Inflammatory Myopathy (Myositis) The Journey Towards Discovery
2011I received a call from one of the puppy owners of the Becker x Alpha litter stating concern about her pup, a Dutch Shepherd male named “Polo”. He acted as if he was stiff, she said, and couldn’t walk properly. She worked as a veterinary technician and the dog was currently at her clinic. Since that clinic was in relative close proximity to my veterinarian Karen Wroblewski DVM of Allen Animal Clinic in Livonia, I went down, picked Polo up and brought him to Dr. Wroblewski.
Upon seeing Polo, my mind went directly to the “mystery disease” I had previously seen crop up in some of the dogs of my obedience training clients. Stiff, no temperature, no appetite… Many of the vets that had seen these dogs were at a loss. What I had found was treatment with Doxyclycline or Tetracycline for 45 days did the trick. Was this a tick borne disease? Perhaps a form of Lymes disease? Toxoplasmosis was first suspected but nothing was apparent, only that the old fashioned antibiotics seemed to work. On speaking with Dr. Wroblewski, she agreed to treat the Dutch puppy “Polo” with antibiotics, and sure enough, he seemed to come around.
During this same time, I was going through treatment for a reoccurrence of breast cancer, so between chemotherapy then daily radiation treatments while still trying to work, I chalked it up to another situation of a dog coming with the “mystery disease”… until I took a call from another client in California with a pup from the same litter. She also had her dog at the vet where he showed the exact same symptoms. Unfortunately, this vet was not so supportive, so I paid $500 toward the vet bill just to get them to put the dog on the antibiotics that seemed to be working for his littermate “Polo”. That dogs’ name was “Zander”. Now, it seemed that there was something going on with this litter, so I began working on transport for “Zander” to come to Michigan for further treatment, but his original owner had given up and signed him over to the Vet clinic. There was no way they were going to ship the dog to us. “Zander” ended up being euthanized in California because he became so affected he could not stand by himself. More concerning, the early signs of recovery we saw in “Polo” were disappearing.
We reached out to all the people who had pups from this litter. Most were fine, but a local owner of male Dutch Shepherd “Achilles” realized that his dog was showing symptoms and then a local female named “Bella” was also affected by now what seemed to be an unstoppable disease.
At this point, Dr. Karen Wroblewski went into overdrive. She called, contacted, wrote and e-mailed other veterinarians from across the country and just about anybody she could think of to try and figure this out. She arranged for tissue samples from the 4 affected dogs to be examined by the University of California, Davis Neuromuscular Lab and although they originally diagnosed 4 different types of muscular dystrophy, this was viewed with skepticism as 4 types would be highly unlikely (especially since 1 affected dog was a female and true muscular dystrophy does not affect females). Muscular Dystrophy was eventually dismissed from consideration by UC Davis, but not before samples were also tested by the Lisa Project in Florida to see if the gene mapping was similar to those mapped in human Muscular Dystrophy. Their research was also abandoned because the myriad focal points of disease did not correlate with the variables in human Muscular Dystrophy. 2012At this point, we were stalled. All the dogs from the Becker x Alpha litter that were affected had been euthanized, and the following litters of closely related dogs showed no signs of this disease. We had considered other possibilities, like a parasite in our dog food (we feed raw) or a mosquito carried disease. Was it a onetime occurrence? Did we cause it? What could be done now?
2013In March 2013, we shared our experience to date with the North American Dutch Shepherd Rescue organization to make them aware (and with hopes they would share with us if any dog they came across was showing similar symptoms).
Believing that this was a genetic disease, I wanted to make certain it was not found in our foundation stock, since “Becker” the sire of the affected litter was a son of our Touché and Rush. I then bred an experimental litter between Becker and his daughter Mafia in October 2013, with the intention of keeping all the pups until muscle biopsies could be performed. I certainly breathed a sigh of relief when all five pups born showed no signs of the disease. That, in my mind, proved that it didn’t come down from the sire, and my attention turned back to the lines that Alpha came from. I reached out to several people that I believed would know a little something about the pedigrees, including an experienced breeder in the Netherlands, but she reported back that no one she contacted seemed to know any dog, or breeder, that had run across this problem in the Netherlands. 2015Feeling confident that Becker was not at fault, I bred him in October 2015 to the beautiful Grand Champion “Chanel” (ROM-XX NGRCH RBIS RBISS UUFCH SI AN UAGI USF CA URO1 GSJCH GRCH Cher Car's Some Like It Hot) who is out of our Lothar and Cayenne. This breeding was the first “line bred” litter I was planning, and was really looking forward to the pups. 2016We did indeed get a very nice litter. (One of those dogs from this litter is in our club, “Multi-HIT URO1 UAGI EV EE EI MN USJCH CA GRCH Cher Car’s I’m Your Blind Spot” a large, blue brindle male that is exactly what I thought I would get with this breeding.) MUCH TO MY DISMAY… one pup in the litter was showing the unmistakable signs of this mystery disease!!! How was that possible? How did it NOT happen in the Becker x Mafia litter? What was this dreaded disease, and what to do next???
This reoccurrence, with the same sire as the affected litter, pointed Dr. Wroblewski directly at this being a genetic disease. Biopsy results from the affected puppy were sent to the University of California, San Diego - Comparative Neuromuscular Laboratory where an inflammatory and necrotizing myopathy of undetermined origin was concluded and a GWAS study (genome-wide association study) was recommended on their report of March 7, 2016.
