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Journal of Virology, July 2007, p. 7710-7715, Vol. 81, No. 14
0022-538X/07/$08.00+0     doi:10.1128/JVI.02750-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Ultracentrifugation of Serum Samples Allows Detection of Hepatitis C Virus RNA in Patients with Occult Hepatitis C{triangledown}

Javier Bartolomé,1,{dagger} Juan Manuel López-Alcorocho,1,{dagger} Inmaculada Castillo,1 Elena Rodríguez-Iñigo,1 Juan Antonio Quiroga,1 Ricardo Palacios,2 and Vicente Carreño1*

Fundación Estudio Hepatitis Virales, Madrid, Spain,1 Laboratorios Diater, San Martín de la Vega, Madrid, Spain2

Received 14 December 2006/ Accepted 23 April 2007

Occult hepatitis C virus (HCV) infection of patients with abnormal liver function tests of unknown origin who are anti-HCV and serum HCV RNA negative but who have HCV RNA in the liver has been described. As HCV replicates in the liver cells of these patients, it could be that the amount of circulating viral particles is under the detection limit of the most sensitive techniques. To prove this hypothesis, serum samples from 106 patients with occult HCV infection were analyzed. Two milliliters of serum was ultracentrifuged over a 10% sucrose cushion for 17 h at 100,000 x gav, where av means average, and HCV RNA detection was performed by strand-specific real-time PCR. Out of the 106 patients, 62 (58.5%) had detectable serum HCV RNA levels after ultracentrifugation, with a median load of 70.5 copies/ml (range, 18 to 192). Iodixanol density gradient studies revealed that HCV RNA was positive at densities of 1.03 to 1.04 and from 1.08 to 1.19 g/ml, which were very similar to those found in the sera of patients with classical chronic HCV infection. Antigenomic HCV RNA was found in the livers of 56 of 62 (90.3%) patients with detectable serum HCV RNA levels after ultracentrifugation, compared to 27 of 44 (61.4%) negative patients (P < 0.001). No differences in the median loads of antigenomic HCV RNA between patients with an those without serum HCV RNA (4.5 x 104 [range, 7.9 x 102 to 1.0 x 106] versus 2.3 x 104 [range, 4.0 x 102 to 2.2 x 105]) were found. Alanine aminotransferase and gamma-glutamyl transpeptidase levels, liver necroinflammatory activity, and fibrosis did not differ between both groups. In conclusion, HCV RNA can be detected in the sera of patients with occult HCV infection after circulating viral particles are concentrated by ultracentrifugation.

* Corresponding author. Mailing address: Fundación para el Estudio de las Hepatitis Virales, Guzmán el Bueno 72, 28015 Madrid, Spain. Phone: 34-91-544-60-13. Fax: 34-91-544-92-28. E-mail: fehvhpa{at}fehv.org

{triangledown} Published ahead of print on 2 May 2007.

{dagger} J.B. and J.M.L.-A. have contributed equally to this work.

Journal of Virology, July 2007, p. 7710-7715, Vol. 81, No. 14
0022-538X/07/$08.00+0     doi:10.1128/JVI.02750-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.



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