We’re Fighting Bloat
The following is from Dr. Michael A. Harkey, chief investigator for the AKC/CHF research study entitled “The Genetics of Bloat in German Shepherd Dogs: The Roles of Immune System Genes and the Gut Microbes”. The American German Shepherd Dog Charitable Foundation is proud to sponsor this research study.
Update From Dr. Harkey (May 2018): We have proposed that genetic tendency to bloat may be caused largely by certain “risk” variants of genes of the immune system. These genes are responsible for distinguishing foreign cells from “self”, and as such, regulate which bacterial species in the gut will survive and which will be targeted for destruction. So these genes regulate the so-called “gut microbiome”. We hypothesized that dogs with a particular set of immune gene variants will maintain an unhealthy microbiome that predisposes them to bloat. Our recent research has supported this hypothesis in Great Danes.
The purpose of the proposed research is to expand the genetic and microbiome analysis of bloat to German Shepherds. We proposed to repeat the genetic and microbiome analysis, described for Great Danes, in this group of German Shepherds. We plan to sequence the 3 immune genes, DLA88, DRB1, and TLR5, and determine if any variants associate with bloat. We will then analyze the gut microbiome profiles of 50 affected dogs and 50 controls to look for changes in the bacterial population that correlate with bloat.
Early Result of Genetic Analysis
With about 60% of the genetic data now in, we have preliminary analysis of the association of genetic variants with bloat. These results already show strong genetic associations with bloat that may lead to genetic tests for risk of this condition in German Shepherds.
DLA88 Gene. For the gene, DLA88, so far no variants associate with bloat. The “Risk” variant of this gene, that associated with bloat in Great Danes, has not been seen in German Shepherds Since the frequency of GDV in German Shepherds is 4-5 fold lower than in Great Danes, the 5101 allele may be correspondingly less frequent in the Shepherds. Even by that logic, we should have seen this allele 3-4 times so far in the German Shepherd data set. Alternatively, the DLA88:05101 may play a breedspecific roll in bloat in Great Danes.
DRB1 Gene. Our data for the DRB1 gene shows a strong association of variant 1201 with bloat. This is the same variant that was identified in Great Danes as a “risk” variant. So DRB1:1201 may be a good genetic marker for bloat in a wide variety of breeds. Additionally, allele 1501 associates with the bloat group in German Shepherds. This allele was relatively rare in Danes (about 3% of all alleles) but is very frequent in Shepherds (30-50%). The risk effect of 1501 is dose-dependent: that is, dogs with two copies of this variant are at higher risk of bloat than dogs with just one copy. Allele 1101 show a significant protective effect, associating with the healthy control group.
TLR5 Gene. In our previous study, this gene had only two variants in Great Danes, designated as “A” and “B”. The “B” allele was found to be a risk allele, with a 4-fold higher frequency in the bloat group. The emerging data for TLR5 in German Shepherds also shows two variants. But, as with DLA88, the risk variant found in Danes is not found in German Shepherds. Instead, we have observed a new variant, designated as “C”, which has not been previously reported in any breed. The “C” variant associates strongly with the control group, defining it as a protective variant. Since only 2 variants exist in the German Shepherds, “A” is a risk variant in this breed. But this same “A” variant was defined as the protective allele in Danes, because the other allele (“B”) associated with the GDV group. So this sets up a possible hierarchy of TLR5 variants, with “C” as the most protective, “B” as the most risky and “A” as relatively neutral, with respect to bloat risk. The predominance of the risky “B” variant in Danes and the protective “C” in Shepherds correlates with the much higher risk of bloat in Great Danes.
Update From Dr. Harkey: We are just moving out of the collection phase and into the analysis phase of the study. So we do not yet have big enough numbers for strong statistical significance. But we do see one interesting number emerging for the DRB1 gene. In Great Danes, the DRB1: 1201 allele was associated with GDV. In the German Shepherds, this allele is already emerging as a significant risk factor. 1201 occurs at a 13 fold higher frequency in GDV dogs than in controls, with a p value of 0.0013. Allele 1101 appears to have a protective effect, being twice as frequent in control dogs with a p value of 0.0089. We don’t have enough data yet to see significant associations with DLA88, and TLR5 shows no trend at all. Interestingly, the B allele of TLR5, which associated with risk for GDV in Great Danes, is completely missing from the German Shepherds. A new allele (“C”), which has never been reported, has been seen 22 times in our GSD study, evenly distributed between GDV and control groups.
The microbiome analysis is still at an early stage. We are just getting the last samples in this week, and much of the processing for bacterial DNA is done. But the high throughput sequencing and analysis are waiting for the last samples. They will all be sequenced together and the analysis should be fairly
The causes of bloat are poorly understood and may involve multiple triggers. Certainly, genetics plays a role. In addition, the scientific literature implicates a variety of factors, including age, gender, diet, exercise, position of the food bowl, stress, and even weather conditions. Except for stress, much of the data is equivocal My working theory is that specific genetic factors shift the gut bacterial population (microbiome) to an unhealthy state (dysbiosis), which predisposes the dog to bloat. Then some non-genetic trigger, such as stress, or an unusual meal can set off an episode of bloat in a genetically predisposed susceptible.
The logic for this theory is complicated, but this is it in a nutshell. Certain genes of the immune system (DLA genes in dogs) are responsible for detecting foreign invaders (like bacteria) and starting the process of destroying those invaders. These genes are highly variable, each having hundreds of variants (alleles) within the larger population of dogs. So each individual dog has a pretty unique array of these alleles, and a pretty unique ability to detect and destroy the thousands of potential invaders. These same genes influence the types of bacteria that are allowed to grow in the gut. So the microbiome of each dog can vary, due to its DLA genes. Our hypothesis is that certain “risk” alleles of the DLA genes cause dysbiosis of the gut microbiome, which in turn, predisposes the dog to bloat.
Our work in Great Danes supports this hypothesis. It showed that each of 3 immune genes has a single “risk” allele that predisposes carriers to bloat. A Great Dane carrier is three times more likely to bloat than a non-carrier. This study is now in press at the American Journal of Veterinary Research. A study of microbiome composition in this same group of dogs showed significant changes in the bacterial population, associated with bloat. These changes included a two-fold change in the ratio of the two most abundant bacterial phyla, Firmicutes and Bacteroidetes. These are major changes, but we do not yet know the biological significance of these changes, but they are clearly associated with the bloat group. The microbiome study is about to be submitted to PLOS ONE
Now, with the help of your support, we are asking if these same factors, DLA genes and gut microbiome, play a role in bloat in German Shepherds. We do not yet know if the data from Great Danes is breed-specific, or if it might apply to all dogs. If we find similar associations in German Shepherds the implication is that our hypothesis holds for dogs in general. If so, we may be able to establish a universal genetic test for predisposition to bloat. The microbiome data may lead to probiotic or dietary therapies to prevent bloat, even in predisposed dogs. So now that we have your financial support, we are excited to start to answer some of these questions.
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