Developing a reliable mastitis challenge infection model is required to test new intramammary antimicrobial preparations, other novel bovine mastitis treatments, and study mastitis pathogenesis. Three treatment groups of Holstein Friesian cows in active lactation were administered two doses (104 and 106 cfu/quarter) on a single occasion with one of the three
Streptococcus uberisstrains (BFR6019, MFF1283 and SA002) suspended in 5 ml of sterile PBS, administered via intramammary inoculation immediately after milking. All quarters that were challenged with S. uberisstrains MLF1283 and BFR6019 showed clinical signs of mastitis on day 1 and 2 after the challenge. Strain SA002 had a lower rate of inducing clinical mastitis which was detected later than day 3 after the challenge. We successfully developed a rapid and reliable model for inducing experimental S. uberismastitis with 100% success rate in cows in active lactation. On the basis of the correlation results between strains, RAPD fingerprinting results, clinical findings, and a 100% success rate of mastitis induction for low and high doses S. uberisstrains MLF1283 and BFR6019, strain virulence seems to be a more important effect than challenge dose in induction of clinical mastitis following experimental challenge.