Many viruses have the capacity to prevent a cell from being infected by a second virus, often termed
superinfection exclusion. Alphaherpesviruses, including the human pathogen herpes simplex virus 1 (HSV-1) and the animal herpesvirus pseudorabies virus (PRV), encode a membrane-bound
glycoprotein, gD, that can interfere with subsequent virion entry. We sought to characterize the timing and mechanism of
superinfection exclusion during HSV-1 and PRV
infection. To this end, we utilized recombinant viruses expressing fluorescent
protein (FP) markers of
infection that allowed the visualization of
viral infections by microscopy and flow cytometry as well as the differentiation of viral progeny. Our results demonstrated the majority of HSV-1- and PRV-infected cells establish
superinfection exclusion by 2 h postinfection. The modification of
viral infections by virion inactivation and
phosphonoacetic acid,
cycloheximide, and
actinomycin D treatments indicated new
protein synthesis is needed to establish
superinfection exclusion. Primary
infection with gene deletion PRV recombinants identified that new gD expression is not required to establish
superinfection exclusion of a secondary viral inoculum. We also identified the timing of
coinfection events during axon-to-cell spread, with most occurring within a 2-h window, suggesting a role for cellular
superinfection exclusion during neuroinvasive spread of
infection. In summary, we have characterized a gD-independent mechanism of
superinfection exclusion established by two members of the alphaherpesvirus family and identified a potential role of exclusion during the pathogenic spread of
infection.
IMPORTANCE:
Superinfection exclusion is a widely observed phenomenon initiated by a primary
viral infection to prevent further viruses from infecting the same cell. The capacity for alphaherpesviruses to infect the same cell impacts rates of interviral recombination and disease. Interviral recombination allows genome diversification, facilitating the development of resistance to
antiviral therapeutics and evasion of
vaccine-mediated immune responses. Our results demonstrate
superinfection exclusion occurs early, through a gD-independent process, and is important in the directed spread of
infection. Identifying when and where in an infected host viral genomes are more likely to coinfect the same cell and generate viral recombinants will enhance the development of effective
antiviral therapies and interventions.