The U.S. Fish and Wildlife Service federal protocol for dual disinfection of fish eggs in 50 mg/L iodine solution for 30 min followed by a secondary disinfection in 100 mg/L iodine for 10 min was investigated during six spawning cycles of Atlantic salmon Salmo salar held at the Richard Cronin National Salmon Station (Sunderland, Massachusetts). This population of salmon had undergone an epizootic of furunculosis, and the surviving fish maintained a persistent infection of Aeromonas salmonicida throughout the course of study. Eggs from 20 individual paired matings of salmon were obtained annually during the first 2 weeks of November in each spawning cycle from 1995 through 2000 except for 1999, when fertilized eggs from 35 pairs of salmon were examined. Aeromonas salmonicida was isolated from 19 of the total 135 groups of fertilized eggs investigated during this study. In those cases, all isolations of the pathogen were made only in fertilized eggs that had not yet undergone disinfection in iodophor. In contrast to the results produced in the field, in vitro assays showed that A. salmonicida was not completely killed when initial concentrations of the bacterium ranged between 1.0 × 10⁷ and 1.2 × 10⁸ colony forming units (CFU)/mL. However, even when bacterial concentrations exceeded 1.0 × 10⁷ CFU/mL, no A. salmonicida remained viable in eggs treated first with 50 mg/L iodine for 30 min and then with 100 mg/L iodine for 10 min, as prescribed in federal policy. Results of the current analysis also provided further evidence that A. salmonicida is not transmitted vertically through intra-ovum infection.