HCM Slowak-Czech Study from 2008-2009 Incidence of hypertrophic cardiomyopathy in the Ragdoll breed
Incidence of hypertrophic cardiomyopathy in the Ragdoll breed
Ľ. HRIB
Veterinary Clinic Jesenice, Prague
Veterinářství 2009;59:268-272.
SUMMARY
Hrib Ľ. Prevalence of hypertrophic cardiomyopathy in the Ragdoll breed.
The article deals with the observation of the incidence of hypertrophic cardiomyopathy in the Ragdoll breed in the Berberis line. So far 34 Ragdolls aged 3 months to 12 years have been genetically tested. Echocardiographic examination was preferentially performed in positive homozygous and heterozygous individuals. From the HCM distribution maps developed so far, by comparing genetic tests with echocardiographic findings, it is clear that the Berberis lineage is not a group at high risk of HCM disease.
SUMMARY
Hrib Ľ. Occurrence of hypertrophic cardiomyopathy in Ragdoll breed.
This paper deals with monitoring of hypertrophic cardiomyopathy in Ragdoll breed of Berberis line. Altogether 34 Ragdoll cats aged from 3 months to 12 years were genetically tested up to now. In preference, one positive homozygote and some heterozygote cats were echocardiographically examined. From maps with HCM spreading it was found by comparison of genetic tests with echocardiographic findings that Berberis line is not highly threatened group with HCM disease.
Introduction
Hypertrophic cardiomyopathy (HCM), is the most commonly diagnosed cardiac muscle disease in cats, in which thickening of the left ventricular wall and interventricular septum occurs in undilated heart chambers. From a genetic perspective, the predominant definition of HCM is that it is an inherited, autosomal dominant disease caused by genes encoding cardiac sarcomeric proteins.1 It has been reported that approximately 11% of the canine and feline population is affected by cardiac disease. In numbers, this translates to 8 to 10 million individuals.2 In approximately 60% of human cases of HCM, it is an autosomal dominant disease with a prevalence of 1 in 500, meaning that one mutant allele is sufficient to cause an outbreak.3 The overall prevalence in humans is approximately 0.2%, making it the most commonly inherited cardiovascular disease. It is estimated that there are approximately 20,000 human patients with HCM in the country, of which about 2000-4000 carry a higher risk of sudden death.1 The prevalence of HCM in cats varies from case to case and depends on the number of animals examined and the time period observed. Studies by Rush indicate a 37.3% prevalence of cats with HCM,4 whereas Ferasin reports 57.5%, Riesen 67.6%,5,6 and Sampedrano only 9.6% positive for HCM in his study group (of which 2 individuals in the genetic test were negative homozygotes N/N, but phenotypically positive! )26 The international breeding database PawPeds.com and The Winn Feline Foundation report that HCM researchers report between 33% and 34% (1,090 of 3,238 Maine Coon cats tested between December 2005 and February 2007) positive HCM test results in Maine Coon cats tested.11 In the Ragdoll population, Dr. Meurs' group reports a 23% prevalence of at least one defective allele of the HCM gene.12
Gene mutations
The gene implicated as causative for HCM in Ragdolls was discovered in mid-2007 by Dr. Meurs' team at the University of Washington in Pullman.7 Samples of 21 Ragdolls diagnosed with HCM from two cat families and samples of 110 control, unaffected cats were examined, including 19 Ragdolls, 6 Sphynx, 12 Maine Coons, 10 Domestic Shorthairs, 4 Pixie Bob, 4 Bengal cats, 7 Scottish Folds, 2 Persians, 4 Rex and 42 crossbreeds. The causative gene is thought to be a mutation in the gene encoding the binding protein for myosin (MYBPC3-C820T). This mutation results in the amino acid arginine, which is found at this site in many species, being replaced by tryptophan. The two amino acids have completely different physical properties and the mutation results in a lack of protein integration in the unit of the muscle fibre responsible for its contraction (the sarcomere).
