P. Pring-akerblom et al., Multiplex polymerase chain reaction for subgenus-specific detection of human adenoviruses in clinical samples, J MED VIROL, 58(1), 1999, pp. 87-92
The polymerase chain reaction (PCR) has been used previously for the detect
ion acid typing of adenoviruses directly in clinical samples. Since under c
linical conditions subgenus-specific identification is often sufficient, we
extended the genus- and type-specific PCR by a subgenus-specific PCR. By s
equencing several loop I, gene regions of the hexon (major adenovirus coat
protein) and comparing them to published sequences, subgenus-specific seque
nces were identified in this region. By using primers targeted to this regi
on and to a conserved hexon gene region, a multiplex, nonnested PCR for the
detection and subgenus-specific identification of adenoviruses could be es
tablished. The six subgenus-specific amplimers are distinguishable by agaro
se gel electrophoresis, and subsequent restriction analysis is not necessar
y. The specificity of the subgenus-specific primer pairs was tested on 23 a
denovirus prototypes, representing all six subgenera, on 9 subgenus B and D
intermediate strains, and on 16 subgenus C genome types. Furthermore, mult
iplex, subgenus-specific PCR was performed directly with 100 clinical speci
mens, including stool samples, ocular swabs, and throat swabs. Adenoviruses
of all subgenera could be detected. Especially for clinical application, t
he rapid, one-step differentiation between subgenus D adenoviruses, causing
the severe and highly contagious epidemic keratoconjunctivitis, and subgen
us B and E adenoviruses, causing relative harmless ocular infections, is of
great importance. The subgenus-specific PCR could also facilitate the prim
ary classification of unknown virus isolates. J. Med Virol. 58:87-92, 1999.
(C) 1999 Wiley-Liss, Inc.