Samples were then forwarded from the University of California, San Diego - Comparative Neuromuscular Laboratory to the University of Minnesota Canine Genetics Laboratory and they identified a gene locus on chromosome 36 as the probable affected locus on the affected dogs. A whole genome association study (WGA study or WGAS) was recommended to study the genetic variants in different individuals to see if any variant is associated with this inflammatory myopathy. Dr. Karen Wroblewski began coordinating with Katie Minor R.N. at the University of Minnesota Canine Genetics Laboratory and they agreed to perform the WGA study. Payment to fund the research was forwarded to the University of Minnesota Department of Veterinary Biosciences in May 2016.
2017The University of Minnesota Canine Genetics Laboratory completed the genome association study and identified the abnormal gene sequence, then began working on a DNA test to allow for breeding dogs to be screened. We then forwarded individual samples from 41 Dutch Shepherd cheek swabs to UMN and they were successful in creating a DNA test. (Although we had already removed Alpha from our breeding program in 2011 and Becker from our breeding program in 2016, test results revealed two other dogs in our then breeding program were identified as carriers and they were immediately removed as well.) On March 13, 2017, the Penn Vet Working Dog Center published a request on their facebook page asking for 24 to 48 additional DNA samples of healthy, unrelated, adult Dutch shepherds to assist UMN in evaluating their DNA test results and identified Dr. Karen Wroblewski as the veterinarian spearheading the collection.
The following synopsis was presented to Dr. Karen Wroblewski on March 24, 2017 from the University of Minnesota Canine Genetics Laboratory:
POLYMYOSITISA previously unknown inflammatory and necrotizing myopathy was characterized in five related Dutch Shepherd dogs. Clinical signs included muscle tremors, pelvic limb stiffness, progressive weakness, and severe muscle atrophy. The serum creatine kinase activity was elevated in all affected dogs. The age of onset was 3 – 9 months; all were euthanized prior to 2 years of age. Histopathologic examination of muscle biopsies revealed multifocal areas of mixed mononuclear cell infiltration, myonecrosis and regeneration. The severity and early onset of clinical signs, coupled with the pedigree knowledge, made a recessive mode of inheritance likely. From the available pedigree we reasoned that the condition was very like an autosomal recessive disease, in which cases contain two copies of the mutation and the parents contain a single copy. This allowed a research strategy where we scan the genome with high density DNA marker arrays for regions of chromosomes that fit the pattern. Homozygosity mapping of 5 cases (4 full-sibs, 1 half-sib) and 5 controls (2 parents, 3 full-sibs) via Illumina CanineHD SNP genotyping BeadChips identified a single chromosome (canine chromosome 36; CFA36) of interest where the gene containing a responsible mutation is expected to be located. However, the issue remained that the critical interval of CFA36 included more than 140 protein coding genes. Whole genome resequencing of one case was performed and identified variants in the critical intervals compared to control genomes from 492 dogs of diverse breeds. Of particular note, a mutation within a gene of the mitochondrial solute carrier transporter gene family, that lies within the critical interval on CFA36, was identified. Follow-up genotyping revealed that all 5 cases were homozygous for the mutant allele, both parents were heterozygous, and ten sibs and first degree relatives were heterozygous or homozygous for the reference allele. The mutation was not identified in 16 population control Dutch Shepherds or 56 other dogs of related breeds (Belgian Malinois, Belgian Shepherd, Belgian Tervuren, German Shepherd). This mitochondrial transporter gene is expressed in brain and skeletal muscle tissues. The literature contains a report of genetically modified mice that do not express this gene that show decreased growth, generalized tremor, ataxia, impaired CNS myelination, and decreased survival. Limited histochemical staining for the mitochondrial specific enzymes succinic dehydrogenase and cytochrome C oxidase did not reveal any structural mitochondrial abnormalities.
These data allow us to suggest that this gene mutation is responsible for the novel myopathy in Dutch Shepherds.
Dr. Karen Wroblewski delivered a Dutch Shepherd Health Seminar at the Dutch Shepherd Dog Club of America National Specialty Show on April 7, 2017 during which she presented these Polymyositis findings and moderated a discussion group.
2018The University of Minnesota Canine Genetics Laboratory concluded the testing of 85 US dogs and a collaborator in the Netherlands tested 72 Dutch Shepherds. These results were used to create a genetic test for the condition formerly known as Polymyositis now identified as IM: Inflammatory Myopathy (Myositis). The UMN College of Veterinary Medicine has updated their Canine Genetic Testing webpage to include submission forms and instructions for submitting samples for Dutch Shepherd Inflammatory Myopathy (Myositis) testing. They have also coordinated with the Orthopedic Foundation for Animals to have the IM test results listed on the OFA website.
The University of Minnesota Canine Genetics Laboratory website also contains a portable document format (pdf) form detailing IM: Inflammatory Myopathy (Myositis) Test Result Interpretation which reveals, amongst other details, that both parents must be carriers to produce affected offspring.
Dr. Karen Wroblewski again conducted a Dutch Shepherd Health Seminar at the Dutch Shepherd Dog Club of America National Specialty Show on March 2, 2018 during which she gave a presentation detailing the 7 year research and discovery process of this disease affecting Dutch Shepherds now called Inflamatory Myopathy and moderated a discussion group afterwards.
In closing, I must say the advances in science absolutely astound me! The fact that a breeder can find out EXACTLY where a genetic problem can be found, that a simple test can be created to first identify and then used to help breed away from a problem is mind boggling. I will forever be indebted to Dr. Karen Wroblewski for her undying devotion to this mission of discovery and to Katie Minor R.N. for coordinating the research project at the University of Minnesota Canine Genetics Laboratory. It will go down in Dutch Shepherd history as a first of many steps to promote the health of our beloved breed.