In humans, there are more than 450 known mutations in 16 genes responsible for HCM at different stages and with different prognoses.8 In cats, only three mutations in the MYBPC3 gene have been identified so far. The first genetic mutation was discovered in the Maine Coon cat in 2005 (A31P).9 The second is the aforementioned mutation responsible for the Ragdoll mutation (C820T).7 At a conference in Montreal in 2007, Dr. Nyberg's team introduced a third mutation, or second mutation, in the Maine Coon cat (A74T).10
Currently, there is not enough information on the Ragdoll breed to clearly define the time of disease onset and course in homozygous (two defective alleles) and heterozygous (one defective allele) individuals.9,13,14 According to studies by Dr. Lefbom between 1997 and 1999, who followed ten Ragdolls with HCM (note that the genetic mutation for HCM in Ragdolls had not yet been discovered), it was clear that the disease manifests at a very early age - an average of 15 months (5 to 24 months).15 Dr. Meurs' research shows that homozygous cats (21 months) develop the disease on average 18 months earlier than heterozygous cats (39 months). From Dr. Meurs' study, it is also possible to infer the probability that the Ragdolls in Dr. Lefbom's study that became ill at a very early age (six cats less than one year old) were homozygous, and the individuals who became ill later may have carried one affected allele (heterozygous). Dr. Meurs' 2007 studies in Ragdolls are also consistent with published data she conducted when monitoring mutations in Maine Coons in 2005. Here, homozygous affected cats develop moderate to severe HCM disease at 4 years of age or earlier, while 3 out of 10 heterozygous cats still survive with a mild form of HCM between 8 and 12 years of age.7 The so-called Munich study is also published, which questions the relevance of the MYBPC3 mutation to practical breeding in Maine Coon cats.16 A 2009 study by Sampedrane also points out that genetically heterozygous individuals do not suffer from cardiac muscle hypertrophy and myocardial dysfunction under all circumstances.26
Berberis lineage and HCM distribution maps
As part of efforts to rehabilitate the gene pool and protect the few breeds, 34 Ragdolls from the Middle-Earth Hobby Ragdoll Breeders Club and Ragdolls from some SCHK breeders have been tested at NCHK from July 2008 to the present. DNA testing was performed on cheek swabs and examined in the German Laboklin laboratory. The sample of Ragdolls tested included animals ranging in age from 3 months to 12 years. Further genetic testing of individual Ragdoll lines is currently underway in both the Czech and Slovak Republics. In the NCHK, the testing itself was preceded by work on mapping the distribution of HCM in the European Ragdoll population from publicly available sources on the Internet and international databases. So far, two HCM distribution maps have been produced. The first map included only positively diagnosed Ragdolls based on echocardiographic examination or autopsy. The second map is a map created only on the basis of DNA testing of Ragdolls who have participated in genetic testing at the NCHK. Both of these maps go back to the founders of the breed. They therefore map a time period of 40 years. Both maps are used exclusively for HCM clinical research and are therefore not freely available on the internet.
Like the German breeders, their Czech colleagues expected clear answers from the genetic tests and a comprehensible tool for breeding practice. Already the first results of the tests were very interesting and it was obvious that this testing is only the beginning of a very long way we still have to go to unravel the mysteries of HCM. The big surprise was the test of Naomi Berberis' 12 year old Ragdolla seal colourpoint cat, which came directly from American parents Chubby Tuftytoes and Pippa Kan-Du. This cat had the N/HCM genotype. The test was repeated with the same result: heterozygote N/HCM. Naomi herself enjoyed good health throughout her life. After her death in August 2008, an autopsy was performed at the SVU in Prague with negative results with regard to HCM. Another direct descendant of these parents was also tested. Shawn, again with a result of N/HCM. At the age of almost 12 years, he is still in excellent health and after an echocardiographic examination in late January 2009, he suffers from a mild asymptomatic form of HCM (IVSd 6.1 mm and LVWd 6.2 mm) (Figs. 1, 2). The sample of Ragdolls examined included other descendants of the first Berberis. It is evident from the HCM distribution maps that both N/N and N/HCM genotypes are present in this lineage. Only one eight-month-old cat, out of 34 examined so far, was found to have the HCM/HCM genotype (positive homozygote). Echocardiographic testing of this homozygote in early December 2008 has so far been negative for HCM (IVSd 5.1 mm and LVWd 5.1 mm) (Figs. 3, 4).
Since December 2008, all genetically tested ragdolls have been examined echocardiographically, according to established standards.17-19 We compare genetic testing with echocardiographic findings and also perform radiographic or ECG examination and therapy according to clinical status. We also follow up any sonographic manifestations of the C820T mutation, age range of occurrence, prevalence or also penetration in case of crossbreeding and examination of offspring including positive individuals. In case the genetic test is negative and the echocardiographic findings are positive, the patient's documents and swabs will be further investigated to detect any new mutation. The use of a relatively new method for the determination of the natriuretic peptide BNP in cats, from the plasma of the animals examined, remains an open question.20-23 Studies have shown that the level of BNP can be used to determine whether HCM-related changes in the cardiac muscle are already present in the early stages of the disease.20-23 Intensive communication with foreign laboratories regarding the determination of BNP is currently underway.
