Summary Study Title The Antimicrobial Activity of Stalosan-F in Moist and Wet Conditions Using S.aureus . MicroChem Laboratory Project ID Numbers 040114-1, 040114-2, 040120-2 In test 1, S.aureus was labeled onto 2x2 cm paper squares to simulate damp straw that might be found in animal quarters, and three paper squares apiece were placed in a sterile plastic petri plates. One set of S. aureus labeled paper squares in petri plates was immediately sprinkled with 0.365g per petri plate of Stalosan-F per label directions using a saltshaker and another set was left untreated as a control. After exposure times of 0.0 minutes, 20.0 minutes, 4.0, 8.0 24.0, and 48.0 hours at ambient temperature, the three paper squares from each petri plate were removed and placed into 10.0 ml neutralizing medium and agitated on a vortex mixer for 60 seconds to release surviving microbes. Serial ten-fold dilutions were made as 1.0 ml into 9.0 ml neutralizing medium, and 0.5 ml portions of the dilutions were placed onto nutrient agar in petri plates to measure the number of surviving CFU of S. aureus at each exposure time. Stalosan-F was found to kill 100% of S.aureus within 8.0 hours at ambient temperature. In test 2 and 3, S.aureus was labeled as 1.0 ml in a sterile plastic petri plate to simulate pooled water or urine. Three petri plates were immediately sprinkled with 0.365g per petri plate per the label directions using a saltshaker and three other petri plates were left untreated as a control. For test 2, after being mixed and held for an exposure time of 20.0 minutes, the petri plates were tilted on their side, 0.8-0.3 ml was removed and placed into a 9.0 ml tube of neutralizing medium. For test 3, after being mixed and held for an exposure time of 20.0 minutes, 4.0 ml of neutralizing medium was placed into the petri plate and mixed for 30 seconds. The petri plates were tilted on their side and 1.0 ml was removed and placed into a 5.0 ml tube of neutralizing medium. The tubes were agitated on a vortex mixer for 30 seconds and serial ten-fold dilutions were made as 1.0 ml into 9.0 ml neutralizing medium. One-half (0.5) ml portions of the dilutions were placed onto nutrient agar in petri plates to measure the number of surviving CFU of S.aureus at an exposure time of 20.0 minutes. Stalosan-F was found to kill 76%-99.8% of S.aureus within 20.0 minutes at ambient temperature. Page 2 of 12
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