IN-SITU PCR LOCALIZATION OF HERPES-SIMPLEX VIRUS-DNA SEQUENCES IN DISSEMINATED NEONATAL HERPES-ENCEPHALITIS

To more precisely define the role of herpes simplex virus (HSV) in dev elopment of nervous system disease in neonates with disseminated infec tion, an in situ polymerase chain reaction (ISPCR) method was used to detect and localize HSV DNA sequences in paraffin sections of neural a nd non-neural autopsy tissues. In subregions of adjacent sections corr esponding to ISPCR-labeled and unlabeled areas, HSV specificity was ve rified using solution PCR and Southern blots. In serial sections, ISPC R results were compared to lesions, HSV antigen and, in selected sampl es, to viral sequence detection by in situ hybridization. By ISPCR, HS V-specific labeling was limited to HSV-infected neonates and experimen tally infected mouse controls. ISPCR-labeled cells corresponded to reg ions that were histologically abnormal and contained HSV antigen or in situ hybridization signal in some foci; in others, labeled cells were in areas with no evident lesions or antigen. Results suggest two rout es of HSV spread to the CNS: (i) blood-borne infection, with HSV DNA i n splenic lymphocytes, circulating cells, meningeal vessel walls and c ells in intraventricular hemorrhage, and (ii) neural spread, with HSV detected in brain stem sensory neurons. In the brain of one neonate su rviving acute infection, detection of HSV nucleic acid sequences sugge sts a latent or persistent viral genome. With other methods, this high ly sensitive ISPCR technique permits a more complete definition of HSV infection in these infants and provides new insights into disease mec hanisms. Fuller understanding of HSV persistence and recurrent neurolo gical disease in survivors will require further studies using these an d other techniques in human tissues and in animal models.