Testing for Equine Genetic Disorders. What, Why & How?

Testing
for
Equine Genetic Disorders
What, Why and How?
©2016 Arreton Arabians

Genetic Disorders
• are the result of inheritable abnormalities (mutations) in a
horse’s genes
• we will be discussing disorders that result from the mutation
of single genes (defined sections of chromosomes)
 Chromosomes occur in pairs (except in sperm and ova),
and are strings of genes (thus, genes occur in pairs too).
 Genes are made up of DNA
 At conception, the embryo receives one half of each pair
of chromosomes from the sire and the other half of the
pair from the dam

Sperm + egg = zygote embryo
Homo means ‘same’ Hetero means ‘different’
Homozygous means that the zygote received two identical copies of a gene (one from
the sperm and the other from the egg). They may be two mutant copies, or two normal
copies of the gene.
Heterozygous means that the zygote received one normal copy of the gene from one
parent and one mutant copy from the other parent.
Some definitions

Genetic Disorders can be
 autosomal and recessive (autosomal means that the
gene responsible is present on any chromosome other than
the X/Y chromosomes)
 autosomal and dominant
 sex-linked (the gene is located on the X/Y
chromosomes)
I don’t know of any sex-linked disorders that have been identified
in the equine, but a human example is Haemophilia; which results
in failure of the blood to clot.

Autosomal Dominant Disorders
• A dominant mutation does precisely what we should expect: it dominates
• Even if a horse inherits one normal copy of the gene, and only ONEmutant copy, the
mutation will dominate and the horse will be affected by the condition.
• Some conditions are termed ‘incomplete dominant’. In these cases, a homozygous
(having two mutant copies of the gene) horse will be more seriously affected by the
condition than a heterozygous horse (who has one mutant copy and one normal copy of
the gene).
• Examples of dominant genetic conditions of horses are
 MH (Malignant Hyperthermia)
 HYPP (Hyperkalemic Periodic Paralysis)

Autosomal Recessive Disorders
• For a horse to be affected by an autosomal recessive genetic disorder it has to inherit
TWO mutant copies of the gene responsible (one copy from each parent).
• Heterozygous horses (who have one normal copy and one mutant copy) are unaffected
by the condition because any effect of the mutation is suppressed by the normal copy of
the gene. These horses are also called ‘Carriers’ – they carry the mutation, but it is not
expressed.
• Carrier horses are physiologically normal and exhibit no symptoms of the disorder.
• Examples of recessive autosomal conditions are
 Lavender Foal Syndrome (LFS)
 Hereditary Equine Regional Dermal Asthenia (HERDA)

Inheritance of Autosomal Recessive Conditions (1)
• Is independent of the gender of the progeny.
• An affected parent (carrying two mutant copies of the gene) will
always pass on the mutation to every one of its progeny.
• Every time a carrier (one normal copy of the gene and one mutant
copy) is mated there is a 50% probability that the mutation will be
inherited by the resultant foal.

• If a carrier is put to a mate that is clear (has two normal copies of the gene), the
progeny will always be either clear or carriers; never affected. No problem.
• If a carrier is bred to a carrier there is
 a 25% chance that the foal will be affected by the disorder (inherit two mutant
copies of the gene). BIG problem.
 a 50% chance that the foal will be a carrier (inherit a mutant copy of the gene from
one parent, and a normal copy from the other).
 a 25% chance that the foal will be clear of the disorder (inherit a normal copy of
the gene from each parent).
• It is important to understand that every mating of carrier to carrier has a
25% chance of resulting in an affected foal. If one affected foal is bred, it
does not mean that the next three foals from the same mating will be
unaffected or carriers.

N = normal copy of the gene n = mutated copy of the gene
NN = unaffected horse Nn/nN = carrier nn = affected horse
Carrier Sire Carrier Dam
Nn Nn
Possible progeny NN Nn nN nn
Potential progeny from each mating:
1 in 4 (25%) chance of a homozygous normal foal
2 in 4 (50%) chance of a carrier
1 in 4 (25%) chance of an affected foal

Genetic conditions that can be tested for include:
Condition Abbreviation Dominant/Recessive Breed first identified in
Malignant Hyperthermia MH Autosomal Dominant Quarter Horse
Polysaccharide Storage Myopathy Type 1 PSSM1 Autosomal Dominant Several
Foal Immunodeficiency Syndrome FIS Autosomal Recessive Fell Pony
Overo Lethal White Syndrome OLWS Autosomal Recessive Several
Cerebellar Abiotrophy CA Autosomal Recessive Arabian
Severe Combined Immune Deficiency SCID Autosomal Recessive Arabian
Lavender Foal Syndrome LFS Autosomal Recessive Arabian
Hereditary Equine Regional Dermal Asthenia HERDA Autosomal Recessive Quarter Horse
Hyperkalemic Periodic Paralysis HYPP Autosomal Dominant Quarter Horse
Glycogen Branching Enzyme Deficiency GBED Autosomal Recessive Several
Hoof Wall Separation Disease HWSD Autosomal Recessive Connemara
Junctional Epidermolysis Bullosa JEB1/JEB2 Autosomal Recessive Draught Horses/Saddlebreds
Friesian Dwarfism Autosomal Recessive Friesian
Friesian Hydrocephalus Autosomal Recessive Friesian
Warmblood Fragile Foal Syndrome WFFS Autosomal Recessive Warmblood

