HSV-1 Virus Isolation in Human Trigeminal Ganglia from Cadavers

Objectives:After primary infection, Herpes simplex virus type 1 (HSV-1) becomes latent in human trigeminal ganglia (TG). Previously, we detected HSV-1 DNA bilaterally by PCR in more than half of 120 cadavers in forensic autopsies. Therefore, we tried to isolate HSV-1 virus from human TG to determine the comparative genotypes bilaterally and superinfection in one side.

Methods:Sixteen TG were obtained bilaterally from 8 cadavers. Half of each sample was used for isolation of HSV-1. An explantation was achieved by the placement of fragments of TG on Vero cell monolayers.　The culture medium was collected on from 2-11 days. The DNA from the 200 µL culture medium was extracted using a DNA extraction Kit. Furthermore, DNA was extracted directly from the remaining half of each sample. Using these DNA, two positions in the stable RL2 region (147 bp) and variable UL3 and UL4 regions (666 bp) were PCR-amplified.

Results:The virus cytopathic effect (CPE) was not observed in any of the 16 samples. The DNA amounts from the culture medium decreased as the culture time passed in almost samples. However, PCR products were amplified in 12 samples in the RL2 region and 8 samples in the UL3 and UL4 regions without regard to the culture time. In the other samples, PCR became difficult as the culture time passed. Using DNA extracted directly from TG, PCR products were amplified in all samples.

Conclusion:In this study, the culture-negative HSV-1 virus and PCR-positive HSV-1 DNA from the culture medium turned out to exist in human TG from cadavers whose time after death was 1~2 days. It is assumed that the latent HSV-1 virus was reactivated and released into the culture medium.