Breed distribution and history of canine mdr1-1∆, a pharmacogenetic mutation that marks the emergence of breeds from the collie lineage Author: Neff, M.W. et.al. Presented by Katheryn McDonald and Todd Mercier

Introduction  Goal of study: To identify breeds at risk for multi-drug sensitivity and level of susceptibility for each breed  Ivermectin 1980’s, parasiticide Causes a charge in the ligand-gated chloride ion channels of the peripheral nervous system, resulting in an influx that paralyzes nematode and arthropod parasites Generally safe in domestic animals due to the blood-brain barrier

Introduction  Ivermectin Key component of the blood-brain barrier is P- glycoprotein –Large protein complex that pumps drugs out of the brain and into bloodstream –Encoded by the multiple drug resistance gene (MDR1) –Lack of P-glycoprotein may lead to neurotoxicosis

Introduction  MDR1 mdr1-1∆ –Causes frameshift with multiple premature stop codons, drastically shortening the P-glycoprotein –Allele probably results in a complete loss of P-glycoprotein function –First discovered in the collie breed

Objective  Determination of genetic linkage of the mdr1-1∆ allele in non-herding breeds to find other at risk canine populations  Determination of ancestral lineage in other breeds not previously thought to be related to the collie

Methods  Four classes of dogs were tested Breeds from the collie lineage based on breed history European herding breeds not thought to be related to the collie Sighthounds and miscellaneous breeds that exhibited drug sensitivities Multibreed panel of over a thousand samples from more than 90 breeds

Methods  DNA samples taken from buccal cells collected by oral swab  Genotyping for MDR1 accomplished with PCR (rapid duplication of DNA)  Genome location mapped for MDR1 using a canine/hamster radiation hybrid panel

Methods  Haplotype analysis Loci were selected based on proximity to MDR1 Primers were produced for 4 markers and MDR1 for PCR Haplotypes reconstructed with PHASE program (assumes all haplotypes from a pop. were created in that population)

Methods  Statistical analysis Deviation from Hardy-Weinberg equilibrium in each test was assessed by a χ 2 test in each breed Allele age determined by linkage disequilibrium (LD) at mdr1-1∆ Distance between mdr1-1∆ and marker loci was 1cM or greater, suggests mutation is a weaker force than recombination However, mutation may result in rare haplotypes

Results  4000 samples surveyed, the mdr1-1∆ allele was present in nine breeds Allele %Genotype % Breed # of dogsmdr1-1∆mdr1-1∆/ mdr1-1∆mdr1-1∆/MDR1MDR1/MDR1 Austr. Shepherd Austr. Shepherd min Collie English Sheepdog Longhaired Whippet McNab Old English Sheepdog Shetland Sheepdog Silken Windhound

Results  mdr1-1∆ present in predicted collie breeds  mdr1-1∆ not found in herding breeds traced back to Europe  mdr1-1∆ also found in two non-herding breeds, the Longhaired Whippet and the Silken Windhound (sighthounds)

Results  High frequency of the mdr1-1∆ allele in the Collie and Longhaired Whippet breeds suggests a founder effect for those breeds  So, separate mutation or are they related?

Results  Palindrome located 9 base pairs upstream of mdr1-1∆ could have served as mutational hotspot~ suggest possible unrelated mutations  MDR1 tested with radiation hybrid analysis, revealed marker alleles strongly associated with mdr1-1∆ were identical at 3 of the 4 loci for herding breeds and sighthounds  Identical allele by descent~ all dogs carrying the mdr1-1∆ allele are descendants of a dog from Great Britain before the genetic isolation of breeds

Discussion  mdr1-1∆ was not present in the Border collie, Bearded collie or the Austr. Cattle dog, yet do exhibit ivermectin sensitivity Possible reasons –mdr1-1∆ present at a lower frequency –Another mutation is responsible

Discussion  Within breeds, genotype frequency is consistent with Hardy-Weinberg equil.  No evidence linking mdr1-1∆ with selection or non-random mating  Allele differences most likely arose from genetic drift and expansion of formal breeds since the 1800’s  Ultimately genetic drift and artificial selection have played a major role in genetic composition of breeds

Questions?