Studies on the microbiome of oral squamous cell carcinoma (OSCC) have been limited to 16S rRNA gene sequencing. Here, laser microdissection coupled with brute-force, deep metatranscriptome sequencing was employed to simultaneously characterize the microbiome and host transcriptomes and predict their interaction in OSCC. The analysis involved 20 HPV16/18-negative OSCC tumor/adjacent normal tissue pairs (TT and ANT) along with deep tongue scrapings from 20 matched healthy controls (HC). Standard bioinformatic tools coupled with in-house algorithms were used to map, analyze, and integrate microbial and host data. Host transcriptome analysis identified enrichment of known cancer-related gene sets, not only in TT versus ANT and HC, but also in the ANT versus HC contrast, consistent with field cancerization. Microbial analysis identified a low abundance yet transcriptionally active, unique multi-kingdom microbiome in OSCC tissues predominated by bacteria and bacteriophages. HC showed a different taxonomic profile yet shared major microbial enzyme classes and pathways with TT/ANT, consistent with functional redundancy. Key taxa enriched in TT/ANT compared with HC wereCutibacterium acnes,Malassezia restricta, Human Herpes Virus 6B, and bacteriophage Yuavirus. Functionally, hyaluronate lyase was overexpressed byC. acnesin TT/ANT. Microbiome-host data integration revealed that OSCC-enriched taxa were associated with upregulation of proliferation-related pathways. In a preliminaryin vitrovalidation experiment, infection of SCC25 oral cancer cells withC. acnesresulted in upregulation of MYC expression. The study provides a new insight into potential mechanisms by which the microbiome can contribute to oral carcinogenesis, which can be validated in future experimental studies.Significance:Studies have shown that a distinct microbiome is associated with OSCC, but how the microbiome functions within the tumor interacts with the host cells remains unclear. By simultaneously characterizing the microbial and host transcriptomes in OSCC and control tissues, the study provides novel insights into microbiome-host interactions in OSCC which can be validated in future mechanistic studies.