A multistate outbreak of listeriosis occurred in the United States in 1998 with illness onset dates between August and December. The outbreak caused illness in 108 persons residing in 24 states and caused 14 deaths and four miscarriages or stillbirths. This outbreak was detected by public health officials in Tennessee and New York who observed significant increases over expected listeriosis cases in their states. Subsequently, the Centers for Disease Control and Prevention (CDC) began laboratory characterization of clinical isolates of Listeria monocytogenes by serotyping and restriction fragment length polymorphism analysis using pulsed-field gel electrophoresis (PFGE). For the purpose of this investigation, outbreak-related isolates were defined as those that had a specific AscI-PFGE pattern and indistinguishable or highly similar (no more than 2 band difference in 26 bands) ApaI-PFGE patterns when their DNA was restricted by AscI and ApaI restriction enzymes. Timely availability of molecular subtyping results enabled epidemiologists to separate outbreak cases from temporally associated sporadic cases in the same geographic areas and facilitated the identification of contaminated hot dogs manufactured at a single commercial facility as the source of the outbreak. During the investigation of this outbreak, a standardized protocol for subtyping L. monocytogenes by PFGE was developed and disseminated to public health laboratories participating with CDC's PulseNet network; these laboratories were requested to begin routine PFGE subtyping of L. monocytogenes.