title: Multiplexed Luminex xMAP assay for detection and identification of five adenovirus serotypes associated with epidemics of respiratory disease in adults.
abstract: Several serotypes of human adenovirus (HAdV) cause acute respiratory disease (ARD) among healthy adults, sometimes generating broad outbreaks with high attack rates and occasional fatalities. Timely serotype identification provides valuable epidemiological information and significantly contributes to prevention (vaccination) strategies. The prevalence of specific serotypes causing ARD varies geographically. HAdV-3, HAdV-4, HAdV-7, HAdV-14, and HAdV-21 are the serotypes most commonly found in adult populations in the Western Hemisphere. Unfortunately, conventional serotype identification is a tedious process which can take a week or longer. For this reason, new molecular methods for serotype identification are needed. Commercially available rapid antigen and PCR assays for the detection of HAdV are universal but do not distinguish between the different serotypes. We describe the development of a sensitive and specific multiplex assay capable of identifying serotypes 3, 4, 7, 14, and 21. Two sets of primers were used for nonspecific (universal) PCR amplification, and serotype-specific probes coupled to Luminex tags were used for target-specific extension (TSE). PCR and TSE primers were designed using known hexon gene sequences of HAdV. The TSE products of HAdV-3, HAdV-4, HAdV-7, HAdV-14, and HAdV-21 were correctly identified using the Luminex xMAP fluid microsphere-based array system. No cross-reactivity with other respiratory pathogens or other HAdV serotypes was observed. This multiplexed assay can be expanded to include more serotypes and will allow broad and rapid detection and identification of adenoviral serotypes in a high-throughput environment.
authors.affiliation: Division of Viral Diseases, Walter Reed Army Institute of Research, Silver Spring, Maryland 20910, USA. Cicely.Washington@us.army.mil
authors.forename: Cicely
authors.initials: C
authors.lastname: Washington
authors.forename: David
authors.initials: D
authors.lastname: Metzgar
authors.forename: Manzour Hernando
authors.initials: MH
authors.lastname: Hazbón
authors.forename: Leonard
authors.initials: L
authors.lastname: Binn
authors.forename: Arthur
authors.initials: A
authors.lastname: Lyons
authors.forename: Carl
authors.initials: C
authors.lastname: Coward
authors.forename: Robert
authors.initials: R
authors.lastname: Kuschner
mesh_terms: D000258:Adenovirus Infections, Human / Q000175:diagnosis* / Q000453:epidemiology; D000260:Adenoviruses, Human / Q000145:classification* / Q000302:isolation & purification*; D000328:Adult; D017931:DNA Primers / Q000235:genetics; D004196:Disease Outbreaks; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D012140:Respiratory Tract Diseases / Q000453:epidemiology / Q000821:virology*; D012680:Sensitivity and Specificity; D014773:Virology / Q000379:methods*
chemical_list: D017931:DNA Primers
references.citation: J Med Virol. 1997 Mar;51(3):198-201
references.pmid: 9139083
references.citation: J Clin Microbiol. 2006 Aug;44(8):2808-15
references.pmid: 16891496
references.citation: J Clin Microbiol. 2008 Dec;46(12):3997-4003
references.pmid: 18923006
references.citation: J Clin Microbiol. 2004 Apr;42(4):1686-93
references.pmid: 15071026
references.citation: J Clin Virol. 2006 Apr;35(4):381-7
references.pmid: 16406799
references.citation: Am J Epidemiol. 1983 Apr;117(4):455-66
references.pmid: 6301263
references.citation: J Clin Microbiol. 2004 Feb;42(2):670-3
references.pmid: 14766835
references.citation: J Med Virol. 1999 Sep;59(1):66-72
references.pmid: 10440810
references.citation: J Clin Microbiol. 2007 Sep;45(9):2965-70
references.pmid: 17596360
references.citation: J Clin Microbiol. 2009 Mar;47(3):697-703
references.pmid: 19109466
references.citation: Ann N Y Acad Sci. 1980;354:16-42
references.pmid: 6261642
references.citation: J Clin Microbiol. 2001 Aug;39(8):2984-6
references.pmid: 11474029
references.citation: J Clin Microbiol. 2005 Nov;43(11):5743-52
references.pmid: 16272512
references.citation: J Clin Microbiol. 2003 Oct;41(10):4636-41
references.pmid: 14532195
references.citation: J Clin Microbiol. 2004 Sep;42(9):3963-9
references.pmid: 15364976
references.citation: J Med Virol. 2008 May;80(5):856-65
references.pmid: 18360899
references.citation: Emerg Infect Dis. 2006 Jun;12(6):921-30
references.pmid: 16707047
references.citation: Acta Microbiol Hung. 1993;40(4):303-23
references.pmid: 7976207
references.citation: Mil Med. 1995 Jun;160(6):300-4
references.pmid: 7659229
references.citation: Arch Virol. 1986;91(3-4):277-90
references.pmid: 3022681
references.citation: J Virol Methods. 1998 Apr;71(2):187-96
references.pmid: 9626952
references.citation: Clin Infect Dis. 2002 Oct 1;35(7):808-18
references.pmid: 12228817
references.citation: J Med Virol. 2001 Dec;65(4):710-8
references.pmid: 11745936
references.citation: Clin Infect Dis. 2002 Mar 1;34(5):577-82
references.pmid: 11803503
references.citation: Lancet Infect Dis. 2003 Feb;3(2):79-86
references.pmid: 12560192
references.citation: Clin Chem. 2005 Aug;51(8):1365-73
references.pmid: 15951322
references.citation: J Clin Microbiol. 2008 Sep;46(9):3056-62
references.pmid: 18632902
references.citation: J Clin Microbiol. 2001 Feb;39(2):498-505
references.pmid: 11158096
references.citation: J Immunol. 1960 Jun;84:562-8
references.pmid: 13841541
references.citation: J Clin Virol. 2007 Nov;40 Suppl 1:S39-46
references.pmid: 18162254
references.citation: Clin Microbiol Rev. 1992 Jul;5(3):262-74
references.pmid: 1323383
references.citation: J Med Virol. 2001 Aug;64(4):537-42
references.pmid: 11468741
references.citation: J Clin Microbiol. 2000 Nov;38(11):4114-20
references.pmid: 11060077
references.citation: J Infect Dis. 1982 May;145(5):678-82
references.pmid: 7042855
references.citation: J Infect Dis. 2007 Nov 15;196(10):1465-73
references.pmid: 18008225