Herpes Simplex Virus 1 Reactivates from Autonomic Ciliary Ganglia Independently from Sensory Trigeminal Ganglia To Cause Recurrent Ocular Disease

Abstract

Herpes simplex virus 1 (HSV-1) and HSV-2 establish latency in sensory and autonomic neurons after ocular or genital infection, but their recurrence patterns differ. HSV-1 reactivates from latency to cause recurrent orofacial disease, and while HSV-1 also causes genital lesions, HSV-2 recurs more efficiently in the genital region and rarely causes ocular disease. The mechanisms regulating these anatomical preferences are unclear. To determine whether differences in latent infection and reactivation in autonomic ganglia contribute to differences in HSV-1 and HSV-2 anatomical preferences for recurrent disease, we compared HSV-1 and HSV-2 clinical disease, acute and latent viral loads, and viral gene expression in sensory trigeminal and autonomic superior cervical and ciliary ganglia in a guinea pig ocular infection model. HSV-2 produced more severe acute disease, correlating with higher viral DNA loads in sensory and autonomic ganglia, as well as higher levels of thymidine kinase expression, a marker of productive infection, in autonomic ganglia. HSV-1 reactivated in ciliary ganglia, independently from trigeminal ganglia, to cause more frequent recurrent symptoms, while HSV-2 replicated simultaneously in autonomic and sensory ganglia to cause more persistent disease. While both HSV-1 and HSV-2 expressed the latency-associated transcript (LAT) in the trigeminal and superior cervical ganglia, only HSV-1 expressed LAT in ciliary ganglia, suggesting that HSV-2 is not reactivation competent or does not fully establish latency in ciliary ganglia. Thus, differences in replication and viral gene expression in autonomic ganglia may contribute to differences in HSV-1 and HSV-2 acute and recurrent clinical disease.

FIG 1

Acute severity and cumulative recurrences in the guinea pig ocular model. (A) Severity of acute infection from 1 to 14 dpi, graphed as the mean lesion score for each group of guinea pigs on each day of observation, based on a scale of 0 to 4 (0, no symptoms; 1, inflammation or redness; 2, 1 or 2 lesions; 3, 3 to 5 lesions; 4, >5 lesions or coalescence of lesions); HSV-1, n = 40; HSV-2, n = 42; P = 0.002 by Mann-Whitney test. (B) Cumulative recurrences per guinea pig for each group during latent infection from 15 to 60 dpi; HSV-1, n = 11; HSV-2, n = 9; P = 0.020 by Mann-Whitney test. (C) Representative images of HSV-1 and HSV-2 acute (day 7) and recurrent (HSV-1, day 22; HSV-2, day 39) ocular disease.

FIG 2

Viral DNA quantities and thymidine kinase gene expression of HSV-1 and HSV-2 in sensory and autonomic ganglia of guinea pigs. HSV-1 and HSV-2 viral DNA extracted from ganglia was quantified by qPCR in sensory trigeminal ganglia (TG) (A), sympathetic superior cervical ganglia (SCG) (B), and parasympathetic ciliary ganglia (CG) (C). HSV-1 and HSV-2 viral gene thymidine kinase (TK) copy number was quantified by qRT-PCR in sensory trigeminal ganglia (D), sympathetic superior cervical ganglia (E), and parasympathetic ciliary ganglia (F). (n = 2 to 4 samples per group per time point.)

FIG 3

Immediate early gene expression in sensory and autonomic ganglia of guinea pigs. HSV-1 and HSV-2 viral immediate early (IE) ICP0 gene expression was quantified by qRT-PCR in sensory trigeminal ganglia (A), sympathetic superior cervical ganglia (B), and parasympathetic ciliary ganglia (C). HSV-1 and HSV-2 viral IE ICP27 gene expression was quantified by qRT-PCR in sensory trigeminal ganglia (D), sympathetic superior cervical ganglia (E), and parasympathetic ciliary ganglia (F). (n = 2 to 4 samples per group per time point.)

FIG 4

LAT gene expression in sensory and autonomic ganglia of guinea pigs. HSV-1 and HSV-2 viral gene LAT copy number was quantified by qRT-PCR in sensory trigeminal ganglia (A), sympathetic superior cervical ganglia (B), and parasympathetic ciliary ganglia (C). (n = 2 or 3 samples per group per time point.)

FIG 5

HSV-1 and HSV-2 in cultured murine primary neurons. (A) Percentage of cultured primary adult murine neurons productively infected in vitro with HSV-1 or HSV-2 at an MOI of 10 for 10 h, immunostained for HSV antigens with polyclonal antisera, in trigeminal ganglia (TG, P = 0.045, n = 30 cultures/virus), superior cervical ganglia (SCG, P = 0.015, n = 8 cultures/virus), and ciliary ganglia (CG, P = 0.880, n = 4 cultures/virus). (B) Cumulative number of reactivating neurons over 3 days ex vivo from ganglia harvested and cultured from adult mice latently infected with GFP-expressing HSV-1 or HSV-2. The graph represents three separate experiments, including 10 mice/group for each experiment.