Discussion

Recurrent airway obstruction (RAO) is indeed 'in the genes'. In contrast to simple genetic disorders of the horse (e.g. 'lethal white foal syndrome'), however, RAO is a complex disease caused by interactions of environmental and different genetic factors.

RAO is a heritable disease:
Family histories and familial aggregation suggestive of a genetic basis of RAO have been reported for over 70 years (Schaeper 1939; Koch 1957). In the 1990s, it was demonstrated in Warmbloods and Lipizzans that the relative risk of developing RAO was significantly increased more than 3-fold in offspring from an affected dam or sire and almost 5-fold when both parents are affected. Parent, age and stable environment had significant additive effects (Gerber 1989; Marti et al. 1991). More recently, Ramseyer et al. (2007) found that the RAO-risk of offspring by affected sires was 4- to 5.5-fold increased, while hay feeding was the dominant environmental factor. Segregation analyses clearly revealed the presence of major genes for RAO, with an autosomal dominant mode of inheritance in one investigated family and autosomal recessive inheritance in the other family (Gerber et al. 2009). Furthermore, we also found that the ability to mount a high IgE-response is also influenced by genetic factors in the horse, and differs between RAO 'high- prevalence families (Scharrenberg et al. 2010). In contrast, the clinical characteristics of RAO completely agreed between the examined families (Laumen et al. 2010).

Multiple genes are involved in RAO and they differ between affected families
A genome scan using microsatellite markers in the same 2 high- prevalence families studied by Ramseyer et al. (2007) indicated 2 chromosome regions (on ECA13 in family 1 and ECA15 in family 2) with a genome-wide significant association with RAO. An additional 11 chromosome regions showed a more modest association (Swinburne et al. 2009). The fact that the chromosomal regions and the mode of inheritance (see above) do not agree between families indicates genetic heterogeneity for RAO: depending on the genetic make-up of the affected horse different genes confer the susceptibility for the disease. Interleukin 4 receptor (IL4R) is so far the major candidate gene
Many possible candidate genes are located in these chromosomal regions. Several further studies using microsatellite and SNP genotyping and association analyses (e.g. Watson et al. 2003; Jost et al. 2007; Klukowska-Rotzler et al. 2011; Shakhsi-Niaei et al.
2012), as well as gene expression studies in broncho-alveolar lavage (BALF) cells (Klukowska-Rotzler et al. 2011) and hay-dust stimulated blood mononuclear cells (unpublished results) indicate that the IL4R gene is the major candidate gene in a subset of RAO-affected horses with a specific genetic background. Molecular pathway analyses (Ingenuity Pathway Analysis) of genomic and proteomic data showed that interactions between IL4R gene products and another candidate gene (SOCS5) could explain how different genes/genotypes can lead to identical clinical RAO phenotypes: these proteins interact upstream of an important cascade involving NFkB (Racine et al. 2011). Important to note, however, that so far no causal mutation has been identified in IL4R and association studies using the Illumina Equine SNP50 BeadChip in >500 unrelated cases and controls have thus far failed to reveal any chromosomal regions or genes that influence the risk for RAO in the general Warmblood population.
Is RAO susceptibility linked to parasite defence and allergic skin diseases?
While the role of IL4R in RAO appears to be limited to a subset of cases, it is interesting that this candidate gene is associated with asthma and other atopic phenotypes in humans and also plays an important role in the defence against parasites. Together with preliminary clinical observations, these comparative aspects have led to the investigation of novel aspects of RAO: RAO-affected Warmblood horses show an increased risk for developing insect bite hypersensitivity and urticaria (unpublished results), but in exchange they seem to be less susceptible to intestinal infestation with strongylid nematodes (Bruendler 2010). Furthermore, Neuhaus et al. 2010 demonstrated that members of the family showing the association of RAO with IL4R had a more than 7-fold decreased risk of shedding >100 EPG of strongylid nematode eggs compared to unrelated pasture mates. Within this family, the RAO-affected horses had yet a significantly decreased risk of parasite egg shedding compared to healthy offspring. Interestingly, IL4R expression of peripheral blood mononuclear cells from RAO-affected horses of this family is not only increased in response to hay-dust, but also to recombinant strongylid antigen (unpublished results). So far it is unclear, however, as to how RAO and egg shedding (Nussbaumer et al. 2011), and RAO and allegic skin diseases are genetically linked.

Take-home-message:
The present findings suggest a strong and complex genetic basis for RAO. The major genes responsible for RAO differ between families and affected horses. Different genotypes must converge within functional immunological pathways to result in a clinically indistinguishable RAO phenotype.
It is therefore suggest that it is unlikely that single gene tests will be diagnostically useful in RAO. Genetic profiling panels, combining several genetic factors with an assessment of environmental risk factors, may have greater value, but much work is still needed to uncover diagnostically useful genetic markers or even causative variants for RAO, allergic skin diseases and susceptibility to intestinal parasites.

Acknowledgements:
The author wishes to acknowledge the contributions of all co-investigators, graduate students and horses' owners. They are too numerous to mention here by name. Major contributions to this work have come from the Institute of Genetics, the Institute of Parasitology of the Vetsuisse-Faculty, University of Berne, the Animal Health Trust, the University of Giessen und Purdue University, and have been funded by The Swiss National Science Foundation, the Berne Equine Lung Group and The Horse Trust.