Quantitation of latent varicella-zoster virus and herpes simplex virus genomes in human trigeminal ganglia

Abstract

Using real-time fluorescence PCR, we quantitated the numbers of copies of latent varicella-zoster virus (VZV) and herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2) genomes in 15 human trigeminal ganglia. Eight (53%) and 1 (7%) of 15 ganglia were PCR positive for HSV-1 or -2 glycoprotein G genes, with means of 2,902 +/- 1,082 (standard error of the mean) or 109 genomes/10(5) cells, respectively. Eleven of 14 (79%) to 13 of 15 (87%) of the ganglia were PCR positive for VZV gene 29, 31, or 62. Pooling of the results for the three VZV genes yielded a mean of 258 +/- 38 genomes/10(5) ganglion cells. These levels of latent viral genome loads have implications for virus distribution in and reactivation from human sensory ganglia.

FIG. 1

Standard curves for QF-PCR assays for VZV genes encoding ORFs 29 and 62 and gB. Shown are the C_t values at which a given input of a VZV DNA-containing plasmid was detected by PCR. Plasmid concentrations tested ranged from 10⁰ to 10⁶ copies per reaction mixture. The Spearman rank coefficient of correlation (r) for lines fitting each standard curve are given.

FIG. 2

Copies of VZV, HSV-1, and HSV-2 genes in human trigeminal ganglia. The mean ± SEM number of copies per 10⁵ cells for each gene in the PCR-positive ganglia are tabulated below the figure. The filled circles show the gene copy numbers for samples that exceeded the assay limit of detection (≥10 copies/500 ng of total DNA). Open circles represent values for samples for which gene copy numbers may have been extrapolated from the QF-PCR results but were below the threshold of reliable DNA detection (<10 copies/500 ng of DNA). The inability to define a precise threshold for positive ganglia on the per-10⁵-cell basis used here reflects the variability in β-actin gene number per 500 ng of total DNA from the individual ganglia.