Why test for recessive conditions?
• Are you willing to take the risk of breeding an affected foal?
• If you test your breeding stock and avoid mating carrier to carrier you will never run
that risk.
• Don’t be complacent: the mutant genes are not restricted to the breeds in which they
were first identified. CA was, for example, found in three breeds tested as part of a
study: a Welsh Pony, two Trakheners, and 3 Bashkir Curlies
Why would you not test?????????

Who to test, and for what?
• Genes do not discriminate. There is the same probability of a mutation being
inherited by the foal, no matter the breed of the other parent.
• The mutations responsible for LFS, CA and SCID were first identified in Arabians but
are not restricted to that breed. Arabians have been used to develop and strengthen
many breeds, all of which may carry the mutations.
• Test your own breeding stock, and when considering a potential mate for a carrier,
check the extended pedigree and consider the possibility that they may be a carrier
of the same condition. If the owner of the prospective mate refuses to disclose test
results or to permit testing, you might consider looking elsewhere.
• Carrying a recessive disorder does not affect a horse’s conformation, temperament or
performance, and should not be considered shameful, nor should the horse be
considered unworthy, BUT they should be bred responsibly.

How to test
• Contact an accredited testing laboratory that offers the tests you require.
• Usually they will require about thirty hairs pulled from the mane or tail,
which must have the hair root attached. If you pull the hair straight out,
the root will almost always come with it. Cut hair is of no use for testing.
Hair shaft (contains very little, if any, DNA)
Hair root (lots of cells containing DNA)

What happens to the hair when it gets to the testing lab? (1)
• DNA is extracted from the hair root, not from the hair itself,
which is why it is important to ensure that you pull hairs cleanly
and ensure that the roots are attached.
• The extracted DNA contains every gene that the horse from which
the hair was pulled carries.
• The extracted DNA is mixed with a variety of chemicals, at least
two of which are uniquely matched to the gene being
investigated, and a dye that makes the gene fluoresce.

• During repeated cycles of heating and cooling, the gene of interest is
copied multiple times if it is present in the sample, to generate enough
material to be analysed.
A thermal cycler. The samples are in the tiny tubes
• Tests are designed so that both the normal and mutated gene of interest
will be copied if they are present.
• Tests are also designed so that the size of the copies of the normal gene is
different to the size of the copies of the mutant gene.

• Next, the mixture, containing copies of the normal and defective genes (if present), is
applied to a thin slab of gel, and an electric current is applied
An electrophoresis tank. The samples are added to small
wells at the top of the gel and an electric current applied.
• The copied genes migrate through the gel under the influence of the electric current,
separating out according to size.
• The smaller the size of the copied gene, the faster and further it moves through the gel
• The DNA fluoresces and is visible under UV light

Interpreting Test Results
Stylised image of a DNA gel exposed to UV light
normal gene
Defective or mutant gene
Unaffected: Carrier: Affected:
Normal gene only Normal and mutant gene Mutant gene only

Genetic disorders could be excluded from the gene pool if breeders
excluded carriers from their breeding programmes BUT
• Carriers of autosomal recessive conditions are physiologically
normal and only identified as being ‘flawed’ if they are subjected
to genetic testing (or breed affected progeny).
• Many fine examples of true to type and form mares and stallions
would then be excluded from breeding programmes, and already
limited gene pools would become even more so.
• As more and more genes are identified it is possible that we will
eventually find a genetic ‘flaw’ in every horse tested.
Why Breed From Carriers of Recessive Disorders?
Waiting for the perfect horse?

Responsible Breeding of Carriers of Autosomal Recessive Conditions:
• Subject breeding stock to genetic testing for all relevant conditions.
• Do not ever breed a carrier to another carrier of the same genetic disorder.
• Do not assume that conditions such as SCID and LFS are restricted to
Arabians. The original mutations occurred many generations ago and
Arabians have been used to improve several breeds since then.
• Owners of carriers should ensure that any prospective breeding partners
are tested and found clear, no matter what breed they may be.

Testing for Equine Genetic Disorders. What, Why & How?

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