Integrating clinical, data-driven and in-vitro approaches to the study of host-pathogen interactions in bovine digital dermatitis

Cattle lameness is a debilitating and painful condition, and is described as one of the clearest indicators of compromised welfare in dairy cattle and one of the most important causes of involuntary removal and replacement of animals. No other common condition is associated with such visible signs of pain and, as such, lameness also damages the public's perception of the industry. Painful foot lesions account for more than 90% of reduced cow mobility and lameness cases. One main cause of cattle lameness is Digital Dermatitis (DD) a painful, infectious, foot skin disease affecting ruminants worldwide. The disease is endemic on more than 90% of UK dairy farms and more than 50% of UK dairy cows are affected annually. In addition to pain and compromised animal welfare, DD is also associated with reduced milk yield, feed intake, and reproductive performance, and estimated to cost the UK dairy industry more than £74 million per year. Bacteria of the genus Treponema are considered the main pathogen associated with DD; however, we still don't fully understand how the disease develops and what the role of the animal's genetics in this is. Current control strategies are mostly generic and lack a substantial evidence base, relying on the empirical use of topical antibiotics and foot-bathing solutions; the latter however often contain heavy metals, such as copper sulphate, or formalin (carcinogen). The use of hazardous chemicals and the increasing evidence for antibiotic resistance in DD treponemes mean current control methods may pose a serious threat to human health and environment.
We hypothesise that animal genetics play a key role in the development of DD with resulting differences in animal-pathogen interactions determining the development and progression of the disease. Our overarching goal is to conduct an interdisciplinary, integrated study to further our understanding of the mechanisms leading to the development of DD, investigate animal-pathogen interactions, and determine the optimum evidence based breeding strategy to enhance animal resistance to DD development and animal ability to recover from DD. More specifically we will perform an in-depth characterisation of the genetic background of animal DD phenotypes in order to identify what are the key genes, mutations and cell populations playing an important role in the disease. Moreover, we will investigate the role of animal-pathogen interactions in DD development and progression utilising novel laboratory models of the disease. Finally, we will use all these novel knowledge to develop advanced strategies to control this debilitating disease. A thorough understanding of these mechanisms will underpin modern breeding programmes aiming to reduce incidence of DD and improve recovery from DD, and inform the development of effective targeted treatments by helping to identify novel vaccine and drug targets. This will improve animal health and welfare, potentially reducing further development of antimicrobial resistance, and support the production of healthy food. Associated decrease in involuntary culling of animals and increase in cattle longevity will indirectly contribute to reduction of greenhouse gas emissions and improve the sustainability of the sector. Mounting welfare and cost issues brought about by increasing prevalence of DD in bovine populations both nationally and worldwide demonstrate the timeliness of the proposed project.

Genotypic, pedigree and phenotypic clinical data are already available for a population of 3,286 Holstein dairy animals. A new population of 500 dairy heifers will be prospectively monitored. We will be inspecting feet monthly and record presence of digital dermatitis (DD) lesions. DD resistance phenotypes and a novel DD progression phenotype will be developed. Blood samples for genotyping, whole genome sequencing and serum harvesting will be collected. Individual animal data will be analysed to determine genetic variants associated with the DD-related phenotypes. We will also derive genomic breeding values for each animal and DD phenotype. Foot skin biopsies will be performed on 24 animals genetically resistant to DD that will remain healthy throughout the study and 48 genetically susceptible animals (including recovered and chronically infected). Foot skin biopsy samples will be processed for total bulk RNA-sequencing in order to define DD resistance and progression transcriptomic signatures. We will perform single cell RNA-Seq in a subset of samples to dissect which cells are the key players driving DD resistance and progression. We will isolate fibroblasts and keratinocytes from cattle foot skin of the three respective biopsied groups and will co-incubate them with both a poly-treponemal mixture and an individual treponemes species. After co-incubation we will undertake RNA sequencing to determine global differential mRNA expression. The cattle foot skin culture model will be combined with a transepithelial cell migration assay to assess migration of granulocytes through bovine foot skin keratinocytes and fibroblasts. We will prioritise candidate genes and genetic variants for DD using a combined data-driven and biology driven approach, and will validate key findings using in vitro techniques. Outcomes from the previous steps will be integrated in a series of simulation studies to determine optimal biology-driven selective breeding strategies for DD control.