Our research has focused on the ecology of commensal populations of big brown bats (Eptesicus fuscus) in Fort Collins, Colorado (USA), in relation to rabies virus (RV) transmission. We captured 35 big brown bats (Eptesicus fuscus) in late summer 2001 and held them captive for 4.8 mo. The bats were initially placed in an indoor cage for 1 mo then segregated into groups of two to six per cage. Two of the bats succumbed to rabies virus (RV) within the first month of capture. Despite group housing, all of the remaining bats were healthy over the course of the investigation; none developed rabies, although one of the rabid bats was observed to bite her cage mates. Reverse transcription-polymerase chain reaction (RT-PCR) and Taqman real-time PCR analysis of the RNA derived from the brain tissue, salivary glands, and oral swab samples confirmed RV infection in the dead bats. Rabies virus was also isolated from the brain tissue upon passage in mouse neuroblastoma cells. Nucleotide sequence analysis of the RV nucleoprotein (N) gene showed 100% identity with the N gene sequence of a 1985 E. fuscus isolate from El Paso County, Colorado. Bat sera obtained six times throughout the study were assayed for RV neutralizing antibodies using the rapid fluorescent focus inhibition test. The RV neutralizing activity in the serum was associated with the IgG component, which was purified by binding to protein G Sepharose. Five bats were RV seropositive prior to their capture and maintained titers throughout captivity. Two adult bats seroconverted during captivity. Two volant juvenile bats had detectable RV antibody titers at the first serum collection but were negative thereafter. Four seronegative bats responded to a RV vaccine administration with high titers of RV antibodies. A serologic survey of big brown bats in the roost from which one of the captive rabid bats had originated showed a significant rise in seroprevalence during 2002.