"Louisiana Catahoula - A Truly Merle Breed" - by Dr. Helena Synková

The following is Helena's translated article that was published in several European canine magazines in 2011 - 2012 and the Catahoula Club EU's website.

A topic of the merle gene presence, its traits, manifestation, and heredity is widely discussed among breeders and owners of various dog breeds which are known to carry it. Therefore we had not been expecting any big revelation, when the testing among Louisiana Catahoula in our club started. There were some doubts, of course, namely when unexpected "merle" phenotypes were born out of solid colored parents. This was also the main reason for us to start looking for an explanation.

A typical merle coat color, i.e. coat with patches of diluted pigment irregularly spread over body and intermingled with undiluted melanin, is known in several breeds, such as Australian Sphepard, Collie, Border Collie, Cardigan Welsh Corgi, Shetland Sheepdog, Dachshund, etc. However, individuals with such a merle coat coloring are not a majority in those breeds. It is probably due to breeding regulations valid for those breeds that prevent combinations of both merle colored parents. The main reason for an existence of such breeding rules is a fact that offsprings of predominantly white color (called double merle) are born from such merle parents and they are known to suffer often from a variety of serious health defects.

However, there is one breed that is famous for its merle colors and leopard patterns - Louisiana Catahoula (LC). It seems that for many breeders from country of its origin, USA, solid colored LC are not so valuable. Thus, a combination of both merle carrying parents is done on purpose to obtain merle carrying offsprings of leopard colors in spite of possible health defects. No surprice then that most dogs of this breed are the merle gene carriers and true solid colored (not carrying the merle gene) are rather rare. Nevertheless, it seems that the breed itself is not endangered by this fact seriously and the occurrence of the merle caused defects is not higher than in other merle carrying breeds.

In 2005, when our first article on this topic was written and published here, the merle gene question and its heredity seemed to be similar to other breeds, so we did not expect any difference in LC. However, the merle gene itself has not been identified yet, so there was no possibility to prove it by any other means than breeding itself. Frankly, there were not so many litters born here at that time and their "results" fitted quite well to our expectation.

The Merle gene

In 2006, an article of Clark et al. (PNAS 103 (5): 1376-81, 2006) was published and the merle gene was finally identified. It opened an era of possible genetical testing of entire dog populations from this point of view. Clark et al. demonstrated that the gene in question is SILV (Silver locus, Pmel17), which is known to be a pigment dilution gene and its significant expression is localized exclusively to the skin and eye. They proved that genetical modifications (mutations) in SILV gene affect its normal function and it results in a coat pattern where a diluted pigment is mixed with patches of normal, undiluted melanin. Non-modified SILV gene is marked as m, while modified versions are marked as M, i.e. the merle gene. Modifications in SILV gene are caused by an insertion of a mobile gene element, called retrotransposon, which can be of various length, as we know now. It seems that the length of this insertion affects the way how the merle gene manifests itself in dog phenotype.

There were two modification of SILV gene described in the original article of Clark et al. The normal M gene includes the insertion of the length of 459-462 bp. It causes the typical merle/leopard coat pattern in dogs, heterozygous for it (mM), and the typical double merle phenotype with predominant white coloring in dogs homozygous (MM).

Another modification referred by Clark et al. contains a shorter insertion of the length of 420-425 bp. This modification of SILV gene is called cryptic (phantom) merle, Mc, and it seemes to have no apparent effect on coat coloring. Both heterozygous (mMc) and homozygous (McMc) individuals appear to be of solid coat color, similarly as homozygous dogs with non-modified SILV gene, i.e. mm. As far as we know now, there is only one way to differentiate them from non-merle gene carriers - by genetical testing.

Both above mentioned modifications of SILV gene were described in the original article of Clark et al. They were recognized in all breeds, studied then. The merle gene seems to be inherited in an autosomal, incompletely dominant way. Offspring gets one allel of this gene from both parents and resulting genotypes are divided according to Mendel′s rules.

Catahoula Merle Gene

Our wide-spread testing for the merle gene presence in LC proved another modification of SILV gene, i.e. the insertion of medium length containing 443 -449 bp, which seems to be specific for LC breed. Scientists from Biofocus laboratories, who carried genetical analyses, have not found such a modification elsewhere until now. For time being, I called it Ma (a = atypical). The length of this insertion is medium to M and Mc and it affects phenotype less than M, but more than Mc. Dogs heterozygous for Ma (i.e. mMa) are born with solid colored phenotype, which seems to change more or less to the merle pattern during ageing. Dogs homozygous for this gene, i.e. MaMa, show the merle pattern since a birth, but it is usually all in diluted colors such as blue or yellow. There is no or only a little white usually, even if they are double merle, too. A combined genotype for M and Ma, i.e. MaM, seems to be a typical for patchwork phenotype, which is specific by larger areas of undiluted, leopard and white colors. Heterozygous dogs for McMa might be born as solid colored, showing most probably some merle pattern later as above mentioned (mMa).

