2000.

3.  Giger U, Haskins ME. Erbkrankheiten in Katzenkrankheiten, Eds. Horzinek M. Lutz H., Enke Verlag, 589-602, 2005.

4.  Haskins ME, Giger U. Lysosomal Storage Diseases, in Clinical Biochemistry of Domestic Animals, 6th eds. In press, 2008.

5.  Ostrander E, Lindbald-Toh K, Giger U. The Dog and its Genome. Cold Spring Harbor Press, Cambridge, 2006.

6.  Pontius JU, et al. (2007) Initial sequence and comparative analysis of the cat genome. Genome Res 17, 1675-1689.

7.  Sewell AC, Haskins ME, Giger U. Inherited Metabolic Disease in Companion Animals: Searching for Nature's Mistakes. Vet J. 174:252-9, 2007.

8.  Werner P, Haskins ME, Giger U. Comparative and Medical Genetics, in Clinical Biochemistry of Domestic Animals, 6th eds. In press, 2008.

Websites

1.  Inherited Diseases in Dogs @ http://www.vet.cam.ac.uk/idid and Mendelian Inheritance in Animals http://www.angis.org.au/Databases/BIRX/omia/

2.  Soon a web site from the WSAVA Hereditary Disease Committee on VIN Associates

Speaker Information
(click the speaker's name to view other papers and abstracts submitted by this speaker)

Urs Giger, DACVIM, DECVIM (Internal Medicine)
University of Pennsylvania
Philadelphia, PA


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