1. Diversity and selection of the MHC class II genes in canids
Research plan seminar
Alina Niskanen

2. Contents Background Major histocompatibility complex (MHC)
Finnish wolf population
Diversity in dog breeds
Objectives
Material
Methods
Results

3. Background - MHC Major histocompatibility complex (MHC):
Gene dense region
Vast amount of immune genes
Highly polymorphic
Associations to parasite infections and immunological diseases
MHC has been studied for decades especially on human and mouse
The most important immune gene region in the genome
Lowered diversity can lead to lowered immunity against diseases and parasites
higher extinction risk

4. Background – MHC class II
MHC genes divided to three classes based on their function
Class II genes recognise extracellular antigens
Extracellular
Transmembrane
Cytoplasmic
Picture: Dukkipati, V. S. R., Blair, H. T., Garrick, D. J. & Murray, A. 2006: ‘Ovar-MHC’ - ovine major histocompatibility complex: structure and gene polymorphism. – Genetics and Molecular Research. 5:

5. Background – MHC class II function
Picture:

6. Background – Selection in MHC
Balancing selection maintains polymorphism by:
Heterozygote advantage
Frequency-based selection
Alleles with low frequency are favoured
Accumulation of recessive harmful alleles around MHC
> Purifying selection against homozygotes
Positive selection for new alleles:
Excess of non-synonymous mutations compared to synonymous mutations
Prenatal selection detected in South Amerindians
Offspring more heterozygous than expected, depends on mother’s genotype

7. Background – Finnish wolf population
Started to decline in the late 19th century
Caused by hunting and human land use
Only few individuals in 1920s
Has followed the changes in the Russian Karelian population until last decade
Around 200 individuals at the moment
Finnish and Russian populations diverged
-> Populations separating?
Picture:

8. Background – Diversity in dog breeds
Direct descendant from wolf
Bred by strict breeding standards for years
Highly inbred
Low genetic diversity within breed, high among breeds
Immunological diseases getting more common

9. Objectives
1. What is the level of MHC diversity in Finnish versus Russian wolf
population?
2. What is the effect of different breeding histories to variation in the
dog breeds?
3. Is it possible to detect selection in MHC class II loci in wolves and
dogs?
4. Is prenatal selection taking place in the dog and the wolf?
5. Are there associations between wolf MHC class II loci and parasite
infections?
6. Is neutral microsatellite heterozygosity correlated with MHC
heterozygosity in dogs and wolves?

10. Material - Wolf Finland: 141 tissue samples Russia: 43 pelt samples
Echinococcus granulosus examined from 92 individuals; 15 infected
Trichinella spp. examined from 99 individuals; 33 infected
Russia: 43 pelt samples
Picture: Aspi et al Genetic structure of the northwestern Russian wolf populations and gene flow between Russia and Finland. Conserv Genet (In Press)

11. Material - Dog Dog samples include:
72 Löwchens
69 Icelandic sheepdogs
100 Finnish hounds
100 Kromfohrländers
80 Jackrussel terriers
Collected from Finland during the last 3 years

12. Material – Loci Three MHC class II loci: Neutral autosomal locus
DLA-DRB1, DLA-DQA1 and DLA-DQB1
Neutral autosomal locus
Dogs: 22 microsatellite loci
Wolves: 10 microsatellite loci from previous study

13. Methods - Diversity Diversity measures:
Observed (Ho) and expected (He) heterozygosity
Inbreeding coefficient (FIS)
Allelic richness
Nucleotide diversity
Differentiation between wolf populations:
FST

14. Methods - Selection Genotype level Sequence level
Ewens-Watterson test for homozygosity
Based on expected homozygosity with given allele number
Sequence level
Tajima’s D
Fu & Li F* and D*
The ratio of non-synonymous to synonymous substitutions (dN/dS)
Whole sequence
Peptide binding regions
Selection parameter ω (based on dN/dS) for each codon in Bayesian settings including recombination

15. Methods – Prenatal selection & recombination
Study units two puppies per litter and the parents
MHC heterozygosity and haplotype frequencies compared to the values expected based on their parents values
MHC supertypes
Groups of alleles, which overlap functionally
Recombination rate monitored from data with 3 generations

16. Methods – Parasite association
Association between MHC and parasite infection:
χ2-test for each allele and both parasites
Effect of heterozygosity on parasite prevalence:
Oneway analysis of variance (ANOVA)

17. Results – Diversity Preliminary results:
Finnish wolf population slightly less diverse than the Russian wolf population
Icelandic sheepdog most diverse, Löwchen least diverse

18. Results - Selection Preliminary results imply balancing selection:
Allele frequencies in Finnish wolf population more equal than expected
Also Tajima’s D, Fu & Li D* and F* significantly positive
And positive selection:
Excess of non-synonymous substitutions in peptide binding regions (DRB1 and DQA1 loci)
Exact codons under selection are expected to be found with the Bayesian approach

19. Results – Prenatal selection
Expected results:
If prenatal selection is taking place, we will find non-Mendelian segregation in the progeny
If functional MHC-supertypes can be detected, the selection might be happening on supertype, rather than allelic, level

20. Results – parasite association
Homozygosity of MHC haplotype is associated with Trichinella spp. infection
Less E. granulosus infections in individuals carrying DQA1*00201 (p = 0,034) and DQB*02901 (p = 0,035) alleles
Less Trichinella spp. Infections in individuals carrying DRB1*05301 (p = 0,007) allele
Mean homozygosity
No infection Infection

21. Objectives
1. What is the level of MHC diversity in Finnish versus Russian wolf
population?
2. What is the effect of different breeding histories to variation in the
dog breeds?
3. Is it possible to detect selection in MHC class II loci in wolves and dogs?
4. Is prenatal selection taking place in the dog and the wolf?
5. Are there associations between wolf MHC class II loci and parasite
infections?
6. Is neutral microsatellite heterozygosity correlated with MHC
heterozygosity in dogs and wolves?