Viral Oral Infections in Immunosuppression and Cancer Section

National Institute Of Allergy And Infectious Diseases

Investigators

Abstract

The oral cavity, a major portal to viral infection, harbors viral lesions and cancers and is central to viral transmission. T Among our major focuses in FYI 2025 was determining the relationships between periodontitis severity and oral viral loads of oncogenic virus such as Epstein-Barr virus by qPCR. This suggests periodontal pathogens are directly involved in driving lytic viral gene expression and subsequent virus production. Determining bacterial factors that contribute to increased viral loads in the oral cavity provides insight into in vivo mechanisms that drive pathogenesis. Furthermore, this provides insights to strategies that could circumvent bacterial-driven viral gene expression/particle production, thereby decreasing viral pathogenesis/tumorigenesis. Our findings demonstrate the importance of bacterial end products derived from pure cultures of oral pathogens as drivers of multiple signaling pathways (as determined by pharmacologic inhibition, immunoblot, phosphokinome analysis, RNA seq) with resultant epigenetic modification occurring at both EBV and cellular promoters (ChIP analysis). Resultant reactivation allowed for full permissive infection in both lymphoid and epithelial cells as determined by RT-qPCR and immunoblot analysis. Viral transfer from the lymphoid to the epithelial compartment occurred as determined by transfer of virus genome-encoded GFP in lymphoid cells to the null GFP epithelial cells. We are currently determining specific activating bacterial components and their role in activating upstream cellular and viral G protein-coupled receptors. Oral oncogenic HPV infection is a significant contributor to head and neck cancers. The described project above is being expanded to assess bacteria-driven HPV. Understanding the role that specific HIV1-encoded proteins may play to increase oncogenic HPV gene expression/viral load in the oral cavity will lead to better monitoring and to decrease development of HPV-associated head and neck cancers in PLWH. We have studied a role for HIV tat, a protein secreted into saliva even in the context of ART, in the late phase HPV replication. Co-transfection of telomerase-immortalized human oral keratinocytes (OKF6tert1) with a HIVtat expression vector and a GFP/HPV16L1 fusion construct was used to determine that HIV tat can relieve inhibition of HPV16 L1 gene expression, potentially promoting increased viral production. We are currently determining whether relief of inhibition is due to increased RNA stability/nuclear export by cellular fractionation and RT-qPCR. We have also assessed the interaction of a wildtype oral HPV isolate with the differentiation machinery of the oral epithelia utilizing OKF6tet1 cells treated with sodium butyrate to induce differentiation. HIV salivary gland disease (HIVSGD) is an AIDS defining illness, characterized by a Sjogren’s disease-like phenotype that contributes to oral pathologies in PLWH. We have previously shown that BK polyomavirus (BKpyV) is frequently found in the saliva of patients with HIVSGD (qPCR), is capable of permissive replication in salivary gland cells (immunofluorescence assay) and is associated with decreased amylase production (immunoblot) in these cells suggesting that it is a likely etiological agent of salivary gland dysfunction in PLWH. Determining the BKpyV-encoded factors that contribute to HIVSGD will aid in the development of strategies to prevent salivary gland dysfunction in PLWH. Toward this end we have generated reagents (myc-tagged Tag, tag, Agnoprotein VP1, VP2, VP3 and expression constructs for the virus-encoded microRNAs) that will allow for systematic analysis of BKV products and salivary gland cell function. These reagents have been validated by immunoblot analysis and RT-qPCR.