Type‐specific identification of human adenovirus 3, 7, and 21 by a multiplex PCR assay†

Abstract Human adenovirus (Ad) serotypes 3, 7, and 21 of DNA cluster B:1 are often associated with severe respiratory illness, particularly in infants and young children and, in addition to Ad4, are among the most important causes of acute respiratory disease syndrome in new military recruits. To address the inherent problems associated with classic typing methods, we developed a multiplex PCR assay for the rapid, specific identification of Ad3, Ad7, and Ad21 field isolates. To design type‐specific primers for our assay, we sequenced the Ad21 hexon gene and compared this sequence with previously published sequences of Ad3, Ad7, and Ad16. The overall nucleotide (nt) and amino acid (aa) identities between Ad21 and Ad3, Ad7, and Ad16 were similar (ranges 78.3–80.8% nt; 84.1–86.2% aa), with significantly greater variability in the regions of the hexon that encode surface loops 1 and 2. Type‐specific primers designed to the hypervariable regions correctly identified Ad3, Ad7, and Ad21 prototype strains and 53 previously typed Ad field isolates. No cross‐reactions with other Ad serotypes were identified. Our multiplex PCR assay for type‐specific identification of Ad3, Ad7, and Ad21 isolates will provide a rapid and convenient tool for the epidemiologic investigation of Ad‐associated respiratory illness. J. Med. Virol. 64:537–542, 2001. © 2001 Wiley‐Liss, Inc.