The presence of LC specific modification of SILV gene, i.e. Ma, is probably a point missing until now. It explains well those examples, where the merle phenotypes were born from solid colored parents. It has been noticed that leopard colors are changing during dog life, some are bleaching, some get darker and sometimes finally leopard color is unrecognizable later in a life. It seems that M a Ma modifications are responsible for it. Thus we experience dogs heterozygous for M, i.e. mM, that are born with the typical merle coat pattern and their colors seem to be becoming darker and patches of diluted colors less apparent with increasing age. Contrary to it, dogs heterozygous for Ma, i.e. mMa, are born as solid colored and merle patterns is more and more visible with their ageing. It is at least our present experience. However, the presence of Mc cannot be detected according to the coat color, but only by genetical tests. We do not know exactly, if Mc affects other merle genes (M, Ma) by its presence, because it seems to be very rare in the population of Czech born LC.

Determination of the merle gene

The assay for the merle gene, which is used in Biofocus laboratories, originates from the methods of Clark et al., i.e. PCR analyses of gene products using specific primers. The analyse itself starts with an isolation of DNA from samples taken from dogs. Although it is possible to use buccal swabs from facial cheeks, our practical experience proved that blood is more reliable starting material for DNA isolation. Buccal swab quality is more dependent on skills of collector and probably also duration of storage and transport before analysis. That′s why it is not possible to obtain always a sufficient amount of DNA for a precise detection of modified SILV gene. Our experience shows that if blood samples were used as starting material, the identification of the merle gene was more accurate.

Benefits for breeding programmes

What are benefits which can the knowledge of the merle genotype bring for further LC breeding? This is a question that comes up to many breeders surely. Testing for the merle gene started among members of our club (CC EU), however, other LC owners sent samples for testing on their own, too. Many of them did not know, why it should be done, and they just followed the suite. Some wanted their dogs to be proved not being the merle gene or double merle carriers. From this poin of view, most of them was very disappointed from results they received.

We would like to thank to all, who joined our club effort, cooperated with us and published results of their dogs, and namely to those, who gave us an open access to details of their analysis. It helped us to elucide or suggest more generally rules of the merle gene heredity and traits. We tested various LC within our club with various family relationships, therefore we can finally make some general conclusions out of it.

It is not possible now to test the first imported LC, i.e. the male, Cats Cradles Car, and the female, So. Sloughs Kathy, that were used as the foundation stock here. We cannot even test all of their offsprings at this moment. However, we can estimate their genotypes from phenotypes of their offsprings and their known combinations. Some of them have been tested already, so there is some clue to determine it better than only according to their known phenotype. Now it is possible to evaluate litters born in the past, namely those born from parents of known (i.e. tested) genotypes. Such an analysis enables to prove supposed heredity and it gives us for the future a possibility to predict more precisely genotypes and phenotypes of expected pups. This has been a sphere of a fiction and wishful thinking until now for this breed. It is important namely because it is better to avoid a production of double merles with predominantly white color present in their coat, if it is possible for other reasons. Now we know that it is a case of homozygous for MM and also MaM, where white coloring is an effect of the merle gene and not probably of another white color coding gene, such as e.g. piebald gene. Most health defected pups are born with those above mentioned genotypes.

Our results also show that even in such a "merle gene grown through" breed, Czech LC population contains probably higher percetage of solid, mm, non-merle dogs than it is usual in other countries. It happened because the foundation stock was of mm (or mMc) and mM genotype and the first generation born here was at least half of mm, solid colored LC. However, further imported LC were mostly of MM or MaM genotypes. This fact means that the merle gene carriers probably exceeded already non-merle, mm dogs. Thus, also majority of solid colored LC is probably of mMa genotype in our present population of LC.

As a byproduct, the knowledge of the merle genotype of one or both parents enables also to prove an accidental breeding, when pups paternity is in question. This is valid namely if sires in question are of different genotypes for the merle gene and phenotypes of pups are impossible to arise only from one of them.

Our Catahoula Club EU is surely going to go on in testing and we surely support also other owners and breeders to do so. Frankly, it makes no sense to keep results of testing for themselves and not to publish it, particularly if only reason is that someone does not like the verdict on the merle gene presence of his/her dog. Moreover, there are not so many possibilities for non-tested or kept secret parents if you know the genotypes of their pups.

We want our breeders to be well informed, therefore we have just started to include the merle gene information of tested parents into pedigree information of their offsprings beside other important tests such as status of hip dysplasia, etc.

Written by RNDr. Helena Synková