The Affect of the Oral Microbiome in Health and Disease on the Uptake of HPV in Oral Keratinocyte Cells

Background: Human papilloma virus oncogenic subtype 16 (HPV 16) entry into oral keratinocytes can result in an enhanced risk for malignant transformation. Although epidemiological evidence suggests several factors that may increase risk of HPV-16 positive oropharyngeal squamous cell carcinoma (OPSCC), there is not an assay that can predict a particular individual’s risk for infection with the virus. Currently, identification of a current HPV 16 identification is used as an indication for future rates of infection of HPV 16 without determination immediate entry functionality. The purpose of this study was to assess immediate entry functionality independent of previous HPV 16 infection. In addition, a broader view of variables that influence HPV 16 entry were explored. Methods: Oral cytology samples were taken from 24 human subjects at various sites in the oral cavity including the oropharynx; with no history, clinical observation or cytopathology characteristics to identify HPV infection. The oral keratinocytes were plated and exposed to HPV-16 pseudoparticle and plasmid (PsV/pShell16) with expression of green fluorescent protein to quantify entry into the cells. Microbial samples were also taken at each site where cells were obtained. Identification of microbes was completed using a 16S rRNA pyrosequencing technique. Results: Subjects who were smokers had a mean entry of 65.5% (+/- 7.2%) at the oropharynx site, which was more than two times that seen in the non-smokers (23.5 +/- 8.0%). Subjects who were smokers and had a diagnosis of moderate to severe periodontitis had significantly less diverse microbial populations with reduced presence of Nesisseria spp., Haemophilus spp., Porphyromonas spp. and Gamellaceae spp.. Subjects with periodontitis who were not smokers demonstrated more diversity of the microbiome compared to non-periodontitis subjects with increased presence of Novoshingobium spp., Prevotella spp., and Rothia spp.. Conclusion: Our results suggest HPV 16 entry may be affected by the oral microbiome and tobacco smoke components in subjects that had no prior history of HPV 16 infection. Tobacco smoke association with enhanced periodontal disease risk also is associated with DNA damage and a generalized change in oral microbiome diversity, number and bacterial genera.