By comparing the map of HCM spread in European diagnosed cases and the map of DNA testing in Ragdolls in the Czech breeding stock, it is clear that the Berberis lineage extends beyond the founders, which are the basis of the positive European animals and animals that have been proven to have died of HCM. The Berberis line is descended from breeders originally united around the founder of the breed A. Baker (Raggedy-Ann) and later organized in TICA. However, the mainstream of European Ragdoll breeding is derived from British breeding, where Ragdolls from the Blossom-Time D kennel were imported. Daytona from the USA. These Ragdolls were based exclusively on acromelanic (Siamese) coloured lines. On the other hand, the ancestors of Berberis were also solid colored and included lilac Balinese. Both groups, however, had common founding ancestors: Josephine, who is not yet considered a Ragdoll, and three founding cats: Raggedy Ann Dady Warbucks, Raggedy-Ann Fugianna, and Buckwheat. The latter was a black, solid color. The main stud cat Daddy Warbucks was later replaced by Raggedy Ann Kyoto, who died relatively young for unknown reasons. The representation of these common ancestors in all pedigrees may suggest the possibility that the gene identified as causative for HCM has been in the Ragdoll population since the beginning of breeding. However, this observation is somewhat complicated by the fact that Ragdolls that have been shown to have HCM have rarely been genetically tested - also because some deaths occurred before the discovery of the C820T mutation in the MYBPC3 gene. Records of the examinations and genetic tests performed on individual Ragdolls, and their pedigrees, can be found in the international online Ragdoll Health Database, or the linked international PawPeds database. From the HCM gene distribution map, it is clear that the breed was able to cope with the heterozygous genotypes and the cats lived to a great age!
Coefficients of relatedness of animals from breeding between Berberis descendants reached quite high values in SCHK in the 1990s (at least in one case even 31.2% ). This strong inbreeding could have shifted the frequency of the defective allele very significantly in one or the other direction by the action of genetic drift. The monitoring of allele frequencies will be the subject of further, accompanying research. However, at present there are not enough genetically tested animals to calculate the frequency of the defective allele in the population with satisfactory accuracy and to compare allele frequencies in subsequent years with this value. It is known that some of the heavily inbred animals are still in good health with no signs of health problems. On the other hand, there are known deaths in the NCHK of young animals aged 4 to 6 years whose parents were interrelated Berberis offspring with inbreeding coefficients of 15, 6 and higher. However, these deaths were not linked to HCM and were thought to be due more to inbreeding depression. In these individuals or HCM was also never confirmed by autopsy.
It is clear from the map of the distribution of HCM that the deaths of very young animals in European breeds occurred in lines derived from purely acromelanic individuals, not in lines where full-blooded animals were also represented. Moreover, according to current knowledge, the early deaths were exclusively the offspring of Ragdolls from the union of two founder Ragdolls originally of British breeding imported from the USA. It is very interesting that the partial coefficients of relatedness in a standard, four-generation pedigree are zero for the parents of affected individuals, while the overall coefficients often exceed 40%.
The experience of Czech breeders is similar to that of German breeders, who find positively tested animals in lines in which HCM has never been reported. So is the gene discovered by the Washington State University team any different? Is testing Ragdolls unnecessary? Also, the course of the disease cannot be accurately compared to human medicine because there is no exact copy of the C820T mutation as in Ragdolls. However, the two close mutations at codons 810 and 820 in humans are interesting. The R810H mutation has been described in two humans with different clinical and echocardiographic findings. A severe form of HCM was found in the homozygote R810H(3B) (arginine for histidine). A heterozygote R810H(10B) had a moderate form of HCM.24 The MYBPC3 Arg820Gln (R820Q) (arginine for glutamine) mutation was the first to be described in MYBPC3, which clinically manifests and is diagnosed as dilated cardiomyopathy in elderly and elderly people, although the gene mutation in MYBPC3 is typical of HCM.25 Do mutations at codon 820 in cats manifest similarly to humans as dilated cardiomyopathy in the elderly? What is the general course of the disease we can expect in Ragdolls?
Conclusion
We believe that despite all the efforts of scientists dedicated to research, our knowledge of HCM is still limited. In this situation, it is a considerable gamble to dismiss the genetic test, as HCM may be only one of the peaks of a related genetic problem. The same is true for keeping individuals of the N/HCM genotype in breeding. We know too little to afford to remove them from the breed unless other problems appear in their litters. However, by the same token, we need to test their progeny and keep as careful a record as possible in the studbook of any problems that may occur. We are at the beginning of a long journey to protect low-numbered populations. We hope that our clinical trials and comparisons with genetic test results will be of benefit to both Siberian and Maine cat breeders, and will also serve to correctly interpret genetic test results and their predictive value.
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Author Address:
MVDr. Ľuboš Hrib
Veterinary Clinic Jesenice
Budějovická 81
252 42 Jesenice - Prague
E-mail: humerus@azet.sk