Mastitis is a major economic and welfare issue costing an estimated £120M/annum to the UK sheep industry. Mastitis is a bacterial infection that can lead to decreased milk yield, premature culling and increased ewe mortality. An intervention study was conducted on one flock in Great Britain, from February to April 2017, to investigate the effect of additional hygiene protocols during indoor lambing on the transmission and persistence of bacteria within ewes and between ewes and lambs at strain level. Hygiene protocols for intervention ewes including use of antibacterial hand gel before handling or providing lambing assistance to ewes, and use of antibacterial bedding powder. Intervention ewes were managed by researchers. Control ewes, managed by farm staff, did not receive these additional hygiene protocols. Ten twin-bearing ewes (5 control and 5 intervention) were swab-sampled on 4 occasions: pre-lambing, immediately post-lambing, 24 hours after lambing and when leaving housing. Swab locations included the ewe’s nasal passage, vagina, both teats, lambs’ mouths, and the handler’s hands. Aseptic milk samples were collected from both udder halves. From 168 cultured samples 793 isolates were identified and analysed by matrix-assisted laser desorption/ionisation time-of-flight mass spectronomy (MALDI-ToF-MS). Unsupervised hierarchical clustering of the resulting spectra was used to assess the similarity between isolates and identify strains. Initial analysis indicates clustering of strains between ewe teat and lamb mouth, between lambs’ mouths, and between the two udder halves (both teat and milk) over time. There is also evidence of strain clustering between lambs’ mouths, and between the udder halves (both teat and milk) at the same time point. Further analysis will explore the possible transmission pathways and persistence of bacteria associated with ewe mastitis and investigate any intervention impact.