Unlike other human adenovirus (HAdV) species, B is divided into subspecies B1 and B2. Originally this was partly based on restriction enzyme (RE) analysis. B1 members, except HAdV-50, are commonly associated with respiratory diseases while B2 members are rarely associated with reported respiratory diseases. Recently two members of B2 have been identified in outbreaks of acute respiratory disease (ARD). One, HAdV-14, has re-emerged after an apparent 52-year absence. Genomic analysis and bioinformatics data are reported for HAdV-14 prototype for use as a reference and to understand and counter its re-emergence. The data complement and extend the original criteria for subspecies designation, unique amongst the adenoviruses, and highlight differences between B1 and B2, representing the first comprehensive analysis of this division. These data also provide finer granularity into the pathoepidemiology of the HAdVs. Whole genome analysis uncovers heterogeneous identity structures of the hexon and fiber genes amongst the HAdV-14 and the B1/B2 subspecies, which may be important in prescient vaccine development. Analysis of cell surface proteins provides insight into HAdV-14 tropism, accounting for its role as a respiratory pathogen. This HAdV-14 prototype genome is also a reference for applications of B2 adenoviruses as vectors for vaccine development and gene therapy.