CVI Accepts, published online ahead of print on 25 February 2009

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Clin. Vaccine Immunol. doi:10.1128/CVI.00470-08
Copyright (c) 2009, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Evidence of prior exposure to Human Bocavirus: a retrospective serological study of 404 adult sera in the United States

Sylvain Cecchini, Alejandro Negrete, Tamas Virag, Barney Graham, Jeffrey I. Cohen, and Robert M. Kotin^*

Laboratory of Biochemical Genetics; National Heart, Lung, and Blood Institute, National Institutes of Health, Viral Pathogenesis Laboratory; Vaccine Research Center, NIAID, NIH, National Institute of Allergy and Infectious Disease, National Institutes of Health, Laboratory of Clinical Infectious Diseases; National Institute of Allergy and Infectious Disease, National Institutes of Health

^* To whom correspondence should be addressed. Email: kotinr{at}nhlbi.nih.gov.

Recently, molecular screening for pathogenic agents has identified a partial genome of a novel parvovirus, called human bocavirus (HBoV). The presence of this newly described parvovirus correlated with upper and lower respiratory tract infections (LRTI) in children. LRTIs are a leading cause of hospital admission in children and the etiological agent has not been identified in up to 39% of these cases. Using baculovirus expression vectors (BEVs) and insect cell system; we produced virus-like-particles (VLP) of HBov. The engineered BEVs express the HBov capsid proteins stoichiometrically from a single open reading frame. Three capsid proteins assemble into the VLP rather than two proteins predicted from the HBoV genome sequence. The denatured capsid proteins, VP1, VP2, and VP3, resolve on silver-stained SDS-polyacrylamide gels as three bands with apparent molecular masses of 72 kDa, 68 kDa, and 62 kDa, respectively. VP2 apparently initiates at a GCT codon (alanine) 273 nucleotides downstream from VP1 start site and 114 nucleotides upstream from the VP3 initiation site. We characterized the stable capsids using physical, biochemical, and serological techniques. We found that the density of the VLP is 1.32g/cm³ and consistent of an icosahedral symmetry with approximately 25 nm diameter. Rabbit anti-serum against the capsid of HBoV, which did not cross-react with adeno-associated virus type 2, was used to develop enzyme-linked immunosorbant assays (ELISA) for anti-HBoV antibodies in human serum. Using ELISA, we tested and 404 human serum samples and established a range of antibody titers in a large United States adult population sample.