Concern over the adequacy of biosafety training and incident-reporting practices within biological laboratories in the United States has risen in recent years due to the increase in research on infectious diseases and the concomitant rise in the number of biocontainment laboratories. Reports of laboratory-acquired infections and delays in reporting such incidents have also contributed to the concern. Consequently, biosafety training and incident-reporting practices are being given considerable attention by both the executive branch and Congress. We conducted a 51-question survey of biosafety professionals in June 2008 to capture information on methods used to train new laboratory workers within biosafety level 2 (BSL-2) laboratories, animal biosafety level 2 (ABSL-2) laboratories, biosafety level 3 (BSL-3) laboratories, and animal biosafety level 3 (ABSL-3) laboratories. The survey results suggest nearly all senior scientists, faculty, staff, and students working in these biocontainment laboratories are required to have biosafety training, and three-quarters of respondents indicated a biosafety or environmental health and safety professional provides explicit instructions on reporting incidents to each new lab worker. Only half of the respondents with BSL-2/ABSL-2 laboratories at their institution and 59% of respondents from institutions with BSL-3/ABSL-3 laboratories indicated custodial or maintenance workers are required to receive biosafety training at the BSL-2/ABSL-2 and BSL-3/ABSL-3 levels, respectively. Opportunities for targeted improvement such as providing training to non-traditional laboratory workers (e.g., custodians, maintenance workers) and posting laboratory incident-reporting protocols on institutional environmental health and safety websites may exist. Variations in biosafety training requirements, incident-reporting practices, and attitudes towards laboratory safety revealed through this survey of biosafety professionals also support the development of core competencies in biosafety practice that could lead to more uniform practices and robust safety cultures.